ASSOCIATION BETWEEN MICROALBUMINURIA AND DIABETIC

i

ASSOCIATION BETWEEN MICROALBUMINURIA AND DIABETIC

RETINOPATHY IN TYPE 2 DIABETES PATIENTS . A ONE YEAR CROSS –

SECTIONAL STUDY

BY

DR. N. RAHUL NAGMBBS

Dissertation Submitted to the

Rajiv Gandhi University of Health Sciences, Bengaluru, Karnataka.

In partial fulfillment

of the requirements for the degree of

DOCTOR OF MEDICINE

in

GENERAL MEDICINE

Under the Guidance of

DR. SIDDALINGA REDDYMD

ASSOCIATE PROFESSOR

DEPARTMENT OF GENERAL MEDICINE

NAVODAYA MEDICAL COLLEGE, HOSPITAL AND

RESEARCH CENTRE, RAICHUR -584103

2011

II

RAJIV GANDHI UNIVERSITY OF HEALTH SCIENCES,

KARNATAKA, BANGALORE

DECLARATION BY THE CANDIDATE

I hereby declare that this dissertation/thesis entitled “ASSOCIATION

BETWEEN MICROALBUMINURIA AND DIABETIC

RETINOPATHY IN TYPE 2 DIABETES PATIENTS – A ONE YEAR

CROSS-SECTIONAL STUDY” is a bonafide and genuine research work carried

out by me under the guidance of Dr. SIDDALINGA REDDY, MD., Associate

Professor, Department of General Medicine, Navodaya Medical College, Hospital &

Research Centre, Raichur.

Date: Dr. N. RAHUL NAG

Place: Raichur

III



CERTIFICATE BY THE GUIDE

This is to certify that the dissertation entitled “ASSOCIATION



CROSS-SECTIONAL STUDY” is a bonafide research work done by Dr. N.

RAHUL NAG in partial fulfillment of the requirement for the degree of DOCTOR

OF MEDICINE IN GENERAL MEDICINE.

Date:

Place: Dr. Siddalinga Reddy MD. (General Medicine)

Associate Professor,

Dept. of General Medicine

Navodaya Medical College, Raichur

IV



CERTIFICATE BY THE CO-GUIDE





RAHUL NAG in partial fulfillment of the requirement for the degree of DOCTOR

OF MEDICINE IN GENERAL MEDICINE.

Date:

Place: Dr. Anupama. T M.S.. (Ophth.)

Professor & Head,

Dept. of Ophthalmology

Navodaya Medical College, Raichur

V



ENDORSEMENT BY THE HOD, PRINCIPAL/ HEAD OF

THE INSTITUTION





RAHUL NAG under the guidance of Dr. SIDDALINGA REDDY, MD. (General

Medicine), Associate Professor, Department of General Medicine, Navodaya Medical

College.

Dr. S.S. ANTIN Dr.Khaja Naseeruddin Professor & Head. Principal

Dept. of General Medicine Navodaya Medical College,

Hospital & Research Centre, Raichur

Date: Date:

Place: Raichur Place: Raichur

VI



COPYRIGHT

DECLARATION BY THE CANDIDATE

I hereby declare that the Rajiv Gandhi University of Health Sciences,

Karnataka shall have the rights to preserve, use and disseminate this dissertation

/thesis in print or electronic format for academic /research purpose.

Date:

Place: Raichur Dr. N. RAHUL NAG

© Rajiv Gandhi University of Health Sciences, Karnataka.

VII

ACKNOWLEDGMENT

At the outset I thank the Lord, Almighty, for giving me the strength to perform all

my duties.

It is indeed a great pleasure to recall the people who have helped me in

completion of dissertation. Naming all the people who have helped me in achieving this

goal would be impossible, yet I attempt to thank a select few, who have helped me in

diverse ways.

It gives me immense pleasure to express my deep sense of gratitude and

indebtedness that I feel towards my teacher and guide Dr. Siddalinga Reddy, MD,

General Medicine, Associate Professor, Department of General

Medicine,Dr.Anupama.T MS Ophthalmology Professor and Head for their valuable

suggestions, guidance, great care and attention to detail that he has so willingly shown in

the preparation of this dissertation.

I acknowledge and express my humble gratitude and sincere thanks to my beloved

teacher Dr.S.S. Antin, MD., General Medicine, Professor and H.O.D., for his constant

help to undertake this study

I owe a great deal of respect and gratitude to Dr.Chaitanya Kumar Swamy, MD,

General Medicine, Associate Professor, Dr. Krishna Prasad, MD, General Medicine,

Associate Professor, Dr.Ajit Kumar , MD, General Medicine, Associate Professor,

Dr.Ramakrishna. M.R., MD, General Medicine, Assistant Professor, Dr.N.S. Javali,

MD, General Medicine, Assistant Professor, Dr. Imran, General Medicine, Assistant

Professor, Dr. Amar S. Patil, General Medicine, Assistant Professor, Dr. Anand

Chaudhuri, General Medicine, Assistant Professor, Dr. S.S. Reddy, General Medicine,

VIII

Assistant Professor, Dr. Vinay, General Medicine, Assistant Professor, Dr. Praveen

Bhadri, General Medicine, Assistant Professor, Navodaya Medical College, Raichur, for

their scholarly suggestions and all round encouragement.

I am extremely grateful to Dr. Khaja Naseeruddin, M.S (ENT)., Principal, and

Dr.S.R.Hegde, M.S.,Medical Director, Navodaya Medical College, Raichur, for their

valuable help and co-operation.

I am also thankful to my colleagues Dr. Novak Gupta, Dr. Deepak, Dr. Kavya,

Dr. Soumya, Dr. Darshana, Dr. Hardhik Gajera, Dr. Aniket Kataria, Dr. Harsha

Totad, Dr. Praveen Paul, Dr. Swathi, Dr. Dasrathi for their constant voluble help and

co-operation and encouragement to complete this dissertation.

I express my sincere thanks to Superintendent and Resident Medical Officers of

Navodaya Medical College Hospital for their valuable help and cooperation.

Finally, I thank all my patients who formed the back bone of this study without

whom this study would not have been possible.

Date:

Place: Dr. N. Rahul Nag

IX

LIST OF ABBREVIATIONS USED

ACE: Angiotensin Converting Enzyme

AER: Albumin Excretion Rate

AGE: Advanced Glycosylation Products

BMI: Body Mass Index

FBS: Fasting Blood Sugar

GAG: Glycosaminoglycans

GFR: Glomerular Filtration rate

HDL: High Density Lipoproteins

HTN: Hypertension

Ig: Immunoglobulin

IL: Interleukin

IGF: Insulin like Growth Factor

IHD: Ischemic Heart Disease

LDL: Low Density Lipoprotein

NIDDM: Non Insulin Dependent Diabetes Mellitus

PAS: Periodic Acid Schiff

PDGF: Platelet Derived Growth Factor

PPBS: Post Prandial Blood Sugar

PVD: Peripheral Vascular Disease

PN: Peripheral Neuropathy

TGL: Triglycerides

VLDL: Very Low Density Lipoprotein

X

XI

ABSTRACT

Background: Diabetes Mellitus is one of the most common chronic disorders and the

commonest metabolic disease affecting man. The incidence of this disease is increasing

all over the world. The magnitude of the problem is compounded by the various

complications targeting vital organs. Diabetic nephropathy is the leading cause of End

Stage Renal Disease in the world, accounting for more than one third of the cases. About

25 to 40% of type 2 diabetics will develop nephropathy eventually. The earliest evidence

of nephropathy is microalbuminuria. The association between microalbuminuria and

diabetic retinopathy is closely related and can be useful in preventing or delaying the

occurrence of diabetic retinopathy. Microalbuminuria is a marker of wide spread

microvascular damage in Type 2 diabetes mellitus. The association between overt

proteinuria and proliferative diabetic retinopathy have been demonstrated and there is

increasing evidence that microalbuminuria could be a marker of early diabetic

retinopathy.

Objectives:

To study the prevalence of microalbuminuria in Type 2 diabetes mellitus.

To study the association between microalbuminuria and diabetic retinopathy in

Type 2 diabetics.

Methods: A cross-sectional study over a period of one year. 100 cases those attending

the OPD and medical wards of Navodaya Medical College, Raichur, were enrolled for

the study.

Results: Out of 100 cases 57 cases were male and 43 cases were female.

Microalbuminuria was found in 38% of patients and diabetic retinopathy was present in

XII

44% of patients.Microalbuminuria and diabetic retinopathy was present in 31% of

patients showing that there is association between microalbuminuria and diabetic

retinopathy.

Microalbuminuria and retinopathy was found more for the age group above 50years

(p=0.053,0.001).

Conclusion:

The present study has shown that there is significant association between the

presence of microalbuminuria and retinopathy.

It has also shown that there is increase in the prevalence of microalbuminuria and

retinopathy with increasing age ,HbA1c >7%,BMI>25 Kg/m2

.

Key words: Diabetic Retinopathy, Microalbuminuria

XIII

TABLE OF CONTENTS

1. INTRODUCTION ............................. 1-2

2. AIMS AND OBJECTIVES ................................ 3

3. REVIEW OF LITERATURE ........................... 4-42

4. MATERIALS & METHODS ......................... 43-47

5. OBSERVATION & RESULTS ......................... 48-59

6. DISCUSSION ......................... 60-64

7. CONCLUSION .............................. 65

8. SUMMARY .............................. 66

9. BIBLIOGRAPHY ......................... 67-80

10. ANNEXURES

* INFORMED CONSENT .............................. 81

* PROFORMA ......................... 82-89

* MASTER CHART ......................... 90-94

XIV

LIST OF TABLES

Sl.

No. Titles

Page

No.

1. Table showing age and sex distribution 48

2. Table showing number of patients with microalbuminuria and

retinopathy 50

3. Association of age with microalbuminuria and retinopathy

51

4. Association of duration of diabetes with microalbuminuria and

retinopathy 53

5. Association of HbA1c with microalbuminuria and retinopathy

55

6. Association of BMI with microalbumiuria and retinopathy

57

7. Association between microalbuminuria and retinopathy

59

XV

LIST OF GRAPHS

Sl.

No. Titles

Page

No.

1. Table showing age and sex distribution 49

2. Table showing number of patients with microalbuminuria and

retinopathy

50

3. Association of age with microalbuminuria and retinopathy 52

4. Association of duration of diabetes with microalbuminuria and

retinopathy

54

5. Association of HbA1c with microalbuminuria and retinopathy 56

6. Association of BMI with microalbumiuria and retinopathy 58

7. Association between microalbuminuria and retinopathy 59

XVI

LIST OF FIGURES

Fig.

No. Titles

Page

No.

1 MICRAL Test Strip – II 32

2 Background Diabetic Retinopathy showing scattered exudates and

haemorrhages near fovea

40

3 Background Retinopathy with severe Maculopathy and hard

exudates at macula

40

4 Proliferative Diabetic Retinopathy with disc Neovascularisation 41

5 Advanced Proliferative Diabetic Retinopathy with Tractional

Retinal detachment

41

6 Proliferative Diabetic Retinopathy with abnormal vessels at optic

disc and retina with early vitreous haemorrhage

42

1

INTRODUCTION

Diabetes mellitus, the most common endocrine disorder is characterised by

metabolic abnormalities and long-term microvascular and macrovascular

complications.

The prevalence of diabetes is on the rise, more alarmingly in the developing

countries. Besides multiplying risk for coronary heart disease, diabetes enhances the

incidences of cerebro vascular accidents. Moreover it is the leading cause of acquired

blindness and accounts for about a quarter of the cases with end stage renal disease as

well as half of the cases of non-traumatic lower limb amputations.

Diabetic nephropathy occurs in as many as 30% of insulin dependant diabetes

mellitus patients and 25% of non-insulin dependent diabetes mellitus patients.

Diabetic nephropathy is a dreaded disease with progressive and continuous

deterioration in glomerular function resulting in irreversible renal failure. Diabetic

nephropathy is an important cause of morbidity and mortality and is now among the

most common cause of end stage renal disease. However there is an early phase of

diabetic renal disease called incipient diabetic nephropathy. In this stage, there is a

rise in urinary excretion of albumin i.e. microalbuminuria. But the rise is detectable

only by use of sensitive assay for urinary albumin. At this stage urine is negative for

macro albumin and renal function is normal by standard clinical tests. The presence of

microalbuminuria precedes the development of overt diabetic nephropathy by 10 to

15 years. It is at this stage that one can hope to reverse diabetic renal disease or

prevent its progression. Therapeutic interventions which reverse microalbuminuria

2

include intensified insulin treatment, dietary protein restriction and control of

hypertension by ACE inhibitors and Beta-blockers.

