Rotonya M. Carr

Reduction of TIP47 improves hepatic steatosis and glucose homeostasis in mice

Lipid droplets in the liver are coated with the perilipin family of proteins, notably adipocyte differentiation-related protein (ADRP) and tail-interacting protein of 47 kDa (TIP47). ADRP is increased in hepatic steatosis and is associated with hyperlipidemia, insulin resistance, and glucose intolerance. We have shown that reducing ADRP in the liver via antisense oligonucleotide (ASO) treatment attenuates steatosis and improves insulin sensitivity and glucose tolerance. We hypothesized that TIP47 has similar effects on hepatic lipid and glucose metabolism. We found that TIP47 mRNA and protein levels were increased in response to a high-fat diet (HFD) in C57BL/6J mice. TIP47 ASO treatment decreased liver TIP47 mRNA and protein levels without altering ADRP levels. Low-dose TIP47 ASO (15 mg/kg) and high-dose TIP47 ASO (50 mg/kg) decreased triglyceride content in the liver by 35% and 52%, respectively. Liver histology showed a drastic reduction in hepatic steatosis following TIP47 ASO treatment. The high dose of TIP47 ASO significantly blunted hepatic triglyceride secretion, improved glucose tolerance, and increased insulin sensitivity in liver, adipose tissue, and muscle. These findings show that TIP47 affects hepatic lipid and glucose metabolism and may be a target for the treatment of nonalcoholic fatty liver and related metabolic disorders.

Absence of Perilipin 2 Prevents Hepatic Steatosis, Glucose Intolerance and Ceramide Accumulation in Alcohol-Fed Mice

Background Perilipin 2 (Plin2) is a lipid droplet protein that has roles in both lipid and glucose homeostasis. An increase in Plin2 in liver is associated with the development of steatosis, glucose intolerance, and ceramide accumulation in alcoholic liver disease. We investigated the role of Plin2 on energy balance and glucose and lipid homeostasis in wildtype and Plin2 knockout (Plin2KO) mice chronically fed a Lieber-DeCarli liquid ethanol or control diet for six weeks. Methods We performed in vivo measurements of energy intake and expenditure; body composition; and glucose tolerance. After sacrifice, liver was dissected for histology and lipid analysis. Results We found that neither genotype nor diet had a significant effect on final weight, body composition, or energy intake between WT and Plin2KO mice fed alcohol or control diets. Additionally, alcohol feeding did not affect oxygen consumption or carbon dioxide production in Plin2KO mice. We performed glucose tolerance testing and observed that alcohol feeding failed to impair glucose tolerance in Plin2KO mice. Most notably, absence of Plin2 prevented hepatic steatosis and ceramide accumulation in alcohol-fed mice. These changes were related to downregulation of genes involved in lipogenesis and triglyceride synthesis. Conclusions Plin2KO mice chronically fed alcohol are protected from hepatic steatosis, glucose intolerance, and hepatic ceramide accumulation, suggesting a critical pathogenic role of Plin2 in experimental alcoholic liver disease.

Pathophysiology of lipid droplet proteins in liver diseases

Cytosolic lipid droplets (LDs) are present in most cell types, and consist of a core comprising neutral lipids, mainly triglycerides and sterol esters, surrounded by a monolayer of phospholipids. LDs are heterogeneous in their structure, chemical composition, and tissue distribution. LDs are coated by several proteins, including perilipins and other structural proteins, lipogenic enzymes, lipases and membrane-trafficking proteins. Five proteins of the perilipin (PLIN) family (PLIN1 (perilipin), PLIN2 (adipose differentiation-related protein), PLIN3 (tail-interacting protein of 47kDa), PLIN4 (S3-12), and PLIN5 (myocardial lipid droplet protein)), are associated with LD formation. More recently, the CIDE family of proteins, hypoxia-inducible protein 2 (HIG2), and patanin-like phospholipase domain-containing 3 (PNPLA3) have also gained attention in hepatic LD biology. Evidence suggests that LD proteins are involved in the pathophysiology of fatty liver diseases characterized by excessive lipid accumulation in hepatocytes. This review article will focus on how hepatic LDs and their associated proteins are involved in the pathogenesis of three chronic liver conditions: hepatitis C virus infection, non-alcoholic fatty liver disease, and alcoholic liver disease.

