本文選題：干細胞 + 間質(zhì)干細胞�。� 參考：《中國組織工程研究》2017年05期

【摘要】：背景:骨髓間充質(zhì)干細胞對肝癌增殖能力和轉(zhuǎn)移能力的影響是研究的熱點,可通過基因工程技術(shù)對肝癌細胞進行改造,使其降低生物活性因子的表達,從而尋找合適的干預(yù)靶點,提高骨髓間充質(zhì)干細胞的干預(yù)效能。目的:經(jīng)轉(zhuǎn)化生長因子β1、骨橋蛋白基因沉默的高轉(zhuǎn)移潛能肝癌細胞(MHCC97-H)與骨髓間充質(zhì)干細胞共培養(yǎng),觀察MHCC97-H細胞侵襲能力的變化,并通過動物模型熒光成像觀察骨髓間充質(zhì)干細胞對MHCC97-H肝癌組織動物模型的干預(yù)作用。方法:(1)將MHCC97-H細胞細胞分為4組,MHCC97-H設(shè)為空白對照組,MHCC97-H基因沉默轉(zhuǎn)染陰性對照為陰性對照組,MHCC97-H-轉(zhuǎn)化生長因子β1為轉(zhuǎn)化生長因子β1基因沉默組,MHCC97-H-骨橋蛋白為骨橋蛋白基因沉默組;(2)Transwells法進行各組細胞與骨髓間充質(zhì)干細胞共培養(yǎng)實驗,48 h后結(jié)晶紫染色,隨機取3個視野,計數(shù);(3)結(jié)合有紅色熒光蛋白基因的各組MHCC97-H細胞株,分別自裸鼠右側(cè)腋下接種復(fù)制皮下移植瘤模型。腫瘤體積到50 mm3開始瘤內(nèi)注射骨髓間充質(zhì)干細胞,4周后進行熒光強度分析;肝癌組織冰凍切片二脒基苯基吲哚細胞核染色后行熒光顯微鏡觀察。結(jié)果與結(jié)論:(1)細胞計數(shù)結(jié)果表明,骨髓間充質(zhì)干細胞使經(jīng)基因沉默后的MHCC97-H細胞遷移數(shù)量明顯減少;結(jié)晶紫染色病理圖片顯示,經(jīng)基因修飾的MHCC97-H細胞遷移能力明顯降低;(2)體內(nèi)熒光成像結(jié)果顯示,骨橋蛋白基因沉默組腫瘤體積明顯縮小,熒光亮度低于其他組別,定量結(jié)果顯示骨橋蛋白基因沉默組吸光度值明顯降低,表明骨髓間充質(zhì)干細胞對經(jīng)骨橋蛋白干擾的MHCC97-H高轉(zhuǎn)移潛能肝癌動物模型干預(yù)效能最佳;(3)病理切片熒光觀察結(jié)果顯示,骨髓間充質(zhì)干細胞主要分布在腫瘤壞死區(qū)附近,骨橋蛋白基因沉默組熒光表達較轉(zhuǎn)化生長因子β1基因沉默組和空白對照組多,表明MHCC97-H細胞經(jīng)骨橋蛋白基因沉默后骨髓間充質(zhì)干細胞在腫瘤中的分布增多;(4)結(jié)果提示,通過抑制骨橋蛋白和轉(zhuǎn)化生長因子β1兩種因子的表達可以降低肝癌細胞侵襲能力,經(jīng)骨橋蛋白沉默可能更有利于基于骨髓間充質(zhì)干細胞的生物治療。
[Abstract]:Background: the effect of bone marrow mesenchymal stem cells (BMSCs) on the proliferation and metastasis of hepatocellular carcinoma (HCC) is a hot topic. In order to find appropriate intervention targets, improve the intervention effectiveness of bone marrow mesenchymal stem cells. Objective: to observe the change of invasion ability of MHCC97-H cells by co-culture of MHCC97-Hand bone marrow mesenchymal stem cells (BMSCs) by transforming growth factor 尾 1, osteopontin gene silencing (TGF- 尾 1) and osteopontin gene silencing (TGF- 尾 1). The effects of bone marrow mesenchymal stem cells (BMSCs) on MHCC97-H liver cancer tissue were observed by fluorescence imaging. Methods MHCC97-H cells were divided into four groups: MHCC97-H as blank control group, MHCC97-H gene silencing as negative control group, MHCC97-H- H- transforming growth factor 尾 _ 1 as transforming growth factor 尾 _ 1 gene silencing group, MHCC97-H- osteopontin as osteopontin group. In the silencing group, the cells were co-cultured with bone marrow mesenchymal stem cells (BMSCs) for 48 h and then stained with crystal violet. MHCC97-H cell lines with red fluorescent protein gene were randomly selected from 3 fields, and subaxillary inoculation of MHCC97-H cell lines with red fluorescent protein gene was carried out to establish subcutaneous transplanted tumor model from right axillary of nude mice. The fluorescence intensity of bone marrow mesenchymal stem cells was analyzed 4 weeks after intratumoral injection of bone marrow mesenchymal stem cells (BMSCs) from 50 mm3 to 50 mm3, and the staining of diamidine phenyl indole nuclei in frozen sections of liver cancer was observed by fluorescence microscope. Results and conclusion the cell count showed that bone marrow mesenchymal stem cells significantly decreased the migration of MHCC97-H cells after gene silencing. The results of fluorescence imaging showed that the tumor volume of osteopontin gene silencing group was significantly smaller and the fluorescence brightness was lower than that of other groups. The quantitative results showed that the absorbance value of osteopontin gene silencing group was significantly decreased, which indicated that bone marrow mesenchymal stem cells had the best intervention effect on MHCC97-H high metastatic potential liver cancer model with osteopontin interference. Bone marrow mesenchymal stem cells (BMSCs) mainly distributed near the necrotic area of tumor. The fluorescence expression of osteopontin gene silencing group was more than that of transforming growth factor 尾 1 gene silencing group and blank control group. The results indicated that the distribution of bone marrow mesenchymal stem cells (BMSCs) increased after osteopontin gene silencing in MHCC97-H cells. The results suggested that the expression of osteopontin and transforming growth factor 尾 1 could decrease the invasion ability of hepatocellular carcinoma cells by inhibiting the expression of osteopontin and transforming growth factor 尾 1. Osteopontin silencing may be more beneficial to the biotherapy based on bone marrow mesenchymal stem cells.
【作者單位】： 解放軍總醫(yī)院第一附屬醫(yī)院放射科;解放軍第九五醫(yī)院放射科;解放軍南京總醫(yī)院醫(yī)學(xué)影像科;
【基金】：國家自然科學(xué)基金資助項目(81271607) 南京軍區(qū)醫(yī)藥衛(wèi)生重點資助項目(11Z035) 中國博士后基金(2015M572810)~~
【分類號】：R735.7

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