2012-02-20 Nano Saliva

8/10/2019 2012-02-20 Nano Saliva

1/49

SALIVASALIVA

NANOTECHNOLOGYNANOTECHNOLOGY

Retty RatnawatiRetty Ratnawati

Physiology Lab.Physiology Lab.

Faculty of MedicineFaculty of Medicine

Brawijaya UniversityBrawijaya University

8/10/2019 2012-02-20 Nano Saliva

2/49

ChemicalChemical

composition andcomposition and

functions of salivafunctions of saliva

8/10/2019 2012-02-20 Nano Saliva

3/49

33Retty Ratnawati 2009Retty Ratnawati 2009

Chronology of definingrono ogy o e n ng

salivary comonents andsalivary comonents and

functionsfunctionsBeginning in !"#$%sBeginning in !"#$%s whole salivawhole salivaevaluatedevaluated&antimicrobial roerties' role in microbial&antimicrobial roerties' role in microbial

attachment' minerali(ation' taste' lubrication)attachment' minerali(ation' taste' lubrication)

*ecretions of*ecretions of major glands

major glands&arotid and&arotid and

submandibular+sublingual)submandibular+sublingual)

,n !"-$%s,n !"-$%s individual comonentsindividual co

monentsisolated andisolated and

biochemically characteri(edbiochemically characteri(ed,n mid!"/$%s beginning to ma,n mid!"/$%s beginning to ma functionalfunctional

domainsdomains&etide synthesis and recombinant&etide synthesis and recombinant

aroaches)aroaches)

8/10/2019 2012-02-20 Nano Saliva

4/49


Major salivary comonentsMajor salivary comonents

Mucin 1 (MG1)Mucin 1 (MG1)

sIgAsIgAMucin 2 (MG2)Mucin 2 (MG2)

LactoferrinLactoferrin

PeroxidasesPeroxidases

AmylasesAmylasesCarbonic anhydrasesCarbonic anhydrases

Prolinerich !roteinsProlinerich !roteinsLyso"ymeLyso"yme

#tatherins#tatherins$istatins$istatins

11 1010 100100 10001000 1000010000

Size (kDa)Size (kDa)

8/10/2019 2012-02-20 Nano Saliva

5/49


Current concets regardingCurrent concets regarding

the functional features ofthe functional features ofsalivary macromoleculessalivary macromolecules

Recent structure+function studies haveRecent structure+function studies have

identifiedidentified general rinciles

general rincilesregardingregarding

functionfunction

Based onBased on in vitroin vitrostudies of urifiedstudies of urified

moleculesmolecules0dditional studies re1uired to evaluate

0dditional studies re1uired to evaluate

concetsconcets in situin situ

8/10/2019 2012-02-20 Nano Saliva

6/49


Conformational re1uirementsConformational re1uirements

Conformation or shae of aConformation or shae of amolecule ismolecule is criticalcriticalfor itsfor itsbiological functionbiological function

23amles23amles Prolinerich roteins interact withProlinerich roteins interact withA.A.

viscosusviscosus andand St. gordoniiSt. gordonii only whenonly whenadsorbedadsorbedonto minerali(ed surfaceonto minerali(ed surface

*tatherins and histatins re1uire*tatherins and histatins re1uire helicalhelicalconformationconformation

4uman salivary amylase re1uire #4uman salivary amylase re1uire #interchaininterchain disulfide bondsdisulfide bonds

