Showing papers by "Mark E. Cooper published in 2004"

Albuminuria, a Therapeutic Target for Cardiovascular Protection in Type 2 Diabetic Patients With Nephropathy

Abstract: Background— Albuminuria is an established risk marker for both cardiovascular and renal outcomes. Albuminuria can be reduced with drugs that block the renin-angiotensin system (RAS). We questioned whether the short-term drug-induced change in albuminuria would predict the long-term cardioprotective efficacy of RAS intervention. Methods and Results— We analyzed data from Reduction in Endpoints in Non-insulin dependent diabetes mellitus with the Angiotensin II Antagonist Losartan (RENAAL), a double-blind, randomized trial in 1513 type 2 diabetic patients with nephropathy, focusing on the relationship between the prespecified cardiovascular end point (composite) or hospitalization for heart failure and baseline or reduction in albuminuria. Patients with high baseline albuminuria (≥3 g/g creatinine) had a 1.92-fold (95% CI, 1.54 to 2.38) higher risk for the cardiovascular end point and a 2.70-fold (95% CI, 1.94 to 3.75) higher risk for heart failure compared with patients with low albuminuria (<1.5 g/g). Amon...

Genome-wide survey of protein kinases required for cell cycle progression.

Abstract: Cycles of protein phosphorylation are fundamental in regulating the progression of the eukaryotic cell through its division cycle. Here we test the complement of Drosophila protein kinases (kinome) for cell cycle functions after gene silencing by RNA-mediated interference. We observed cell cycle dysfunction upon downregulation of 80 out of 228 protein kinases, including most kinases that are known to regulate the division cycle. We find new enzymes with cell cycle functions; some of these have family members already known to phosphorylate microtubules, actin or their associated proteins. Additionally, depletion of several signalling kinases leads to specific mitotic aberrations, suggesting novel roles for familiar enzymes. The survey reveals the inter-digitation of systems that monitor cellular physiology, cell size, cellular stress and signalling processes with the basic cell cycle regulatory machinery.

Advanced glycation end product-interventions reduce diabetes-accelerated atherosclerosis

Abstract: Advanced glycation end product (AGE) formation may contribute to the progression of atherosclerosis, particularly in diabetes. The present study explored atherosclerosis in streptozotocin-induced diabetic apolipoprotein E-deficient (apoE-/-) mice that were randomized (n = 20) to receive for 20 weeks no treatment, the AGE cross-link breaker ALT-711, or the inhibitor of AGE formation aminoguanidine (AG). A sixfold increase in plaque area with diabetes was attenuated by 30% with ALT-711 and by 40% in AG-treated mice. Regional distribution of plaque demonstrated no reduction in plaque area or complexity within the aortic arch with treatment, in contrast to the thoracic and abdominal aortas, where significant attenuation was seen. Diabetes-associated accumulation of AGEs in aortas and plasma and decreases in skin collagen solubility were ameliorated by both treatments, in addition to reductions in the vascular receptor for AGE. Collagen-associated reductions in the AGEs carboxymethyllysine and carboxyethyllysine were identified with both treatments. Diabetes was also accompanied by aortic accumulation of total collagen, specifically collagens I, III, and IV, as well as increases in the profibrotic cytokines transforming growth factor-beta and connective tissue growth factor and in cellular alpha-smooth muscle actin. Attenuation of these changes was seen in both treated diabetic groups. ALT-711 and AG demonstrated the ability to reduce vascular AGE accumulation in addition to attenuating atherosclerosis in these diabetic mice.

ACE2, a new regulator of the renin-angiotensin system.

