Heart failure and chronic kidney disease (CKD) share several mediators of cardiac pathological remodelling. Akin to heart failure, this remodelling sets in motion a vicious cycle of progressive ...pathological hypertrophy and myocardial dysfunction in CKD. Several decades of heart failure research have shown that beta blockade is a powerful tool in preventing cardiac remodelling and breaking this vicious cycle. This phenomenon remains hitherto untested in CKD. Therefore, we set out to test the hypothesis that beta blockade prevents cardiac pathological remodelling in experimental uremia.
rats had subtotal nephrectomy or sham surgery and were followed up for 10 weeks. The animals were randomly allocated to the beta blocker metoprolol (10 mg/kg/day) or vehicle. In vivo and in vitro cardiac assessments were performed. Cardiac tissue was extracted, and protein expression was quantified using immunoblotting. Histological analyses were performed to quantify myocardial fibrosis. Beta blockade attenuated cardiac pathological remodelling in nephrectomised animals. The echocardiographic left ventricular mass and the heart weight to tibial length ratio were significantly lower in nephrectomised animals treated with metoprolol. Furthermore, beta blockade attenuated myocardial fibrosis associated with subtotal nephrectomy. In addition, the Ca
- calmodulin-dependent kinase II (CAMKII) pathway was shown to be activated in uremia and attenuated by beta blockade, offering a potential mechanism of action. In conclusion, beta blockade attenuated hypertrophic signalling pathways and ameliorated cardiac pathological remodelling in experimental uremia. The study provides a strong scientific rationale for repurposing beta blockers, a tried and tested treatment in heart failure, for the benefit of patients with CKD.
Molecular recognition reagents are key tools for understanding biological processes and are used universally by scientists to study protein expression, localisation and interactions. Antibodies ...remain the most widely used of such reagents and many show excellent performance, although some are poorly characterised or have stability or batch variability issues, supporting the use of alternative binding proteins as complementary reagents for many applications. Here we report on the use of Affimer proteins as research reagents. We selected 12 diverse molecular targets for Affimer selection to exemplify their use in common molecular and cellular applications including the (a) selection against various target molecules; (b) modulation of protein function in vitro and in vivo; (c) labelling of tumour antigens in mouse models; and (d) use in affinity fluorescence and super-resolution microscopy. This work shows that Affimer proteins, as is the case for other alternative binding scaffolds, represent complementary affinity reagents to antibodies for various molecular and cell biology applications.
Oestrogen influences autonomic function via actions at classical nuclear oestrogen receptors α and β in the brain, and recent evidence suggests the orphan G protein-coupled receptor GPR30 may also ...function as a cytoplasmic oestrogen receptor. We investigated the expression of GPR30 in female rat brains throughout the oestrous cycle and after ovariectomy to determine whether GPR30 expression in central autonomic nuclei is correlated with circulating oestrogen levels. In the nucleus of the solitary tract (NTS), ventrolateral medulla (VLM) and periaqueductal gray (PAG) GPR30 mRNA, quantified by real-time PCR, was increased in proestrus and oestrus. In ovariectomised (OVX) rats, expression in NTS and VLM appeared increased compared to metoestrus, but in the hypothalamic paraventricular nucleus and PAG lower mRNA levels were seen in OVX. GPR30-like immunoreactivity (GPR30-LI) colocalised with Golgi in neurones in many brain areas associated with autonomic pathways, and analysis of numbers of immunoreactive neurones showed differences consistent with the PCR data. GPR30-LI was found in a variety of transmitter phenotypes, including cholinergic, serotonergic, catecholaminergic and nitrergic neurones in different neuronal groups. These observations support the view that GPR30 could act as a rapid transducer responding to oestrogen levels and thus modulate the activity of central autonomic pathways.