Microalbuminuria thus is an important warning sign for both the physician and

the patient which if ignored can lead to irreversible renal damage.

Microalbuminuria is most commonly associated with other microvascular

complications of diabetes namely retinopathy, neuropathy, and ischemic heart disease.

So, microalbuminuria may be a marker for widespread microvascular damage in a

patient of diabetes mellitus.

The aim was to study the occurrence of microalbuminuria in patients with

non-insulin dependant diabetes mellitus and also to find out its association with the

duration of diabetes mellitus and retinopathy.

3

OBJECTIVES

To study the prevalence of microalbuminuria and retinopathy in type 2

diabetes mellitus.

To study the association between microalbuminuria and retinopathy in type2

diabetes mellitus.

4

REVIEW OF LITERATURE

HISTORICAL REVIEW:

1. Diabetes mellitus:

The knowledge of diabetes dates back to centuries before Christ. Polyuric

disease, resembling diabetes was described as early as 150 BC in ancient Egyptian

records discovered by George Beers. Celsius (30BC-50AD) had recognized the

disease. Diabetes, a Greek term, which literally means to „run thru‟ or a „siphon‟ was

initially used by Aretaeus in first century AD for the generic description of a

condition causing increased urine output1. Roman physicians thought of diabetes as a

“wonderful affection, not very frequent among men, being melted down of flesh and

limbs into urine. The patient never stopped making water, but the flow is incessant as

if from a opening of aqueducts- Aretaeus, the Cappadocian2,3

.

The association of polyuria with a sweet tasting substance in the urine was

first reported in Sanskrit literature dating from fifth to sixth centuries AD at the time

of two noted Indian physicians Susruth and Charaka.

It was in the seventeenth century that Thomas Willis (1621-1675) made the

observation “as if imbibed with honey and sugar about the diabetic urine”. A century

after Willis, Mathew Dobson (1735-1784) demonstrated that the sweetness of urine

was indeed due to sugars. It was John Rollo who was one of the first to use the

adjective mellitus (mellitus = honey) to distinguish it from other polyuric states in

which the urine was unsavory (Greek – insipidus). Over the centuries, gradually the

causes and complications of this disease were recognized. Aricanne, an Arab

physician at around the tenth century had described gangrene2,3

.

5

The diabetes world was overwhelmed with joy in 1921 when young physician

and surgeon Fredrick Grant Banting (1891-1941) and Charles.H.Best, his graduate

student assistant, working in Toronto through the summer on an almost non-existent

budget in a lab loaned to them temporarily by a vacating professor, prepared active

extracts of pancreas which lowered the elevated level of sugars in diabetic dogs. The

first patient to be treated with pancreatic extract was Leonard Thomson in 1922.The

long acting insulin preparation (isophane) was introduced in 1936 by Hans Christian

Hagedorn and his colleagues. The testing of Sulfonylureas was done by Auguste

Loubatieries in 1944.The first therapeutic use of a Biguanide was done by G.Ungar in

1957.The efficacy of insulin in preventing the complications and retarding

multisystem involvement was heralded by the fact that the untreated cases in the pre-

insulin era had a high mortality rate which was mostly due to diabetic ketoacidosis2,3

.

2. Diabetic nephropathy:

Diabetes was for many years regarded as the disease of the kidneys. This was

the opinion of Aretaeus, Capadcian in second century AD. The view was still held by

Erasmus Darwin in 1801. The presence of proteinuria in diabetes mellitus had long

been known. Contunniues (1770), Rollo (1798), Darwin (1801), Rayer (1840), Van

Noorden (1912), all had described the association of dropsy with diabetes.

Vacuolization of tubular epithelium was observed by Armani (1875) and Ebstein

(1881) and was shown to be due to glycogen infiltration by Ehlrich in 1888.

Kimmelstiel and Wilson were the first to attribute specific glomerular lesions

entitled inter capillary lesions in the glomeruli of kidney. These peripheral hyaline

masses are known as Kimmelstiel Wilson lesions. These histological features were

6

associated with clinical features of diabetes, hypertension, nephrotic syndrome and

renal failure. Fahr demonstrated soon after the diffuse lesions in 1942.

Several major studies among diabetics have been undertaken (Tuft et al 1956,

Farqahar et al 1959, Gellman et al 1959, Hatch et al 1961, O Sullivan et al, Thomsal)

and proteinuria especially when diffuse changes are considered. On the whole,

advanced clinical disease is accompanied by severe glomerular lesions5.

3. Microalbuminuria:

In 1963,Keen and Chlouvervakis developed sensitive and specific

Radioimmunoassay for detecting human albumin in low concentration i.e.

microalbuminuria, which indicate earliest stage of diabetic renal disease. Later

various other methods were developed for detection of microalbuminuria6.

DEFINITION AND PATHOPHYSIOLOGY

Microalbuminuria:

This means significant increase in albumin excretion rate (AER). Albumin

excretion in healthy individuals ranges from 1.5 to 20 mcg/min with geometric mean

in the range of 6.5mcg/min, these have been termed normoalbuminuria.

Microalbuminuria thus defines the wide substantial range of albumin hyper secretion

ranging between 20to 200mcg/min (30 to 300mg/day)7. Normal persons excrete less

than 30 mg /day. Microalbuminuria is not detected by reagent sticks for urinary

protein which generally becomes positive only when proteinuria is greater than 550

mg/day. This degree of leakage is termed macroalbuminuria8,9

.

7

In 1963, Keen and Chlouvervakis developed sensitive specific

radioimmunoassay for detecting human albumin in low concentration in urine to

study the sub clinical increase in urine albumin excretion rate (AER), which might be

the earliest pointer to the development of diabetic renal disease6.

A significant increase in mean AER was demonstrated in non insulin

dependant diabetic subjects newly detected in population survey by Keen in 1969.

This AER was an average three to four times above the normal level. Later

Mongensen et al in 1971 found increased AER in newly diagnosed youngsters with

insulin dependent diabetes mellitus as did Parving et al in short term insulin

dependent diabetes mellitus patients whose control had been deliberately worsened by

withdrawal of insulin for some days. In established conventionally treated adult

insulin dependent diabetes mellitus, patients known to be diabetic between 6 months

and 39 years, albumin and IgG excretion rates were found to be raised in 30 to 45%

patients especially those patients with duration of diabetes between 10 and 20 years8.

Term Synonym Urinary Albumin Excretion

Normoalbuminuria ----- <20 mcg/min

Microalbuminuria Incipient nephropathy 20-200 mcg/min

Macroalbuminuria

Clinical Nephropathy/

Overt Nephropathy

>200 mcg/min

Histopathology of kidney in normal individuals and diabetes10

:

The main pathological features of diabetic kidney disease occur in the

glomerulus.

8

Normal subject:

Glomerulus consists of tuft of 20 –40 capillary loops, which arise from an

afferent and drain into an efferent arteriole. Mesangial tissue, comprising both cellular

and matrix components, support lobules of capillaries. Electron microscopy of the

loops shows that each loop consists of a basement membrane lined by fenestrated

endothelium and covered by visceral epithelial cells (podocytes) carrying foot

processes that interdigitate along the membrane, leaving spaces (filtration slits or

pores) between the processes. The Bowman‟s capsule encloses the glomerular tuft,

which is continuous with the tubular basement membrane and binds the urinary space.

Filtration of plasma proceeds from the capillary, across the endothelium (probably via

the fenestrae),the basement membrane, through the slit pores of the epithelium and

into the urinary space and the proximal tubule11

.

Changes in diabetes:

In diabetes, the volume of the whole kidney and of individual glomeruli is

increased at the time of diagnosis12

and glomeruli continue to enlarge later in the

disease12,13

. Early glomerular enlargement is probably due to enhanced basement

membrane production leading to an increased in filtration surface area while later

expansion may be caused due to mesangial expansion14

. The increase in total renal

volume is likely to be caused by tubular tissue. In diabetic nephropathy, renal size is

therefore usually normal or large even when end stage renal failure develops, which is

in contrast to most causes of chronic renal failure in which renal size tends to decrease

with advancing disease. In some cases however, concomitant to a reduction renal

artery stenosis or upper urinary tract infections may contribute in overall renal size15

.

9

Basement membrane thickening has long been recognized as a pathological

hallmark of diabetes16

. Thickening can be detected within two years of the detection

of diabetes mellitus17

. Marked thickening occurs in patients with diabetes for duration

more than 10 years18

. Mesangial expansion seems to occur after the thickening of the

glomerular basement membrane, although this may not be the true sequence of events

because it is technically easier to detect changes in basement membrane thickness

than in the mesangium19,20

. Matrix accumulation rather than cellular increase accounts

for most mesangial expansion21

. Unlike basement membrane thickening, mesangial

volume may be normal in some patients after 25 years of diabetes, although those

with established nephropathy invariably have mesangial expansion18

.

Nodular lesions consisting of ovoid accumulations of periodic acid schiff

(PAS) positive material, often occupying the central mesangium of a lobule, is almost

pathognomonic of diabetes22

. When advanced, these changes are referred to as the

Kimmelstiel-Wilson kidney. Hyaline deposits also occur as eosinophilic, acellular

material but are non- specific and are found in several other renal conditions. They

can be present inside Bowman‟s capsule („capsular drop‟), between the endothelial

cell and basement membrane („fibrin cap‟), and in the afferent and efferent arterioles.

Global glomerular sclerosis or occlusion, caused by mesangial expansion or ischemia

secondary to afferent arteriolar blockage, is a feature of patients with declining GFR.

As in other types of progressive renal disease, the tubules and interstitium may show a

variety of non-specific changes22,23

.

The origin and determinants of microalbuminuria24

:

Of the small quantity of albumin and IgG normally filtered at the glomerulus,

95to 97% is reabsorbed by proximal tubules. The reabsorptive process works near

10

maximal capacity. So that moderate increase in filtered protein is reflected in an

elevation of its urinary excretory rate. If reabsorption is proportional to the filtered

load, then the excretion of albumin and IgG in the urine will change proportionally

with amount filtered, when the tubular reabsorption capacity is saturated, urinary

protein excretion will parallel filtration.

The fact that the microalbuminuria and microIgGuria of the diabetes both at

rest and response to exercise is glomerular in origin and associated with normal

tubular is supported by the findings of a normal β2 microglobulin (molecular weight

11,800 Daltons, stokes radius 16Ao) a sensitive indicator of tubular reabsorptive

capacity.

Tubular proteinuria is characterised by large increase in β2-microglobulin

excretion with small changes in albumin excretion. The moderate increases of β2-

microglobulin excretion that have been reported in diabetes have been under

conditions of extremely poor metabolic control and ketosis.

The glomerular capillary blood urine barrier can be regarded functionally as

membrane, perforated by pores of average size 555Ao and uniformly coated by

negative electric charge. Therefore the size and charge of circulating molecules as

well as transglomerular pressure gradient, the force during glomerular filtration of

macromolecules will determine the passage of protein across the barrier. In

microalbuminuric diabetic subjects, the clearance of albumin, a polyanion (PL, 48

mol.wt 49,000 stokes radius 36Ao) and IgG, a larger but electrically neutral molecule

(PL 7.5-7.8 mol.wt. 1,60,000 stokes radius 55Ao) are both increased, these early

increases are likely to be a consequence of alteration in glomerular haemodynamics,

and in particular of transglomerular pressure gradient.

11

In a diabetic patient,the enlarged filtration surface area would probably result

in greater number of pores (no larger pores have been shown at this stage of diabetes)

and influences the filtration of plasma proteins.

As microalbuminuria becomes persistent and increases in degree, the

sensitivity index i.e. clearance of IgG/clearance of albumin starts to fall reaching its

lowest values when albumin excretion is around 90 mcg/min or higher. This is due to

disproportionate increase in filtration of albumin compared with IgG, and it marks a

new selective stage of glomerular leakage of anionic albumin. At this stage, medium

sized pores are unchanged and the likely reason for the increased glomerular albumin

filtration is a loss of fixed negative electrical charge on the membrane. This would

permit increased permeation of anionic albumin but have little influence on IgG, a

neutral molecule the filtration of which is regulated by pore radius or number and

intraglomerular pressure.

