骨髓来源的非粘附基质干细胞在造血重建和遗传缺陷纠正中的作用
发布时间：2018-08-18 19:48
【摘要】：本文假设成体骨髓中存在一类处于非粘附状态的、悬浮的、与造血细胞类似的多潜能骨髓基质干细胞,它们虽具有贴壁潜能,但比贴壁的基质干细胞更原始,称为骨髓来源的非粘附基质干细胞(non-adherent bone marrow-derived stromal cell,NA-BM-SC)。这类细胞能够经血液循环转运到全身各器官,在特定的微环境下贴壁,增殖并分化为各器官的终末分化细胞,参与正常组织的再生及损伤组织的修复。 为了验证这一假设,从以下三个部分进行实验研究: 第一部分采用反复转移非贴壁骨髓细胞(non-adherent bone marrow cells,NA-BMC)体外培养法,取6周龄小鼠股骨和胫骨来源的骨髓总细胞(bone marrow cells,BMC)培养10-18天,每天转移一次非贴壁骨髓细胞至新的培养皿,即“pour off”,至少4次。对一天后贴壁的细胞加入新鲜培养液继续培养。实验结果证实了不仅总BMC可以产生成纤维细胞克隆形成单位(Fibroblastoid Colony forming unit,CFU-F),非贴壁骨髓细胞也能产生CFU-F;地塞米松可以抑制CFU-F的形成效率,而1.25(OH)_2D_3则可以部分拮抗地塞米松的抑制作用。为了确定NA-BMC来源的基质干细胞(NA-BM-SC)是否具有自我更新能力和多分化潜能,采用5-溴脱氧尿核苷(Bromodeoxyuridine,BrdU)掺入法、免疫细胞化学法和流式细胞仪法证实了NA-BM-SC不但具有自我更新能力,而且在体外不同条件下培养,能分化为成骨细胞、软骨细胞、脂肪细胞和神经胶质细胞。 第二部分采用维生素D受体(vitamin D receptor,VDR)基因敲除小鼠,经致死剂量照射后,移植野生型小鼠的NA-BM-SC,通过VDR基因的示踪作用,观察NA-BM-SC是否具有造血重建和参与辐射损伤修复的作用。结果显示,单纯照射组的小鼠由于造血系统损伤、肠组织和多脏器坏死等原因,两周内全部死亡;然而经过NA-BM-SC移植治疗的照射移植组小鼠,在移植后30、60和90天的存活率分别达到75%、65%和60%。外周血细胞在照射移植后30天基本恢复到正常水平。通过PCR和免疫组织化学法检测,VDR基因在照射移植组存活小鼠的多个器官表达,包括骨髓、脾脏和小肠。结果提示NA-BM-SC不仅可以重建小鼠的造血系统,而且可
[Abstract]:In this paper, we hypothesize that there is a class of non-adherent, suspended, hematopoietic-like multipotent bone marrow stromal cells (NA-BM-SC) in adult bone marrow. Although they have adherent potential, they are more primitive than adherent stromal cells. These cells can be transported through blood circulation to all organs of the body, adhere to the wall in a specific microenvironment, proliferate and differentiate into terminal differentiation cells of various organs, and participate in the regeneration of normal tissues and repair of damaged tissues.
In order to test this hypothesis, the following three parts are studied:
In the first part, non-adherent bone marrow cells (NA-BMC) from femur and tibia of 6-week-old mice were cultured in vitro for 10-18 days. Non-adherent bone marrow cells were transferred to a new culture dish, i.e. pour off, at least four times a day. Adherent cells were further cultured in fresh medium. The results showed that not only total BMC could produce fibroblast cloning unit (CFU-F), but also non-adherent bone marrow cells could produce CFU-F. Dexamethasone could inhibit the formation of CFU-F and 1.25 (OH) _2D_3 could partially antagonize dexamethasone. To determine whether NA-BM-SC has the ability of self-renewal and multi-differentiation, 5-bromodeoxyuridine (BrdU) incorporation, immunocytochemistry and flow cytometry confirmed that NA-BM-SC not only has the ability of self-renewal, but also under different conditions in vitro. Under culture, they can differentiate into osteoblasts, chondrocytes, adipocytes and glial cells.
