Emerging spectrum of post-COVID-19 syndrome.

'Post-COVID-19 syndrome' refers to symptoms in the convalescent phase following initial COVID-19 infection. This term encompasses a wide array of presentation involving lungs, heart and the neuromuscular system. Pulmonary manifestations include post-COVID-19 fibrosis, which is akin to post acute respiratory distress syndrome fibrosis and may reflect the permanent damage to the lungs following an initial bout of infection. Cardiovascular system is often involved, and the presentation can be in terms of acute coronary syndrome, myocarditis and heart failure. Clinical manifestations are often varied and non-specific, which entails a detailed workup and a multidisciplinary approach. Post-COVID-19 syndrome adds to the overall disease morbidity and leads to a prolonged hospital stay, greater healthcare utilisation and loss of productivity marring the country's dwindling economy. Thus, it is imperative that post-COVID-19 syndrome be prevented and identified early followed by a prompt treatment.

Management of cardiovascular emergencies during the COVID-19 pandemic.

BACKGROUND: It has been reported that patients attending the emergency department with other pathologies may not have received optimal medical care due to the lockdown measures in the early phase of the COVID-19 pandemic. METHODS: This was a retrospective study of patients presenting with cardiovascular emergencies to four tertiary regional emergency departments in western India during the government implementation of complete lockdown. RESULTS: 25.0% of patients during the lockdown period and 17.4% of patients during the pre-lockdown period presented outside the window period (presentation after 12 hours of symptom onset) compared with only 6% during the pre-COVID period. In the pre-COVID period, 46.9% of patients with ST elevation myocardial infarction underwent emergent catheterisation, while in the pre-lockdown and lockdown periods, these values were 26.1% and 18.8%, respectively. The proportion of patients treated with intravenous thrombolytic therapy increased from 18.4% in the pre-COVID period to 32.3% in the post-lockdown period. Inhospital mortality for acute coronary syndrome (ACS) increased from 2.69% in the pre-COVID period to 7.27% in the post-lockdown period. There was also a significant decline in emergency admissions for non-ACS conditions, such as acute decompensated heart failure and high degree or complete atrioventricular block. CONCLUSION: The COVID-19 pandemic has led to delays in patients seeking care for cardiac problems and also affected the use of optimum therapy in our institutions.

RGS4 is a negative regulator of insulin release from pancreatic beta-cells in vitro and in vivo.

Therapeutic strategies that augment insulin release from pancreatic beta-cells are considered beneficial in the treatment of type 2 diabetes. We previously demonstrated that activation of beta-cell M(3) muscarinic receptors (M3Rs) greatly promotes glucose-stimulated insulin secretion (GSIS), suggesting that strategies aimed at enhancing signaling through beta-cell M3Rs may become therapeutically useful. M3R activation leads to the stimulation of G proteins of the G(q) family, which are under the inhibitory control of proteins known as regulators of G protein signaling (RGS proteins). At present, it remains unknown whether RGS proteins play a role in regulating insulin release. To address this issue, we initially demonstrated that MIN6 insulinoma cells express functional M3Rs and that RGS4 was by far the most abundant RGS protein expressed by these cells. Strikingly, siRNA-mediated knockdown of RGS4 expression in MIN6 cells greatly enhanced M3R-mediated augmentation of GSIS and calcium release. We obtained similar findings using pancreatic islets prepared from RGS4-deficient mice. Interestingly, RGS4 deficiency had little effect on insulin release caused by activation of other beta-cell GPCRs. Finally, treatment of mutant mice selectively lacking RGS4 in pancreatic beta-cells with a muscarinic agonist (bethanechol) led to significantly increased plasma insulin and reduced blood glucose levels, as compared to control littermates. Studies with beta-cell-specific M3R knockout mice showed that these responses were mediated by beta-cell M3Rs. These findings indicate that RGS4 is a potent negative regulator of M3R function in pancreatic beta-cells, suggesting that RGS4 may represent a potential target to promote insulin release for therapeutic purposes.

Effect of Flow Rate and Ionic Strength on the Stabilities of YOYO-1 and YO-PRO-1 Intercalated in DNA Molecules.

