Modulation of β-Catenin Promotes WNT Expression in Macrophages and Mitigates Intestinal Injury

Overview

Journal Cell Commun Signal

Publisher Biomed Central

Specialties Biochemistry
Cell Biology

Date 2025 Feb 11

PMID 39934819

Authors

Rishi Man Chugh

Payel Bhanja

Ryan Zitter

Sumedha Gunewardena

Rajeev Badkul

Subhrajit Saha

Affiliations

Soon will be listed here.

Abstract

Background: Macrophages are the major source of WNT ligands. However, the regulation of WNT expression in macrophages has not been studied. In the present study, we have discovered that activation of canonical β-Catenin signaling suppresses WNT expression in macrophages. EVs from these pre-conditioned macrophages promoted intestinal stem cell regeneration and mitigated intestinal injury.

Method: ChIP-seq analysis and validation studies using recombinant DNA construct expressing Luciferase reporter under WNT promoter (e.g. WNT5a and WNT9b) were conducted to demonstrate the involvement of β-Catenin in the transcriptional regulation of WNT expression. The regulatory role of β-Catenin in WNT expression in macrophages was examined by treating these cells with a Tankyrase inhibitor. In addition, the gene expressing β-Catenin was deleted in macrophages using Csf1r.iCre; Ctnnb1 mice model. Both pharmacological and genetically modulated macrophages were examined for WNT expression and activity by qPCR and TCF/LEF luciferase assay respectively. Additionally, Csf1r.iCre; Ctnnb1 mice were exposed to irradiation to compare the radiosensitivity with their wildtype littermate. Extracellular vesicles (EVs) were isolated from pre-conditioned WNT-enriched macrophages and infused in irradiated C57BL/6 and Lgr5/eGFP-IRES-Cre-ERT2; R26-ACTB-tdTomato-EGFP mice to determine the regenerative response of intestinal stem cell (ISC) and epithelial repair. Regenerative effects of EVs were also examined in mice model DSS induced colitis.

Result: ChIP-seq analysis and subsequent validation study suggested physical association of β-Catenin with WNT promoters to suppress WNT expression. Macrophage specific deletion of gene expressing β-Catenin or pharmacological inhibition of Tankyrase improves the WNT expression in macrophages several folds compared to control. Transfusion of these preconditioned macrophages or EVs from these cells delivers optimum level of morphogenic WNT to injured epithelium, activates ISC regeneration and mitigated radiation induced intestinal injury. Intestinal epithelium in Csf1r.iCre; Ctnnb1 mice also showed radioresistance compared to wild type littermate. Moreover, EVs derived from WNT enriched macrophages can mitigate intestinal injury in mice model of DSS induced acute colitis.

Conclusion: The study provides substantial evidence that macrophage-targeted modulation of canonical WNT signaling induces WNT expression in macrophages. Treatment with preconditioned macrophage derived WNT-enriched EVs can be a promising therapeutic approach against intestinal injury.

References

Saidu N, Bonini C, Dickinson A, Grce M, Inngjerdingen M, Koehl U . New Approaches for the Treatment of Chronic Graft-Versus-Host Disease: Current Status and Future Directions. Front Immunol. 2020; 11:578314. PMC: 7583636. DOI: 10.3389/fimmu.2020.578314. View

Leibowitz B, Wei L, Zhang L, Ping X, Epperly M, Greenberger J . Ionizing irradiation induces acute haematopoietic syndrome and gastrointestinal syndrome independently in mice. Nat Commun. 2014; 5:3494. PMC: 4327858. DOI: 10.1038/ncomms4494. View

Potten C, Booth C, Pritchard D . The intestinal epithelial stem cell: the mucosal governor. Int J Exp Pathol. 1997; 78(4):219-43. PMC: 2694540. DOI: 10.1046/j.1365-2613.1997.280362.x. View

Sato T, van Es J, Snippert H, Stange D, Vries R, van den Born M . Paneth cells constitute the niche for Lgr5 stem cells in intestinal crypts. Nature. 2010; 469(7330):415-8. PMC: 3547360. DOI: 10.1038/nature09637. View

Tashiro S, Nakamura M, Okano H . Regenerative Rehabilitation and Stem Cell Therapy Targeting Chronic Spinal Cord Injury: A Review of Preclinical Studies. Cells. 2022; 11(4). PMC: 8870591. DOI: 10.3390/cells11040685. View

