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There are currently 3 established techniques employed routinely to determine the risk of foot ulceration in the patient with diabetes mellitus. These are the assessment of circulation, neuropathy, and foot pressure. These assessments are widely used clinically as well as in the research domain with an aim to prevent the onset of foot ulceration. Routine neuropathic evaluation includes the assessment of sensory loss in the plantar skin of the foot using both the Semmes Weinstein monofilament and the biothesiometer. Thermological measurements of the foot to assess responses to thermal stimuli and cutaneous thermal discrimination threshold are relatively uncommon. Indeed, there remains uncertainty regarding the importance of thermal changes in the development of foot ulcers. Applications of thermography and thermometry in lower extremity wounds, vascular complications, and neuropathic complications have progressed as a result of improved imaging software and transducer technology. However, the uncertainty associated with the specific thermal modality, the costs, and processing times render its adaptation to the clinic. Therefore, wider adoption of thermological measurements has been limited. This article reviews thermal measurement techniques specific to diabetic foot such as electrical contact thermometry, cutaneous thermal discrimination thresholds, infrared thermography, and liquid crystal thermography. [ABSTRACT FROM AUTHOR]

Study Design: Secondary analysis of an observational cohort study., Objectives: To determine if the perceived overall limitation of neuropathic pain on activities (NP limitation) covaries in 26 specific life activities in people with SCI, taking into account the severity of injury., Setting: Community-based in Canada., Methods: Secondary analysis of a cohort (N = 1481) was performed using questionnaires to rate NP frequency and limitation on activities as well as participation in 26 life activities. Relative risks (RR) analyses using Poisson regression were used to examine the data., Results: Most participants (N = 1158; 78%) reported living with NP (from once a year to every day). When NP limitation was described as "not at all" or "very little" (N = 394; 34%), there was no statistically significant RR (p > 0.0019), suggesting no additional risk of not participating "as much as wanted" in any of activities compared to participants with no NP. When NP limitation was described as "to some extent" (N = 411; 35%), a significantly higher risk was observed for 5 of the 26 activities (1.34 < RR < 1.62), and for 23 activities with a large range (1.24 < RR < 3.20) when NP limitation was rated as "to a great extent or more" (N = 353; 31%)., Conclusion: The variation of RR observed across the 26 activities suggests that the NP limitation may not be general but rather related to specific activity characteristics, which should be taken into account when evaluating NP limitations with a specific focus on the life activities being affected.

The TetraGrip is an inertial measurement unit-controlled surface upper limb FES device developed for improving hand functions of people with tetraplegia. The reliability of the control system and the repeatability and reproducibility of the device were assessed by analysing the results obtained when 14 able-bodied volunteers used the device. These volunteers were able to generate the control signals effectively once they had sufficient training. The two tetraplegic volunteers participated in a 12-week long clinical study (exercise, 4 weeks; functional tasks, 8 weeks), where they used the device to perform functional tasks. Outcome measures used were the grasp release test, the grip strength test, and the box and block test. Both tetraplegic volunteers showed improvement in performing the tasks specified in all outcome measures. The TetraGrip performed as intended when the able-bodied volunteers used it, and it improved the hand functions of both volunteers with tetraplegia. However, a larger clinical study is necessary to assess the performance of the device with a wider range of people with tetraplegia such as those with C5 complete/incomplete.

Cutaneous sun exposure is an important determinant of circulating vitamin D. Both sun exposure and vitamin D have been inversely associated with risk of autoimmune disease. In juvenile idiopathic arthritis (JIA), low circulating vitamin D appears common, but disease-related behavioral changes may have influenced sun exposure. We therefore aimed to determine whether predisease sun exposure is associated with JIA. Using validated questionnaires, we retrospectively measured sun exposure for 202 Caucasian JIA case-control pairs born in Victoria Australia, matched for birth year and time of recruitment. Measures included maternal sun exposure at 12 weeks of pregnancy and child sun exposure across the life-course prediagnosis. We converted exposure to UVR dose and looked for case-control differences using logistic regression, adjusting for potential confounders. Higher cumulative prediagnosis UVR exposure was associated with reduced risk of JIA, with a clear dose-response relationship (trend P = 0.04). UVR exposure at 12 weeks of pregnancy was similarly inversely associated with JIA (trend P = 0.011). Associations were robust to sensitivity analyses for prediagnosis behavioral changes, disease duration and knowledge of the hypothesis. Our data indicate that lower UVR exposure may increase JIA risk. This may be through decreased circulating vitamin D, but prospective studies are required to confirm this., (© 2018 The American Society of Photobiology.)

