探究PRDM5通过调控Wnt/β-catenin信号通路在克罗恩病肠上皮屏障损伤中的作用及其分子机制。

采用回顾性病例对照研究设计，从医院病历档案系统调取2021年1月至2023年12月于张家港市第一人民医院确诊的50例克罗恩病患者（L2/L3型，未使用生物制剂，克罗恩病组）及50例健康对照组（肠道息肉切除术后正常肠黏膜组织，经病理学证实无炎症改变），采用免疫组织化学H-score评估肠道组织PRDM5及active-caspase3共定位。构建三硝基苯磺酸（TNBS）诱导BALB/c小鼠克罗恩病模型（TNBS组与乙醇溶媒组各5只），评估疾病活动指数（DAI）、病理组织学评分（HAI）及PRDM5阳性细胞比例，Western blot检测β-catenin、c-Myc、Cyclin D1、TCF4表达；体外采用IFN-γ（50 ng/mL）刺激HT-29细胞建立炎症模型，分为空白对照组、IFN-γ组及siRNA-PRDM5干预组，通过Western blot检测LC3II/I、p62、active-caspase3及Bax/Bcl-2比值，流式细胞术分析凋亡率，免疫荧光观察β-catenin核转位，并进一步检测紧密连接蛋白（如claudin-2、occludin、ZO-1）表达及跨上皮电阻（TEER）和旁细胞通透性（FITC-Dextran）。

克罗恩病组PRDM5阳性率、H-score及与active-caspase3共定位率均显著高于健康对照组（均P<0.001）。TNBS模型组DAI、大体形态评分、HAI、PRDM5阳性细胞比例显著高于乙醇溶媒组（均P<0.001），且PRDM5与β-catenin、c-Myc、Cyclin D1表达以及TCF4核转位比例呈显著负相关（r值分别为－0.852、－0.774、－0.823、－0.791，均P<0.01）。敲低PRDM5后，HT-29细胞自噬活性、β-catenin表达及细胞活力显著提升（均P<0.001），凋亡率及Bax/Bcl-2比值显著降低（均P<0.001）。此外，敲低PRDM5可显著上调claudin-2、occludin、ZO-1等紧密连接蛋白的表达水平（P<0.005），并显著提高跨上皮电阻值（P<0.001），同时降低旁细胞通透性（FITC-Dextran通透率下降，P<0.001），表明其对肠上皮屏障功能具有保护作用。

PRDM5通过抑制Wnt/β-catenin通路活性，介导肠上皮细胞自噬-凋亡失衡及屏障功能损伤，其靶向干预可能为克罗恩病黏膜修复提供潜在治疗策略。

To explore the role and molecular mechanism of PRDM5 in Crohn's disease intestinal epithelial barrier injury by regulating Wnt/β-catenin signaling pathway.

A retrospective case-control study design was adopted. A total of 50 patients with Crohn's disease (L2/L3 subtypes, without biological agent administration, assigned to the Crohn's disease group) who were diagnosed in Zhangjiagang First People's Hospital from January 2021 to December 2023, as well as 50 healthy controls (with normal intestinal mucosal tissues after intestinal polypectomy and pathologically confirmed to be free of inflammatory changes), were retrieved from the hospital's medical record archive system. Immunohistochemical H-score was used to evaluate the co-localization of PRDM5 and active-caspase3 in intestinal tissues. Crohn's disease model of BALB/c mice induced by TNBS was established (5 mice in TNBS group and 5 mice in ethanol solvent group). The disease activity index (DAI), histopathological score (HAI) and the proportion of PRDM5 positive cells were evaluated, and the expressions of β-catenin, c-Myc, Cyclin D1 and TCF4 were detected by Western blotting. In vitro, HT-29 cells were stimulated by IFN-γ (50 ng/ml) and divided into blank control group, IFN-γ group and siRNA-PRDM5 intervention group. The LC3Ⅱ/Ⅰ, p62, active-caspase3 and Bax/Bcl-2 ratio were detected by Western blotting, and the apoptosis rate was analyzed by flow cytometry. The translocation of β-catenin nucleus was observed by immunofluorescence, and the expression of tight junction proteins (such as claudin-2, occludin, ZO-1), transepithelial resistance (TEER) and paracellular permeability (FITC-Dextran) were further detected.

The positive rate of PRDM5, H-score and co-location rate with active-caspase3 in Crohn's disease group were significantly higher than those in the healthy control group (all P<0.001). The proportion of DAI, gross morphological score, HAI and PRDM5 positive cells in the TNBS model group was significantly higher than that in the ethanol solvent group (all P<0.001), and PRDM5 was negatively correlated with the expression of β-catenin, c-Myc, Cyclin D1 and the nuclear translocation rate of TCF4 (r values were -0.852, -0.774, -0.823 and -0.791, all P<0.01). After knocking down PRDM5, the autophagy activity, β-catenin expression and cell viability of HT-29 cells were significantly increased (all P<0.001), while apoptosis rate and Bax/Bcl-2 ratio were significantly decreased (all P<0.001). In addition, knocking down PRDM5 could significantly increase the expression levels of claudin-2, occludin, ZO-1 and other tight junction proteins (P<0.005), significantly increase the TEER value (P<0.001), and reduce the permeability of paracellular cells (FITC-Dextran permeability decreased, P<0.001), indicating that it has a protective effect on the intestinal epithelial barrier function.

PRDM5 mediates autophagy-apoptosis imbalance and barrier function damage of intestinal epithelial cells by inhibiting Wnt/β-catenin pathway activity, and its targeted intervention may provide potential therapeutic strategies for mucosal repair of Crohn's disease.