Wei L, Han Z J, Xu L, Li J W
Department of Thoracic Surgery, Henan Provincial People's Hospital, Zhengzhou 450003, China.
Zhonghua Yi Xue Za Zhi. 2018 Jul 3;98(25):2019-2023. doi: 10.3760/cma.j.issn.0376-2491.2018.25.011.
To establish the ischemia reperfusion injury model in rat after lung transplantation(LT) and explore the expression of high mobility group box 1 protein(HMGB1) after intravenous injection with bone marrow mesenchymal stem cells(MSCs). Forty healthy 8-10 weeks male SD rats were randomly divided into four groups including the sham-operated group, ischemia-reperfusion (IR), Saline-IR and MSC-IR group. The sham-operated rats were only conducted thoracotomy without lung transplantation and the rest groups were respectively conducted with the left LT, left LT followed by 1 ml saline and left LT followed by 1 ml MSCs (1.0×10(7)/ml). Four groups of rats were killed at 24 h after reperfusion. The blood and left lung tissue were collected. Oxygenation index(OI) and the ratio of wet/dry in four groups were detected and histological sections stained with hematoxylin and eosin (HE) were made. HMGB1 levels in serum were detected with ELISA. Real-time PCR and Western blot were performed to detect the expression of HMGB1 in mRNA and protein levels. The OI in four groups were respectively 383±15, 174±24, 170±30 and 217±21.OI in IR and Saline-IR group decreased compared with the sham-operated group , all <0.01. The OI increased after injection with MSCs compared with IR group, <0.01. The histological images showed the marked inflammatory infiltrates and interalveolar septal thickening in IR group. Treatment with MSCs reduced inflammatory injury.The ratio of wet/dry in IR group and Saline-IR group increased compared with the sham-operated group((5.38±0.19), (5.24±0.15) vs (4.16±0.12), all <0.05). Ratio in MSC-IR group decreased compared with the IR group (4.47±0.14) vs (5.38±0.19), <0.05. ELISA results showed that HMGB1 level increased significantly in IR group (287±37)ng/ml, Saline-IR group (260±24)ng/ml and MSC-IR group (101±14)ng/ml when compared with the sham-operated group (41±5) ng/ml. The serum HMGB1 level in IR group was positively correlated with the OI (=0.759, <0.05) and wet/dry ratio (=0.725, <0.05). RT-PCR showed that HMGB1 mRNA level in sham-operated group was the lowest and increased significantly in IR group, while decreased significantly in MSC-IR group compared with IR group and Saline-IR group(<0.01). The results of HMGB1 expression at protein level by Western blot were consistent with the mRNA level. Lung transplantation can induce the expression of HMGB1 but HMGB1 level of lung tissue decreased significantly after the treatment with MSCs, which indicated that MSCs might play an important role in protecting transplanted lung via HMGB1.
建立大鼠肺移植（LT）后缺血再灌注损伤模型，并探讨静脉注射骨髓间充质干细胞（MSCs）后高迁移率族蛋白B1（HMGB1）的表达。将40只健康的8 - 10周龄雄性SD大鼠随机分为四组，包括假手术组、缺血再灌注（IR）组、生理盐水 - IR组和MSC - IR组。假手术大鼠仅行开胸手术，不行肺移植，其余组分别行左肺移植、左肺移植后注射1 ml生理盐水及左肺移植后注射1 ml MSCs（1.0×10⁷/ml）。再灌注24小时后处死四组大鼠，采集血液和左肺组织。检测四组的氧合指数（OI）和湿干比，制作苏木精 - 伊红（HE）染色的组织切片。用酶联免疫吸附测定（ELISA）检测血清中HMGB1水平。进行实时聚合酶链反应（Real - time PCR）和蛋白质免疫印迹法（Western blot）检测HMGB1在mRNA和蛋白质水平的表达。四组的OI分别为383±15、174±24、170±30和217±21。IR组和生理盐水 - IR组的OI与假手术组相比降低，均P<0.01。与IR组相比，注射MSCs后OI升高，P<0.01。组织学图像显示IR组有明显的炎性浸润和肺泡间隔增厚。MSCs治疗减轻了炎性损伤。IR组和生理盐水 - IR组的湿干比与假手术组相比升高（（5.38±0.19），（5.24±0.15）对（4.16±0.12），均P<0.05）。MSC - IR组的湿干比比IR组降低（4.47±0.14）对（5.38±0.19），P<0.05。ELISA结果显示，与假手术组（41±5）ng/ml相比，IR组（287±37）ng/ml、生理盐水 - IR组（260±24）ng/ml和MSC - IR组（101±14）ng/ml的HMGB1水平显著升高。IR组血清HMGB1水平与OI呈正相关（r = 0.759，P<0.05），与湿干比呈正相关（r = 0.725，P<0.05）。逆转录 - 聚合酶链反应（RT - PCR）显示，假手术组HMGB1 mRNA水平最低，IR组显著升高，而与IR组和生理盐水 - IR组相比，MSC - IR组显著降低（P<0.01）。蛋白质免疫印迹法检测HMGB1在蛋白质水平的表达结果与mRNA水平一致。肺移植可诱导HMGB1表达，但MSCs治疗后肺组织中HMGB1水平显著降低，这表明MSCs可能通过HMGB1在保护移植肺中发挥重要作用。