Abstract: Angiotensin-converting enzyme (ACE) is a zinc metalloproteinase and a key regulator of the renin-angiotensin system (RAS) ACE2 is a newly described enzyme identified in rodents and humans with a more restricted distribution than ACE, and is found mainly in heart and kidney ACE2 cleaves a single residue from angiotensin I (Ang I) to generate Ang 1-9, and degrades Ang II, the main effector of the RAS, to the vasodilator Ang 1-7 The importance of ACE2 in normal physiology and pathophysiological states is largely unknown ACE2 might act in a counter-regulatory manner to ACE, modulating the balance between vasoconstrictors and vasodilators within the heart and kidney, and playing a significant role in regulating cardiovascular and renal function

Improved islet morphology after blockade of the renin-angiotensin system in the ZDF rat

Abstract: The renin-angiotensin system (RAS) has an important role in the endocrine pancreas. Although angiotensin II has significant effects on cell proliferation and apoptosis, the contribution of the RAS to changes in islet structure and function associated with type 2 diabetes is yet to be defined. This study examined the specific effects of RAS blockade on islet structure and function in diabetes. Thirty-six male Zucker diabetic fatty (ZDF) rats, 10 weeks of age, were randomized to receive the angiotensin-converting enzyme inhibitor perindopril (8 mg/l in drinking water; n = 12), irbesartan (15 mg/kg via gavage; n = 12), or no treatment (n = 12) for 10 weeks. Results were compared with lean littermates (ZL) (n = 12) studied concurrently. ZDF rats had increased intra-islet expression of components of the RAS correlating with increased intraislet fibrosis, apoptosis, and oxidative stress. Disordered islet architecture, seen in ZDF rats, was attenuated after treatment with perindopril or irbesartan. Islet fibrogenesis was also diminished, as measured by picrosirius staining and expression of collagens I and IV. Gene expression of transforming growth factor-beta1 was increased in the ZDF pancreas (ZL, 1.0 +/- 0.1; ZDF, 2.0 +/- 0.3; P < 0.05) and reduced after blockade of the RAS (ZDF + P, 1.3 +/- 0.2; ZDF + I, 1.5 +/- 0.1; vs. ZDF, both P < 0.05). Improvements in structural parameters were also associated with functional improvements in first-phase insulin secretion. These findings provide a possible mechanism for the reduced incidence of new-onset diabetes that has been observed in clinical trials of RAS blockade.

Advanced Glycation End Products Induce Tubular Epithelial-Myofibroblast Transition through the RAGE-ERK1/2 MAP Kinase Signaling Pathway

Abstract: Advanced glycation end products (AGEs) have been shown to play a role in tubular epithelial-myofibroblast transdifferentiation (TEMT) in diabetic nephropathy, but the intracellular signaling pathway remains unknown. We report here that AGEs signal through the receptor for AGEs (RAGE) to induce TEMT, as determined by de novo expression of a mesenchymal marker (α-smooth muscle actin, α-SMA) and loss of epithelial marker (E-cadherin), directly through the MEK1-ERK1/2 MAP kinase pathway, which is TGF-β independent. This is supported by the following findings: AGEs induced de novo α-SMA mRNA expression as early as 2 hours followed by a loss of E-cadherin before TGF-β mRNA expression at 24 hours and occurred in the absence of TGF-β and AGE-induced activation of ERK1/2 MAP kinase at 15 minutes and TEMT at 24 hours were completely blocked by a neutralizing RAGE antibody, a soluble RAGE receptor, an ERK1/2 MAP kinase inhibitor (PD98059), and DN-MEK1, but not by a neutralizing TGF-β antibody. Thus, this study demonstrates that AGEs activate the RAGE-ERK1/2 MAP kinase pathway to mediate the early TEMT process. The findings from this study suggest that targeting the RAGE or the ERK MAP kinase pathway may provide new therapeutic strategies for diabetic nephropathy and shed new light on the pathogenesis of diabetic nephropathy.

Advances in membrane receptor screening and analysis.

