Osteoclast/ dental courses


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Osteoclast/ dental courses

  1. 1. INDIAN DENTAL ACADEMY Leader in continuing Dental Education www.indiandentalacademy.com
  2. 2. References ₪Tencate, s - Oral Histology ₪Development structure and Function ₪Orbans Oral Histology – 13th edition ₪Osteoclast Differentiation ₪ G. David Roodman ₪ Department of Medicine, Division of Hematology, University of Texas Health Science Center and the Audie L. Murphy Memorial Veterans Hospital, Research Service, San Antonio, Texas www.indiandentalacademy.com
  3. 3. References ₪Identification and characterization of osteoclast-like cells and their progenitors in cultures of feline marrow mononuclear cells ₪ KENNETH. IBBOTSON, G. DAVID ROODMAN, LINDA M . McMANUS, and GREGORY R. MUNDY ₪ The Medical Service of the Audie L. Murphy Memorial Veterans Administration Hospital and Departments of Medicine and Pathology of the University of Texas Health Science Center, San Antonio, Texas 78284 www.indiandentalacademy.com
  4. 4. References ₪OSTEOCLAST FUNCTION AND ITS CONTROL ₪ MONE ZAIDI, MICHAEL PAZIANAS, VIJAI S. SHANKAR,BRIDGET E. BAX, CHRISTOPHER M. R. BAX, PETER J. R. BEVIS, CLIFFORD STEVENS*, CHRISTOPHER L.-H. HUANGt, DAVID R. BLAKE*, BALJIT S. MOONGA AND A. S. M. TOWHIDUL ALAM ₪ Department of Cellular and Molecular Sciences, Bone Research Unit, St George's Hospital Medical School, London SW17 ORE, ₪ Bone and Joint Research Unit, the London Hospital Medical School, London EC, ₪ Physiological Laboratory, University of Cambridge, Cambridge CB2 3EG ₪ (MANUSCRIPT RECEIVED 14 SEPTEMBER 1992, ACCEPTED 2 MARCH 1993) www.indiandentalacademy.com
  5. 5. Learning Objectives • At the end of the seminar the learner should be able to know about ₪ Osteoclast cells and its features ₪ Origin of Osteoclast cell ₪ Differentiating markers of osteoclast ₪ Mechanism of action of Osteoclast cell.www.indiandentalacademy.com
  6. 6. Osteoclast (OCL) ₪ Bone is a dynamic tissue that under goes constant remodelling through balancing bone formation and resorption. ₪Formation  Osteoblast cells ₪ Resorption  Osteoclast cells www.indiandentalacademy.com
  7. 7. Osteoclast (OCL) ₪ The osteoclast (OCL) is the primary bone resorbing cell. ₪ The osteoclast were first recognised in 1873 by Albert Kolliker as a multinuclear cell that digest bone. ₪ It is a highly specialized multinucleated cell whose primary function is to help in the control of calcium homeostasis. www.indiandentalacademy.com
  8. 8. Osteoclast (OCL) ₪ Increase in the degree of multinucleation enhance the capacity of OCL to resorb bone. ₪ More no. of nuclei  more bone resorption ₪ Less no. of nuclei  less bone resorption www.indiandentalacademy.com
  9. 9. Osteoclast (OCL) ₪ Compare to other bone cells and their respective precursors, osteoclast are much larger cells. ₪ And thus because of their size they can be easily identified under light microscope and often are seen in clusters. Identification Features : www.indiandentalacademy.com
  10. 10. Osteoclast (OCL) www.indiandentalacademy.com
  11. 11. Osteoclast (OCL) ₪ The osteoclast is an extremely large cell, ranging in size from 50 -100 µm in diameter. ₪ It is multinucleated and may contain 2-100 nuclei per cell, but usually contains between 10 and 20. Identification Features : www.indiandentalacademy.com
  12. 12. Osteoclast (OCL) ₪ Typically osteoclast are found against the surface of bone, occupying the hollow out depression known as HOWSHIP , s LACUNAE. ₪ These are the shallow trough which shows the activity and mobility of osteoclast cell during active resorption. Identification Features : www.indiandentalacademy.com
