TMEM219 signaling promotes intestinal stem cell death and exacerbates colitis
Francesca D’Addio, Giovanni Amabile, Emma Assi, Anna Maestroni, Adriana Petrazzuolo, Cristian Loretelli, Ahmed Abdelasalam, Moufida Ben Nasr, Ida Pastore, Maria Elena Lunati, Vera Usuelli, Monica Zocchi, Andy Joe Seelam, Domenico Corradi, Stefano La Rosa, Virna Marin, Monique Zangarini, Marta Nardini, Stefano Porzio, Filippo Canducci, Claudia Nardini, Basset El Essawy, Manuela Nebuloni, Jun Yang, Massimo Venturini, Giovanni Maconi, Franco Folli, Silvio Danese, Gianvincenzo Zuccotti, Gianluca M. Sampietro, Sandro Ardizzone, Paolo Fiorina
Francesca D’Addio, Giovanni Amabile, Emma Assi, Anna Maestroni, Adriana Petrazzuolo, Cristian Loretelli, Ahmed Abdelasalam, Moufida Ben Nasr, Ida Pastore, Maria Elena Lunati, Vera Usuelli, Monica Zocchi, Andy Joe Seelam, Domenico Corradi, Stefano La Rosa, Virna Marin, Monique Zangarini, Marta Nardini, Stefano Porzio, Filippo Canducci, Claudia Nardini, Basset El Essawy, Manuela Nebuloni, Jun Yang, Massimo Venturini, Giovanni Maconi, Franco Folli, Silvio Danese, Gianvincenzo Zuccotti, Gianluca M. Sampietro, Sandro Ardizzone, Paolo Fiorina
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Research Article Gastroenterology

TMEM219 signaling promotes intestinal stem cell death and exacerbates colitis

Abstract

Mechanisms by which mucosal regeneration is abrogated in inflammatory bowel disease (IBD) are still under investigation, and a role for an intestinal stem cell (ISC) defect is now emerging. Herein, we report an abnormal ISC death that occurs in Crohn’s disease, which exacerbates colitis, limits ISC-dependent mucosal repair, and is controlled through the death factor Transmembrane protein 219 (TMEM219). Large alterations in TMEM219 expression were observed in patients with Crohn’s disease, particularly in those with active disease and/or those who were nonresponders to conventional therapy, confirming that TMEM219 signaling is abnormally activated and leads to failure of the mucosal regenerative response. Mechanistic studies revealed a proapoptotic TMEM219-mediated molecular signature in Crohn’s disease, which associates with Caspase-8 activation and ISC death. Pharmacological blockade of the IGFBP3/TMEM219 binding/signal with the recombinant protein ecto-TMEM219 restored the self-renewal abilities of miniguts generated from patients with Crohn’s disease in vitro and ameliorated DSS-induced and T cell-mediated colitis in vivo, ultimately leading to mucosal healing. Genetic tissue-specific deletion of TMEM219 in ISCs in newly generated TMEM219fl/flLGR5cre mice revived their mucosal regenerative abilities both in vitro and in vivo. Our findings demonstrate that a TMEM219-dependent ISC death exacerbates colitis and that TMEM219 blockade reestablishes intestinal self-renewal properties in IBD.

Authors

Francesca D’Addio, Giovanni Amabile, Emma Assi, Anna Maestroni, Adriana Petrazzuolo, Cristian Loretelli, Ahmed Abdelasalam, Moufida Ben Nasr, Ida Pastore, Maria Elena Lunati, Vera Usuelli, Monica Zocchi, Andy Joe Seelam, Domenico Corradi, Stefano La Rosa, Virna Marin, Monique Zangarini, Marta Nardini, Stefano Porzio, Filippo Canducci, Claudia Nardini, Basset El Essawy, Manuela Nebuloni, Jun Yang, Massimo Venturini, Giovanni Maconi, Franco Folli, Silvio Danese, Gianvincenzo Zuccotti, Gianluca M. Sampietro, Sandro Ardizzone, Paolo Fiorina

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Figure 6

TMEM219 genetic deletion in ISCs ameliorates acute colitis in vivo.

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TMEM219 genetic deletion in ISCs ameliorates acute colitis in vivo. (A)....
(A). Genetic approach used to generate the Tmem219fl/fl EGFP-Lgr5cre mouse, namely the ISC-Tmem219–/– mouse. (B–D). Flow plot and bar graph quantifying TMEM219 protein (B and C) and mRNA (D) expression in EpCam+EGFP-LGR5+ intestinal cells isolated from the Tmem219fl/fl EGFP-Lgr5cre mouse, in which Tmem219 was deleted through tamoxifen injection (ISC-Tmem219–/–, n = 3), compared with the ISC-B6 mice, in which Cre was not activated by tamoxifen injection (n = 3). (E and F). Normalized mRNA expression of Lgr5 and Casp8 in colons of ISC-Tmem219–/– (n = 4) compared with ISC-B6 mice (n = 3). (G). Bar graphs showing ex vivo–generated 8-day miniguts from crypts of ISC-Tmem219–/– (n = 4) and of ISC-B6 controls (n = 5) cultured with/without IGFBP3 (50 ng/mL). (H). Experimental design of the DSS acute prevention model conducted in ISC-Tmem219–/– mice. (I–K). DAI score, colon length, and histological score quantified in ISC-Tmem219–/– mice and in the ISC-B6 control (n = 8–10) with or without treatment with oral DSS (2.5%) in the prevention acute study model described in H. (L). H&E staining of colons obtained from mice as described in I. Arrows highlight inflammation, infiltrating leukocytes. Original magnification ×10; scale bar: 200 μm. (M and N). Flow cytometric analysis of infiltrating CD45+ cells performed in colon samples of ISC-Tmem219–/– mice and of ISC-B6 control mice with or without treatment with oral DSS, (n = 5). (O and P). Ex vivo–generated 8-day miniguts from crypts of ISC-Tmem219–/– mice and of ISC-B6 control control mice with or without treatment with DSS, (n = 6–7). Original magnification ×10; scale bar: 200 μm. Mean ± SEM. 1-way ANOVA followed by Šidák’s post hoc test, 2-sided t test.