Exploring the early molecular pathogenesis of osteoarthritis using differential network analysis of human synovial fluid.

The molecular mechanisms that drive the onset and development of osteoarthritis (OA) remain largely unknown. In this exploratory study, we used a proteomic platform (SOMAscan assay) to measure the relative abundance of over 6000 proteins in synovial fluid (SF) from knees of human donors with healthy or mildly degenerated tissues and knees with late stage OA from patients undergoing knee replacement surgery. Using linear mixed effects model, we estimated the differential abundance of 6251 proteins between the three groups. We found 583 proteins upregulated in the late-stage OA, including MMP1, MMP13 and IL6. Further, we selected 760 proteins (800 aptamers) based on absolute fold changes between healthy and mild degeneration group. To those, we applied Gaussian Graphical Models (GGMs) to analyze the conditional dependence of proteins and to identify key proteins and subnetworks involved in early OA pathogenesis. After regularization and stability selection, we identified 102 proteins involved in GGM networks. Notably, network complexity was lost in the protein graph for mild degeneration when compared to controls, suggesting a disruption in the regular protein interplay. Furthermore, among our main findings were several downregulated (in mild degeneration versus healthy) proteins with unique interactions in healthy group, one of which, SLCO5A1, have not previously been associated with OA. Our results suggest that this protein is important for healthy joint function. Further, our data suggests that SF proteomics, combined with GGMs, can reveal novel insights into the molecular pathogenesis and identification of biomarker candidates for early stage OA.

Digital Screening for Cognitive Impairment - A Proof of Concept Study.

BACKGROUND: Due to an ageing demographic and rapid increase of cognitive impairment and dementia, combined with potential disease-modifying drugs and other interventions in the pipeline, there is a need for the development of accurate, accessible and efficient cognitive screening instruments, focused on early-stage detection of neurodegenerative disorders. OBJECTIVE: In this proof of concept report, we examine the validity of a newly developed digital cognitive test, the Geras Solutions Cognitive Test (GCST) and compare its accuracy against the Montreal Cognitive Assessment (MoCA). METHODS: 106 patients, referred to the memory clinic, Karolinska University Hospital, due to memory complaints were included. All patients were assessed for presence of neurodegenerative disorder in accordance with standard investigative procedures. 66% were diagnosed with subjective cognitive impairment (SCI), 25% with mild cognitive impairment (MCI) and 9% fulfilled criteria for dementia. All patients were administered both MoCA and GSCT. Descriptive statistics and specificity, sensitivity and ROC curves were established for both test. RESULTS: Mean score differed significantly between all diagnostic subgroups for both GSCT and MoCA (p<0.05). GSCT total test time differed significantly between all diagnostic subgroups (p<0.05). Overall, MoCA showed a sensitivity of 0.88 and specificity of 0.54 at a cut-off of <=26 while GSCT displayed 0.91 and 0.55 in sensitivity and specificity respectively at a cut-off of <=45. CONCLUSION: This report suggests that GSCT is a viable cognitive screening instrument for both MCI and dementia.

Long-term changes in adipose tissue gene expression following bariatric surgery.

OBJECTIVE: Patients undergoing bariatric surgery present long-term metabolic improvements and reduced type 2 diabetes risk, despite long-term weight regain. We hypothesized that part of these protective effects could be linked to altered gene expression in white adipose tissue (WAT). METHODS: Transcriptomic profiling by gene microarray was performed in abdominal subcutaneous WAT from women before (n = 50) and two (n = 49) and five (n = 38) years after Roux-en-Y gastric bypass (RYGB) surgery as well as in 28 age-matched nonoperated women. RESULTS: In the obese women, the average body weight decrease was 38 kg 2 years postsurgery followed by an 8 kg weight regain between 2 and 5 years. Most of the long-term changes in WAT gene expression occurred during the first 2 years. However, a subset of genes encoding proteins involved in inflammation displayed a continued decrease between baseline, 2 and 5 years, respectively; that is an expression pattern independent of body weight regain. Expression of 71 of these genes correlated with measurements of adipocyte morphology or serum adipokine levels. CONCLUSION: The continuous improvement in WAT inflammatory gene expression, despite body weight relapse, may contribute to the sustained effects on adipose morphology after bariatric surgery.

