Triglyceride synthesis in normal and hyperapoB fibroblasts

The goal of this thesis was to examine the regulation of intracellular triglyceride synthesis from normals and HyperapoB patients using cultured skin fibroblasts as an experimental model. In a medium supplemented with lipoprotein-deficient serum, triglyceride synthesis and cholesterol esterification were reduced in the HyperapoB fibroblasts by 40% and 44% respectively as compared to the normal fibroblasts. These differences were due to differences in de novo synthesis, and not due to re-esterification or hydrolysis. However, unexpectedly, there was no difference in triglyceride synthesis between the cells from normals and HyperapoB when a serum-free supplemented medium was used. The stimulatory factor in lipoprotein-deficient serum was then isolated from human plasma and characterized. The factor responsible is nondialyzable and trypsin sensitive, and its effects are both concentration and time dependent. The protein responsible has been purified to homogeneity and is a small (MW 14,000), basic (pI 9.0) protein. It has been named Acylation Stimulating Protein (ASP) because it markedly stimulates triglyceride synthesis and cholesterol esterification by 80% and 42% in normal human skin fibroblasts. Further ASP is specifically bound with a KD of 9.9 $\times$ 10\sp{-7} and is internalized and degraded by normal human skin fibroblasts. In contrast, ASP has much less effect on triglyceride synthesis in HyperapoB fibroblasts with an average of only 25% triglyceride stimulation. This is consistent with its reduced maximal binding (51% of normal) and subsequent internalization into these cells. Although both cell groups demonstrate the same binding affinity for ASP (Scatchard analysis slope = $-.0725$ ug\sp{-1} normal and $-.080$R ug\sp{-1} HyperapoB) the maximal number of apparent binding sites is reduced in HyperapoB (1.217 mg\sp{-1} normal vs 0.621 mg\sp{-1} HyperapoB). The ASP effect on cells from patients with Familial Hypercholesterolemia, however, is normal with regard to ASP stimulation of triglyceride synthesis (average 55% stimulation) and ASP binding to cells. This research therefore has led to the description of a previously unrecognized protein in human plasma. This protein, ASP, appears to be the most potent stimulant of intracellular triglyceride synthesis yet describe

Adipocyte size as a determinant of metabolic disease and adipose tissue dysfunction

Obesity is a heterogeneous disease and is associated with comorbidities such as type 2 diabetes mellitus, cardiovascular disease and cancer. Several studies have examined the role of dysfunctional adipose tissue in the pathogenesis of obesity, highlighting the contrasting properties and impact of distinct fat compartments, sometimes with contradictory results. Dysfunctional adipose tissue involves enlargement, or hypertrophy, of pre-existing fat cells, which is thought to confer increases in cardiometabolic risk, independent of the level of obesity per se . In this article, we critically analyze available literature that examined the ability of adipocyte cell size to predict metabolic disease and adipose tissue dysfunction in humans. Many studies demonstrate that increased fat cell size is a significant predictor of altered blood lipid profiles and glucose–insulin homeostasis independent of adiposity indices. The contri- bution of visceral adiposity to these associations appears to be of particular importance. However, available studies are not unanimous and many fat depot-specific aspects of the relationship between increased fat cell size and cardiometabolic risk or parameters of adipose tissue dysfunction are still unresolved. Methodological factors such as the approach used to express the data may represent significant confounders in these studies. Additional studies should consider the fact that the relationship between fat cell size and common adiposity indices is non-linear, particularly when reaching the obese range. In conclusion, our analysis demonstrates that fat cell size is a significant predictor of the cardiometabolic alterations related to obesity. We propose that adipocyte hypertrophy, especially in the visceral fat compartment, may represent a strong marker of limited hyperplasic capacity in subcutaneous adipose tissues, which in turn is associated with the presence of numerous cardiometabolic alterations

Acylation stimulating protein is higher in Inuit from Nunavik compared to a southern Quebec population

Objectives. The Inuit of Nunavik in northern Quebec have a lower risk for ischemic heart disease (IHD) compared to Caucasian populations. Acylation stimulating protein (ASP), which is involved in the storage of dietary fat, may play a role. The objective of the study was to determine plasma concentration of ASP in an Inuit and a southern Quebec Caucasian population. Study design. This is a cross-sectional study evaluating the relationship between ASP and dietary factors, such as retinol, whose intake is higher in the Inuit. As well, concentrations of ASP were evaluated in relationship to components of the metabolic syndrome. Methods. Medical history was collected via a questionnaire and anthropometric measurements and blood samples were collected. Results. ASP was significantly higher in both the Inuit men and women compared to Caucasian men (66.1±4.1 nM vs 27.5±2.5 nM, p<0.0001) and women (71.8±3.8 nM vs 29.4±1.3 nM, p<0.0001). In addition, ASP significantly correlated with total retinol (r=0.17, p=0.02) and free retinol (r=0.15, p=0.04) in Inuit men but not with other distinctive dietary markers such as omega3 fatty acids. Conclusions. Inuit men and women have higher ASP which was unrelated to the number of risk factors for IHD that were present

