Histological study of gonadal tissues of adult Artemia salina (Linnaeus 1758) and immunohistochemistry by Caspase 3 and HSP70 to detect specific apoptosis markers on gonadal tissues after exposure to TBTCl.

Background: Several types of research have been recently carried out on the biological effects of TBTs, including investigations of genitals in invertebrates in response to exposure to TBTs in marine water. Aim: The objective of this research was to investigate the acute effects of tributyltin chloride (TBTCl) on gonads in the adult stage of Artemia salina by use normal histology and immunohistochemistry (IHC) (Caspase 3 and HSP70) to see specific apoptosis markers. Methods: After exposure of A. salina to different concentrations of TBTCl (25, 50, 100, 200, and 300 ng.l-1), 50 adult A. salina (25 male and 25 female) were selected randomly from each concentration to histologically study the gonads. The gonad tissue was sectioned (5 µm) and some slides were stained with hematoxylin and eosin and others were stained with IHC avidin-biotin complex, and were examined under a light microscope. Results: The results showed significant differences (p < 0.05) in histological lesions between different concentrations of TBTCl. The histological lesions in the testis and ovary section were undifferentiated cells, degenerating yolk globules, and follicle cells enveloping the oocyte which was then compared with control tissue, and these effects were found to be increased in females more than in males with the highest concentration of TBTCl. Immunohistochemistry (IHC) showed that positive immunostaining was observed in the testis and ovary as brownish deposits to Caspase 3 and HSP70 antibody after exposure to TBTCl, while the testis and ovary section in control tissue had no immunoreactivity to Caspase 3 and HSP70 antibody; these effects were profoundly increased with the highest concentration of TBTCl in females more than in males. Finally, the histological lesions and IHC (Caspase 3 and HSP70) revealed that the apoptosis and immune system stress of A. salina gonad tissue damage in females were more sensitive to TBTCl toxicity as compared to white males. Conclusion: In general, the present study aimed to observe the effects TBTCl on A. salina gonads by using histological sections and IHC (Caspase 3 and HSP70), which were evaluated for the first time and have been proven to possess an important function in apoptosis marker and immune system stress in Artemia. Finally, the specific mechanisms through which TBTCl affects A. salina Caspase 3 and HSP70 expression need further investigation.

Effects of tributyltin chloride on cell structures of epithelial layer in different stages of Artemia salina (Linnaeus, 1758).

Background: Tributylin chloride (TBTCl) has been demonstrated to be acutely toxic to aquatic organisms. Aim: This study was conducted to investigate the effects of TBTCl on epithelial cell of gut Artemia salina in different stages (Nauplii, Juveniles, and Adults). Methods: Samples of A. salina used were cultivated in incubators for hatching. Nauplii were harvested at 24 hours of age, while the juveniles and adults were harvested at 21 and 35 days of age, respectively. These three stages of A. salina were exposed to different concentrations of TBTCl (1 ng.L-1 to 500 ng.L-1) for 24 hours. For nauplii, juveniles, and adults, 100 individuals were exposed, and those that survived in the exposure test were harvested for histological analysis. Results: The histological examinations revealed significant differences (p < 0.05) in type of lesions associated with different TBTCl concentrations and at different stages. The predominant lesions associated with different stages and different concentrations of TBTCl were epithelial cell necroasis, degeneration, cell loss, disruption, piknosis, and submucosal necrosis. Cell scoring was a significant difference (p < 0.05) between the groups of different TBTCl concentrations and different life stages. Conclusion: Overall, in this study, the generality of the lesion scores showed that the adults are relatively more susceptible to the effects of TBTCl compared to the juvenile and the nauplii.