胞外基质材料木糖葡聚糖对HepG2细胞的黏附及相关功能的影响  

Effect of Xyloglucan on Attachment and Function of HepG2 Cells as a New Extracellular Matrix

作　　者：陆胜利[1] 祁超[2] 

机构地区：[1]安庆医药高等专科学校药学系,安徽安庆246052 [2]华中师范大学生命科学学院,湖北武汉430079

出　　处：《武汉大学学报（医学版）》2017年第4期525-529,559,共6页Medical Journal of Wuhan University

基　　金：国家自然科学基金资助项目(编号:20772040;31272645;31672284);教育部中央高校自主项目(编号:CCNU14F01006)

摘　　要：目的:研究胞外基质(ECM)材料木糖葡聚糖(XG)对人肝癌细胞系HepG2细胞在细胞培养板上的贴壁性及对微囊化HepG2细胞的清蛋白合成速率、NH_4^+清除速率、细胞增殖速率和间隙连接蛋白(Cx32)、细胞黏附分子(E-cadherin)表达的影响。方法:将HepG2细胞接种于不同浓度XG(0,0.5,1,4,6mg/ml)包被的细胞培养板上,孵育4h后收集贴壁细胞,BCA法测贴壁细胞总蛋白量进而测定细胞贴壁率;利用自制装置制备包被HepG细胞的海藻酸-多聚赖氨酸-海藻酸(APA)微囊并在培养微囊第6,10,14天以Human Serum Albumin Elisa Kit、Ammonia Assay Kit分别测定培养液中人血清白蛋白(HSA)和处理溶液中NH_4^+的浓度,检测微囊化HepG2细胞的清蛋白合成速率和NH_4^+清除速率;微囊培养1,2,3d后裂解微囊收集HepG2细胞并以RT-PCR方法检测细胞内Cx32和E-cadherin的表达情况。结果:HepG2细胞在XG包被的细胞培养板上的贴壁率随XG浓度增大呈递增趋势,同时微囊内细胞数随XG浓度增大、培养时间延长而增加,说明XG可促进细胞/基质间相互作用,促进细胞生长增殖;微囊化HepG2细胞的清蛋白合成速率与NH_4^+清除速率在XG浓度为1 mg/ml时达到最大且较空白组高,说明XG对细胞合成及代谢功能有一定促进作用;加入XG的微囊内细胞表达Cx32、E-cadherin的时间较空白组提前,说明XG的存在促进了HepG2细胞肝脏特有功能的维持和体现。结论:XG对微囊化HepG2细胞的增殖及肝脏特有功能的体现都具有一定促进作用。Objective： To test the effects of xyloglucan （XG）, used as a new synthetic extracellular ma trix （ECM） for adhesion of HepG2 cells and the albumin secretion of micro capsule HepG2 cells, ammonia elimination, cell proliferation and Cx32, E-cadherin gene expression of encapsulated HepG2 cells. Methods： HepG2 cells were seeded on different concentrations of XG coated surface and incubated for 4 h, then the adherent cells were collected and quantified by BCA method for determining the adhesioin ratio. Micro capsules were prepared by self-made device and cultured for 6 d, 10 d or 14 d, then the Human Serum Albumin Elisa kit and Ammonia Assay Kit were applied to determine the sythetic rate of HSA and elimination rate of NH＋ , respectively. The expression of Cx32 and E-cadherin genes in capsulated HepG2 cells at separate days was detected by reverse transcriptase-polymerase chain reaction （RT-PCR）. Results： The adhesion ratio of HepG2 ceils onto coated surface gave a rising tendency and the cell number increased when the concentration of xyloglucan increased, indicating the promotion effects of xyloglucan on cell/matrix interactions and cell proliferation. The rates of albumin secretion and ammonia elimination were highest at t mg/ml of xyloglucan, and higher than that of blank groups, suggesting the promotion effects of xyloglucan on liver-specific functions of encapsulated HepG2 cells. The Cx32 and E-cadherin genes in alginate （AL）/XG capsules were more rapidly expressed, than in AL ones, respectively. Conclusion： Multicellular spheroid formation of HepG2 cells can enhance the liver-specific functions in the three-dimensional space in the presence of XG as a new synthetic ECM.

关 键 词：肝组织工程 木糖葡聚糖 海藻酸 微囊化 

分 类 号：R318.08[医药卫生—生物医学工程]