Single-molecule DNA studies have improved our understanding of the DNAs' structure and their interactions with other molecules. A variety of DNA labeling dyes are available for single-molecule studies, among which the bis-intercalating dye YOYO-1 and mono-intercalating dye YO-PRO-1 are widely used. They have an extraordinarily strong affinity toward DNA and are bright with a high quantum yield (>0.5) when bound to DNAs. However, it is still not clear how these dyes behave in DNA molecules under higher ionic strength and strong buffer flow. Here, we have studied the effect of ionic strength and flow rate of buffer on their binding in single DNA molecules. The larger the flow rate and the higher the ionic strength, the faster the intercalated dyes are washed away from the DNAs. In the buffer with 1 M ionic strength, YOYO-1 and YO-PRO-1 are mostly washed away from DNA within 2 min of moderate buffer flow.

Neuronal M3 muscarinic acetylcholine receptors are essential for somatotroph proliferation and normal somatic growth.

The molecular pathways that promote the proliferation and maintenance of pituitary somatotrophs and other cell types of the anterior pituitary gland are not well understood at present. However, such knowledge is likely to lead to the development of novel drugs useful for the treatment of various human growth disorders. Although muscarinic cholinergic pathways have been implicated in regulating somatotroph function, the physiological relevance of this effect and the localization and nature of the receptor subtypes involved in this activity remain unclear. We report the surprising observation that mutant mice that selectively lack the M(3) muscarinic acetylcholine receptor subtype in the brain (neurons and glial cells; Br-M3-KO mice) showed a dwarf phenotype associated with a pronounced hypoplasia of the anterior pituitary gland and a marked decrease in pituitary and serum growth hormone (GH) and prolactin. Remarkably, treatment of Br-M3-KO mice with CJC-1295, a synthetic GH-releasing hormone (GHRH) analog, rescued the growth deficit displayed by Br-M3-KO mice by restoring normal pituitary size and normal serum GH and IGF-1 levels. These findings, together with results from M(3) receptor/GHRH colocalization studies and hypothalamic hormone measurements, support a model in which central (hypothalamic) M(3) receptors are required for the proper function of hypothalamic GHRH neurons. Our data reveal an unexpected and critical role for central M(3) receptors in regulating longitudinal growth by promoting the proliferation of pituitary somatotroph cells.

A chemical-genetic approach to study G protein regulation of beta cell function in vivo.

Impaired functioning of pancreatic beta cells is a key hallmark of type 2 diabetes. beta cell function is modulated by the actions of different classes of heterotrimeric G proteins. The functional consequences of activating specific beta cell G protein signaling pathways in vivo are not well understood at present, primarily due to the fact that beta cell G protein-coupled receptors (GPCRs) are also expressed by many other tissues. To circumvent these difficulties, we developed a chemical-genetic approach that allows for the conditional and selective activation of specific beta cell G proteins in intact animals. Specifically, we created two lines of transgenic mice each of which expressed a specific designer GPCR in beta cells only. Importantly, the two designer receptors differed in their G protein-coupling properties (G(q/11) versus G(s)). They were unable to bind endogenous ligand(s), but could be efficiently activated by an otherwise pharmacologically inert compound (clozapine-N-oxide), leading to the conditional activation of either beta cell G(q/11) or G(s) G proteins. Here we report the findings that conditional and selective activation of beta cell G(q/11) signaling in vivo leads to striking increases in both first- and second-phase insulin release, greatly improved glucose tolerance in obese, insulin-resistant mice, and elevated beta cell mass, associated with pathway-specific alterations in islet gene expression levels. Selective stimulation of beta cell G(s) triggered qualitatively similar in vivo metabolic effects. Thus, this developed chemical-genetic strategy represents a powerful approach to study G protein regulation of beta cell function in vivo.

Efficient k-means clustering and greedy selection-based reduction of nodal search space for optimization of sensor placement in the water distribution networks.

Monitoring of water distribution network (WDN) requires placement of sensors at strategic locations to detect maximum contamination events at the earliest. The multi-objective optimization (MOO) of sensor placement is a complicated problem owing to its combinatorial nature, interconnected and large WDN sizes, and temporal flows producing complex outcomes for a given set of contamination events. In this study, a new method is proposed to reduce the complexity of the problem by condensing the nodal search space. This method first segregates the nodes based on intrusion events detected, using k-means clustering, followed by selecting nodes from each group based on the improvement observed in the objectives, namely, contamination event detection, expected detection time, and affected population. The selected nodes formed the decision variable space for the MOO study. The developed strategy was tested on two benchmark networks: BWSN Network1 and C-town network, and its performance is compared with the traditional method in terms of hypervolume contribution rate (CR) indicator and the number of Pareto points. The optimal subset of nodes generated twice the number of Pareto points than the complete set of nodes set for placing 20 sensors and had 10% more than CR indicator than the traditional method. For the placement of 5 sensors, the proposed solutions were better at the higher detection likelihood values, which is required to achieve maximum detection. The proposed sensor placement algorithm can be easily scaled to large WDNs. It is expected to provide a better optimal sensor placement solution irrespective of network size as compared to the traditional approach.