Nonalcoholic Fatty Liver Disease: Pathophysiology and Management.

Nonalcoholic fatty liver disease (NAFLD) is an important cause of morbidity and mortality worldwide and is rapidly becoming the leading cause of end-stage liver disease and liver transplant. With a prevalence of 30% in the United States, it has reached epidemic proportions. The clinical syndrome of NAFLD spans from bland steatosis to steatohepatitis, which can progress to fibrosis and cirrhosis. The pathogenesis includes the roles of hormones, nutritional and intestinal dysbiosis, insulin resistance, lipotoxicity, hepatic inflammation, and genes. Noninvasive testing and liver biopsy indications are reviewed. Approved and investigational therapies for NAFLD and nonalcoholic steatohepatitis are outlined in this article.

Temporal effects of ethanol consumption on energy homeostasis, hepatic steatosis, and insulin sensitivity in mice.

BACKGROUND Alcoholic liver disease (ALD) progresses from steatosis to inflammation, fibrosis, and cirrhosis. Although ALD has been associated with insulin resistance, it is unclear whether insulin resistance coincides with the development of steatosis. METHODS We studied the temporal relationship of steatosis and glucose homeostasis in mice fed a Lieber-DeCarli liquid control or ethanol (EtOH) diet for 2, 4, or 8 weeks. We studied the effects of alcohol consumption on energy balance, body composition, and hepatic lipids. Glucose tolerance test was performed, and insulin sensitivity was evaluated with hyperinsulinemic-euglycemic clamp. RESULTS EtOH-fed mice developed hepatic steatosis over time as compared with control-fed mice despite similar energy intake and expenditure, and gain in body weight and fat. EtOH-fed mice developed glucose intolerance as early as 2 weeks, while insulin resistance developed at 4 weeks. A hyperinsulinemic clamp study at 8 weeks revealed both hepatic and peripheral insulin resistance in EtOH-fed mice. Insulin resistance was associated with hepatic steatosis, increased ceramide levels, and Perilipin 2 expression. CONCLUSIONS Chronic EtOH consumption leads to the development of hepatic steatosis, impaired glucose tolerance, and insulin resistance. These changes are independent of energy intake or expenditure, weight, whole body fat content, and inflammation. A better understanding of the processes linking EtOH-induced steatosis and abnormal glucose homeostasis may lead to novel therapies targeting the progression of ALD.

FXR Agonists as Therapeutic Agents for Non-alcoholic Fatty Liver Disease

Non-alcoholic fatty liver disease (NAFLD) is the hepatic manifestation of the metabolic syndrome and a risk factor for both cardiovascular and hepatic related morbidity and mortality. The increasing prevalence of this disease requires novel therapeutic approaches to prevent disease progression. Farnesoid X receptors are bile acid receptors with roles in lipid, glucose, and energy homeostasis. Synthetic farnesoid X receptor (FXR) agonists have been developed to specifically target these receptors for therapeutic use in NAFLD patients. Here, we present a review of bile acid physiology and how agonism of FXR receptors has been examined in pre-clinical and clinical NAFLD. Early evidence suggests a potential role for synthetic FXR agonists in the management of NAFLD; however, additional studies are needed to clarify their effects on lipid and glucose parameters in humans.

Insulin resistance in clinical and experimental alcoholic liver disease

Alcoholic liver disease (ALD) is the number one cause of liver failure worldwide; its management costs billions of healthcare dollars annually. Since the advent of the obesity epidemic, insulin resistance (IR) and diabetes have become common clinical findings in patients with ALD; and the development of IR predicts the progression from simple steatosis to cirrhosis in ALD patients. Both clinical and experimental data implicate the impairment of several mediators of insulin signaling in ALD, and experimental data suggest that insulin‐sensitizing therapies improve liver histology. This review explores the contribution of impaired insulin signaling in ALD and summarizes the current understanding of the synergistic relationship between alcohol and nutrient excess in promoting hepatic inflammation and disease.

Reducing colorectal cancer risk among African Americans.

