Neurotoxicity of ß-HgS differs from environmental mercury pollutants (MeHgCl and HgCl2) in Neuro-2a cell.

ß-HgS, differing from environmental mercury pollutants (MeHgCl and HgCl2) in chemical form, is used as traditional medicine in Asian countries for thousands of years. In this study, Neuro-2a cells were exposed to ß-HgS, MeHgCl and HgCl2 (5 µM) for 6-24 h. The cell viability of ß-HgS was higher than MeHgCl with 25.9% and 72.4% in 12 h and 24 h respectively. As the incubation time increased, MeHgCl had obvious damage to cell morphology, decreased the ratio of Bcl-2 and Bak and increased the expressions of TNF-α, IL-6 and IL-1ß significantly. Furthermore, the expressions of IL-1ß and IL-6 in HgCl2 group were increased significantly in 6 h and 24 h. The apoptotic rates in MeHgCl and HgCl2 group were respectively higher than ß-HgS with 32.2% and 7.30% in 24 h. Our findings indicate that ß-HgS is much less neurotoxicity than MeHgCl and HgCl2 in Neuro-2a cells.

Effects of ß-HgS on cell viability and intracellular oxidative stress in PC-12 cells.

Traditional Tibetan medicines containing ß-HgS have been used to treat chronic ailments for thousands of years. However, there has recently been speculation regarding the safety of these medicines due to their high mercury content. Although the toxic effect of ß-HgS has been previously investigated in vivo, the mechanism underlying the toxicity of this compound remains unclear. In this study, we investigate the mechanism of ß-HgS cytotoxicity via experiments performed on rat adrenal gland tumor cells (PC-12). Specifically, we analyze the viability and intracellular oxidative stress state of PC-12 cells treated with varying concentrations of ß-HgS. For comparison purposes, the effects of MeHgCl and HgCl2, two Hg-based compounds, on ROS generation and MDA, GSH/GSSG, Nrf2, NQO-1, and HO-1 levels are also determined. It should be noted that we used the small-molecule thiols of cell culture medium, such as cysteine, to increase the solubility of ß-HgS and prepare a ß-HgS solution to treat PC-12 cells. The obtained results show that ß-HgS inhibits cell viability at concentrations of 200-1000 ng Hg mL-1 (48 h treatment). In the concentration range of 200-600 ng Hg mL-1 (24 h treatment), the inhibitory effect of ß-HgS is stronger than that of MeHgCl; however, this trend is reversed at higher concentrations (800-1000 ng mL-1) and longer exposure times (48 h). Moreover, ß-HgS significantly promotes MDA, but has no appreciable influence on cell apoptosis and ROS generation in PC-12 cells, which suggests that its inhibitory effect on cell viability might be related to the stimulation of ROS-independent oxidative stress. Notably, ß-HgS and HgCl2 significantly increase the GSH content, GSH/GSSG ratio, NQO-1 mRNA expression, and HO-1 protein expression in PC-12 cells, indicating that the antioxidant protection against these compounds is triggered by Nrf2 activation. HPLC-AFS analysis shows that in ß-HgS and HgCl2 solutions, mercury exists in the same form of Hg2+, but the cytotoxicity of the former is greater. This is probably due to the additional oxidative damage induced by the S2- ion in ß-HgS. In conclusion, ß-HgS induces ROS-independent oxidative stress in PC-12 cells, and thus, is obviously cytotoxic. At the same time, it promotes the antioxidant capacity of cells by activating the Nrf2 pathway.

Hormesis of methylmercury-human serum albumin conjugate on N9 microglia via ERK/MAPKs and STAT3 signaling pathways.

Methylmercury (MeHg+) is an extremely toxic organomercury cation that can induce severe neurological damage. Once it enters the body, methylmercury binds to amino acids or proteins containing free sulfhydryl groups. In particular, methylmercury is known to bind with human serum albumin (HSA) in human plasma; however, the effects of methylmercury-HSA conjugate (MeHg-HSA) on the central nervous system (CNS) are not fully understood. In the present study, we used the microglial cell line N9 as the target cells to evaluate the effect of MeHg-HSA on physiological function of the CNS preliminarily. The various factors in the cell culture were monitored by MTT assay, total lactate dehydrogenase assay, ELISA, qPCR, Western blot and flow cytometry techniques. The results showed that low-dose treatment with MeHg-HSA activated N9 cells, promoting cell proliferation and total cell number, enhancing NO and intracellular Ca2+ levels, and suppressing the release of TNFα and IL1ß without cytotoxic effects; while high-dose MeHg-HSA exhibited cytotoxic effects on N9 cells, including promoting cell death and increasing the secretion of TNFα and IL1ß. These results indicate that MeHg-HSA causes hormesis in microglia N9 cells. Furthermore, ERK/MAPKs and STAT3 signaling pathways related to the hormesis of MeHg-HSA on N9 cells. In addition, low dose of MeHg-HSA might be viewed as something very close to a lowest observed adverse effect level (LOAEL) for N9 cells. These findings will be useful for investigating the hormesis mechanism of MeHg+ and exploring the specific functions of MeHg-sulfhydryl conjugates on the central nervous system.

Hormesis of mercuric chloride-human serum albumin adduct on N9 microglial cells via the ERK/MAPKs and JAK/STAT3 signaling pathways.

Mercury chloride (HgCl2), a neurotoxicant that cannot penetrate the blood-brain barrier (BBB). Although when the BBB are got damaged by neurodegenerative disorders, the absorbed HgCl2, mainly in form of Hg (II)-serum albumin adduct (Hg-HSA) in human plasma, can penetrate BBB and affect central nervous system (CNS) cells. Current study planned to evaluate the effect of Hg-HSA on the physiological function of N9 microglial cells. At low dosage (15 ng/mL) of Hg-HAS, the observed outcomes was: promoted cell propagation, Nitric Oxide (NO) and intracellular Ca2+ levels enhancement, suppressed the release of TNF-α and IL-1ß and inhibited cell proliferation. At high dosage (15 µg/mL) we observed decline in NO and intracellular Ca2+ levels, and increment in the release of TNF-α and IL-1ß. These biphasic effects are similar to hormesis, and the hormesis, in this case, was executed through ERK/MAPKs and JAK/STAT3 signaling pathways. Study of quantum chemistry revealed that Hg2+ could form stable coordination structures in both Asp249 and Cys34 sites of HSA. Although five-coordination structure in Asp249 site is more stable than four-coordination structure in Cys34 site but four-coordination structure is formed easily in-vivo in consideration of binding-site position in spatial structure of HSA.