| ObjectiveIn this study we detected the expression of HSP90 protein in stromal tumor tissues and corresponding normal tissues, analyzed the correlation between the expression and clinicopathological parameters of HSP90 proteins in glioma patients, followed by detection of HSP90 influence on glioma cells in cell proliferation and apoptosis, we also studied the mechanism of action of curcumin in the regulation of HSP90 targeted inhibition of glioma cell proliferation and invasion, it aims to provide a theoretical basis for the treatment of glioma and new ideas, but also to find new HSP90 inhibitors foundation in glioma.Methods(1) We collected 50 cases of glioma tissue samples, including 28 male patients, female 22 cases. All patients enrolled in the age range between 23 to 78 years, with an average age of (37.55 ± 11.2) years; all glioma patients, I grade glioma 6 cases, II grade 8 cases, III grade 17 cases, IV grade 19 cases; at the same time set up by CT and MRI and other imaging tests confirmed no tumor diseases tissue samples of normal brain due to trauma or intracranial pressure and cerebral hemorrhage 10 patients served as controls. Western blot were used to detect the expression of the above samples, and analysised the expression of HSP90 protein with clinical and pathological parameters correlation.(2) We transfected HSP90 siRNA treatment T98G glioma cells to inhibit the expression of HSP90, the expression of HSP90 mRNA was detected by quantitative PCR after transfection; cell proliferation was detected by MTT assay after transfection; apoptotic was detected by flow cytometry after transfection.(3) The inhibitory effect of anacardic acid on Hsp90 was assessed with in vitro ATPase inhibition assay and ATP-sepharose binding assay. MTT assay was used to detect the growth inhibition induced by Curcumin in T98G cells. Transwell assays were used to evaluate T98G cell invasion. Western blotting was performed to assess the effect of Curcumin in triggering the degradation of MMP-9 and Hsp90.Result(1) HSP90 protein expression in glioma tissues was significantly higher than the expression (P<0.01) in normal brain tissue, and with the increase of its pathological expression level rise; software analysis using Spearman glioma patients correlation between clinical and pathological grading and HSP90 expression, the result is:the correlation coefficient HSP90 expression of r= 0.713 (P<0.001), showed that glioma cancer tissues PHSP90 protein expression was positively correlated with pathological grades, namely high high expression levels of the patient group, a low level of cases in the low expression group.(2) With time, after HSP90 siRNA T98G cell proliferation effect gradually decreased. When the effect of 3,4,5,6 days, siHSP90 group T98G cell proliferation compared with control cells are decreased, which reduces the amplitude of the first day 4,5,6 significant difference was statistically significant (P<0.05); control group, apoptosis rate was (8.35 ± 1.27)%, siHSP90 group apoptosis rate (17.88 ± 0.13)%, compared to the difference between the two groups was statistically significant (P <0.01).(3) Curcumin is able to inhibit the ATPase activity of recombinant protein HSP90, HSP90 ATPase activity of curcumin inhibition IC50 value of 83.3 ?/mol; curcumin can inhibit HSP90 protein binding and ATP-agarose gel strains, and this combined inhibition with increasing curcumin concentration increased; T98G cell proliferation increased capacity with curcumin concentration decreased, that the higher the concentration of curcumin to inhibit the proliferation of T98G cells more obvious dose-dependent manner; invasion of T98G cells with increasing curcumin concentration decreased; the expression of MMP-9 protein with increasing curcumin concentration decreased in a dose-dependent manner, while the expression of HSP90 protein did not change significantly.ConclusionCurcumin can significantly inhibit the proliferation and invasion of T98G cells, the mechanism of which may involve the inhibition of Hsp90 ATPse activity and down-regulation of MMP-9 expression. |
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