Ano letivo 2012-2013 Biologia celular e molecular II

1. Ano letivo 2012-2013 Biologia celular e molecular II Cellular and molecular mechanisms in Osteogenesisimperfecta Luís Baptista, turma 2 Tânia Alves, turma 1 Tiago Barbosa, turma 2 Tiago Capela, turma 2

2. OsteogenesisImperfecta • Osteogenesisimperfecta (OI) is a congenital bone disorder. • Causedbymutationsaffectingtype I collagen.

3. Collagen Themolecule Fibrousproteinsfoundinallmulticellularanimals Triple-strandedhelicalstructure Repeating sequence of three amino acids glycine-X-Y (X and Y are often proline and hydroxyproline) 28 types of collagen identified and described 90% of the collagen in the body is of type I Main types found in connective tissue: types I, II, III, V and VI Collagen fibrils. These collagen fibrils are present in the joint capsule tissue that surrounds the knee.

4. Collagen formation

5. What genes are related to osteogenesisimperfecta? Mutations in the COL1A1, COL1A2, CRTAP, and LEPRE1 genes, as well as others.

6. COL1A1 and COL1A2 genes The COL1A1 gene is located on the long (q) arm of chromosome 17 at position 21.33. The COL1A2 gene is located on the long (q) arm of chromosome 7 at position 22.1.

7. CRTAP and LEPRE1 genes The CRTAP gene is located on the short (p) arm of chromosome 3 at position 22.3. The LEPRE1 gene is located on the short (p) arm of chromosome 1 at position 34.1.

8. How do peopleinheritOsteogenesisimperfecta? • Most cases of cases have an autosomal dominant pattern of inheritance; • Sporadic mutations; • Less commonly, osteogenesisimperfecta has an autosomal recessive pattern of inheritance.

9. Molecular defects OI type I • Over 90% ofpatientshavemutationsthatreducethesynthesisof pro-α1 chains to aboutonehalf; • Mutationsthatreducethesynthesisofpro-α2 chainsproduceslightly more severephenotypes;

10. OI types II, III and IV • Mutationsthatproducestructurally abnormal pro-αchainsthathavecompromisedassemblyor abnormal foldingofthe triple helix;

11. “Procollagensuicide” • Intracellularefects One abnormal chaininteractswithtwo normal chains Foldingisprevented Accumulationofunfoldedchains in the ER Enzimaticdegradation Unfolded protein response (UPR) is estimulated

12. Intracellularefects

13. Glycine’sreplacement • Intracellularefects Replacementofglycineby a bulkieraminoacid Production of poorly formed and unstable triple helices Disruption to helix folding Increasing of posttranslational hydroxylation and glycosylation of lysines

14. Intracellularefects

15. N-propeptide’s persistence • Extracellularefects Abnormal pro-α chains prevent normal processing of the N-propeptides N-propeptides persist Incorrectfibrilassembly

16. Extracellularefects

17. Collagencontaininghelixmutation can forminsolubleaggregatesin the ER that are degradedbytheautophagosome-endosomesystem;

18. Glycinesubstitutions in theN-terminalregiontend to producemilderphenotypes; • Raresubstitutionsofchargedaminoacidsor a branchedaminoacid in X- or Y- positions are lethal;

19. What determines the patients’ phenotype? • Hypothesis: gradient of phenotypic severity

20. Symptoms

21. Type I (Mild)

22. Type II (Perinatal Lethal)

23. Type III (Progressive Deforming)

24. Type IV

25. Diagnosis • Diagnosis is primarily based on clinical evidence; • Themostfrequentexamsused to identifyingthedisease are: • X rays • Laboratorytesting • DualEnergy X-ray Absorptiometry (DXA) • BoneBiopsy

26. Treatments Currently prescribed • orthopaedic surgery ; • scoliosis management ; • rehabilitation, including water therapy and physical activity ; • Develop healthy lifestyle diet and exercise habits ;

27. Treatments Currentlybeinginvestigated: Treatments that are ineffective and are no longer prescribed: PharmacologicTherapy • vitamin C • sodium fluoride • magnesium • anabolic steroids • calcitonin Bisphosphonates Teriparatide GrowthHormone Inhibition of receptor activator of nuclear factor-kappaB ligand (RANKL) CellularandGeneticTherapy Bonemarrowtransplantation Gene therapy

28. Bone formation and bone resorption

29. Pharmacologictherapy Biphosphonates Synthetic analogues of pyrophosphate, like Pamidronate, Risedronate, Alendronate and zoledronic acid ; Inhibit osteoclast-mediated bone resorption on the endosteal surface of bone by:

30. Teriparatide (Forteo) A synthetic form of the natural human Parathyroid Hormone (PTH) ; Intermittent administration of these drug would: • Increase bone mineral density and bone strength • Reduce the chance of getting a fracture In same cases, mainly in chilhood, there is an increase in the incidence of osteosarcoma, a malignant bone tumor

31. Growth Hormone(GH) The effects of GH on bone cells are mediated through the functional GH receptors (GHRs) on osteoblast cells ; But, it also stimulates the insulin-like growth factor type 1 (IGF-1) production in liver and bone Osteoblast formation is increased and osteoblast apoptosis is inhibited Increase in bone turnover and formation

32. Inhibition of receptor activator of nuclear factor-kappaB ligand (RANKL) It’s a member of the tumor necrosis factor (TNF) cytokine family ; It’s a ligand for osteoprotegerin(OPG) ; Key factor for osteoclast differentiation and activation ; The first FDA-approved RANKL inhibitor was Denosumab

33. CellularandGeneticTherapy Bonemarrowtransplantation contains both hematopoietic stem cells and mesenchymal stem cells (MSCs) latter precursors of osteoblasts

34. CellularandGeneticTherapy Gene Therapy

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