Tumor necrosis factor alpha and palmitate simulate bovine fatty liver disease in vitro when using abattoir-derived primary bovine hepatocytes isolated by a novel nonperfusion method

Hepatic lipidosis (i.e., fatty liver) is a common periparturient disease in high-producing dairy cattle affecting nearly 50% of cows to some degree and costing an estimated 60 million dollars annually. Large animal studies are costly, labor intensive, and are not well suited to mechanistic studies. Traditionally, mechanistic studies employ in vitro methodologies, utilizing established cell lines or primary cell culture methods. However, with dairy cattle, established hepatic cell lines do not exist, and methods for primary cell culture studies typically involve complicated procedures that often utilize very young animals (typically bull calves). Several previously published papers have used abattoir-derived tissues as a source of primary cells; however, a simple method utilizing simple culture media has yet to be presented. In addition, we sought to develop a way to replicate the syndrome of fatty liver disease "in a dish" using adult cattle that should more closely represent the physiology of the periparturient dairy cow. Herein we present a non-perfusion-based method that results in robust growth and proliferation of abattoir-derived bovine hepatocytes that demonstrate lipid loading, elevated lactate dehydrogenase leakage, and cytotoxicity as demonstrated by elevated caspase 3/7 expression consistent with in vivo physiology of the periparturient dairy cow with fatty liver disease.