Background: Various types of limitations on community participation are experienced by people with spinal cord injury (SCI)., Objectives: To determine: 1) the perceived influence of six intrinsic/extrinsic factors (i.e. physical impairment, emotional condition, thinking skills, environment, lack of assistance, discrimination) on participation in 26 life activities, 2) if this influence varied based on extent of participation, and 3) if personal or environmental characteristics influenced perceptions., Methods: Secondary analysis of a cohort (SCI Community Survey, n = 1508) using the SCI Person-Perceived Participation in Daily Activities Questionnaire. Frequency tables, Fisher's exact tests and correspondence analyses., Results: Respectively, 79.6% and 38.5% of respondents perceived that their physical impairment and the natural and/or built environment were the main factors that limited participation across all activities. Considering participation between three groups (no participation; less than wanted; as much as wanted), significant differences (p < 0.001) of perceptions were observed in 65% of the combinations (26 activities x 6 factors). The hypothesis that respondents who did not participate would perceive the highest proportion of limitations was confirmed in 41% of the combinations. Perceived influence of the intrinsic/extrinsic factors on participation was not significantly influenced by other personal or environmental characteristics., Conclusion: A majority of people with SCI perceived that their participation is limited by one or more of intrinsic/extrinsic factors. Perceptions regarding which factors influence participation differ between activities and these perceptions appear related to the extent of participation suggesting that those who actively participate could be the most sensitive to limitations in certain activities., (Copyright © 2018 Elsevier Inc. All rights reserved.)

Elevated urinary albumin excretion (microalbuminuria) is an early marker of diabetic nephropathy, but there is an unmet need for better biomarkers that capture the individuals at risk with higher accuracy and earlier than the current markers do. We performed an untargeted metabolomic study to assess baseline differences between individuals with type 1 diabetes who either developed microalbuminuria or remained normoalbuminuric. A total of 102 individuals progressed to microalbuminuria during a median follow-up of 3.2 years, whereas 98 sex-, age- and body mass index (BMI) matched non-progressors remained normoalbuminuric during a median follow-up of 7.1 years. Metabolomic screening identified 1,242 metabolites, out of which 111 differed significantly between progressors and non-progressors after adjustment for age of diabetes onset, baseline glycosylated hemoglobin A1c (HbA 1c ), and albumin excretion rate (AER). The metabolites that predicted development of microalbumiuria included several uremic toxins and carnitine metabolism related molecules. Iterative variable selection indicated erythritol, 3-phenylpropionate, and N-trimethyl-5-aminovalerate as the best set of variables to predict development of microalbuminuria. A metabolomic index based on these metabolites improved the prediction of incident microalbuminuria on top of the clinical variables age of diabetes onset, baseline HbA 1c and AER (ROC AUC  = 0.842 vs 0.797), highlighting their ability to predict early-phase diabetic nephropathy.

Context: Renal disease in type 2 diabetes mellitus (T2DM) is associated with excess morbidity/mortality. Although estimated glomerular filtration rate (eGFR) and albuminuria are routine for assessing renal impairment, novel biomarkers could improve risk stratification and prediction., Objective: To identify specific biomarkers of progression of renal dysfunction., Design: Prospective observational., Setting: Academic diabetes clinics., Patients: A total of 286 T2DM patients (age, 62 ± 8 y; glycosylated hemoglobin, 7.2 ± 0.9%; eGFR, 85 ± 20 mL · min(-1) · 1.73 m(2))., Interventions: None., Main Outcome Measures: Progression of eGFR and albuminuria., Results: We performed screening metabolomics in serum and urine samples by gas chromatography/mass spectroscopy (MS) and ultra-high performance liquid chromatography/MS/MS. Biomarker identification was performed by random forest using an eGFR cutoff of < 60 mL · min(-1) · 1.73 m(2) or an albumin/creatinine ratio (ACR) cutoff ≥ 30 mg/g as response variables. At follow-up, eGFR had declined by 16 [9] (median [interquartile ratio]) mL · min(-1) · 1.73 m(2), and ACR had increased by 41 [135] mg/g in patients in the respective top quartile of changes from baseline. Clinical parameters (gender, age, fasting glucose, and baseline eGFR) predicted outcome, with receiver operator characteristics curve (ROC) = 0.671. The five serum metabolites best correlated with either eGFR < 60 or ACR ≥ 30 at baseline were tested for their ability to improve clinical prediction. The sum of C-glycosyl tryptophan, pseudouridine, and N-acetylthreonine (MetIndex) raised the ROC to 0.739 (P < .0001). eGFR decline was predicted by the top MetIndex quartile (odds ratio = 5.48 [95% confidence interval, 2.23-14.47]). MetIndex also predicted an ACR increase with an odds ratio of 2.82 [1.20-7.03] and a ROC of 0.750. Top urine metabolites did not add significant predictivity., Conclusions: A limited number of circulating intermediates of amino acid and nucleotide pathways carry clinically significant predictivity for deterioration of renal function in well-controlled T2DM.

