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We had previously reported that addition of putrescine to the culture medium was reported to reduce methylmercury toxicity in C17.2 neural stem cells. Here, we have examined the inhibition of methylmercury-induced cytotoxicity by putrescine using ODC1 -overexpressing C17.2 cells. We established stable ODC1 -overexpressing C17.2 cells and evaluated methylmercury-induced apoptosis by examining the TUNEL assay and cleaved caspase-3 levels. Mitochondria-mediated apoptosis was also evaluated by reduction of mitochondrial membrane potential and recruitment of Bax and Bak to the mitochondria. ODC is encoded by ODC1 gene, and putrescine levels in ODC1 -overexpressing cells were significantly higher than in control cells. Overexpression of ODC1 and addition of putrescine to the culture medium suppressed DNA fragmentation and caspase-3 activation, which are observed when apoptosis is induced by methylmercury. Moreover, mitochondrial dysfunction and reactive oxygen species (ROS) generation, caused by methylmercury, were also inhibited by the overexpression of ODC1 and putrescine; pretreatment with ODC inhibitor, however, promoted both ROS generation and apoptosis by methylmercury. Finally, we found that Bax and Bak, the apoptosis-promoting factors, to be increased in mitochondria, following methylmercury treatment, and the same was inhibited by overexpression of ODC1. These results suggest that overexpression of ODC1 may prevent mitochondria-mediated apoptosis by methylmercury via increase of putrescine levels. Our findings provide important clues to clarify mechanisms involved in the defense against methylmercury toxicity and suggest novel biological functions of putrescine. [ABSTRACT FROM AUTHOR]

Drug-induced acute renal failure (ARF) is a public health concern that hinders optimal drug therapy. However, pathological mechanisms of drug-induced ARF remain to be elucidated. Here, we show that a pathological process of drug-induced ARF is mediated by proinflammatory cross-talk between kidney tubular cells and macrophages. Both polymyxin B and colistin, polypeptide antibiotics, frequently cause ARF, stimulated the ERK and NF-κB pathways in kidney tubular cells, and thereby upregulated M-CSF and MCP-1, leading to infiltration of macrophages into the kidneys. Thereafter, the kidney-infiltrated macrophages were exposed to polypeptide antibiotics, which initiated activation of the NLR family pyrin domain containing 3 (NLRP3) inflammasome. Interestingly, blockade of the NLRP3 activation clearly ameliorated the pathology of ARF induced by polypeptide antibiotics, suggesting that a combination of the distinct cellular responses to polypeptide antibiotics in kidney tubular cells and macrophages plays a key role in the pathogenesis of colistin-induced ARF. Thus, our results provide a concrete example of how drugs initiate ARF, which may give insight into the underlying pathological process of drug-induced ARF., (Copyright © 2024 by The American Association of Immunologists, Inc.)

trans-Fatty acids (TFAs) are unsaturated fatty acids with at least one carbon-carbon double bond in trans configuration. TFA consumption has been epidemiologically associated with neurodegenerative diseases (NDs) including Alzheimer's disease. However, the underlying mechanisms of TFA-related NDs remain unknown. Here, we show a novel microglial signaling pathway that induces inflammation and cell death, which is dramatically enhanced by elaidic acid (EA), the most abundant TFA derived from food. We found that extracellular ATP, one of the damage-associated molecular patterns (DAMPs) leaked from injured cells, induced activation of the apoptosis signal-regulating kinase 1 (ASK1)-p38 pathway, which is one of the major stress-responsive mitogen-activated protein (MAP) kinase signaling pathways, and subsequent caspase-3 cleavage and DNA ladder formation (hallmarks of apoptosis) in mouse microglial cell lines including BV2 and MG6 cells. Furthermore, we found that in these microglial cell lines, EA, but not its cis isomer oleic acid, facilitated extracellular ATP-induced ASK1/p38 activation and apoptosis, which was suppressed by pharmacological inhibition of either p38, reactive oxygen species (ROS) generation, P2X purinoceptor 7 (P2X 7 ), or Ca 2+ /calmodulin-dependent kinase II (CaMKII). These results demonstrate that in microglial cells, extracellular ATP induces activation of the ASK1-p38 MAP kinase pathway and ultimately apoptosis downstream of P2X 7 receptor and ROS generation, and that EA promotes ATP-induced apoptosis through CaMKII-dependent hyperactivation of the ASK1-p38 pathway, in the same manner as in macrophages. Our study may provide an insight into the pathogenesis of NDs associated with TFAs.