Abstract: During the last decade there has been significant progress in the development of analytical techniques for the screening of ligand binding to membranes and membrane receptors This review focuses on developments using label-free assays that facilitate ligand-membrane-receptor screening without the need for chemical-, biological- or radiological-labelled reagents These assays include acoustic, optical surface plasmon resonance biosensing, sedimentation (analytical ultracentrifugation), chromatographic assays, isothermal titration calorimetry and differential scanning calorimetry The merits and applications of cell-based screening systems and of different model membrane systems, including planar supported lipid layers, bead-supported membranes and lipid micro-arrays, are discussed Recent advances involving more established techniques including intrinsic fluorescence, FRET spectroscopy, scintillation proximity assays and automated patch clamping are presented along with applications to peripheral membrane proteins, ion channels and G protein-coupled receptors Novel high-throughput assays for determination of drug- and protein-partitioning in membranes are also highlighted To aid the experimenter, a brief synopsis of the techniques commonly employed to purify and reconstitute membranes and membrane receptors is included

Irbesartan but Not Amlodipine Suppresses Diabetes-Associated Atherosclerosis

Abstract: Background—It remains controversial whether specific blockade of the renin-angiotensin system confers superior antiatherosclerotic effects over other antihypertensive agents in diabetes. Therefore, the aim of this study was to compare equihypotensive doses of the angiotensin II subtype 1 (AT1) receptor blocker irbesartan with the calcium antagonist amlodipine on diabetes-induced plaque formation in the apolipoprotein E (apoE)–null mouse and to explore molecular and cellular mechanisms linked to vascular protection. Methods and Results—Diabetes was induced by injection of streptozotocin in 6-week-old apoE-null mice. Diabetic animals were randomized to no treatment, irbesartan, or amlodipine for 20 weeks. Diabetes was associated with an increase in plaque area and complexity in the aorta in association with a significant increase in aortic AT 1 receptor expression, cellular proliferation, collagen content, macrophage- and -smooth muscle actin–positive cell infiltration, as well as an increased expression of platelet-derived growth factor-B (PDGF-B), monocyte chemoattractant protein-1 (MCP-1), and vascular cell adhesion molecule-1 (VCAM-1). Irbesartan but not amlodipine treatment attenuated the development of atherosclerosis, collagen content, cellular proliferation, and macrophage infiltration as well as diabetes-induced AT1 receptor, PDGF-B, MCP-1, and VCAM-1 overexpression in the aorta despite similar blood pressure reductions by both treatments. Conclusions—Diabetes-associated atherosclerosis is ameliorated by AT1 receptor blockade but not by calcium channel antagonism, providing further evidence for the vascular renin-angiotensin system playing a pivotal role in the development and acceleration of atherosclerosis in diabetes. (Circulation. 2004;109:1536-1542.)

Mapping As You Go

Abstract: The advent of high throughput molecular technologies has led to an expectation that breeding programs will use marker-trait associations to conduct marker-assisted selection (MAS) for traits. Many challenges exist with this molecular breeding approach for so-called complex traits. A major restriction to date has been the limited ability to detect and quantify marker-trait relationships, especially for traits influenced by the effects of gene-by-gene and gene-by-environment interactions. A further complication has been that estimates of quantitative trait loci (QTL) effects are biased by the necessity of working with a limited set of genotypes in a limited set of environments, and hence the applications of these estimates are not as effective as expected when used more broadly within a breeding program. The approach considered in this paper, referred to as the Mapping As You Go (MAYG) approach, continually revises estimates of QTL allele effects by remapping new elite germplasm generated over cycles of selection, thus ensuring that QTL estimates remain relevant to the current set of germplasm in the breeding program. Mapping As You Go is a mapping-MAS strategy that explicitly recognizes that alleles of QTL for complex traits can have different values as the current breeding material changes with time. Simulation was used to investigate the effectiveness of the MAYG approach applied to complex traits. The results indicated that greater levels of response were achieved and these responses were less variable when estimates were revised frequently compared with situations where estimates were revised infrequently or not at all.