Proteomic comparison of osteoarthritic and reference human menisci using data-independent acquisition mass spectrometry.

OBJECTIVE: Recent research in knee osteoarthritis (OA) highlights the role of the meniscus in OA pathology. Our aim was to compare the proteomes of medial and lateral menisci from end-stage medial compartment knee OA patients, with reference menisci from knee-healthy deceased donors, using mass spectrometry. DESIGN: Tissue plugs of Ø3 mm were obtained from the posterior horns of the lateral and medial menisci from one knee of 10 knee-healthy deceased donors and 10 patients undergoing knee replacement. Proteins were extracted and prepared for mass spectrometric analysis. Statistical analysis was conducted on abundance data that was log2-transformed, using a linear mixed effects model and evaluated using pathway analysis. RESULTS: We identified a total of 835 proteins in all samples, of which 331 were included in the statistical analysis. The largest differences could be seen between the medial menisci from OA patients and references, with most proteins showing higher intensities in the medial menisci from OA patients. Several matrix proteins, e.g., matrix metalloproteinase 3 (MMP3) (4.3 times higher values [95%CI 1.8, 10.6]), TIMP1 (3.5 [1.4, 8.5]), asporin (4.1 [1.7, 10.0]) and versican (4.4 [1.8, 10.9]), all showed higher abundance in medial menisci from OA patients compared to medial reference menisci. OA medial menisci also showed increased activation of several pathways involved in inflammation. CONCLUSION: An increase in protein abundance for proteins such as MMP and TIMP1 in the medial menisci from OA patients suggests simultaneous activation of both catabolic and anabolic processes that warrants further attention.

Adipose lipid turnover and long-term changes in body weight.

The worldwide obesity epidemic1 makes it important to understand how lipid turnover (the capacity to store and remove lipids) regulates adipose tissue mass. Cross-sectional studies have shown that excess body fat is associated with decreased adipose lipid removal rates2,3. Whether lipid turnover is constant over the life span or changes during long-term weight increase or loss is unknown. We determined the turnover of fat cell lipids in adults followed for up to 16 years, by measuring the incorporation of nuclear bomb test-derived 14C in adipose tissue triglycerides. Lipid removal rate decreases during aging, with a failure to reciprocally adjust the rate of lipid uptake resulting in weight gain. Substantial weight loss is not driven by changes in lipid removal but by the rate of lipid uptake in adipose tissue. Furthermore, individuals with a low baseline lipid removal rate are more likely to remain weight-stable after weight loss. Therefore, lipid turnover adaptation might be important for maintaining pronounced weight loss. Together these findings identify adipose lipid turnover as an important factor for the long-term development of overweight/obesity and weight loss maintenance in humans.

Insulin action is severely impaired in adipocytes of apparently healthy overweight and obese subjects.

OBJECTIVE: Many overweight/obese subjects appear metabolically healthy with normal in vivo insulin sensitivity. Still, they have increased long-term risk of developing type 2 diabetes. We hypothesized that adipose tissue dysfunction involving decreased insulin action in adipocytes is present in apparently healthy overweight/obese subjects. DESIGN/METHODS: Subjects with normal metabolic health according to Adult Treatment Panel-III or Framingham risk score criteria were subdivided into 67 lean, 32 overweight and 37 obese according to body mass index. They were compared with 200 obese individuals with metabolic syndrome. Insulin sensitivity and maximum action on inhibition of lipolysis and stimulation of lipogenesis was determined in subcutaneous adipocytes. Gene expression was determined by micro-array and qPCR. DNA methylation was assessed by array, pyrosequencing and reporter assays. RESULTS: Compared with lean, adipocytes in overweight/obese displayed marked reductions in insulin sensitivity in both antilipolysis and lipogenesis as well as an attenuated maximum lipogenic response. Among these, only antilipolysis sensitivity correlated with whole-body insulin sensitivity. These differences were already evident in the overweight state, were only slightly worse in the unhealthy obese state and were not related to fat cell size. Adipose tissue analyses linked this to reduced expression of the insulin signalling protein AKT2, which associated with increased methylation at regulatory sites in the AKT2 promoter. CONCLUSIONS: Apparently healthy subjects have severely disturbed adipocyte insulin signalling already in the overweight state which involves epigenetic dysregulation of AKT2. This may constitute an early defect in insulin action that appears even upon modest increases in fat mass.

