骨髓单个核细胞移植对肾缺血再灌注后肾小管坏死及细胞凋亡的影响

目的：观察和分析外源性骨髓单个核细胞（BMMNCs）对肾缺血再灌注诱发的肾小管坏死及凋亡的影响。方法：密度梯度离心法分离获取BMMNCs并行DAPI标记。制作SD大鼠肾缺血再灌注损伤模型，通过下腔静脉进行BMMNCs移植。分别于缺血再灌注后不同时相获取肾脏标本，荧光显微镜下观察DAPI的标记情况，HE染色做肾组织学检测，TUNEL法检测肾组织凋亡细胞，免疫组织化学染色观察增殖细胞核抗原（PCNA）的表达。结果：BMMNCs移植组大鼠肾脏组织中可见DAPI荧光细胞，部分存在于肾小管上皮组织中，未见明显细胞坏死及变性征象，其凋亡细胞计数显著减少，而PCNA阳性细胞数增多。结论：外源性BMMNCs移植可减少缺血再灌注诱发的肾小管上皮细胞的变性、坏死和凋亡，促进缺血再灌注损伤后肾小管上皮细胞的增殖，从而提示BMMNCs移植有助于缺血再灌注损伤后肾小管的修复及其结构完整性的维持。

Effect of bone marrow mononuclear cells on necrotic and apoptotic cell death induced by renal ischemia-reperfusion in rats

AIM: To investigate the effect of bone marrow mononuclear cells (BMMNCs) on necrotic and apoptotic cell death and to test whether BMMNCs can restore the renal tissues with ischemia-reperfusion injury to tubular structural integrity in rats. METHODS: BMMNCs were isolated from the femurs and tibiae of male SD rats with Percoll density gradient centrifugation and labeled with DAPI in vitro. All model rats were subjected to clamping bilateral renal pedicles for 45 min, followed by injection of BMMNCs or saline into the inferior vena cava, and were sacrificed to collect kidneys at 1, 2, 7, 14 d after operation, respectively. Fluorescence microscope was performed to detect the DAPI labels in the renal paraffin slices. All kidney samples were examined by HE staining under light microscope. Apoptosis was detected by terminal deoxynucleotidyl transferase mediated nick end labeling (TUNEL). Immunohistochemical staining was used to examine the expression of proliferating cell nuclear antigen (PCNA). RESULTS: There were lots of cells with blue fluorescence in the injured kidney samples under fluorescence microscope, some of which lay in the renal tubular epithelium. Tubular cell necrosis and degeneration were observed in kidneys of control group, no necrosis and significant tubular cell degeneration was observed in kidneys of transplantation group. Decreased numbers of apoptotic cells were detected in kidney samples of transplantation group as compared with control group. The numbers of PCNA-positive cells in kidney samples of transplantation group were higher than those in control group. CONCLUSION: Transplanted BMMNCs migrate into the injured kidney, some of which might directly reconstitute the tubular epithelium. Transplantation of these cells attenuates necrotic and apoptotic cell death and accelerates the proliferation of the renal tubular epithelium after ischemia-reperfusion injury. It is suggested that these cells might contribute to restore injured kidney to the tubular structural integrity.