Duchenne muscular dystrophy (DMD) is a fatal muscle‐wasting disease arising from mutations in the dystrophin gene. Upregulation of utrophin to compensate for the missing dystrophin offers a potential ...therapy independent of patient genotype. The first‐in‐class utrophin modulator ezutromid/SMT C1100 was developed from a phenotypic screen through to a Phase 2 clinical trial. Promising efficacy and evidence of target engagement was observed in DMD patients after 24 weeks of treatment, however trial endpoints were not met after 48 weeks. The objective of this study was to understand the mechanism of action of ezutromid which could explain the lack of sustained efficacy and help development of new generations of utrophin modulators. Using chemical proteomics and phenotypic profiling we show that the aryl hydrocarbon receptor (AhR) is a target of ezutromid. Several lines of evidence demonstrate that ezutromid binds AhR with an apparent KD of 50 nm and behaves as an AhR antagonist. Furthermore, other reported AhR antagonists also upregulate utrophin, showing that this pathway, which is currently being explored in other clinical applications including oncology and rheumatoid arthritis, could also be exploited in future DMD therapies.
Chemische Proteomik und phänotypisches Profiling zeigen, dass der Aryl‐Kohlenwasserstoff‐Rezeptor (AhR) ein Target von Ezutromid ist, dem ersten Utrophin‐Modulator zur Behandlung von Duchenne‐Muskeldystrophie. Ezutromid bindet an AhR mit einem effektiven KD von 50 nm und fungiert als AhR‐Antagonist. Andere AhR‐Antagonisten bewirken ebenfalls die Hochregulierung von Utrophin, was zeigt, dass dieser Weg auch in zukünftigen DMD‐Therapien genutzt werden könnte.
Until recently, it had been thought that trypanosomes lack glutathione peroxidase activity. Here we report the subcellular localization and biochemical properties of a second glutathione-dependent ...peroxidase from Trypanosoma cruzi (TcGPXII). TcGPXII is a single-copy gene which encodes a 16 kDa protein that appears to be specifically dependent on glutathione as the source of reducing equivalents. Recombinant TcGPXII was purified and shown to have peroxidase activity towards a narrow substrate range, restricted to hydroperoxides of fatty acids and phospholipids. Analysis of the pathway revealed that TcGPXII activity could be readily saturated by glutathione and that the peroxidase functioned by a Ping Pong mechanism. Enzyme reduction was shown to be the rate-limiting step in this pathway. Using immunofluorescence, TcGPXII was shown to co-localize with a homologue of immunoglobulin heavy-chain binding protein (BiP), a protein restricted to the endoplasmic reticulum and Golgi. As the smooth endoplasmic reticulum is the site of phospholipid and fatty acid biosynthesis, this suggests that TcGPXII may play a specific role in the T. cruzi oxidative defence system by protecting newly synthesized lipids from peroxidation.
Long-term exposure to primary traffic pollutants may be harmful for health but few studies have investigated effects on mortality. We examined associations for six primary traffic pollutants with ...all-cause and cause-specific mortality in 2003–2010 at small-area level using linear and piecewise linear Poisson regression models. In linear models most pollutants showed negative or null association with all-cause, cardiovascular or respiratory mortality. In the piecewise models we observed positive associations in the lowest exposure range (e.g. relative risk (RR) for all-cause mortality 1.07 (95% credible interval (CI) = 1.00–1.15) per 0.15 μg/m3 increase in exhaust related primary particulate matter ≤2.5 μm (PM2.5)) whereas associations in the highest exposure range were negative (corresponding RR 0.93, 95% CI: 0.91–0.96). Overall, there was only weak evidence of positive associations with mortality. That we found the strongest positive associations in the lowest exposure group may reflect residual confounding by unmeasured confounders that varies by exposure group.
Display omitted
•Evidence of association between primary traffic pollutants and mortality is scarce.•We examined this in a large city using most recent small-area statistical methods.•Overall, there was only weak evidence of positive associations with mortality.
Overall, there was only weak evidence of positive associations between long-term exposure to primary traffic pollutants and mortality for all, cardiovascular or respiratory causes.