Aim: To address the effect of heat-shock protein 90 (HSP90) inhibitors on the release of the hepatitis C virus (HCV), a cell culture-derived HCV (JFH1/HCVcc) from Huh-7 cells was examined., Methods: We quantified both the intracellular and extracellular (culture medium) levels of the components (RNA and core) of JFH-1/HCVcc. The intracellular HCV RNA and core levels were determined after the JFH1/HCVcc-infected Huh-7 cells were treated with radicicol for 36 h. The extracellular HCV RNA and core protein levels were determined from the medium of the last 24 h of radicicol treatment. To determine the possible role of the HSP90 inhibitor in HCV release, we examined the effect of a combined application of low doses of the HSP90 inhibitor radicicol and the RNA replication inhibitors cyclosporin A (CsA) or interferon. Finally, we statistically examined the combined effect of radicicol and CsA using the combination index (CI) and graphical representation proposed by Chou and Talalay., Results: We found that the HSP90 inhibitors had greater inhibitory effects on the HCV RNA and core protein levels measured in the medium than inside the cells. This inhibitory effect was observed in the presence of a low level of a known RNA replication inhibitor (CsA or interferon-α). Treating the cells with a combination of radicicol and cyclosporin A for 24 h resulted in significant synergy (CI < 1) that affected the release of both the viral RNA and the core protein., Conclusion: In addition to having an inhibitory effect on RNA replication, HSP90 inhibitors may interfere with an HCV replication step that occurs after the synthesis of viral RNA, such as assembly and release.

To elucidate the mechanism of toxicity of methylmercury (MeHg), we searched for factors that determine the sensitivity of yeast cells to MeHg and found that overexpression of Cdc34 or Rad23, both proteins related to the ubiquitin-proteasome (UP) system, induces resistance to MeHg toxicity. The acquisition of resistance to MeHg in Cdc34-overexpressing yeast cells requires the ubiquitin-conjugating activity of Cdc34 and the proteolytic activity of proteasomes. Therefore, it seems likely that certain as-yet-unidentified proteins that increase MeHg toxicity might exist in cells and that the toxicity of MeHg might be reduced by the enhanced degradation of such proteins through the UP system when Cdc34 is overexpressed. Unlike Cdc34, Rad23 suppresses the degradation of ubiquitinated proteins by proteasomes. This activity of Rad23 might be involved in the acquisition of resistance to MeHg toxicity when Rad23 is overexpressed. Overexpression of Rad23 might induce resistance to MeHg by suppressing the degradation of proteins that reduce the MeHg toxicity. Moreover, when we overexpressed Cdc34 in normal and Rad23-defective yeasts, resistance to MeHg was enhanced to almost the same extent in both lines of yeast cells. Thus it is possible that the binding of Rad23 to ubiquitinated proteins might be regulated by a mechanism that involves the recognition of substrate proteins and that the functions of Rad23 might not affect the protein-degradation system in which Cdc34 is involved. Many proteins that reduce or enhance MeHg toxicity and are ubiquitinated might exist in cells. The UP system and related proteins might determine the extent of MeHg toxicity by regulating the cellular concentrations of these various proteins.

The mechanism responsible for the toxic effects of methylmercury (MeHg), an important environmental pollutant, is poorly understood. We have identified a gene, CDC34, that confers resistance to MeHg in Saccharomyces cerevisiae by screening a yeast genomic DNA library. CDC34 encodes a ubiquitin-conjugating enzyme, Cdc34, which is involved in ubiquitin-dependent proteolysis. Overexpression of Cdc34 results in significant resistance to MeHg both in yeast and human cells, and it increases the cellular level of ubiquitinated proteins. The ubiquitin-conjugating activity of Cdc34 is essential for the Cdc34-mediated resistance to MeHg, and the protective effect of the overexpression of Cdc34 is depressed by inhibition of proteasome activity. Our results support the hypothesis that MeHg induces the cellular accumulation of a certain protein(s) that causes cell damage and that this protein(s) is degraded after its ubiquitination in proteasomes.