Interactions between Angiotensin II and NF-κB–Dependent Pathways in Modulating Macrophage Infiltration in Experimental Diabetic Nephropathy

Abstract: NF-kappaB-dependent pathways play an important role in macrophage infiltration and kidney injury. NF-kappaB is regulated by angiotensin II (AII). However, the role of this pathway in diabetic nephropathy has not been clearly delineated. First, the activation of NF-kappaB, monocyte chemoattractant protein-1 (MCP-1), and macrophage infiltration in the diabetic kidney were explored, in a temporal manner. The active subunit of NF-kappaB, p65, was elevated in the diabetic animals in association with increased MCP-1 gene expression and macrophage infiltration. Second, the effects of treatment for 4 wk with the AII type 1 receptor antagonist valsartan, the AII type 2 receptor antagonist PD123319, or pyrrolidine dithiocarbamate, an inhibitor of NF-kappaB and on these parameters were assessed. These treatments were associated with a reduction in p65 activation, MCP-1 gene expression, and macrophage infiltration. These findings demonstrate a role for activation of NF-kappaB, in particular the p65 subunit, in the pathogenesis of early renal macrophage infiltration in experimental diabetes. In the context of the known proinflammatory effects of AII, it is postulated that the renoprotection conferred by angiotensin II receptor antagonism is at least partly related to the inhibition of NF-kappaB-dependent pathways.

Attenuation of Extracellular Matrix Accumulation in Diabetic Nephropathy by the Advanced Glycation End Product Cross-Link Breaker ALT-711 via a Protein Kinase C-α−Dependent Pathway

Abstract: This study investigated the role of advanced glycation end products (AGEs) in mediating protein kinase C (PKC) isoform expression in diabetic nephropathy. In vitro, vascular smooth muscle cells incubated in a high-glucose (25-mmol/l) medium demonstrated translocation and increased expression of PKC-α as compared with those from a low-glucose (5-mmol/l) environment. Coincubation with the cross-link breaker ALT-711 and, to a lesser extent, with aminoguanidine, an inhibitor of AGE formation, attenuated the increased expression and translocation of PKC-α. Streptozotocin-induced diabetic rats were randomized to no treatment, treatment with ALT-711, or treatment with aminoguanidine. Diabetes induced increases in PKC-α as well as in the -βI, -βII, and -e isoforms. Treatment with ALT-711 and aminoguanidine, which both attenuate renal AGE accumulation, abrogated these increases in PKC expression. However, translocation of phosphorylated PKC-α from the cytoplasm to the membrane was reduced only by ALT-711. ALT-711 treatment attenuated expression of vascular endothelial growth factor and the extracellular matrix proteins, fibronectin and laminin, in association with reduced albuminuria. Aminoguanidine had no effect on VEGF expression, although some reduction of fibronectin and laminin was observed. These findings implicate AGEs as important stimuli for the activation of PKC, particularly PKC-α, in the diabetic kidney, which can be directly inhibited by ALT-711.

Imatinib Attenuates Diabetes-Associated Atherosclerosis

Abstract: Objective— Diabetes is associated with accelerated atherosclerosis, the major factor contributing to increased mortality and morbidity in the diabetic population. The molecular mechanisms by which diabetes promotes atherosclerosis are not fully understood. Platelet-derived growth factor has been shown to play a major role in the pathology of vascular diseases, but whether it plays a role in atherosclerosis associated with diabetes remains unknown. The aims of this study were to assess whether platelet-derived growth factor–dependent pathways are involved in the development of diabetes-induced atherosclerosis and to determine the effects of platelet-derived growth factor receptor antagonism on this disorder. Methods and Results— Diabetes was induced by injection of streptozotocin in 6-week-old apolipoprotein E knockout mice. Diabetic animals received treatment with a tyrosine kinase inhibitor that inhibits platelet-derived growth factor action, imatinib (STI-571, 10 mg/kg per day), or no treatment for 20 w...