Beneficial metabolic effects of M3 muscarinic acetylcholine receptor deficiency.

Most animal models of obesity and hyperinsulinemia are associated with increased vagal cholinergic activity. The M3 muscarinic acetylcholine receptor subtype is widely expressed in the brain and peripheral tissues and plays a key role in mediating the physiological effects of vagal activation. Here, we tested the hypothesis that the absence of M3 receptors in mice might protect against various forms of experimentally or genetically induced obesity and obesity-associated metabolic deficits. In all cases, the lack of M3 receptors greatly ameliorated impairments in glucose homeostasis and insulin sensitivity but had less robust effects on overall adiposity. Under all experimental conditions tested, M3 receptor-deficient mice showed a significant elevation in basal and total energy expenditure, most likely due to enhanced central sympathetic outflow and increased rate of fatty-acid oxidation. These findings suggest that the M3 receptor may represent a potential pharmacologic target for the treatment of obesity and associated metabolic disorders.

A critical role for beta cell M3 muscarinic acetylcholine receptors in regulating insulin release and blood glucose homeostasis in vivo.

One of the hallmarks of type 2 diabetes is that pancreatic beta cells fail to release sufficient amounts of insulin in the presence of elevated blood glucose levels. Insulin secretion is modulated by many hormones and neurotransmitters including acetylcholine, the major neurotransmitter of the peripheral parasympathetic nervous system. The physiological role of muscarinic acetylcholine receptors expressed by pancreatic beta cells remains unclear at present. Here, we demonstrate that mutant mice selectively lacking the M3 muscarinic acetylcholine receptor subtype in pancreatic beta cells display impaired glucose tolerance and greatly reduced insulin release. In contrast, transgenic mice selectively overexpressing M3 receptors in pancreatic beta cells show a profound increase in glucose tolerance and insulin release. Moreover, these mutant mice are resistant to diet-induced glucose intolerance and hyperglycemia. These findings indicate that beta cell M3 muscarinic receptors play a key role in maintaining proper insulin release and glucose homeostasis.

Distinct muscarinic acetylcholine receptor subtypes contribute to stability and growth, but not compensatory plasticity, of neuromuscular synapses.

Muscarinic acetylcholine receptors (mAChRs) modulate synaptic function, but whether they influence synaptic structure remains unknown. At neuromuscular junctions (NMJs), mAChRs have been implicated in compensatory sprouting of axon terminals in paralyzed or denervated muscles. Here we used pharmacological and genetic inhibition and localization studies of mAChR subtypes at mouse NMJs to demonstrate their roles in synaptic stability and growth but not in compensatory sprouting. M(2) mAChRs were present solely in motor neurons, whereas M(1), M(3), and M(5) mAChRs were associated with Schwann cells and/or muscle fibers. Blockade of all five mAChR subtypes with atropine evoked pronounced effects, including terminal sprouting, terminal withdrawal, and muscle fiber atrophy. In contrast, methoctramine, an M(2/4)-preferring antagonist, induced terminal sprouting and terminal withdrawal, but no muscle fiber atrophy. Consistent with this observation, M(2)(-/-) but no other mAChR mutant mice exhibited spontaneous sprouting accompanied by extensive loss of parental terminal arbors. Terminal sprouting, however, seemed not to be the causative defect because partial loss of terminal branches was common even in the M(2)(-/-) NMJs without sprouting. Moreover, compensatory sprouting after paralysis or partial denervation was normal in mice deficient in M(2) or other mAChR subtypes. We also found that many NMJs of M(5)(-/-) mice were exceptionally small and reduced in proportion to the size of parental muscle fibers. These findings show that axon terminals are unstable without M(2) and that muscle fiber growth is defective without M(5). Subtype-specific muscarinic signaling provides a novel means for coordinating activity-dependent development and maintenance of the tripartite synapse.

Muscarinic acetylcholine receptors: mutant mice provide new insights for drug development.

Muscarinic acetylcholine receptors (mAChRs), M(1)-M(5), regulate the activity of numerous fundamental central and peripheral functions. The lack of small-molecule ligands that can block or activate specific mAChR subtypes with high selectivity has remained a major obstacle in defining the roles of the individual receptor subtypes and in the development of novel muscarinic drugs. Recently, phenotypic analysis of mutant mouse strains deficient in each of the five mAChR subtypes has led to a wealth of new information regarding the physiological roles of the individual receptor subtypes. Importantly, these studies have identified specific mAChR-regulated pathways as potentially novel targets for the treatment of various important disorders including Alzheimer's disease, schizophrenia, pain, obesity and diabetes.