8/10/2019 2012-02-20 Nano Saliva

7/49

MultifunctionalityMultifunctionality

SalivarySalivary

FamiliesFamilies

Anti-Anti-

BacterialBacterialBufferingBuffering

DigestionDigestion

MineralMineral

i"ationi"ation

LubricatLubricat

ion %&iscoion %&isco

elasticityelasticity

'issue'issue

CoatingCoating

Anti-Anti-

FungalFungal

Anti-Anti-

ViralViral

Carbonic anhydrases,Carbonic anhydrases,

HistatinsHistatins

Amylases,Amylases,

Mucins, LipaseMucins, Lipase

Cystatins,Cystatins,

Histatins, Proline-Histatins, Proline-rich proteins,rich proteins,

StatherinsStatherins

Mucins, StatherinsMucins, Statherins

Amylases,Amylases,

Cystatins, Mucins,Cystatins, Mucins,

Proline-rich proteins, StatherinsProline-rich proteins, Statherins

HistatinsHistatins

Cystatins,Cystatins,

MucinsMucins

Amylases, Cystatins,Amylases, Cystatins,

Histatins, Mucins,Histatins, Mucins,PeroxidasesPeroxidases

adapted from M.J. Levine, 1993adapted from M.J. Levine, 1993

8/10/2019 2012-02-20 Nano Saliva

8/49


RedundancyRedundancy

Saliva has built-inSaliva has built-in redundancyredundancyin regard to itsin regard to itsprotective unctions!protective unctions!

"#a$ple - %any salivary $olecules can inhibit"#a$ple - %any salivary $olecules can inhibit

the precipitation o calciu$ phosphate salts!the precipitation o calciu$ phosphate salts! strong inhibitors such as statherin and acidicstrong inhibitors such as statherin and acidic

proline-rich proteinsproline-rich proteins

$oderate inhibitors such as histatins and cystatins$oderate inhibitors such as histatins and cystatins

wea& inhibitors such as $ucins and a$ylasewea& inhibitors such as $ucins and a$ylase

8/10/2019 2012-02-20 Nano Saliva

9/49


0mhifunctionality0mhifunctionality

0 molecule may have both rotective and detrimental roerties 0 molecule may have both rotective and detrimental roerties

55doubleedged sworddoubleedged sword6.6.

May deend on molecule%sMay deend on molecule%s locationlocationor site of actionor site of action

0mylases0mylases

,n solution' they facilitate clearance of viridans stretococci,n solution' they facilitate clearance of viridans stretococci

0dsorbed to tooth surface' they can romote adherence of0dsorbed to tooth surface' they can romote adherence ofthese bacteria and digest starch to dietary maltose andthese bacteria and digest starch to dietary maltose and

roduction of acidroduction of acid

*tatherin and acidic rolinerich roteins*tatherin and acidic rolinerich roteins

0t enamel surface' they lay an imortant role in minerali(ation0t enamel surface' they lay an imortant role in minerali(ation

by inhibiting the formation of rimary and secondary calciumby inhibiting the formation of rimary and secondary calciumhoshate salts. 7hen adsorbed to the enamel surface' theyhoshate salts. 7hen adsorbed to the enamel surface' they

romote attachment of cariogenic microorganisms.romote attachment of cariogenic microorganisms.

8/10/2019 2012-02-20 Nano Saliva

10/49

'0'0Retty Ratnawati 2009Retty Ratnawati 2009

Comle3ingComle3ing

Functional relationshis e3ist between differentFunctional relationshis e3ist between different

molecules in salivamolecules in saliva

8wo tyes of comle3ing &covalent and noncovalent)8wo tyes of comle3ing &covalent and noncovalent)

homotyic &between similar molecules)homotyic &between similar molecules) heterotyic &between different molecules)heterotyic &between different molecules)

23amle9 Mucins23amle9 Mucins

homotyic comle3es necessary for lubrication andhomotyic comle3es necessary for lubrication and

viscoelastic roertiesviscoelastic roerties heterotyic comle3es with s,g0' lyso(yme andheterotyic comle3es with s,g0' lyso(yme and

cystatins concentrate these antimicrobials at tissuecystatins concentrate these antimicrobials at tissue