Little is known about the prevalence of protozoan parasites in the muscles of rock pigeons (Columbia livia). The muscles from 54 (heart from 45 and breast from 54) rock pigeons were examined for DNA ...of Toxoplasma gondii, Neospora caninum, and Sarcocystis species using PCR. Twenty-four were female and 30 were males. The birds were part of flocks of pigeons housed at the tombs of saints in Lahore, Pakistan. Birds that died or were euthanized due to poor health were submitted for necropsy at the Department of Parasitology, University of Veterinary and Animal Sciences, Lahore, Pakistan, where DNA isolations and PCR were conducted. Nineteen (35.1%) of the birds were positive for T. gondii DNA. Seven males and 12 females were positive. Breast tissue was always infected in T. gondii positive birds, while the heart was infected in 13 (28.8%) of breast positive birds. Five (9.2%) of the pigeons, 2 males and 3 females, were positive for N. caninum. The distribution of N. caninum DNA was more variable in the muscles of pigeons than T. gondii and was found only in the heart of 1 (female), heart and breast muscle of 2 (male), and only the breast muscle of 2 birds (female). One of the 54 rock pigeons (female) was positive for both T. gondii (heart and breast) and N. caninum (heart only). Two of the positive Neospora caninum amplicons were sequenced and had 97% nucleotide identity with N. caninum isolates. Sarcocystis DNA was not found in any bird. The prevalence of T. gondii in rock pigeons and their predation by cats suggest that they may play an unrecognized role in maintaining environmental contamination with T. gondii oocysts by cats. Our study indicates that rock pigeons are intermediate hosts of N. caninum and this information will aid in understanding the epidemiology of N. caninum.
AIM:To investigate the effect of Tenascin C(TNC)on the expression of pro-inflammatory cytokines and matrixmetalloproteinases in human cardiac myofibroblasts(CMF).METHODS:CMF were isolated and ...cultured from patients undergoing coronary artery bypass grafting.Cultured cells were treated with either TNC(0.1μmol/L,24 h)or a recombinant protein corresponding to different domains of the TNC protein;fibrinogen-like globe(FBG)and fibronectin typeⅢ-like repeats(TNⅢ5-7)(both 1μmol/L,24 h).The expression of the proinflammatory cytokines;interleukin(IL)-6,IL-1β,TNFαand the matrix metalloproteinases;MMPs(MMP1,2,3,9,10,MT1-MMP)was assessed using real time RT-PCR and western blot analysis.RESULTS:TNC increased both IL-6 and MMP3(P<0.01)mR NA levels in cultured human CMF but had no significant effect on the other markers studied.The increase in IL-6 mR NA expression was mirrored by an increase in protein secretion as assessed by enzymelinked immunosorbant assay(P<0.01).Treating CMF with the recombinant protein FBG increased IL-6mR NA and protein(P<0.01)whereas the recombinant protein TNⅢ5-7 had no effect.Neither FBG nor TNⅢ5-7 had any significant effect on MMP3 expression.The expression of toll-like receptor 4(TLR4)in human CMF was confirmed by real time RT-PCR,western blot and immunohistochemistry.Pre-incubation of cells with TLR4neutralising antisera attenuated the effect of both TNC and FBG on IL-6 mR NA and protein expression.CONCLUSION:TNC up-regulates IL-6 expression in human CMF,an effect mediated through the FBG domain of TNC and via the TLR4 receptor.
Two commercial preparations of Bacillus thuringiensis var. israelensis VectoBac WDG and Mousticide WP were evaluated and compared for larval control of Anopheles stephensi a malaria vector in ...Pakistan under laboratory and semi field conditions. An. stephensi larvae were susceptible to VectoBac WDG and Mousticide WP but susceptibility was higher for VectoBac WDG. In semi field experiment VectoBac WDG at 0.4 mg/L (1X LC100) and1.6 mg/L (4X LC100) remained more effective against An. stephensi L3 larvae up to day 7 whereas larval mortality dropped below 50% at day 14 of post VectoBac WDG application. Mousticide WP at 1.22 mg/L (1X LC100) produced 79.33% mean mortality of An. stephensi L3 at 1 day post treatment. The larval mortality dropped to 30% at day 14 of post treatment with Mousticide WP. No significant difference was seen in larval mortality between two dose rates of VectoBac WDG and Mousticide WP. Copyright 2014 Friends Science Publishers
The development of novel protein-targeted MRI contrast agents crucially depends on the ability to derivatise suitable targeting moieties with a high payload of relaxation enhancer (
, gadolinium(iii) ...complexes such as Gd-DOTA), without losing affinity for the target proteins. Here, we report robust synthetic procedures for the preparation of trivalent Gd-DOTA reagents with various chemical handles for site-specific modification of biomolecules. The reagents were shown to successfully label proteins through isothiocyanate ligation or through site-specific thiol-maleimide ligation and strain-promoted azide-alkyne cycloaddition.
