Theories of mineralization

1st BDS class

1. Theories of Mineralization Dr. P. Poornima Dept. Of Oral Pathology & Microbiology

2. Mineralization Process of deposition of insoluble calcium salts in a tissue. Mechanism of calcification: Normally, the concentration of Ca & Phosphate ions are insufficient to form crystals. When certain local factors raises the concentration of these ions, then initiation of crystals starts.

3. The mineral component of all hard tissues of our body are cheifly Hydroxyapatite crystals. Mineralization was thought to be based on precipitation dynamics, with initially the only ions needed being calcium and phosphate. Though the ECF is supersaturated with calcium and phosphate, yet the mineralization is not a common phenomenon. It may be due to: Presence of inhibitors of mineralization Formation of unstable or insufficient amount of crystals to proceed

4. Thus for precipitation to occur, Interplay of various factors in the osteoid matrix may contribute to conditions that may favor mineralization. They are: Presence of increased concentration of Extracellular Ca & P ions which facilitate mineralization – Homogenous nucleation Presence of nucleating substance – Nucleating substances like collagen which will act as a template over which mineralization occurs even with Ca & P ion concentration of ECF is not elevated – Heterogenous nucleation Absence of Inhibitors of Mineralization.

5. Theories of Mineralization There are 3 main theories of mineralization. They are:

6. Alkaline phosphatase theory Proposed by Robinson in 1923 This enzyme present in the organic matrix of calcifying tissues ↓ Hrdrolyses organic phosphates → Inorganic phosphate ↓ ↑ P ion concentration – boosting effect & combines with Ca ions ↓ Amorphous Calcium phosphate ↓ HA crystal formation It acts as a marker for active tissue mineralization.

7. Supporting When slices of cartilage from animals affected by rickets are placed in Alk . Phosphatase , there is evidence of mineralization More Alkaline Phosphatase in calcifying cartilage than non calcifying cartilage Contrasting Alkaline Phosphatase also seen in other tissues that do not calcify. Inhibitors of some enzymes other than Alk . Phosphatase can inhibit mineralization The organic phosphates present in tissue fluid alone are insufficient to produce sufficient number of inorganic p ions.

8. Nucleation theory Neumann and Neumann in 1953 put forward this theory. Also called as seeding theory or collagen template theory Some nucleating substances act as a template upon which crystals can be laid down. Collagen acts as most important seeding substance. Lysine and Hydroxylysine – Phosphate binding site. Carboxyl ions – Calcium binding site

9. Gaps between the collagen fibril ↓ First Proteoglycans attaches to the collagen ↓ Enzymatic degradation of proteoglycans releases Ca which attaches to collagen fibril ↓ Later phosphoproteins will come and attaches to the collagen and by the action of Alkaline phosphatase , phosphate is released and attaches to Ca to form apatite crystal in the gap zone of collagen.

10. Supporting Electron microscopic studies supported parallel arrangement of Hydroxyapatite crystals and collagen fibers Contrasting Enamel is a highly mineralized tissue – No collagen at all. So, some other mechanism is playing a role.

11. So, all the tissues which contains collagen should mineralize. But this is not happening in soft tissues. WHY? In soft tissues, the charged site may be covered by some ground substance material which prevent the attachment of ions to initiate mineralization. This substance is called - Crystal poison – Pyrophosphate .

12. Collagen shows intrafibrillar pores through which Ca & P ions should pass to reach nucleating sites located inside the fibril. Diameter of P ion – 0.4 nm Gap between tropocollagen molecules in Calcifiable tissues – 0.6 nm Gap between tropocollagen molecules in Non- Calcifiable tissues – 0.3 nm

13. Matrix Vesicle theory Rich in phospholipids - Phosphatidyl serine - high affinity for Ca ions. Vesicles also contain annexins . Annexins in the vesicles form a calcium channel, thus incorporating the ion within the matrix vesicles. Their membranes furnish binding sites for the nucleation of hydroxyapatite crystals.

14. Matrix Vesicle theory Matrix vesicles are small membrane bound structures 25 to 250 nm in diameter, lying free in the matrix, where calcification is going to occur. Are rounded outgrowths of cell membrane that bud from osteoblasts , chondrocytes and odontoblasts . There are 2 types of matrix vesicles

15. Matrix Vesicle theory Rich in phospholipids - Phosphatidyl serine - high affinity for Ca ions. Vesicles also contain annexins . Annexins in the vesicles form a calcium channel, thus incorporating the ion within the matrix vesicles. Their membranes furnish binding sites for the nucleation of hydroxyapatite crystals.

16. Once the HA crystal is formed ↓ After attaining certain thickness, they will be released into ECM by action of Phospholipases which disintegrates membrane of Matrix vesicles ↓ Progression of mineralization occurs as spheres or globular masses

17. Inhibitors of Calcification Pyrophosphate Diphosphonates Adenosine triphosphate Citrate, magnesium, and proteins like albumin also acts as inhibitors

18. Summary All the 3 mechanisms are involved in mineralization. Alkaline phosphatase helps in providing more phosphate ions and in removing crystal poisons or inhibitors of calcification. Collagen acts as a seed – helps in intrafibrillar calcification or linear calcification Matrix vesicles – helps in extrafibrillar calcification or globular calcification and

19. THANK YOU