interfacesinterfaces

8/10/2019 2012-02-20 Nano Saliva

11/49

''''Retty Ratnawati 2009Retty Ratnawati 2009

*alivary Protein Functions*alivary Protein Functions

Oral function Problem Protein function

Acts as an airway Air-born organisms Dehydration

Anti-bacterialsystems

Water-retainingglycoproteins

Speech Need for lubrication Lubrication system

Taste -- Gustin

Entry-point for foodmastication,swallowing

Food-bornorganisms

Soft and hard tissueabrasion

Food toxins


Lubrication;mucins, statherin

Toxin-neutralization


Acts as an airway Air-born organisms Dehydration


Water-retainingglycoproteins

Speech Need for lubrication Lubrication system

Taste -- Gustin

Entry-point for foodmastication,swallowing

Food-bornorganisms

Soft and hard tissueabrasion

Food toxins


Lubrication;mucins, statherin

Toxin-neutralization

8/10/2019 2012-02-20 Nano Saliva

12/49



&cont%d)&cont%d)


Control of in-digenous & in-

vading bacteria,fungi and viruses

Colonization &infection

Controllingpathogens andcommensals

Adhesion of bac-teria versus their

detection


Immunoglobu-lins, histatins,glycoproteins,lysozyme, sia-loperoxidase,

lactoferrin Adhesion-modulating pro-teins


Control of in-digenous & in-

vading bacteria,fungi and viruses

Colonization &infection

Controllingpathogens andcommensals

Adhesion of bac-teria versus their

detection


Immunoglobu-lins, histatins,glycoproteins,lysozyme, sia-loperoxidase,

lactoferrin Adhesion-modulating pro-teins

8/10/2019 2012-02-20 Nano Saliva

13/49



&cont%d)&cont%d)


Digestion __ Starch & fathydrolysis:

amylase andlingual lipase Protection &

repair of softtissues

Toxins,carcinogens,degradative

proteases

Mucin-richprotectivebarrier film

Proteaseinhibitors,cystatins, tissuegrowth factors


Digestion __ Starch & fathydrolysis:amylase andlingual lipase

Protection &repair of softtissues

Toxins,carcinogens,degradative

proteases

Mucin-richprotectivebarrier film

Proteaseinhibitors,cystatins, tissuegrowth factors

8/10/2019 2012-02-20 Nano Saliva

14/49



&cont%d)&cont%d)Oral function Problem Protein function

Protection & re-pair of hard tis-sues

Enamel mineralis potentiallysoluble; acid-

damagedenamel requiresremineralization

Biologicallycontrolled pro-tective & re-

pairative inor-ganic environ-ment, stabilizedby statherin,acidic proline-

rich and pellicleproteins Pellicle forma-

tion__ __

Plaque acid

formation

Plaque pH con-

trol

Basic amino

acids & peptides


Protection & re-pair of hard tis-sues

Enamel mineralis potentiallysoluble; acid-

damagedenamel requiresremineralization

Biologicallycontrolled pro-tective & re-

pairative inor-ganic environ-ment, stabilizedby statherin,acidic proline-

rich and pellicleproteins Pellicle forma-

tion__ __

Plaque acid

formation

Plaque pH con-

trol

Basic amino

acids & peptides

8/10/2019 2012-02-20 Nano Saliva

15/49


MucinsMucins

(ac& precise olded structure o globular proteins(ac& precise olded structure o globular proteins

)sy$$etrical $olecules with open* rando$ly)sy$$etrical $olecules with open* rando$ly

organi+ed structureorgani+ed structure

,olypeptide bac&bone apo$ucin. with /1 side-,olypeptide bac&bone apo$ucin. with /1 side-

chainschainsSide-chains $ay end in negatively charged groups*Side-chains $ay end in negatively charged groups*

such as sialic acid and bound sulatesuch as sialic acid and bound sulate

ydrophillic* entraining water resists dehydration.ydrophillic* entraining water resists dehydration.

niue rheological properties e!g!* high elasticity*niue rheological properties e!g!* high elasticity*adhesiveness* and low solubility.adhesiveness* and low solubility.

wo $aor $ucins %' and %2.wo $aor $ucins %' and %2.

