Ethanol-induced Cell Apoptosis Research Articles

Benzydamine hydrochloride ameliorates ethanol-induced inflammation in RAW 264.7 macrophages by stabilizing redox homeostasis

Objective: To evaluate the protective effect of benzydamine hydrochloride against ethanol-induced oxidative stress and inflammation in RAW 264.7 macrophages. Methods: RAW 264.7 macrophages were treated with ethanol (100 mM) and benzydamine hydrochloride (7.5 μM). The inflammatory status was confirmed by measuring pro-(TNF-α and IL-6) and anti-inflammatory (IL-10) cytokines through ELISA and RT-PCR assays. Reactive oxygen species generation and mitochondrial membrane potential were investigated to study the protective role of benzydamine hydrochloride against ethanol-induced oxidative stress. Apoptosis detection was also investigated using flow cytometry and acridine orange/ethidium bromide staining. Results: Benzydamine hydrochloride significantly decreased the secretion of TNF-α and IL-6, as well as the generation of reactive oxygen species inside the cells, thereby stabilizing the mitochondrial membrane potential and reducing DNA fragmentation. The ethanol-induced cellular necrosis was also reversed by the administration of benzydamine hydrochloride. Conclusions: Benzydamine hydrochloride ameliorates ethanol-induced cell apoptosis and inflammation in RAW macrophages.

Protective Effect and Potential Mechanism of the Traditional Chinese Medicine Shaoyao-Gancao Decoction on Ethanol-Induced Gastric Ulcers in Rats.

Background Shaoyao-Gancao decoction (SGD) is a classic prescription in traditional Chinese medicine. SGD is effective in the treatment of gastric and duodenal ulcers. However, the biological activity and possible mechanisms of SGD in the treatment of gastric ulcers have not been fully elucidated. The purpose of this study was to scientifically evaluate the protective effect and potential mechanism of SGD against ethanol-induced gastric ulcers in rats. Methods A single gavage of 10 mL/kg of 75% ethanol was used to establish a rat gastric ulcer model. A histopathological examination of the gastric tissue was performed. The levels of TNF-α, EGF, PGE2, SOD, and TBARS in gastric tissue were measured by ELISA. Cellular apoptosis in gastric tissues was assessed by TUNEL assay. The expression levels of caspase-3 and Bcl-2 were determined by immunohistochemistry. The potential mechanism of SGD in treating gastric ulcers was further studied using a network pharmacology research method. Results The gastric tissue of rats with ethanol-induced gastric ulcers had obvious injury throughout the mucosal layer, which was significantly weakened in rats treated with SGD. Furthermore, treatment with SGD significantly increased the levels of EGF, PGE2, SOD, and Bcl-2 and decreased the levels of TNF-α, TBARS, and caspase-3 in the gastric tissue of rats with ethanol-induced gastric ulcers. SGD reduced ethanol-induced cell apoptosis in gastric tissue from rats with gastric ulcers. A traditional Chinese medicine-based network pharmacology study revealed that SGD exerts its anti-gastric ulcer effect by acting on multiple pathways. Conclusions The above results indicate that SGD can improve gastric ulcers induced by ethanol. Moreover, this study demonstrated multicomponent, multitarget, and multipathway characteristics of SGD in the treatment of gastric ulcers and provided a foundation for further drug development research.

Hepato-protective effect of fucoidan extracted from acid-processed Sargassum fusiformis in ethanol-treated Chang liver cells and in a zebrafish model

In our previous study, the anticancer effect of the active fucoidan (JHCF4) isolated from acid-processed Sargassum fusiformis was evaluated. In this study, the liver-protective effects of JHCF4 against ethanol-induced Chang liver cell damage and apoptosis-related responses were investigated. Furthermore, the low cytotoxicity and high cell viability of JHCF4 against ethanol-induced cell damage, as well as its protective effect against ethanol-induced cell apoptosis, were observed via nuclear staining with Hoechst 33342 in Chang liver cells. Additionally, the treatment of the 72 h post-fertilization (hpf) zebrafish model with JHCF4 increased the ethanol-stimulated survival rates as well as decreased oxidative stress, lipid peroxidation, and cell death levels. JHCF4 was found to significantly decrease steatosis production in the 128 hpf zebrafish model by Oil Red O staining, as well as attenuate the malondialdehyde and increase the glutathione contents, compared with the untreated group. These results demonstrate that JHCF4 has a potential hepato-protective effect against ethanol-induced damage both in vitro and in vivo.

NO66 overexpression rescues ethanol-induced cell apoptosis in human AC16 cardiomyocytes by suppressing PTEN and activating the PI3K/Akt signaling.

