Saliva – salivary glands

–

lectures 1 and 2 and 3 Oral biology

Dr. Varga Gábor

2016

Composition and functions of saliva

No saliva - then what?

Sjögren syndrome

Irradiation induced atrophy of the acinar

parenchyma

The

digestive

tract

WATER FLUXES THROUGH THE

INTESTINE

No saliva

• Dry lips, dry mouth

• Difficult to swallow

• Difficult to chew

• Difficult to speak

• Difficult to taste

WATER 98%

MUCINS

GUSTIN

Most of the fundamental work on nervous

innervation of the salivary glands, stomach

and pancreas came from the work of Pavlov

and his students

Pancreatic fistula with pancreatic juice

Beaker

Nobel prize -1904 - Pavlov

Nobel prize – 1974 - Palade

Nobel prize – 1974 - Palade

Nobel prize – 1974 - Palade

SALIVARY GLANDS

Parotid gland anterior to ear

Accessory parotid gland

Parotid duct

Sublingual gland below tongue

Submandibular gland below lower jaw

1) Intrinsic Glands (Buccal glands): Inside oral cavity

2) Extrinsic Glands: Outside oral cavity; connected via ducts

More than 600

minor salivary

glands

Major glands

• Parotid: so-called watery serous saliva rich in

amylase, proline-rich proteins

– Stenson’s duct

• Submandibular gland: more mucinous

– Wharton’s duct

• Sublingual: viscous saliva

– ducts of Rivinus; duct of Bartholin

Minor glands

• Minor salivary glands are not found within

gingiva and anterior part of the hard palate

• Serous minor glands = von Ebner glands :

below the sulci of the circumvallate and folliate

papillae of the tongue

• Glands of Blandin-Nuhn: ventral tongue

• Palatine, glossopalatine glands are pure mucus

• Weber glands

Embryonic development

• The parotid: ectoderm (4-6 weeks of embryonic life)

• The sublingual-submandibular glands: endoderm

• The submandibular gland around the 6th week

• The sublingual and the minor glands develop around the 8-12 week

• Differentiation of the ectomesenchyme

• Development of fibrous capsule

• Formation of septa that divide the gland into lobes and lobules

Stages of salivary gland development –

epithelial/mesenchymal interactions Schematic showing prebud, initial bud, pseudoglandular,

canalicular and terminal bud stage of development in the SMG.

Prebud Initial bud Pseudo-

glandular Canalicular Terminal bud

Antisense oligonucleotides to keratinocyte

growth factor receptor (KGFR/Bek) decrease

branching morphogenesis of E12 SMGs

Individual FGFs and BMPs

have distinct morphological

effects on isolated epithelium

cultured in growth factor-

reduced Matrigel for

44 hours.

FGF1-, FGF4- and FGF10-

treated epithelium

form duct-like structures,

whereas FGF2 and FGF7

promote bud formation.

Epithelium treated with FGF8,

BMP4 or BMP7 alone do not

grow.

SMGs treated with rFGFR2b (which binds

differentiation factors) for 44 hours show decreased

epithelial cell proliferation (A-C) and increased

mesenchyme apoptosis (D-F)

A model of how FGF7 and

FGF10 signaling through

FGFR2b regulates

morphogenesis.

The model summarizes our

findings, and the dotted lines

show other potential

mechanisms: MMP2 may

regulate FGFR1 cleavage;

FGF1 expression may

stimulate both FGFR1 and

FGFR2; cofactors or co-

receptors may specify the

localization of FGF binding

and, therefore, where

proliferation occurs.

Signaling events likely propagate during

SMG development

A kinetic model incorporating unknown

transcription factor (TFx) activation by Eda/Edar

signaling

The cellular structure of the developed

salivary glands:

acini and ducts

Captured phase-contrast microscope photographs from

a 24-hour video showing the dynamics of acinotubular

structure formation on the surface of BME.

Captured phase-contrast microscope photographs from

a 24-hour video showing the dynamics of acinotubular

structure formation on the surface of BME

BME 6 mg/ml

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Claudin 1 expression indicating tight junction

formation of huSMG cells grown on Transwell filters (red propidium iodide staining shows cell nuclei)

Tran et al., Tissue Eng.

2005

Tissue organization of salivary

glands

• Acinus: serosus, mucinosus, mixed

• Duct: ductus intercalaris, ductus striatus, ductus

excretorius, main exretory duct

Acinus Striated duct Excretory duct

Intercalat

-ed duct

Parotid – serous gland

Sublingual – mucinous gland

Submandibular – mixed gland

STRUCTURAL ORGANIZATION OF

SALIVARY GLANDS

http://www.lab.anhb.uwa.edu.au/mb140/CorePages/Epithelia/Epithel.htm#Simple

Serous acini: well-stained, secretory vesicles visible, the nuclei are round or slightly ovoid, contain large amounts of rough ER. These acini produce a „watery” secretion.