Improved metabolism and body composition beyond normal levels following gastric bypass surgery: a longitudinal study.

BACKGROUND: The cardiometabolic risk profile improves following bariatric surgery. However, the degree of improvement in relation to weight-stable control subjects is unknown. OBJECTIVES: To study the differences in cardiometabolic risk profile between formerly obese patients following Roux-en-Y gastric bypass (RYGB) surgery and control subjects. METHODS: Subjects undergoing RYGB and reaching a BMI <30 kg m-2 2 years postsurgery were matched with control subjects regarding age, sex and BMI. The following examinations were performed: insulin sensitivity measured by hyperinsulinaemic-euglycaemic clamp, insulin clearance, homeostatic model assessment of insulin resistance (HOMA-IR), lipid profile, inflammatory marker levels, dual-energy X-ray absorptiometry and subcutaneous adipose tissue cellularity (fat cell size and number). RESULTS: Sixty-nine subjects undergoing RYGB were matched to a control subject. Insulin sensitivity measured by hyperinsulinaemic-euglycaemic clamp, blood pressure, inflammatory status and glucose, triglyceride and HDL cholesterol levels were comparable to values of control subjects. However, HOMA-IR (1.0 ± 0.5 vs. 1.3 ± 0.7, P = 0.005), insulin clearance (0.38 ± 0.08 vs. 0.34 ± 0.08 µL m-2  min-1 , P < 0.0001) and circulating levels of insulin (31 ± 15 vs. 37 ± 17 pmol L-1 , P = 0.008), total cholesterol (4.1 ± 0.7 vs. 4.8 ± 0.9 mmol L-1 , P < 0.0001) and LDL cholesterol (2.1 ± 0.6 vs. 2.9 ± 0.8 mmol L-1 , P < 0.0001) were improved beyond the levels in matched control subjects. Furthermore, formerly obese subjects had higher lean and lower fat mass as well as a more benign type of adipose cellularity (hyperplasia with many small fat cells) compared to control subjects. CONCLUSIONS: Subjects who underwent RYGB and reached a postobese state demonstrated a beneficial body composition, slightly increased insulin sensitivity as indirectly measured by HOMA-IR and higher insulin clearance, lower atherogenic lipid/lipoprotein levels and benign adipocyte morphology compared with control subjects who had never been obese. In line with previous results, our findings may in part explain why RYGB confers long-term protection against metabolic complications.

Family history of diabetes is associated with enhanced adipose lipolysis: Evidence for the implication of epigenetic factors.

AIMS: Type 2 diabetes is associated with insulin resistance, adipose hypertrophy and increased lipolysis. The heritability of these traits has been determined by associating them with a family history of diabetes. METHODS: Abdominal subcutaneous fat biopsies were obtained from 581 subjects in a cross-sectional study. Fat cells were isolated, and the difference between measured and expected fat-cell volume was used to determine adipose morphology (degree of hypertrophy or hyperplasia). Spontaneous lipolytic activity was determined in explants of adipose tissue by measuring glycerol release. Insulin-stimulated lipogenesis was assessed by measuring the incorporation of radiolabelled glucose into fat-cell lipids. Information on parental history of diabetes was gathered by a questionnaire. RESULTS: Adipose morphology correlated positively with lipolysis (P<0.0001) and inversely with insulin-stimulated lipogenesis (P<0.008). Also, 24% of probands had a family history of diabetes, which was associated with higher body mass index (BMI) scores, and more insulin resistance (HOMAIR) and adipose hypertrophy. Lipolytic activity was increased, and insulin-stimulated lipogenesis decreased, in probands with a parental history of diabetes. The results for HOMAIR, lipolysis and adipose morphology remained significant after adjusting for proband BMI. A maternal history of diabetes was associated with increased adipose lipolytic activity in probands. CONCLUSION: A family history of diabetes is independent of proband BMI, but associated with adipocyte hypertrophy and enhanced lipolysis, which suggests that these factors are genetically linked to diabetes. Moreover, the influence on lipolysis was only observed in probands with a maternal history of diabetes, thereby supporting an epigenetic impact.

