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(2012) The role of AKT1 and autophagy in the protective effect of hydrogen sulphide against hepatic ischemia/reperfusion injury in mice, Autophagy, 8:6, 954-962, DOI: 10.4161/auto.19927 To link to this article: https://doi.org/10.4161/auto.19927 Do not distribute.The role of AKT1 and autophagy in the protective effect of hydrogen sulphide against hepatic ischemia/reperfusion injury in mice Keywords: hydrogen sulphide, liver, ischemia-reperfusion injury, autophagy, mouseAbbreviations: I/R, ischemia-reperfusion; H 2 S, hydrogen sulphide; A/R, anoxia/reoxygenation; NaHS, sodium hydrosulfide; GPT, glutamic-pyruvate transaminase/alanine aminotransferase; GOT1, glutamic-oxaloacetic transaminase 1, soluble/aspartate aminotransferase; TNF, tumor necrosis factor(-a); IL6, interleukin 6; PtdIns3K, phosphatidylinositol 3-kinase; LC3, microtubule-associated protein 1 light chain 3; 3MA, 3-methyladenine; TUNEL, TdT-mediated dUTP nick-end labeling;ELISA, enzyme-linked immunosorbent assayHydrogen sulphide (H 2 S) exerts a protective effect in hepatic ischemia-reperfusion (I/R) injury. However, the exact mechanism of H 2 S action remains largely unknown. This study was designed to investigate the role of the PtdIns3K-AKT1 pathways and autophagy in the protective effect of H 2 S against hepatic I/R injury. Primary cultured mouse hepatocytes and livers with or without NaHS (a donor of H 2 S) preconditioning were exposed to anoxia/reoxygenation (A/R) and I/R, respectively. In certain groups, they were also pretreated with LY294002 (AKT1-specific inhibitor), 3-methyladenine (3MA, autophagy inhibitor) or rapamycin (autophagy enhancer), alone or simultaneously. Cell viability, expression of P-AKT1, T-AKT1, LC3 and BECN1 were examined. The severity of liver injury was measured by the levels of serum aminotransferase and inflammatory cytokine, apoptosis and histological examination. GFP-LC3 redistribution and transmission electron microscopy were used to test the activity of autophagy. H 2 S preconditioning activated PtdIns3K-AKT1 signaling in hepatocytes. LY294002 could abolish the AKT1 activation and attenuate the protective effect of H 2 S on hepatocytes A/R and hepatic I/R injuries. H 2 S suppressed hepatic autophagy in vitro and in vivo. Further reducing autophagy by 3MA also diminished the protective effect of H 2 S, while rapamycin could reverse the autophagy inhibitory effect and enhance the protective effect of H 2 S against hepatocytes A/R and hepatic I/R injuries, consequently. Taken together, H 2 S protects against hepatocytic A/R and hepatic I/R injuries, at least in part, through AKT1 activation but not autophagy. An autophagy agonist could be applied to potentiate this hepatoprotective effect by reversing the autophagy inhibition of H 2 S.

Methylated SEPT9 showed relatively low sensitivity in detecting early stage colorectal cancer (CRC) and advanced adenomas (AA) in plasma. Combination of multiple biomarkers was an effective strategy to improve sensitivity in early stage cancer diagnosis and screening. A new qPCR‐based assay combining the detection of methylated SEPT9 and SDC2 (ColoDefense test) was used. Methylation statuses of SEPT9 and SDC2 were examined in 40 sets of cancer tissues and paired adjacent tissues, 10 adenomatous polyps and 3 hyperplastic polyps (HP). Then evaluated with 384 plasma samples, including 117 CRC patients, 23 AA patients, 78 small polyps patients, and 166 normal individuals. The limit of detection of ColoDefense was about 25 pg per reaction. Both SEPT9 and SDC2 were shown by ColoDefense to be heavily methylated in CRC tissues when compared to paired paracancerous tissues and HP (P < .01). The sensitivities for detecting AA and stage I CRC by plasma SEPT9 methylation alone were 12.1% and 65.0%, and those by plasma SDC2 methylation alone were 43.5% and 55.0%. In comparison, the sensitivities to detect AA and stage I CRC by ColoDefense improved to 47.8% and 80.0%. The overall sensitivity of ColoDefense in detecting CRC was 88.9% (95% CI: 81.4%‐93.7%) with a specificity of 92.8% (95% CI: 87.4%‐96.0%). Detection of the combinatorial biomarker of methylated SEPT9 and/or SDC2 is a powerful, convenient and highly effective strategy for early CRC screening with high sensitivity and specificity.

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