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DRG1 Maintains Intestinal Epithelial Cell Junctions and Barrier Function by Regulating RAC1 Activity in Necrotizing Enterocolitis

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Abstract

Background

An immature intestine is a high-risk factor for necrotizing enterocolitis (NEC), which is a serious intestinal disease in newborns. The regulation of developmentally regulated GTP-binding protein 1 (DRG1) during organ development suggests a potential role of DRG1 in the maturation process of the intestine.

Aim

To illustrate the function of DRG1 during the pathogenesis of NEC.

Methods

DRG1 expression in the intestine was measured using immunohistochemistry and q-PCR. Immunoprecipitation coupled with mass spectrometry was used to identify the interacting proteins of DRG1. The biological functions of the potential interactors were annotated with the Database for Annotation, Visualization and Integrated Discovery. Caco2 and FHs74Int cells with stable DRG1 silencing or overexpression were used to investigate the influence of DRG1 on cell junctions and intestinal barrier permeability and to elucidate the downstream mechanism.

Results

DRG1 was constitutively expressed during the intestinal maturation process but significantly decreased in the ileum in the context of NEC. Protein interaction analysis revealed that DRG1 was closely correlated with cell junctions. DRG1 deficiency destabilized the E-cadherin and occludin proteins near the cell membrane and increased the permeability of the epithelial cell monolayer, while DRG1 overexpression prevented lipopolysaccharide-induced disruption of E-cadherin and occludin expression and cell monolayer integrity. Further investigation suggested that DRG1 maintained cell junctions, especially adherens junctions, by regulating RAC1 activity, and RAC1 inhibition with NSC23766 attenuated intestinal injury and led to improved barrier integrity in experimental NEC.

Conclusions

Our findings illustrate the mechanism underlying the effect of DRG1 deficiency on epithelial cell permeability regulation and provide evidence supporting the application of RAC1 inhibitors for protection against NEC.

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Funding

This work was supported by the Shanghai Pu Jiang Program (19PJ1409000), a grant from Shanghai Jiao Tong University (YG2015ZD13), the Natural Science Foundation of Shanghai (18ZR1431300), and the National Natural Science Foundation of China (81503089).

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Authors and Affiliations

  1. Department of General Surgery, Shanghai Children’s Hospital, Shanghai Jiaotong University, Shanghai, 200040, China

    Li Lu, Weijue Xu, Jiangbin Liu, Liping Chen, Qingfeng Sheng & Zhibao Lv

  2. Department of Clinical Laboratory, Shanghai Children’s Hospital, Shanghai Jiaotong University, Shanghai, 200040, China

    Shaohua Hu

  3. Clinical Pharmacy Laboratory, Chinese PLA General Hospital, Beijing, 100853, China

    Minghua Zhang

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  1. Li Lu
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Contributions

All authors participated in the acquisition of data and the revision of the manuscript. All authors read and approved the final manuscript. LL conducted all the studies, analyzed the data, and drafted the manuscript. ZL was responsible for the conception and design of the study and revised the manuscript critically for important intellectual content. WX and JL supplied the clinical samples and information. LC and SH helped to construct the NEC mouse model and performed the HE and IHC assays. QS and MZ participated in the data collection and analysis.

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Correspondence to Zhibao Lv.

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Lu, L., Xu, W., Liu, J. et al. DRG1 Maintains Intestinal Epithelial Cell Junctions and Barrier Function by Regulating RAC1 Activity in Necrotizing Enterocolitis. Dig Dis Sci 66, 4237–4250 (2021). https://doi.org/10.1007/s10620-020-06812-3

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