Hydrogen sulfide suppresses the proliferation of intestinal epithelial cells through cell cycle arrest

The pathogenesis of chronic kidney disease (CKD) is closely related to the changes in the intestinal microbiota and integrity. Our previous studies have shown the accumulation of hydrogen sulfide (H₂S)-producing bacterial family, Desulfovibrionacea, in the colon of a murine model of CKD, suggesting that the increased H₂S contributes to the impaired intestinal integrity in CKD. Here, we investigated the anti-proliferative effect of H₂S in the intestinal epithelial cells. A slow- H₂S releasing molecule GYY4137 ((p-methoxyphenyl)morpholino-phosphinodithioic acid) reduced the proliferation of Caco-2 and IEC-6 cells. Flow cytometric analysis demonstrated that GYY4137 accumulated Caco-2 cells in the S phase fraction, suggesting that H₂S arrested the cell cycle at G2 and/or M phases. The RNA sequencing analysis demonstrated that GYY4137 modulated the mRNA expression of the genes involved in the G2/M and the spindle assembly checkpoints; increased mRNA levels of Cdkn1a, Gadd45a, and Sfn and decreased mRNA levels of Cdc20, Pttg1, and Ccnb1 were observed. These alterations were confirmed by quantitative reverse transcription-polymerase chain reaction and Western blot analyses. Besides, studies exploring the MEK inhibitor indicated that MEK activation is involved in the GYY4137-mediated increase in the Sfn expression. Altogether, our data showed that H₂S reduced the proliferation of intestinal epithelial cells through transcriptional regulation in G2/M and the spindle assembly checkpoints. This may be one of the underlying mechanisms for the observed impaired intestinal integrity in CKD.