川楝素对人胃癌细胞MKN-45中CTPS细胞蛇形成的影响及其机制*

目的:本研究旨在对川楝素(TSN)与胃癌细胞MKN-45中CTPS细胞蛇形成的联系进行初步探究。方法:以人胃癌细胞MKN-45为实验材料,设置7个处理组分别为:0、20、40、60、80、100、120 nmol/L TSN。每组3次重复,分别作用24 h、48 h、72 h,利用CCK-8法检测川楝素对MKN-45细胞增殖抑制作用,使用免疫荧光检测之后再通过激光共聚焦显微镜观察细胞内CTPS细胞蛇形态,qRT-PCR检测川楝素对MYC基因表达的影响。另外设置2个处理组为1 mmol/L DON和1 mmol/L MPA,每组3次重复,作用6 h然后采用免疫荧光检测细胞蛇形态。结果:免疫荧光结果显示,分别利用1 mmol/L DON和1 mmol/L MPA 处理MKN-45细胞后CTPS形成丝状的细胞蛇结构,意味着该细胞具有形成CTPS细胞蛇的能力;川楝素处理组的细胞增殖率明显低于0 nmol/L TSN组(P＜ 0.01);免疫荧光结果显示80 nmol/L的川楝素作用72 h时MKN-45细胞中CTPS细胞蛇形成数量最多;qRT-PCR检测结果显示,80 nmol/L川楝素作用24 h细胞内MYC表达明显降低(P＜0.05),48 h后细胞内MYC的表达量明显增多(P＜0.01)随后表达降低。结论:川楝素可能通过调节MYC的表达影响细胞内细胞蛇的组装。

Effects of Toosendanin on the formation of CTPS cytoophidium in human gastric cancer cell MKN-45 and its mechanism

Objective: To investigate the relationship between toosendanin(TSN) and CTP synthase(CTPS) cytoophidium formation in gastric cancer MKN-45 cells. Methods: In this study, the experimental material is MKN-45 human gastric cancer cells. It contains 7 treatment groups of 0, 20, 40, 60, 80, 100, and 120 nmol/L TSN. Each group was treated in triplex privately for 24、48 and 72 hours. Cell counting kit-8 (CCK8) was used to detect the inhibitory effect of TSN on the proliferation of MKN-45 cells. After immunofluorescence detection, the morphology of CTPS cells was observed by a laser confocal microscope. The effect of TSN on MYC gene expression was detected by qRT-PCR. In addition, it contains 2 treatment groups of 1 mmol/L DON and 1 mmol/L MPA, each group was treated in triplex privately for 6 hours and then the cytoophidium morphology was detected by immunofluorescence. Results: The results of immunofluorescence showed that CTPS formed a filamentous cytoophidium structure after treating MKN-45 cells with 1 mmol/L DON and 1 mmol/L MPA, which means that the cells have the ability to form CTPS cytoophidium; The cell proliferation rate of TSN treatment group was significantly lower than that of 0 nmol / L TSN group (P＜0.01); Immunofluorescence results showed that CTPS cytoophidium was the most abundant in MKN-45 cells after treated with 80 nmol/L TSN for 72 h. The results of qRT-PCR showed that MYC expression in cells was significantly decreased after treated with 80 nmol/L TSN for 24 h (P＜0.05), and MYC expression was significantly increased after 48 h (P＜0.01), and then decreased. Conclusion: Toosendanin may affect intracellular cytoophidium assembling by regulating the expression of MYC.