Objective: The objective was to test the clinical utility of Quantose M(Q) to monitor changes in insulin sensitivity after pioglitazone therapy in prediabetic subjects. Quantose M(Q) is derived from fasting measurements of insulin, α-hydroxybutyrate, linoleoyl-glycerophosphocholine, and oleate, three nonglucose metabolites shown to correlate with insulin-stimulated glucose disposal., Research Design and Methods: Participants were 428 of the total of 602 ACT NOW impaired glucose tolerance (IGT) subjects randomized to pioglitazone (45 mg/d) or placebo and followed for 2.4 years. At baseline and study end, fasting plasma metabolites required for determination of Quantose, glycated hemoglobin, and oral glucose tolerance test with frequent plasma insulin and glucose measurements to calculate the Matsuda index of insulin sensitivity were obtained., Results: Pioglitazone treatment lowered IGT conversion to diabetes (hazard ratio = 0.25; 95% confidence interval = 0.13-0.50; P < .0001). Although glycated hemoglobin did not track with insulin sensitivity, Quantose M(Q) increased in pioglitazone-treated subjects (by 1.45 [3.45] mg·min(-1)·kgwbm(-1)) (median [interquartile range]) (P < .001 vs placebo), as did the Matsuda index (by 3.05 [4.77] units; P < .0001). Quantose M(Q) correlated with the Matsuda index at baseline and change in the Matsuda index from baseline (rho, 0.85 and 0.79, respectively; P < .0001) and was progressively higher across closeout glucose tolerance status (diabetes, IGT, normal glucose tolerance). In logistic models including only anthropometric and fasting measurements, Quantose M(Q) outperformed both Matsuda and fasting insulin in predicting incident diabetes., Conclusions: In IGT subjects, Quantose M(Q) parallels changes in insulin sensitivity and glucose tolerance with pioglitazone therapy. Due to its strong correlation with improved insulin sensitivity and its ease of use, Quantose M(Q) may serve as a useful clinical test to identify and monitor therapy in insulin-resistant patients.

Functional Electrical Stimulation (FES) is a technique that uses electricity to activate the nerves of a muscle that is paralysed due to hemiplegia, multiple sclerosis, Parkinson's disease or spinal cord injury (SCI). FES has been widely used to restore upper limb functions in people with hemiplegia and C5-C7 tetraplegia and has improved their ability to perform their activities of daily living (ADL). At the time of writing, a detailed literature review of the existing upper limb FES devices and their man-machine interfaces (MMI) showed that only the NESS H200 was commercially available. However, the rigid arm splint doesn't fit everyone and prevents the use of a tenodesis grip. Hence, a robust and versatile upper limb FES device that can be used by a wider group of people is required.

Objective: The purpose of this study was to determine whether implementation of a Pressure Ulcer Prevention Initiative (PUPI) changed the assessment and treatment of patients with a traumatic spinal cord injury (SCI) in an acute care setting, and improved patient outcomes., Method: The success of implementation was evaluated by examining the percentage of patients with completed occupational therapist (OT) skin care assessments and prescriptions for therapeutic support surfaces (TSS; i.e., mattresses) before implementation (historical, cohort 1) and after implementation (experimental, cohort 2). Patient outcomes were evaluated by examining changes in PU incidence, severity, timing and recurrence, as well as PU prevalence and satisfaction with life in the community., Results: Final analysis included 70 patients in cohort 1 and 73 in cohort 2. OT skin care assessment documentation (31% to 60%; p<0.001) and TSS prescriptions (31% to 60%; p=0.02) significantly increased following the implementation. The PU incidence based on patient charts (both nursing and OT assessments) did not increase significantly (26% to 36%; p=0.2). However, documented PU incidence according to OT assessments showed a substantial increase (14% to 33%; p=0.002). No effect of the PUPI was seen on immediate or long-term patient outcomes during the study period., Conclusion: PUPI was successful in changing clinical practice in PU prevention but no statistically significant improvements in PU-related patient outcomes were demonstrated. Results from this study identified facilitators and barriers to implementation and highlighted the complexity and difficulty of instituting effective preventative or therapeutic interventions for this population in an acute care setting. This information will assist with refinements of the PUPI and inform similar future initiatives.