The mechanism of this transition from low to high levels of microalbuminuria

is unknown; but may be a combination of deleterious effects of long-term elevation of

intraglomerular pressure and flow with cumulative metabolic abnormality of synthesis

of electronegative membrane glycosialoprotein and proteoglycans. Moreover the

recent reports indicate that the glycosylated proteins including albumin undergo the

preferential transport across the glomerular membranes. The reason for this facilitated

flux of glycosylated macromolecules through the glomerular barrier is unknown. The

transition to high selectivity proteinuria signifies the advent of heavier proteinuria.

This indicates the critical importance of loss of charge barrier in unfolding sequence

of pathogenic events.

12

Concomitants of microalbuminuria24

:

In diabetic humans, there is no correlation between glycosuria and urinary

albumin excretion. Whether acute worsening of glycemia by glucose ingestion or

infusion increases AER remains controversial. Glucose ingestion has been reported to

increase albuminuria in normal subjects but not in diabetic patients. A number of

studies have failed to show any acute effects on either oral or intra venous glucose on

urinary albumin excretion. Mild metabolic acidosis has no effect on urinary albumin

excretion but increases the excretion of β2 microglobulin. Neither growth hormone

nor glucagons affect albumin excretion in normal or in diabetic subjects.

A consistent association of microalbuminuria is found with higher level of

arterial pressure. Many investigators have confirmed a positive linear and independent

correlation between arterial pressure and AER. Furthermore, changes in blood

pressure have been shown to be positively correlated to changes in AER in a

prospective study. The association is much more closer than between AER and blood

glucose levels and independent of number of other variables including age, sex,

duration of diabetes, body mass index and blood glucose levels itself.

The observation of higher arterial pressure in microalbuminuric patients

without reduced GFR speaks against the assumption that higher blood pressure is

consequence of renal dysfunction and argues in favour of more complex relationship.

This raises the possibility that either the rise in blood pressure could be contributory

to renal disease or alternatively that microalbuminuria and higher blood pressure may

be related to a common determinant. It is of interest that microalbuminuric patients

with elevation of arterial pressure show significantly more marked mesangial

expansion than patients with lower AER and arterial pressure.

13

Macroproteinuria:

The appearance of macroproteinuria in an insulin dependent diabetic subject

initiates the phase leading through progressively declining renal function to end stage

renal failure. The GFR may be normal or even raised at the onset of macroproteinuria,

but will proceed relentlessly to fall, its rate of decline ranging from 0.6 to 2.4

ml/min/month. Urinary protein increases as the GFR falls and a highly significant

negative correlation has been found between GFR on one hand and clearance of

albumin and the other. The 24-hour excretion of albumin correlates positively with

the urinary total protein excretion.

By the stage of macroproteinuria, albumin represents approximately 50% of

total urinary proteins. This is much higher proportion than that found in subjects with

normoproteinuria (upto11%) or microproteinuria (up to 22%). In individuals with

macroproteinuric phase,excretion of total protein varies considerably from day to day,

but the fractional clearance (i.e. clearance of a protein per unit of GFR) of individual

protein is much more stable and represents a more accurate index of residual

glomerular performance. The fractional clearance of albumin and IgG increases

progressively with time and in parallel with decline in GFR.

Origin of macroproteinuria and relation to glycemia:

The increasing proteinuria is primarily glomerular in origin, tubular

reabsorptive capacity for albumin soon becomes completely saturated and changes in

urinary excretion are then a more accurate reflection of changes in transglomerular

flux. The urinary excretion rates of low molecular weight proteins, such as β2

microglobulin or lysozyme, does not go up until glomerular function is markedly

depressed and their plasma levels have risen. Saturation of tubular reabsorptive

14

capacity in functioning nephrons with protein overload may cause the rising urinary

excretion rate rather than primary tubular dysfunction. Urinary clearance of β2

microglobulin rises exponentially as GFR falls below 40ml/min/1.73m2.

In contrast to microproteinuria, the degree of macroproteinuria shows no

relationship with current level of diabetic control. Neither the mean plasma glucose

concentration nor glycosylated haemoglobin levels correlated significantly with

clearance and excretion rates of different proteins. Long term correction of

hyperglycaemia by an intensified treatment regimens, failed to stop or significantly

slow the progressive increase in fractional clearance of albumin and IgG in insulin

dependent diabetic subjects with renal failure over a period of 2 years observation24

.

ETIOPATHOGENESIS

1. Hyperglycemia:

There is overwhelming evidence linking hyperglycemia to diabetic micro

vascular and macro vascular complications. In the kidney, histological changes such

as mesangial expansion may be reversed by transplantation of diabetic kidney in to a

normal animal or by correcting diabetes with islet cell transplantation.

2. Non-enzymatic glycosylation:

One possible link between elevated glucose levels and diabetic nephropathy is

through non-enzymatic glycosylation of cellular proteins.

There are three possible mechanisms through which non-enzymatic glycation

may contribute to diabetic complications.

15

1) AGE may alter the structure and functions of extracellular matrix by cross-

linking matrix proteins.

2) AGE may affect the activity of signals such as cytokines, growth factors and

free radicals, by interacting with AGE receptors on various tissues.

3) Glycation may directly affect the functions of enzymes and other key

intracellular proteins1,25

.

3. The polyol pathway:

Sorbitol is produced in cells from glucose by reaction catalysed by aldolase

reductase. In the normal kidney, aldolase reductase is present in the papilla,

glomerular epithelial cells, distal tubular cells and also in mesangial cells. In the renal,

medullary cells of the kidney the primary role of aldolase reduction is in the

generation of Sorbitol, an organic osmolyte in response to high salinity in medullary

interstitium. Sorbitol would aid in preventing the osmotic stress. Chronic

hyperglycemia may lead to Sorbitol accumulation in a variety of tissues including

renal tubules and glomeruli. Sorbitol accumulation could cause tissue damage perhaps

by disturbing cellular osmoregulation and depleting intracellular myoinositol.

Depletion of phospoinositidase may result in reduced hydrolysis of

phosphotidylinositol bi phosphate and decreased diacylglycerol formation.

Diacylglycerol is a major endogenous cellular mediator of protein kinase C activation

which itself has been implicated in pathogenesis of diabetic renal disease26

.

4. Biochemical abnormalities of extracellular matrix:

Diabetic glomerulopathy is characterised by excessive accumulation of

glomerular basement membrane and mesangial matrix. The activity of lysyl

hydroxylase, an enzyme involved in the hydroxylation of peptide bound lysine during

16

collagen biosynthesis is found to be increased in the glomeruli of diabetic rats.

Glycosaminoglycans (GAG), polysaccharides account for approximately 90% of total

carbohydrate component of glomerular basement membrane with sialoprotein

constituting remainder.

The principal GAG in the glomerular basement membrane is heparan sulphate

that together with sialic acid contributes to negative charge of glomerular capillary

wall and thereby to charge selective properties of the filtration barrier. In diabetes,

there is reduced de novo synthesis of glomerular heparan sulphate and the total GAG

content in the glomerulus and the glomerular basement membrane is reduced. The

heparan sulphate content of glomerular basement membrane has been found to be

decreased in patients with IDDM with nephropathy. Sialoglycoproteins are highly

negatively charged and coat glomerular epithelial cells, their foot process and

epithelial slit diaphragm. A loss of negative charge of glomerular membrane may be

responsible for foot process fusion, with consequent obliteration of the slit diaphragm

and could partly explain the albuminuria of diabetic nephropathy27

.

5. Glucotoxicity:

Glucose itself may have direct toxic effects on cells. Lorenzi et al have

demonstrated cultured human endothelial cells that are chronically exposed to high

glucose concentrations show important abnormalities in cell function, which cannot

be ascribed to polyol pathway activity. Abnormalities include alteration in cell

replication and maturation associated with evidence of damage to DNA. High glucose

levels also lead to increased expression and synthesis of collagen, fibronectin and

laminin, which may partly explain the enhanced products of extracellular matrix

observed in diabetic kidneys. Mesangial cells in high glucose levels induce

17

transcription and secretion of TGF-B, which is unique among the cytokines in that it

stimulates the matrix synthesis and inhibits its degradation. Abnormalities in

endothelial cell function have been implicated in the increased frequency of

cardiovascular disease which is a feature of diabetic nephropathy. It has been shown

that such abnormalities evidenced by raised plasma levels of Von Willebrand factor

and decreased release of tissue plasminogen activator in response to exercise are

present even before overt nephropathy develops28,29,30

.

6. Haemodynamic and hypertrophic pathways:

Glomerular hemodynamic disturbances with elevation of renal blood flow and

GFR occur early in the course of diabetes have been suggested directly responsible

for development of glomerulosclerosis and attendant proteinuria. Several observations

support the notion that renal hyper perfusion and hyper filtration contribute to renal

damage. Elevated intraglomerular pressure via increased mechanical stress and shear

forces may damage the endothelial surface and disrupt the normal structure of

glomerular barrier, and could eventually lead to mesangial proliferation, increased

production of extracellular matrix and thickening of glomerular basement

membrane26

.

7. Familial and genetic pathways:

Diabetes induces important metabolic, hormonal and growth factor changes.

These changes that are related in part to the degree of glycemic control, occur in

virtually all patients, but till now it has been impossible to isolate a subset of

individuals in whom the severity of these environmental perturbations is convincingly

linked to development of these complications. On the contrary, there is ever growing

evidence that the diabetic control is only a necessary component but is not linearly

18

related to the development of renal failure. To explain the susceptibility of renal

failure in a subgroup of patients who develop renal failure alternative hypothesis has

been advocated taking into account the host response to diabetic induced

environmental disturbances.

Familial clustering of diabetic kidney disease has been reported. In IDDM

83% of siblings of proband with diabetic nephropathy have evidence of nephropathy,

compared with only 17% of diabetic sibling of probands without nephropathy31

.

Familial influence on development of nephropathy has been described in Pima

Indians with NIDDM. A familial predisposition to raised arterial pressure has been

suggested by some reports as possible contributing factor to susceptibility to

nephropathy in diabetics32

.

Sodium Lithium counter transport:

Genetically determined red cell sodium-lithium counter transport, a cell

membrane cation transport system whose elevated levels are associated with essential

hypertension has given insight into predisposition to diabetic renal disease and also

attendant cardiovascular disease33

. The rate of sodium-lithium counter transport has

been found to be higher in proteinuric patients with diabetes than in

normoalbuminuric controls. Microalbuminuric diabetic patients have also been found

to have higher sodium-lithium counter transport activity. Higher rate of counter

transport were associated with elevated LDL cholesterol, total and VLDL,

triglycerides and reduced HDL cholesterol concentrations.

The mechanism of association between sodium lithium counter transport

activity, hypertension and lipid abnormalities and susceptibility to diabetic renal and

vascular disease could be insulin resistant state. These associations (i.e. albuminuria,

19

left ventricular and renal hypertrophy and insulin resistance) were independent of

actual level of blood pressure or duration of arterial hypertension. This combination

of risk factors may not be confined to diabetic population but may be a manifestation

of syndrome in general population (Syndrome X)34,35.

Sodium-hydrogen antiporter:

Sodium-hydrogen antiporter is a cell membrane cation exchanger that

catalyses the electroneural exchange of extracellular sodium ions for intracellular

hydrogen ions with a stociometry of 1:1. Molecular ionic studies have so far revealed

the presence of five subtypes of sodium hydrogen exchangers36

.

The most widely studied sodium isoform is referred to as NHE-1, is expressed

ubiquitously. The gene for NHE-1is located on short arm of chromosome 1; and

encodes a protein of B15 amino acid with two distinct domains.

Increased sodium-hydrogen antiport activity has been reported in leucocytes

of IDDM patients with nephropathy as well as patients with essential hypertension

and on red cells from IDDM patients with microalbuminuria. In serially passaged skin

fibroblasts, Trusian et al demonstrated significantly greater sodium hydrogen antiport

activity in cells from patients with nephropathy. These findings are consistent with the

view that cells of diabetic patients who develop nephropathy have intrinsic enhanced

capacity to proliferate and this phenomenon is associated with high rates of sodium

hydrogen exchange activity. The activity of sodium hydrogen antiport seems to act as

an indicator of some mechanism possibly genetically determined controlling cell

growth and hypertrophy on one hand and intracellular homeostasis on the other.