The purpose of this study was to record the seroprevalence of Toxoplasma gondii antibodies in buthchers and buffaloes at Lahore Pakistan. Blood samples of 50 butchers and 50 buffaloes were taken from ...main abattoir and local buffalo farms around Lahore and assessed for seroprevalence of T. gondii by using a latex agglutination test (LAT). In the present study 10% of butchers and 14% buffaloes were found seropositive for T. gondii infection. Butchers ranging in age from 51-60 (n=12) years and above exhibited the highest seropositivity 25% followed by those aging 41- 50 years (n=10) 20%. Similarly in buffaloes 14% was also related to age by using LAT.
Abstract
Background and Aims
Heart failure (HF) and uremia share several potent cardiac hypertrophic stimuli such as wall stress, sympathetic activation, renin angiotensin system activation and ...oxidative stress. In heart failure, induction of fetal gene program is one of the major processes through which such stimuli lead to progressive pathological hypertrophy and myocardial dysfunction. The fetal genes are activated by the transcription factor, myocyte enhance factor 2 (MEF2) which is a downstream target of the Ca++- Calmodulin Dependent Kinase II (CAMKII) pathway. The CAMKII pathway has been shown to be the major transduction pathway of the hypertrophic stimuli in HF 1, 2. We hosphorzed that such mechanisms are also at play in uremic cardiomyopathy and tested the hypothesis in a rodent model of experimental uremia.
Method
Wistar rats (n = 18) had subtotal nephrectomy (STNx) or sham surgery (sham) and were followed up for 10 weeks. In vivo and in vitro cardiac assessments were performed. Cardiac tissue was extracted, and protein expression of CAMKII and hosphor-CAMKII were quantified using immunoblotting. Protein expression of MEF2 and its target fetal genes were also studied. Histological analyses were performed to quantify myocardial fibrosis. Data was analysed using an independent sample t-test with Welch's correction and expressed as mean± SEM.
Results
STNx resulted in higher echocardiographic left ventricular mass, heart weight to tibia length ratio and myocardial fibrosis in the STNx group (Fig. 1) demonstrating cardiac pathological hypertrophy. CaMKII signalling was activated and the expression of MEF2 was also increased following STNx (Fig. 1). There was trend towards increased expression of fetal genes (Fig. 1). However, the difference did not reach statistical significance.
Conclusion
The study shows that experimental uremia induces cardiac pathological hypertrophy along with activation of intracellular pathways involved in fetal gene induction. Further studies evaluating fetal genes at an earlier time course in the evolution of pathological hypertrophy will be worthwhile.
Abstract
Background and Aims
Fifty years of heart failure research has shown that pathological cardiac remodelling forms a vicious cycle with myocardial dysfunction leading to progressive heart ...failure (HF) Circulation, 102
IV14-23, 2000. Fetal gene induction is associated with this process and beta blocker therapy has been shown to prevent it. Although chronic kidney disease (CKD) and HF share similar mediators of cardiac remodelling, the benefits of beta blocker therapy in CKD has not been studied. We, therefore, tested the hypothesis that beta blocker therapy prevents fetal gene induction and pathological cardiac remodelling in experimental uraemia.
Method
Wistar rats (n=32) had subtotal nephrectomy (STNx) Frontiers in physiology, 10
1365, 2019 or sham surgery and were followed up for 10 weeks. The animals were randomly allocated to metoprolol (10mg/kg/day) or vehicle. In vivo and in vitro cardiac assessments were performed, and changes in myocardial fetal gene expression were also studied.
Results
Heart rate was significantly lower in metoprolol groups compared to untreated groups demonstrating effective beta blockade (Fig 1A). Echocardiographic LV mass was significantly higher in untreated STNx group compared to the metoprolol group (896.4 vs 632.2g, P=0.0004). Similar changes were seen with heart weight to tibia ratio (Fig 1B). There was no significant difference in blood pressure (BP) between treated and untreated STNx animals (123 vs 119 mmHg, P=0.359) (Fig 1A). STNx increased mRNA expression of fetal genes and there was a trend towards attenuation of this increase with beta blocker therapy (Fig 1C).
Conclusion
Beta blocker therapy ameliorates uraemic pathological cardiac remodelling irrespective of changes to BP. This benefit appears be associated with a reduction of induced fetal gene expression. Further translational research on the benefits of beta blockade in the treatment of uraemic cardiomyopathy is required.
PDF