Colorectal cancer (CRC) burden is not equal among populations in the United States. African Americans have the highest CRC incidence and mortality of all US populations, and rates are not decreasing to the levels of non-Hispanic Whites.1 In addition to increased cancer risk, adenoma risk is also higher in African Americans, and both adenomas and cancers occur more frequently in the proximal colon and at younger ages in African Americans.2 Reasons for population differences are multifactorial and include differences in tumor biology and behavior, genetic risk, access to health care, and screening rates.3,4 As demonstrated by the Delaware CRC screening program, strategies to maximize screening hold significant promise for correcting CRC disparities.5 Current US Multisociety Task Force guidelines recommend CRC screening for all populations at average risk beginning at age 50 years, and individuals at increased risk (such as those with family history, inherited genetic syndromes or inflammatory bowel disease) are recommended to begin screening earlier.6 Owing to increased and earlier neoplasia risk, some professional organizations recommend screening in African Americans starting at age 45.3 Others raise concerns about the impact of complicating existing standardized guidelines and the unclear benefit of earlier age screening in African Americans despite an increased proportion of CRC under the age of 50 years. They recommend that efforts should focus instead on improving screening efforts in African Americans starting at age 50. Given this controversy, it is timely to examine how our profession can take the lead in reducing CRC disparities among African Americans. Several strategies should be considered when prioritizing our efforts (Table 1). Table 1 Strategies to Decrease Disparities in Colorectal Cancer (CRC) Among African Americans (AAs) African Americans are less knowledgeable about CRC and screening guidelines compared with Caucasians,7 and are less likely to transmit a family history of cancer.4 Both lack of knowledge about screening benefits and fatalistic views about cancer are associated with reduced likelihood of screening among African Americans.8 Interventions designed to educate patients about CRC and screening guidelines can improve screening rates and attitudes,9,10 and those that contain culturally sensitive materials have been shown to boost screening among African Americans.9 These and other studies suggest that lack of knowledge about CRC screening benefits is a surmountable barrier, but challenges remain. For example, ≤40% of African Americans aged 65 years and older in some US areas are estimated to read below a 5th-grade level,9 limiting the use of some CRC screening materials. In addition, standardized patient education approaches may not work in all populations and age groups, potentially necessitating individualized interventions and inclusion of personnel to engage in community-based education and outreach. The impact of provider endorsement on screening rates cannot be under-estimated. Lack of provider recommendation is an important barrier to screening in African Americans.11 However, studies that evaluate the impact of provider education on CRC screening in African Americans are lacking. Continuing medical education seminars can increase CRC knowledge, but whether this translates to improved screening rates is not clear.12 Just as there are no standardized approaches for patient education, there are no standardized strategies to improve provider education. Moreover, providers cite insufficient time as a barrier to recommending CRC screening to patients,13 potentially causing additional delay in timely CRC screening for this higher risk population. Strategies focused on physician education about the increased CRC burden among African Americans may improve CRC screening, but more research is needed to demonstrate this. Patient navigation is a proven strategy for increasing CRC screening rates in African Americans and also improves no show rates and bowel preparation.3,14 A randomized trial in older African Americans of phone navigation and printed material versus printed material alone found a 53% increase in endoscopic screening in the navigation group with health literate subjects showing a stronger effect from navigation.14 Financial modeling based on a program in New York City found patient navigation to be cost effective,15 whereas a randomized trial noted greater costs for tailored navigation.16 Implementation of patient navigation from research studies into the “real world” can be complex and requires flexibility and cooperation among stakeholders.17 Thus, although patient navigation can increase screening among African Americans, logistics and cost are major barriers to widespread adoption. Efforts should focus on overcoming these barriers through education, research, and advocacy for patient navigation in CRC screening. A more controversial strategy is to lower the initial screening age recommendation for African Americans. Arguments supporting this strategy include increased rates of significant neoplasia, higher stage of CRC at younger ages, and proximal location of tumors among African Americans.3 Lieberman et al2 showed that the rate of high risk polyps (>9 mm) was increased 17%–38% in African American men ages 50-69 years and 25%– 50% in African American women ages 50-64 years compared with Caucasian men and women in these age groups. Although the risk of large polyps was not statistically different for African American men and women <50, the sample size was small, limiting power to detect true differences in this subgroup. A simulation study argued in favor of earlier screening in African Americans as well as shortening screening intervals by 1 year given significant life-years gained.18 Moreover, higher prevalence of proximal neoplasia in African Americans argues for colonoscopy as the preferred endoscopic screening modality. If we consider race as a heritable factor,4 changing screening recommendations is akin to how we already risk stratify based on family history, and has the potential to prevent CRC among younger African Americans who currently are not detected early enough. Population-specific screening also raises awareness of the increased CRC burden in African Americans and could increase screening overall. Arguments against changing initial screening age guidelines for African Americans include the lack of definitive evidence for increased neoplasia risk and screening benefit before age 50 years.2,19 To address this, a comparative effectiveness trial enrolling those starting screening before age 50 and at age 50 could be performed. Another argument is that the current screening guidelines are already complex and that population-based screening recommendations would further confuse patients and physicians. Further modifications to guidelines could potentially negatively impact screening knowledge and adherence; however, there are no studies to support this notion. Although changes in breast and prostate cancer screening led to some public confusion,20,21 this was largely based on less intensified and reduced screening, not more. Given that African Americans have been shown to prefer non–endoscopy-based screening examinations,22 perhaps focusing less on structural examinations and simplifying the message to encourage screening of any kind that is acceptable to the patient is the most appropriate initial comprehensive strategy. Increasing screening rates by 5%–10% has been argued to be as or more effective than reducing the screening age.19 Although screening rates have been improving among African Americans and the gap with Caucasians is closing, there is certainly more work to be done. Finally, insurance coverage and access to endoscopy resources for earlier screening is a real concern and likely will only be changed if there is consensus among all bodies authoring screening guidelines. CRC burden among African Americans is unacceptably high, representing a major health disparity, and there are surmountable barriers to reduce this disparity. Patient and physician education as well as implementation of patient navigators can increase screening rates and should receive considerable attention. Whether implementing new population-specific screening guidelines will augment these efforts remains unclear; however, it is now time for our profession to establish consensus on this issue by establishing an expert panel to review the data and formulate updated evidence-based guidelines to reduce CRC disparities in African Americans, including a formal recommendation regarding the optimal screening age and modality. Such efforts will help us to make progress on health equity for all populations at risk for CRC and form a basis for an approach to care in this high risk population.

