Chen Rui, Lv Chengjie, Zhao Yun, Gu Weizhong, Zhang Luyin, Shi Bo, Tou Jingfa
Department of Neonatal Surgery, Children's Hospital Affiliated to Medical College of Zhejiang University, National Center for Clinical Medicine of Children's Health and Disease, Hangzhou, China.
Department of Pathology, Children's Hospital Affiliated to Medical College of Zhejiang University, National Center for Clinical Medicine of Children's Health and Disease, Hangzhou, China.
Front Pediatr. 2022 Jul 25;10:836128. doi: 10.3389/fped.2022.836128. eCollection 2022.
To investigate the expression and possible role of Sirtuin1 or Silent mating-type information regulation 2 homolog-1 (SIRT1) in post-necrotizing enterocolitis stricture.
The expression characteristics of SIRT1 and TGF-β1 in post-necrotizing enterocolitis stricture were detected by immunohistochemistry. The siRNA-SIRT1 was used to inhibit the expression of SIRT1 in intestinal epithelial cells-6 (IEC-6), and qRT-PCR, WB, and ELISA were utilized to detect the changes of Transforming growth factor-β1 (TGF-β1), nuclear factor (NF)-κB, tumor necrosis factor-α (TNF-α), tight junction protein-1 (ZO-1), and vascular endothelial growth factor (VEGF) expressions. The IEC-6 cell proliferation and migration ability were tested CCK8 kit and Transwell test. The expression of E-cadherin and Vimentin in cells was detected by immunofluorescence.
The CRP, IL-6, IL-10, and IFN-γ in the serum of Necrotizing enterocolitis (NEC) intestinal stenosis patients were significantly higher than the reference values. The SIRT1 protein was under-expressed and the TGF-β1 protein was overexpressed in NEC intestinal stenosis tissue. And the expression of SIRT1 was negatively correlated with TGF-β1. At the time of diagnosis of NEC, the expression of SIRT1 decreased in children with respiratory distress syndrome and CRP level increased. After inhibiting the expression of SIRT1 in IEC6 cells, the expression levels of TGF-β1, Smad3, and NF-κB were decreased, and the expression of ZO-1 was also decreased. The proliferation and migration ability of IEC6 cells was decreased significantly, and the expression of E-cadherin and Vimentin proteins in IEC6 cells did not change significantly.
Promotion of intestinal fibrosis by inflammation may be the mechanism of post-necrotizing enterocolitis stricture. SIRT1 may be a protective protein of NEC. The probable mechanism is that SIRT1 can regulate intestinal fibrosis and can protect the intestinal mucosal barrier function to participate in the process of post-necrotizing enterocolitis stricture.
探讨沉默信息调节因子1（Sirtuin1，SIRT1）在坏死性小肠结肠炎后狭窄中的表达及可能作用。
采用免疫组化法检测坏死性小肠结肠炎后狭窄组织中SIRT1和转化生长因子-β1（TGF-β1）的表达特征。利用小干扰RNA（siRNA）-SIRT1抑制肠上皮细胞-6（IEC-6）中SIRT1的表达，采用实时定量聚合酶链反应（qRT-PCR）、蛋白质免疫印迹法（WB）及酶联免疫吸附测定（ELISA）检测转化生长因子-β1（TGF-β1）、核因子（NF）-κB、肿瘤坏死因子-α（TNF-α）、紧密连接蛋白-1（ZO-1）及血管内皮生长因子（VEGF）表达的变化。使用CCK8试剂盒和Transwell实验检测IEC-6细胞的增殖和迁移能力。通过免疫荧光检测细胞中E-钙黏蛋白和波形蛋白的表达。
坏死性小肠结肠炎（NEC）肠狭窄患者血清中的C反应蛋白（CRP）、白细胞介素-6（IL-6）、白细胞介素-10（IL-10）和干扰素-γ（IFN-γ）显著高于参考值。NEC肠狭窄组织中SIRT1蛋白表达下调，TGF-β1蛋白表达上调。且SIRT1的表达与TGF-β1呈负相关。在NEC诊断时，呼吸窘迫综合征患儿的SIRT1表达降低，CRP水平升高。抑制IEC6细胞中SIRT1的表达后，TGF-β1、Smad3和NF-κB的表达水平降低，ZO-1的表达也降低。IEC6细胞的增殖和迁移能力显著降低，IEC6细胞中E-钙黏蛋白和波形蛋白的表达无明显变化。
炎症促进肠纤维化可能是坏死性小肠结肠炎后狭窄的机制。SIRT1可能是NEC的一种保护蛋白。可能机制是SIRT1可调节肠纤维化，并能保护肠黏膜屏障功能，参与坏死性小肠结肠炎后狭窄的过程。