Cardiovascular complications and its impact on outcomes in COVID-19.

BACKGROUND: Coronavirus disease 2019 (COVID-19) has led to a widespread morbidity and mortality. Limited data exists regarding the involvement of cardiovascular system in COVID-19 patients. We sought to evaluate the cardiovascular (CV) complications and its impact on outcomes in symptomatic COVID-19 patients. METHODS: This was a single center observational study among symptomatic COVID-19 patients. Data regarding clinical profile, laboratory investigations, CV complications, treatment and outcomes were collected. Cardiac biomarkers and 12 lead electrocardiograms were done in all while echocardiography was done in those with clinical indications for the same. Corrected QT-interval (QTc) at baseline and maximum value during hospitalization were computed. RESULTS: Of the 108 patients, majority of them were males with a mean age of 51.2 ± 17.7 years. Hypertension (38%) and diabetes (32.4%) were most prevalent co-morbidities. ECG findings included sinus tachycardia in 18 (16.9%), first degree AV block in 5 (4.6%), VT/VF in 2 (1.8%) and sinus bradycardia in one (0.9%). QTc prolongation was observed in 17.6% subjects. CV complications included acute cardiac injury in 25.9%, heart failure, cardiogenic shock and acute coronary syndrome in 3.7% each, "probable" myocarditis in 2.8% patients. Patients with acute cardiac injury had higher mortality than those without (16/28 [57.1%] vs 14/78 [17.5%]; P < 0.0001). Multivariate logistic regression analysis showed that acute cardiac injury (OR: 11.3), lymphopenia (OR: 4.91), use of inotropic agents (OR: 2.46) and neutrophil-lymphocyte ratio (OR:1.1) were independent predictors of mortality. CONCLUSIONS: CV complications such as acute cardiac injury is common in COVID-19 patients and is associated with worse prognosis.

Single molecule protein patterning using hole mask colloidal lithography.

The ability to manipulate single protein molecules on a surface is useful for interfacing biology with many types of devices in optics, catalysis, bioengineering, and biosensing. Control of distance, orientation, and activity at the single molecule level will allow for the production of on-chip devices with increased biological activity. Cost effective methodologies for single molecule protein patterning with tunable pattern density and scalable coverage area remain a challenge. Herein, Hole Mask Colloidal Lithography is presented as a bench-top colloidal lithography technique that enables a glass coverslip to be patterned with functional streptavidin protein onto patches from 15-200 nm in diameter with variable pitch. Atomic force microscopy (AFM) was used to characterize the size of the patterned features on the glass surface. Additionally, single-molecule fluorescence microscopy was used to demonstrate the tunable pattern density, measure binding controls, and confirm patterned single molecules of functional streptavidin.

Antimicrobial studies, synthesis and characterization of novel 1-nitro-10H-phenothiazine bearing sulfone/nucleoside moieties.

The current research article involves one pot synthesis of novel substituted 1-nitro-10H-phenothiazines via Smiles rearrangement. These substituted phenothiazines undergo oxidation to yield 10H-phenothiazine-5,5-dioxides (sulfones) while on treatment with ß-D-ribofuranose-1-acetate-2,3,5-tribenzoate yield ribofuranosides. These compounds were screened for their antimicrobial vitalities (in vitro) against selected strains of bacteria and fungi. The characterization of synthesized compounds was done by elemental and spectral studies.

Kinetic Model under Light-Limited Condition for Photoinitiated Thiol-Ene Coupling Reactions.

Thiol-ene click chemistry has become a powerful paradigm in synthesis, materials science, and surface modification in the past decade. In the photoinitiated thiol-ene reaction, an induction period is often observed before the major change in its kinetic curve, for which a possible mechanism is proposed in this report. Briefly, light soaking generates radicals following the zeroth-order reaction kinetics. The radical is the reactant that initializes the chain reaction of thiol-ene coupling, which is a first-order reaction. Combining both and under the light-limited conditions, a surprising kinetics represented by a Gaussian-like model evolves that is different from the exponential model used to describe the first-order reaction of the final product. The experimental data are fitted well with the new model, and the reaction kinetic constants can be pulled out from the fitting.

Conditional disruption of IkappaB kinase 2 fails to prevent obesity-induced insulin resistance.