Accelerated Nephropathy in Diabetic Apolipoprotein E-Knockout Mouse: Role of Advanced Glycation End Products

Abstract: . Hyperlipidemia not only may be relevant to cardiovascular disease in diabetes but may also play a role in the development and progression of diabetic nephropathy. Furthermore, there is increasing evidence that advanced glycation end products (AGE) play an important role in diabetic renal disease. The objectives of this study were first to characterize renal injury in diabetic apolipoprotein E knockout (apo E-KO) mice and second to explore the role of AGE in the development and progression of renal disease in this model. Diabetes was induced by injection of streptozotocin in 6-wk-old apo E-KO mice. Diabetic animals received no treatment or treatment with the inhibitor of AGE formation aminoguanidine (1 g/kg per d) or the cross-link breaker [4,5-dimethyl-3-(2-oxo2-phenylethyl)-thiazolium chloride] ALT-711, which cleaves preformed AGE (20 mg/kg per d) for 20 wk. Nondiabetic apo E-KO mice as well as nondiabetic and diabetic C57BL/6 mice served as controls. Compared with nondiabetic apo E-KO mice, induction of diabetes in apo E-KO mice resulted in accelerated renal injury characterized by albuminuria and glomerular and tubulointerstitial injury. These abnormalities were associated with increased expression of collagen type I and type IV and transforming growth factor-β1 (TGF-β1), increased α-smooth muscle actin immunostaining and macrophage infiltration, and increased serum and renal AGE. The two treatments, which attenuated renal AGE accumulation in a disparate manner, were associated with less albuminuria, structural injury, macrophage infiltration, TGF-β1, and collagen expression. The accelerated renal injury that was observed in diabetic apo E-KO mice was attenuated by approaches that inhibit renal AGE accumulation.

Heparanase Is Involved in the Pathogenesis of Proteinuria as a Result of Glomerulonephritis

Abstract: The beta-D-endoglycosidase heparanase has been proposed to be important in the pathogenesis of proteinuria by selectively degrading the negatively charged side chains of heparan sulfate proteoglycans within the glomerular basement membrane. A loss of negatively charged heparan sulfate proteoglycans may result in alteration of the permselective properties of the glomerular basement membrane, loss of glomerular epithelial and endothelial cell anchor points, and liberation of growth factors. In this study, therefore, the role of heparanase in passive Heymann nephritis (PHN) was examined. Normal glomeruli showed low-level heparanase expression as determined by immunohistochemistry and Western blot analysis. Days 5, 14, and 28 of PHN were associated with an increase in endothelial and glomerular epithelial cell heparanase. Reverse transcription-PCR confirmed a significant increase in mRNA at day 21 of disease (P < 0.0004). Furthermore, urinary and glomerular heparanase activities were significantly increased at days 5 and 21 of disease, respectively. Western blot analysis of isolated glomeruli separated into membrane- and cytosol-enriched protein fractions showed that the active 58-kD heparanase species was increased but restricted to the cytosol of diseased glomeruli at day 21. The inactive 65-kD precursor, however, was found in membrane and cytosol-diseased fractions, suggesting cell membrane processing. Complement depletion prevented glomerular heparanase expression; in addition, administration of a polyclonal anti-heparanase antibody significantly reduced urinary protein excretion at day 5 of disease to 62 +/- 11 mg/d compared with 203 +/- 43 and 159 +/- 18 mg/d in the normal rabbit serum- and normal saline-treated experimental groups, respectively (P < 0.002). Proteinuria was reduced in the absence of any altered glomerular C5b-9 activity, sheep IgG deposition, or rat anti-sheep antibody titers. These data suggest that heparanase contributes to the pathogenesis of proteinuria in PHN.

