Effects Of Inflammatory Response On The Expression Of HIF-1α Induced By Hypoxia

1. Pre-treatment of Lipopolysaccharide on the expression of HIF-1α andVEGF induced by CoCl2in Raw264.7macrophagesObjective To explore the effects of hypoxia environments simulated by cobaltchloride(CoCl2) on the expression of hypoxia-inducible factor-1α(HIF-1α) and vascularendothelial growth factor(VEGF) in murine macrophage-Raw264.7pre-treated by usingLPS and the potential mechanism involved.Methods Divided the Raw264.7cells into four groups as follows:(1) controlgroup;(2) LPS(1mg/L) group;(3) CoCl2(100μmol/L) group;(4) LPS pre-treatedgroup(LPS-Pre,CoCl2was added when cells were pretreated by LPS8hours before).First,At different time point, LPS or CoCl2induced cellular proliferation were observedby use of CCK8test;and then,the mRNA expression of HIF-1α and VEGF weredetected by RT-PCR.And final, the protein expression of HIF-1α and VEGF wereanalyzed by the methods of Western blot and immunocytochemistry.Results(1)The CCK8data showed that the proliferation of Raw264.7tend to be stableafter treated by LPS and CoCl28h.(2)The results of RT-PCR showed that the mRNA expression of HIF-1α and VEGFincreased in CoCl2, LPS and LPS-pretreated groups when compared with control group(P<0.05), especially, the expression in LPS-pretreated group were higher than CoCl2group(P<0.05).(3)The results of Western Blot and immunocytochemistry indicated that the proteinexpression of HIF-1α and VEGF increased in CoCl2, LPS and LPS-pretreated groupswhen compared with control group(P<0.05), especially, the expression inLPS-pretreated group were higher than CoCl2group(P<0.05). Conclusion Pre-treatment with LPS can up-regulate the protein of HIF-1α andVEGF at the post-transcriptional level in macrophages under hypoxia environment.2. Pre-treatment of Lipopolysaccharide on the expression of HIF-1α in mouseunder hypoxia environmentObjective To observe the mRNA and protein expression of HIF-1α in pneumoniamouse pre-treated by using LPS when they were under hypoxia condition and thepotential mechanism involved.Methods Male mice were randomly divided to four groups as follows: controlgroup，LPS group，hypoxia group,and LPS-pretreated group。Mice of hypoxia groupwere inhaled intranasally with50ul of N.S, while LPS group and LPS-pretreated groupwere administrated by50ul of LPS (1g/L).2.5hours before all the mice were sacrificedat the third day, mice in hypoxia and LPS-pretreated groups experienced hypoxiaenvironment. The lungs were separated and first weighed to analyze lung coefficientand then embedded and sliced to5um sections, which were then stained by HE todetermine the severity of inflammation. The mRNA expression of HIF-1α was detectedby RT-PCR and the protein expression of HIF-1α was analyzed by the methods ofWestern blot.Results(1)The data showed that lungs coefficient were elevated in pneumonia mouseinduced by LPS and the significant inflammation can be clearly observed in the lungswith the recruitment of neutrophils, macrophages and lymphocytes.(2) The results of RT-PCR showed that the mRNA expression of HIF-1α increasedsignificently in LPS-pretreated group when compared with control group (P<0.05),especially, the expression in LPS-pretreated group were higher than LPS and hypoxiagroups (P<0.05).(3) The results of Western Blot indicated that the protein expression of HIF-1αincreased in LPS and LPS-pretreated groups when compared with control group(P<0.05), especially, the expression in LPS-pretreated group were higher than LPS andhypoxia group (P<0.05). Conclusion The mRNA and protein expressions of HIF-1α under hypoxiaenvironment can be up-regulated in the inflammatory lung tissues after the mice werepre-treated by LPS。...