白血病抑制因子 (Leukemia inhibitory factor, LIF) 是一种调节细胞生长、增殖和分化的重要因子。骨髓间充质干细胞 (Bone marrow mesenchymal stem cells, BM‑MSCs) 在组织损伤修复、自身免疫疾病治疗中具有广阔的应用前景，对大熊猫等濒危动物的疾病治疗和遗传保护方面有着巨大的潜力。本研究旨在构建大熊猫LIF过表达载体，应用脂质体转染大熊猫骨髓间充质干细胞 (Panda bone marrow mesenchymal stem cells, PDBM‑MSCs)，研究LIF对PDBM‑MSCs生长的影响。本研究从PDBM‑MSCs cDNA 中扩增LIF基因CDS区，无缝克隆连接至pCDH‑CMV‑MCS‑EF1‑GFP+puro过表达载体上，构建 LIF过表达重组质粒，利用Lipo3000转染重组质粒至PDBM‑MSCs中。研究结果显示，转染48 h后PDBM‑MSCs表达绿色荧光，转染72 h后分别提取细胞总RNA，反转录成cDNA，经q‑PCR检测发现，转染后LIF过表达组相比阴性对照组LIF的mRNA表达水平显著提高。同时，LIF过表达后能够显著上调细胞周期调控基因CCNB2、CDK7在mRNA水平上的表达，显著下调凋亡基因caspase3、P53、P16、P21在mRNA水平上的表达；转染72 h后流式检测显示转染LIF过表达重组质粒的PDBM‑MSCs细胞活率显著高于阴性对照组。本研究成功构建大熊猫LIF过表达载体并转染至PDBM‑MSCs中，显著提高细胞活率、抑制凋亡，表明LIF对PDBM‑MSCs起到促进生长抑制凋亡的作用，为LIF在大熊猫中的作用机制研究奠定了基础。

Leukemia inhibitory factor (LIF) is an important factor regulating cell growth, proliferation and differentiation. Bone marrow mesenchymal stem cells (BM-MSCs) have broad application prospects in the repair of tissue damage as well as the treatment of autoimmune diseases, and, as a result, have great potential in the disease treatment and genetic conservation of rare and endangered animals, such as giant pandas. The aim of this study was to construct a giant panda LIF overexpression vector and transfect panda bone marrow mesenchymal stem cells (PDBM-MSCs) with liposome to study the effect of LIF on the growth of PDBM-MSCs. The CDS region of the LIF gene was amplified from the cDNA of PDBM-MSCs, and seamlessly cloned onto the pCDH-CMV-MCS-EF1-GFP+puro overexpression vector to construct the LIF overexpression recombinant plasmid. Lipo3000 was used to transfect recombinant plasmid into PDBM-MSCs. The results showed that PDBM-MSCs expressed green fluorescence 48 h after transfection, and total RNA was extracted and reverse-transcribed into cDNA 72 h after transfection. A q-PCR detection showed that the mRNA expression level of LIF in the LIF overexpression group was significantly higher than that in the negative control group. Moreover, overexpression of LIF could significantly up-regulate the mRNA expressions of CCNB2 and CDK7, and significantly down-regulate the mRNA expressions of apoptosis genes caspase3, P53, P16 and P21. 72 h after transfection, flow detection showed that the viability of PDBM-MSCs transfected with LIF overexpression recombinant plasmid was significantly higher than that of the negative control group. In this study, the panda LIF overexpression vector was successfully constructed and transfected into PDBM-MSCs, which significantly improved cell viability and inhibited apoptosis, indicating that LIF can promote growth and inhibit apoptosis of PDBM-MSCs, laying a foundation for the study of the mechanism of LIF in giant pandas.