尿源干细胞在治疗慢性肾病大鼠模型中的应用

发布时间：2018-05-27 11:41

本文选题：尿源干细胞 + 细胞疗法　；参考：《第二军医大学》2014年博士论文

【摘要】：背景：慢性肾病是一种可以导致尿毒症和心血管系统并发症的常见疾病，且发病率一直处于上升趋势。全世界慢性肾病患者约占总人口的13%，使之成为一个重大的公共卫生问题。我国慢性肾病的流行病学也不容乐观，慢性肾病患者约占总人口的10.25%左右，且发病率逐年升高。慢性肾病为进展性疾病，可发展至终末期肾病，给患者身体上及心理上造成巨大的创伤。与此同时，随之而来的经济负担也十分沉重。2007年，仅美国的终末期肾病支出就高达230多亿，占其全部医疗支出的5.8%。目前对于终末期肾病的治疗手段仍十分有限，主要依赖透析和肾移植。透析成本高昂、并发症多、且降低患者的生活质量。此外，透析仅仅替代肾脏的滤过功能，而不能替代其内分泌等功能。另一方面，供体的短缺严重制约了肾移植的开展，，且免疫排斥和免疫抑制药物对患者造成的影响也难以解决。因此，临床上需要新的治疗慢性肾病的手段。细胞疗法近年来逐渐受到重视，尤其在干细胞方面，由于其强大的自我更新和多向分化能力，在各个研究领域均取得了长足的进展。尿源干细胞是一种新的干细胞来源，属于间充质干细胞，可从新鲜尿液中获取，其获取手段无创，可反复收集从而得到大量的起始细胞。间充质干细胞对肾功能有改善作用，而尿源干细胞最可能的来源是肾脏，组织来源上的优势使之可能对肾功能具备更明显的改善作用。研究目的：研究尿源干细胞是否对慢性肾病具有治疗作用，并初探其机制。研究方法： 1.从健康供体新鲜尿液中提取尿源干细胞，通过自我更新能力、多向分化能力、表面标志物情况对尿源干细胞进行鉴定和确认。 2.向裸小鼠肾脏中注射尿源干细胞，在注射细胞1周、2周、3周、4周后处死小鼠、收集肾脏，观察尿源干细胞的分布情况，并通过观察其表面标志物的表达情况初探其分化为肾脏细胞的可能性。 3.实验动物分为3组，第一组为年龄匹配的正常对照，第二组为接受注射尿源干细胞的慢性肾病大鼠模型，第三组为注射磷酸盐缓冲液的慢性肾病大鼠模型，通过对其血清肌酐、肾小球滤过率的随访判断尿源干细胞对肾功能的改善作用，并对3组肾脏进行组织形态学的评估。结果： 1.新鲜尿液中获得的细胞克隆可自我更新，能够分化成为脂肪细胞和骨细胞。尿源干细胞表达间充质干细胞的表面标志物：CD24、CD29、CD44、CD73、CD90、CD105、CD146等；不表达造血细胞表面抗原：CD31、CD34、CD45等。 2.尿源干细胞可以在肾脏组织内存活，在有限的随访时间内，显微镜下未发现与肾脏组织结构的融合，且尿源干细胞始终表达间充质干细胞的表面抗原，分化的可能性较小。 3.尿源干细胞对肾功能具有改善作用，相较于注射磷酸盐缓冲液的对照组，注射尿源干细胞的动物模型血清肌酐降低、肾小球滤过率升高。尿源干细胞对肾脏的组织学形态有改善作用，相较于注射磷酸盐缓冲液的对照组，注射尿源干细胞的动物模型健康肾单元的数量增加、一型胶原和三型胶原数量减少、肾脏间质炎性细胞浸润减轻。结论：尿源干细胞可能对慢性肾病患者的肾功能有改善作用，可能成为无创的、自体的治疗慢性肾病的新的细胞来源，其作用机制和优化的治疗手段仍需要进一步的研究。
[Abstract]:Background :

Chronic kidney disease is a common disease that can lead to uremia and cardiovascular system complications . The incidence of chronic kidney disease in the world is increasing . The epidemiology of chronic kidney disease in China is about 10 . 25 % of the total population .

Cell therapy has been paid more and more attention in recent years , especially in stem cells , due to its strong self - renewal and multi - directional differentiation ability , it has made great progress in various research fields . The urine source stem cells are a new source of stem cells , which can be obtained from fresh urine , and can be collected repeatedly to obtain a large number of initial cells .

Purpose of study :

To study whether urine source stem cells have therapeutic effect on chronic kidney disease and explore its mechanism .

Study method :

1 . The urine source stem cells are extracted from fresh urine of healthy donors , and the urine source stem cells are identified and confirmed by self - updating capability , multi - directional differentiation ability and surface markers .

2 . To inject urine source stem cells into the kidney of nude mice . After 1 week , 2 weeks , 3 weeks and 4 weeks of injection , mice were sacrificed , the kidney was collected , the distribution of urine source stem cells was observed , and the possibility of differentiation into kidney cells was investigated by observing the expression of their surface markers .

3 . The experimental animals were divided into three groups , the first group is the normal control group matched with age , the second group is the rat model of chronic kidney disease receiving the injection urine source stem cell , the third group is the rat model of chronic kidney disease injected with phosphate buffer solution , and the function of improving the renal function of the urine source stem cell is judged by the follow - up of serum creatinine and glomerular filtration rate , and the three groups of kidneys are evaluated .

Results :

1 . The cell clones obtained in fresh urine can be self - updated to differentiate into adipocytes and osteocytes . Surface markers of mesenchymal stem cells expressed by urine derived stem cells : CD24 , CD29 , CD44 , CD73 , CD90 , CD105 , CD146 , etc . ;
Do not express hematopoietic cell surface antigen : CD31 , CD34 , CD45 , etc .

2 . The urine source stem cells can survive in the renal tissue , and in a limited follow - up time , there is no fusion with the kidney tissue structure under the microscope , and the urine source stem cells always express the surface antigen of the mesenchymal stem cells , and the possibility of differentiation is small .

3 . Urine derived stem cells have an improved effect on renal function . Compared with the control group injected with phosphate buffer solution , the serum creatinine of the animal model injected with urine source stem cells is decreased , and the glomerular filtration rate is increased . The number of healthy kidney cells in the animal model of the injected urine source stem cells is increased compared with the control group injected with phosphate buffer solution , the number of the type I collagen and the type III collagen is reduced , and the infiltration of the renal interstitial inflammatory cell is reduced .

Conclusion :

Urine derived stem cells may improve the renal function of patients with chronic kidney disease , and may be non - invasive , autologous therapy for chronic kidney disease . The mechanism of action and optimization of treatment still need further research .
【学位授予单位】：第二军医大学
【学位级别】：博士
【学位授予年份】：2014
【分类号】：R692;R-332

【共引文献】