Basic Study
Copyright ©The Author(s) 2022. Published by Baishideng Publishing Group Inc. All rights reserved.
World J Gastrointest Surg. Dec 27, 2022; 14(12): 1329-1339
Published online Dec 27, 2022. doi: 10.4240/wjgs.v14.i12.1329
Hydrogen gas and preservation of intestinal stem cells in mesenteric ischemia and reperfusion
Ryo Yamamoto, Sayuri Suzuki, Koichiro Homma, Shintaro Yamaguchi, Tomohisa Sujino, Junichi Sasaki
Ryo Yamamoto, Sayuri Suzuki, Koichiro Homma, Junichi Sasaki, Department of Emergency and Critical Care Medicine, Keio University School of Medicine, Tokyo 1608582, Japan
Shintaro Yamaguchi, Division of Endocrinology, Metabolism and Nephrology, Department of Internal Medicine, Keio University School of Medicine, Tokyo 1608582, Japan
Tomohisa Sujino, Division of Gastroenterology and Hepatology, Department of Internal Medicine, Keio University School of Medicine, Tokyo 1608582, Japan
Author contributions: Yamamoto R, Suzuki S, Homma K, Yamaguchi S, and Sujino T designed the experiment; Yamamoto R, Suzuki S, and Homma K performed the experiment and analyzed the data; Sasaki J supervised the experiment; Yamamoto R and Suzuki S wrote the original manuscript; Yamamoto R, Suzuki S, and Homma K revised the manuscript; all authors reviewed the manuscript.
Institutional animal care and use committee statement: The protocol used in this study was approved by the Research Council and Animal Care and Use Committee of the Research Institute of Keio University in Tokyo, Japan (approval number 21013-0) and was performed in accordance with the guidelines for the care and use of laboratory animals established by the Japanese Pharmacological Society and the National Institutes of Health.
Conflict-of-interest statement: All the authors report no relevant conflicts of interest for this article.
Data sharing statement: Technical appendix, statistical code, and dataset available from the corresponding author at homma@keio.jp.
ARRIVE guidelines statement: The authors have read the ARRIVE guidelines, and the manuscript was prepared and revised according to the ARRIVE guidelines.
Open-Access: This article is an open-access article that was selected by an in-house editor and fully peer-reviewed by external reviewers. It is distributed in accordance with the Creative Commons Attribution NonCommercial (CC BY-NC 4.0) license, which permits others to distribute, remix, adapt, build upon this work non-commercially, and license their derivative works on different terms, provided the original work is properly cited and the use is non-commercial. See: https://creativecommons.org/Licenses/by-nc/4.0/
Corresponding author: Koichiro Homma, MD, PhD, Assistant Professor, Department of Emergency and Critical Care Medicine, Keio University School of Medicine, 35 Shinanomachi, Shinjuku-ku, Tokyo 1608582, Japan. homma@keio.jp
Received: August 25, 2022
Peer-review started: August 25, 2022
First decision: September 2, 2022
Revised: September 11, 2022
Accepted: November 7, 2022
Article in press: November 7, 2022
Published online: December 27, 2022
Processing time: 124 Days and 2 Hours
Abstract
BACKGROUND

Patients with mesenteric ischemia frequently suffer from bowel necrosis even after revascularization. Hydrogen gas has showed promising effects for ischemia-reperfusion injury by reducing reactive oxygen species in various animal and clinical studies. We examined intestinal tissue injury by ischemia and reperfusion under continuous initiation of 3% hydrogen gas.

AIM

To clarify the treatment effects and target cells of hydrogen gas for mesenteric ischemia.

METHODS

Three rat groups underwent 60-min mesenteric artery occlusion (ischemia), 60-min reperfusion following 60-min occlusion (reperfusion), or ischemia-reperfusion with the same duration under continuous 3% hydrogen gas inhalation (hydrogen). The distal ileum was harvested. Immunofluorescence staining with caspase-3 and leucine-rich repeat-containing G-protein-coupled 5 (LGR5), a specific marker of intestinal stem cell, was conducted to evaluate the injury location and cell types protected by hydrogen. mRNA expressions of LGR5, olfactomedin 4 (OLFM4), hairy and enhancer of split 1, Jagged 2, and Neurogenic locus notch homolog protein 1 were measured by quantitative polymerase chain reaction. Tissue oxidative stress was analyzed with immunostaining for 8-hydroxy-2'-deoxyguanosine (8-OHdG). Systemic oxidative stress was evaluated by plasma 8-OHdG.

RESULTS

Ischemia damaged the epithelial layer at the tip of the villi, whereas reperfusion induced extensive apoptosis of the cells at the crypt base, which were identified as intestinal stem cells with double immunofluorescence stain. Hydrogen mitigated such apoptosis at the crypt base, and the LGR5 expression of the tissues was higher in the hydrogen group than in the reperfusion group. OLFM4 was also relatively higher in the hydrogen group, whereas other measured RNAs were comparable between the groups. 8-OHdG concentration was high in the reperfusion group, which was reduced by hydrogen, particularly at the crypt base. Serum 8-OHdG concentrations were relatively higher in both reperfusion and hydrogen groups without significance.

CONCLUSION

This study demonstrated that hydrogen gas inhalation preserves intestinal stem cells and mitigates oxidative stress caused by mesenteric ischemia and reperfusion.

Keywords: Hydrogen molecule; Intestinal ischemia; Ischemia-reperfusion injury; Tissue protection; Non-operative management; Leucine-rich repeat-containing G-protein-coupled 5

Core Tip: Distal ileum of rats was observed after 60-min mesenteric artery occlusion (ischemia), 60-min reperfusion following 60-min occlusion (reperfusion), or ischemia-reperfusion with the same duration under continuous 3% hydrogen gas inhalation (hydrogen). Immunofluorescence staining with caspase-3 and leucine-rich repeat-containing G-protein-coupled 5 (LGR5) (a specific marker of intestinal stem cell) identified ischemia damaged the epithelial layer at the tip of the villi, whereas reperfusion induced extensive apoptosis of intestinal stem cells that was mitigated by hydrogen. In addition, quantitative polymerase chain reaction revealed the LGR5 expression of the tissues was higher in rats with hydrogen inhalation than in those with reperfusion injury.