Repin1 deficiency in adipose tissue improves whole-body insulin sensitivity, and lipid metabolism.

BACKGROUND/OBJECTIVES: Replication initiator 1 (Repin1) gene encodes for a zinc-finger protein and has been implicated in the regulation of adipocyte cell size and glucose transport in vitro. Here, we investigate the consequences of reduced adipose tissue (AT) Repin1 expression in vivo. SUBJECTS/METHODS: We have inactivated the Repin1 gene in adipose tissue (iARep-/-) at an age of 4 weeks using tamoxifen-inducible gene targeting strategies on the background of C57BL/6NTac mice. Furthermore, we differentiated human primary adipocytes derived from subcutaneous AT in vitro and knocked down REPIN1 using siRNA technique to measure glycerol release. RESULTS: Conditional Repin1 inactivation results in decreased AT mass, smaller adipocytes in both, subcutaneous and epigonadal AT compared to controls. Compared to controls, iARep-/- mice were more insulin sensitive, had better glucose tolerance and lower LDL-, HDL- and total cholesterol. Significantly lower AT expression of the Repin1 target genes Cd36 and Lcn2 may contribute to the phenotype of iARep-/- mice. Knockdown of REPIN1 in human in vitro differentiated adipocytes revealed an increased glycerol release. CONCLUSIONS: In conclusion, deficiency of Repin1 in AT causes alterations in AT morphology and function, which may underlay lower body weight and improved parameters of insulin sensitivity, glucose and lipid metabolism.

Abdominal subcutaneous adipose tissue cellularity in men and women.

BACKGROUND/OBJECTIVE: Differences in subcutaneous abdominal adipose tissue (SAT) fat cell size and number (cellularity) are linked to insulin resistance. Men are generally more insulin resistant than women but it is unknown whether there is a gender dimorphism in SAT cellularity. The objective was to determine SAT cellularity and its relationship to insulin sensitivity in men and women. METHODS: In a cohort study performed at an outpatient academic clinic in Sweden, 798 women and 306 men were included. Estimated SAT mass (ESAT) was derived from measures of dual-energy X-ray absorptiometry and a formula. SAT biopsies were obtained to measure mean fat cell size; SAT adipocyte number was obtained by dividing ESAT with mean fat cell weight. Fat cell size was also compared with level of insulin sensitivity in vivo. RESULTS: Over the entire range of body mass index (BMI) both fat cell size and number correlated positively with ESAT in either sex. On average, fat cell size was larger in men than in women, which was driven by significantly larger fat cells in non-obese men compared with non-obese women; no gender effect on fat cell size was seen in obese subjects. For all subjects fat cell number was larger in women than men, which was driven by a gender effect among non-obese individuals (P<0.0001). The relationship between fat cell size and insulin resistance was significant in both genders (P<0.0001) but steeper in men than in women (F=19, P<0.0001). CONCLUSIONS: Although both fat cell size and number determine SAT mass, adipocyte number contributes more and size less in women than in men and this is most evident in non-obese subjects. Over the entire BMI range, fat cell size contributes stronger to insulin resistance in men.

Cardiovascular risk score is linked to subcutaneous adipocyte size and lipid metabolism.

