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  1. 1. Select here for the Lecture Notes Inflammatory Periodontal Disease Dr. Robert J. Boackle
  2. 2. I . BACTERIA Percent Shift from Gram Positive (facultative) to Gram Negative -Anaerobes in Supragingival Plaque in Gingivitis. In Periodontitis - “SUBGINGIVAL PLAQUE” <ul><li>Porphyromonas gingivalis </li></ul><ul><li>Actinobacillus actinomycetemcomitans </li></ul>Two Very Important GRAM NEGATIVE Bacteria Closely Associated with with Periodontal Disease:
  3. 3. <ul><li>Porphyromonas gingivalis </li></ul><ul><li>http://biology. kenyon .edu/Microbial_Biorealm/bacteria/bacteroidete_chlorob_group/Porphyromonas/porphyromonas. htm </li></ul>May travel between cells ( intercellular movement ) via bacterial protease action and/or Into cells (e.g., into and through epithelial cells) termed intracellular movement and multiply in those host cells http: //iai . asm .org/cgi/content/full/69/4/2700 causing release of pro-inflammatory cytokines
  4. 4. Other Implicated Periopathogens: <ul><li>Bacteroides forsythus Fusobacterium nucleatum Prevotella intermedia Campylobacter rectus </li></ul><ul><li>Eikenella corrodens </li></ul><ul><li>But these Periopathogens may be detected both in inflammatory and in non-inflammatory sites of periodontal patients, as well as in the healthy subjects. </li></ul>
  5. 5. “ Porphyromonas gingivalis , one of the major causative agents of periodontal diseases, produces large amounts of arginine- and lysine-specific cysteine proteinases in cell-associated and secretory forms, which are now referred to as Arg-gingipain (abbreviated Rgp or Arg-X) and Lys-gingipain (abbreviated Kgp or Lys-X), respectively. …..With respect to the physiology of the bacterium, Rgp and Kgp are indispensable for it to obtain nutrients from the environment, since Porphyromonas gingivalis cannot utilize saccharides as carbon/energy sources for growth and totally depends on peptides and amino acids that are provided from environmental proteins by Rgp and Kgp. Furthermore, proteolytic activities of Rgp and Kgp contribute to processing/maturation of various cell-surface proteins of P. gingivalis , such as fimA fimbrilin (a subunit of major fimbriae), 75-kDa protein (a subunit of minor fimbriae), hemagglutinins, and the hemoglobin receptor protein, which are important for the bacterium to colonize and proliferate in the gingival crevice and to invade the periodontium. These findings strongly indicate critical roles of Rgp and Kgp in the virulence of P. gingivalis.” Taken from, Kadowaki et al., Journal of Biochemistry, Vol 128, 153-159, 2000
  6. 6. Porphyromonas gingivalis with chronic and severe adult periodontitis Actinobacillus actinomycetemcomitans with Localized Aggressive Periodontitis (LAP) (formally termed Advanced Onset Periodontitis and localized juvenile periodontitis ) Treponema denticola and Prevotella intermedia with acute necrotizing gingivitis
  7. 7. Coronary Heart Disease and Periodontal Disease http: //atvb . ahajournals .org/cgi/content/full/25/3/e17 High-serum antibody levels to major periodontal pathogens are associated with subclinical, prevalent, and future incidence of coronary heart disease (CHD). Periodontal pathogens or host response against them may contribute to the pathogenesis of CHD. Pussinen PJ, Nyyssonen K, Alfthan G, Salonen R, Laukkanen JA, Salonen JT. Serum antibody levels to Actinobacillus actinomycetemcomitans predict the risk for coronary heart disease. Arterioscler Thromb Vasc Biol. 2005, 25: 833-838. Pro-atherogenic properties of lipopolysaccharide from the periodontal pathogen Actinobacillus actinomycetemcomitans http://www.musc. edu/~boacklrj/ Porph-Atherosclerosis . pdf
  8. 8. Periodontal disease and bacterial vaginosis increase the risk for adverse pregnancy outcome. Oittinen J, Kurki T, Kekki M, Kuusisto M, Pussinen P, Vilkuna-Rautiainen T, Nieminen A, Asikainen S, Paavonen J. Infect Dis Obstet Gynecol. 2005 13:213-216. Will future reports substantiate this paper concerning women attempting to become pregnant? Please keep up with the Literature!