8/10/2019 2012-02-20 Nano Saliva

16/49


Mucin FunctionsMucin Functions

issue /oatingissue /oating ,rotective coating about hard and sot tissues,rotective coating about hard and sot tissues

,ri$ary role in or$ation o acuired pellicle,ri$ary role in or$ation o acuired pellicle

/oncentrates anti-$icrobial $olecules at $ucosal/oncentrates anti-$icrobial $olecules at $ucosal

interaceinterace

(ubrication(ubrication )lign the$selves with direction o low)lign the$selves with direction o low

characteristic o asy$$etric $olecules.characteristic o asy$$etric $olecules. 7ncreases lubricating ualities il$ strength.7ncreases lubricating ualities il$ strength.

il$ strength deter$ines how eectively opposedil$ strength deter$ines how eectively opposed

$oving suraces are &ept apart$oving suraces are &ept apart

8/10/2019 2012-02-20 Nano Saliva

17/49

''Retty Ratnawati 2009Retty Ratnawati 2009

%ucin unctions cont:d.%ucin unctions cont:d.

)ggregation o bacterial cells)ggregation o bacterial cells ;acterial adhere to $ucins $ay result in surace;acterial adhere to $ucins $ay result in surace

attach$ent* orattach$ent* or

%ucin-coated bacteria $ay be unable to attach to%ucin-coated bacteria $ay be unable to attach to

suracesurace

;acterial adhesion;acterial adhesion

%ucin oligosaccharides $i$ic those on $ucosal cell%ucin oligosaccharides $i$ic those on $ucosal cellsuracesurace

React with bacterial adhesins* thereby bloc&ing the$React with bacterial adhesins* thereby bloc&ing the$

8/10/2019 2012-02-20 Nano Saliva

18/49


)$ylases)$ylases

Calcium metalloen(ymeCalcium metalloen(yme4ydroly(es4ydroly(es &!:) bonds of starches such as&!:) bonds of starches such as

amylose and amyloectinamylose and amyloectin

*everal salivary isoen(ymes*everal salivary isoen(ymes

Maltose is the major endroduct &;$< is glucose)Maltose is the major endroduct &;$< is glucose)550ears6 to have digestive function0ears6 to have digestive function

7hy is it also resent in tears' serum' bronchial'7hy is it also resent in tears' serum' bronchial'

and male and female urogenital secretions=and male and female urogenital secretions=

0 role in modulating bacterial adherence=0 role in modulating bacterial adherence=

8/10/2019 2012-02-20 Nano Saliva

19/49


(ingual (ipase(ingual (ipase

Secreted by von "bner:s glands o tongueSecreted by von "bner:s glands o tongue

7nvolved in irst phase o at digestion7nvolved in irst phase o at digestion

ydroly+es $ediu$- to long-chain triglyceridesydroly+es $ediu$- to long-chain triglycerides7$portant in digestion o $il& at in new-born7$portant in digestion o $il& at in new-born

nli&e other $a$$alian lipases* it is highlynli&e other $a$$alian lipases* it is highly

hydrophobic and readily enters at globuleshydrophobic and readily enters at globules

8/10/2019 2012-02-20 Nano Saliva

20/49


*tatherins*tatherins

Calcium hoshate salts of dental enamel areCalcium hoshate salts of dental enamel aresoluble under tyical conditions of 4 andsoluble under tyical conditions of 4 and

ionic strengthionic strength

*uersaturation of calcium hoshates*uersaturation of calcium hoshates

maintain enamel integritymaintain enamel integrity

*tatherins revent reciitation or*tatherins revent reciitation or

crystalli(ation of suersaturated calciumcrystalli(ation of suersaturated calcium

hoshate in ductal saliva and oral fluidhoshate in ductal saliva and oral fluidProduced by acinar cells in salivary glandsProduced by acinar cells in salivary glands

0lso an effective lubricant0lso an effective lubricant

8/10/2019 2012-02-20 Nano Saliva

21/49

2'2'Retty Ratnawati 2009Retty Ratnawati 2009

Prolinerich Proteins &PRPs)Prolinerich Proteins &PRPs)