OBJECTIVE: Although white adipose tissue mass and distribution correlates with cardiovascular disease, the fat cell-specific perturbations underlying this association are not known. We determined the relationship between adipocyte size and lipid metabolism with cardiovascular risk. DESIGN/SUBJECTS: Adipocyte size as well as spontaneous (basal) and hormone-stimulated effects on adipocyte lipid metabolism (lipolysis and lipogenesis) were investigated in abdominal subcutaneous adipose tissue of 304 men and 775 women. Subjects were classified into five categories according to Adult Treatment Panel III (ATPIII) metabolic syndrome criteria. RESULTS: Adipocyte size increased with increasing ATPIII score (P < 0.0001). For lipolysis, there was a gradual increase in basal and catecholamine-stimulated lipolysis and a decrease in insulin-mediated inhibition of stimulated lipolysis with ATPIII (P < 0.0001). In contrast, the lipolytic action of atrial natriuretic peptide was similar between ATPIII classes. Basal and insulin-stimulated lipogenesis decreased with increasing score (P < 0.0001). Circulating free fatty acid levels were 50% higher in the top risk category (4-5) compared with the lowest score (P < 0.0001). Fat cell size correlated positively with increasing ATPIII score and lipolysis but negatively with lipogenesis. All these differences were independent of age, sex and body weight status (P < 0.0001 to 0.02 after correction). When all functional measures were put together, maximum insulin-stimulated lipogenesis, insulin-antilipolytic sensitivity and basal lipolysis together explained about 20% in the variation of ATPIII in score. CONCLUSIONS: Independently of sex, age and body weight status, a high cardiovascular risk score associates with increased circulating free fatty acid levels and hormone-specific alterations of lipolysis/lipogenesis in enlarged subcutaneous fat cells.

Depot-specific differences in fatty acid composition and distinct associations with lipogenic gene expression in abdominal adipose tissue of obese women.

Cardiometabolic diseases are primarily linked to enlarged visceral adipose tissue (VAT). However, some data suggest heterogeneity within the subcutaneous adipose tissue (SAT) depot with potential metabolic differences between the superficial SAT (sSAT) and deep SAT (dSAT) compartments. We aimed to investigate the heterogeneity of these three depots with regard to fatty acid (FA) composition and gene expression. Adipose tissue biopsies were collected from 75 obese women undergoing laparoscopic gastric bypass surgery. FA composition and gene expression were determined with gas chromatography and quantitative real-time-PCR, respectively. Stearoyl CoA desaturase-1 (SCD-1) activity was estimated by product-to-precursor FA ratios. All polyunsaturated FAs (PUFA) with 20 carbons were consistently lower in VAT than either SAT depots, whereas essential PUFA (linoleic acid, 18:2n-6 and α-linolenic acid, 18:3n-3) were similar between all three depots. Lauric and palmitic acid were higher and lower in VAT, respectively. The SCD-1 product palmitoleic acid as well as estimated SCD-1 activity was higher in VAT than SAT. Overall, there was a distinct association pattern between lipid metabolizing genes and individual FAs in VAT. In conclusion, SAT and VAT are two distinct depots with regard to FA composition and expression of key lipogenic genes. However, the small differences between sSAT and dSAT suggest that FA metabolism of SAT is rather homogenous.

Impaired atrial natriuretic peptide-mediated lipolysis in obesity.

BACKGROUND: Catecholamines and natriuretic peptides (NPs) are the only hormones with pronounced lipolytic effects in human white adipose tissue. Although catecholamine-induced lipolysis is well known to be impaired in obesity and insulin resistance, it is not known whether the effect of NPs is also altered. METHODS: Catecholamine- and atrial NP (ANP)-induced lipolysis was investigated in abdominal subcutaneous adipocytes in vitro and in situ by microdialysis. RESULTS: In a cohort of 122 women, both catecholamine- and ANP-induced lipolysis in vitro was markedly attenuated in obesity (n=87), but normalized after substantial body weight loss (n=52). The impairment of lipolysis differed between the two hormones when expressing lipolysis per lipid weight, the ratio of stimulated over basal (spontaneous) lipolysis rate or per number of adipocytes. Thus, while the response to catecholamines was lower when expressed as the former two measures, it was higher when expressed per cell number, a consequence of the significantly larger fat cell size in obesity. In contrast, although ANP-induced lipolysis was also attenuated when expressed per lipid weight or the ratio stimulated/basal, it was similar between non-obese and obese subjects when expressed per cell number suggesting that the lipolytic effect of ANP may be even more sensitive to the effects of obesity than catecholamines. Obesity was characterized by a decrease in the protein expression of the signaling NP A receptor (NPRA) and a trend toward increased levels of the clearance receptor NPRC. The impairment in ANP-induced lipolysis observed in vitro was corroborated by microdialysis experiments in situ in a smaller cohort of lean and overweight men. CONCLUSIONS: ANP- and catecholamine-induced lipolysis is reversibly attenuated in obesity. The pro-lipolytic effects of ANP are relatively more impaired compared with that of catecholamines, which may in part be due to specific changes in NP receptor expression.

