转录因子Twist调控巨噬细胞异质性促进肾间质纤维化的机制研究

Renal interstitial fibrosis is a common pathological characteristic of various CKD, and macrophage infiltration and activation play an important role in the occurrence of renal fibrosis.Our previous studies have confirmed that transcription factor Twist is an important molecule that leads to renal fibrosis: the up-regulation of Twist causes the transdifferentiation of renal tubular epithelial cells.Twist may be involved in the regulation of macrophages in addition to the damage of the tubular epithelial cells.We also found that the expression level of Twist in renal macrophages was positively correlated with the degree of renal fibrosis.After unilateral ureteral obstruction in macrophage specific twist gene knockout mouse, renal fibrosis was significantly relieved, macrophages infiltration decreased, and M2-type cells reduced. Thus, we predicted that twist regulates the activation and function of macrophages, and accelerates renal fibrosis. This research proposed using the technology of specific gene knockout in macrophages to explore the effect of transcription factor Twist on the regulation mechanism of macrophage plasticity and its effect on renal interstitial fibrosis, to provide a theoretical basis for comprehensively elucidate Twist as a new therapeutic target for renal fibrosis.factor Twist on the regulation mechanism of macrophage plasticity and its effect on renal interstitial fibrosis, to provide a theoretical basis for comprehensively elucidate Twist as a new therapeutic target for renal fibrosis.

肾间质纤维化是各种CKD最终的共同病理改变，巨噬细胞浸润和活化在肾纤维化发生中发挥重要作用。我们前期研究证实，转录因子Twist是促进肾纤维化发生的重要分子；Twist除了介导肾小管上皮细胞损伤外，还可能参与巨噬细胞的调控。我们发现：小鼠肾纤维化模型中，高表达Twist的肾小管周围有大量巨噬细胞浸润，这些巨噬细胞中有Twist表达，且其表达水平与肾纤维化程度正相关；巨噬细胞特异性Twist基因敲除鼠UUO后，肾纤维化明显减轻，巨噬细胞浸润减少，M2型细胞比例降低。由此推测，Twist可能调控巨噬细胞迁移、活化及功能，进而加速肾纤维化，但其分子机制还不清楚。本课题拟利用巨噬细胞特异性基因剔除等技术，探讨转录因子Twist对巨噬细胞可塑性调控的分子机制及对肾间质纤维化的影响，为转录因子Twist成为肾纤维化新的治疗靶点提供理论依据。

肾间质纤维化是各种CKD最终的共同病理改变，巨噬细胞浸润和活化在肾纤维化中发挥重要作用。本研究发现在临床肾活检标本和小鼠单侧输尿管结扎（unilateral ureteral obstruction，UUO）肾纤维化模型中，巨噬细胞中Twist的表达与巨噬细胞浸润量及肾纤维化严重程度呈正相关。为进一步探讨巨噬细胞中Twist在肾纤维化中的作用，我们成功构建了巨噬细胞特异性剔除Twist基因小鼠（简称敲除鼠）。在UUO肾纤维化模型小鼠中，我们发现：与野生鼠相比，敲除鼠UUO术后肾组织中巨噬细胞浸润减少，纤维化的程度减轻，肾小管上皮损伤较轻，表明巨噬细胞Twist1特异性敲除可以显著减轻UUO小鼠的肾纤维化程度。进而我们研究Twist分子如何调控巨噬细胞参与肾纤维化的机制，结果发现：与野生鼠相比，敲除鼠UUO术后3天、7天及14天纤维化肾组织中M2型巨噬细胞显著减少，M1型巨噬细胞浸润数量无明显差异；进一步研究显示，下调巨噬细胞中Twist表达可以抑制巨噬细胞趋化和迁移能力。上述结果表明：Twist通过影响巨噬细胞的趋化、迁移，使其在肾组织浸润、聚集，并促进巨噬细胞向M2极化，加剧肾间质纤维化。为阐明Twist调控巨噬细胞异质性的分子机制，我们利用高通量表达谱芯片，筛选出肾脏巨噬细胞差异显著的基因- Lgals 3 （蛋白半乳糖素-3（galectin3）编码基因），报告基因及染色质共沉淀实验证实转录因子Twist直接转录激活Lgals3 表达，进一步细胞生物学实验显示galectin-3（Lgals3编码蛋白）介导Twist1调控巨噬细胞的M2极化。综上所述，巨噬细胞中 Twist1 转录激活Lgals3表达，进而调控肾组织中巨噬细胞向M2 型活化，促进肾间质纤维化进展。因此，靶向阻断巨噬细胞中 Twist可能为临床治疗慢性肾脏病提供新的靶点。