Reference and type strains of well-known bacteria have been a cornerstone of microbiology research for decades. The sharing of well-characterized isolates among laboratories has run in parallel with ...research efforts and enhanced the reproducibility of experiments, leading to a wealth of knowledge about trait variation in different species and the underlying genetics. Campylobacter jejuni strain NCTC 11168, deposited at the National Collection of Type Cultures in 1977, has been adopted widely as a reference strain by researchers worldwide and was the first Campylobacter for which the complete genome was published (in 2000). In this study, we collected 23 C. jejuni NCTC 11168 reference isolates from laboratories across the UK and compared variation in simple laboratory phenotypes with genetic variation in sequenced genomes. Putatively identical isolates, identified previously to have aberrant phenotypes, varied by up to 281 SNPs (in 15 genes) compared to the most recent reference strain. Isolates also display considerable phenotype variation in motility, morphology, growth at 37 °C, invasion of chicken and human cell lines, and susceptibility to ampicillin. This study provides evidence of ongoing evolutionary change among C. jejuni isolates as they are cultured in different laboratories and highlights the need for careful consideration of genetic variation within laboratory reference strains. This article contains data hosted by Microreact.
Transforming growth factor-beta (TGF-beta) has been implicated in the pathogenesis of a number of kidney diseases characterized by glomerulosclerosis and tubulointerstitial fibrosis. TGF-beta is ...secreted in a latent form requiring extracellular modification to become biologically active. TGF-beta inducible gene-h3 (beta ig-h3) is a recently identified TGF-beta-induced gene product. The present study sought to examine beta ig-h3 expression in normal and diabetic rats.
Beta ig-h3, TGF-beta1 and alpha1 (IV) collagen gene expression were assessed by Northern blot analysis and in situ hybridization in 20 Sprague Dawley rats, randomly assigned to receive streptozotocin (diabetic, N = 11) or citrate buffer alone (control, N = 9) and sacrificed eight months later. The effect of exogenous TGF-beta1 on beta ig-h3 expression was also assessed in cultured proximal tubular cells.
In situ hybridization localized beta ig-h3 gene expression to the juxtaglomerular apparatus and the pars recta (S3 segment) of proximal tubules in both control and diabetic animals. Kidney TGF-beta 1, beta ig-h3 and alpha1 (IV) collagen mRNA from diabetic rats were increased two- to threefold compared with controls (P < 0.01). There was a significant correlation between TGF-beta1 and beta ig-h3 gene expression in kidneys from diabetic rats (r = 0.73, P = 0.01). In addition, beta ig-h3 mRNA increased in response to exogenous TGF-beta1 in a dose-dependent fashion in cultured proximal tubular cells.
These findings support the hypothesis that biologically active TGF-beta plays a pathogenetic role in diabetic kidney disease and suggest that beta ig-h3 may be a useful index of TGF-beta1 bioactivity in the kidney.
There is accumulating evidence that local renin-angiotensin systems (RASs) influence cell growth and organ function in a variety of tissues including the ovary. The first aim of this study was to ...characterise the cellular location of RAS components in the rat ovary. This was facilitated by the use of the hypertensive transgenic (mRen-2)27 rat which overexpresses renin and angiotensin in extra-renal tissues. Comparisons were made with normal Sprague-Dawley (SD) rats. The second aim was to determine if the upregulated RAS of the transgenic (mRen-2)27 rat and infusion of angiotensin II (ANG II) in SD rats influences follicle number and litter size. Gene expression, immunohistochemical and autoradiographic techniques were used to identify a discrete RAS including ANG II receptors in the ovarian stroma, follicles (particularly atretic) and to a lesser extent corpora lutea. The RAS at these sites was most abundant in homozygous (HMZ) followed by heterozygous (HTZ) (mRen-2)27 rats and then SD rats. Large antral and preovulatory follicles and litter size were reduced in (mRen-2)27 rats. In HMZ (mRen-2)27 rats and SD rats infused with ANG II, angiotensin 1a (AT(1a)) receptor mRNA in the ovarian stroma was lower than control SD rats and was associated with a reduction in large antral and preovulatory follicles. These findings indicate that upregulation of the ovarian RAS in the rat influences follicular development and, potentially, reproductive capacity.