Retinal Expression of Vascular Endothelial Growth Factor Is Mediated by Angiotensin Type 1 and Type 2 Receptors

Abstract: Angiotensin II is a known stimulus for the expression of vascular endothelial growth factor (VEGF). This action of angiotensin II is mediated by the angiotensin type 1 (AT1) receptor. However, the role of the angiotensin type 2 (AT2) receptor subtype in inducing VEGF expression has been controversial. The aim of the present study was to assess the effects of AT2 receptor blockade on VEGF expression in the retina, initially in experimental diabetic rats induced by injection of streptozotocin. The AT1 receptor antagonist, valsartan, or the AT2 receptor antagonists, PD123319, were administered to diabetic rats for 4 weeks. Increased gene and protein expressions of VEGF, as assessed by real-time reverse transcription-polymerase chain reaction and immunostaining, respectively, were observed in the retina in diabetic rats. Treatment with either valsartan or PD123319 attenuated retinal VEGF expression. To further explore the link between angiotensin receptor subtypes and VEGF expression, valsartan, or PD123319 were administered to rats that were infused with angiotensin II for 2 weeks. VEGF expression was also increased in the retina from angiotensin II infused rats, and this was attenuated by valsartan and PD123319. These findings suggest that VEGF expression is modulated by AT1 and AT2 receptors, thereby implicating angiotensin II receptor subtypes in retinal diseases such as diabetic retinopathy.

Renin Angiotensin Aldosterone System Blockade and Renal Disease in Patients with Type 2 Diabetes: An Asian perspective from the RENAAL study

Abstract: OBJECTIVE —Asia is predicted to have the largest population of patients with diabetes who are at high risk for renal disease. In the Reduction of Endpoints in NIDDM with the Angiotensin II Antagonist Losartan (RENAAL) study, ∼17% of patients were Asians. In this subgroup analysis, we examined the characteristics, response, and adherence to treatment of the Asian population, as well as their baseline predictors of risk of renal end points. RESEARCH DESIGN AND METHODS —A total of 252 Asian patients were enrolled in the RENAAL study, which compared losartan (50 mg titrated to 100 mg) to placebo in addition to conventional antihypertensive medications in type 2 diabetic patients with nephropathy. Mean follow-up was 3.2 years. The effect of losartan therapy on renal and cardiovascular outcomes was examined, and baseline predictors of risk were determined using a Cox proportional hazards model with prespecified baseline covariates. RESULTS —Losartan reduced the risk of the primary composite end point composed of a doubling of serum creatinine, end-stage renal disease, or all-cause mortality in Asian patients by 35% ( P = 0.02). No difference between losartan and placebo was observed for the cardiovascular composite outcomes. Losartan reduced the level of proteinuria by 47% ( P P P CONCLUSIONS —In this subgroup analysis of the RENAAL study, losartan conferred significant renal benefits and was well tolerated in Asian patients with type 2 diabetes and clinical nephropathy. Baseline proteinuria and low Hb were strong predictors of risk of renal outcomes.

Changes in Pedigree Backgrounds of Pioneer Brand Maize Hybrids Widely Grown from 1930 to 1999

Abstract: Surveys of pedigree usage indicate that breeding programs can generate genetic diversity in time. There are also concerns that genetic diversity is being lost. Previous studies of pedigree usage in U.S. maize (Zea mays L.) production have reviewed usage of publicly bred inbred lines. However, proprietary inbred lines were already contributing over 90% of U.S. maize production by 1985. This chronological study of germplasm usage compliments previous studies of genetic diversity in U.S. maize production by reporting the pedigree backgrounds of 68 proprietary, widely used maize hybrids released by Pioneer Hi-Bred International during the period 1930 to 1999. Objectives are to identify founder sources for the hybrids and to reveal changes in founder contributions through time. We also compare founder usage with pedigrees of widely used U.S. public inbred lines to measure the extent of their reliance on common germplasm, backgrounds. Pedigrees of the hybrids collectively traced to at least 61 founders. Several founders of the hybrids had complex pedigrees. Founder backgrounds of the hybrids could be traced to at least 22 landraces with additional contributions from other populations or landraces. Public lines used for comparison traced to 14 founders. Nine founders of the public lines were common in the era hybrids; five were unique to the public lines. Differences in founder contributions were evident for the era hybrids and the public lines. Significant contributions from both the private and public sectors were evident in the pedigrees of the era hybrids. Diversity in time was evident. Hybrids tended to associate by decade of initial release. Most new founder contributions occurred in the 1940s (35%), 1960s (36%), and 1980s (20%). Breeding networks have allowed germplasm that was once exotic to the central Corn Belt to contribute improved productivity in that region.