Low early B-cell factor 1 (EBF1) activity in human subcutaneous adipose tissue is linked to a pernicious metabolic profile.

AIM: Recently, in both human and murine white adipose tissue (WAT), transcription factor early B-cell factor 1 (EBF1) has been shown to regulate adipocyte differentiation, adipose morphology and triglyceride hydrolysis (lipolysis). This study investigated whether EBF1 expression and biological activity in WAT is related to different metabolic parameters. METHODS: In this cross-sectional study of abdominal subcutaneous WAT, EBF1 protein levels were examined in 18 non-obese subjects, while biological activity was determined in 56 obese and non-obese subjects. Results were assessed by anthropometric measures and blood pressure as well as by plasma lipid levels and insulin sensitivity. RESULTS: EBF1 protein levels were negatively associated with waist circumference (r=-0.56; P=0.015), but not with body mass index (BMI) or body fat (P=0.10-0.29). Biological activity of EBF1 correlated negatively with plasma triglycerides (r=-0.46; P=0.0005) and plasma insulin (r=-0.39; P=0.0027), but positively with plasma HDL cholesterol (r=0.48; P=0.0002) and insulin sensitivity, as assessed by intravenous insulin tolerance test (r=0.64; P<0.0001). These relationships, except for plasma insulin, remained statistically significant after adjusting for BMI and adipose morphology. EBF1 activity was not associated with age, systolic/diastolic blood pressure or total plasma cholesterol (P=0.17-0.48). In contrast to EBF1 activity, after adjusting for BMI, EBF1 mRNA levels displayed only an association with plasma triglycerides. CONCLUSION: Low EBF1 protein expression and activity in abdominal subcutaneous WAT is a BMI-independent marker for several traits associated with the metabolic syndrome. However, whether EBF1 constitutes a novel treatment target remains to be demonstrated.

The fat cell epigenetic signature in post-obese women is characterized by global hypomethylation and differential DNA methylation of adipogenesis genes.

BACKGROUND/OBJECTIVES: Obese subjects have increased number of enlarged fat cells that are reduced in size but not in number in post-obesity. We performed DNA methylation profiling in fat cells with the aim of identifying differentially methylated DNA sites (DMS) linked to adipose hyperplasia (many small fat cells) in post-obesity. SUBJECTS/METHODS: Genome-wide DNA methylation was analyzed in abdominal subcutaneous fat cells from 16 women examined 2 years after gastric bypass surgery at a post-obese state (body mass index (BMI) 26±2 kg m(-2), mean±s.d.) and from 14 never-obese women (BMI 25±2 kg m(-2)). Gene expression was analyzed in subcutaneous adipose tissue from nine women in each group. In a secondary analysis, we examined DNA methylation and expression of adipogenesis genes in 15 and 11 obese women, respectively. RESULTS: The average degree of DNA methylation of all analyzed CpG sites was lower in fat cells from post-obese as compared with never-obese women (P=0.014). A total of 8504 CpG sites were differentially methylated in fat cells from post-obese versus never-obese women (false discovery rate 1%). DMS were under-represented in CpG islands and surrounding shores. The 8504 DMS mapped to 3717 unique genes; these genes were over-represented in cell differentiation pathways. Notably, 27% of the genes linked to adipogenesis (that is, 35 of 130) displayed DMS (adjusted P=10(-8)) in post-obese versus never-obese women. Next, we explored DNA methylation and expression of genes linked to adipogenesis in more detail in adipose tissue samples. DMS annotated to adipogenesis genes were not accompanied by differential gene expression in post-obese compared with never-obese women. In contrast, adipogenesis genes displayed differential DNA methylation accompanied by altered expression in obese women. CONCLUSIONS: Global CpG hypomethylation and over-representation of DMS in adipogenesis genes in fat cells may contribute to adipose hyperplasia in post-obese women.

Adipose tissue morphology predicts improved insulin sensitivity following moderate or pronounced weight loss.

