Wan Huang

Depletion of Liver Kupffer Cells Prevents the Development of Diet-Induced Hepatic Steatosis and Insulin Resistance

OBJECTIVE Increased activity of the innate immune system has been implicated in the pathogenesis of the dyslipidemia and insulin resistance associated with obesity and type 2 diabetes. In this study, we addressed the potential role of Kupffer cells (liver-specific macrophages, KCs) in these metabolic abnormalities. RESEARCH DESIGN AND METHODS Rats were depleted of KCs by administration of gadolinium chloride, after which all animals were exposed to a 2-week high-fat or high-sucrose diet. Subsequently, the effects of these interventions on the development of hepatic insulin resistance and steatosis were assessed. In further studies, the effects of M1-polarized KCs on hepatocyte lipid metabolism and insulin sensitivity were addressed. RESULTS As expected, a high-fat or high-sucrose diet induced steatosis and hepatic insulin resistance. However, these metabolic abnormalities were prevented when liver was depleted of KCs. In vitro, KCs recapitulated the in vivo effects of diet by increasing hepatocyte triglyceride accumulation and fatty acid esterification, and decreasing fatty acid oxidation and insulin responsiveness. To address the mechanisms(s) of KC action, we inhibited a panel of cytokines using neutralizing antibodies. Only neutralizing antibodies against tumor necrosis factor-α (TNFα) attenuated KC-induced alterations in hepatocyte fatty acid oxidation, triglyceride accumulation, and insulin responsiveness. Importantly, KC TNFα levels were increased by diet in vivo and in isolated M1-polarized KCs in vitro. CONCLUSIONS These data demonstrate a role for liver macrophages in diet-induced alterations in hepatic lipid metabolism and insulin sensitivity, and suggest a role for these cells in the etiology of the metabolic abnormalities of obesity/type 2 diabetes.

CCAAT/Enhancer-binding Protein β Deletion Reduces Adiposity, Hepatic Steatosis, and Diabetes in Leprdb/db Mice

CCAAT/enhancer-binding protein β (C/EBPβ) plays a key role in initiation of adipogenesis in adipose tissue and gluconeogenesis in liver; however, the role of C/EBPβ in hepatic lipogenesis remains undefined. Here we show that C/EBPβ inactivation in Leprdb/db mice attenuates obesity, fatty liver, and diabetes. In addition to impaired adipogenesis, livers from C/EBPβ-/- x Leprdb/db mice had dramatically decreased triglyceride content and reduced lipogenic enzyme activity. C/EBPβ deletion in Leprdb/db mice down-regulated peroxisome proliferator-activated receptor γ2 (PPARγ2) and stearoyl-CoA desaturase-1 and up-regulated PPARα independent of SREBP1c. Conversely, C/EBPβ overexpression in wild-type mice increased PPARγ2 and stearoyl-CoA desaturase-1 mRNA and hepatic triglyceride content. In FAO cells, overexpression of the liver inhibiting form of C/EBPβ or C/EBPβ RNA interference attenuated palmitate-induced triglyceride accumulation and reduced PPARγ2 and triglyceride levels in the liver in vivo. Leptin and the anti-diabetic drug metformin acutely down-regulated C/EBPβ expression in hepatocytes, whereas fatty acids up-regulate C/EBPβ expression. These data provide novel evidence linking C/EBPβ expression to lipogenesis and energy balance with important implications for the treatment of obesity and fatty liver disease.

