Histopathology is an essential biomarker for evaluating the effects of aquatic environmental factors on fish health(M et al. 2019). For instance, fluoride exposure can result in several visible histopathological changes, such as disordered kidney structure, narrow lumen, atrophy of kidney tubule with lymphocyte invasion, loss of the brush border(J et al. 2015). At the same time, loss of the brush border and cast formation in the kidney was induced after heat stress(H et al. 2022). In this study, we found that heat stress caused kidney structural disturbances and infiltration of lymphocytes in A.baerii. Severe damage occurred at 28℃, with massive infiltration of inflammatory cells in the glomeruli and interstitium, suggesting that heat stress induces kidney injury in A. baerii, and the kidney damage is more severe with increasing intensity of heat stress. Previous studies in our laboratory found that heat stress at 24℃ and 28℃ caused severe damage to the gills and intestine of A. baerii. The epithelial cells of gill filaments degenerated and hyperostosis occurred after heat stress at 24℃ and 28℃, particularly at 28℃ where necrosis appeared(S et al. 2023). Similarly, mucosal epithelial cells showed marked signs of necrosis after heat stress at 24℃ and 28℃, and shedding cells were seen in the lumen of the valve intestine accompanied by a large amount of inflammatory cell infiltration(S et al. 2022). It can be observed that under the same heat stress treatment, the damage of the kidney is relatively mild compared with other tissues, which may be related to tissue specificity. Besides, studies on the kidneys of other fish have found that sesamin can protect against fluoride-induced oxidative stress and apoptosis in the kidney of carp (Cyprinus carpio) via the JNK signaling pathway(J et al. 2015), and herbicide paraquat dichloride causes necrosis of glomeruli and damage to the collecting duct in acute exposure(IA et al. 2020).
Numerous studies have shown that apoptosis in fish cells occurs after exposure to heat stress(IM et al. 2014). Sales(CF et al. 2019) found that heat stress at the blastopore closure stage increases caspase3-mediated apoptosis in P. lineatus embryos. Liu(E et al. 2022) revealed that the level of apoptosis in the Pikeperch (Sander lucioperca) liver tissue significantly increased with the rising temperature. In this study, we found that heat stress caused apoptosis in kidney cells, and the rate of apoptosis was significantly higher at 28℃ than in the control group. Therefore, the kidney of A.baerii showed severe damage at 28℃.
Innate immunity is the earliest immune mechanism with a non-specific character, which can be induced by the external environment and then respond by affecting the expression of cytokines such as IL-1β and TNF-α(MM et al. 2020; S et al. 2013). Therefore, the expression levels of cytokines such as IL-1β and TNF-α have been used as a tool to measure the immune response in fish(M et al. 2020). Increased expression of IL-1β in A.baerii suggests that heat stress may activate inflammatory signal transduction. However, the expression of TNF-α in the kidney of A.baerii decreased under heat stress, which contradicts the study on the pikeperch liver(E et al. 2022). The reason for the different expression of TNF-α after heat stress may be that the intensity of heat stress in this study was weak and did not trigger a violent immune response, or perhaps the kidney was not the main target organ of heat stress. Notably, there was no significant difference in the expression levels of IL-1β and TNF-α mRNA after heat stress compared with the control group, indicating that heat stress has a relatively minor effect on the immune response in the kidney of A.baerii.
Heat Shock Proteins (HSPs) play a critical role in coping with adverse environmental conditions, such as heat stress and hypoxic stress(Immunology 2002; P et al. 2023; Y et al. 2021b). Multiple studies have demonstrated the key role of HSP70 in the heat resistance of fish(F and L 2020; T et al. 2022; Y et al. 2017). Moreover, HSP90 is involved in many cellular processes beyond protein folding, including DNA repair, development, and immune response, while GRP75 plays a cytoprotective role in maintaining cellular energy metabolism balance(FH et al. 2017; P et al. 2018). In this study, we investigated the mRNA expression levels of HSP70, HSP90, and GRP75 in A. baerii following heat stress. The results indicate that the expression of these genes in the kidney increased significantly after heat stress, with the maximum expression level observed at 28℃. These findings are consistent with previous studies conducted on gill, liver, and spleen(Yang et al. 2021). Notably, the expression level of GRP75 mRNA significantly increased at 28℃, unlike other HSPs. This finding suggests that GRP75 may play a more significant role than other HSPs in mitigating heat stress in A. baerii. Therefore, our results demonstrate that A. baerii produces a robust stress response at 28℃. Furthermore, our study reiterates the critical role of HSPs in responding to heat stress conditions in fishes, as seen in other species such as Paralichthys olivaceus(Y et al. 2017), Channa striatus(GK et al. 2014) and rainbow trout(NM et al. 2014).