Differential methylation of inflammatory and insulinotropic genes after metabolic surgery in women

Context: Biliopancreatic diversion with duodenal switch (BPD-DS), a metabolic bariatric operation, induces durable loss of excess weight and reduced cardiometabolic risk. Altered epigenetic marks are mechanistically associated with environment-driven phenotypic variations. Objective: The current study aimed to compare gene methylation levels before and after BPD-DS to identify epigenetic marks potentially linked to metabolic improvements induced by BPD-DS. Design and patients: Metabolic risk factors and gene methylation levels of 20 women studied mean 12 years (range 4-22) after BPD-DS were compared to those of 20 severely obese surgical candidates as controls, matched for pre-surgical age, body mass index and dyslipidemia and hypertension prevalences. Whole-genome blood DNA methylation analysis enabled between-group differential methylation analyses. We calculated correlations between methylation levels of the most differentially methylated CpG sites and plasma glucose and insulin levels and HOMA-IR. Results: Differential methylation analysis identified 15,343 genes demonstrating at least one differentially methylated CpG site (p<1.43x10-7). Diabetic and inflammation/immune functions were among the most overrepresented from the 200 genes exhibiting the largest group differences in methylation levels. CpG sites methylation levels of genes related to insulin action correlated significantly with fasting insulin levels and homeostatic model of insulin resistance (p≤0.002 for all). Conclusion: These findings suggest that differential methylation levels in obese controls versus treated women may partially explain the durable metabolic improvements after BPD-DS

Circulating steroid levels as correlates of adipose tissue phenotype in premenopausal women

Background: Obesity-related alterations in the circulating steroid hormone profile remain equivocal in women. Our objective was to identify circulating steroid levels that relate to increased adiposity and altered adipose phenotype in premenopausal women. Materials and methods: In a sample of 42 premenopausal women (age 46±3 years; BMI 27.1±4.2 kg/m2 ), 19 plasma steroids were quantified by ESI-LC-MS/MS. Body composition and fat distribution were assessed by dual-energy X-ray absorptiometry and computed tomography, respectively. Markers of adipose tissue function including adipocyte size distributions, radiological attenuation, and macrophage infiltration were also analyzed in surgically obtained visceral and subcutaneous fat samples. Results: Many negative correlations were observed between adiposity measurements such as BMI, body fat percentage or total abdominal adipose tissue area and plasma levels of androstenedione (r=-0.33 to -0.39, p≤0.04), androsterone (r=-0.30 to -0.38, p≤0.05) and plasma levels of steroid precursor pregnenolone (r=-0.36 to -0.46, p≤0.02). Visceral adipocyte hypertrophy was observed in patients with low pregnenolone concentrations (p<0.05). Visceral adipose tissue radiologic attenuation, a potential marker of adipocyte size, was also positively correlated with pregnenolone levels (r=0.33, p<0.05). Low levels of pregnenolone were related to increased number of macrophages infiltrating visceral and subcutaneous adipose tissue (p<0.05). Conclusion: Plasma levels of androgens and their precursors are lower in women with increased adiposity and visceral adipocyte hypertrophy. Low circulating pregnenolone concentration may represent a marker of adipose tissue dysfunction

17β-HSD type 2 activity and localization in human adipose tissue

Testosterone can be converted into androstenedione (4-dione) by 17β-hydroxysteroid dehydrogenase (HSD) activity likely performed by 17β-HSD type 2. Our objective was to evaluate the rate of testosterone conversion to 4-dione as well as expression and localization of 17β-HSD type 2 in omental (OM) vs. subcutaneous (SC) adipose tissues of men. Formation of 4-dione from testosterone was significantly higher in homogenates (p ≤ 0.001) and explants (p ≤ 0.01) of OM than SC tissue. Microscopy analyses and biochemical assays in cell fractions localized the enzyme in the vasculature/endothelial cells of adipose tissues. Conversion of testosterone to 4-dione was weakly detected in most OM and/or SC preadipocyte cultures. Positive correlations were found between 17β-HSD type 2 activity in whole tissue and BMI or SC adipocyte diameter. We conclude that conversion of testosterone to 4-dione detected in abdominal adipose tissue is caused by 17β-HSD type 2 which is localized in the vasculature of the adipose compartment