BACKGROUND: Cross-sectional studies show that white adipose tissue hypertrophy (few, large adipocytes), in contrast to hyperplasia (many, small adipocytes), associates with insulin resistance and increased risk of developing type 2 diabetes. We investigated if baseline adipose cellularity could predict improvements in insulin sensitivity following weight loss. METHODS: Plasma samples and subcutaneous abdominal adipose biopsies were examined in 100 overweight or obese individuals before and 10 weeks after a hypocaloric diet (7±3% weight loss) and in 61 obese subjects before and 2 years after gastric by-pass surgery (33±9% weight loss). The degree of adipose tissue hypertrophy or hyperplasia (termed the morphology value) in each individual was calculated on the basis of the relationship between fat cell volume and total fat mass. Insulin sensitivity was determined by homeostasis model assessment-estimated insulin resistance (HOMAIR). RESULTS: In both cohorts at baseline, subjects with hypertrophy displayed significantly higher fasting plasma insulin and HOMAIR values than subjects with hyperplasia (P<0.0001), despite similar total fat mass. Plasma insulin and HOMAIR were normalized in both cohorts following weight loss. The improvement (delta insulin or delta HOMAIR) was more pronounced in individuals with hypertrophy, irrespective of whether adipose morphology was used as a continuous (P=0.0002-0.027) or nominal variable (P=0.002-0.047). Absolute adipocyte size associated (although weaker than morphology) with HOMAIR improvement only in the surgery cohort. Anthropometric measures at baseline (fat mass, body mass index, waist-to-hip ratio or waist circumference) showed no significant association with delta insulin or delta HOMAIR. CONCLUSIONS: In contrast to anthropometric variables or fat cell size, subcutaneous adipose morphology predicts improvement in insulin sensitivity following both moderate and pronounced weight loss in overweight/obese subjects.

Regional variations in the relationship between arterial stiffness and adipocyte volume or number in obese subjects.

BACKGROUND: Cardiovascular disease is associated with multiple risk factors including stiff arteries and large adipocytes. Whether the latter two are interrelated is unknown. We aimed to determine whether arterial stiffness is associated with fat cell size and number in subcutaneous or visceral white adipose tissue (WAT). METHODS: A cross-sectional study of 120 obese subjects scheduled for bariatric surgery in whom WAT mass and distribution was assessed by dual-X-ray absorptiometry. Biopsies from visceral (greater omentum) and subcutaneous (abdominal) WAT were obtained to calculate fat cell volume and number. Arterial stiffness was determined as aortic pulse wave velocity (PWV). RESULTS: Visceral adipocyte volume, but not number, was strongly (P<0.0001) and positively correlated with PWV, explaining 20% of the inter-individual variations in this parameter. This relationship remained significant after correction for clinical confounders. PWV correlated positively (r=0.38, P<0.0001) with visceral (but not subcutaneous) WAT mass. Furthermore, PWV was also positively associated with subcutaneous adipocyte volume (r=0.20, P=0.031) and negatively with fat cell number (r=-0.26, P=0.006). However, the relationships between PWV and visceral WAT mass or subcutaneous fat cell size/number became non-significant when controlling for visceral fat cell volume. In a multiple regression analysis to determine the factors that explain variations in PWV, only visceral fat cell volume, age, pulse rate and diastolic blood pressure entered the model, together explaining 42% of the variation in PWV. CONCLUSIONS: Visceral fat cell volume was the only WAT parameter that constituted an independent and significant, positive regressor for arterial stiffness determined by PWV. Although a causal relationship is not established, visceral fat cell volume may explain the well-known correlation between central fat mass, arterial stiffness and cardiovascular risk, at least in severely/morbidly obese subjects.

Semaphorin 3C is a novel adipokine linked to extracellular matrix composition.