Impaired Activation of Phosphatidylinositol 3-Kinase by Leptin Is a Novel Mechanism of Hepatic Leptin Resistance in Diet-induced Obesity

Obesity is associated with the development of leptin resistance. However, the effects of leptin resistance on leptin-regulated metabolic processes and the biochemical defects that cause leptin resistance are poorly understood. We have addressed in rats the effect of dietinduced obesity (DIO), a situation of elevated tissue lipid levels, on the well described lipid-lowering effect of leptin in liver, an action that is proposed to be important for the prevention of tissue lipotoxicity and insulin resistance. In addition, we have addressed the role of phosphatidylinositol 3-kinase (PI 3-kinase) in mediating the acute effects of leptin on hepatic lipid levels in lean and DIO animals. A 90-min leptin (∼10 ng/ml) perfusion of isolated livers from lean animals decreased triglyceride levels by 42 ± 5% (p = 0.006). However, leptin concentrations ranging from ∼10 to ∼90 ng/ml had no effect on triglyceride levels in livers from DIO animals. The acute lipid-lowering effect of leptin on livers from lean animals was mediated by a PI 3-kinase-dependent mechanism, because wortmannin and LY294002, the PI 3-kinase inhibitors, blocked the effects of leptin on hepatic triglyceride levels and leptin increased liver PI 3-kinase activity by 183 ± 6% (p = 0.003) and insulin receptor substrate 1 tyrosine phosphorylation by 185 ± 30% (p = 0.02) in the absence of PI 3-kinase inhibitors. Contrary to the effects of leptin in lean livers, leptin did not activate PI 3-kinase in livers from DIO rats. These data present evidence for a role for 1) leptin resistance in contributing to the excessive accumulation of tissue lipid in obesity, 2) PI 3-kinase in mediating the acute lipid-lowering effects of leptin in liver, and 3) defective leptin activation of PI 3-kinase as a novel mechanism of leptin resistance.

Associations Between Serum Biomarkers and Pain and Pain‐Related Function in Older Adults with Low Back Pain: A Pilot Study

To examine the relationship between serum biomarkers and self‐reported pain intensity and pain‐related function, in addition to the contribution of magnetic resonance imaging (MRI) findings of lumbar spine degenerative changes, in older adults with chronic low back pain.

Identification of Distinct Monocyte Phenotypes and Correlation With Circulating Cytokine Profiles in Acute Response to Spinal Cord Injury: A Pilot Study

BACKGROUND Macrophage infiltration to the injury site during the acute response to traumatic spinal cord injury (SCI) is not uniform. Macrophage phenotype has been characterized as either proinflammatory (M1) or anti-inflammatory (M2). Results of animal studies suggest that M1 or M2 dominance at the site of injury relates to spontaneous recovery after SCI. OBJECTIVE To investigate whether the phenotype of circulating macrophage precursors-monocytes (MO) is altered in the acute phase of SCI and corresponds to circulating inflammatory cytokines. STUDY DESIGN A prospective observational cohort study. SETTING A single academic medical center in Pennsylvania. PATIENTS A cohort of 27 subjects with complete or incomplete traumatic SCI enrolled within 7 days after SCI injury. METHODS The MO phenotype was defined within the first week after SCI by using flow cytometry and was compared with that of historic uninjured controls. Concentrations of 25 cytokines and/or chemokines were assessed by using Luminex in serial blood samples up to 2 weeks after SCI. An analysis of variance was used to determine the correlations between the phenotypes and the cytokine profiles. RESULTS Patient subsets were identified with either M1- or M2-dominant circulating MOs distinct from the uninjured controls. The M1 dominant was associated with higher circulating levels of proinflammatory mediators interleukin (IL)12p70 and interferon gamma-induced protein 10 kDa (IP-10/CXCL10), and lower levels of anti-inflammatory cytokines IL-10, IL-15, and IL-7, whereas the M2 dominant exhibited the opposite cytokine profiles with significantly higher IL-10 and IL-7. CONCLUSION In the acute phase after SCI, at comparable injury severity, subgroups of patients exhibit distinct M1 or M2 MOs dominance and the phenotype is correlated with M1- or M2-specific cytokine and/or chemokine profiles. Although further studies are needed to determine how these observed phenotypic differences relate to functional recovery, our findings (1) provide the first evidence, to our knowledge, that indicates the possible individual differences in the immune responses to the comparable traumatic SCI, with potential implications for management of acute SCI and rehabilitation; and (2) may represent easily accessible biomarkers with prognostic utility.

