A Study Relating To The Mechanism About Hypoix Induced HIF-1? Regulating LDHA In Osteosarcoma

Objective: to explore the regulate relationship between HIF-1? and LDHA in osteosarcoma under hypoxia condition and whether the LDHA can influence the function of osteosarcoma cells.Method:1.Detected and analyzed LDHA in 18 cases of osteosarcoma samples by immunohistochemistry.2.Inspect the expression status about LDHA in osteosarcoma under hypoxia condition or normoxia condition by RT-PCR.Inspect the expression status about HIF-1? and LDHA under hypoxia condition or normoxia condition by western-blot.3.Constructed plasmid which contain of the promoter of LDHA,by given the hypoxia condition to induce endogenous HIF-1? in MG63 cells and explored the regulate relationship between HIF-1? and LDHA,and determined the specific binding site;and induced exogenous HIF-1? by expression plasmid to further study this relationship.4.Constructed plasmid to knockdown the expression of HIF-1? in MG63 cells,compare the expression of LDHA under hypoxia and normoxia condition.5.Constructed plasmid to knockdown the expression of LDHA,and by the way of scratch assay to inspect the migration of MG63 cells which under hypoxia condition.6.Knock down the expression of LDHA in MG63 cells and inspected which proliferation by MTT under hypoxia.Result : 1.We found that LDHA is always overexpression in osteosarcoma,in our study there are 4 cases were weak expression,8cases were moderate expression and 6 cases were intensity expression,and the positive ratio about 80%;2.By given different oxygen condition,we found that hypoxia can induce LDHA overexpression in osteosarcoma;3.Under hypoxia condition,by luciferase assay we found that hypoxia induce LDHA expression depend on HIF-1? binding to the HER-D which in the promoter of LDHA;4.Using RNA interfere technique to knockdown the expression of HIF-1? will suppression LDHA expression;5.After knockdown LDHA,scratch assay found that the migration ability of MG63 cells was been suppression under hypoxia condition;6.The proliferation activity of MG63 cell was been suppression a little after knockdown LDHA,but this difference is not obvious.Conclusion: our study found that osteosarcoma always in heavy hypoxia,hypoxia condition lead HIF-1? overexpression,and HIF-1? will binding to HER-D and induce LDHA overexpression.Meanwhile,LDHA can promote osteosarcoma cells migration,but have little influence on the cells proliferation.