Fast and slow releasing sulphide donors engender distinct transcriptomic alterations in Atlantic salmon hepatocytes

Abstract

Hydrogen sulphide (H₂S) is a naturally occurring compound generated either endogenously or exogenously and serves both as a gaseous signalling molecule and an environmental toxicant. Though it has been extensively investigated in mammalian systems, the biological function of H₂S in teleost fish is poorly identified. Here we demonstrate how exogenous H₂S regulates cellular and molecular processes in Atlantic salmon (Salmo salar) using a primary hepatocyte culture as a model. We employed two forms of sulphide donors: the fast-releasing salt form, sodium hydrosulphide (NaHS) and the slow-releasing organic analogue, morpholin-4-ium 4-methoxyphenyl(morpholino) phosphinodithioate (GYY4137). Hepatocytes were exposed to either a low (LD, 20 µg/L) or high (HD, 100 µg/L) dose of the sulphide donors for 24 hrs, and the expression of key sulphide detoxification and antioxidant defence genes were quantified by qPCR. The key sulphide detoxification genes sulfite oxidase 1 (soux) and the sulfide: quinone oxidoreductase 1 and 2 (sqor) paralogs in salmon showed pronounced expression in the liver and likewise responsive to the sulphide donors in the hepatocyte culture. These genes were ubiquitously expressed in different organs of salmon as well. HD-GYY4137 upregulated the expression of antioxidant defence genes, particularly glutathione peroxidase, glutathione reductase and catalase, in the hepatocyte culture. To explore the influence of exposure duration, hepatocytes were exposed to the sulphide donors (i.e., LD versus HD) either transient (1h) or prolonged (24h). Prolonged but not transient exposure significantly reduced hepatocyte viability, and the effects were not dependent on concentration or form. The proliferative potential of the hepatocytes was only affected by prolonged NaHS exposure, and the impact was not concentration dependent. Microarray analysis revealed that GYY4137 caused more substantial transcriptomic changes than NaHS. Moreover, transcriptomic alterations were more marked following prolonged exposure. Genes involved in mitochondrial metabolism were downregulated by the sulphide donors, primarily in NaHS-exposed cells. Both sulphide donors influenced the immune functions of hepatocytes: genes involved in lymphocyte-mediated response were affected by NaHS, whereas inflammatory response was targeted by GYY4137. In summary, the two sulphide donors impacted the cellular and molecular processes of teleost hepatocytes, offering new insights into the mechanisms underlying H₂S interactions in fish.

Keywords: Cellular stress; Fish health; GYY4137; Hydrogen sulphide toxicity; Immunity.