  9. 9. Note: Supragingival plaque , even from dental hygienists, is a natural habitat for periodontitis-associated bacteria in healthy persons. Colonization with: Actinobacillus actinomycetemcomitans Bacteroides forsythus Campylobacter rectus And is mostly stable in spite of better than average personal plaque control of dental hygienists. Supragingival Plaque
  10. 10. Bacterial colonization by; Actinobacillus actinomycetemcomitans (serotype b) and/or Porphyromonas gingivalis appear to be Associated With Early Onset Periodontitis (Juvenile Periodontitis) Now termed - Localized Aggressive Periodontitis (LAP) seen in young people, is a rare condition characterized by severe periodontal destruction around first molars and incisorswith little or no accumulation of visible plaque and/or calculus. It was previously named Localized Juvenile Periodontitis (LJP) and recently reclassified as Localized Aggressive Periodontitis (LAP).
  11. 11. BACTERIAL PROTEASES i.e., from Porphyromonas gingivalis A percentage of these proteases remains associated with the surface of the organism while the rest is liberated into the surroundings <ul><li>Collagenases ---------Directly damage the Periodontal Ligament which is made of Collagen </li></ul><ul><li>Gingipain proteases ----The Lys-X (Lys-gingipain) and Arg-X (Arg-gingipain.. RGP ) cysteine proteases of PORPHYROMONAS GINGIVALIS -Destroy intercellular “glue-like proteins” </li></ul><ul><li>Antibodies directed to the amino-terminal region of the catalytic domain of gingipains R are capable of inducing a protective immune response against P. gingivalis infection in the mouse chamber model. </li></ul>
  12. 12. The Bacterial Gingipain proteases -- cysteine proteases The Arg-X (Arg-gingipain.. RGP ) cysteine proteases & The Lys-X (Lys-gingipain) cysteine proteases of PORPHYROMONAS GINGIVALIS a) Destruction of host intercellular glue-like proteins: E-cadherin Beta1-Integrin and Occludin
  13. 13. <ul><li>b) Non-productive complement consumption and removal of deposited complement components (C3b) from the bacterial surface </li></ul><ul><li>c) Bradykinin release from kininogens- </li></ul><ul><li>Crevicular Fluid (Exudate) Flow Increases </li></ul><ul><li>(Bradykinin is normally excised from kininogens by the enzyme kallikrein) </li></ul>Other Actions of Bacterial Gingipain proteases ---- The Arg-X (Arg-gingipain.. RGP ) and Lys-X (Lys-gingipain) cysteine proteases of PORPHYROMONAS GINGIVALIS
  14. 14. &quot;Black pigmenting anaerobic bacilli” Bind many cells (via its Hemagglutinin B ) including epithelial cells, and degrade erythrocytes (one reason for being “Black Pigmented” ) Human Beta-Defensin 1 Interacts with Hemagglutinin B from Porphyromonas gingivalis Purified recombinant Hemagglutinin B (rHag B) is being used in animal models to induce a protective immune response and information may support the potential usefulness of this component of P. gingivalis in the development of a vaccine against adult periodontitis. Porphyromonas gingivalis
  15. 15. CH2 CH3 CH2 CH3 Serum IgA1
  16. 16. CH2 CH3 CH2 CH3 IgA1 IgA antibodies co-deposited with IgG antibodies that are on antigens or in the blood will pick-up “accept” complement fragments in areas of periodontal inflammation – where complement activation is occurring iC3b C3b
  17. 17. CH2 CH3 CH2 CH3 IgA1 And will “arm” PMNs Macrophages and Immature Dendritic Cells These cells have complement receptors for C3b iC3b and or C3d fragments PMN iC3b C3b
  18. 18. CH2 CH3 CH2 CH3 IgA1 And will “arm” PMN Macrophages and Immature Dendritic Cells These cells have complement receptors for C3b iC3b and / or C3d fragments PMN CR1 CR3 iC3b C3b
  19. 19. CH2 CH3 CH2 CH3 iC3b C3b BUT: What if IgA1 proteases released from Periopathogens e.g., P. gingivalis Cleave IgA1 before the complement is deposited IgA1 PMN CR1 CR3
  20. 20. CH2 CH3 CH2 CH3 Fc fragments have No ability to bind antigen IgA1
  21. 21. CH2 CH3 CH2 CH3 C3d C3d Antigen Presenting Cells (B cell or Immature Dendritic Cell or Macrophage CR2 CR2 Thus this entire process is not allowed to happen. Nor will the properly directed stimulation of antibody responses occur. Also in the absence o the intact intercepting IgA1, inadvertent deposition of complement directly on dendritic cells could non-specifically stimulate the immune system and lead to IgG2 production. P. Gingivalis Antigens IgA1
  22. 22. Effects of the IgA1 Proteases released from periodontal pathogens and the released of free Fc alpha1 and of free Fab fragments to compete with intact antibodies: A lower degree of Specific B (and T) cell responses, leading to a continued infection and more non-specific polyclonal antibody responses due to the release of polyclonal activators (i.e., LPS) and Superantigens.