20

The environmental changes brought about by diabetes could lead to

dysregulation of these mechanisms in susceptible individuals and induces cell

hypertrophy and hyperplasia contributing to glomerular hypertrophy and mesangial

expansion in the kidneys as well as tubular hypertrophy and hyperplasia. Increased

renal sodium reabsorption would augment transmitted systemic and renal perfusion

pressure to maintain sodium balance. The increased perfusion pressure of the

glomerular capillaries is because of generalised vasodilatation present in diabetes.

This would lead to increased intra glomerular pressure, which determines at least in

part, and increase in GFR may be responsible for disruption of glomerular

permeability properties generating proteinuria.

On the other hand progressive mesangial expansion would lead to

glomerulosclerosis and further disruption of glomerular basement membrane

permeability selective properties. The insulin resistance associated with excessive

growth and the consequent hyperinsulinemia may cause lipid abnormalities, that in

the setting of vascular hyperpermeability characteristic of diabetic microvascular

disease, would further aggravate the renal histological damage and contribute in

combination with hypertension, and accelerated atherosclerosis of diabetic renal

failure37,38,39

.

PATHOLOGY

Gross Appearance:

The kidneys are of usually the normal size. They may be enlarged in the early

stages, but later becomes contracted with granular surface. The cut surface is usually

pale and the renal arteries may show arteriosclerosis in later stages.

21

Light microscopy:

Glomerular lesions:1,40,41,42

:

1. Nodular

2. Diffuse

3. Exudative

4. Glomerular hyalinization

Nodular lesions:

The nodular lesions described by Kimmelstiel and Wilson in 1936 have been

considered for a long time virtually specific for diabetes.

The nodules are well-demarcated hard masses, eosinophilic, and periodic acid

schiff positive, located in the central regions of peripheral lobules. When not

acellular, they contain pyknotic nuclei and infrequently foam cells can be seen

surrounding them. Relatively homogenous when stained with haematoxylin, their

structure is laminated when viewed in preparation with PAS or reticulin stains. They

are characteristically irregular in size and distribution, both within and between

glomerular loops and located away from the hilus. A rim of mesangial cells can

sometimes be seen between them and adjoining capillary, which is often distended.

Although when present, this lesion is pathognomonic for diabetes, it is not a

universal finding. Its incidence varies considerably from 12-46% in different series,

which included both IDDM and NIDDM cases. Nodules have been found in 55% of

an autopsy series of Pima Indian patients with NIDDM. Nodules are not seen in the

absence of diffuse lesions, and this reflects their appearance only after a long period

of disease.

22

Diffuse glomerular lesions:

Diffuse glomerular lesions comprise an increase in mesangial area and

capillary wall thickening with the mesangial matrix extending to involve the capillary

loops.

As the distribution of diffuse lesion is non-uniform, both among lobules of the

same glomerulus and between different glomeruli, leading to appearance suggestive

of transition to nodule formation. The thickening of capillary walls also tends to be

non-uniform, and this is particularly evident when the histological changes are not

very severe. The diffuse lesions represent earlier stage in the evolution of the disease.

In a large autopsy study, however changes compatible with diabetic

glomerulosclerosis were found in as many as 90% of patients with IDDM with

disease duration of more than 10 years. In patients with NIDDM, the reported

prevalence of these changes ranges between 25-51%.

Exudative Glomerular lesions:

Exudative lesions are highly eosinophilic rounded homogenous structures seen

in capsular space overlying a capillary loop (fibrin cap) or lying on the inside of

Bowman‟s capsule (capsular drop). They are non-specific, containing various proteins

and sometimes lipid materials.

Glomerular hyalinization:

As a consequence of above lesions, increasing number of glomeruli becomes

hyalinised in advanced cases. In some of the ischemic glomeruli, the tufts shrink with

fibrous thickening of the inner surface of Bowman‟s capsule.

23

Arterial lesions:

Diffuse intimal fibrosis in these vessels has been found to be more frequent

and advanced in autopsies on diabetics (Bell 1952,Warren et al 1966)

Arteriolar lesions:

Arteriolar lesions are prominent in diabetics with hyaline material

progressively replacing the entire wall structure. Bell first underlined that both

afferent and efferent arterioles could be affected. Bell also established lesions were

often present in absence of hypertension and the involvement of efferent vessel was

highly specific for diabetes. These arteriolar changes may be the first change

detectable by light microscopy in the diabetic kidney as judged by their recurrence at

2 years in non-diabetic kidneys transplanted into diabetic patients.

Tubular and interstitial changes:

Tubules and interstitium may show a variety of changes that are non-specific

and similar to those seen in other forms of progressive renal disease. Armani-Ebstein

lesions are the result of accumulation of glycogen in tubular cells of the

corticomedullary region in patients with profound glycosuria. More subtle tubular

changes consisting of vacuolization, a decrease in the intercellular spaces normally

present between the macula densa and a significant increase in the contact area

between them and extraglomerular mesangial cell43

.

Immunopathology:

Westberg and Michael confirmed previous observation of linear staining of

glomerular basement membrane for IgG, IgM, albumin and fibrinogen in kidneys of

24

patients with IDDM. The findings were later extended for IgG and albumin not only

in the glomerular basement membrane but also in the Bowman‟s capsule and

especially in the outer aspect of tubular basement membrane were considered specific

for diabetes.

Immunofluoroscence techniques have shown increased mesangial amounts of

type I, IV, V and VI collagens. Immunochemical analysis has also showed reduced

levels of laminin and markedly decreased amounts of heparan sulphates, proteoglycan

where as levels of fibronectin were normal in diabetic mesangium.

Electron microscopy:

Salient features are:

1) Thickening of glomerular basement membrane

2) Maintenance of fine detail of epithelial foot process and abundant epithelial

cytoplasm containing enlarged mitochondria.

3) Accumulation of basement membrane like material within the mesangium.

4) Fibrin deposition in the mesangium and along endothelial aspect of capillary

basement membrane.

Structure-function relationship:

In early phases of IDDM, the increase in luminal volume and filtration surface

area may explain the increase in GFR.

With advancing renal disease, a close association is observed between the

functional changes and mesangial expansion but not thickness of glomerular basement

membrane. Mesangial expansion also correlates inversely with capillary filtration

25

surface area, a variable closely associated with glomerular filtration rate from levels

of hyperfiltration to markedly reduce renal function. Therefore it has been suggested

that expansion of mesangium with attendant reduction in glomerular filtration surface

area that is responsible for the progressive loss of renal function in IDDM.

In IDDM patients, with low levels of microalbuminuria (i.e. AER of 20-

30mcg/min) no consistent glomerular abnormalities have been found. Above these

levels of urinary albumin excretion,the fractional volume of mesangium on average

increases significantly, and minor reduction in creatinine clearance and rise in blood

pressure are observed. Similar findings have been reported in NIDDM patients with

microalbuminuria and proteinuria42

.

INCIDENCE AND PREVALENCE OF MICROALBUMINURIA

The prevalence of microalbuminuria in IDDM patients has been reported in

range from 5-37% in different population based and diabetic clinic based studies. In

NIDDM patients, prevalence of microalbuminuria have been reported in between 8%

and 46% in Europeans and 47% in Pima Indians. Microalbuminuria is found not only

in patients with diabetes but also in patients with impaired glucose tolerance.

Age:

There is no correlation that has been found between AER and age

Sex:

There is male preponderance in IDDM patients. There is no correlation

between microalbuminuria and sex in NIDDM patients.

26

Duration of diabetes:

Prevalence increase with duration of IDDM with distinct variation in the rate

of increase. Incidence of microalbuminuria is very high during the first 3 years after

diagnosis of diabetes, declines at the end of first decade of diabetes and then increase

again to a peak around 12-15 years duration. The prevalence is 8% in patients with

IDDM of only 1-3 years of duration. The prevalence of microalbuminuria levels off

after 10 years (prevalence of 20%) and then assumes its steep climb of around 32%

after 30 years of post pubertal duration of diabetes.

Glycemia:

The level of glycemic control seems to be the strongest factor influencing

transition from normoalbuminuria to microalbuminuria. In recent observational study

of the dose response relationship between intensity of hyperglycemia (measured as

average glycosylated hemoglobin HbA1C level during a 2-4 year period) and the rise

to microalbuminuria was determined in a large cohort of IDDM patients.

A threshold effect of hyperglycemia on the development of microalbuminuria

was found. Below a HbA1c of 10.1%, the risk of persistent microalbuminuria varied

little. In contrast to above ,a threshold HbA1c of 10.1%, the risk of microalbuminuria

rose steeply with increasing levels of HbA1c. In comparison, the risk of

microalbuminuria increases six fold faster between HbA1c levels of 11 and 12% and

that between 8 and 9%. This relationship between HbA1c and levels of

microalbuminuria was independent of the effect of duration of IDDM.

27

Exercise:

Moderately strenuous exercise can provoke an exaggerated rise in AER in

patients with diabetes whose resting values are normal. The severity of exercise

induced albuminuria seems to be related to duration of diabetes and is modulated by

level of glucose control.

Blood pressure and heart rate:

Significant positive associations are found between microalbuminuria and

diastolic blood pressure and resting heart rate.

Other factors influencing the risk of developing microalbuminuria include

cigarette smoking, elevated levels of serum LDL cholesterol44,45

.

STAGES OF DIABETIC NEPHROPATHY

Diabetic nephropathy can conveniently be categorized into different stages,

which differ with respect to renal haemodynamic, systemic blood pressure, urinary

findings and susceptibility to therapeutic interventions.

28

Stages of diabetic nephropathy- typical findings44

Stage Glomerular

filtration

Albuminuri

a

Blood

Pressure

Years After

Diagnosis

Renal hyperfunction Elevated Absent Normal At

Diagnosis

Clinical latency High

Normal Absent Normal

At

Diagnosis

Microalbuminuria

(incipient nephropathy)

Within

normal

Range

20-

200mcg/min

(30-

300mg/day)

Rising within

or above

normal

5-15

Macroalbuminuria or

persisting proteinuria

(clinically manifest

nephropathy)

Decreasing

>200mcg/mi

n

(>300mg/day

)

Increased 10-15

End stage nephropathy Diminished Massive Increased 15-30

PROGNOSTIC SIGNIFICANCE OF MICROALBUMINURIA

Development of persistent proteinuria and overt nephropathy:

Prognostic significance of microalbuminuria for development of persistent

proteinuria and overt nephropathy has been demonstrated by five longitudinal cohort

studies of IDDM patients. These investigations have all suggested the existence of

threshold of AER above, which the risk of progression to clinical nephropathy

increases by about twenty fold. The overall findings of these studies are remarkably

similar, the differences in method of urine collection and length of follow up probably

being responsible for different risk levels of AER.

29

Table: Predictive value of albumin excretion rate (AER) for persistent clinical

proteinuria in patients with IDDM

Group No. of

subjects

Baseline

AER

mcg/min

Follow

up

Years

Type of

Collection

Predictive

value

Viberti et al45

63 >30 14 Over night 88%

Parving et al46

23 >28 6 24 hours 63%

Mathiesen et al47

71 >70 6 24 hours 100%

Mongensen &

Christensen et al48

43 >15 10 Short term 86%

Microalbuminuria is unlikely to be a marker for susceptibility to the

development of clinical nephropathy bur it is more likely to be a sign of early disease.

This interpretation has been recently corroborated by the finding that patients with

persistent microalbuminuria have more severe histological lesions than do patients

with normal AER.

Microalbuminuria and atherosclerotic diseases:

NIDDM is associated with two to three fold increased mortality mainly from

cardiovascular disease. This propensity for vascular disease among NIDDM patients

cannot be explained by co-existent conventional cardiovascular risk factors such as

hypertension or dyslipidemia since the effect of diabetes persists even after

controlling the confounding effects and other risk factors. Nor it can be entirely

attributed to hyperglycemic state, since the available data on the relationship between

glycaemic and cardiovascular complications in NIDDM if not conflicting, is certainly

inconclusive.

30

Mongensen in Denmark and Jarret et al in Britain were the first to explore the

role of microalbuminuria as a marker in patients with NIDDM. Both reported

independently in 1984 that microalbuminuria predicted all cause mortality chiefly

from cardiovascular diseases in NIDDM subjects. Three retrospective studies have

been examined to predict the prognostic significance of microalbuminuria in cohorts

of patients with NIDDM. Over an interval of 10-14 years, clinical proteinuria has

shown a significantly increased risk of death. They have also shown an increased risk

of cardiovascular death in these patients. Another prospective study of 3 years

duration has confirmed the greater incidence of cardiovascular events in patients with

NIDDM.

