Whole genome sequencing identifies a deletion mutation in the unknown-functional KCNG2 from familial sick sinus syndrome.

Sick sinus syndrome (SSS) is a term used for a variety of disorders defined by abnormal cardiac impulse formation and by abnormal propagation from the heart's sinoatrial node. In this study, we present a case from a Chinese family in which two closely related individuals had the symptoms and electrocardiographic evidence of SSS. We hypothesized that multiple individuals affected by the disease in the family was an indication of its genetic predisposition, and thus performed high-throughput sequencing for the participants from the family to detect potential disease-associated variants. One of the potential variants that was identified was a KCNG2 gene variant (NC_000018.9: g.77624068_77624079del). Further bioinformatic analysis showed that the observed variant may be a pathogenic mutation. The results of protein-protein docking and whole cell patch-clamp measurements implied that the deletion variant in KCNG2 could affect its binding the KV2.1 protein, and finally affect the function of Kv channel, which is an important determinant in regulation of heartbeat. Therefore, we inferred that the variable KCNG2 gene may affect the function of Kv channel by changing the binding conformation of KCNG2 and KV2.1 proteins and then adversely affect propagation from the sinoatrial node and cardiac impulse formation by changing the action potential repolarization of heart cells. In summary, our findings suggested that the dominant KCNG2 deletion variant in the examined Chinese family with SSS may be a potential disease-associated variant.

Hepatoprotective Effect of Neoagarooligosaccharide via Activation of Nrf2 and Enhanced Antioxidant Efficacy.

Neoagarooligosaccharides (NAOS) are generated by ß-agarases, which cleave the ß-1,4 linkage in agarose. Previously, we reported that NAOS inhibited fat accumulation in the liver and decreased serum cholesterol levels. However, the hepatoprotective effect of NAOS on acute liver injury has not yet been investigated. Thus, we examined whether NAOS could activate nuclear factor (NF)-E2-related factor 2 (Nrf2)-antioxidant response element (ARE) and upregulates its target gene, and has hepatoprotective effect in vivo. In hepatocytes, phosphorylation and subsequent nuclear translocation of Nrf2 are increased by treatment with NAOS, in a manner dependent on p38 and c-Jun N-terminal kinase (JNK). Consistently, NAOS augmented ARE reporter gene activity and the antioxidant protein levels, resulting in increased intracellular glutathione levels. NAOS antagonized tert-butylhydroperoxide-induced reactive oxygen species (ROS) generation. Moreover, NAOS inhibited acetaminophen (APAP)-induced serum alanine aminotransferase (ALT) and aspartate aminotransferase (AST) and significantly decreased hepatocyte degeneration and inflammatory cell infiltration. Moreover, ROS production and glutathione depletion by APAP were reversed by NAOS. APAP-mediated apoptotic signaling pathways were also inhibited in NAOS-treated mice. Upregulalted hepatic expression of genes related to inflammation by APAP were consistently diminished by NAOS. Collectively, our results demonstrate that NAOS exhibited a hepatoprotective effect against APAP-mediated acute liver damage through its antioxidant capacity.

Effects of streptozotocin on pancreatic islet ß-cell apoptosis and glucose metabolism in zebrafish larvae.

Type 1 diabetes is characterized by an increase in blood glucose levels resulting from damage to ß cells in pancreatic islets and the consequent absolute insufficiency of insulin. Animal models of type 1 diabetes were usually established using drugs toxic to ß cells, such as streptozotocin (STZ). To assess the application of zebrafish larvae in diabetes research, we explore the effects of STZ on pancreatic islets and glucose metabolism in zebrafish larvae. STZ was microinjected into the pericardial cavity of zebrafish larvae on alternate days for three times. At 2 days after the whole series of STZ injection (12 dpf), free-glucose level in larvae tissue shows a significant increase, and the fluorescence signal in immunohistochemistry, which indicates the insulin expression, was significantly weaker compared with the solution-injected control. Obvious apoptosis signals were also observed in the location of pancreatic islet, and insulin content decreased to be undetectable in STZ-injected larvae. Gene expression level of ins decreased to half of the solution injection control and that of casp3a was upregulated by 2.20-fold. Expression level of glut2 and gck decreased to 0.312-fold and 0.093-fold, respectively. pck1 was upregulated by 2.533-fold in STZ-injected larvae. By tracking detection, we found the free-glucose level in STZ-injected larvae gradually approached the level of the solution injection control and the insulin content recovered at 6 days post-STZ injection (16 dpf). Consistent with the change of the glucose level, the regeneration rate of the caudal fin in the STZ-injected group decreased initially, but recovered and accelerated gradually finally at 8 days post-amputation (20 dpf). These results indicate the generation of a transient hyperglycemia model due to ß-cell apoptosis caused by STZ, which is abated by the vigorous regeneration ability of ß cells in zebrafish larvae.

Mechanism of isoniazid-induced hepatotoxicity in zebrafish larvae: Activation of ROS-mediated ERS, apoptosis and the Nrf2 pathway.

Isoniazid (INH) is a first-line anti-tuberculosis drug. INH has been detected in surface waters which may create a risk to aquatic organisms. In this study, the hepatotoxicity of INH was elucidated using zebrafish. The liver morphology, transaminase level, redox-related enzyme activity, reactive oxygen species (ROS) content and mRNA levels of liver injury-related genes were measured. The results showed that INH (4, 6 mM) significantly caused liver atrophy and increased levels of alanine aminotransferase (ALT) and aspartate aminotransferase (AST) in zebrafish. INH (6 mM) led to decreased catalase (CAT) activity, glutathione peroxidase (GPx) activity and glutathione (GSH) content but increased ROS and malondialdehyde (MDA) levels. Moreover, INH (6 mM) decreased expression levels of miR-122 and pparα but increased mRNA levels of ap-1 and c-jun. Furthermore, mRNA levels of factors related to endoplasmic reticulum stress (ERS) (grp78, atf6, perk, ire1, xbp1s and chop), apoptosis (bax, cyt, caspase-3, caspase-8 and caspase-9) and the Nrf2 signalling pathway (nrf2, ho-1, nqo1, gclm and gclc) were significantly upregulated. INH may act on hepatotoxicity in zebrafish by increasing ROS content, which weakens the antioxidant capacity, leading to ERS, cell apoptosis and liver injury. In addition, the Nrf2 signalling pathway is activated as a stress compensation mechanism during INH-induced liver injury, but it is not sufficient to counteract INH-induced hepatotoxicity.