Chronic Alcohol Exposure of Cells Using Controlled Alcohol-Releasing Capillaries

Abstract

Alcohol is one of the main causes of liver diseases such as fatty liver, alcoholic hepatitis, and chronic hepatitis with liver fibrosis or cirrhosis. To reproduce the conditions of alcohol-induced liver diseases and to identify the disease-causing mechanisms at the cellular level, several methods have been used to expose the cells to ethanol. As ethanol evaporates easily, it is difficult to mimic chronic alcohol exposure conditions at the cellular level. In this study, we developed a glass capillary system containing ethanol, which could steadily release ethanol from the polyethylene tubing and hydrogel portion at both sides of the capillary. The ethanol-containing capillary could release ethanol in the cell culture medium for up to 144 h, and the concentration of ethanol in the cell culture medium could be adjusted by controlling the number of capillaries. A long-term exposure to ethanol by the capillary system led to an increased toxicity of cells and altered the cellular physiologies, such as increasing the lipid accumulation and hepatic transaminase release in cells, as compared to the traditional direct ethanol addition method. Ethanol capillaries showed different gene expression patterns of lipid accumulation- or chronic alcoholism-related genes. Our results suggest that our ethanol-containing capillary system can be used as a valuable tool for studying the mechanism of chronic alcohol-mediated hepatic diseases at the cellular level.

Establishment of a Simple and Versatile Evaporation Compensation Model for In Vitro Chronic Ethanol Treatment: Impact on Neuronal Viability

“…Borgs et al compared the effectiveness of sealing culture plates with parafilm, tapes and a plate-sealing clamp system which was found to be a much more effective measure than parafilm or tapes in preventing evaporative loss of ethanol during treatment [39]. Kim et al reported the use of gel-filled capillary tubes as a source of ethanol for in vitro delivery in a 6-well culture format [40]. When in-culture ethanol concentrations were evaluated over a period of 144 h, a bell-shaped curve was observed with a build-up time of approximately 36 h at the front to reach a concentration between 150 and 200 mM that lasted for approximately 48 h followed by a linear decline to nearly 0 mM by 144 h [40].…”

“…Kim et al reported the use of gel-filled capillary tubes as a source of ethanol for in vitro delivery in a 6-well culture format [40]. When in-culture ethanol concentrations were evaluated over a period of 144 h, a bell-shaped curve was observed with a build-up time of approximately 36 h at the front to reach a concentration between 150 and 200 mM that lasted for approximately 48 h followed by a linear decline to nearly 0 mM by 144 h [40].…”

Ethanol‐exposed lung fibroblasts cause airway epithelial barrier dysfunction

“…In parallel, the published literature shows that investigators have used a variety of models for chronic ethanol exposure in vitro. Examples include chronic ethanol exposure of 5 days for hepatocytes (Kim et al, 2021) and 60 days for intestinal epithelial cells (Wood et al, 2013). Although we previously determined that just 24 h of ethanol exposure induces fibronectin expression in LF (Roman et al, 2005), we empirically chose 2 weeks of treatment in this study to model more “chronic” ethanol exposure.…”

Mimicking chronic alcohol effects through a controlled and sustained ethanol release device

“…The most frequently employed method involves providing ethanol by changing the medium every 24 hours [ 18 ]. However, this approach results in fluctuating alcohol concentrations throughout cultivation, potentially causing medium contamination, thereby compromising the study’s validity and leading to results misinterpretation [ 17 ]. An alternative method to maintain cell exposure to ethanol is to saturate the chamber atmosphere above the culture media with ethanol or seal the culture dishes with Parafilm® [ 15 ].…”