Perilipin Staining Distinguishes Between Steatosis and Nonalcoholic Steatohepatitis in Adults and Children

*Perelman School of Medicine, Department of Medicine, Gastroenterology and Hepatology Division, Division of Endocrinology, Diabetes and Metabolism, University of Pennsylvania, Philadelphia, Pennsylvania; School of Medical Sciences, University of Hyderabad, Hyderabad, India; kDivision of Gastroenterology and Hepatology, Indiana University School of Medicine, Indianapolis, Indiana; Division of Endocrinology, Diabetes and Metabolism, Johns Hopkins University School of Medicine, Baltimore, Maryland

Ethanol and C2 ceramide activate fatty acid oxidation in human hepatoma cells

Obesogenic lipids and the sphingolipid ceramide have been implicated as potential cofactors in alcoholic liver disease (ALD) patients. However, the mechanisms by which these lipids modulate lipid trafficking in ethanol-treated human liver cells to promote steatosis, an early stage of ALD, are poorly understood. We measured fatty acid (FA) uptake, triglyceride export, FA synthesis and FA oxidation in human hepatoma (VL-17A) cells in response to ethanol and the exogenous lipids oleate, palmitate and C2 ceramide. We found that in combination with ethanol, both oleate and palmitate promote lipid droplet accumulation while C2 ceramide inhibits lipid droplet accumulation by enhancing FA oxidation. Further, using both a pharmacologic and siRNA approach to reduce peroxisome proliferator-activated receptors α (PPARα) gene expression, we demonstrate that C2 ceramide abrogates ethanol-mediated suppression of FA oxidation through an indirect PPARα mechanism. Together, these data suggest that lipids interact differentially with ethanol to modulate hepatocellular lipid droplet accumulation and may provide novel targets for preventing the earliest stage of alcoholic liver disease, alcoholic steatosis.