The inhibitor of NF-kappaB (IkappaB) kinases (IKK1[alpha] and IKK2[beta]), the catalytic subunits of the IKK complex, phosphorylate IkappaB proteins on serine residues, targeting them for degradation and thus activating the transcription factor NF-kappaB. More recently, IKK2 has been implicated in mediation of insulin resistance caused by obesity, lipid infusion, and TNF-alpha stimulation, since salicylate and aspirin, known inhibitors of IKK activity, can reverse insulin resistance in obese mouse models. To further genetically elucidate the role of IKK2 in obesity-mediated insulin resistance, we have conditionally inactivated the mouse IKK2 gene in adult myocytes by Cre-loxP-mediated recombination in vivo. We have investigated the development of obesity-induced insulin resistance in muscle-specific IKK2 knockout mice and mice exhibiting a 50% reduction of IKK2 expression in every tissue and have found that, after gold thioglucose treatment, wild-type and mutant mice developed obesity to a similar extent. Surprisingly, no difference in obesity-induced insulin resistance was detectable, either at a physiological or at a molecular level. Moreover, impaired glucose tolerance resulting from a high-fat diet occurred to the same degree in control and IKK2 mutant mice. These data argue against a substantial role for muscular IKK2 in mediating obesity-induced insulin resistance in these models in vivo.

Control of glycinergic input to spinal dorsal horn neurons by distinct muscarinic receptor subtypes revealed using knockout mice.

Muscarinic acetylcholine receptors (mAChRs) play an important role in the tonic regulation of nociceptive transmission in the spinal cord. However, how mAChR subtypes contribute to the regulation of synaptic glycine release is unknown. To determine their role, glycinergic spontaneous inhibitory postsynaptic currents (sIPSCs) were recorded in lamina II neurons by using whole-cell recordings in spinal cord slices of wild-type (WT) and mAChR subtype knockout (KO) mice. In WT mice, the mAChR agonist oxotremorine-M dose-dependently decreased the frequency of sIPSCs in most neurons, but it had variable effects in other neurons. In contrast, in M3-KO mice, oxotremorine-M consistently decreased the glycinergic sIPSC frequency in all neurons tested, and in M2/M4 double-KO mice, it always increased the sIPSC frequency. In M2/M4 double-KO mice, the potentiating effect of oxotremorine-M was attenuated by higher concentrations in some neurons through activation of GABA(B) receptors. In pertussis toxin-treated WT mice, oxotremorine-M also consistently increased the sIPSC frequency. In M2-KO and M4-KO mice, the effect of oxotremorine-M on sIPSCs was divergent because of the opposing functions of the M3 subtype and the M2 and M4 subtypes. This study demonstrates that stimulation of the M2 and M4 subtypes inhibits glycinergic inputs to spinal dorsal horn neurons of mice, whereas stimulation of the M3 subtype potentiates synaptic glycine release. Furthermore, GABA(B) receptors are involved in the feedback regulation of glycinergic synaptic transmission in the spinal cord. This study revealed distinct functions of mAChR subtypes in controlling glycinergic input to spinal dorsal horn neurons.

M1-M3 muscarinic acetylcholine receptor-deficient mice: novel phenotypes.

The five muscarinic acetylcholine receptors (M1-M5 mAChRs) mediate a very large number of important physiological functions (Caulfield, 1993; Caulfield and Birdsall, 1998; Wess, 2004). Because of the lack of small molecule ligands endowed with a high degree of receptor subtype selectivity and the fact that most tissues or cell types express two or more mAChR subtypes, identification of the physiological and pathophysiological roles of the individual mAChR subtypes has proved to be a challenging task. To overcome these difficulties, we recently generated mutant mouse lines deficient in each of the five mAChR genes (M1R-/- mice, M2R-/- mice, M3R-/- mice, etc. [Wess, 2004]). Phenotyping studies showed that each of the five mutant mouse lines displayed characteristic physiological, pharmacological, behavioral, biochemical, or neurochemical deficits (Wess, 2004). This chapter summarizes recent findings dealing with the importance of the M2mAChR for cognitive processes and the roles of the M1 and M3 mAChRs in mediating stimulation of glandular secretion.

Awareness, treatment adherence and risk predictors of uncontrolled hypertension at a tertiary care teaching hospital in Western India.

INTRODUCTION: Hypertension still remains poorly controlled. RESULT: Adequate BP control was achieved in 37.4% of patients and significant attributes for poor control were BMI, marital status, literacy, socioeconomic status, smoking, medication adherence, absence of side effects, number of drugs, number of years on drug therapy and co-morbid conditions.