UNC-CH School of Medicine, Chapel Hill-- For the first time, scientists at the University of North Carolina at Chapel Hill have used gene knockout technology to firmly identify a key molecular player in alcohol-induced liver disease . A report of the study appears October 1 in the journal Gastroenterology. The findings carry implications for targeting new treatments aimed at preventing alcohol-induced hepatitis and cirrhosis.
"Having this knock-out technology will help us develop drug therapies for this devastating disease which affects 11 million people in the United States alone," says senior study author Dr. Ronald G. Thurman, professor of pharmacology at UNC-CH School of Medicine.
The report points to tumor necrosis factor-alpha (TNF-alpha) as the liver injury culprit. This hormone-like protein is produced in the liver and cells throughout the body. An immune system cytokine, it is a central pro-inflammatory molecule.
The study offers the first solid proof that TNF-alpha figures importantly in the development of early liver injury associated with long-term alcohol consumption. Previously, higher levels of TNF-alpha were found in alcoholics with hepatitis and laboratory studies have shown that antibodies to TNF-alpha attenuated alcohol-induced liver injury. But the cytokine's role in the disease remained unclear.
"This major increase in our understanding of mechanisms of liver injury brings us one step closer to therapies for alcoholic liver disease," says Dr. Enoch Gordis, director, National Institute on Alcohol Abuse and Alcoholism, a component of the National Institutes of Health.
Thurman explains the production of TNF-alpha in the liver occurs in phagocytes called Kupffer cells. These white cells help kill bacteria , including endotoxin - pieces of raw, gram-negative bacteria that get into the liver via the intestines. Studies suggest that alcohol intake increases the gut's permeability, thus releasing increased levels of endotoxin into circulation. This, in turn, may activate Kupffer cells to release many potent cytokines, including TNF-alpha. The resultant output could lead to liver injury. The UNC study was led by Dr. Ming Yin, a research associate in Thurman's Laboratory of Hepatobiology and Toxicology. The researchers studied mice lacking receptor sites for TNF-alpha on their liver cell surfaces. These "knocked-out" receptors were TNF-R1 and TNF-R2.
Yin and his colleagues successfully adapted a feeding system to this mouse model that would allow a continuous infusion of a high-fat diet with alcohol directly into the stomach via a tube, without curtailing the animals' mobility. "The use of animals lacking TNF receptors provides a direct way to test the hypothesis that this cytokine is important in initiation and progression of alcohol-induced liver injury," the researchers state.
Compared to "wild-type" mice and those lacking TNF-R2 , the TNF-R1 knockouts fared significantly better on measures of liver injury after four weeks of continuous alcohol (ethanol) exposure. These included measures of liver enzymes, steatosis (fatty liver), and liver pathology.
"Ethanol caused severe liver injury in wild-type mice and TNF-R2 knockout mice but not in TNF-R1 knockout mice," the report states. "Moreover, the long-term enteral ethanol feeding technique we described for the first time for knockout mice provides a useful new tool for alcohol research."
Says Thurman: "The next step will be to try out our knockout technology for [liver] fibrosis. We would use the same model, just go longer with it." He adds: "Alcoholic liver disease is more common in men than women but the most sensitive group are overweight females. So we're hypothesizing that estrogen plays a role in the mechanism of pathology." In that regard, Thurman plans to apply the study's technology to estrogen receptor knockout mice.
The UNC study was supported in part by grants from the National Institute on Alcohol Abuse and Alcoholism.
Journal
GASTROENTEROLOGY