• TRAP(+) multi nucleated bone resorbing cell, derived from fusion of TRAP(-) monocyte precursors (hematopoetic lineage)
• TRAP = Tartrate Resistant Acid Phosphatase
• Related to Foreign Body Giant Cells - they both surround/wall off material, but osteoclasts are specialized to resorb bone
• Definitive proof of the lineage: Infantile Osteopetrosis is treated with bone marrow transplantation, providing monocyte lineage precursors which became mature osteoclasts
• Various factors involved in its sequential differentiation; Mutation of some of the factors can block osteoclast formation
• Resorption Lacunae: feature that resembles a secondary lysosome between the cell and the bone
• where resorption takes place – contains a sealing zone and a ruffled border
• Sealing Zone: area of cellular attachment to bone that forms the resorption space (resorption lacunae)
• also referred to as Clear zone due to its clear appearance in transmission electron microscopy as intracellular organelles are excluded because packed full with dense cytoskeleton
• cytoskeleton attaches to bone via integrins (transmemb proteins that bind to extracellular bone matrix molec)
• binding can be disrupted by peptides that contain the integrin recognition sequence (RGD)
• Ruffled Border: involuted cellular membrane within sealing zone that secretes protons and lysosomal enzymes into the resorption lacunae
• Protons are pumped into resorption lacunae by ATPase pump
• reduce to pH 5-6 which dissolves bone mineral
• Carbonic Anhydrase Type II generates protons from CO₂ + H₂O « HCO₃^- + H⁺
• Removed by Cl/HCO₃ exchanger to maintain cytoplasmic pH; hydrogen transported to lacuna with Cl
• Defective CA II Þ Osteopetrosis with renal tubular acidosis. RBC unaffected because it also has CA I
• integrins with the RGD sequence trigger the acid release at the sealing zone
• Lysosomal enzymes are directed into the lacunae in addition to lysosomes via mannose-6-P tag
• TRAP is an osteoclast-specific lysosomal enzyme that can be used to identify osteoclasts

Osteoblasts – make bone via hydroxyapatite

• Related to fibroblast: they both secrete collagen, but osteoblasts are specialized to make bone
• Derived from the mesenchymal lineage
• multipotent mesenchymal precursors Þ Preosteoblasts (committed step) Þ mature osteoblasts Þ osteocytes
• Mature Osteoblasts exist in 2 inter-convertible forms:
• Active osteoblasts (produce the bone matrix Þ fill in the holes)
• Lining Cells (quiescent, flattened cells that line inactive bone surfaces (periosteum, etc))
• Osteocytes – osteoblasts that have become entrapped in mineral
• Bone Matrix:
• Consists of:
• (1) Type I collagen (mostly)
• (2) Proteoglycans: these are smaller than those in cartilage and do not aggregate
• (3) Non-collagenous proteins (NCP): some identified but most are unidentified
• Osteocalcin: most osteoblast specific, but is also produced by megakeryocytes
• Each NCP is produced at specific developmental stages for specific functions
• i.e., regulation of cell attachment, spreading and activity; collagen fibril formation and mineralization
• Mediates Mineralization:
• Geometry of the mineral lattice confers structural properties to bone
• Collagen Type I acts as a scaffold for the geometric arrangement of these minerals
• The acidic groups of collagen attract Ca²⁺ which then attracts PO₄^- etc.
• In osteogenisis imperfecta, the Type I collagen gene is mutated, therefore the geometric arrangement is abnormal and bones are fragile rather than strong - Often mistaken for child abuse
• Alkaline phosphatase functions to make PO₄^- available by cleaving it from proteins in local milieu
• some of these proteins are inhibitory to mineralization when PO₄^- is bound to them
• therefore alkaline phosphatase also functions to promote mineralization by removing inhibition
• osteomalacia from reduced mineralization in hypophosphatasia is due to mutations in alkaline phosphatase gene
• Many of these proteins and molecule can be markers for resorption and formation, thus can indicate disease:
• Formation: Serum Levels of: osteocalcin, alkaline phosphatase, procollagen extension peptide (type 1)
• Resorption: Plasma: TRAP; Urine (breakdown products): Ca²⁺, hydroxyproline, hydroxylysine, glycoside

