Experimental Study Of Inducing-differentiation Effect Of Valproate And Combined With Chemotherapeutic Drug Against Human Glioma

[Objective]Valproate(VPA) is a well established clinical drug in the long-term therapy of epilepsy. Recently , it has become evident that VPA can induce kinds of tumor cells to differentiate. However, the therapeutic effects of VPA alone and combined with chemotherapeutic drug on brain glioma in vivo has not been evaluated, this study was designed to investigate the associated effects of VPA and chemotherapeutic drug Nimustine(ACNU) against glioma. Furthermore, the Tob gene and the HDAC1 gene were selected to detect their expression by RT-RCR .thereinto, the Tob gene was one of the 28 glioma inducing-differentiation relative genes according to the former cDNA microarray experimental taken by our laboratory and HDAC1 was the focus of inducing-differentiation research at present, this part of experiment could be useful to elucidate the mechanism of VPA and provide foundation for further gene therapy. [Method] (1) lower-differentiated human brain glioma cell line SHG-44 were xenografted subcutaneously on NC nude mice to create the solid tumor model. (2)VPA was applied alone and combined with ACNU to tumor-bearing mice(VPA 400mg/ kg, qdX40d;ACNU 30mg/kg, q4dX4), control group was applied with 0.9%NS(25ml/kg, qdX40d). (3) tumor size was measured by vernier caliper .pathological change was observed by HE dyeing .different phase of cell cycle of tumor cell proliferation and expression of GFAP were detected by FCM , the gene expression of HDAC1 and Tob was determined by RT-PCR. [Result](1) the tumor was always accelerate to growth in the control group, even canker was occurred on the surface of skin, the growth of ACNU group' stumor was reduced in the early stage, which was no difference with VPA + ACNU group, but once ACNU was withdrawn, the tumor resume rapid growth, the growth of ACNU group' s tumor between the control group and the ACNU group was moderate without accelerate stage. The inducing-differentiation effects of VPA was demonstrated by lower growth rate of tumor and the low cell heteromorphism under the microscope ,tumor cell proliferation arrested in G0/G1 phase by FCM, the expression level of Glial fibrillary acidic protein (GFAP) was also upregulated. (2)if combined with chemotherapeutic drug ACNU, the inhibition effect was more obvious and more persistent than VPA or ACNU alone. (3) HDAC1 genes was high expressed in control tumor group, but decreased in VPA group and further decreased in VPA combined with ACNU group, On the contrary, expressions of TOB in the therapy group increased compared with the control group. [Conclusion] (1) VPA can definitely inhibit the cell proliferation of transplanted human glioma and induce tumor cell toward benign , otherwise, the VPA' s effective drug concentration was in the safe range and VPA wan no obvious toxic effects during administration, so will it have a good prospect in the future. (2) VPA combined with ACNU can obviously enhance ACNU' s anti-glioma effects and reduce it' s toxic side-effect and ACNU can promote the inducing-differentiation effect of VPA. but which can not eliminate the tumor completely, so the next step research need to aim at the molecular target of glioma. (3)PT-PCR revealed that the expression of HDAC1 enchanced and the expression of Tob reduced , which proved HDAC1 and Tob were the target of VPA and controlled the genesis and evolvement of glioma. If knockou or interfere the genes respectively, it will improve the therapeutic effects radically.