AIMS/HYPOTHESIS: Alterations in white adipose tissue (WAT) function, including changes in protein (adipokine) secretion and extracellular matrix (ECM) composition, promote an insulin-resistant state. We set out to identify novel adipokines regulated by body fat mass in human subcutaneous WAT with potential roles in adipose function. METHODS: Adipose transcriptome data and secretome profiles from conditions with increased/decreased WAT mass were combined. WAT donors were predominantly women. In vitro effects were assessed using recombinant protein. Results were confirmed by quantitative PCR/ELISA, metabolic assays and immunochemistry in human WAT and adipocytes. RESULTS: We identified a hitherto uncharacterised adipokine, semaphorin 3C (SEMA3C), the expression of which correlated significantly with body weight, insulin resistance (HOMA of insulin resistance [HOMAIR], and the rate constant for the insulin tolerance test [KITT]) and adipose tissue morphology (hypertrophy vs hyperplasia). SEMA3C was primarily found in mature adipocytes and had no direct effect on human adipocyte differentiation, lipolysis, glucose transport or the expression of ß-oxidation genes. This could in part be explained by the significant downregulation of its cognate receptors during adipogenesis. In contrast, in pre-adipocytes, SEMA3C increased the production/secretion of several ECM components (fibronectin, elastin and collagen I) and matricellular factors (connective tissue growth factor, IL6 and transforming growth factor-ß1). Furthermore, the expression of SEMA3C in human WAT correlated positively with the degree of fibrosis in WAT. CONCLUSIONS/INTERPRETATION: SEMA3C is a novel adipokine regulated by weight changes. The correlation with WAT hypertrophy and fibrosis in vivo, as well as its effects on ECM production in human pre-adipocytes in vitro, together suggest that SEMA3C constitutes an adipocyte-derived paracrine signal that influences ECM composition and may play a pathophysiological role in human WAT.

Provitamin A carotenoids are independently associated with matrix metalloproteinase-9 in plasma samples from a general population.

BACKGROUND AND AIM: Carotenoids in plasma are inversely associated with cardiovascular risk. Low levels can be explained by low dietary intake but also by a number of other factors including inflammatory activity. Given that matrix metalloproteinase (MMP)-9 has an important role in inflammation and cardiovascular disease, we hypothesized that circulating MMP-9 levels would be inversely related to total or single carotenoids in a general population cohort. METHODS: A well-characterized population-based cohort of 285 Swedish men and women (45-69 years) was used for the present study. The intake of carotenoid-rich fruits and vegetables was estimated from a food frequency questionnaire. Levels of MMP-9, C-reactive protein (CRP), interleukin (IL)-6 and six major carotenoids [ß-cryptoxanthine, α-carotene, ß-carotene, lutein (+zeaxanthin) and lycopene] were determined in plasma. RESULTS: Lower plasma levels of total and single carotenoids were associated with lower dietary intake of carotenoids, older age, male sex, lower physical activity, higher alcohol consumption, higher body mass index (BMI), higher systolic and diastolic blood pressures, lower levels of total cholesterol and HDL cholesterol and higher levels of CRP, IL-6 and MMP-9. After multivariate adjustments, plasma levels of total carotenoids and provitamin A carotenoids (ß-cryptoxanthine, α-carotene and ß-carotene) remained independently associated with sex, dietary intake of carotenoids, BMI, HDL cholesterol and MMP-9, whilst associations with CRP and IL-6 were not maintained. Neither dietary intake of carotenoid-rich fruits and vegetables, nor vitamin supplement use was associated with MMP-9, CRP or IL-6 levels. CONCLUSION: Plasma carotenoids were associated with a variety of factors including age, sex, dietary intake and metabolic variables. A new finding was the independent relationship in plasma between low provitamin A carotenoids and high MMP-9, suggesting a link between these carotenoids, matrix turnover and arterial remodelling.

Liver X receptors and fat cell metabolism.

Liver X receptors (LXRs) are members of the nuclear receptor family and are present in two isoforms, α and ß, encoded by two separate genes. Originally described in the liver, LXRs have in the last 15 years been implicated in central metabolic pathways, including bile acid synthesis, lipid and glucose homeostasis. Although the vast majority of studies have been performed in non-adipose cells/tissues, results in recent years suggest that LXRs may have important modulatory roles in adipose tissue and adipocytes. Although several authors have published reviews on LXR, there have been no attempts to summarize the effects reported specifically in adipose systems. This overview gives a brief introduction to LXR and describes the sometimes-contradictory results obtained in murine cell systems and in rodent adipose tissue. The so far very limited number of studies performed in human adipocytes and adipose tissue are also presented. It should be apparent that although LXR may impact on several different pathways in metabolism, the clinical role of LXR modulation in adipose tissue is still not clear.