CAJ | 학술논문

目的应用小鼠肝星状细胞(HSC)与小鼠成体肝脏干细胞(AHPC)体外间接共培养模型,观察HSC旁分泌途径对AHPC增殖及代谢功能的影响.方法分别建立HSC及AHPC细胞分离及培养模型.进一步选择Transwell 6孔板,建立小鼠HSC与AHPC间接共培养模型.在共培养24 h和96 h后,计算AHPC克隆数及克隆形成率;共培养14 d后,行5-溴脱氧尿嘧啶核苷(5-BrdU)增殖实验,计算AHPC细胞克隆内BrdU阳性的细胞数,评估细胞增殖能力.同时,收集细胞上清液行酶联免疫吸附试验(ELISA)检测细胞代谢产物以评估细胞的代谢能力.通过以上方法比较HSC组和空白对照组中AHPC在克隆形成率、细胞增殖及代谢功能上的差异.结果 HSC组AHPC的克隆形成率为(26.88±3.31)％,对照组为(20.11±2.34)％(P＜0.05).BrdU增殖实验提示,对照组内BrdU/白蛋白双阳性细胞为(69.50±18.81)/高倍视野,较HSC组增多61.29％ (P＜0.01).代谢功能方面,HSC组AHPC细胞上清液中白蛋白(Albumin)和尿素(Urea)的浓度分别为(14.17 ±3.14) mg/(L·d)和(34.83 ±11.41) mg/(L·d),比对照组分别提高72.17％和66.89％(P＜0.05).结论一定数量的HSC在体外培养时能提高AHPC克隆形成率,促进细胞增殖分裂,细胞代谢功能亦相应增强.
목적 응용소서간성상세포(HSC)여소서성체간장간세포(AHPC)체외간접공배양모형,관찰HSC방분비도경대AHPC증식급대사공능적영향.방법 분별건립HSC급AHPC세포분리급배양모형.진일보선택Transwell 6공판,건립소서HSC여AHPC간접공배양모형.재공배양24 h화96 h후,계산AHPC극륭수급극륭형성솔;공배양14 d후,행5-추탈양뇨밀정핵감(5-BrdU)증식실험,계산AHPC세포극륭내BrdU양성적세포수,평고세포증식능력.동시,수집세포상청액행매련면역흡부시험(ELISA)검측세포대사산물이평고세포적대사능력.통과이상방법비교HSC조화공백대조조중AHPC재극륭형성솔、세포증식급대사공능상적차이.결과 HSC조AHPC적극륭형성솔위(26.88±3.31)％,대조조위(20.11±2.34)％(P＜0.05).BrdU증식실험제시,대조조내BrdU/백단백쌍양성세포위(69.50±18.81)/고배시야,교HSC조증다61.29％ (P＜0.01).대사공능방면,HSC조AHPC세포상청액중백단백(Albumin)화뇨소(Urea)적농도분별위(14.17 ±3.14) mg/(L·d)화(34.83 ±11.41) mg/(L·d),비대조조분별제고72.17％화66.89％(P＜0.05).결론 일정수량적HSC재체외배양시능제고AHPC극륭형성솔,촉진세포증식분렬,세포대사공능역상응증강.
Objective Appling co-culture model to observe the effect of hepatic stellate cells (HSC) paracrine pathway on the proliferation and metabolism of adult hepatic progentior cell (AHPC).Methods Establishing HSC and AHPC indirect co-culture model with Transwell six-hole plate.At 24 h and 96 h after co-culture,AHPC clone formation rate was analyzed; 14 days after co-culture,BrdU proliferation assay was done to assess cell proliferation ability.In addition,the cell supernatant was collected for enzyme linked immunosorbent assay (ELISA) to detection of cell metabolism.Results In HSC group,the clone rate of AHPC was averaged (26.88 ± 3.31) ％,compared with the control group was (20.11 ±2.34) ％ (P ＜ 0.05).In control group,BrdU/Albumin double positive cells were (69.50 ± 18.81)/FOV,and HSC group increased 61.29％ (P ＜0.01).In HSC group,the metabolic concentration of Albumin and Urea of the AHPC supernatant were (14.17 ± 3.14) mg/(L· d) and (34.83 ± 11.41) mg/(L· d),increased by 72.17％ (P ＜0.01) and 66.89％ (P ＜0.05),respectively,compared to the control group.Conclusion A certain numbers of HSC would improve clone formation rate,promote the proliferation and cell metabolism function of AHPC.