  23. 23. http://www. musc . edu/~boacklrj/BacterialProteases . pdf Please read the references below regarding proteases and periodontal disease http://www. musc . edu/~boacklrj/regulationGingipains . pdf Please Use this site to search for these papers
  24. 24. BACTERIAL ANTIGENS and BACTERIAL MITOGENS <ul><li>Induce Mitosis of host lymphocytes </li></ul><ul><li>B cell activation (Specific and/or Polyclonal) </li></ul><ul><li>Differentiation into PLASMA CELLS </li></ul><ul><li>[Note: a relatively large number of plasma cells is present in inflamed periodontal tissues] </li></ul>
  25. 25. Contact with BACTERIAL ENDOTOXINS (Lipopolysaccharides--LPS) Several different Toll-Like Receptors (TLR) for LPS are on host cells— activation of nuclear factor-  B (NF-  B) indicates the production of Proinflammatory Cytokines from those host cells. http://www. musc .edu/~boacklrj/ Cytokines-TLR-Periodontal.pdf http://www.musc. edu/~boacklrj/ osteoclast-Bcell . pdf h ttp:// http://www. musc . edu/~boacklrj/hyporesponsiveness . pdf <ul><li>INDIRECTLY Activate OSTEOCLASTS-mediated Bone resorption via the stimulation of the production of the above mentioned pro-inflammatory cytokines e.g., Interleukin 1 (IL-1) and IL-6 </li></ul><ul><li>Or via direct interactions of antigen stimulated B Cells with Osteoclasts http://www. musc . edu/~boacklrj/ osteoclast-Bcell .pdf </li></ul><ul><li>Also the Coating of periodontal tissues with Bacterial Products like endotoxin may disrupt a timely healing. </li></ul><ul><li>Endotoxins are neutralized by Complement, if sufficient amounts of complement are present </li></ul>
  26. 26. Direct BACTERIAL CHEMOTACTIC FACTORS ---from living bacteria n-formyl-methionine residue <ul><li>Attract PMN (if PMNs are normal) </li></ul>
  27. 27. II. HOST RESPONSE Inflamed Gingival Tissues : Crevicular Fluid Flow Rate Increases
  28. 28. Crevicular Fluid is a serum exudate HOST RESPONSE Crevicular Fluid Flow Rate Increases
  29. 29. HOST RESPONSE 1) Human Leukocyte Elastase from PMN and Macrophages 2) Human Matrix Metalloproteinases (MMP) (IL1- induced) from PMN, macrophages and fibroblasts 3) Lysozyme digests peptidoglycan, a part of the backbone of gram positive Bacterial cell walls In the Inflamed Gingival Tissues : Important digestive enzymes released from Our (HOST) PMNs, macrophages and Fibroblasts
  30. 30. <ul><li>A powerful host protease released from PMNs (termed HLE) and Macrophages (also termed MMP-12) </li></ul><ul><li>Important Substrates of Elastase in inflammatory situations like periodontal disease: </li></ul><ul><li>Elastin (matrix destruction) </li></ul><ul><li>C1 Inhibitor (deregulates Classical and Lectin complement pathways) </li></ul><ul><li>Alpha-1-proteinase inhibitor (alpha1-PI) </li></ul>Human Leukocyte Elastase
  31. 31. IL1- induced MMPs released from PMN, macrophages and fibroblasts (Certain Tetracyclines may inhibit MMP activity). some MMPs are Zn ++ dependent We Destroy our own Collagen (e.g., periodontal ligament) C1 (collagen part) Mannan Binding Lectin (collagen part) Human Matrix Metalloproteinases These are Host Collagenases
  32. 32. SPECIFIC ANTIBODY Neutralizes BACTERIAL SUBSTANCES e.g., bacterial proteases “Antibodies to gingipain promote uptake of P. gingivalis by PMNs” Gibson et al. Gingipain-specific IgG in the sera of patients with periodontal disease is necessary for opsonophagocytosis of Porphyromonas gingivalis. J Periodontol. 2005 Oct;76(10):1629-36. In the inflamed periodontal tissues, both IgG and IgA are produced in situ. This means that plasma cells in the periodontal tissues are liberating IgG or IgA1, then later IgA2 antibody producing plasma cells appear. While the IgG (especially IgG3 and IgG1) activates the classical pathway of complement, serum IgA1 and perhaps IgA2 antibody appear to have a regulatory and transport mechanism in complement activation and preserve IgG1 binding. IgG2 is a weaker complement activator that is produced “out of proportion” in response to Actinobacillus actinomycetemcomitans. http://www. musc . edu/~boacklrj/ Actinobacillus-specific-IgG . pdf