Association of adipocyte genes with ASP expression: a microarray analysis of subcutaneous and omental adipose tissue in morbidly obese subjects

<p>Abstract</p> <p>Background</p> <p>Prevalence of obesity is increasing to pandemic proportions. However, obese subjects differ in insulin resistance, adipokine production and co-morbidities. Based on fasting plasma analysis, obese subjects were grouped as Low Acylation Stimulating protein (ASP) and Triglyceride (TG) (LAT) vs High ASP and TG (HAT). Subcutaneous (SC) and omental (OM) adipose tissues (n = 21) were analysed by microarray, and biologic pathways in lipid metabolism and inflammation were specifically examined.</p> <p>Methods</p> <p>LAT and HAT groups were matched in age, obesity, insulin, and glucose, and had similar expression of insulin-related genes (InsR, IRS-1). ASP related genes tended to be increased in the HAT group and were correlated (factor B, adipsin, complement C3, p < 0.01 each). Differences between LAT and HAT group were almost exclusively in SC tissue, with little difference in OM tissue. Increased C5L2 (p < 0.01), an ASP receptor, in HAT suggests a compensatory ASP pathway, associated with increased TG storage.</p> <p>Results</p> <p>HAT adipose tissue demonstrated increased lipid related genes for storage (CD36, DGAT1, DGAT2, SCD1, FASN, and LPL), lipolysis (HSL, CES1, perilipin), fatty acid binding proteins (FABP1, FABP3) and adipocyte differentiation markers (CEBPα, CEBPβ, PPARγ). By contrast, oxidation related genes were decreased (AMPK, UCP1, CPT1, FABP7). HAT subjects had increased anti-inflammatory genes TGFB1, TIMP1, TIMP3, and TIMP4 while proinflammatory PIG7 and MMP2 were also significantly increased; all genes, p < 0.025.</p> <p>Conclusion</p> <p>Taken together, the profile of C5L2 receptor, ASP gene expression and metabolic factors in adipose tissue from morbidly obese HAT subjects suggests a compensatory response associated with the increased plasma ASP and TG.</p

Racial difference in Acylation Stimulating Protein (ASP) correlates to triglyceride in non-obese and obese African American and Caucasian women

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Hepatitis B Vaccine Antibody Response and the Risk of Clinical AIDS or Death

Whether seroresponse to a vaccine such as hepatitis B virus (HBV) vaccine can provide a measure of the functional immune status of HIV-infected persons is unknown.This study evaluated the relationship between HBV vaccine seroresponses and progression to clinical AIDS or death. (HR 3.40; 95% CI, 1.39–8.32).

Oval Cell Response Is Attenuated by Depletion of Liver Resident Macrophages in the 2-AAF/Partial Hepatectomy Rat

BACKGROUND/AIMS: Macrophages are known to play an important role in hepatocyte mediated liver regeneration by secreting inflammatory mediators. However, there is little information available on the role of resident macrophages in oval cell mediated liver regeneration. In the present study we aimed to investigate the role of macrophages in oval cell expansion induced by 2-acetylaminofluorene/partial hepatectomy (2-AAF/PH) in rats. METHODOLOGY/PRINCIPAL FINDINGS: We depleted macrophages in the liver of 2-AAF/PH treated rats by injecting liposome encapsulated clodronate 48 hours before PH. Regeneration of remnant liver mass, as well as proliferation and differentiation of oval cells were measured. We found that macrophage-depleted rats suffered higher mortality and liver transaminase levels. We also showed that depletion of macrophages yielded a significant decrease of EPCAM and PCK positive oval cells in immunohistochemical stained liver sections 9 days after PH. Meanwhile, oval cell differentiation was also attenuated as a result of macrophage depletion, as large foci of small basophilic hepatocytes were observed by day 9 following hepatectomy in control rats whereas they were almost absent in macrophage depleted rats. Accordingly, real-time polymerase chain reaction analysis showed lower expression of albumin mRNA in macrophage depleted livers. Then we assessed whether macrophage depletion may affect hepatic production of stimulating cytokines for liver regeneration. We showed that macrophage-depletion significantly inhibited hepatic expression of tumor necrosis factor-α and interleukin-6, along with a lack of signal transducer and activator of transcription 3 phosphorylation during the early period following hepatectomy. CONCLUSIONS: These data indicate that macrophages play an important role in oval cell mediated liver regeneration in the 2-AAF/PH model