Leptin Augments the Acute Suppressive Effects of Insulin on Hepatic Very Low-Density Lipoprotein Production in Rats

It is well established that leptin increases the sensitivity of carbohydrate metabolism to the effects of insulin. Leptin and insulin also have potent effects on lipid metabolism. However, the effects of leptin on the regulation of liver lipid metabolism by insulin have not been investigated. The current study addressed the effects of leptin on insulin-regulated hepatic very low-density lipoprotein (VLDL) metabolism in vivo in rats. A 90-min hyperinsulinemic/euglycemic clamp (4 mU/kg x min(-1)) reduced plasma VLDL triglyceride (TG) by about 50% (P < 0.001 vs. saline control). Importantly, a leptin infusion (0.2 microg/kg x min(-1)) in combination with insulin reduced plasma VLDL-TG by about 80% (P < 0.001 vs. insulin alone). These effects did not require altered skeletal muscle lipoprotein lipase activity but did include differential effects of insulin and leptin on liver apolipoprotein (apo) B and TG metabolism. Thus, insulin decreased liver and plasma apoB100/B48 levels (approximately 50%, P < 0.01), increased liver TGs (approximately 20%, P < 0.05), and had no effect on fatty acid oxidation. Conversely, leptin decreased liver TGs (approximately 50%, P < 0.01) and increased fatty acid oxidation (approximately 50%, P < 0.01) but had no effects on liver or plasma apoB levels. Importantly, the TG-depleting and prooxidative effects of leptin were maintained in the presence of insulin. We conclude that leptin additively increases the suppressive effects of insulin on hepatic and systemic VLDL metabolism by stimulating depletion of liver TGs and increasing oxidative metabolism. The net effect of the combined actions of insulin and leptin is to decrease the production and TG content of VLDL particles.

Skeletal muscle as a regulator of the longevity protein, Klotho

Klotho is a powerful longevity protein that has been linked to the prevention of muscle atrophy, osteopenia, and cardiovascular disease. Similar anti-aging effects have also been ascribed to exercise and physical activity. While an association between muscle function and Klotho expression has been previously suggested from longitudinal cohort studies, a direct relationship between circulating Klotho and skeletal muscle has not been investigated. In this paper, we present a review of the literature and preliminary evidence that, together, suggests Klotho expression may be modulated by skeletal muscle activity. Our pilot clinical findings performed in young and aged individuals suggest that circulating Klotho levels are upregulated in response to an acute exercise bout, but that the response may be dependent on fitness level. A similar upregulation of circulating Klotho is also observed in response to an acute exercise in young and old mice, suggesting that this may be a good model for mechanistically probing the role of physical activity on Klotho expression. Finally, we highlight overlapping signaling pathways that are modulated by both Klotho and skeletal muscle and propose potential mechanisms for cross-talk between the two. It is hoped that this review will stimulate further consideration of the relationship between skeletal muscle activity and Klotho expression, potentially leading to important insights into the well-documented systemic anti-aging effects of exercise.

Biomarker Development for Musculoskeletal Diseases

More than one in 4 Americans has a musculoskeletal (MSK) disorder that requires medical diagnosis and treatment. Imaging tools are able to demonstrate structural changes but cannot reflect the disease activity or symptom severity of MSK conditions. This is of paramount concern in the aging population, in which imaging findings have poor correlation with symptoms, and multiple pain generators frequently coexist. Because levels of inflammatory and matrix breakdown products address disease activity, evaluation of biomarkers has the potential to provide assessment of active pain generators above and beyond the changes observable on imaging studies. This fact has stimulated research interest in the search for novel biomarkers of disease activity and response to treatment in body fluids. The goal is to develop panels of multi‐biomarkers, which could be used independently or in conjunction with the imaging tools, for the diagnosis, prognosis, and treatment validation in MSK diseases. The current review of MSK biomarkers is organized into 3 mechanistic categories: the metabolites of extracellular matrix of MSK tissues; inflammatory cytokines and chemokines; and pain‐related neuropeptides and/or chemicals. Although some representative biomarkers could be used alone, the fact that MSK diseases are multi‐tissue disorders that involve the muscles, bones, cartilage, and nerves suggests that panels of biomarkers may have greater potential than any single biomarker used in isolation. As advances in biotechnology make this a reality, multi‐biomarker panels that include all 3 categories of biomarkers, used either alone or in combination with imaging tools, has the potential to revolutionize the clinical approach to MSK diseases.