  33. 33. Complement <ul><li>Neutralizes BACTERIAL SUBSTANCES via the classical complement pathway (immune complexes) and via the alternative pathway. Covalent C3b binding changes the nature of the bacterial LPS-endotoxin and detoxifies it. </li></ul><ul><li>C5a --Chemotactic factor (high level of PMN in diseased periodontal tissues). PMNs release oxygen radicals to kill microbes in the phagolysozome) but these radicals when released (leaked) outside the PMNs can cause damage to surrounding host tissues. </li></ul><ul><li>Complement can damage host tissue ( bystander membrane attack complex). </li></ul><ul><li>Our periodontal host cells (fibroblasts, epithelial cells etc.) up-regulate their expression of membrane complement inhibitors (DAF and Protectin) to protect themselves against inadvertently deposited C3b and MAC and resist bystander damage by the membrane attack complex of complement. </li></ul>
  34. 34. Lymphocytes <ul><li>The adaptive immune response, and in particular stimulated CD4 positive T cells and the pro-inflammatory cytokines that they secrete (IL1 and IL6), are important effectors of bone loss consequent to Porphyromonas gingivalis oral infection. In animal studies, mice genetically lacking the cytokine &quot;gamma interferon&quot; or &quot;interleukin-6&quot; demonstrated decreased bone loss during the infection. </li></ul><ul><li>Cytotoxic T Lymphocytes may also release lymphotoxins, which kill host cells, perforins , which can damage host cell membranes and release chemotactic factors (interleukins) attracting PMNs. </li></ul><ul><li>Endotoxin induced Tolerance to suppress the specific immune response? http://www. musc . edu/~boacklrj/ hyporesponsiveness .pdf </li></ul><ul><li>Lower number of Toll-Like Receptors? </li></ul>
  35. 35. HUMAN EPITHELIAL CELLS --- Eikenella corrodens and “intracellular” habitation by P. gingivalis induces the human epithelial cells to secrete pro-inflammatory cytokines (IL-6 and IL-8) which serve as an early signaling system to host immune and inflammatory cells in underlying connective tissues. Involvement of HUMAN GINGIVAL FIBROBLASTS Human Matrix Metalloproteinases (IL1- induced) liberated from fibroblasts (and PMN) in the inflamed tissue When contacting Human gingival fibroblasts There is an &quot;additive&quot; effect of both IL-1 along with P. gingivalis - LPS on IL-6 production by Osteoblasts Result Bone Resorption
  36. 36. &quot;DEFECTS&quot; in PMN / lymphocyte function Advanced Onset Periodontitis (Localized Juvenile Periodontitis) Diabetes ------ Prolonged high glucose levels lead to altered energy metabolism (PMN dysfunction), and to abnormal glycosylation of serum proteins and “many cell-surface receptors” leading to cell malfunction and a weaker immune response. Secondary PMN and lymphocyte dysfunction may occur in uncontrolled diabetes. AIDS Patients ---Develop Periodontal Disease
  37. 37. <ul><ul><ul><ul><li>PLASMA CELLS </li></ul></ul></ul></ul><ul><li>High levels of plasma cells in gingival tissue with production of </li></ul><ul><li>A) Specific Ab and </li></ul><ul><li>B) Non-specific Ab </li></ul>
  38. 38. Important points concerning Specific IgG, IgA, complement and periodontal disease: Specific IgG, IgA1 and IgA2 antibody to subgingival Periodontal Pathogens are produced in situ by local plasma cells in the inflamed periodontal tissues. “ Antibodies to gingipain promote uptake of P. gingivalis by PMNs” Gibson et al. Gingipain-specific IgG in the sera of patients with periodontal disease is necessary for opsonophagocytosis of Porphyromonas gingivalis. J Periodontol. 2005 Oct;76(10):1629-36. SPECIFIC ANTIBODY PRODUCED in situ Some Periodontopathic Bacteria ( P. gingivalis ) produce IgA1 proteases specific only for IgA1 Why does this protease help the periodontopathic bacteria to survive in the subgingival tissues? Potential Exam question?
  39. 39. Non-specific salivary Ab Salivary IgA Antibody to Sheep erythrocytes is present in 1/2 of the severe periodontal disease cases that diminishes after successful periodontal surgery Why?
  40. 40. Please read the ABSTRACTS in the back of your Handout. These abstracts will help with your National Boards (Part 1 and especially Part 2).