Li>e statherin' PRPs are also highlyLi>e statherin' PRPs are also highly

asymmetricalasymmetrical

,nhibitors of calcium hoshate crystal,nhibitors of calcium hoshate crystalgrowthgrowth

,nhibition due to first ?$ residues of,nhibition due to first ?$ residues of

negativelycharged aminoterminal endnegativelycharged aminoterminal endPresent in the initially formed enamelPresent in the initially formed enamel

ellicle and in 5mature6 elliclesellicle and in 5mature6 ellicles

8/10/2019 2012-02-20 Nano Saliva

22/49


Role of PRPsRole of PRPs

in enamel ellicle formationin enamel ellicle formation

0c1uired enamel ellicle is $.!!.$ @m thic> layer0c1uired enamel ellicle is $.!!.$ @m thic> layer

of macromolecular material on the dentalof macromolecular material on the dental

mineral surfacemineral surface

Pellicle is formed by selective adsortion ofPellicle is formed by selective adsortion of

hydro3yaatitereactive salivary roteins' serumhydro3yaatitereactive salivary roteins' serum

roteins and microbial roducts such asroteins and microbial roducts such as

glucans and glucosyltransferaseglucans and glucosyltransferasePellicle acts as a diffusion barrier' slowing bothPellicle acts as a diffusion barrier' slowing both

attac>s by bacterial acids and loss of dissolvedattac>s by bacterial acids and loss of dissolved

calcium and hoshate ionscalcium and hoshate ions

8/10/2019 2012-02-20 Nano Saliva

23/49


Re$inerali+ation o ena$el andRe$inerali+ation o ena$el and

calciu$ phosphate inhibitorscalciu$ phosphate inhibitors

2arly caries are reaired desite resence of2arly caries are reaired desite resence of

minerali(ation inhibitors in salivaminerali(ation inhibitors in saliva

*ound surface layer of early carious lesion*ound surface layer of early carious lesion

forms imermeable barrier to diffusion of highforms imermeable barrier to diffusion of high

mol.wt. inhibitors.mol.wt. inhibitors.

*till ermeable to calcium and hoshate ions*till ermeable to calcium and hoshate ions

,nhibitors may encourage minerali(ation by,nhibitors may encourage minerali(ation byreventing crystal growth on the surface ofreventing crystal growth on the surface of

lesion by >eeing ores oenlesion by >eeing ores oen

8/10/2019 2012-02-20 Nano Saliva

24/49


Calculus formation andCalculus formation and

calcium hoshate inhibitorscalcium hoshate inhibitors

Calculus forms in la1ue desite inhibitoryCalculus forms in la1ue desite inhibitory

action of statherin and PRPs in salivaaction of statherin and PRPs in saliva

May be due to failure to diffuse intoMay be due to failure to diffuse intocalcifying la1uecalcifying la1ue

Proteolytic en(ymes of oral bacteria or lysedProteolytic en(ymes of oral bacteria or lysed

leu>ocytes may destroy inhibitory roteinsleu>ocytes may destroy inhibitory roteins

Pla1ue bacteria may roduce their ownPla1ue bacteria may roduce their own

inhibitorsinhibitors

8/10/2019 2012-02-20 Nano Saliva

25/49


Calcium hoshate reciitationCalcium hoshate reciitation

inhibitors and la1ueinhibitors and la1ue

*tatherin and PRPs might be e3ected to occur*tatherin and PRPs might be e3ected to occur

in la1ue' have not been detectedin la1ue' have not been detected

Pla1ue bacteria roduce calcium hoshatePla1ue bacteria roduce calcium hoshate

inhibitorsinhibitors

Might be necessary to revent calcification ofMight be necessary to revent calcification of

bacteria haens with dead cellsbacteria haens with dead cells

,mmobili(ed crystal growth inhibitors can,mmobili(ed crystal growth inhibitors canfunction as nucleators of crystal growthfunction as nucleators of crystal growth