Mucous acini: weakly stained, empty-looking vesicles give these cells a distinct "foamy" or "frothy" appearance, the nuclei are darker and smaller than the nuclei of serous cells, they seem to be "pressed" against the basal limit of the cells and may look flattened with an angular ("edgy") outline. They produce a rather „slimy” secretion.

Parotid gland H&E

Within the lobules and between the acini of the parotid there are two types of ducts. Striated ducts are lined by a simple tall columnar epithelium. Intercalated ducts are lined by a simple cuboidal epithelium and connect individual acini to the striated ducts. The main excretory duct conveys the secretory product to one of the external surfaces of the body.

http://www.lab.anhb.uwa.edu.au/mb140/CorePages/Epithelia/Epithel.htm#Simple

SALIVARY CONTROL/food

Stimulation of chemoreceptors and mechanoreceptors Increased salivation

(watery saliva)

Activation of parasympathetic motor neuron

Thinking Smelling Tasting

Stress / salivation

Stress / excitment

Increasing salivation - viscous

small volume High in proteins

Sympathetic motor neuron activation/β-adrenergic action

PIP2

IP3

DAG Ca2

+ Cl-

M3

Gq

PLC

cAMP

ATP

AC

GS

β adr

cAMP

Protein

Composition - inorganic

Na +

6 - 80 mmol/L

Cl -

17 - 30 mmol/L

K +

20 - 30 mmol/L

Ca 1 - 2 mmol/L

P 2 - 23 mmol/L

HCO3+

2 - 80 mmol/L

Electrolyte concentrations in basal

and stimulated mixed saliva

Plasma Stimulated Basal

Na+ (mmol/l) 145 5 20–80

K+ (mmol/l) 4 22 20

Ca2+ (mmol/l) 2,2 1–4 1–4

Cl− (mmol/l) 120 15 30–100

HCO3

− (mmol/l) 25 5 15–80

phospate (mmol/l) 1,2 6 4

Mg2+ (mmol/l) 1,2 0.2 0.2

SCN− (mmol/l) <0.2 2,5 2

NH3 (mmol/l) 0.05 6 3

(NH2)2CO (mmol/l) 2–7 3,3 2–4

Protein (g/l) 70 3 3

Saliva – water (almost…)

•Osmolality

•Extracellular

•How to secrete it

How to secrete water

• Actively move ions

• Sodium and chloride (active anion

transport)

• If possible, conserve sodium (and

chloride) by reabsorption

Salivation –

two-stage hypothesis

• Az acinar cells produce isoosmotic primary

saliva

• Passing through the ductal system

reabsorption of electolytes happens without

water movement resulting hypoosmotic fluid

• The composition of saliva depends on the

rate of salivary secretion (flow rate)

Acini Duct Primary secretion

Isotonic

FLOW RATE CURVES OF SALIVA AND

THE TWO-STAGE HYPOTHESIS

Secondary ductal modification Hypotonic

H2O

K+

HCO3ˉ

H2O

Na+

Cl-

Cl-

Na+

HCO3ˉ

K+

Saliva

HCO3ˉ

Plasma Flow ml/min

Concentration

mEq/l

K+

HCO3ˉ

K+

Na+

Cl-

Saliva Na+

Cl-

Main transporters-channels-pumps

• Primary pumps : ATP supplied energy liberation supports ion movements against gradient

• Facilitating transporters: carry various ions or uncharged molecules driven by concentration or electrochemical gradients. Based on the number and direction of moved particules, we may differentiate between uniporters, antiporters and cotransporters

• Ion channels: in open stage selectively allows certain cations to pass through membranes towards electrochemical gradients

• Water channels: allows to move water passively through membranes

Transporters of salivary glands

Acinar transporters –

Primary secretion

Na+

K+

2Cl-

K+

Na+

K+

Na+

H+

Cl-

HCO3-

Cl -

HCO3 -

H2O

AQP

H2O

Na+

CO2 CA

H2CO3

H+

HCO3 -

Na +

NBC

AE

NKCC

1

NA-K

ATPase

NHE

HCO3-

H2O

Cl- Na+

HCO3- H2O

Az acinar cell transporters Basolateral side

• Na+/K+-ATPase

• cation/chloride-

cotransporters:

– Na+/K+/2Cl- cotransporter

– Na+/Cl- cotransporter

– K+/Cl- cotransporter

– unknown substrate specificity transp.

