[關(guān)鍵詞]
[摘要]
目的 研究胃復(fù)春膠囊對(duì)人胃癌細(xì)胞株MKN-49P的作用,為胃復(fù)春膠囊在抗消化道腫瘤方面的應(yīng)用提供新的依據(jù)。方法 將SD大鼠隨機(jī)分為2組,即對(duì)照組、胃復(fù)春膠囊組。胃復(fù)春膠囊組取膠囊內(nèi)容物,以生理鹽水混懸,按300 g/(kg·d)劑量ig給藥,對(duì)照組大鼠ig 2 mL生理鹽水,共5 d。末次給藥后2 h,在無(wú)菌條件下,ip 10%水合氯醛(350 mg/kg)麻醉,腹主動(dòng)脈取血,分離血清,經(jīng)滅活處理后,置于—20℃冰箱保存?zhèn)溆?。用不同質(zhì)量濃度(0、2.5、5、10、20、30、50、80、100 μg/mL)的胃復(fù)春膠囊含藥血清處理MKN-49P細(xì)胞,采用MTT法檢測(cè)細(xì)胞活力。加入各質(zhì)量濃度(25、50、100 μg/mL)胃復(fù)春膠囊含藥血清,流式細(xì)胞術(shù)Annexin V/PI雙染色檢測(cè)細(xì)胞凋亡情況,Hoechst染色法檢測(cè)細(xì)胞形態(tài),細(xì)胞劃痕實(shí)驗(yàn)測(cè)定細(xì)胞的遷移能力,Western blotting法檢測(cè)信號(hào)傳導(dǎo)與轉(zhuǎn)錄激活因子(STAT)蛋白、核轉(zhuǎn)錄因子-κB(NF-κB)信號(hào)通路及與細(xì)胞凋亡相關(guān)的蛋白表達(dá)情況。結(jié)果 胃復(fù)春膠囊含藥血清能夠劑量和時(shí)間相關(guān)地降低MKN-49P細(xì)胞的活力,并可誘導(dǎo)MKN-49P細(xì)胞凋亡。Hoechst染色顯示細(xì)胞膜通透性增強(qiáng);細(xì)胞劃痕實(shí)驗(yàn)結(jié)果顯示,與對(duì)照組(0 μg/mL)相比,不同濃度胃復(fù)春膠囊含藥血清(25、50、100 μg/mL)可濃度相關(guān)性地抑制MKN-49P細(xì)胞的遷移。MKN-49P細(xì)胞經(jīng)過(guò)25、50、100 μg/mL胃復(fù)春膠囊含藥血清分別處理24 h后,MKN-49P細(xì)胞中STAT3的磷酸化減弱,NF-κB信號(hào)通路相關(guān)的蛋白表達(dá)量下調(diào),抗凋亡蛋白表達(dá)量下降而促凋亡蛋白表達(dá)量增加。結(jié)論 胃復(fù)春膠囊含藥血清能抑制胃癌細(xì)胞的增殖、誘導(dǎo)細(xì)胞凋亡,并顯著降低細(xì)胞遷移能力,其作用機(jī)制推測(cè)與NF-κB信號(hào)通路有關(guān)。
[Key word]
[Abstract]
Objective To study the effect of Weifuchun Capsules on human gastric cancer cell line MKN-49P, and to provide a new basis for the application of Weifuchun Capsules in anti-digestive tract tumor. Methods SD rats were randomly divided into two groups:control group and Weifuchun Capsules group. Weifuchun Capsules group was given the Weifuchun Capsules contents, suspended with normal saline, ig at 300 g/(kg∙d) dose, and control group was ig 2 mL normal saline, for a total of 5 d. 2 h after the last administration, under aseptic conditions, 10% chloral hydrate (350 mg/kg) was injected intraperitoneally for anesthesia. Blood was taken from abdominal aorta and the serum was separated. After inactivation treatment, the serum was stored in a refrigerator at −20℃ for future use. MKN-49P cells were treated with Weifuchun Capsules containing serum at different concentrations (0, 2.5, 5, 10, 20, 30, 50, 80, and 100 µg/mL). Cell viability was detected by MTT assay. Add the drug containing serum of Weifuchun Capsules at different concentrations (25, 50, 100 µg/mL), apoptosis was detected by Annexin V/PI double staining by flow cytometry. Cell morphology was detected by Hoechst staining. The cell migration ability was measured by cell scratch assay. The expression of signal transduction and transcription activator (STAT) protein, nuclear transcription factor-κB (NF-κB) signaling pathway and apoptosis-related proteins were detected by Western blotting. Results The serum containing Weifuchun Capsules could decrease the viability of MKN-49P cells, and induce apoptosis of MKN-49P cells in a dose and time dependent manner. Hoechst staining showed enhanced cell membrane permeability. Compared with control group (0 μg/mL), different concentrations of Weifuchun Capsules containing serum (25, 50, and 100 μg/mL) could inhibit the migration of MKN-49P cells in concentration-dependent manner. After MKN-49P cells were treated with 25, 50, and 100 μg/mL serum containing Weifuchun Capsules for 24 h, the phosphorylation of STAT3 were decreased. The protein expression of NF-κB signaling pathway was down-regulated. The expression of anti-apoptotic protein decreased while that of pro-apoptotic protein increased. Conclusion The drug-containing serum of Weifuchun Capsules can inhibit the proliferation and induce apoptosis of gastric cancer cells, and significantly reduce the cell migration ability. The mechanism of action is speculated to be related to NF-κB signaling pathway.
[中圖分類(lèi)號(hào)]
R979.1
[基金項(xiàng)目]