Dennis E. Lopatin, Ph.D. Page no. 1 Anti-microbial activities of saliva

Dennis E. Lopatin, Ph.D. Page no. 2 Time course of sIgA appearance 3m6m2y?1m2-4w8w11w19w26w Gestation Birth SCBronchialEpithel-ium Peyer’sPatches SCSalivaryGland IgACells Saliva: Adult SC No IgA SalivasIgA Salivary Antibody to Initial Oral and Gut Flora Early IgA Peak ToothEruption Many Salivary IgA Concentrations in Adult Range AdultConcen-trations Adapted from Taubman & Smith, 1993

Dennis E. Lopatin, Ph.D. Page no. 3 Non-immunoglobulin anti-microbial agents in whole saliva

Dennis E. Lopatin, Ph.D. Page no. 4 Lactoferrin Binds ferric iron (Fe 3+ ) and unavailable for microbes Binds ferric iron (Fe 3+ ) and unavailable for microbes Nutritional immunity Nutritional immunity Some microorganisms (e.g., E. coli) have adapted to this mechanism by producing enterochelins. Some microorganisms (e.g., E. coli) have adapted to this mechanism by producing enterochelins. –bind iron more effectively than lactoferrin –iron-rich enterochelins are then reabsorbed by bacteria Lactoferrin, with or without iron, can be degraded by some bacterial proteases. Lactoferrin, with or without iron, can be degraded by some bacterial proteases. In unbound state, a direct bactericidal effect In unbound state, a direct bactericidal effect

Dennis E. Lopatin, Ph.D. Page no. 5 Lysozyme Present in numerous organs and most body fluids Present in numerous organs and most body fluids Oral LZ is derived from at least four sources Oral LZ is derived from at least four sources –major and minor salivary glands, phagocytic cells and gingival crevicular fluid (GCF) Biological function Biological function –Classic concept of anti-microbial activity of LZ is based on its muramidase activity (hydrolysis of  (1-4) bond between N- acetylmuramic acid and N-acetylglucosamine in the peptidoglycan layer. –Gram negative bacteria generally more resistant than gram positive because of outer LPS layer

Dennis E. Lopatin, Ph.D. Page no. 6 Other anti-microbial activities of LZ Muramidase activity (lysis of peptidoglycan layer) Muramidase activity (lysis of peptidoglycan layer) Cationic-dependent activation of bacterial autolysins Cationic-dependent activation of bacterial autolysins –strongly cationic protein (pI ) –disrupts membranes Aggregation of bacteria Aggregation of bacteria Inhibition of bacterial adhesion to tooth surfaces Inhibition of bacterial adhesion to tooth surfaces Inhibition of glucose uptake and acid production Inhibition of glucose uptake and acid production De-chaining of streptococci De-chaining of streptococci

Dennis E. Lopatin, Ph.D. Page no. 7 Salivary peroxidase systems Sialoperoxidase (SP, salivary peroxidase) Sialoperoxidase (SP, salivary peroxidase) –Produced in acinar cells of parotid glands –Also present in submandibular saliva –Readily adsorbed to various surfaces of mouth »enamel, salivary sediment, bacteria, dental plaque Myeloperoxidase (MP) Myeloperoxidase (MP) –From leukocytes entering via gingival crevice –15-20% of total peroxidase in whole saliva

Dennis E. Lopatin, Ph.D. Page no. 8 Components of the peroxidase anti-microbial system Peroxidase enzymes (SP or MP) Peroxidase enzymes (SP or MP) Hydrogen peroxide (H 2 O 2 ) Hydrogen peroxide (H 2 O 2 ) –oral bacteria (facultative aerobes/catalase negative) produce large amounts of peroxide »S. sanguis, S. mitis, S. mutans Thiocyanate ion (SCN - ) which is converted to hypothiocyanite ion (OSCN - ) by peroxidase Thiocyanate ion (SCN - ) which is converted to hypothiocyanite ion (OSCN - ) by peroxidase –salivary concentration is related to diet and smoking habits

Dennis E. Lopatin, Ph.D. Page no. 9 Thiocyanate reactions The pK for HOSCN/OSCN - is 5.3 The pK for HOSCN/OSCN - is 5.3 More acid favors HOSCN More acid favors HOSCN Due to uncharged nature, HOSCN penetrates bacterial cell envelope better Due to uncharged nature, HOSCN penetrates bacterial cell envelope better H 2 O 2 + SCN - OSCN - +H 2 O SP and/or MP HOSCN OSCN - + H+ Hypothiocianous acid Hypothiocyanite ion Acid/Base Equilib.

Dennis E. Lopatin, Ph.D. Page no. 10 HOSCN/OSCN - -mediated cell damage can oxidize sulfhydryl groups of enzymes can oxidize sulfhydryl groups of enzymes block glucose uptake block glucose uptake inhibit amino acid transport inhibit amino acid transport damage inner membrane, leading to leakage of cell damage inner membrane, leading to leakage of cell disrupt electrochemical gradients disrupt electrochemical gradients

Dennis E. Lopatin, Ph.D. Page no. 11 Regulation of oral microorganisms by SP/MP Unstimulatedbacteria Inhibitedbacteria Activebacteria Food Ingestion carbohydrates StimulationStimulation MetabolismMetabolism H+H+H+H+ RecoveryRecovery O2O2O2O2 thiols InhibitionInhibition Salivary Glands SCN - + H 2 O 2 OSCN - /HOSCN Autoinhibition spontaneous +SP

Dennis E. Lopatin, Ph.D. Page no. 12 Histatins A group of small histidine-rich proteins A group of small histidine-rich proteins Potent inhibitors of Candida albicans growth Potent inhibitors of Candida albicans growth Histatin 1, which is phosphorylated modulates precipitation of calcium phosphates Histatin 1, which is phosphorylated modulates precipitation of calcium phosphates

Dennis E. Lopatin, Ph.D. Page no. 13 Amylases Well-known function as a digestive enzyme Well-known function as a digestive enzyme –Calcium metalloenzyme, which hydrolyses the  (1-4) bonds of starches, such as amylose and amylopectin. Anti-microbial activity Anti-microbial activity –potent inhibitor and specific inhibitor of N. gonorrheoeae and Legionella pneumonophila in vitro. –modulates adhesion of certain oral species to teeth and other body surfaces

Dennis E. Lopatin, Ph.D. Page no. 14 Cystatins Are inhibitors of cysteine-proteases Are inhibitors of cysteine-proteases Are ubiquitous in many body fluids Are ubiquitous in many body fluids Considered to be protective against unwanted proteolysis Considered to be protective against unwanted proteolysis –bacterial proteases –lysed leukocytes May play inhibit proteases in periodontal tissues May play inhibit proteases in periodontal tissues Also have an effect on calcium phosphate precipitation Also have an effect on calcium phosphate precipitation