Understanding the genetic risk of JIA, a relatively rare chronic disease, is a challenging task, but recent research in this field has shown great advances. This review summarizes the current understanding of the genetic architecture of JIA susceptibility and proposes where this work is heading in the coming years. Insights into how we might progress this relatively understudied field will be provided, highlighting how the field will move towards the ultimate goals of predicting long-term disease outcomes at onset, predicting drug response, and move towards more targeted treatment options for children with JIA.

Objectives: Genome-wide association studies have facilitated the identification of over 30 susceptibility loci for rheumatoid arthritis (RA). However, evidence for a number of potential susceptibility genes have not so far reached genome-wide significance in studies of Caucasian RA., Methods: A cohort of 4286 RA patients from across Europe and 5642 population matched controls were genotyped for 25 SNPs, then combined in a meta-analysis with previously published data., Results: Significant evidence of association was detected for nine SNPs within the European samples. When meta-analysed with previously published data, 21 SNPs were associated with RA susceptibility. Although SNPs in the PTPN2 gene were previously reported to be associated with RA in both Japanese and European populations, we show genome-wide evidence for a different SNP within this gene associated with RA susceptibility in an independent European population (rs7234029, P = 4.4×10(-9))., Conclusions: This study provides further genome-wide evidence for the association of the PTPN2 locus (encoding the T cell protein tyrosine phosphastase) with Caucasian RA susceptibility. This finding adds to the growing evidence for PTPN2 being a pan-autoimmune susceptibility gene.

Immobilization of antimicrobial peptides (AMPs) holds potential for creating surfaces with bactericidal properties. In order to successfully incorporate AMPs into desired materials, increased fundamental understanding of the relationship between AMP immobilization and the efficacy of bound peptides as antibacterial agents is required. In this study, we characterize the relationship between surface binding of the AMP and subsequent ability of the peptide to kill bacteria. Surface immobilization of the AMP chrysophsin-1 (CHY1) via a flexible linker is studied in real-time, using a quartz crystal microbalance with dissipation monitoring (QCM-D). Depending on whether the AMP is physically adsorbed to the surface or attached covalently via a zero-length or flexible cross-linker, changes could be observed in AMP orientation, surface density, flexibility, and activity against bacteria. Covalent surface binding of CHY1 led to the formation of solvated monolayers of vertically positioned peptide molecules, while the physical adsorption of CHY1 led to the deposition of rigid monolayers of horizontally positioned peptide molecules on the sensor surface. Covalently bound peptides were not removed by extensive washing and did not leach from the surface. Zero-length immobilization of the peptide decreased its ability to kill E. coli to 34% ± 7% of added bacteria, while binding via a flexible linker resulted in 82% ± 11% of bacteria being killed by the AMP.

Background: The functional polymorphism that explains the established association of the androgen receptor (AR) with androgenetic alopecia (AGA) remains unidentified, but Copy Number Variation (CNV) might be relevant. CNV involves changes in copy number of large segments of DNA, leading to the altered dosage of gene regulators or genes themselves. Two recent reports indicate regions of CNV in and around AR, and these have not been studied in relation to AGA. The aim of this preliminary case-control study was to determine if AR CNV is associated with AGA, with the hypothesis that CNV is the functional AR variant contributing to this condition., Methodology/principal Findings: Multiplex Ligation-dependent Probe Amplification was used to screen for CNV in five AR exons and a conserved, non-coding region upstream of AR in 85 men carefully selected as cases and controls for maximal phenotypic contrast. There was no evidence of CNV in AR in any of the cases or controls, and thus no evidence of significant association between AGA and AR CNV., Conclusions/significance: The results suggest this form of genomic variation at the AR locus is unlikely to predispose to AGA.