Previously, Nucleolar protein 66 (NO66) was reported to be closely associated with alcohol exposure-induced injury. However, the role of NO66 in alcohol-induced cytotoxicity remains unclear. In this study, we explored the potential effect and mechanism of NO66 on ethanol-induced apoptosis in human AC16 cardiomyocytes. The AC16 cell lines with NO66 and phosphatase and tensin homolog (PTEN) overexpression were constructed. Cell counting kit-8 (CCK-8), lactate dehydrogenase (LDH) assay, Annexin V-FITC/PI staining, and flow cytometry were used to evaluate the cell viability, membrane damage, and apoptosis, respectively. Quantitative real-time PCR (qRT-PCR) and western blot analysis were applied to measure mRNA and protein expression. The results showed that acute ethanol exposure markedly augmented cytotoxicity and reduced NO66 level in AC16 cardiomyocytes. Overexpression of NO66 partially reversed ethanol-induced apoptosis. NO66 upregulation reversed the decrease in phosphorylation of protein kinase B (Akt) and B-cell lymphoma-2/Bcl-2-associated x (Bcl-2/Bax) ratio and the increase in PTEN, p53, and caspase-3 activity induced by ethanol treatment. Meanwhile, the application of PI3K inhibitor (LY294002) and PTEN overexpression attenuated the inhibition efficiency of NO66 on cell apoptosis. In addition, PTEN overexpression weakened the effect of NO66 on PI3K/Akt activation, without affecting the level of NO66. Our data suggested that NO66 overexpression might play an anti-apoptotic role in ethanol-induced cell injury via reducing PTEN and upregulating the PI3K/Akt pathway.

Chronic ethanol exposure induces SK-N-SH cell apoptosis by increasing N-methyl-D-aspartic acid receptor expression and intracellular calcium.

It has been identified that chronic ethanol exposure damages the nervous system, particularly neurons. There is scientific evidence suggesting that neuronal loss caused by chronic ethanol exposure has an association with neuron apoptosis and intracellular calcium oscillation is one of the primary inducers of apoptosis. Therefore, the present study aimed to investigate the inductive effects of intracellular calcium oscillation on apoptosis in SK-N-SH human neuroblastoma cells and the protective effects of the N-methyl-D-aspartic acid receptor (NMDAR) antagonist, memantine, on SK-N-SH cell apoptosis caused by chronic ethanol exposure. SK-N-SH cells were treated with 100 mM ethanol and memantine (4 µM) for 2 days. Protein expression of NR1 was downregulated by RNA interference (RNAi). Apoptosis was detected by Annexin V/propidium iodide (PI) double-staining and flow cytometry and cell viability was detected using an MTS kit. Fluorescence dual wavelength spectrophotometry was used to determine the intracellular calcium concentration and the levels of NR1 and caspase-3 were detected using western blotting. NR1 mRNA levels were also detected using qPCR. It was found that chronic ethanol exposure reduced neuronal cell viability and caused apoptosis of SK-N-SH cells, and the extent of damage in SK-N-SH cells was associated with ethanol exposure concentration and time. In addition, chronic ethanol exposure increased the concentration of intracellular calcium in SK-N-SH cells by inducing the expression of NMDAR, resulting in apoptosis, and memantine treatment reduced ethanol-induced cell apoptosis. The results of the present study indicate that the application of memantine may provide a novel strategy for the treatment of alcoholic dementia.

Protective effect of dieckol isolated from Ecklonia cava against ethanol caused damage in vitro and in zebrafish model

In the present study, the protective effects of phlorotannins isolated from Ecklonia cava against ethanol-induced cell damage and apoptosis were investigated both in vitro and in vivo. Three phlorotannin compounds, namely phloroglucinol, eckol and dieckol, were successively isolated and identified from the extract. Dieckol showed the strongest protective effect against ethanol-induced cell apoptosis in Chang liver cells, with the lowest cytotoxicity. It was observed that dieckol reduced cell apoptosis through activation of Bcl-xL and PARP, and down-regulation of Bax and caspase-3 in Western blot analyses. In the in vivo study, the protective effect of ethanol induced by dieckol was investigated in a zebrafish model. The dieckol treated group scavenged intracellural reactive oxygen species and prevented lipid peroxidation and ethanol induced cell death in the zebrafish embryo. In conclusion, dieckol isolated from E. cava might possess a potential protective effect against ethanol-induced liver diseases.

Role of Notch-1 signaling in ethanol induced PC12 apoptosis

Chronic alcoholic dementia has crucial role in progress of neurodegenerative disease and affects a large portion of our aging population. Neuronal cell apoptosis may be a contributing factor of neurodegenerative disease (ND) and Alzheimer's disease (AD). Previous researches have indicated that Notch-1 signaling pathway is related with learning and memory ability. Mutations of Notch-1 are reported to accelerate the onset of AD and PD. Despite much investigation, very little is known about the exact role of Notch-1 in ND. In our present study, we used neuronal PC12 cell line to study the regulation role of Notch-1 in ethanol-induced cell apoptosis. 3-(4,5-Dimethythiazol-2-yl)-2,5-diphenyl tetrazolium bromide (MTT) assay was used for detection of the proliferation of PC12 cells upon ethanol treatment. Changes in PC12 cell nuclear morphology was detected with Hoechst 33258/propidium iodide (PI) double-staining. Western blot was used for detecting the level of cell apoptosis-related protein. At the same time, Notch-1 signaling activity was detected through enzymatic assay and Western blot. Our results demonstrate that PC12 cell apoptosis was induced by ethanol modulated by Notch-1 signaling pathway. Keywords: Neuronal PC12 cell, neurodegenerative disease, ethanol, Notch-1