Connective tissue growth factor is up-regulated in the diabetic retina: amelioration by angiotensin-converting enzyme inhibition.

Abstract: Connective tissue growth factor (CTGF) has been postulated to have prosclerotic and angiogenic properties. The aim of this present study was to characterize retinal CTGF expression in the absence and presence of diabetes and in the context of treatment with the angiotensin-converting enzyme (ACE) inhibitor, perindopril. Retinas were obtained from control, diabetic, and diabetic plus perindopril-treated (3 mg/d) rats. CTGF gene expression was quantitated by RT-PCR and localized by in situ hybridization. CTGF protein expression was analyzed by Western blotting and localized by immunohistochemistry. Diabetes was associated with a greater than 2-fold increase in CTGF mRNA levels, which was attenuated by perindopril treatment. CTGF immunoreactivity was increased almost 2-fold in diabetes and was ameliorated by the ACE inhibitor perindopril. By in situ hybridization and immunohistochemistry, the major site of CTGF gene expression in the retina of diabetic rats was the ganglion cell layer. Based on the known in vivo effects of CTGF, it is postulated that this growth factor plays a pivotal role in mediating diabetes-associated retinal pathology. Furthermore, the protective effects of ACE inhibitors on retinal pathology may partly be mediated via effects on retinal CTGF expression.

Identification of angiotensin converting enzyme 2 in the rodent retina.

Abstract: Purpose. An active renin-angiotensin system has been found in the retina of rats and humans. Angiotensin-converting enzyme 2 (ACE2) is a recently discovered enzymatic homologue of Angiotensin-converting enzyme (ACE) that may be an important new component of the renin-angiotensin system (RAS). This study assesses the involvement of ACE2 in the normal and diabetic rodent retina and its modulation by ACE inhibition.Methods. Sprague-Dawley rats were randomised into three groups, control, diabetes, and diabetes plus ramipril, with diabetes induced with the β-cell toxin streptozocin and the study run for 24 weeks. ACE2 and ACE gene levels were measured using quantitative real-time polymerase chain reaction (QRT-PCR), ACE2 protein expression was confirmed by Western blotting, and ACE and ACE2 catalytic activity were measured using specific activity assays in the rat retina. Localisation of ACE2 mRNA and protein were determined by in situ hybridisation and immunohistochemistry, respectively.Results. ACE mRNA leve...

A novel method of resistance for influenza against a channel-blocking antiviral drug.

Abstract: Effective antivirals are few and far between, and as such, the appearance of resistance toward such treatments is an obvious medical concern In this article, we analyze the mechanism by which influenza attains resistance toward amantadine, a blocker of the viral M2 H(+) channel Binding analyses of amantadine to M2 peptides from different viral strains showed that the virus has developed two alternate routes to avoid blockage of its channel: (1) a conventional route, in which the channel no longer binds the blocker and, hence, the blocker cannot exert its inhibitory function; and (2) a novel mechanism, in which binding of the blocker is retained, yet the function of the protein is unaffected Pore diameter profiles revealed the molecular mechanism by which the virus may attain this novel type of resistance: an increase in the size of the channel Thus, despite the drug binding the channel, it may not be able to block the pore, since the channel diameter has increased Our findings may have broad ramifications in the design of new antivirals, and of novel blockers against malfunctioning human channels implicated in disease

Superior renoprotective effects of combination therapy with ACE and AGE inhibition in the diabetic spontaneously hypertensive rat.