PPARgamma-activating thiazolidinediones and carboxylic acids such as farglitazar exert their anti-diabetic effects in part in PPARgamma rich adipose. Both pro- and anti-adipogenic PPARgamma ligands promote glucose and lipid lowering in animal models of diabetes. Herein, we disclose representatives of an array of 160 farglitazar analogues with atypical inverse agonism of PPARgamma in mature adipocytes.

Androgenetic alopecia, or male pattern baldness, is a complex condition with a strong heritable component. In 2001, we published the first significant evidence of a genetic association between baldness and a synonymous coding SNP (rs6152) in the androgen receptor gene, AR. Recently, this finding was replicated in three independent studies, confirming an important role for AR in the baldness phenotype. In one such replication study, it was claimed that the causative variant underlying the association was likely to be the polyglycine (GGN) repeat polymorphism, one of two apparently functional triplet repeat polymorphisms located in the exon 1 transactivating domain of the gene. Here, we extend our original association finding and present comprehensive evidence from approximately 1,200 fathers and sons drawn from 703 families of the Victorian Family Heart Study, a general population Caucasian cohort, that neither exon 1 triplet repeat polymorphism is causative in this condition. Seventy-eight percent of fathers (531/683) and 30% of sons (157/520) were affected to some degree with AGA. We utilised statistical methods appropriate for the categorical nature of the phenotype and familial structure of the cohort, and determined that whilst SNP rs6152 was strongly associated with baldness (P < 0.0001), the GGN triplet repeat was not (P = 0.13). In the absence of any other known common functional coding variants, we argue that the causative variant is likely to be in the non-coding region, and yet to be identified. The identification of functional non-coding variants surrounding AR may have significance not only for baldness, but also for the many other complex conditions that have thus far been linked to AR.

In the course of a high throughput screen to search for ligands of peroxisome proliferator activated receptor-gamma (PPARgamma), we identified GW9662 using a competition binding assay against the human ligand binding domain. GW9662 had nanomolar IC(50) versus PPARgamma and was 10- and 600-fold less potent in binding experiments using PPARalpha and PPARdelta, respectively. Pretreatment of all three PPARs with GW9662 resulted in the irreversible loss of ligand binding as assessed by scintillation proximity assay. Incubation of PPAR with GW9662 resulted in a change in the absorbance spectra of the receptors consistent with covalent modification. Mass spectrometric analysis of the PPARgamma ligand binding domain treated with GW9662 established Cys(285) as the site of covalent modification. This cysteine is conserved among all three PPARs. In cell-based reporter assays, GW9662 was a potent and selective antagonist of full-length PPARgamma. The functional activity of GW9662 as an antagonist of PPARgamma was confirmed in an assay of adipocyte differentiation. GW9662 showed essentially no effect on transcription when tested using both full-length PPARdelta and PPARalpha. Time-resolved fluorescence assays of ligand-modulated receptor heterodimerization, coactivator binding, and corepressor binding were consistent with the effects observed in the reporter gene assays. Control activators increased PPAR:RXR heterodimer formation and coactivator binding to both PPARgamma and PPARdelta. Corepressor binding was decreased. In the case of PPARalpha, GW9662 treatment did not significantly increase heterodimerization and coactivator binding or decrease corepressor binding. The experimental data indicate that GW9662 modification of each of the three PPARs results in different functional consequences. The selective and irreversible nature of GW9662 treatment, and the observation that activity is maintained in cell culture experiments, suggests that this compound may be a useful tool for elucidation of the role of PPARgamma in biological processes.

The findings of clinical pilot study (n = 9 subjects) using a new laser Doppler sensor for assessing blood flux in plantar skin tissue are described. Cutaneous blood perfusion was recorded under the first metatarsal head (right foot) in standing and walking. The sensor was located in a measurement shoe custom made for each test subject. The test group comprised diabetic patients (type II) with vascular (n = 3) or neuropathic (n = 3) complications and three controls. All subjects were Caucasian males and in the age range considered particularly at risk of diabetic foot problems (mean 61 years, 51-72 years). Following static loading for 2, 3 and 4 min the blood.flux response increased rapidly in the control (mean = 10 s) and neuropathic (mean = 18 s) groups to a well-defined, peak blood flow. For the vascular group. the blood flux response was typically a slower rise (mean = 30 s) to a poorly defined peak blood flow value. Due to movement artifact a reliable signal could only be obtained for the swing phase of gait during which blood flux was observed to increase linearly. This was interpreted as reperfusion of the tissue following unloading. The rates of reperfusion expressed in arbitrary units (of blood flux) per millisecond (au ms(-1)) were 6.1-7.9 au ms(-1) for the control, 4-6.2 au ms- for the vascular and 2.3-4.5 au ms(-1) for the neuropathic groups. The feasibility of assessing the microcirculation of the plantar skin under conditions of static and dynamic loading, with the foot in-shoe, has been demonstrated for the first time. The results suggest that abnormal responses may be obtained from asymptomatic feet of diabetic patients with vascular and/or neuropathic complications. This method of assessment could be of use in predicting the occurrence of ulceration in the diabetic foot.

