Glycolytic Regulation of Intestinal Stem Cell Self-Renewal and Differentiation

Chang Li; Yuning Zhou; Ruozheng Wei; Dana L. Napier; Tomoko Sengoku; Michael C. Alstott; Jinpeng Liu; Chi Wang; Yekaterina Y. Zaytseva; Heidi L. Weiss; Qingding Wang; B. Mark Evers

doi:10.1016/j.jcmgh.2022.12.012

Glycolytic Regulation of Intestinal Stem Cell Self-Renewal and Differentiation

Chang Li
, Yuning Zhou
, Ruozheng Wei
, Dana L. Napier
, Tomoko Sengoku
, Michael C. Alstott
, Jinpeng Liu
, Chi Wang
, Yekaterina Y. Zaytseva
, Heidi L. Weiss
, Qingding Wang
, B. Mark Evers

Research output: Contribution to journal › Article › peer-review

45 Scopus citations

Abstract

Background and Aims: The intestinal mucosa undergoes a continual process of proliferation, differentiation, and apoptosis. An imbalance in this highly regimented process within the intestinal crypts is associated with several intestinal pathologies. Although metabolic changes are known to play a pivotal role in cell proliferation and differentiation, how glycolysis contributes to intestinal epithelial homeostasis remains to be defined. Methods: Small intestines were harvested from mice with specific hexokinase 2 (HK2) deletion in the intestinal epithelium or LGR5⁺ stem cells. Glycolysis was measured using the Seahorse XFe96 analyzer. Expression of phospho-p38 mitogen-activated protein kinase, the transcription factor atonal homolog 1, and intestinal cell differentiation markers lysozyme, mucin 2, and chromogranin A were determined by Western blot, quantitative real-time reverse transcription polymerase chain reaction, or immunofluorescence, and immunohistochemistry staining. Results: HK2 is a target gene of Wnt signaling in intestinal epithelium. HK2 knockout or inhibition of glycolysis resulted in increased numbers of Paneth, goblet, and enteroendocrine cells and decreased intestinal stem cell self-renewal. Mechanistically, HK2 knockout resulted in activation of p38 mitogen-activated protein kinase and increased expression of ATOH1; inhibition of p38 mitogen-activated protein kinase signaling attenuated the phenotypes induced by HK2 knockout in intestinal organoids. HK2 knockout significantly decreased glycolysis and lactate production in intestinal organoids; supplementation of lactate or pyruvate reversed the phenotypes induced by HK2 knockout. Conclusions: Our results show that HK2 regulates intestinal stem cell self-renewal and differentiation through p38 mitogen-activated protein kinase/atonal homolog 1 signaling pathway. Our findings demonstrate an essential role for glycolysis in maintenance of intestinal stem cell function.

Original language	English
Pages (from-to)	931-947
Number of pages	17
Journal	CMGH
Volume	15
Issue number	4
DOIs	https://doi.org/10.1016/j.jcmgh.2022.12.012
State	Published - Jan 2023

Bibliographical note

Publisher Copyright:
© 2023 The Authors

Funding

The authors thank Dr Nissim Hay for providing HK2 f/f mice; Donna Gilbreath for manuscript preparation; and the Biospecimen Procurement and Translational Pathology Shared Resource Facility, the Flow Cytometry & Immune Monitoring Shared Resource Facility, Redox Metabolism Shared Resource Facility, and the Biostatistics and Bioinformatics Shared Resource Facility of the University of Kentucky Markey Cancer Center (supported by National Cancer Institute grant P30 CA177558 , to BME). Funding This work was supported by National Institutes of Health grants R01 DK48498 (to BME) and P30 CA177558 (to BME). The funding agencies had no role in the study design, collection, analysis, or interpretation of data. Funding This work was supported by National Institutes of Health grants R01 DK48498 (to BME) and P30 CA177558 (to BME). The funding agencies had no role in the study design, collection, analysis, or interpretation of data.The authors thank Dr Nissim Hay for providing HK2f/f mice; Donna Gilbreath for manuscript preparation; and the Biospecimen Procurement and Translational Pathology Shared Resource Facility, the Flow Cytometry & Immune Monitoring Shared Resource Facility, Redox Metabolism Shared Resource Facility, and the Biostatistics and Bioinformatics Shared Resource Facility of the University of Kentucky Markey Cancer Center (supported by National Cancer Institute grant P30 CA177558, to BME). Chang Li, PhD (Formal analysis: Equal; Investigation: Equal), Yuning Zhou, MD (Formal analysis: Equal; Investigation: Equal), Ruozheng Wei, PhD (Investigation: Equal), Dana L. Napier, PhD (Investigation: Equal), Tomoko Sengoku, PhD (Investigation: Equal), Michael C. Alstott, PhD (Investigation: Equal), Jinpeng Liu, PhD (Formal analysis: Equal), Chi Wang, PhD (Formal analysis: Equal; Methodology: Equal), Yekaterina Y. Zaytseva, PhD (Investigation: Equal), Heidi L. Weiss, PhD (Formal analysis: Equal; Methodology: Equal), Qingding Wang, PhD (Conceptualization: Equal; Formal analysis: Equal; Methodology: Equal; Visualization: Equal; Writing – original draft: Equal), B. Mark Evers, MD (Conceptualization: Equal; Formal analysis: Equal; Funding acquisition: Lead; Methodology: Equal; Visualization: Equal; Writing – original draft: Equal)

