Ethanol is a neuroteratogen and neurodegeneration is the most devastating consequence of developmental exposure to ethanol. The mechanisms underlying ethanol-induced neurodegeneration are complex. Ethanol exposure produces reactive oxygen species (ROS) which cause oxidative stress in the brain. We hypothesized that ethanol would activate autophagy to alleviate oxidative stress and neurotoxicity. Our results indicated that ethanol increased the level of the autophagic marker Map1lc3-II (LC3-II) and upregulated LC3 puncta in SH-SY5Y neuroblastoma cells. It also enhanced the levels of LC3-II and BECN1 in the developing brain; meanwhile, ethanol reduced SQSTM1 (p62) levels. Bafilomycin A1, an inhibitor of autophagosome and lysosome fusion, increased p62 levels in the presence of ethanol. Bafilomycin A1 and rapamycin potentiated ethanol-increased LC3 lipidation, whereas wortmannin and a BECN1-specific shRNA inhibited ethanol-promoted LC3 lipidation. Ethanol increased mitophagy, which was also modulated by BECN1 shRNA and rapamycin. The evidence suggested that ethanol promoted autophagic flux. Activation of autophagy by rapamycin reduced ethanol-induced ROS generation and ameliorated ethanol-induced neuronal death in vitro and in the developing brain, whereas inhibition of autophagy by wortmannin and BECN1-specific shRNA potentiated ethanol-induced ROS production and exacerbated ethanol neurotoxicity. Furthermore, ethanol inhibited the MTOR pathway and downregulation of MTOR offered neuroprotection. Taken together, the results suggest that autophagy activation is a neuroprotective response to alleviate ethanol toxicity. Ethanol modulation of autophagic activity may be mediated by the MTOR pathway.

Subchronic exposure to arsenic increases the incidence of human cancers such as skin, lung, colon, and rectal cancer. The mechanism for arsenic-induced tumorigenesis is still not clear. It is generally believed that DNA damage and genomic instability, generated by arsenic-promoted oxidative stress, account largely for this process. The major sources of reactive oxygen species (ROS) are arsenic-damaged mitochondria. Autophagy is a catabolic process functioning in turnover of long-lived proteins and dysfunctional organelles such as mitochondria. Defects of autophagy under stress conditions promote genomic instability and increase the risk of tumorigenesis. In the present study using a human bronchial epithelial cell line, BEAS-2B cells, we investigated the role of autophagy in arsenic-induced cell transformation, an important step in arsenic tumorigenesis. Our results show that subchronic arsenic exposure induces BEAS-2B cell transformation accompanied with increased ROS generation and autophagy activation. However, the patterns for ROS and autophagy alteration are different. Arsenic exposure generated a prolonged and steady increase of ROS levels, whereas the activation of autophagy, after an initial boost by arsenic administration, decreases in response to subchronic arsenic exposure, although the activity is still higher than a nontreated control. Further stimulation of autophagy increases mitochondria turnover and decreases ROS generation and arsenic-induced cell transformation. Contrarily, inhibition of autophagy activity decreases mitochondria turnover and enhances arsenic-induced ROS generation and cell transformation. In addition, the mammalian target of rapamycin signaling pathway is involved in arsenic-mediated autophagy activation. Our results suggest that autophagy is a cell self-protective mechanism against arsenic-induced cell transformation.

Chronic inflammation has been implicated as an etiologic factor in cancer, whereas autophagy may help preserve cancer cell survival but exert anti-inflammatory effects. How these phenomenas interact during carcinogenesis remains unclear. We explored this question in a human bronchial epithelial cell–based model of lung carcinogenesis that is mediated by subchronic exposure to arsenic. We found that sustained overexpression of the pro-inflammatory IL6 promoted arsenic-induced cell transformation by inhibiting autophagy. Conversely, strategies to enhance autophagy counteracted the effect of IL6 in the model. These findings were confirmed and extended in a mouse model of arsenic-induced lung cancer. Mechanistic investigations suggested that mTOR inhibition contributed to the activation of autophagy, whereas IL6 overexpression was sufficient to block autophagy by supporting Beclin-1/Mcl-1 interaction. Overall, our findings argued that chronic inflammatory states driven by IL6 could antagonize autophagic states that may help preserve cancer cell survival and promote malignant progression, suggesting a need to uncouple inflammation and autophagy controls to enable tumor progression. Cancer Res; 74(14); 3740–52. ©2014 AACR.

Neuronal loss is a prominent etiological factor for fetal alcohol spectrum disorders. The cerebellum is one of the areas in the developing central nervous system that is most sensitive to ethanol, especially during the temporal window of ethanol vulnerability. MicroRNAs are small, non-coding RNAs capable of regulating diverse cellular functions including apoptosis. Ethanol exposure has been shown to interfere with the expression of microRNAs. However, the role of microRNAs in ethanol neurotoxicity is still not clear. In the present study, we identified a particular microRNA, miR-29b, as a novel target of ethanol in the developing cerebellar granule neurons. We discovered that ethanol exposure suppressed miR-29b and induced neuronal apoptosis. Overexpression of miR-29b rendered neurons protection against ethanol-induced apoptosis. Furthermore, our data indicated that miR-29b mediated ethanol neurotoxicity through the SP1/RAX/PKR cascade. More importantly, the expression of miR-29b is developmentally regulated, which may account for, at least partially, the temporal window of ethanol sensitivity in the developing cerebellum.

Lung adenocarcinoma is one of the most common malignant tumors worldwide. The purpose of this study was to construct a stable immune gene signature for prediction of prognosis (IGSPP) and response to immune checkpoint inhibitors (ICIs) therapy in LUAD patients. Five genes were screened by weighted gene coexpression network analysis, Cox regression and LASSO regression analyses and were used to construct the IGSPP. The survival rate of the IGSPP low-risk group was higher than that of the IGSPP high-risk group. Multivariate Cox regression analysis showed that IGSPP could be used as an independent prognostic factor for the overall survival of LUAD patients. IGSPP genes were enriched in cell cycle pathways. IGSPP gene mutation rates were higher in the high-risk group. CD4 memory-activated T cells, M0 and M1 macrophages had higher infiltration abundance in the high-risk group, which was associated with poor overall survival. In contrast, the abundance of resting CD4 memory T cells, monocytes, resting dendritic cells and resting mast cells associated with a better prognosis was higher in the low-risk group. TIDE scores and the expressions of different immune checkpoints showed that patients in the high-risk IGSPP group benefited more from ICIs treatment. In short, an IGSPP of LUAD was constructed and characterized. It could be used to predict the prognosis and benefits of ICIs treatment in LUAD patients.