Lastly the association between microalbuminuria and cardiovascular risk

factor is not confined to diabetic individuals since it has recently been shown also to

extend to the general population45,46,47,48

.

METHODS OF MEASURING MICROALBUMINURIA

Small concentration of albumin in the urine can be measured qualitatively by

several methods.

Radioimmunoassay was the first and most widely used method. Various

methods to determine microalbuminuria are given in the table49.

31

Method sensitivity Time of assay

Single radio immune diffusion(mannil 1965) 1.25mg/ml 1 day

Electroimmuno assay (Laurel1966) 5mg/l 4-6hrs

Immunoturbidimetric assay (Teppor1982) 5mg/l 20-30min

Radio immuno assay (Keen and

Chlouvervakis, 1963)

6.2mcg/ml 1-2days

ELISA (Filding 1983) 250mcg/l 12-18min

Fluorescent immuno assay (Chavers 1984) 500mcg/l 4-6hrs

Latex agglutinates immuno nephelometry

(Vasquez 1984)

750mcg/l

6hrs

Immuno chemical semi quantitative dipstic

(MICRAL)

20-300mg/l 5sec-5min

In our study we have used Micral test for estimation of microalbuminuria.

Micral test (Boehringer Mannheim, Germany) is dipstick method of estimation of

microalbuminuria. Test principle is immunochemical in nature. Sensitivity of Micral

test was 93% ad its specificity was 93% when compared to radioimmunoassay in a

study by Gilbert PE et al50

. Micral test has also been compared with

immunoturbidimetricassay and radioimmunoassay methods. In all studies, Micral test

is comparable in sensitivity and specificity to the other methods of estimation of

microalbuminuria.

32

MICRAL TEST

Fig: 1 Micral II Test Strip

TREATMENT OF INCIPIENT NEPHROPATHY

Microalbuminuria indicates early stage of development of diabetic

nephropathy and also a marker of increased mortality from cardiovascular risk factors.

Microalbuminuria is also associated with other microvascular and macrovascular

complications of diabetes. It is a warning sign for the patient, which should not be

neglected. Progress of microalbuminuria to macroalbuminuria or overt nephropathy

can be reversed or delayed by intervention at this stage.

The strategies for treatment at this stage include1,5,7,8,51

:

1. Optimum glycemic control-diet, intensified insulin treatment, oral

hypoglycemic agents.

2. Blood pressure control and ACE inhibitors.

3. Dietary treatment.

4. Newer treatment modalities which are under study

Aldolase reductase inhibitors

Glycosaminoglycans

33

5. Correction of cardiovascular risk factors and other diabetic complications.

Stopping of cigarette smoking

Correction of dyslipidemia

Optimum glycemic control

In patients with IDDM with microalbuminuria, strict metabolic control by

continuous subcutaneous insulin infusion has been effective in reducing AER. Similar

reduction in AER is seen after multiple injection therapy, provided that similar levels

of blood glucose control are achieved, a finding suggesting that it is the attained

blood glucose control concentration rather than the modality of treatment that

matters52

.

Blood pressure control:

Blood pressure is generally above normal in microalbuminric stage of

nephropathy. In earlier studies multiple drug therapy of hypertension was used,

including β blockers, diuretics, vasodilators and calcium channel blockers. During last

few years, a angiotensin converting enzyme (ACE) inhibitors have shown great

promise not only in patients with established diabetic nephropathy but also in non

hypertensive patients with microalbuminuria. ACE inhibitors may be specifically

protective for renal function because of their ability to reduce efferent arteriolar

vasoconstriction and thus reduce the intra glomerular pressures. This “specific” effect

has been claimed to be separate from any effect on systemic blood pressure53

.

Antihypertensive treatment especially with ACE inhibitors therefore delays

the progression of established diabetic nephropathy and if started at the stage of

34

microalbuminuria may even prevent or at least retard the onset of clinically overt

renal disease54,55,56,57

.

Dietary and Behavioural modification:

Reductions of dietary protein by approximately 50% has been shown to reduce

the fractional clearance of albumin in patients with microalbuminuria, and to lower

GFR in patents with hyperfiltration, independently of changes in glucose control and

pressure. Diets restricted 0.5-0.6gm of protein /kg body weight per day is ideal and

does not have long-term detrimental effect on nutritional status of an individual51

.

Other treatment modalities:

Aldolase reductase inhibitors, which have been studied in a few studies, show

reduction in GFR and decrease in AER in IDDM patients who had either a normal

AER or microalbuminuria. But study in subjects with NIDDM with

microalbuminuria, it failed to show any effects on renal function. Further studies are

needed to confirm the finding.

Recent observation suggests that oral administration of sulodexide (a naturally

occurring glycosaminoglycan) extracted from pig intestinal mucosa, containing a fast

moving heparan like fraction (80%) and a dermatan sulphate fraction (20%) along

with ACE inhibitors seems to retard progression from incipient to overt

nephropathy in NIDDM patients. Mechanism is possibly by restoring glomerular

basement membrane charge and size selectivity to albumin molecules as well as

reducing glomerular capillary pressure. Further studies are needed to confirm this

finding58

.

35

Correlation of cardiovascular risk factors and other complications:

A detailed cardiovascular examination is necessary early in the course of

diabetic nephropathy. Hypertension must be treated energetically. Left ventricular

hypertrophy and function should be assessed echocardiographically at the stage of

microalbuminuria and thereafter every 6–12 months. Effective antihypertensive

therapy can reverse left ventricular hypertrophy. In addition, cardiac assessment

should include electrocardiography, stress testing, coronary angiography and Holter

monitoring is indicated whenever needed. Ischemic heart disease should be treated

aggressively. Peripheral vascular disease must be assessed and treated as necessary.

Doppler flow studies and arteriography may be useful to assess the severity of the

disease59,60

.

Microalbuminuria is frequently associated with hyperlipidemia and lipid

profile is an essential investigation and dyslipidemia should be treated61

.

Testing vibration perception threshold and thermal discrimination may

identify the risk of neuropathic ulceration. The test should be repeated regularly as

sensation may become impaired later, during the course of nephropathy. Autonomic

dysfunction is very common in nephropathic patients. The important manifestations

are postural hypotension and incomplete bladder emptying which predisposes to

urinary tract infection62

.

Microalbuminuria is frequently associated with retinopathy. Retinopathy

almost always accompanies diabetic nephropathy. Early and regular ophthalmic

review and prompt treatment is necessary to prevent blindness63,64

.

36

Diabetic retinopathy

Diabetic retinopathy is a well-characterized, sight-threatening, chronic

microvascular complication that eventually afflicts virtually all patients with diabetes

mellitus65

.

Diabetic retinopathy is characterized by gradually progressive alterations in

the retinal microvasculature,leading to areas of retinal non-perfusion,increased

vasopermeability, and pathologic intraocular proliferation of retinal vessels.

Epidemiology of diabetic retinopathy

All patients with type 1 diabetes and more than 60% of patients with type 2

diabetes develop some degree of retinopathy after 20 years. In patients with type 2

diabetes, approximately 20% have retinopathy at the time of diagnosis of diabetes and

most have some degree of retinopathy over subsequent decades. About 4% of patients

younger than 30 years of age at diagnosis and nearly 2% of patients older than 30

years of age at diagnosis were legally blind.Approximately 25% of patients with type

1 diabetes have retinopathy after 5 years, with this figure increasing to 60% and 80%

after 10 and 15 years, respectively66

.

Pathophysiology

The earliest histologic effects of diabetes mellitus in the eye include loss of

retinal vascular pericytes (supporting cells for retinal endothelial cells), thickening of

vascular endothelium basement membrane, and alterations in retinal blood flow. With

increasing loss of retinal pericytes, the retinal vessel wall develops outpouchings

(microaneurysms) and becomes fragile. With time, increasing sclerosis and

endothelial cell loss lead to narrowing of the retinal vessels, which decreases vascular

37

perfusion and may ultimately lead to obliteration of the capillaries and small vessels,

the resulting retinal ischemia is a potent inducer of angiogenic growth factors. These

factors promote the development of new vessel growth and retinal vascular

permeability. Proliferating new vessels in diabetic retinopathy have a tendency to

bleed, which results in preretinal and vitreous hemorrhages and later macular edema.

Risk factors

1. Duration of diabetes is closely associated with onset and severity of diabetic

Retinopathy.

2. Lack of glycemic control.

3. Renal disease, as manifested by microalbuminuria and Protienuria67

.

4. Hypertension68

.

5. Elevated serum lipid levels are associated with extravasated lipid in the retina

(hard exudates) and visual loss69

.

Clinical findings

Clinical findings associated with early and progressing diabetic retinopathy

include haemorrhages or microaneurysms (H/Ma), cotton-wool spots (CWSs), hard

exudates, intraretinal microvascular abnormalities (IRMAs), and venous calibre

abnormalities (VCABs), such as venous loops, venous tortuosity, and venous beading.

The intraretinal hemorrhages can be “flame-shaped” or “dot/blot” like in

appearance. IRMAs are either new vessel growth within the retinal tissue itself or

shunt vessels through areas of poor vascular perfusion. It is common for IRMAs to be

adjacent to CWSs, which are caused by micro infarcts in the nerve fiber layer.

VCABs are a sign of severe retinal hypoxia. In some cases of extensive vascular loss,

38

however, the retina may actually appear free of non-proliferative lesions. Such areas

are termed “featureless retina” and are a sign of severe retinal hypoxia.

Symptoms of diabetic retinopathy

1. Patient complaints of blurred vision, usually central vision and

metamorphosia as a result of maculopathy with foveal involvement.

2. Black spots, floaters or sudden visual loss may be experienced by patients

with vitreous haemorrhage, depending on quantum of bleed.

Kanski classification of diabetic retinopathy70

1. Background diabetic retinopathy

a. Haemorrhages (Dot and Blot)

b. Microaneurysms (Located in inner nuclear layer)

c. Hard exudates ( Located in between inner plexiform and inner nuclear

layer)

d. Retinal edema (Located between outer plexiform and inner nuclear

layer)

2. Pre proliferative

a. Vascular changes(beading , looping)

b. Dark blot haemorrhages

c. Cottonwool spots

d. Intraretinal microvascular abnormalities

e. Shunt vessels

3. Proliferative

a. Neo vascularisation

39

b. Fibrous proliferation

c. Vitreous detachment and haemorrhages

4. Maculopathy

a. Focal

b. Diffuse

c. Ischaemic

Classification of diabetic maculopathy

Intraretinal

Macular edema

Macular hard exudates

Macular ischaemia

Pre retinal and vitreo- retinal

Thick and post hyaloids

Thick and pre retinal membrane

Tractional detachment of macula

Macular ectodia

Monitoring and treatment of diabetic retinopathy

Appropriate clinical management of diabetic retinopathy has been defined by

results of four major, randomized, multicentered clinical trials the Diabetic

Retinopathy Study (DRS), the ETDRS, the Diabetic Retinopathy Vitrectomy Study

(DRVS) and the DCCT.

40

Figure 2: Background diabetic retinopathy showing scattered exudates and

haemorrhages near fovea. Normal vision (6/6). Laser photocoagulation indicated

Figure 3: Background diabetic retinopathy with severe maculopathy. Hard

exudates at macula. Vision 6/60. Central vision permanently lost.

41

Figure 4: Proliferative diabetic retinopathy with disc neovascularization. Vision

6/6 Laser photocoagulation required

Figure 5: Advanced proliferative diabetic retinopathy with traction retinal

detachment resulting from preretinal scar tissue. Vision: Hand movement. Too

late for treatment

42

Figure 6: Proliferative diabetic retinopathy with abnormal vessels at optic disc

and retina with early vitreous haemorrhage. Eye in danger of being blind and

requires laser photocoagulation

43

METHODOLOGY

One hundred patients of diabetes (NIDDM) who are eligible and attending

outpatient department and admitted in medical wards of Navodaya medical college

hospital and research centre were chosen for the study based on random selection.

Patients were considered to be diabetic based on WHO criteria for diagnosis of

diabetes mellitus which is: -

1. Symptoms of diabetes mellitus plus a random glucose concentration >200

(11.1mmol/l). The classical symptoms of diabetes mellitus include polyuria,

polydipsia ,polyphagia and unexplained weight loss

OR

2. Fasting blood glucose >126 mg/dl (7.0mmol/l). Fasting is defined as no

caloric intake for at least 8 hours.