Takahashi T, Fujishima K, Kengaku M . Modeling Intestinal Stem Cell Function with Organoids. Int J Mol Sci. 2021; 22(20). PMC: 8535974. DOI: 10.3390/ijms222010912. View

Liao Y, Smyth G, Shi W . featureCounts: an efficient general purpose program for assigning sequence reads to genomic features. Bioinformatics. 2013; 30(7):923-30. DOI: 10.1093/bioinformatics/btt656. View

Gross J, Zelarayan L . The Mingle-Mangle of Wnt Signaling and Extracellular Vesicles: Functional Implications for Heart Research. Front Cardiovasc Med. 2018; 5:10. PMC: 5850280. DOI: 10.3389/fcvm.2018.00010. View

Chassaing B, Aitken J, Malleshappa M, Vijay-Kumar M . Dextran sulfate sodium (DSS)-induced colitis in mice. Curr Protoc Immunol. 2014; 104:15.25.1-15.25.14. PMC: 3980572. DOI: 10.1002/0471142735.im1525s104. View

10.

Barker N, van Oudenaarden A, Clevers H . Identifying the stem cell of the intestinal crypt: strategies and pitfalls. Cell Stem Cell. 2012; 11(4):452-60. DOI: 10.1016/j.stem.2012.09.009. View

11.

Kim J, Jenrow K, Brown S . Mechanisms of radiation-induced normal tissue toxicity and implications for future clinical trials. Radiat Oncol J. 2014; 32(3):103-15. PMC: 4194292. DOI: 10.3857/roj.2014.32.3.103. View

12.

Sato T, Vries R, Snippert H, van de Wetering M, Barker N, Stange D . Single Lgr5 stem cells build crypt-villus structures in vitro without a mesenchymal niche. Nature. 2009; 459(7244):262-5. DOI: 10.1038/nature07935. View

13.

Rios C, Cassatt D, DiCarlo A, Macchiarini F, Ramakrishnan N, Norman M . Building the strategic national stockpile through the NIAID Radiation Nuclear Countermeasures Program. Drug Dev Res. 2014; 75(1):23-8. PMC: 8365557. DOI: 10.1002/ddr.21163. View

14.

Ahangari F, Kaminski N . WNT5a in Extracellular Vesicles - A New Frontier for Pulmonary Fibrosis. Am J Respir Crit Care Med. 2018; . DOI: 10.1164/rccm.201807-1321ED. View

15.

Bhanja P, Saha S, Kabarriti R, Liu L, Roy-Chowdhury N, Roy-Chowdhury J . Protective role of R-spondin1, an intestinal stem cell growth factor, against radiation-induced gastrointestinal syndrome in mice. PLoS One. 2009; 4(11):e8014. PMC: 2777375. DOI: 10.1371/journal.pone.0008014. View

16.

El-Kadiry A, Rafei M, Shammaa R . Cell Therapy: Types, Regulation, and Clinical Benefits. Front Med (Lausanne). 2021; 8:756029. PMC: 8645794. DOI: 10.3389/fmed.2021.756029. View

17.

Dawson P, Huxley S, Gardiner B, Tran T, McAuley J, Grimmond S . Reduced mucin sulfonation and impaired intestinal barrier function in the hyposulfataemic NaS1 null mouse. Gut. 2009; 58(7):910-9. DOI: 10.1136/gut.2007.147595. View

18.

Sato T, Stange D, Ferrante M, Vries R, van Es J, van den Brink S . Long-term expansion of epithelial organoids from human colon, adenoma, adenocarcinoma, and Barrett's epithelium. Gastroenterology. 2011; 141(5):1762-72. DOI: 10.1053/j.gastro.2011.07.050. View

19.

Stanley E . Murine bone marrow-derived macrophages. Methods Mol Biol. 2011; 5:299-302. DOI: 10.1385/0-89603-150-0:299. View

20.

Love M, Huber W, Anders S . Moderated estimation of fold change and dispersion for RNA-seq data with DESeq2. Genome Biol. 2014; 15(12):550. PMC: 4302049. DOI: 10.1186/s13059-014-0550-8. View