CAJ | 학술논문

目的探讨输尿管上皮种子细胞的分离培养和增殖规律,以及构建种植种子细胞输尿管组织工程网状支架的可行性.方法采用自制输尿管翻转分离器,分离实验用小型猪输尿管上皮种子细胞.体外培养并鉴定输尿管上皮种子细胞.观察输尿管种子细胞形态变化,噻唑蓝(MTT)比色法检测原代、第3代、第5代和第7代种子细胞增殖特性.后将输尿管种子细胞种植到电纺丝输尿管网状支架,于支架孵育第1、3、5、7、9天检测种子细胞增殖活性,并绘制细胞生长曲线.结果成功分离培养输尿管上皮种子细胞,角蛋白抗体和尿路上皮特异抗体UroplakinⅡ证实为输尿管尿路上皮细胞.输尿管上皮种子细胞接种24 h基本贴壁,呈多角形团簇样生长,8d时细胞融合70％以上,呈铺路石状排列.输尿管上皮种子细胞在电纺丝输尿管网状支架上生长增殖良好.结论改良输尿管上皮细胞分离方法可获得大量纯化的输尿管上皮种子细胞,其在输尿管网状支架上生长增殖良好,成功构建输尿管组织工程网状支架有望应用于输尿管损伤替代修复.
목적 탐토수뇨관상피충자세포적분리배양화증식규률,이급구건충식충자세포수뇨관조직공정망상지가적가행성.방법 채용자제수뇨관번전분리기,분리실험용소형저수뇨관상피충자세포.체외배양병감정수뇨관상피충자세포.관찰수뇨관충자세포형태변화,새서람(MTT)비색법검측원대、제3대、제5대화제7대충자세포증식특성.후장수뇨관충자세포충식도전방사수뇨관망상지가,우지가부육제1、3、5、7、9천검측충자세포증식활성,병회제세포생장곡선.결과 성공분리배양수뇨관상피충자세포,각단백항체화뇨로상피특이항체UroplakinⅡ증실위수뇨관뇨로상피세포.수뇨관상피충자세포접충24 h기본첩벽,정다각형단족양생장,8d시세포융합70％이상,정포로석상배렬.수뇨관상피충자세포재전방사수뇨관망상지가상생장증식량호.결론 개량수뇨관상피세포분리방법가획득대량순화적수뇨관상피충자세포,기재수뇨관망상지가상생장증식량호,성공구건수뇨관조직공정망상지가유망응용우수뇨관손상체대수복.
Objective To investigate the isolation and proliferation of ureter epithelial cells,and the feasibility of construction of tissue engineering ureter.Methods The ureters of minipigs were excised under sterile conditions.Homemade ureteral cell dissection tools were made and used to turn the ureteral mucosal surface outside.Ureteral epithelial cells were isolated and resuspended in Defined Keratinocyte SFM.Immunohistochemistry was used to identify cells.Morphology,growth and proliferation of the primary ureteral epithelial cells and cells from passage 3,5 and 7 were observed.Cells were then seeded onto an electropspun polylactic acid scaffold to investigate the feasibility of construction of tissue engineering ureter.At the 1st,3rd,5th,7th and 9th day of incubation,MTT assay was adopted to get a cell growth curve.Results Immunohistochemistry and morphology demonstrated that the cultured cells were exclusively ureteral epithelial cells and retained the properties of normal urothelium.Most of the cells attached to the culture dish and began to spread after 24 h.The cultured ureteral epithelial cells possessed a typical appearance of cobblestone and their confluence reached up to 70％ at 8th day.The seeded cells on the scaffold grew well and had a similar growth curve as cells growing on a culture dish.Conclusion The method of isolation can harvest a larger number of homogeneous ureteral epithelial cells.The cells can be used as seed cells for constructing tissue engineering ureter which can be used for reconstruction of injured ureter.