Background. We have previously reported that severe glomerulosclerosis progressively develops in the streptozotocin (STZ) diabetic transgenic (mRen‐2)27 rat. In this diabetic model, monotherapy with ...either angiotensin converting enzyme inhibition (ACEI) or angiotensin type 1 (AT1) receptor blockade is largely renoprotective. The objective of the present study was to determine if a combination therapy at lower doses than monotherapy would confer greater renoprotection. Methods. At 6 weeks of age, non‐diabetic control and STZ diabetic female heterozygous Ren‐2 rats were randomized to receive vehicle, the AT1 receptor blocker valsartan (V, 20 mg/kg/day), the ACEI perindopril (P, 6 mg/kg/day), or a combination of low‐dose V+P (V, 3 mg/kg/day plus P, 0.5 mg/kg/day) for 12 weeks. Results. Systolic blood pressure was lowered with all treatments, but the greatest reductions were observed with V monotherapy and combination V+P therapy. All treatments reduced albuminuria, the decline in glomerular filtration rate, and cortical collagen staining, to the same extent. The glomerulosclerotic index was increased with diabetes and reduced with V and P monotherapy. However, the low‐dose combination therapy was more effective than single therapy and reduced severe glomerulosclerosis to levels observed in non‐diabetic controls. Conclusions. Monotherapy with either V or P reduced blood pressure and retarded the decline in renal function and glomerulosclerosis in the diabetic Ren‐2 rat. Combination therapy has the additional benefit of requiring only low doses of AT1 receptor blockade and ACEI to achieve superior renoprotective effects in this diabetic nephropathy model.
Transvaginal ultrasound ovarian volume was moderately correlated with
volume measured by pathology report and not statistically significantly
correlated with sex-steroid hormone levels in ...post-menopausal women.
Brief recurrent apneas in preterm infants and adults can precipitate rapid and severe arterial O(2) desaturation for reasons that remain unclear.
We tested a mathematically derived hypothesis that ...when breathing terminates apnea, mixed-venous hypoxemia continues into the subsequent apnea; as a result, there is a surge in pulmonary O(2) uptake that rapidly depletes the finite alveolar O(2) store, thereby accelerating arterial O(2) desaturation.
Recurrent apneas were simulated in an experimental lamb model. Pulmonary O(2) uptake was calculated from continuously measured arterial and mixed-venous O(2) saturation and cardiac output.
Direct measurements revealed that asynchrony in the desaturation and resaturation of arterial and venous blood gave rise to dips and surges in O(2) uptake. After desaturation to 50%, a typical nadir in preterm infants, O(2) uptake surged to a peak of 176.9 ± 7.8% of metabolic rate. During subsequent apneas, desaturation rate was increased two- to threefold greater than during isolated apneas, in direct proportion to the magnitude of the surge in O(2) uptake (P < 0.001; R(2) = 0.897). Application of our mathematical model to a published recording of cyclic apneas in a preterm infant precisely reproduced the accelerated desaturation rates of up to 15% · s(-1) observed clinically.
Rapid depletion of alveolar O(2) stores by surges in O(2) uptake almost completely explains the acceleration of desaturation that occurs during recurrent apnea. This powerful mechanism is likely to explain the severity of intermittent hypoxemia that is associated with neurocognitive and cardiovascular morbidities in preterm infants and adults.
The transgenic (mRen-2)27 rat (TGR) is a high tissue renin, high angiotensin (Ang) II model of hypertension. When administered streptozotocin (STZ), TGRs develop a rapidly progressive diabetic ...nephropathy with renal failure over 12 weeks. Bradykinin (BK) and Ang II are potent vasoactive peptides that may participate in the vascular and metabolic abnormalities of diabetes.
TGR and Sprague-Dawley (SD) rats were administered STZ (diabetic) or citrate buffer (nondiabetic) at six weeks of age. Diabetic rats received daily ultralente insulin to maintain moderate hyperglycemia (∼18 m M). Rats were sacrificed four- and eight-weeks post-STZ or vehicle.
Diabetes did not modify the blood pressure of either SD rats or TGRs. Diabetes increased levels of BK-(1-9) and its metabolite BK-(1-7) in kidney, aorta, and heart of both SD rats and TGRs. Diabetes did not influence Ang II levels in plasma, kidney, aorta, heart, or adrenal gland of SD rats, but reduced to normal the elevated Ang II levels in plasma, kidney, aorta, and adrenal gland of TGRs.
STZ-induced diabetes was associated with elevated tissue levels of BK-(1-9) and “normal” circulating and tissue levels of Ang II. The increased BK-(1-9) levels were consistent with the participation of this peptide in the vascular and metabolic abnormalities of diabetes. However, the rapidly progressive nephropathy of diabetic TGRs was not associated with BK-(1-9) and Ang II levels in target organs that differed from those of diabetic SD rats.
PDF