In the second part, we used vitamin D receptor (VDR) knockout mice. After lethal dose irradiation, NA-BM-SC was transplanted into wild type mice to observe whether NA-BM-SC could reconstruct hematopoiesis and participate in the repair of radiation injury. However, the survival rates of the irradiated mice treated with NA-BM-SC were 75%, 65% and 60% at 30, 60 and 90 days after transplantation, respectively. Peripheral blood cells returned to normal level at 30 days after transplantation. VDR gene was detected by PCR and immunohistochemistry. The expression of NA-BM-SC in multiple organs including bone marrow, spleen and small intestine of surviving mice in irradiated transplantation group suggested that NA-BM-SC could not only reconstruct the hematopoietic system of mice, but also reconstruct the hematopoietic system.
【学位授予单位】：苏州大学
【学位级别】：博士
【学位授予年份】：2005
【分类号】：R329
本文编号：2190500
[Abstract]:In this paper, we hypothesize that there is a class of non-adherent, suspended, hematopoietic-like multipotent bone marrow stromal cells (NA-BM-SC) in adult bone marrow. Although they have adherent potential, they are more primitive than adherent stromal cells. These cells can be transported through blood circulation to all organs of the body, adhere to the wall in a specific microenvironment, proliferate and differentiate into terminal differentiation cells of various organs, and participate in the regeneration of normal tissues and repair of damaged tissues.
In order to test this hypothesis, the following three parts are studied:
In the first part, non-adherent bone marrow cells (NA-BMC) from femur and tibia of 6-week-old mice were cultured in vitro for 10-18 days. Non-adherent bone marrow cells were transferred to a new culture dish, i.e. pour off, at least four times a day. Adherent cells were further cultured in fresh medium. The results showed that not only total BMC could produce fibroblast cloning unit (CFU-F), but also non-adherent bone marrow cells could produce CFU-F. Dexamethasone could inhibit the formation of CFU-F and 1.25 (OH) _2D_3 could partially antagonize dexamethasone. To determine whether NA-BM-SC has the ability of self-renewal and multi-differentiation, 5-bromodeoxyuridine (BrdU) incorporation, immunocytochemistry and flow cytometry confirmed that NA-BM-SC not only has the ability of self-renewal, but also under different conditions in vitro. Under culture, they can differentiate into osteoblasts, chondrocytes, adipocytes and glial cells.
In the second part, we used vitamin D receptor (VDR) knockout mice. After lethal dose irradiation, NA-BM-SC was transplanted into wild type mice to observe whether NA-BM-SC could reconstruct hematopoiesis and participate in the repair of radiation injury. However, the survival rates of the irradiated mice treated with NA-BM-SC were 75%, 65% and 60% at 30, 60 and 90 days after transplantation, respectively. Peripheral blood cells returned to normal level at 30 days after transplantation. VDR gene was detected by PCR and immunohistochemistry. The expression of NA-BM-SC in multiple organs including bone marrow, spleen and small intestine of surviving mice in irradiated transplantation group suggested that NA-BM-SC could not only reconstruct the hematopoietic system of mice, but also reconstruct the hematopoietic system.
【学位授予单位】：苏州大学
【学位级别】：博士
【学位授予年份】：2005
【分类号】：R329
【引证文献】
相关期刊论文 前1条
1 万建美;罗蔚锋;李厚怀;包仕尧;刘春风;张志琳;李冰燕;;激光扫描共聚焦显微镜在骨髓基质细胞移植中的应用[J];激光杂志;2010年05期
，本文编号：2190500
本文链接：https://www.wllwen.com/yixuelunwen/binglixuelunwen/2190500.html
最近更新
教材专著