Repression of gene transcription by nuclear receptors is mediated by interactions with co-repressor proteins such as SMRT and N-CoR, which in turn recruit histone deacetylases to the chromatin. Aberrant interactions between nuclear receptors and co-repressors contribute towards acute promyelocytic leukaemia and thyroid hormone resistance syndrome. The binding of co-repressors to nuclear receptors occurs in the unliganded state, and can be stabilized by antagonists. Here we report the crystal structure of a ternary complex containing the peroxisome proliferator-activated receptor-alpha ligand-binding domain bound to the antagonist GW6471 and a SMRT co-repressor motif. In this structure, the co-repressor motif adopts a three-turn alpha-helix that prevents the carboxy-terminal activation helix (AF-2) of the receptor from assuming the active conformation. Binding of the co-repressor motif is further reinforced by the antagonist, which blocks the AF-2 helix from adopting the active position. Biochemical analyses and structure-based mutagenesis indicate that this mode of co-repressor binding is highly conserved across nuclear receptors.

[structure] Two general routes to 1,4-disubstituted-2,3,4, 5-tetrahydro-1H-3-benzazepines are described. Both routes utilize an appropriately functionalized phenethylamino alcohol as the penultimate intermediate: the first route makes use of the reductive amination of a benzyl alkyl ketone with alpha-(aminomethyl)benzyl alcohol, while the second route utilizes the addition of a Grignard reagent to the oxazolidine derived from a substitued phenylacetaldehyde and alpha-(methylaminomethyl)benzyl alcohol. In all cases studied, the cis-1,4-disubstituted-2,3,4, 5-tetrahydro-1H-3-benzazepine was obtained as the major product.

The discovery that peroxisome proliferator-activated receptor (PPAR)-gamma was the molecular target of the thiazolidinedione class of antidiabetic agents suggested a key role for PPAR-gamma in the regulation of carbohydrate and lipid metabolism. Through the use of high-throughput biochemical assays, GW1929, a novel N-aryl tyrosine activator of human PPAR-gamma, was identified. Chronic oral administration of GW1929 or troglitazone to Zucker diabetic fatty (ZDF) rats resulted in dose-dependent decreases in daily glucose, free fatty acid, and triglyceride exposure compared with pretreatment values, as well as significant decreases in glycosylated hemoglobin. Whole body insulin sensitivity, as determined by the euglycemic-hyperinsulinemic clamp technique, was significantly increased in treated animals. Comparison of the magnitude of glucose lowering as a function of serum drug concentrations showed that GW1929 was 2 orders of magnitude more potent than troglitazone in vivo. These data were consistent with the relative in vitro potencies of GW1929 and troglitazone. Isolated perfused pancreas studies performed at the end of the study confirmed that pancreata from vehicle-treated rats showed no increase in insulin secretion in response to a step change in glucose from 3 to 10 mmol/l. In contrast, pancreata from animals treated with GW1929 showed a first- and second-phase insulin secretion pattern. Consistent with the functional data from the perfusion experiments, animals treated with the PPAR-gamma agonist had more normal islet architecture with preserved insulin staining compared with vehicle-treated ZDF rats. This is the first demonstration of in vivo efficacy of a novel nonthiazolidinedione identified as a high-affinity ligand for human PPAR-gamma. The increased potency of GW1929 compared with troglitazone both in vitro and in vivo may translate into improved clinical efficacy when used as monotherapy in type 2 diabetic patients. In addition, the significant improvement in daily meal tolerance may impact cardiovascular risk factor management in these patients.

