Inhibitory Effect Of Inositol Phosphate And Inositol On Proliferation Of Tumor Cells

Objective:To observe the inhibitory effect of both IP6 and inositol individually and in combination on different cancer cells, and examine whether the combination of IP6 and inositol will enhance the inhibitory effect on xenograft tumor.Method: we cultivated thehuman cell lines Hep G2, HT-29, 7901, MCF-7 and A549 in 96 well plates with the intervention of both IP6 and inositol individually and in combination(the volume ratio 1:1) at four concentrations: 0.5mmol/L、1mmol/L、2mmol/L、4mmol/L for 48 h.Then calculating the inhibition ratio and IC50(half maximal inhibitory concentration) using optical density(OD) from MTT assay.and testing the inhibitory effect of the combination of IP6 and Ins on two cancer cells which were more sensitive to IP6 and Ins. What’s more, forty male BALB nude mice were chosen to establish the model of transplanted tumor under aseptic condition with 0.2ml cell suspension(2×106 cells) by hypodermic injection. When it comes the obvious node, the forty nude mice were randomly divided into four groups named control group, IP6 group, Ins group and IP+Ins grouop according to the volume of node. Every mouse in each group will receive the respectively administration of normal saline, IP6, Ins, IP6+Ins by intraperitoneal injection, and the weighing up every four days were also included. After 20 days, all nude mice will be put to death, the simple of tumor, liver, kidney, blood will be conserved for detection. The HE staining will be used to observe the condition of tumor; and we will use the immunohistochemical method to detect the protein expression of PCNA, Cyclin D 1, CDK4; and the expression of Rb, E2F1 were detected by Real-time PCR.Result:Compared with the control group, results from MTT assay showed IP6 and/or inositol inhibited growth of all kinds of cancer cells at all concentrations significantly(F=8.253~15.292,t=6.871~8.537,P<0.05). When three drugs were used on HT-29、7901、MCF-7、A549 cells respectively, we confirmed that the inhibitory effect was no significant difference on four kinds of cancer cells(F=1.195~2.408,t=0.978~2.133,P>0.05). But when the concentration ups to 4mmol/L, IP6+Ins shows higher inhibition rateto Hep G2(t=7.462,P<0.05). the data also showed the IC50 was different between five tested cancer cells, the IC50 of Hpe G2 and HT-29 was much lower than others. Compared with the control group,IP6, Ins, IP6+Ins showed no effect on nude mice’s metabolic activity(P>0.05). After measure the weight and volume of tumors, data showed that the tumors from experimental groups were average smaller than control groups’ with the smallest tumors from IP6+Ins(P>0.05). The HE staining showed that the control tumor cells arranged closely, have visible fission phase, and no obvious apoptosis phenomenon. However, the three experimental groups have different degrees of apoptosis morphological features, such as severe endoplasmic reticulum expansion, cytoplasm edge set, etc. Immunohistochemical results show that compared with control group, the expression of PCNA protein was lower, and IP6+Ins shared the minimum expression level, but there was no significant difference between the four groups(P>0.05). Experiment found that the experimental group could reduce the expression of Cyclin D1 and CDK4 protein and IP6, IP6+Ins group share significant difference compared with control group(P < 0.05). The inhibitory effect of Ins to the expression of Cyclin D1 and CDK4 protein is not obvious(P > 0.05). the date from Rt-PCR showed that Compared with control group, IP6 and Ins group share significantly increase of the expression of Rb m RNA(P < 0.05); however, when it came to E2F1, IP6 and IP6+Ins showed no obvious effect on the expression of E2F1 m RNA, Ins could reduce the expression of the E2F1 m RNA(P < 0.05).Conclusion: It was different between IP6 and/or inositol on inhibitory effect of different cancer cells, and the combination of IP6 with inositol failed to enhance the inhibitory effect to HT-29 、 7901 、 MCF-7 、 A549 cancer cells. But when the concentration ups to 4mmol/L, the IP6 combined with Inositol can obviously enhance the inhibitory effect to Hep G2. IP6 and IP6+Ins showed no significant inhibitory effect on nude mice tumors, but IP6 and the combination of IP6 with Ins can obviously reduce the expression of Cyclin D 1,CDK4 protein, and increase the expression of Rb m RNA.