Nutritional model of steatohepatitis and metabolic syndrome in the Ossabaw miniature swine

Nonalcoholic fatty liver disease (NAFLD) is one of the most common chronic liver diseases in humans.1-4 Its histology is broadly categorized into simple steatosis and nonalcoholic steatohepatitis (NASH).1-4 Simple steatosis is generally believed to be benign and is histologically characterized by macrovesicular steatosis without additional signs of liver injury. However, NASH is considered to be a progressive condition and can cause advanced fibrosis, cirrhosis, and liver failure.1-4 Histologically, it is characterized by macrovesicular steatosis along with varying degrees of lobular inflammation, balloon degeneration, and pericellular fibrosis.5,6 It remains unclear why some patients exhibit only simple steatosis whereas others with a comparable risk profile exhibit steatohepatitis. A two-hit hypothesis was previously postulated as a conceptual framework for understanding the pathogenesis of NASH,7 in which a first hit leads to the development of steatosis, and subsequently one or more second hits lead to the development of steatohepatitis. However, the validity of this oversimplified concept and the very relevance of macrovesicular steatosis in the development of steatohepatitis have been questioned recently.8,9 Ossabaw Island, off the coast of Georgia near Savannah, is home to a feral breed of swine that were left by Spaniards nearly 500 years ago (Ossabaw pigs).10 In the wild, Ossabaw pigs are typically miniature in size and exhibit a thrifty genotype, which allows them to store large amounts of fat during feasting and to survive long periods of famine.10 To protect the island’s loggerhead turtles, the Georgia Department of Natural Resources had adopted a policy in 2000 to eradicate all of its Ossabaw pigs.10 Recognizing the scientific value of these pigs, one of the authors (M.S.) led an expedition that trapped and exported 26 disease-free animals to the mainland.10 Sturek and his colleagues from Indiana and Purdue Universities have subsequently established a breeding colony and published a number of seminal studies to show that Ossabaw pigs serve as an excellent model for investigating metabolic syndrome, progression to type 2 diabetes, and long-term complications including coronary artery disease.10,11 In general, Ossabaw pigs when fed high-fat and high-calorie diets develop obesity, insulin resistance, glucose intolerance, dyslipidemia, and hypertension—in other words, metabolic syndrome—in a relatively consistent fashion.10,11 Because obesity and insulin resistance are known risk factors for NAFLD, and the entire constellation of components of metabolic syndrome is so closely linked to human NAFLD, we examined the histology of frozen liver samples from 38 pigs that participated in two previous experiments. In these experiments, Ossabaw pigs consumed for 55 weeks either standard chow or an excess calorie atherogenic diet composed of 6% to 8% kcal from protein, 19% kcal from complex carbohydrates, and 46% to 75% kcal from hydrogenated soy bean oil (predominantly trans fatty acids) and 2% cholesterol and 0.7% cholate by weight. All animals were sacrificed at the conclusion of the study, and as published previously, animals receiving atherogenic diet exhibited marked metabolic syndrome.12,13 However, to our surprise, despite these striking metabolic abnormalities, pigs on the atherogenic diet had normal serum liver biochemistries and normal liver histology (data not shown). This indicated that hydrogenated soybean oil–containing atherogenic diets without fructose do not cause liver injury in Ossabaw pigs. Therefore, we conducted a study in which Ossabaw pigs were subjected to different diets for 24 weeks, and our primary objective was to induce liver injury mimicking human NASH in association with metabolic syndrome.