OR

3. 2 hour plasma glucose > 200mg/dl (11.1 mmol/l) during an oral glucose

tolerance test. Among diabetics, the above criteria were considered to include

the patients for the study.

Selection criteria

Inclusion criteria

1. The American Diabetic Association criteria for the diagnosis of Type 2

diabetes mellitus are FPG > 126 mg/dl with symptoms of diabetes and PPPG >

200 mg/dl on two different occasions.

44

OR

Two hour plasma glucose > 200 mg/dl during an oral glucose tolerance test.

2. Diagnosis of DM after 30 years of age.

Exclusion criteria

1) Patients with macroalbuminuria

2) Patients with congestive cardiac failure, urinary tract infection.

3) Ketonuria

4) Pregnant patients

5) Patients with overt diabetic nephropathy

6) Serum creatinine > 1.5 mg/dl

7) Diabetes with any acute stressors like infections, MI, etc.,

8) Diabetes with prior hypertension or within 5years of detection of type 2

diabetes mellitus.

The selected patients were studied in detail with history and physical

examination

History

Patient‟s characteristics like age, sex, age of onset and duration of diabetes.

All details regarding the presenting complaints were noted.

Total duration of diabetes, the drugs the patient was taking and the dosages

were noted. The regularity of the treatment taken by the patients was also

noted. The family history regarding diabetes was taken.

45

Personal history regarding smoking, alcohol consumption, bowel and bladder

habits and drug intake were noted.

A complete clinical examination was carried out in each patient. Height and

weight were measured in all cases and body mass index (BMI) was calculated by

weight in kg / height in m2.

The following investigations were done in all the patients.

Microalbuminuria was estimated by Micral test in all the cases.

Fasting Blood sugar and Postprandial blood sugar

Glycosylated hemoglobin

Blood urea and serum creatinine

Fasting lipid profile

Urine routine and culture

Electrocardiogram

Ultrasonography of the abdomen, echocardiogram and chest x-ray were done in

selected cases only.

Estimation of Microalbuminuria by Micral test:

All patients having overt macroalbuminuria detected by albustik were

excluded from study. Micral test, a immunological rapid dip stick semi qualitative

technique for detection of microalbuminuria, was used for estimation of

microalbuminuria.

46

Micral test components:

1 test strip contains monoclonal antibodies against human albumin

(immunoglobulin G) labelled with colloid gold 2.2mg, fixed albumin 7.7 mg

Test principle

There is a serial arrangement of several reagent pads, which are in fluid

communication by a reaction controlling chromatographic process. This step

combines one step handling with a complex chemistry. The single reaction steps are

as follows:

Urine of the sample is transported through the wick fleece to the buffer fleece,

where acidic urine is adjusted to proper pH

Upon entering the conjugate fleece, the antigen – antibody reaction takes

place. Albumin of the sample is specifically bound to a soluble conjugate of

antibodies and marker enzyme resulting in an antigen – conjugate complex

The excess antibodies are bound to immobilized albumin on the capture

matrix and removed from the sample in this way.

Only the complex of conjugate with sample-albumin reaches the substrate pad.

Here the colour reaction takes place, the marker enzyme B-Galactosidase

cleaves off the purple dye chlorophenol red from the Yellow substrate

(chlorophenol red galactoside) in a kinetic reaction. The intensity of the colour

produced is proportional to the albumin concentration in the urine.

47

Method of data collection:

All patients were afebrile during the course of collection of urine and were

kept at rest during the collection of urine. Urine of the patient was first tested for

albumin by albustik method. Patients who were negative for albumin by the albustik

method were only included in this study.

First morning mid stream urine sample that was collected in a sterile container

was used for determining microalbuminuria. Test strip was immersed in urine such

that fluid level was between the two black bars provided on the strips. Strip was

withdrawn after 5 seconds. Strip was placed horizontally across the urine vessel and

colour change in the test zone was compared with colour scale after one minute.

Sensitivity of the kit is 0.4ng/ml and measuring range is 0.8 to 10ng/ml.

Occular examination:

Detail ophthalmic examination including visual acuity and slit lamp

examination was done for all patients. Fundus examination under mydriasis with

tropicamide + phenylephrine eye drops was done for all patients for the presence of

retinopathy.

Fundus photography and fluorescein angiography was performed in indicated

patients.

48

RESULTS

Table 1 shows the age and sex distribution of patients

Table1: Age and sex distribution

Age in yrs

Male Female Total

No % No % No

40-50 23 40.26 16 37.21 39

51-60 15 26.32 16 37.21 31

61-70 14 24.56 6 13.95 20

>70 5 8.77 5 11.63 10

Total 57 100 43 100 100

Mean age±SD 54.82±12.38 54.82±11.08 54.87±11.59

P value Mean age between male and female=0.965

39 patients were in the age group between 40-50 years, among whom 23 were

male,16 were female patients. 31 patients were in the age group between 51 and 60

years, among whom 15 were male and 16 were female patients. 20 patients were in

the age group between 61 and 70 years, among whom 14 were male and 6 were

female patients. 10 patients were in the age group greater than 70 years, among whom

5 were male and 5 were female patients. The mean age of male patients in the study

was 54.82 ± 12.38 years and that of the female patients was 54.82 ±11.08 years. The

mean age of detection of diabetes mellitus among the male patients was 48.84± 10.11

years and in the patients was 48.75 ± 8.68 years.

The mean age between male and female is not statistically significant with

p=0.964.

49

Graph – 1: Graph showing age and sex distribution of patients

39 patients were in the age group between 40-50 years, among whom 23 were

male(40.26%),16 were female patients(37.21%). 31 patients were in the age group

between 51 and 60 years, among whom 15 were male(26.32%) and 16 were female

patients(37.21%). 20 patients were in the age group between 61 and 70 years, among

whom 14 were male(24.56%) and 6 were female patients(13.95%). 10 patients were

in the age group greater than 70 years, among whom 5 were male(8.77%) and 5 were

female patients(11.63%).

50

Table 2: Number of patients with microalbuminuria and retinopathy

Microalbuminuria Retinopathy

- + - +

62 38 56 44

There were 62 patients negative for microalbuminuria.38 patients were positive

for microalbuminuria. There were 56 patients negative for retinopathy, 44 patients

were positive for retinopathy.

Graph – 2 : Pie diagram showing percentage of patients with microalbuminuria

and retinopathy

51

Table - 3:Association of age with microalbuminuria and retinopathy

Age in

years

Number of

patients


- + - +

40-50 39 31 8 29 10

51-60 31 20 11 19 12

61-70 20 10 10 8 12

>70 10 1 9 - 10

Among 39 patients in the age group between 40-50 years,8 patients had

microalbuminuria and 10 patients had retinopathy.31 patients were in age group

between 51-60 years, among them 11 patients had microalbuminuria and 12 patients

had retinopathy.20 patients were in the age group between 61 -70 years, among whom

10 patients had microalbuminuria and 12 patients had retinopathy.10 patients were in

the age group of above 70 years , among whom 9 patients had microalbuminuria and

all 10 were positive for retinopathy.

Incidence of microalbuminuria is more likely for the age group above 50 years of age

as compared to the age group <50 years of age with p=0.053.

Incidence of retinopathy is more likely for the age group above 50 years of age as

compared to the age group <50 years of age with p=0.001.

52

Graph – 3 : Association of age with microalbuminuria and retinopathy

0

20

40

60

80

100

120

_ + _ +

Pe

rce

nta

ge o

f p

atie

nts

microalbuminuria retinopathy

41-50

51-60

61-70

>70

Among 39 patients in the age group between 40-50 years,8 patients had

microalbuminuria (20.5%)and 10 patients had retinopathy(25.64%).31 patients were

in age group between 51-60 years, among them 11 patients had

microalbuminuria(35.48%) and 12 patients had retinopathy(38.70%).20 patients were

in the age group between 61 -70 years, among whom 10 patients had

microalbuminuria (50%)and 12 patients had retinopathy(60%).10 patients were in the

in the age group of above 70 years , among whom 9 patients had

microalbuminuria(90%) and all 10 patients(100%) had retinopathy

53

Table 4 :Association of Duration of diabetes mellitus with microalbuminuria

and retinopathy

Duration of

diabetes

mellitus

Number of

patients


- + - +

< 5 39 31 8 29 10

6-10 31 20 11 19 12

11-15 20 10 10 8 12

16-20 10 1 9 - 10

Among 39 patients with duration of diabetes <5years ,8 patients had

microalbuminuria and 10 patients had retinopathy.Among 31 patients with duration of

diabetes between 6-10 years, 11 patients had microalbuminuria and 12 patients had

retinopathy.Among 20 patients with duration of diabetes between 11-15years, 10

patients had microalbuminuria and 12 patients had retinopathy.Among 10 patients

with duration of diabetes between 16-20 years , 9 patients had microalbuminuria and

all 10 were positive for retinopathy.

Incidence of microalbuminuria is more likely with duration of diabetes above 6 years

as compared to the duration of diabetes below 6 years.age.

Incidence of retinopathy is more likely with duration of diabetes above 6 years as

compared to the duration of diabetes below 6 years.age.

Graph – 4 : Association of Duration of diabetes mellitus with microalbuminuria and

retinopathy

Among 39 patients with duration of diabetes <5years,8 patients had microalbuminuria

(20.5%)and 10 patients had retinopathy(25.64%).31 patients with duration of diabetes

between 6-10 years, among them 11 patients had microalbuminuria(35.48%) and 12 patients

had retinopathy(38.70%).20 patients with duration of diabetes between 11-15 years, among

whom 10 patients had microalbuminuria (50%)and 12 patients had retinopathy(60%).10

patients with duration of diabetes above 16 years, among whom 9 patients had

microalbuminuria(90%) and all 10 patients(100%) had retinopathy

54

0

20

40

60

80

100

120

_ + _ +

Pe

rce

nta

ge o

f p

atie

nts

microalbuminuria retinopathy

≤5

06-1 0

1 1-15

16-20

55

Table 5:Association of HbA1c with microalbuminuria and retinopathy

HbA1c No of

patients


- + - +

<6.5 22 16 6 18 4

6.5-7 22 17 5 18 4

7-7.5 16 12 4 8 8

>7.5 40 17 23 12 28

22 patients had HbA1c values less than 6.5% among them 6 were positive for

microalbuminuria and 4 were positive for Retinopathy. 22 patients had HbA1c

between 6.5% and 7%among them 5 were positive for microalbuminuria and 4 were

positive for retinopathy.

16 patients had HbA1c levels between 7.0% and 7.5% among them 4 were

positive for microalbuminuria and 8 were positive for retinopathy.40 patients had

HbA1c values more than 7.5%,among them 23 were positive for microalbuminuria

and 28 were positive for retinopathy.

The association of microalbuminuria and retinopathy with HbA1c is significant with

P values 0.016 and 0.001 respectively

56

Graph - 5:Association of HbA1c with microalbuminuria and retinopathy

22 patients had HbA1c values less than 6.5% among them 6 were positive for

microalbuminuria(27.27%) and 4 were positive for Retinopathy(18.18%). 22 patients

had HbA1c between 6.5% and 7%among them 5 were positive for microalbuminuria

(22.72%)and 4 were positive for retinopathy(18.18%).

16 patients had HbA1c levels between 7.0% and 7.5% among them 4 were

positive for microalbuminuria(25%) and 8 were positive for retinopathy(50%).40

patients had HbA1c values more than 7.5%,among them 23 were positive for

microalbuminuria(57.5%) and 28 were positive for retinopathy(70%).

0

10

20

30

40

50

60

70

80

90

_ + _ +

per

cen

tage

of p

atie

nts


HbA1c

<6.5

6.5-7

7-7.5

>7.5

57

Table:6 Association of BMI with microalbuminuria and retinopathy

Body mass

index(kg/m2)

Number of

patients


- + - +

<25 78 53 25 49 29

>25 22 9 13 7 15

78 patients had a BMI less than 25kg/m2, out of them 25 patients were positive for

microalbuminuria and 29 patients were positive for retinopathy.

22 patients had a BMI above 25 kg/m2, out of them 13were positive for

microalbuminuria and 15 were positive for retinopathy

The association of microalbuminuria and retinopathy with BMI is significant with P

values 0.02 and 0.01 respectively

58

Graph -6 :Showing Association of BMI with microalbuminuria and retinopathy:

78 patients had a BMI less than 25kg/m2, out of them 25 patients were positive for

microalbuminuria(32.05%) and 29 patients were positive for retinopathy(37.17%).

