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
×××××××××
×××
×××
0%
20%
40%
60%
80%
100%
120%
samples
% o
f p
lastic
plastic BME (12.8 mg/ml)
×××××××××
×××
×××
0%
20%
40%
60%
80%
100%
120%
samples
% o
f p
lastic
plastic BME (12.8 mg/ml)
×××××××××
×××
×××
0%
20%
40%
60%
80%
100%
120%
samples
% o
f p
lastic
plastic BME (12.8 mg/ml)
A
C
BME 17.1 mg/ml
B
D
E F G
A
C
BME 17.1 mg/ml
B
D
E F G
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