Effects of elevated temperature on prooxidant-antioxidant homeostasis and redox status in the American oyster: Signaling pathways of cellular apoptosis during heat stress

Abstract

Increasing seawater temperature affects growth, reproduction, development, and various other physiological processes in aquatic organisms, such as marine invertebrates, which are especially susceptible to high temperatures. In this study, we examined the effects of short-term heat stress (16, 22, 26, and 30 °C for 1-week exposure) on prooxidant-antioxidant homeostasis and redox status in the American oyster (Crassostrea virginica, an edible and commercially cultivated bivalve mollusk) under controlled laboratory conditions. Immunohistochemical and real-time quantitative PCR (qRT-PCR) analyses were performed to examine the expression of heat shock protein-70 (HSP70, a biomarker of heat stress), catalase (CAT, an antioxidant), superoxide dismutase (SOD, an antioxidant), dinitrophenyl protein (DNP, a biomarker of reactive oxygen species, ROS), and 3-nitrotyrosine protein (NTP, an indicator of reactive nitrogen species, RNS), in the gills and digestive glands of oysters. In situ TUNEL assay was performed to detect cellular apoptosis in tissues. Histological analysis showed an increase in mucus secretion in the gills and digestive glands of oysters exposed to higher temperatures (22, 26, and 30 °C) compared to control (16 °C). Immunohistochemical and qRT-PCR analyses showed significant increases in HSP70, DNP and NTP protein, and mRNA expressions in tissues at higher temperatures. Cellular apoptosis was also significantly increased at higher temperatures. Thus, heat-induced oxidative and nitrative stress likely occur due to overproduction of ROS and RNS. Interestingly, expression of CAT and SOD increased in oysters exposed to 22 and 26 °C, but was at or below control levels in the highest temperature exposure (30 °C). Collectively, these results suggest that elevated seawater temperatures cause oxidative/nitrative stress and induce cellular apoptosis through excessive ROS and RNS production, leading to inhibition of the antioxidant defense system in marine mollusks.