  13. 13. Osteoclast (OCL) ₪The osteoclast is located at ₪ Endosteal bone surfaces, ₪ Within the haversian system, ₪ And on periosteal surfaces beneath the periosteum. ₪Osteoclasts are most abundant at sites of active bone cell activity, such as metaphyses of growing bone. Identification Features : www.indiandentalacademy.com
  14. 14. Osteoclast (OCL) ₪ In electron microscopy osteoclasts have several unique ultrastructural features. ₪The characteristic ruffled border ₪Forms at the site where the osteoclast is attached to bone matrix ₪Is a convolution of plasma membrane with many long cytoplasmic processes. Identification Features : www.indiandentalacademy.com
  15. 15. Osteoclast (OCL) ₪The plasma membrane is closely apposed to the bone surface with an adjacent organelle free area, the clear zone, which is rich in actin like filaments ₪ Actin ₪ Vinculin ₪ Talin Identification Features : www.indiandentalacademy.com
  16. 16. ₪ A many of cell membrane infolding making up the ruffled border ₪ Sealing zone of osteoclast showing striation and granularity representing the concentration of contractile protein in this region. www.indiandentalacademy.com
  17. 17. Osteoclast (OCL) ₪This clear zone or sealing zone ₪Attaches the cells to the mineralized surface ₪Isolate the microenvironment between them and bone surface. Identification Features : www.indiandentalacademy.com
  18. 18. Osteoclast (OCL) ₪Lamina Limitans (electron dense interfacial matrix layer) often observed between the sealing zone and calcified tissue surface. Identification Features : ₪ Immunocytochemical preparation of osteopontin showing black dots of osteopontin on the bone surface at sealing zone. www.indiandentalacademy.com
  19. 19. Osteoclast (OCL) ₪This layer consist of RGD(arg - gly- aspartic acid ) containing molecules ₪Bone sialoprotein ₪Osteopontin ₪ That facilitate the osteoclast adhesion and formation of sealing zone via αvβ3 mediated mechanism. Identification Features : www.indiandentalacademy.com
  20. 20. Osteoclast (OCL) ₪Other ultrastructural characteristics of osteoclasts include ₪Large numbers of lysosomes, ₪Numerous pleomorphic mitochondria, ₪Extensive golgi complexes located around the nucleus Identification Features : www.indiandentalacademy.com
  21. 21. ₪Osteoclast possess numerous golgi complexes located near the nuclei and mitochondriawww.indiandentalacademy.com
  22. 22. Osteoclast (OCL) ₪ The cytoplasm is characterized by dense granules. ₪In addition have many prominent vesicles, especially in the region of the ruffled border. ₪The multiple nuclei are centrally located, but have a highly variable shape and contain one prominent nucleolus per nucleus. Identification Features : www.indiandentalacademy.com
  23. 23. Osteoclast (OCL) ₪ Cytochemically they are identified by possessing TRAP (tartrate resistantant acid phosphatase) with in the cytoplasmic vesicles and vacuoles. Identification Features : Photomicrograph showing the red staining associated with the presence of tartrate-resistant acid phosphatase (TRAP) in the cytoplasm of a multinucleated, culture-generated mouse osteoclast. Bar 5 -20 mm. www.indiandentalacademy.com
  24. 24. Osteoclast (OCL) ₪ A variety of studies have clearly indicated that the osteoclast is hematopoietic in origin and not bone derived. ₪ But still the origin of the osteoclast is a subject of intense investigation and controversy. origin: www.indiandentalacademy.com
  25. 25. Osteoclast (OCL) ₪ Walker et al. reported that transplantation of spleen and marrow cells into osteotropic mice results in removal of the excess bone present in these animals. origin: www.indiandentalacademy.com
  26. 26. Osteoclast (OCL) ₪ Marks has shown that transplantation of spleen cells into the osteopetrotic rat can cure the disease. ₪ In contrast to untreated rats whose osteoclasts lack ruffled borders, infusion of mononuclear spleen cells resulted in the formation of osteoclasts with ruffled borders. origin: www.indiandentalacademy.com