The peroxisome proliferator-activated receptors (PPARs) are nuclear hormone receptors that regulate glucose and lipid homeostasis. The PPARgamma subtype plays a central role in the regulation of adipogenesis and is the molecular target for the 2, 4-thiazolidinedione class of antidiabetic drugs. Structural studies have revealed that agonist ligands activate the PPARs through direct interactions with the C-terminal region of the ligand-binding domain, which includes the activation function 2 helix. GW0072 was identified as a high-affinity PPARgamma ligand that was a weak partial agonist of PPARgamma transactivation. X-ray crystallography revealed that GW0072 occupied the ligand-binding pocket by using different epitopes than the known PPAR agonists and did not interact with the activation function 2 helix. In cell culture, GW0072 was a potent antagonist of adipocyte differentiation. These results establish an approach to the design of PPAR ligands with modified biological activities.

We previously reported the identification of (2S)-((2-benzoylphenyl)amino)-3-¿4-[2-(5-methyl-2-phenyloxazol-4-y l)e thoxy]phenyl¿propanoic acid (2) (PPARgamma pKi = 8.94, PPARgamma pEC50 = 9.47) as a potent and selective PPARgamma agonist. We now report the expanded structure-activity relationship around the phenyl alkyl ether moiety by pursuing both a classical medicinal chemistry approach and a solid-phase chemistry approach for analogue synthesis. The solution-phase strategy focused on evaluating the effects of oxazole and phenyl ring replacements of the 2-(5-methyl-2-phenyloxazol-4-yl)ethyl side chain of 2 with several replacements providing potent and selective PPARgamma agonists with improved aqueous solubility. Specifically, replacement of the phenyl ring of the phenyloxazole moiety with a 4-pyridyl group to give 2(S)-((2-benzoylphenyl)amino)-3-¿4-[2-(5-methyl-2-pyridin-4-yloxazol+ ++- 4-yl)ethoxy]phenyl¿propionic acid (16) (PPARgamma pKi = 8.85, PPARgamma pEC50 = 8.74) or a 4-methylpiperazine to give 2(S)-((2-benzoylphenyl)amino)-3-(4-¿2-[5-methyl-2-(4-methylpiperazin+ ++- 1-yl)thiazol-4-yl]ethoxy¿phenyl)propionic acid (24) (PPARgamma pKi = 8.66, PPARgamma pEC50 = 8.89) provided two potent and selective PPARgamma agonists with increased solubility in pH 7.4 phosphate buffer and simulated gastric fluid as compared to 2. The second strategy took advantage of the speed and ease of parallel solid-phase analogue synthesis to generate a more diverse set of phenyl alkyl ethers which led to the identification of a number of novel, high-affinity PPARgamma ligands (PPARgamma pKi's 6.98-8.03). The combined structure-activity data derived from the two strategies provide valuable insight on the requirements for PPARgamma binding, functional activity, selectivity, and aqueous solubility.

3-¿4-[2-(Benzoxazol-2-ylmethylamino)ethoxy]phenyl¿-(2S)-((2- benzoylph enyl)amino)propionic acid (1) and (2S)-((2-benzoylphenyl)amino)-3-¿4-[2-(5-methyl-2-phenyloxazol-4-y l)e thoxy]phenyl¿propionic acid (2) are peroxisome proliferator-activated receptor gamma (PPARgamma) agonists and have antidiabetic activity in rodent models of type 2 diabetes. As part of an effort to develop the SAR of the N-2-benzoylphenyl moiety of 1 and 2, a series of novel carboxylic acid analogues, 23-66, modified only in the N-2-benzoylphenyl moiety were synthesized from L-tyrosine and evaluated as PPARgamma agonists. In general, only modest changes in the N-2-benzoylphenyl moiety of 1 and 2 are tolerated. More specifically, the best changes involve bioisosteric replacement of one of the two phenyl rings of this moiety. Addition of substituents to this moiety generally produced compounds that are less active in the cell-based functional assays of PPARgamma activity although binding affinity to PPARgamma may be maintained. A particularly promising set of analogues is the anthranilic acid esters 63-66 in which the phenyl ring in the 2-benzoyl group of 1 and 2 has been replaced by an alkoxy group. In particular, (S)-2-(1-carboxy-2-¿4-[2-(5-methyl-2-phenyloxazol-4-yl)ethoxy]phen yl¿ ethylamino)benzoic acid methyl ester (63) has a pKi of 8.43 in the binding assay using human PPARgamma ligand binding domain and a pEC50 of 9.21 in the in vitro murine lipogenesis functional assay of PPARgamma activity. Finally, 63 was found to normalize glycemia when dosed at 3 mg/kg bid po in the Zucker diabetic fatty rat model of type 2 diabetes.