• Modes of Action
• Paracrine – cells release cytokine to other cells within the same proximity.
• Autocrine – cell’s cytokines work on itself.
• Matricrine – cytokine is in ECM, gets released when ECM breaks down.
• Juxtacrine – cytokine is on surface of cell, responding cell is in contact with producing cell.
• Cytokines work in Networks
• Interactions: cytokines may regulate production/activity of other cytokines.
• Pleiotropic: cytokines may be multifunctional.
• Redundant: cytokines may have activities similar to other cytokines.
• Synergistic: cytokines may enhance each other’s activity.

• Agents that directly inhibit osteoclasts
• Ionic regulation – ions down-regulate OC activity by high concentrations of calcium or phosphate, and high pH (less protons present to acidify sealing zone).
• Calcitonin – also down-regulates OC activity; activates both cAMP/PKA and PKC signal transduction
• By Osteoblasts (OB)
• Agents that stimulate resorption (PTH, Vit D, TGF-b , etc) act via OBs!!! i.e. these molecules first interact with OBs, prompting them to release cytokines which affect OCs.
• Osteoclast Differentiating Factor (ODF), also known as RANK ligand, is a juxtacrine cytokine produced by OBs which prompts OC differentiation. Binding happens via RANK receptor.
• M-CSF – another cytokine produced by OBs, has same effect as ODF.
• Mechanisms by which OBs increase OC activity/formation:
• Exposing mineral surface by retraction or by secreting proteases. This broken down surface releases ions and cytokines that were trapped there (Matricrine action).
• Cytokine secretion (autocrine/paracrine): IL-6 and M-CSF are involved.
• Cell surface molecules (juxtacrine): Adhesion molecules, Cell-surface cytokines (M-CSF).
• ECM molecules (matricrine): Collagen, proteoglycans, matrix-associated cytokines (M-CSF), Bone matrix proteins (osteocalcin, osteopontin).
• Bisphosphonates (-nate) are drugs that inhibit resorption. They are stabilized pyrophosphate analogues that block the ability of OB cytokines to stimulate OC activity/formation.
• Other sources of cytokines
• Source: Immune system, other cell types.
• Examples:
• Infection in bone – excessive release of cytokines can attract OCs Þ destroy bone.
• Multiple Myeloma – same (release local IL-1, IL-6, TNF-b )
• Humoral Hypercalcemia of Malignancy – PTH-related protein acts like PTH to degrade bone.
• Estrogen – preserves bone mass (receptors on both OC and OB)
• Duel effects: keeps PTH low, reduces cytokines made by OBs/Monocytes.
• While women get menopausal osteoporosis (Ý resorption), men get senile osteoporosis (ß formation).
• androgens in men have similar effects as estrogen in women
• Clinical Correlation: bone around orthopaedic implants gets resorbed because of cytokine release by Mf s.

• OB proliferation, production, and signal transduction are not clearly correlated with changes in bone formation.
• Factors thought to influence bone formation:
• Physical factors: Mechanical, Electrical, Ultrasound.
• Chemical factors: Fluoride.
• Hormonal factors: PTH, Insulin, Glucocorticoids, Vitamin D.
• Growth Factors: see below.
• Bone Morphogenetic Proteins (BMPs) – in the TGF-b family, effects on many cells, induce ectopic endochondral ossification.
• Coupling of Resorption and Formation:
• As OCs break down the bone cytokines that were trapped in the matrix get released (matricrine) and activate OBs.
• But how do the OBs know where to act? Some lysosomal enzymes released by OCs have mannose-6-phosphate sugar residues on them. These enzymes now coat the newly destroyed surface. The receptors on the OB cells mistake these residues for IGF molecules, bind them, and proceed with their action.