Exploring differential item functioning in the Western Ontario and McMaster Universities osteoarthritis index (WOMAC)

Background: The Western Ontario and McMaster Universities Osteoarthritis Index (WOMAC) is a widely used patient reported outcome in osteoarthritis. An important, but frequently overlooked, aspect of validating health outcome measures is to establish if items exhibit differential item functioning (DIF). That is, if respondents have the same underlying level of an attribute, does the item give the same score in different subgroups or is it biased towards one subgroup or another. The aim of the study was to explore DIF in the Likert format WOMAC for the first time in a UK osteoarthritis population with respect to demographic, social, clinical and psychological factors. Methods: The sample comprised a community sample of 763 people with osteoarthritis who participated in the Somerset and Avon Survey of Health. The WOMAC was explored for DIF by gender, age, social deprivation, social class, employment status, distress, body mass index and clinical factors. Ordinal regression models were used to identify DIF items. Results: After adjusting for age, two items were identified for the physical functioning subscale as having DIF with age identified as the DIF factor for 2 items, gender for 1 item and body mass index for 1 item. For the WOMAC pain subscale, for people with hip osteoarthritis one item was identified with age-related DIF. The impact of the DIF items rarely had a significant effect on the conclusions of group comparisons. Conclusions: Overall, the WOMAC performed well with only a small number of DIF items identified. However, as DIF items were identified in for the WOMAC physical functioning subscale it would be advisable to analyse data taking into account the possible impact of the DIF items when weight, gender or especially age effects, are the focus of interest in UK-based osteoarthritis studies. Similarly for the WOMAC pain subscale in people with hip osteoarthritis it would be worthwhile to analyse data taking into account the possible impact of the DIF item when age comparisons are of primary interest

Structure-Function Analysis of Diacylglycerol Acyltransferase Sequences from 70 Organisms

<p>Abstract</p> <p>Background</p> <p>Diacylglycerol acyltransferase families (DGATs) catalyze the final and rate-limiting step of triacylglycerol (TAG) biosynthesis in eukaryotic organisms. Understanding the roles of DGATs will help to create transgenic plants with value-added properties and provide clues for therapeutic intervention for obesity and related diseases. The objective of this analysis was to identify conserved sequence motifs and amino acid residues for better understanding of the structure-function relationship of these important enzymes.</p> <p>Results</p> <p>117 DGAT sequences from 70 organisms including plants, animals, fungi and human are obtained from database search using tung tree DGATs. Phylogenetic analysis separates these proteins into DGAT1 and DGAT2 subfamilies. These DGATs are integral membrane proteins with more than 40% of the total amino acid residues being hydrophobic. They have similar properties and amino acid composition except that DGAT1s are approximately 20 kDa larger than DGAT2s. DGAT1s and DGAT2s have 41 and 16 completely conserved amino acid residues, respectively, although only two of them are shared by all DGATs. These residues are distributed in 7 and 6 sequence blocks for DGAT1s and DGAT2s, respectively, and located at the carboxyl termini, suggesting the location of the catalytic domains. These conserved sequence blocks do not contain the putative neutral lipid-binding domain, mitochondrial targeting signal, or ER retrieval motif. The importance of conserved residues has been demonstrated by site-directed and natural mutants.</p> <p>Conclusions</p> <p>This study has identified conserved sequence motifs and amino acid residues in all 117 DGATs and the two subfamilies. None of the completely conserved residues in DGAT1s and DGAT2s is present in recently reported isoforms in the multiple sequences alignment, raising an important question how proteins with completely different amino acid sequences could perform the same biochemical reaction. The sequence analysis should facilitate studying the structure-function relationship of DGATs with the ultimate goal to identify critical amino acid residues for engineering superb enzymes in metabolic engineering and selecting enzyme inhibitors in therapeutic application for obesity and related diseases.</p

Paradoxical Increase in TAG and DAG Content Parallel the Insulin Sensitizing Effect of Unilateral DGAT1 Overexpression in Rat Skeletal Muscle

