Efficacy and oncological safety of bone marrow mesenchymal stem cell transplantation for colitis in inflammatory bowel disease models in mice

doi:10.3969/j.issn.2095-4344.2014.23.015

Abstract

Abstract:

BACKGROUND: Transfusion of bone marrow mesenchymal stem cells may become a novel and effective biological therapy for inflammatory bowel disease in clinical practice. Nevertheless, the oncological safety of the treatment is worrisome, and is a key to determine whether mesenchymal stem cells can be widely used in treatment of inflammatory bowel disease, and deserves further investigation.

OBJECTIVE: To evaluate the therapeutic effect of bone marrow mesenchymal stem cell transfusion against inflammatory bowel disease in mouse models, and to clarify the effects of mesenchymal stem cells on tumorigenesis of inflammatory bowel disease.

METHODS: Mouse model of colitis was established using Balb/c (H-2d) mice exposed to dextran sulfate sodium. Syngeneic bone marrow mesenchymal stem cells were transfused into mouse model through caudal vein. The therapeutic effect of mesenchymal stem cells was compared and observed, and pathological remission of colitis was evaluated. Mouse model of colitis-driven colon carcinogenesis was established using Balb/c (H-2d) mice exposed to dextran sulfate sodium and azoxymethane. Tumor formation within the murine colon was compared and observed after transfusion of mesenchymal stem cells.

RESULTS AND CONCLUSION: In models of dextran sulfate sodium-induced colitis, weight loss and fecal occult blood were lessened in the bone marrow mesenchymal stem cell group compared with the phosphate buffered saline group. Histological damage score of colitis was less in the bone marrow mesenchymal stem cell group: mucosal structure of distal colon was almost intact under microscope, and there was small area of epithelial defects and cryptal defects. Inflammatory cell infiltration, proliferation of capillary and small vessels could be observed in mucosa and submucosa. Homing and colonization of mesenchymal stem cells in submucosa of inflamed colon could also be observed by in vivo tracing. In the dextran sulfate sodium/azoxymethane model of colitis-driven colon carcinogenesis, the number of intestinal tumors and tumor load were obviously less in the bone marrow mesenchymal stem cell group than in the control group. Results indicated that transfusion of bone marrow mesenchymal stem cells can apparently improve colitis lesions of mice with inflammatory bowel disease and inhibit carcinogenesis of colitis, which may provide theoretical support for the biological safety of mesenchymal stem cells transplantation for inflammatory bowel disease.

中国组织工程研究杂志出版内容重点：干细胞；骨髓干细胞；造血干细胞；脂肪干细胞；肿瘤干细胞；胚胎干细胞；脐带脐血干细胞；干细胞诱导；干细胞分化；组织工程

全文链接：

Key words: colitis, neoplasms, gastrointestinal neoplasms, mesenchymal stem cells, mesenchymal stem cell transplantation

CLC Number:

R394.2

He Xiao-wen, Chen Ze-xian, Zhang Long-juan, He Xiao-sheng, Lian Lei, Ke Jia, Lin Xu-tao, Chen Xi, Wu Xiao-jian, Lan Ping. Efficacy and oncological safety of bone marrow mesenchymal stem cell transplantation for colitis in inflammatory bowel disease models in mice[J]. Chinese Journal of Tissue Engineering Research, 2014, 18(23): 3696-3701.

Figures/Tables 1

2.1 实验动物数量分析实验选用40只小鼠，分别构建实验性结肠炎模型和小鼠结肠炎癌变模型，并进入分组实验，所有小鼠无意外死亡，均进入实验结果分析阶段。 2.2 骨髓间充质干细胞的生物学特性鉴定骨髓间充质干细胞经扩增传代3代以后，获得形态比较均一的梭形细胞，平行排列或旋涡样单层集落生长，也可见多角形和扁平状细胞。相差显微镜下细胞核浆比大，胞浆少，核大，核仁明显。应用流式细胞检测显示，骨髓间充质干细胞不表达CD34，CD45和MHCⅡ类分子，高表达CD29，CD44和MHCⅠ类分子。体外经成骨诱导培养两至三周后，大多数细胞发生形态变化，由原来的梭形变为多角形或不规则形，细胞呈结节状生长，并有钙盐沉积，茜素红S染色证实有骨钙结节形成(图1A)；成脂诱导分化2周后，细胞内可见融合成团、被油红O染成红色的脂滴(图1B)。 2.3 输注间充质干细胞从临床和组织学水平有效控制小鼠结肠炎尽管对照组结肠炎小鼠在停止饮用葡聚糖硫酸钠后症状趋向逐渐好转，但治疗组小鼠恢复更加明显，腹泻、便血明显好转，实验结束时体质量丧失、粪便黏稠度(不成形黏稠大便)、大便潜血较对照组均有减轻。镜下，间充质干细胞治疗组小鼠远端结肠黏膜结构基本完整，有小范围上皮缺损或隐窝缺损，黏膜层和黏膜下层较多炎症细胞浸润，可见毛细血管和小血管增生。而对照组小鼠远端结肠黏膜结构较紊乱，有较大范围上皮缺损或部分隐窝消失，见少量隐窝脓肿，黏膜及黏膜下层有大量炎性细胞浸润，包括中性粒细胞、淋巴细胞、巨噬细胞和浆细胞(图2)。间充质干细胞治疗组小鼠结肠远端组织炎症损伤严重度评分低于对照组(P < 0.05；图3)。 2.4 输注的间充质干细胞向炎症肠道组织归巢定殖体外CM-DiI标记间充质干细胞，显示细胞膜和胞浆质膜被标上染料，胞核无标记，标记率在90%以上。组织标本冰冻切片可见有CM-DiI标记的间充质干细胞向结肠远端炎症病变处归巢定殖，且主要位于黏膜下层(图4)，同一小鼠结肠近端及小肠组织内未见CM-DiI标记的细胞。 2.5 输注间充质干细胞对小鼠结肠炎癌变具有抑制作用建立Balb/c(H-2d)小鼠的炎症性肠病癌变模型，在建模周期的前3周经尾静脉输注同系小鼠骨髓间充质干细胞3次。建模结束后观察肠道肿瘤形成的情况，如图5，结果显示经间充质干细胞治疗的小鼠肠道肿瘤数量及肿瘤负荷(单只小鼠肿瘤的直径之和)明显少于对照组(P < 0.05)。"

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