Funders	Funder number
The Markey Biostatistics and Bioinformatics Shared Resource Facility
Michael C. Alstott
National Institutes of Health (NIH)	R01 DK48498
National Childhood Cancer Registry – National Cancer Institute	P30 CA177558
University of Kentucky Markey Comprehensive Cancer Center

Keywords

Glycolysis
HK2
Intestinal Stem Cells
Metabolism

ASJC Scopus subject areas

Hepatology
Gastroenterology

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10.1016/j.jcmgh.2022.12.012

Cite this

@article{59d536ba5ee847bc9ac6925d7ab82880,

title = "Glycolytic Regulation of Intestinal Stem Cell Self-Renewal and Differentiation",

abstract = "Background and Aims: The intestinal mucosa undergoes a continual process of proliferation, differentiation, and apoptosis. An imbalance in this highly regimented process within the intestinal crypts is associated with several intestinal pathologies. Although metabolic changes are known to play a pivotal role in cell proliferation and differentiation, how glycolysis contributes to intestinal epithelial homeostasis remains to be defined. Methods: Small intestines were harvested from mice with specific hexokinase 2 (HK2) deletion in the intestinal epithelium or LGR5+ stem cells. Glycolysis was measured using the Seahorse XFe96 analyzer. Expression of phospho-p38 mitogen-activated protein kinase, the transcription factor atonal homolog 1, and intestinal cell differentiation markers lysozyme, mucin 2, and chromogranin A were determined by Western blot, quantitative real-time reverse transcription polymerase chain reaction, or immunofluorescence, and immunohistochemistry staining. Results: HK2 is a target gene of Wnt signaling in intestinal epithelium. HK2 knockout or inhibition of glycolysis resulted in increased numbers of Paneth, goblet, and enteroendocrine cells and decreased intestinal stem cell self-renewal. Mechanistically, HK2 knockout resulted in activation of p38 mitogen-activated protein kinase and increased expression of ATOH1; inhibition of p38 mitogen-activated protein kinase signaling attenuated the phenotypes induced by HK2 knockout in intestinal organoids. HK2 knockout significantly decreased glycolysis and lactate production in intestinal organoids; supplementation of lactate or pyruvate reversed the phenotypes induced by HK2 knockout. Conclusions: Our results show that HK2 regulates intestinal stem cell self-renewal and differentiation through p38 mitogen-activated protein kinase/atonal homolog 1 signaling pathway. Our findings demonstrate an essential role for glycolysis in maintenance of intestinal stem cell function.",

keywords = "Glycolysis, HK2, Intestinal Stem Cells, Metabolism",

author = "Chang Li and Yuning Zhou and Ruozheng Wei and Napier, \{Dana L.\} and Tomoko Sengoku and Alstott, \{Michael C.\} and Jinpeng Liu and Chi Wang and Zaytseva, \{Yekaterina Y.\} and Weiss, \{Heidi L.\} and Qingding Wang and Evers, \{B. Mark\}",

note = "Publisher Copyright: {\textcopyright} 2023 The Authors",

year = "2023",

month = jan,

doi = "10.1016/j.jcmgh.2022.12.012",

language = "English",

volume = "15",

pages = "931--947",

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TY - JOUR

T1 - Glycolytic Regulation of Intestinal Stem Cell Self-Renewal and Differentiation

AU - Li, Chang

AU - Zhou, Yuning

AU - Wei, Ruozheng

AU - Napier, Dana L.

AU - Sengoku, Tomoko

AU - Alstott, Michael C.

AU - Liu, Jinpeng

AU - Wang, Chi

AU - Zaytseva, Yekaterina Y.