We have identified a novel series of antidiabetic N-(2-benzoylphenyl)-L-tyrosine derivatives which are potent, selective PPARgamma agonists. Through the use of in vitro PPARgamma binding and functional assays (2S)-3-(4-(benzyloxy)phenyl)-2-((1-methyl-3-oxo-3-phenylpropenyl)+ ++amin o)propionic acid (2) was identified as a structurally novel PPARgamma agonist. Structure-activity relationships identified the 2-aminobenzophenone moiety as a suitable isostere for the chemically labile enaminone moiety in compound 2, affording 2-((2-benzoylphenyl)amino)-3-(4-(benzyloxy)phenyl)propionic acid (9). Replacement of the benzyl group in 9 with substituents known to confer in vivo potency in the thiazolidinedione (TZD) class of antidiabetic agents provided a dramatic increase in the in vitro functional potency and affinity at PPARgamma, affording a series of potent and selective PPARgamma agonists exemplified by (2S)-((2-benzoylphenyl)amino)-3-¿4-[2-(methylpyridin-2-ylamino+ ++)ethoxy ]phenyl¿propionic acid (18), 3-¿4-[2-(benzoxazol-2-ylmethylamino)ethoxy]phenyl¿-(2S)-((2- benzoylph enyl)amino)propanoic acid (19), and (2S)-((2-benzoylphenyl)amino)-3-¿4-[2-(5-methyl-2-phenyloxazol-4-y l)e thoxy]phenyl¿propanoic acid (20). Compounds 18 and 20 show potent antihyperglycemic and antihyperlipidemic activity when given orally in two rodent models of type 2 diabetes. In addition, these analogues are readily prepared in chiral nonracemic fashion from L-tyrosine and do not show a propensity to undergo racemization in vitro. The increased potency of these PPARgamma agonists relative to troglitazone may translate into superior clinical efficacy for the treatment of type 2 diabetes.

The peroxisome proliferator-activated receptor-gamma (PPAR-gamma) is a ligand-dependent transcription factor that is important in adipocyte differentiation and glucose homeostasis and which depends on interactions with co-activators, including steroid receptor co-activating factor-1 (SRC-1). Here we present the X-ray crystal structure of the human apo-PPAR-gamma ligand-binding domain (LBD), at 2.2 A resolution; this structure reveals a large binding pocket, which may explain the diversity of ligands for PPAR-gamma. We also describe the ternary complex containing the PPAR-gamma LBD, the antidiabetic ligand rosiglitazone (BRL49653), and 88 amino acids of human SRC-1 at 2.3 A resolution. Glutamate and lysine residues that are highly conserved in LBDs of nuclear receptors form a 'charge clamp' that contacts backbone atoms of the LXXLL helices of SRC-1. These results, together with the observation that two consecutive LXXLL motifs of SRC-1 make identical contacts with both subunits of a PPAR-gamma homodimer, suggest a general mechanism for the assembly of nuclear receptors with co-activators.

This investigation sought to determine if cycling power could be accurately modeled. A mathematical model of cycling power was derived, and values for each model parameter were determined. A bicycle-mounted power measurement system was validated by comparison with a laboratory ergometer. Power was measured during road cycling, and the measured values were compared with the values predicted by the model. The measured values for power were highly correlated (R 2 = .97) with, and were not different than, the modeled values. The standard error between the modeled and measured power (2.7 W) was very small. The model was also used to estimate the effects of changes in several model parameters on cycling velocity. Over the range of parameter values evaluated, velocity varied linearly (R 2 > .99). The results demonstrated that cycling power can be accurately predicted by a mathematical model.

The biological activity of amylin is reported to vary widely depending on the source and purity of the material. Three commercial samples of rat amylin were compared for structural differences. The samples were nearly identical using most of the available analytical measures--amino acid analysis, HPLC retention, even Edman sequencing data. When the samples were compared by ion spray ionization mass spectrometry, the molecular mass of one sample was 200 daltons higher than anticipated. Careful analysis of the sample, including atomic emission spectrometry, revealed that a mercury atom was associated with the polypeptide. The mercury presumably resulted from a deprotection step in the synthesis, involving the removal of an acetamidomethyl group from cysteine.