BACKGROUND: The involvement of muscle triacylglycerol (TAG) storage in the onset of insulin resistance is questioned and the attention has shifted towards inhibition of insulin signalling by the lipid intermediate diacylglycerol (DAG). The enzyme 1,2-acylCoA:diacylglyceroltransferase-1 (DGAT1) esterifies a fatty acyl-CoA on DAG to form TAG. Therefore, the aim of the present study was to investigate if unilateral overexpression of DGAT1 in adult rat Tibialis anterior (TA) muscle will increase conversion of the lipid intermediate DAG into TAG, thereby improving muscle insulin sensitivity. METHODOLOGY/PRINCIPAL FINDINGS: The DGAT1 gene construct was injected in the left TA muscle of male rats on chow or high-fat (45% kcal) diet for three weeks, followed by application of one 800 V/cm and four 80 V/cm pulses, using the contralateral leg as sham-electroporated control. Seven days after electroporation, muscle specific insulin sensitivity was assessed with a hyperinsulinemic euglycemic clamp using 2-deoxy-[3H]glucose. Here, we provide evidence that unilateral overexpression of DGAT1 in TA muscle of male rats is associated with an increased rather than decreased DAG content. Strikingly, this increase in DAG content was accompanied by improved muscle insulin sensitivity. Interestingly, markers of muscle lipolysis and mitochondrial function were also increased in DGAT1 overexpressing muscle. CONCLUSIONS/SIGNIFICANCE: We conclude that unilateral DGAT1 overexpression can rescue insulin sensitivity, possibly by increasing DAG and TAG turnover in skeletal muscle. In case of a proper balance between the supply and oxidation of fatty acids in skeletal muscle, the lipid intermediate DAG may not exert harmful effects on insulin signalling

Nanovesicles from adipose-derived mesenchymal stem cells inhibit T lymphocyte trafficking and ameliorate chronic experimental autoimmune encephalomyelitis

Cell based-therapies represent promising strategies for the treatment of neurological diseases. We have previously shown that adipose stem cells (ASC) ameliorate chronic experimental autoimmune encephalomyelitis (EAE). Recent evidence indicates that most ASC paracrine effects are mediated by extracellular vesicles, i.e. micro- and nanovesicles (MVs and NVs). We show that preventive intravenous administration of NVs isolated from ASC (ASC-NVs) before disease onset significantly reduces the severity of EAE and decreases spinal cord inflammation and demyelination, whereas therapeutic treatment with ASC-NVs does not ameliorate established EAE. This treatment marginally inhibits antigen-specific T cell activation, while reducing microglial activation and demyelination in the spinal cord. Importantly, ASC-NVs inhibited integrin-dependent adhesion of encephalitogenic T cells in vitro, with no effect on adhesion molecule expression. In addition, intravital microscopy showed that encephalitogenic T cells treated with ASC NVs display a significantly reduced rolling and firm adhesion in inflamed spinal cord vessels compared to untreated cells. Our results show that ASC-NVs ameliorate EAE pathogenesis mainly by inhibiting T cell extravasation in the inflamed CNS, suggesting that NVs may represent a novel therapeutic approach in neuro-inflammatory diseases, enabling the safe administration of ASC effector factors

Peroxisome proliferator-activated receptor-γ coactivator-1 and insulin resistance: acute effect of fatty acids

Aims/hypothesis Peroxisome proliferator-activated receptor (PPAR)-&gamma; coactivator-1 (PPARGC1), a coactivator regulating the transcription of genes involved in oxidative metabolism, is downregulated in patients with type 2 diabetes and in their first-degree relatives. Whether this downregulation is a cause or effect of early aberrations in the development of insulin resistance, such as disturbances in fat metabolism, is unknown. We examined whether lipid-induced insulin resistance was associated with downregulation of expression of skeletal muscle genes involved in oxidative metabolism and mitochondrial biogenesis in humans.Materials and methods Nine healthy lean male subjects underwent a 6-h hyperinsulinaemic&ndash;euglycaemic clamp with simultaneous infusion of either a lipid emulsion or glycerol as a control. Blood was sampled at regular time points and muscle biopsies were taken before and after every test. Intramuscular triacylglycerol (IMTG) content was determined by Oil Red O staining and gene expression was measured by quantitative PCR.Results Lipid infusion resulted in a &sim;2.7-fold increase in plasma NEFA levels and a 31&plusmn;6% decrease in insulin sensitivity (p=0.001). The infusion of lipids resulted in a &sim;1.6-fold increase in IMTG (p=0.02), whereas during the clamp with glycerol infusion IMTG tended to decrease to &sim;53% of preinfusion levels (p=0.065). Lipid infusion decreased PPARGC1A, PPARGC1B and PPARA expression to &sim;61, 77 and &sim;52% of basal values respectively, whereas expression of uncoupling protein 3 was upregulated 1.8-fold (all p&lt;0.05).Conclusions/interpretation Acute elevation of plasma NEFA levels, leading to muscular fat accumulation and insulin resistance, downregulates PPARGC1A, PPARGC1B and PPARA expression, suggesting that the decrease in PPARGC1 expression observed in the (pre)diabetic state may be the result, rather than the cause of lipid-induced insulin resistance.<br /