,mmobili(ation may occur in la1ue',mmobili(ation may occur in la1ue'

encouraging calculus formationencouraging calculus formation

8/10/2019 2012-02-20 Nano Saliva

26/49


,nteraction of oral bacteria with,nteraction of oral bacteria with

PRPs and other ellicle roteinsPRPs and other ellicle roteins

*everal*everalsalivary roteins aear to be involved insalivary roteins aear to be involved in

reventing or romoting bacterial adhesion to oral softreventing or romoting bacterial adhesion to oral soft

and hard tissuesand hard tissues

PRPs are strongPRPs are strong romotersromotersof bacterial adhesionof bacterial adhesion

0mino terminal9 control calcium hoshate chemistry0mino terminal9 control calcium hoshate chemistry

Carbo3y terminal9 interaction with oral bacteriaCarbo3y terminal9 interaction with oral bacteria

,nteractions are highly,nteractions are highly secificsecific

Aeends on rolineglutamine carbo3yterminalAeends on rolineglutamine carbo3yterminaldietidedietide

PRPs in solution do not inhibit adhesion of bacteriaPRPs in solution do not inhibit adhesion of bacteria

8/10/2019 2012-02-20 Nano Saliva

27/49


hese anti-$icrobial proteinshese anti-$icrobial proteins

Secretory 7$$unoglobulinsSecretory 7$$unoglobulins

(actoerrin(actoerrin

(yso+y$e(yso+y$eSialopero#idaseSialopero#idase

/ystatins/ystatins

istatinsistatins

8/10/2019 2012-02-20 Nano Saliva

28/49


/linical ighlights/linical ighlights

nderstanding o salivary $echanis$s atnderstanding o salivary $echanis$s at

unda$ental level a prereuisite orunda$ental level a prereuisite or eective treat$ent o salivary glandeective treat$ent o salivary gland

dysunctionsdysunctions $odulation o bacterial coloni+ation$odulation o bacterial coloni+ation

develop$ent o artiicial saliva other

8/10/2019 2012-02-20 Nano Saliva

29/49


8/10/2019 2012-02-20 Nano Saliva

30/49


8/10/2019 2012-02-20 Nano Saliva

31/49


8/10/2019 2012-02-20 Nano Saliva

32/49


8/10/2019 2012-02-20 Nano Saliva

33/49


8/10/2019 2012-02-20 Nano Saliva

34/49


8/10/2019 2012-02-20 Nano Saliva

35/49


8/10/2019 2012-02-20 Nano Saliva

36/49


8/10/2019 2012-02-20 Nano Saliva

37/49


8/10/2019 2012-02-20 Nano Saliva

38/49


8/10/2019 2012-02-20 Nano Saliva

39/49


8/10/2019 2012-02-20 Nano Saliva

40/49


8/10/2019 2012-02-20 Nano Saliva

41/49


8/10/2019 2012-02-20 Nano Saliva

42/49


8/10/2019 2012-02-20 Nano Saliva

43/49


8/10/2019 2012-02-20 Nano Saliva

44/49

8/10/2019 2012-02-20 Nano Saliva

45/49


8/10/2019 2012-02-20 Nano Saliva

46/49


8/10/2019 2012-02-20 Nano Saliva

47/49


8/10/2019 2012-02-20 Nano Saliva

48/49


(aurent Seugnet et al* 2006(aurent Seugnet et al* 2006

,dentification of biomar>er for,dentification of biomar>er for

slee drive in flies and humanslee drive in flies and human

$R>)-)$ylase $ening&at pada &eadaan aga$R>)-)$ylase $ening&at pada &eadaan aga

$ening&atnya sleep-debt. pada lalat$ening&atnya sleep-debt. pada lalat

?rosophila $icroarray studies.* identi& dg salah?rosophila $icroarray studies.* identi& dg salah

satu $R>) species dari 3000 sp. di salivasatu $R>) species dari 3000 sp. di saliva$anusia$anusia

8/10/2019 2012-02-20 Nano Saliva

49/49

2012-02-20 Nano Saliva

Documents

Transcript of 2012-02-20 Nano Saliva