22 patients had a BMI above 25 kg/m2, out of them 13were positive for

microalbuminuria (59.09%)and 15 were positive for retinopathy(68.18%).

0

10

20

30

40

50

60

70

80

microalbuminuria - microalbuminuria+ retinopathy- retinopathy+

p

e

r

c

e

n

t

a

g

e

s

<25

>25

59

Table.7: Association between microalbuminuria and retinopathy

Microalbuminuria + Microalbuminuria -

Retinopathy + 31 13

Retinopathy - 7 49

31 subjects had evidence of both microalbuminuria and retinopathy,13 patients had

retinopathy without microalbuminuria,7 patients had only microalbuminuria without

retinopathy

The association between microalbuminuria and diabetic retinopathy is significant with

P value of <0.001.

Graph-7: Association between microalbuminuria and retinopathy

31 subjects had evidence of both microalbuminuria and retinopathy,13 patients had

retinopathy without microalbuminuria,7 patients had only microalbuminuria without

retinopathy

0

10

20

30

40

50

Microalbuminuria + Microalbuminuria -

Association between microalbuminuria and retinopathy Retinopathy + Retinopathy -

60

DISCUSSION

The prevalence of microalbuminuria:

In the present study the prevalence of microalbuminuria is 38%.

Similar results were shown in the following studies,

In a study conducted by Unnikrishnan RI et al in Chennai, South India, the prevalence

of microalbuminuria is 27%.71

Gupta et al reported a prevalence of 26.6% in 65 type 2 north Indian patients.72

John et al reported a prevalence of 19.7% from a tertiary hospital in Vellore, south

India.73

S.No Year Investigator Location Prevalence

1 1991 John et al

Vellore 19.7%

2 1991 Gupta et al North India 26.6%

3 2007 Unnikrishnan et al Chennai 27%

4 2010 Present study Raichur 38%

Various epidemiological studies have reported marked variation in the

prevalence of microalbuminuria ranging from 7%to 42 %.The prevalence in our study

is within this range.

The prevalence of retinopathy:

The prevalence of retinopathy in our study is 44%.Various studies on the

prevalence of diabetic retinopathy in India are as follows:

61

S. No Year Investigator Location Prevalence

1 1996 Rema et al 74

Chennai 34.1%

2 1999 Dandona et al 75

Hyderabad 22.6%

3 1999 Ramachandran et al76

Chennai 23.7%

4 2002 Narendran et al77

Palakkad 26.8%

Age distribution of microalbuminuria:

In our study among 39 patients in the age group between 40-50 years, 8(20.5)

patients had microalbuminuria.31 patients were in age group between 51-60 years,

among them 11(35.4%) patients had microalbuminuria.20 patients were in the age

group between 61 -70 years, among whom 10(50%) patients had microalbuminuria

and 10 patients were in the in the age group of above 70 years , among whom 9(90%)

patients had microalbuminuria.

The prevalence of microalbuminuia increases with that of the age of the

patients. The increasing age was reported as one of the risk factors for the

development of microalbuminuria in various studies conducted by John et al, Vijay et

al, Klein R et al in the Wisconsin study and Nelson et al in Pima Indians78,79,80

.

Deterioration in the beta cell function, which occurs parripassu with increasing

age, is likely to contribute to worsening glycaemic control.

Association of duration of diabetes with microalbuminuria and retinopathy:

In our study, it has been observed that longer the duration of diabetes the

higher the prevalence of microalbuminuria and retinopathy. In patients with diabetes

62

for less than 5 year duration , 20.5% had microalbuminuria and 25.6% had

retinopathy. In patients with diabetes for more than 15 years, 90% had

microalbuminuria and 100% had retinopathy.Similar findings were observed in a

study conducted by Varghese et al in which 30.4% patients with diabetes for less than

5 year duration had microalbuminuria and 96% patients with diabetes for more than

than 15 year duration had microalbuminuria.81

Chronic hyperglycemia is thought to be the primary cause of diabetic

retinopathy. Evidence in support for this hypothesis has come from the Diabetes

Control and Complications Trial (DCCT), which found that intensive insulin therapy,

achieving a mean hemoglobin A1C (A1C) of 7.9 percent, reduced the incidence of

new cases of retinopathy by as much as 76 percent compared with conventional

therapy. The reduction was directly related to the degree of glycemic control as

estimated from haemoglobin A1C values (mean A1C with conventional therapy was

approximately 9.9 percent); progressive retinopathy was uncommon in patients with

A1C values below seven percent.82

Association between microalbuminuria and diabetic retinopathy :

Our study showed that in addition to HbA1c, BMI and the duration of illness,

microalbuminuria is a contributing factor in the degree of retinopathy (p < 0.001) and

the association between microalbuminuria and diabetic retinopathy observed in the

present study could be explained by the view that microalbuminuria might represent a

state of generalized vascular dysfunction [Deckert et al]. Enzymes involved in the

metabolism of anionic components of the extracellular matrix (e.g. heparan sulphate

proteoglycan) vulnerable to hyperglycemia, seem to constitute the primary cause of

albuminuria and its associated complications.83

63

Estacio RO et al evaluated the correlation between albuminuria and

retinopathy in 815 patients with type 2 diabetes (144 Hispanics and 671 whites]. The

presence of albuminuria, defined as urinary albumin excretion >200 mcg/min, was a

significant predictor for retinopathy (as detected via stereoscopic retinal fundus

photographs) among the Hispanic (odds ratio 11.1). 84

Similar association observed in

studies conducted by Lunetta et al, Manaviat et al.85,86

Association of HbA1c with microalbuminuria and retinopathy:

In our study, only 11 out of 44 patients who had a normal HbA1c (< 7.0%)

manifested microalbuminuria, whereas with HbA1c values more than 7, 27 out of 56

(nearly 50%) had microalbuminuria. Also nearly 64% (36/56) patients with Hb A1c

>7 had evidence of retinopathy. It is seen from the above result that even small

increments of HbA1cmore than 7.0% result in almost doubling of the incidence of

microalbuminuria and retinopathy.

Poor glycemic control as a risk factor for microalbuminuria was reported in

various studies including UKPDS, DCCT trial and a study conducted by

Unnikrishnan et al.71,82,87

The mechanism by which lack of glycemic control predisposes to vascular

disease is incompletely understood. Two proposed contributing factors are advanced

glycosylation end products and sorbitol; protein kinase C and other factors also may

contribute.

In addition to systemic factors, organ-specific factors also appear to be

important. In the kidney, for example, stimulation of mesangial matrix production by

64

hyperglycemia, and activation of protein kinase Cmay contribute to the glomerular

injury.88,89

.

Association of BMI with microalbuminuria and retinopathy:

This study has also brought out a significant association of microalbuminuria

with body mass index of more than 25kg/m2. Of the 22 patients with BMI of more

than 25, 13had microalbuminuria (52%). 15 out of 22 (68%) patients with BMI > 25

had retinopathy . Similar findings have been brought forth by Patel Kl et al 90

, Taneja

V et al 91

, Jadhav UM et al 92

.

The possible explanation for this could be

Increasing body mass index is a reflection of insulin resistance which in turn leads

to endothelial dysfunction and microalbuminuria.

Poor glycemic control which in turn is an outcome of insulin resistance is also

held responsible.

Study limitations

It was done in a small group it may not represent the entire population.

65

CONCLUSION

The present study has shown that there is significant association between the

presence of microalbuminuria and retinopathy.

It has also shown that there is increase in the prevalence of microalbuminuria

and retinopathy with increasing age, HbA1c >7%,BMI>25 Kg/m2.

66

SUMMARY

Out of 100 cases,57 cases were male and 43 cases were female.

Microalbuminuria was found in 38 % of patients and diabetic retinopathy was

present in 44 % of patients..

Microalbuminuria and retinopathy show a direct relationship with increasing

age of patients, HbA1c levels and BMI.

HbA1c value above 7 % is associated with increasing incidence of

microalbuminuria and retinopathy.

Patients with a BMI of more than 25 kg/m2

have significant increase in the

incidence of microalbuminuria and retinopathy.

Microalbuminuria and diabetic retinopathy were present together in 31% of

patients showing that there is a significant association between them.

67

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47. Diabetic Nephropathy. Position statement: American diabetic association.

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81

ANNEXURE - I

INFORMED CONSENT

ASSOCIATION BETWEEN MICROALBUMINURIA AND DIABETIC

RETINOPATHY IN TYPE 2 DIABETES PATIENTS – A ONE YEAR CROSS-

SECTIONAL STUDY

Consent: I, ................................. willingly and in my full give sense, give my consent

to take part in this study. The risk and benefits of which have been explained to me in

my own vernacular language.

Witness

Date Signature of the

patient/ relative

82

ANNEXURE – II

PROFORMA

Name: IP/OP No:

Age: Date of admission:

Sex: Date of discharge:

Religion: Marital status:

Address:

Occupation:

Education: Signature of the Patient

Income group: High/middle/poor

CHIEF COMPLAINTS:

Polyuria Polydipsia Blurring of vision

Weight loss Tiredness Polyphagia

Pruritus Aches and pains Skin infections

HISTORY OF PRESENTING ILLNESS

1) Cardiovascular symptoms:

Angina myocardial infarction exertional dyspnea orthopnea

PND palpitations syncope sweating

83

Swelling of feet

2) Peripheral vascular symptoms:

Intermittent claudication Gangrene Impotence thrombophlebitis

3) Cerebrovascular symptoms:

Giddiness headache vomiting transient ischemic attacks Stroke

4) Visual symptoms:

Blurring of vision Progressive loss of vision sudden loss of vision

5) Renal symptoms:

a) Symptoms of urinary tract infection:

Dysuria Fever with chills Flank pain

b) Symptoms of nephropathy:

Swelling of feet Puffiness of face Distension of abdomen

Increasing sensitivity of insulin

6) Symptoms of neurological complications:

a) Symptoms of polyneuropathy:

Tingling Numbness Paraesthesia Nocturnal pain

Sensory ataxia Trophic ulcers Charcot joints

b) Symptoms of mononeuropathy:

Wrist drop Foot drop

c) Cranial nerves:

Diplopia Squint Deviation of the angle of mouth

Inability to close the eyes

d) Symptoms of radiculopathy:

e) Symptoms of autonomic neuropathy

Dysphagia vomiting Nocturnal diarrhea Syncope

84

f) Symptoms of amyotrophy:

Paraesthesia of the anterior thigh Thinning of proximal muscle

Weakness of proximal muscles

7) Symptoms of Diabetic skin complications:

Recurrent skin infections Abscesses Carbuncles

8) Gastro intestinal symptoms:

Dysphagia Abdominal pain Nausea/Vomiting Diarrhea

Weight loss Loss of appetite

9) Genito urinary symptoms:

Polyuria/Nocturia Balanoprosthitis Impotence Dysuria

Discharge per urethra Discharge per vagina

DIABETES HISTORY

Diabetes diagnosed in the year:

Duration of Diabetes:

Diabetes onset at the age of:

Mode of onset of Diabetes:

Family history of Diabetes:

Drugs used for Diabetes: OHA / INSULIN/ INSULIN + OHA

PAST HISTORY

History of hypertension

History of Angina / Myocardial infarction

History of Transient ischemic attack / stroke

PERSONAL HISTORY

HABITS YES NO STOPPED

Smoking

85

Sweets

Alcohol

Tobacco

Appetite /bowel /bladder

FAMILY HISTORY

MEMBERS PRESENT ABSENT

Father

Mother

Sisters

Sons

Daughters

Spouse

Others: History of Hypertension, Ischemic heart disease , Cerebro vascular

accidents

Obesity and Sudden death

OBSTETRIC AND GYNAECOLOGICAL HISTORY

Menarche/ Menopause

Gravida/Para

Abortion

History of stillbirths

History of delivery of large babies

GENERAL PHYSICAL EXAMINATION

1) BUILT: Well / moderately /poor/ emaciated

2) WEIGHT:

HEIGHT:

86

BODY MASS INDEX:

3) PULSE:

4) BLOOD PRESSURE:

5) PALLOR:

EDEMA, PEDAL:

CLUBBING/KOILONYCHIA

CYANOSIS:

ICTERUS:

LYMPHADENOPATHY:

6) JUGULAR VENOUS PULSE

7) PERIPHERAL PULSE

8) SKIN CHANGES:

Diabetic dermopathy

Acanthosis nigricans

Scleroderma

Xanthoma

Fungal infection

Skin tags

9) EYES:

Normal/ Xanthoma / Arcus / Cataract

10) FUNDUS

SYSTEMIC EXAMINATION

1) CARDIOVASCULAR SYSTEM

INSPECTION

PALPATION

87

PERCUSSION

AUSCULTATION

OTHER FINDINGs

2) RESPIRATORY SYSTEM

INSPECTION

PALPATION

PERCUSSION

AUSCULTATION

OTHER FINDINGS

3) ABDOMEN

INSPECTION

PALPATION

PERCUSSION

AUSCULTATION

OTHER FINDINGS

4) CENTRAL NERVOUS SYSTEM

HIGHER MENTAL FUNCTIONS

CRANIAL NERVES

MOTOR SYSTEM

NUTRITION

TONE

POWER/REFLEXES

CO-ORDINATION

INVOLUNTARY MOVEMENTS

88

SENSORY SYSTEM

TOUCH / PAIN / TEMPERATURE/VIBRATION /

POSITION SENSE /

DIABETIC RETINOPATHY: PRESENT/ ABSENT

INVESTIGATIONS

1) URINE:

Macroalbuminuria

Microalbuminuria

Sugar

Microscopy

Ketone bodies

2) BLOOD:

Fasting blood glucose

Post prandial blood glucose

Glycosylated hemoglobin:

Blood urea:

Serum creatinine:

Lipid profile

1. Total cholesterol

2. Triglycerides

3. HDL

4. LDL

3) ELECTROCARDIOGRAM

4) ECHOCARDIOGRAM (selected cases only)

5) ABDOMINAL ULTRASONOGRAPHY

89

DIAGNOSIS

TREATMENT GIVEN:

1) Diet

2) Diet + OHA

3) Diet + OHA + insulin

4) Diet + insulin

Signature of Candidate

Signature of Co-guide Signature of Guide

90

ANNEXURE - III

Master chart

HbA1cFBS PPBS B/U

Sr.Cre RP Htn BMIS.No

Name IP/OP No

Age

Sex Micro Duration of DMAlb

1 Hanumanthaya 14494 65 M + 20.1 270 355 36 1 + - 10.1 12years2 Venkanna 10957 74 M + 16.4 221 291 22 0.9 + - 9.8 16years3 Shankarappa 12888 60 M - 17.8 173 212 38 0.8 + - 7.8 13years4 Chandrasekhar 4326 56 M - 22.5 164 250 33 1.2 - - 6.8 7years5 Padmavathi 12782 50 F - 18.2 160 280 24 1.2 - - 7.6 4years6 Baburao 13618 64 M + 26.3 201 294 26 1.4 + - 9.3 12years7 Kamalamma 23320 54 F + 30.1 230 289 27 1 + - 10.2 8years8 Gouramma 29070 43 F - 18.4 214 261 25 0.8 - - 6.9 5years9 Balappa 13729 47 M - 22.5 210 280 22 0.8 - - 6.8 5years10 Veerabhadrappa 3952 40 M - 26.3 170 250 34 1.1 + - 7.2 5years11 Hussain bee 137690 70 F + 22.6 190 286 26 0.9 + - 8.1 12years12 Veeranagouda 5710 48 M - 29.7 185 270 38 1.3 + - 8.4 5years13 Nagappa 12343 60 M + 21.9 170 250 34 1.2 + - 7.8 8years14 Abdul mohd 27092 65 M + 24.3 210 290 29 1.1 + - 8.4 11years15 Sharanappa 14466 60 M - 23 165 260 30 1.2 - - 7.6 6years16 Shashaidevi 29718 41 F - 28.3 192 280 24 1.1 - - 9.2 5years17 Thayanna 5353 45 M - 21.8 160 247 29 1.2 - - 6.8 5years18 Laxmanna 27221 45 M - 26.2 180 240 24 0.9 - - 7 4years19 Veerabadrappa 12938 55 M + 26.3 180 212 34 1.1 - - 6.6 7years

HbA1cFBS PPBS B/U


Name IP/OP No

Age

Sex

Micro Duration of DMAlb

HbA1cFBS PPBS B/U


Name IP/OP No

Age

Sex

20 Thippaya 28561 63 M + 19.8 189 232 23 1 + - 6.8 13years21 Radhamma 182273 60 F - 22.6 180 256 24 1.1 - - 7.1 6years22 Khaja bee 19650 60 F + 22.9 196 272 28 0.9 - - 8.1 6years23 Khaja moinuddi 21462 48 M - 20.3 154 230 20 0.8 - - 6.4 5years24 Shivamma 19630 42 F - 21.9 146 212 28 0.9 - - 6.3 4years25 Bheemavva 19411 48 F - 22 149 219 29 1 - - 6.4 4years26 Laxmi devi 3823 75 F + 18.2 214 356 29 1.2 + - 10.2 17years27 Bhemmayya 19363 62 M - 22.6 241 317 34 1.3 + - 9.5 12years28 Khaja bee 19650 65 F - 30.1 200 290 42 0.9 - - 7.8 11years29 Ismail 19412 53 M - 24.2 210 290 26 1.1 - - 7.4 6years30 Basavaraj 32752 50 M + 23.3 140 195 29 1 - - 6.1 5years31 Virupakshappa 30710 62 M - 23 156 240 20 1.1 - - 7.4 11years32 Amarappa 30730 55 M - 22.5 148 240 29 1.1 - - 7 6years33 Budappa 4 40 M - 21.1 150 226 31 1.2 - - 5.9 4years34 Narsamma 26914 46 F - 31.2 136 200 28 1 - - 6.1 4years35 Khaja moinuddi 21462 48 M - 20.8 158 250 29 0.9 - - 7.2 5years36 Thipamma 19654 55 F - 23.3 170 300 33 1 - - 8.4 6years37 Damayanthi bai 21417 65 F - 22.4 223 247 25 0.9 + - 7.2 11years38 Md.Hussain 21441 72 M - 20.8 220 280 25 0.8 + - 8 16years39 Khaja bee 4274 70 F + 24.5 180 260 29 1.2 + - 7.8 16years40 Hanumanthu 108095 64 M - 20.9 220 280 34 0.7 - - 8 11years41 Anasuya bai 6141 55 F - 21.6 212 250 26 0.9 - - 8.2 6years


HbA1cFBS PPBS B/U


Name IP/OP No

Age

Sex

42 Ashiya 7774 80 F + 32.5 280 340 26 0.8 + - 10.8 17years43 Huligappa 33630 72 M + 22.5 158 267 34 0.8 + - 9.8 16years44 Laxmidevi 23497 45 F - 21.3 190 235 40 1.2 - - 7.6 4years45 Kristappa 28728 61 M - 23 180 240 36 1 - - 7.2 11years46 Chandrasekhar 9590 65 M - 18.8 164 232 32 1.2 + - 7 11years47 Channabasayya 93924 48 M + 23.1 200 280 20 0.9 + - 8.1 4years48 Vidyasagar 135282 53 M - 21.6 162 240 27 1.2 - - 6.9 6years49 Shankarappa 102741 48 M - 26.3 201 258 30 1 - - 7.3 4years50 Mahadevi 142188 53 F + 24.9 156 242 29 1.3 - - 6.7 6years51 Thikkayya 28233 63 M + 24.3 149 232 27 0.7 - - 6.2 11years52 Neelakanta 2269 70 M + 30.4 186 260 31 0.8 + - 7.1 16years53 Tamizuddin 22925 55 M - 25 188 239 27 1.1 - - 6.8 6years54 Bheemsen 14003 55 M + 32.5 181 252 28 1.2 + - 7.2 6years55 Veerabadrappa 3952 40 M + 21.9 129 189 29 0.7 - - 6.1 3years56 Devamma 22389 51 F - 26.4 130 198 23 1.1 + - 6 6years57 Banamma 4694 50 F + 28.2 163 280 39 0.9 + - 7 5years58 Ramappa 108673 66 M - 21.9 166 250 34 1.2 - - 6.8 11years59 Siddalingamma 138346 45 F - 21.5 160 240 28 0.9 - - 6.6 4years60 Saranamma 9563 65 F + 19.8 205 380 27 1.3 + - 8.2 12years61 Satyamma 19544 54 F + 24.5 174 250 19 1.1 + - 7.3 8years62 Lakshmi 27019 55 F + 21.3 235 310 18 1.2 + - 8.9 8years63 S A Kumar 19397 40 M - 19.8 127 203 18 0.8 - - 5.9 4years


HbA1cFBS PPBS B/U


Name IP/OP No

Age

Sex

64 Sharawan kuma 28250 44 M - 25 130 201 32 0.9 + - 5.9 4years65 Satyamma 19544 54 F - 21.9 140 180 23 0.8 - - 6.1 6years66 Banubayi 153507 42 F + 26.2 210 257 22 0.8 + - 7.2 5years67 Nagamma 10 60 F - 22.5 190 280 29 1.1 + - 8.1 6years68 Veerendra 1393 40 M + 31.2 149 190 32 1.1 + - 5.9 5years69 Narsa reddy 28253 48 M - 26.3 200 251 22 0.9 + - 7.8 5years70 Hanumanthappa 21459 55 M - 22.5 140 242 27 1.2 - - 6.2 6years71 Sarasamma 9563 65 F - 20.8 168 250 40 1.1 - - 6.6 11years72 khaja bee 4274 72 F + 29.7 178 212 42 1.3 + - 6 16years73 Laxmidevi 3823 78 F + 20.3 220 320 22 1.3 + - 10.7 18years74 Hanumavva 21472 60 F - 20.1 129 190 33 1 + - 6 7years75 Basamma 2392 60 F - 22.6 145 184 22 1.2 - - 5.9 6years76 Sharawan kuma 28250 44 M - 20 180 263 23 1 - - 7.2 3years77 Khaja sab 11841 60 M - 18.8 170 219 21 0.9 - - 6.4 6years78 Nagayya 18563 75 M + 22.4 272 348 29 1.1 + - 9.2 16years79 Eshwaramma 21967 75 F + 21.1 200 326 32 0.9 + - 8.8 16years80 Mahanthamma 28380 55 F - 19.8 172 250 22 1.1 - - 7.4 7years81 Jayamma 152238 45 F - 24.2 195 240 31 1.3 - - 6.4 4years82 Gouramma 12951 50 F - 21.9 165 230 33 1.2 - - 6.8 5years83 Zathamagala 152232 45 F - 19.5 190 230 39 0.9 - - 6.6 3years84 Shamshad 14773 40 F - 20.9 182 270 42 1 - - 7.2 3years85 Narsappa 23855 42 M - 21.9 105 230 22 1.2 - - 6.8 3years


HbA1cFBS PPBS B/U


Name IP/OP No

Age

Sex

86 Madhavamma 12459 55 F + 24.2 215 321 41 1 + - 8.9 8years87 Basavaraj 13008 50 M - 22.1 141 210 33 1.1 - - 6 5years88 Thulijabai 27926 54 F + 31.2 195 260 39 0.9 + - 8 8years89 Somayya 9595 74 M + 26.7 210 292 43 1.3 + - 9.2 16years90 Banubai 153507 42 F - 25 155 230 29 0.9 - - 6.8 3years91 Parvathi 145441 46 F + 22 167 198 44 1.4 + - 6.3 5years92 Somappa 5290 67 M - 225 200 250 32 1.2 - - 6.8 11years93 Basangouda 22034 41 M - 32.5 170 250 38 1.2 + - 7.2 5years94 Ananth rao 21675 48 M - 21.9 165 291 28 1.1 - - 7.7 4years95 Hanumanthu 22381 40 M - 23.3 161 245 22 1.1 - - 6.8 3years96 Mahaveer 5214 58 M + 24 176 290 40 0.9 + - 8 8years97 Dharam singh 11234 56 M - 23.2 190 290 39 1.1 - - 7.9 6years98 Rangappa 28119 53 M - 23 166 270 26 1.3 - - 8 6years99 Taranath 29672 40 M - 22.5 145 190 28 0.8 - - 5.8 3years100 Sharangouda 6127 65 M + 24.7 210 290 29 1.3 + - 8.2 12years

Documents

ASSOCIATION BETWEEN MICROALBUMINURIA AND DIABETIC