MMP7在腹膜透析超滤中的作用及其机制研究

Ultra-filtration failure (UFF) is a major reason of peritoneal dialysis (PD) failure that caused re-hospitalization and associated with significant mortality. Recent studies have demonstrated that aquaporin-1 (APQ1) is closely related to PD ultra-filtration. .Matrix metalloproteinase 7 (MMP7), also known as matrilysin, is a secreted, zinc- and calcium-dependent endopeptidase that degrades a broad range of extracellular matrix substrates. Its expression is up-regulated in a variety of tumors and in regulating tissue remodeling in many organs, such as liver, lung, and kidney, after chronic injury. MMP7 regulating a diverse array of cellular processes, including extracellular matrix turnover and remodeling, cell proliferation and apoptosis, inflammation, and epithelial-mesenchymal transition. Our previous studies showed that MMP7 was well expressed in peritoneal mesothelial cells, and was significantly up-regulated after high glucose stimulated. Also, in PD patients, MMP7 was closely associated with ultra-filtration. Further studies showed that stimulated with MMP7 could significantly up-regulate AQP1 protein expression. These data suggested that MMP7 up-regulate AQP1 and ultra-filtration. .Studies had shown MAPK could up-regulate AQP1 expression in respiratory system, while down-regulate AQP1 expression in renal. MMP7 was closely associated with MAPK. Our studies also showed that MMP7 could up-regulate p38 phosphorylation in peritoneal mesothelial cells, suggesting that MMP7 might promote AQP1 expression and ultra-filtration via p38 phosphorylation..We are going to validate these hypotheses by the following studies. Firstly, we are going to detect the changes of AQP1 in MMP7 over-expression and knock-down peritoneal mesothelial cells. Secondly, we are going to examine the changes of AQP1 in MMP7 over-expression cells when inhibiting MAPK phosphorylation or MMP7 knock-down cells when activating MAPK. At last, we are going to confirm these changes in MMP7 gene knock-out mice.

腹膜透析超滤衰竭(UFF)增加住院率和死亡率，是腹膜透析失败的常见原因。水孔蛋白1（AQP1）是调控水分子的主要通道蛋白，与腹膜透析超滤密切相关。基质金属蛋白酶7（MMP7）是一种基质溶解因子，具有多种生物学功能，但其在腹膜透析中的作用不清。我们发现，MMP7在腹膜间皮细胞及腹透液中均有丰富表达，且腹透液MMP7浓度与腹透超滤呈正相关；应用MMP7刺激腹膜间皮细胞后，AQP1表达显著增加，提示MMP7上调AQP1。研究表明，MAPK磷酸化对AQP1有双向调节作用。我们应用MMP7刺激细胞后，p53磷酸化水平显著升高。因此，我们假设：MMP7通过激活MAPK，上调AQP1，调控腹透超滤。我们将分别上调、下调MMP7明确其对腹膜AQP1的作用及其机制；建立MMP7基因敲除小鼠，验证其通过上调AQP1调控UFF的机制。本研究将深入揭示腹透超滤衰竭的机制，为临床防治UFF提供新靶点。

研究背景：腹膜透析（PD）超滤衰竭与溶质清除障碍是腹膜功能损伤的主要形式，增加了患者住院率与全因死亡率，也是PD失败的常见原因，然而当前治疗手段非常有限，根本原因在于其内在机理不清。水孔蛋白1（AQP1）、钠磷共转运体（SLC34A1）是腹膜转运的主要通道蛋白，在腹膜间皮细胞稳定表达，参与腹膜转运功能，然而其调控机理仍不清。基质金属蛋白酶7（MMP7）在晚期肾脏病患者高水平表达，具有多种生物学活性。本研究旨在探讨MMP7在腹膜功能损伤中的作用与机理。.主要研究内容和重要结果、关键数据：1、检测PD患者血清与腹透液，发现MMP7在血清与腹透液中均有高水平表达，与PD超滤显著负相关，与充血性心力衰竭发生相关；上调腹膜间皮细胞MMP7，AQP1表达显著上调，细胞通透性增加，体积增大；同时ERK磷酸化水平显著上升；抑制ERK磷酸化逆转上述改变。以上提示MMP7可通过特异激活ERK，上调AQP1，改变细胞通透性，调控PD超滤；2、在PD人群，MMP7还与腹膜磷清除显著相关；上调腹膜间皮细胞MMP7上游分子β-catenine，细胞间质转分化（MMT）被激活，SLC34A1表达下降，FOXO6表达上调；抑制β-catenine则逆转上述改变；在小鼠模型得到相似结果；特异敲除小鼠腹膜间皮细胞β-catenine基因，FOXO6表达显著下降，腹膜MMT被逆转，SLC34A1表达被恢复；进一步特异敲除小鼠腹膜间皮细胞FOXO6基因，腹膜MMT程度显著下降，SLC34A1表达上升。以上结果提示，β-catenine-MMP7-FOXO6信号途径可调控腹膜MMT，抑制SCL34A1，抑制腹膜磷转运，加重腹膜功能损伤。.科学意义：我们的研究肯定了MMP7在PD患者的高水平表达，发现了MMP7通过ERK-AQP1途径调控水分子转运，通过β-cateniene-FOXO6途径调节钠磷转运及腹膜MMT的作用机制，阐明了MMP7在腹膜功能损伤中的作用与内在机理，为腹膜功能保护提供了潜在的防治靶点。另外，我们还利用队列数据，展示了PD患者血清磷与血钾控制现状，以及血磷血钾控制与患者预后的内在联系，揭示了PD患者管理现状与突出问题，为腹膜透析临床管理提供了证据与策略。