• Cl-/HCO3- (anion exchangers,

AEs, SLC26s)

• Na+/HCO3--cotransporter

• Na+/H+ exchanger (NHE)

• Ca2+-activated K+-channels

Apical side

Cl- -channels • intracellular Ca2+-level

sensitive

• cAMP-level sensitive

• extracellular ATP activated

• Hyperpolarization acivated

• Channels activated by cell swelling

Aquaporin water channels (AQP-5)

Transporters in acinar secretion

• A Na-pump

• A Na+/K+/2Cl--cotransporter

• A Ca2+ -activated K+ - and Cl--channels

• A Na+ follows paracellularly, and water follows transcellularly

Modell 1

Transporters in acinar secretion

• A Cl- ion through HCO3-

/Cl—exchanger

• Carbonic-anhydrase

facilitated bicarbonate

and H+ ion production

• A Na+/H+-antiporter -

(pH)ic regulation

Modell 2

Transporters in acinar secretion

• Luminal exit of HCO3-

instead of Cl-

secretion

Modell 3

Salivary gland transporters

Ductal transporters -

Electrolyte rescue (eNaC has a key role)

Lumen Interstitium

Na+

Cl-

Cl-

HCO3-

H+

H+

Na+

K+

Cl-

K+

H+

3Na+

2K+

Na+

Ductal reabsoroption mechanizms

Isosmotic

primary

secretion

Hipos-

motic

saliva

Na

Cl

n

a

3 Na

2 K

3 Na

2 K

Cl

Na

H

H K

- a ducts are impermeable for water

- NaCl reabsorption by (Na – K –

pump, eNaC és Cl – channel

participationl)

- luminal Na ions exchange for protons

(secondary active transport)

-luminalis H ions a exchange to K ions

(tertiery active transport)

- Hyposmotic saliva

K+

2K+

3Na+

Na+

H+ CO2

CO2

HCO3- H+

HCO3-

H2O

H2O HCO3-

CA

Cl-

CO2 CO2 H+ HCO3

-

CA

Cl-

Cl-

H+ Na+

3Na+ 2K+

K+

K+ 3Na+

2K+ Na+

2Cl- K+

Na+

H+

Cl-

Cl-

Na+

K+

H+

3Na+

2K+ 2K+

3Na+

Na+

H2O

H2O

Acini Primary secretion - isotonic fluid -

Duct Secondary ductal modification

- hypotonic fluid -

Composition - organic

TOTAL PROTEIN 1400 – 2000 mg/L

TOTAL MUCIN

CARBOHYDRATE

110 - 300 mg/L

MG1

MG2

Organic components of mixed saliva quantity Main function

Full protein 1400-2000 mg/l

Prolin-rich proteins 1000-1400 mg/l Caries protective

Lysozime 109 mg/l Antimicrobial

Lactoferrin na Antimicrobial

Sialoperoxidase 3 mg/l Antimicrobial

Secretoros IgA 194mg/l Antimicrobial

IgG 14 mg/l Antimicrobial

IgM 2 mg/l Antimicrobial

Statherin na Caries protective

Gustin ~ 42-60mg/l Taste sensation facilitation

Histatins na Antimicrobial

Cystatins na Tissue regeneration

Amylase 380 mg/l Digestion

Lipase (lingual gland origin) na Digestion

Urea 2-6 mmol/l Acid neutralization

Glucose 0.05 mmol/l „plaque feeding”

Aminoacids 1-2 mmol/l ?

No saliva

• Rampant dental caries

Acid buffering

HCO3-

NH4

(urea and aminoacids)

No saliva

• Erosion of Enamel

• No remineralisation Ca and PO4

Ca-binding proteins

proline-rich proteins

statherin

No saliva

• Extensive and rapid dental caries

Antibacterial factors

Antibodies (IgA)

Sialoperoxidase + SCN

Lactoferrin

No saliva

• Candida infections

• Tissue damage Histatins

Cystatins

No Saliva

• Digestive problems

Amylase

Lingual Gland Lipase

Most important causes for

hyposalivation disorders • Sjögren syndrome and othe and other autoimmun

disorders - frequently antiserum against M3 receptors – acinar parenchyme distruction

• Radiotherapy induced distruction – acinar parenchyme

• Systemic diseases and their treatment diabetes mellitus, antihypertensive and anxiolytic drugs

• Xerostomia – frequently only subjective feeling - more frequent in older ages (especially in women after menopausa)

Saliva as a diagnostic fluid

-

future perspectives

VERY IMPORTANT LINK, part of the preparation for the

exam

http://www.hspp.ucla.edu/wonglab/

Thank you for your attention