AU - Weiss, Heidi L.

AU - Wang, Qingding

AU - Evers, B. Mark

PY - 2023/1

Y1 - 2023/1

N2 - Background and Aims: The intestinal mucosa undergoes a continual process of proliferation, differentiation, and apoptosis. An imbalance in this highly regimented process within the intestinal crypts is associated with several intestinal pathologies. Although metabolic changes are known to play a pivotal role in cell proliferation and differentiation, how glycolysis contributes to intestinal epithelial homeostasis remains to be defined. Methods: Small intestines were harvested from mice with specific hexokinase 2 (HK2) deletion in the intestinal epithelium or LGR5+ stem cells. Glycolysis was measured using the Seahorse XFe96 analyzer. Expression of phospho-p38 mitogen-activated protein kinase, the transcription factor atonal homolog 1, and intestinal cell differentiation markers lysozyme, mucin 2, and chromogranin A were determined by Western blot, quantitative real-time reverse transcription polymerase chain reaction, or immunofluorescence, and immunohistochemistry staining. Results: HK2 is a target gene of Wnt signaling in intestinal epithelium. HK2 knockout or inhibition of glycolysis resulted in increased numbers of Paneth, goblet, and enteroendocrine cells and decreased intestinal stem cell self-renewal. Mechanistically, HK2 knockout resulted in activation of p38 mitogen-activated protein kinase and increased expression of ATOH1; inhibition of p38 mitogen-activated protein kinase signaling attenuated the phenotypes induced by HK2 knockout in intestinal organoids. HK2 knockout significantly decreased glycolysis and lactate production in intestinal organoids; supplementation of lactate or pyruvate reversed the phenotypes induced by HK2 knockout. Conclusions: Our results show that HK2 regulates intestinal stem cell self-renewal and differentiation through p38 mitogen-activated protein kinase/atonal homolog 1 signaling pathway. Our findings demonstrate an essential role for glycolysis in maintenance of intestinal stem cell function.

AB - Background and Aims: The intestinal mucosa undergoes a continual process of proliferation, differentiation, and apoptosis. An imbalance in this highly regimented process within the intestinal crypts is associated with several intestinal pathologies. Although metabolic changes are known to play a pivotal role in cell proliferation and differentiation, how glycolysis contributes to intestinal epithelial homeostasis remains to be defined. Methods: Small intestines were harvested from mice with specific hexokinase 2 (HK2) deletion in the intestinal epithelium or LGR5+ stem cells. Glycolysis was measured using the Seahorse XFe96 analyzer. Expression of phospho-p38 mitogen-activated protein kinase, the transcription factor atonal homolog 1, and intestinal cell differentiation markers lysozyme, mucin 2, and chromogranin A were determined by Western blot, quantitative real-time reverse transcription polymerase chain reaction, or immunofluorescence, and immunohistochemistry staining. Results: HK2 is a target gene of Wnt signaling in intestinal epithelium. HK2 knockout or inhibition of glycolysis resulted in increased numbers of Paneth, goblet, and enteroendocrine cells and decreased intestinal stem cell self-renewal. Mechanistically, HK2 knockout resulted in activation of p38 mitogen-activated protein kinase and increased expression of ATOH1; inhibition of p38 mitogen-activated protein kinase signaling attenuated the phenotypes induced by HK2 knockout in intestinal organoids. HK2 knockout significantly decreased glycolysis and lactate production in intestinal organoids; supplementation of lactate or pyruvate reversed the phenotypes induced by HK2 knockout. Conclusions: Our results show that HK2 regulates intestinal stem cell self-renewal and differentiation through p38 mitogen-activated protein kinase/atonal homolog 1 signaling pathway. Our findings demonstrate an essential role for glycolysis in maintenance of intestinal stem cell function.

KW - Glycolysis

KW - HK2

KW - Intestinal Stem Cells

KW - Metabolism

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UR - https://www.scopus.com/pages/publications/85148349018#tab=citedBy

U2 - 10.1016/j.jcmgh.2022.12.012

DO - 10.1016/j.jcmgh.2022.12.012

M3 - Article

C2 - 36584817

AN - SCOPUS:85148349018

VL - 15

SP - 931

EP - 947

JO - CMGH

JF - CMGH

IS - 4

ER -

Glycolytic Regulation of Intestinal Stem Cell Self-Renewal and Differentiation

Abstract

Bibliographical note

Funding

Keywords

ASJC Scopus subject areas

Access to Document

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