Kupffer cells facilitate the acute effects of leptin on hepatic lipid metabolism

Leptin has potent effects on lipid metabolism in a number of peripheral tissues. In liver, an acute leptin infusion (~120 min) stimulates hepatic fatty acid oxidation (~30%) and reduces triglycerides (TG, ~40%), effects that are dependent on phosphoinositol-3-kinase (PI3K) activity. In the current study we addressed the hypothesis that leptin actions on liver-resident immune cells are required for these metabolic effects. Myeloid cell-specific deletion of the leptin receptor (ObR) in mice or depletion of liver Kupffer cells (KC) in rats in vivo prevented the acute effects of leptin on liver lipid metabolism, while the metabolic effects of leptin were maintained in mice lacking ObR in hepatocytes. Notably, liver TG were elevated in both lean and obese myeloid cell ObR, but the degree of obesity and insulin resistance induced by a high-fat diet was similar to control mice. In isolated primary hepatocytes (HEP), leptin had no effects on HEP lipid metabolism and only weakly stimulated PI3K. However, the coculture of KC with HEP restored leptin action on HEP fatty acid metabolism and stimulation of HEP PI3K. Notably, leptin stimulated the release from KC of a number of cytokines. However, the exposure of HEP to these cytokines individually [granulocyte macrophage colony-stimulating factor, IL-1α, IL-1β, IL-6, IL-10, and IL-18] or in combination had no effects on HEP lipid metabolism. Together, these data demonstrate a role for liver mononuclear cells in the regulation of liver lipid metabolism by leptin.

Poster 113: Association of Clinical Characteristics and Response to Lumbar Epidural Steroid Injections in Subjects with Axial Low Back Pain

Disclosures: Stephen Schaaf: I Have No Relevant Financial Relationships To Disclose Objective: Despite the frequency of interventional procedures performed for axial low back pain, selection criteria remain unclear and suboptimal to predict those that will have an improved outcome. The goal of the study was to examine the association of baseline clinical characteristics and pain improvement following lumbar epidural steroid injections (LESI) in individuals with axial low back pain. Design: Prospective Cohort Study Setting: Academic Medical Center. Participants: Subjects (n1⁄448) were eligible if they had primarily axial low back pain without radiating symptoms. Interventions: Patients recruited had already consented for and then underwent a LESI as part of their routine clinical care. Main Outcome Measures: Gender, age, body mass index, race, education, employment status, smoking status, Oswestry Disability Index, Roland Morris Disability, McGill Pain Questionnaire (MPQ), generalized anxiety disorder, Patient Health Questionnaire (PHQ-9), 10m-walking speed, fear avoidance beliefs questionnaire, catastrophizing, cumulative illness rating scale, prior treatments and exercise, treatment expectation, and medications were collected for baseline clinical characteristics. Pain was scored on 0-10 numeric rating scale. Pain score was taken at pre-injection and two-week follow-up. Responders to injection were defined as those who had at least a 50% reduction in their pain score at follow up. Results: At follow up, 17 subjects reported 50% or greater reduction in their pain score. Responders had a significantly lower PHQ total score, MPQ sensitive score, MPQ affective score, MPQ total score, more frequent exercise, and were less unsure about expecting pain relief following the LESI at baseline. No other significant associations were observed. Conclusions: Certain baseline clinical characteristics represent a potential opportunity to improve the clinical ability to predict response to treatment for LESI. A larger sample size with randomized study design to evaluate the ability of baseline clinical characteristics to improve clinical decision making will be needed. Level of Evidence: Level II