| Glioma is the most common primary tumors of the central nervous system,Malignant glioma are the most common and aggressive brain tumors in intracranial malignant tumors. Despite current advances in multimodality therapies, such as surgery, radiotherapy, and chemotherapy, the outcome for patients with high-grade glioma remains fatal. Although intensive basic and clinical studies and the development of various therapeutic modalities, the average survival of patients with malignant glioma post-diagnosis remains less than 15 months. As an oral alkylating antineoplastic agent in recent twenty years, Temozolomide has become a major drug of glioma. However, due to drug resistance in tumors, the effective rate of temozolomide of glioma is less than 45%. Isoliquiritigenin whose anti-tumor properties have multiple targets, multiple mechanisms, which has significant effect in inhibiting tumor cell growth and encouraging tumor cells apoptosis.In this paper, we try to extend anti-tumor effect and its mechanism of isoliquiritigenin and provide theoretical basis of isoliquiritigenin in treating human gliomas.ObjectiveWe investigated the inhabiting effect of isoliquiritigenin on the cellular proliferation of the human glioma cell SHG44 and its anti-tumor mechanism. Then,we discussed influences of isoliquiritigenin on the mechanism of inhibiting glioma cells proliferation of temozolomide, which might provide some basis for the effectiveness of isoliquiritigenin as a traditional chinese medicine in treatment of glioma.Methods1.Study on the mechanism which isoliquiritigenin uses to inhibit the human glioma cell SHG44 cellular proliferation: with different concentrations of isoliquiritigeninin, under different time, we use CCK-8 method to understand the effect of isoliquiritigeninin on the cellular proliferation of glioma cell line SHG44; we also utilize flow cytometric method Annexin V-IP markers to examine the situation of cell apoptosis 48 h after using different concentrations of isoliquiritigenin(0、20、40、60、80μmol/L); we observe the structure change of cells by transmission electron microscopy; we use immunofluorescence to observe autophagy and apoptosis of cells;we detect Bcl-2, LC3 B, Caspase-3 trend of cells by western blot.2.Study on the inhibition mechanism of isoliquiritigenin in combination with temozolomide: in vitro experiments observation, we observe the combination effect of isoliquiritigenin, temozolomide isoliquiritigenin and, Temozolomide in inhibating SHG44 glioma cellar proliferation; we use CCK-8 method to find the inhibition rate of each group; we also use flow cytometric method Annexin V-IP markers to examine the situation of cell apoptosis of each group; we observe the structure change of cells under the effect of isoliquiritigenin and temozolomide by transmission electron microscopy; we use immunofluorescence and Western blot to observe autophagy and apoptosis of cells of each group.Results1. Study on the mechanism which isoliquiritigenin uses to inhibit human glioma cell SHG44 proliferation. The inhibition effects depend on time and dose: half inhibition appears after 72 h.we also use flow cytometric method Annexin V-IP markers to examine the situation of cell apoptosis of each group. Varied concentrations of isoliquiritigenin(0、20、40、60、80μmol/L) have different early apoptosis rates on glioma cells after 48 h, which are(1.54±0.85)%、(6.72±2.81)%、(11.82±3.21)%、(16.61±3.79)% and(26.15±3.82)%. Apoptotic bodies, autophagy and autophagy lysosome are seen with transmission electron microscopy.Under the fluorescence microscope, SHG44 cells show apoptosis autophagy and fluorescent particles. In the Western blot trial, 48 h after applying isoliquiritigenin on SHG44 cells, Caspase-3, Beclin1 and LC3- II / I ratio of protein expression levels increase,while the protein expression level of Bcl-2 decrease.2. Study on the inhibition mechanism of isoliquiritigenin in combination with temozolomide. The inhibition mechanism on glioma cells is dose-dependent: the combined group show more significant effect than either isoliquiritigenin or temozolomide group(P<0.05). With flow cytometric method, the combined effect that temozolomide(200 μmol/L) and isoliquiritigenin(80 μmol/L) have on glioma cells after 48 h is that early apoptosis rate significantly increases(38.31±4.97)%,while early apoptosis rates of either temozolomide or isoliquiritigenin is just(12.62±3.12)% and(24.86±4.01)%. For the combination group, a large number of autophagic and apoptotic bodies are seen under transmission electron microscopy. In Western blot test, compared with either single group, for the combined group,Caspase-3, Beclin1 and LC3- II / I ratio of protein expression levels increase, while the protein expression level of Bcl-2 decrease.Conclusions1.Isoliquiritigenin can effectively inhibit proliferation of human glioma cell SHG44, and induce glioma cell apoptosis and autophagy. Meanwhile, Isoliquiritigenin can inhibit the expression of Bcl-2, increase the expression of apoptosis related molecules caspase-3 and autophagy related molecule LC3- II.2. Isoliquiritigenin can enhance the growth-suppressing effects of temozolomide on human glioma cell SHG44, which is mostly to do with the combined effects of them in inducing apoptosis and autophagy of glioma cells. |
PDF Full Text Request