  27. 27. Osteoclast (OCL) ₪ All these studies are conclude that the osteoclast precursor is a mononuclear cell that is hematopoietic in origin. ₪ The osteoclast forms by fusion of mononuclear precursors rather than endomitosis. origin: www.indiandentalacademy.com
  28. 28. Osteoclast (OCL) ₪ Multinucleated osteoclast arise from the hematopoietic precursors of the monocyte /macrophage lineage ₪ Stromal cells in the marrow cavity and osteoblast modulate the differentiation of osteoclast. origin: www.indiandentalacademy.com
  29. 29. Osteoclast (OCL) ₪ Stromal cells and osteoblast modulate osteoclast differentiation via ₪Various molecules ₪Direct Cell to cell interaction origin: www.indiandentalacademy.com
  30. 30. Osteoclast (OCL) ₪ Stromal cells and oasteoblast expresses RANKL(receptor-activated nuclear factorκB) over their plasma membrane. ₪ RANKL binds to the RANK presented on the progenitors of osteoclast. origin: www.indiandentalacademy.com
  31. 31. Osteoclast (OCL) ₪ This attachment cause induction of signalling cascade leading to induction of ₪Osteoclast differentiation ₪Fusion of osteoclast precursors cell ₪Leading to the increased survival of activity of osteoclast origin: www.indiandentalacademy.com
  32. 32. Osteoclast (OCL) origin: www.indiandentalacademy.com
  33. 33. Osteoclast (OCL) origin: www.indiandentalacademy.com
  34. 34. Osteoclast (OCL) Differentiation markers ₪ Histochemical studies have shown that osteoclasts contain a Tartrate-resistant acid phosphatase (TRAP) activity that is found in ₪Lysosomes, ₪Golgi complexes, ₪Extracellular channels of the ruffled border, ₪ And the space between cells and bones. www.indiandentalacademy.com
  35. 35. Osteoclast (OCL) Differentiation markers ₪ Tartrate-resistant acid phosphatase appears to be a marker for osteoclasts in bone sections. ₪ Other acid hydrolases are present in osteoclasts, including ₪Leucine aminopeptidase, ₪β-d-galactosidase, ₪β-d-glucosidase. All are non specific for the osteoclast.www.indiandentalacademy.com
  36. 36. Osteoclast (OCL) Differentiation markers ₪ Calcitonin receptors have been suggested as a good maker for mammalian osteoclast differentiation. ₪ This differentiate osteoclast cells from other cells of the mononuclear phagocyte series. www.indiandentalacademy.com
  37. 37. Osteoclast (OCL) Differentiation markers ₪ Gay and co-workers have examined carbonic- anhydrase activity in chicken osteoclasts by electron microscopy. ₪ By using histochemical technique they found positive stains for carbonic-anhydrase in ₪Cytosol, ₪Golgi apparatus and other vesicles, ₪Plasma membrane and its Ruffled border, ₪Bone surface beneath the osteoclast. www.indiandentalacademy.com
  38. 38. Osteoclast (OCL) Differentiation markers ₪ Monoclonal antibody techniques have examined the surface phenotype of osteoclasts and identify unique differentiation markers for osteoclasts. ₪121F ₪ 23c6 and 13c2 (directed against the vitronectin receptor present on osteoclasts.) www.indiandentalacademy.com
  39. 39. Osteoclast (OCL) Differentiation markers ₪ Other markers for osteoclasts are ₪Matrix Metalloproteinase (MMP)-9, ₪Cathepsin K, ₪Carbonic Anhydrase II (CAII), ₪a3 Subunit Of The Vacuolar [H+]-ATPase (proton pump), ₪Chloride Channel 7 (ClC-7), www.indiandentalacademy.com
  40. 40. Osteoclast (OCL) Factors affecting differentiation of OCL ₪ Factors affecting the differentiation of osteoclasts are ₪ 1,25-Dihydroxyvitamin D3 ₪Parathyroid Hormone and Parathyroid Hormone- Related Peptide ₪GM-CSF ₪CSF-1 ₪Interleukin-3, -1, -6. ₪Transforming Growth Factor-α and -β www.indiandentalacademy.com
  41. 41. Osteoclast (OCL) Factors affecting differentiation of OCL ₪ Factors affecting the differentiation of osteoclasts are ₪Tumor Necrosis Factor-α and -β ₪Gamma Interferon ₪Calcitonin ₪Osteoinductive Factor ₪1,25D3 Lactone ₪Prostaglandins www.indiandentalacademy.com
