CAJ | 학술논문

目的建立大鼠体外多器官细胞共培养方法.方法采用胶原酶消化法、支气管灌洗分离法、两步消化法等方法分离大鼠原代肝细胞、肾细胞、心肌细胞、肺泡巨噬细胞和真皮成纤维细胞,用锥虫蓝染色法检测细胞分离成活率,用单层贴壁法分别培养;采用飞片法,将5种细胞的飞片放入同一培养皿,用体积分数为10%的FBS DMEM培养基培养7 d,用噻唑蓝(MTT)比色法比较原代细胞在单独培养和共培养时的生长情况.结果建立的大鼠原代肝细胞、肾细胞、心肌细胞、肺泡巨噬细胞、真皮成纤维细胞分离方法稳定,细胞分离成活率均达90%,其中胶原酶消化法培养肝细胞的存活率达90.3%,两步消化法培养肾细胞的细胞存活率达91.9%,心肌细胞的胶原酶消化法细胞存活率达93.0%.3～15 d的心肌细胞搏动频率比较,差异无统计学意义(P>0.05),差速贴壁法培养的肺泡巨噬细胞存活率可达90.8%,以胶原酶消化法培养的原代真皮细胞存活率达92.7%,细胞生长状态良好.5种原代细胞的多器官细胞共培养结果显示,共培养时细胞生长增殖情况良好,单独培养与共培养细胞生长曲线基本重合.结论所建立的大鼠多器官细胞共培养方法是成功的.
목적 건립대서체외다기관세포공배양방법.방법 채용효원매소화법、지기관관세분리법、량보소화법등방법분리대서원대간세포、신세포、심기세포、폐포거서세포화진피성섬유세포,용추충람염색법검측세포분리성활솔,용단층첩벽법분별배양;채용비편법,장5충세포적비편방입동일배양명,용체적분수위10%적FBS DMEM배양기배양7 d,용새서람(MTT)비색법비교원대세포재단독배양화공배양시적생장정황.결과 건립적대서원대간세포、신세포、심기세포、폐포거서세포、진피성섬유세포분리방법은정,세포분리성활솔균체90%,기중효원매소화법배양간세포적존활솔체90.3%,량보소화법배양신세포적세포존활솔체91.9%,심기세포적효원매소화법세포존활솔체93.0%.3～15 d적심기세포박동빈솔비교,차이무통계학의의(P>0.05),차속첩벽법배양적폐포거서세포존활솔가체90.8%,이효원매소화법배양적원대진피세포존활솔체92.7%,세포생장상태량호.5충원대세포적다기관세포공배양결과현시,공배양시세포생장증식정황량호,단독배양여공배양세포생장곡선기본중합.결론 소건립적대서다기관세포공배양방법시성공적.
Objective To establish the integrated discrete multiple organ cell culture (IdMOC) system. Methods Rat primary cell of hepatocyte, nephrocyte, cadiocyte, alveolar macrophage, dermal fibroblasts were isolated by collagenase digestion, separation of bronchial lavage, two-step digestion method and cultred respectively, with monolayer culture. To establish the integrated discrete multiple organ cell culture (IdMOC) system, glass slides of five different cells were used to the same dish with 10% FBS DMEM medium cultured 7d, using MTT comparison primary cells cultured alone and cocultured when growth. Results Established rat hepatocytes, renal cell, cardiomyocyte, alveolar macrophages, dermal fibroblasts separation method was stable, cell separation survival rate was about 90.0%. Hepatocytes separation survival rate 90.3% ,renal cell separation survival rate 91.9% , cardiomyocyte separation survival rate 93.0% and beating rate indifference curve among 3d-15d, alveolar macrophages cell separation survival rate 90.8%, dermal fibroblasts cell separation survival rate 92.7% . Five primary cells multiple organ cells coculture showed cocultured cell growth proliferation well, cultured alone and cocultured cells growth curve basic coincide. Conclusion Established rat multiple organ cell co-culture is successful.