Krüppel-like factor 5 activates chick intestinal stem cell and promotes mucosal repair after impairment

ABSTRACT

The mucosal renewal, which depends on the intestinal stem cell (ISC) activity, is the foundation of mucosal repairment. Importantly, activation of reserve ISCs (rISCs) plays a vital role in initiating mucosal repair after injury. However, the underlying regulatory mechanism of rISCs activation in chickens remains unclear. In this study, immediately after lipopolysaccharide (LPS) challenge, mitochondrial morphological destruction and dysfunction appeared in the crypt, accompanied by decreased epithelial secretion (decreased Muc2 mRNA abundance and LYSOZYME protein level). However, immediately after mucosal injury, the mucosal renewal accelerated, as indicated by the increased BrdU positive rate, proliferating cell nuclear antigen (PCNA) protein level and mRNA abundance of cell cycle markers (Ccnd1, Cdk2). Concerning the ISCs activity, during the early period of injury, there appeared a reduction of active ISCs (aISCs) marker Lgr5 mRNA and protein, and an increasing of rISCs marker Hopx mRNA and protein. Strikingly, upon LPS challenge, increased mRNA transcriptional level of Krüppel-like factor 5 (Klf5) was detected in the crypt. Moreover, under LPS treatment in organoids, the KLF5 inhibitor (ML264) would decrease the mRNA and protein levels of Stat5a and Hopx, the STAT5A inhibitor (AC-4-130) would suppress the Lgr5 mRNA and protein levels. Furthermore, the Dual-Luciferase Reporter assay confirmed that, KLF5 would bind to Hopx promoter and activate the rISCs, STAT5A would trigger Lgr5 promoter and activate the aISCs. Collectively, KLF5 was upregulated during the early period of injury, further activate the rISCs directly and activate aISCs via STAT5A indirectly, thus initiate mucosal repair after injury.

KEYWORDS:

• Krüppel-like factor 5
• intestinal stem cell
• repair
• chicken

Acknowledgements

This study was supported by the National Natural Science Foundation of China (No. 31972630), and the Basic Public Welfare Research Program of Zhejiang Provincial (No. LGN21C180003). We are grateful to Weidong Zeng (Zhejiang University) and the Experimental Teaching Center (College of Animal Sciences, Zhejiang University) for help in the experiments.

Disclosure statement

No potential conflict of interest was reported by the author(s).

Data availability statement

The authors confirm that the data supporting the findings of this study are available within the article and its supplementary materials. The datasets supporting the conclusions of this article are available in the NCBI BioProject no: PRJNA995058 (https://www.ncbi.nlm.nih.gov/bioproject/?term=PRJNA995058).

Author Contribution

L.Y. conducted the experiments, collected and analyzed the data, and wrote the manuscript. S.Q., G.W., X.R., D.L. and M.Z. were responsible for part of animal experiment and intestinal organoids culture. Y.M. and C.Z. were responsible for data interpretation and manuscript revision. J.L. was responsible for the conception and design of the study, analysis and interpretation of data, and final approval of the version submitted. All authors reviewed the manuscript.

Supplementary material

Supplemental data for this article can be accessed online at https://doi.org/10.1080/15384101.2023.2278938

Additional information

Funding

The work was supported by the National Natural Science Foundation of China [31972630]; Basic Public Welfare Research Program of Zhejiang Provincial [LGN21C180003].