  42. 42. Osteoclast (OCL) Mechanism of action of bone resorption : ₪ An osteoclast resorbs bone through the formation of a sealing zone of close adhesion between itself and the bone. ₪The adjacent plasma membrane is thrown into complex folds, the 'ruffled border', which consists of finger-like membrane projections. www.indiandentalacademy.com
  43. 43. Osteoclast (OCL) Mechanism of action of bone resorption : ₪ The 'ruffled border' and 'clear zone' constitute a system for bone resorption unique to the osteoclast. ₪ The expression of these structures generally depends upon the level of cellular activity. www.indiandentalacademy.com
  44. 44. Osteoclast (OCL) Mechanism of action of bone resorption : ₪ Thus, regulators of osteoclast activity rapidly increase or decrease the size of the 'ruffled border' and 'clear zone'. ₪Parathyroid hormone, ₪ 1,25-dihydroxy vitamin D3 ₪ Prostaglandin E2 ₪ Calcitonin www.indiandentalacademy.com
  45. 45. Osteoclast (OCL) Mechanism of action of bone resorption : ₪ Role of mineral ₪ Osteoclast activation requires contact with bone mineral. ₪ Osteoclasts can resorb calcified cartilage, enamel, dentine or cementum in addition to mineralized bone www.indiandentalacademy.com
  46. 46. Osteoclast (OCL) Mechanism of action of bone resorption : ₪ Role of mineral ₪ Uncalcified cartilage is not resorbed by osteoclasts ₪The osteoid or related unmineralized collagenous matrix could prevent effective osteoclastic activation. www.indiandentalacademy.com
  47. 47. Osteoclast (OCL) Mechanism of action of bone resorption : ₪ Role of mineral ₪ Osteoclasts, normally resting in the periosteal and endosteal mesenchyme, ₪ For its activation it must cross a cellular (osteoblast) barrier and possibly a second osteoid barrier to gain contact with mineral. www.indiandentalacademy.com
  48. 48. Osteoclast (OCL) Mechanism of action of bone resorption : ₪ Role of osteoblast ₪ The osteoblast is the primary target cell in hormonal activation of the osteoclast as it possess specific surface receptors for ₪parathyroid hormone ₪parathyroid hormone-related protein (PTHrP), ₪1,25-dihydroxyvitamin D3. www.indiandentalacademy.com
  49. 49. Osteoclast (OCL) ₪ It is proven that ₪cell contact ₪soluble products released by the osteoblast is required for osteoblast-induced osteoclastic stimulation to take place. ₪ The nature of this soluble factor is still unclear 0.5-1 kDa to over 25 kDa. Mechanism of action of bone resorption : www.indiandentalacademy.com
  50. 50. Osteoclast (OCL) ₪ Osteoclast adhesion to bone ₪ In order to resorb bone, the osteoclast must effectively adhere to the bone surface. ₪ It contain characterized adhesive receptor, osteoclast functional antigen (OFA), Mechanism of action of bone resorption : www.indiandentalacademy.com
  51. 51. Osteoclast (OCL) ₪ Osteoclast adhesion to bone ₪ OFA comprises of ₪140 kDa δ-chain (immunologically cross-reacts with the vitronectin δ-chain receptor) ₪85 kDa β-chain related to the platelet gp III-a molecule (β3, integrin). Mechanism of action of bone resorption : www.indiandentalacademy.com
  52. 52. Osteoclast (OCL) ₪ Osteoclast adhesion to bone ₪ OFA expression on bone cells is restricted to osteoclasts and preosteoclasts. Mechanism of action of bone resorption : www.indiandentalacademy.com
  53. 53. Osteoclast (OCL) ₪ Osteoclast adhesion to bone ₪ Integrins, such as the vitronectin receptor, are transmembrane proteins that act as linkers between ₪Extracellular ligands and cytoskeletal components, such as vinculin, talin, δ-actinin and F-actin Mechanism of action of bone resorption : www.indiandentalacademy.com
