We report a previously unidentified role of tight junction protein claudin 7 in intestinal epithelial stem cell function and regulation by using 2 independent claudin 7 knockout mouse models. Our data show that claudin 7 controls intestinal crypt stem cell survival, self-renewal, and epithelial differentiation through Wnt/b-catenin signaling.BACKGROUND & AIMS: Claudin-7 (Cldn7) is a tight junction (TJ) membrane protein located at the apical TJ and basolateral side of intestinal epithelial cells. Deletion of Cldn7 by gene targeting leads to the inflammatory bowel disease-like phenotype in mice, which includes weight loss, diarrhea, mucosa ulceration, and severe intestinal epithelial damage. In this study, we test our hypothesis that Cldn7 plays a critical role in regulating intestinal crypt stem cell functions. METHODS:Gene expression microarray, quantitative reversetranscription polymerase chain reaction, in situ hybridization, histologic examinations, immunoblotting, 3-dimensional organoid culture, and various treatments to rescue Cldn7-deficient organoid defects were conducted using global Cldn7 knockout mice and inducible, conditional Cldn7 knockout mice. RESULTS:Gene deletion of Cldn7 in intestines showed significant alteration of expression profiles with striking downregulation of intestinal crypt stem cell markers such as Olfm4, dislocated proliferative cells, and disrupted epithelial cell differentiation. In addition, the isolated Cldn7-deficient crypts where the stem cells reside were either unable to survive at all or formed defective spheroids, highlighting the functional impairment of crypt stem cells in the absence of Cldn7. Remarkably, the Cldn7-expressing organoids with buddings underwent rapid cell degeneration within days after turning off Cldn7 expression in the culture. We identified that activation of Wnt/b-catenin signaling rescued the organoid defects caused by Cldn7 deletion. CONCLUSIONS:In this study, we show that Cldn7 is indispensable in controlling Wnt/b-catenin signaling-dependent intestinal epithelial stem cell survival, self-renewal, and cell differentiation. This study could open a door to study roles of TJ proteins in stem cell regulations in other tissues and organs.
Objective: To determine if distance from our Neonatal Intensive Care Unit (NICU) follow-up clinic exacerbated risk of clinic nonattendance in high-risk groups defined by socioeconomic status or medical complexity, as geographical distance from the hospital can affect attendance rates at NICU follow-up clinics. Patients and Methods: We retrospectively identified infants born between January 2014 and June 2018, and subsequently discharged from our 50-bed level IV NICU, which serves a predominantly rural population. Patients were included in our study if they had at least one NICU clinic follow-up visit scheduled at discharge. Distance to the clinic was calculated based on family ZIP code. Mixed-effects logistic regression analysis of attendance at each scheduled visit was used to identify independent associations and interactions with distance among study covariates. Results: We included 576 patients in our study, with 74% missing at least one clinic appointment, and 30% not attending any of the three appointments. Median distance between our hospital and families was 53 km. On multivariable analysis, neither distance nor other infant or family characteristics were associated with clinic non-attendance. Only interfacility transfer had a statistically significant interaction with distance and this association only reached statistical significance for patients living furthest from our center. Conclusions: NICU follow-up is important, but clinic attendance is poor. For families living furthest away, transfers of care during the infant's hospitalization may be associated with lower completion of recommended post-discharge follow-up. Further research is needed to understand how clinics can mitigate barriers to attendance.
Tight junctions are one of the physical barriers protecting the body from invasion of intestinal lumen pathogens. Disruption of tight junctions has been reported in a variety of intestinal disorders, such as intestinal bowel disease, necrotizing enterocolitis, and colon rectal cancer. Claudin‐7 is one of the tight junction membrane proteins and is essential in maintaining intestinal epithelial integrity. However, the function of claudin‐7 in colonic epithelial homeostasis remains to be determined. Here we report that claudin‐7 is required for maintaining the balance of intestinal epithelial cell proliferation and differentiation. To dissect the role of claudin‐7 in colonic epithelium, we used our global (gCldn7−/−) and intestinal‐specific (cCldn7−/−) claudin‐7 knockout (KO) mice that are generated recently as our model systems. Using the histological analysis, we found that enterocytes and enteroendocrine cells are significantly reduced in postnatal day 4 gCldn7−/− and 2–4 months old cCldn7−/− mouse colons. Although the absolute number of Goblet cells and tuft cells are reduced in claudin‐7 KO colon, the ratio of Goblet cells or tuft cells versus the total number of epithelial cells remains unchanged. These results are confirmed by qRT‐PCR and Western blotting, suggesting that epithelial cell differentiation was significantly altered after claudin‐7 deletion. In addition, the ki67 and PCNA positive proliferative cells are significantly increased in claudin‐7 KO colons and no longer limited to the basal‐lateral crypt region, where epithelial stem cells and fate‐committed progenitor cells are located. Genome‐wide gene‐expression microarray analysis of Cldn7+/+ and gCldn7−/− colons identified a list of gene expression changes related to cell proliferation and differentiation. Notch and Hippo signaling pathway are suppressed after claudin‐7 deletion. These findings suggest that claudin‐7 may have a novel, unidentified function regulating epithelial cell differentiation and self‐renewal in mouse colon.Support or Funding InformationThis study is supported by the National Institute of Health grant DK103166.This abstract is from the Experimental Biology 2018 Meeting. There is no full text article associated with this abstract published in The FASEB Journal.
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