Inhibitory Effects of Ligustrazine Plus Emodin on Expression of VEGF-related Factors in Ascites Carcinoma Walker-256 Cells / 中国实验方剂学杂志

ABSTRACT

Objective:To investigate the effects of ligustrazine combined with emodin on angiogenesis of ascites carcinoma Walker-256 cells by observing their inhibition against nuclear factor-<italic>κ</italic>B （NF-<italic>κ</italic>B）， hypoxia-inducible factor-1<italic>α</italic> （HIF-1<italic>α</italic>）， and vascular endothelial growth factor-C （VEGF-C） in HIF signaling pathway. Method:Fifty SD rats were randomly divided into sham operation group， model group， ligustrazine group， emodin group and ligustrazine combined with emodin group. Following the in situ injection of rat ascites carcinoma Walker-256 cells into the liver of normal rats， they were grouped and administered with ligustrazine （10 mg·kg^-1）， emodin （10 mg·kg^-1）， and ligustrazine （10 mg·kg^-1） plus emodin （10 mg·kg^-1） for seven days. Afterwards， the tumor-inoculated liver tissue was sampled from the experimental group and prepared into pathological sections for investigating tumor cell survival and VEGF expression. The <italic>in vitro</italic> hypoxia and hypoglycemia model （oxygen-glucose deprivation model）， hypoxia model， and hypoglycemia model of Walker-256 cells were constructed respectively. In the ligustrazine group， emodin group， and ligustrazine combined with emodin group， three consecutive concentrations that did not affect the proliferation of Walker-256 cells were selected for investigation. The drugs were administered before modeling， and the model treatment lasted for 4 h. The levels of HIF-1<italic>α</italic>， VEGF-C， and NF-<italic>κ</italic>B in the cell culture supernatant of each group were tested. Result:After the rat liver was inoculated with Walker-256 cells， the total liver mass was significantly increased（<italic>P</italic><0.05）， higher than that in the ligustrazine group， the emodin group， or the ligustrazine combined with emodin group（<italic>P</italic><0.05）. Histopathological examination showed that the response of VEGF expression in the liver tissue of each administration group was lower than that of the model group. At the cellular level， the levels of HIF-1<italic>α</italic>， VEGF-C， and NF-<italic>κ</italic>B in oxygen-glucose deprivation model of the ligustrazine group and the ligustrazine combined with emodin group were significantly reduced（<italic>P</italic><0.05）， exhibiting a certain dose-dependent response， followed by the reduction in the hypoxia model. The levels of HIF-1<italic>α</italic> and NF-<italic>κ</italic>B in the oxygen-glucose deprivation model and the hypoglycemia model of the emodin（1×10^-2，1×10^-3 mol·L^-1） group and the ligustrazine combined with emodin（1×10^-2，1×10^-3 mol·L^-1） group were significantly reduced， but there was no significant change in VEGF-C level of the hypoxia model of all the administration groups. Conclusion:Ligustrazine or emodin alone or their combination inhibits the abnormal increase in the weight of rat liver after inoculation with Walker-256 cells and the expression of VEGF in the liver tissue. Ligustrazine and emodin inhibit the protein expression of NF-<italic>κ</italic>B and HIF-1<italic>α</italic>， thereby reducing the gene and protein expression of metastasis-related target VEGF-A activated by HIF-1<italic>α</italic> transcription， restricting tumor cell neovascularization， and inhibiting the invasion and spread of ascites carcinoma cells. Among them， ligustrazine has the most significant effect against hypoxia. Glucose interferes with the effect of ligustrazine. The combination of ligustrazine with emodin is conducive to diminishing the intervention of glucose and stabilizing the inhibition against tumor cells.