  54. 54. Osteoclast (OCL) ₪ Osteoclast adhesion to bone ₪ The ligands in bone containing the RGD (arginine- glycine-aspartic acid) tripeptide recognition sequence that could potentially bind to OFA. ₪ These include ₪Osteopontin, ₪Thrombospondin ₪Bone sialoprotein. Mechanism of action of bone resorption : www.indiandentalacademy.com
  55. 55. Osteoclast (OCL) ₪ Mineral dissolution ₪ Osteoclasts secrete acid ,lactate and citrate and thus maintain a low pH (values of 4.5 ) in the resorptive hemivacuole. ₪ Active H+ion secretion from the osteoclast was demonstrated by using Acridine Orange as an indicator of extracellular acidification. Mechanism of action of bone resorption : www.indiandentalacademy.com
  56. 56. Osteoclast (OCL) ₪ Mineral dissolution ₪ Acridine Orange fluorescence was increased by ₪Parathyroid hormone ₪Prostaglandin E2, ₪ And decreased by calcitonin Mechanism of action of bone resorption : www.indiandentalacademy.com
  57. 57. Osteoclast (OCL) ₪ Mineral dissolution ₪ It has been reported that the apical membrane of the 'ruffled border' contain proton pumps that may be involved in the acidification of the extracellular resorptive hemivacuole. Mechanism of action of bone resorption : www.indiandentalacademy.com
  58. 58. Osteoclast (OCL) ₪ Mineral dissolution ₪ Two types of proton pumps have been implicated in the acidification process. ₪ V-type or Electrogenic vacuolar type proton pump (H+-ATPase){predominantly involved} ₪P-type plasma membrane pump (H+- K+-ATPase). Mechanism of action of bone resorption : www.indiandentalacademy.com
  59. 59. Osteoclast (OCL) ₪ Role of Carbonic anhydrases ₪ The proton pump in the osteoclast is supplied by protons generated by the action of carbonic anhydrase isoenzyme II ₪ Immunocytochemical techniques have localized the enzyme to the 'ruffled border' of active osteoclasts. Mechanism of action of bone resorption : www.indiandentalacademy.com
  60. 60. Osteoclast (OCL) ₪ Role of Carbonic anhydrases ₪ It catalyses the conversion of CO2 (generated from cell respiration) to HCO3 - and H+. CO2  HCO3 - + H+ ₪ Shows that bone resorption is critically dependent upon CO2 concentration. Mechanism of action of bone resorption : www.indiandentalacademy.com
  61. 61. Osteoclast (OCL) ₪ Role of Carbonic anhydrases ₪ Meanwhile, basolateral exchange of HCO3-ions for Cl- by anion exchanger provides Cl- ions required for the intense acidification occurring in the resorption lacuna. Mechanism of action of bone resorption : www.indiandentalacademy.com
  62. 62. Osteoclast (OCL) ₪ Matrix degradation ₪ Osteoclastic lysosomal enzymes are destined for secretion and transported in a specific direction via primary lysosomes and are released into the resorptive hemivacuole at the 'ruffled border'. Mechanism of action of bone resorption : www.indiandentalacademy.com
  63. 63. Osteoclast (OCL) ₪ Matrix degradation ₪ Lysosomal hydrolases, exhibiting optimal activity at an acid pH, resorb the organic matrix of bone. ₪ Collagen can be digested by a number of lysosomal cysteine-proteinases in the acid environment. Mechanism of action of bone resorption : www.indiandentalacademy.com
  64. 64. Osteoclast (OCL) ₪ Matrix degradation ₪ The three types of collagenolytic cysteine-proteinases found in bone ₪cathepsins L, ₪cathepsins B, ₪cathepsins N. Mechanism of action of bone resorption : www.indiandentalacademy.com
  65. 65. Osteoclast (OCL) ₪ Matrix degradation ₪ Increased levels of acid phosphatase activity in plasma occur during physiological and pathological states of increased bone remodelling. ₪ This isoenzyme is not inhibited by dextro-rotatory tartaric acid, (a competitive inhibitor of prostatic and lysosomal acid phosphatases) Mechanism of action of bone resorption : www.indiandentalacademy.com
  66. 66. Osteoclast (OCL) www.indiandentalacademy.com
  67. 67. www.indiandentalacademy.com
  68. 68. www.indiandentalacademy.com
  69. 69. www.indiandentalacademy.com