Abstract: Diabetic renal disease has been postulated to progress as a result of an interaction between metabolic and haemodynamic pathways. Our aim was to assess the functional, structural, molecular and cellular aspects of renal disease in an experimental model of diabetes with associated hypertension. Streptozotocin-induced diabetic spontaneously hypertensive rats were randomised to no treatment, the ACE inhibitor, perindopril (2 mg/l), the AGE formation inhibitor, aminoguanidine (1 g/l) and a combination of both agents and were followed for 32 weeks. Diabetes was associated with a considerable increase in albumin excretion rate. Both aminoguanidine and perindopril retarded the increase in albuminuria, which was completely abrogated by combination therapy. Glomerulosclerosis and tubulointerstitial damage was reduced by both monotherapies with further renoprotection afforded by combination therapy in both cases. Combination therapy was also associated with a superior restoration in diabetes-induced nephrin protein depletion compared to either monotherapy. TGFβ1 expression as assessed by in situ hybridisation was increased in the diabetic rats and reduced by perindopril and aminoguanidine. These findings indicate that in the context of diabetes-related renal injury, blocking both the renin-angiotensin and advanced glycation pathways offers superior renoprotection and could be considered as a therapeutic strategy in the prevention and retardation of progressive-diabetic renal injury.

Simulating the effects of dominance and epistasis on selection response in the CIMMYT wheat breeding program using QuCim

Abstract: Plant breeders use many different breeding methods to develop superior cultivars. However, it is difficult, cumbersome, and expensive to evaluate the performance of a breeding method or to compare the efficiencies of different breeding methods within an ongoing breeding program. To facilitate comparisons, we developed a QU-GENE module called QuCim that can simulate a large number of breeding strategies for self-pollinated species. The wheat breeding strategy Selected Bulk used by CIMMYT's wheat breeding program was defined in QuCim as an example of how this is done. This selection method was simulated in QuCim to investigate the effects of deviations from the additive genetic model, in the form of dominance and epistasis, on selection outcomes. The simulation results indicate that the partial dominance model does not greatly influence genetic advance compared with the pure additive model. Genetic advance in genetic systems with overdominance and epistasis are slower than when gene effects are purely additive or partially dominant. The additive gene effect is an appropriate indicator of the change in gene frequency following selection when epistasis is absent. In the absence of epistasis, the additive variance decreases rapidly with selection. However, after several cycles of selection it remains relatively fixed when epistasis is present. The variance from partial dominance is relatively small and therefore hard to detect by the covariance among half sibs and the covariance among full sibs. The dominance variance from the overdominance model can be identified successfully, but it does not change significantly, which confirms that overdominance cannot be utilized by an inbred breeding program. QuCim is an effective tool to compare selection strategies and to validate some theories in quantitative genetics.

The time has come to target connective tissue growth factor in diabetic complications

Abstract: es that occur in cells and tissues in diabetes. As growth factors commonly control essential biological functions, it follows that they may be critical in causing the end-organ complications of diabetes [1]. Consequently, identifying the key growth factor(s) that cause a particular type and stage of diabetic tissue damage is of potential importance in preventing and treating diabetes complications. In this context, connective tissue growth factor (CTGF), also known as CCN2 [2], is a prime candidate. Following its identification in 1991, this 38-Mr cysteine-rich secreted protein has been shown to exhibit a broad spectrum of biological activities, often specific to cell and tissue type. Many of the effects of CTGF reflect cellular changes that, characteristically, occur more frequently than normal in diabetic complications. The main effects of CTGF are the regulation of extracellular matrix (ECM) accumulation [3], cell hypertrophy and mitogenesis, and in some cases, apoptosis [4], cellular adhesion and chemotaxis [5], and cell trans-differentiation [3]. The complex biological process of angiogenesis can also be promoted by CTGF [6]. CTGF gene expression and protein levels are upregulated by diabetes in tissues that are susceptible to end-organ injury. The kidney has been the main focus of CTGF research in diabetes. CTGF was found to be increased both in rodent models of diabetic renal disease, in the glomerulus [7, 8] and the tubulo-interstitium [9], and in human diabetes, in the same sites, involving the resident cells of the kidney [10]. Increases in CTGF renal expression correspond with the extent of glomerulosclerosis and tubulo-interstitial fibrosis,