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.

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.

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.

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.

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.

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.

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.

Ileosigmoid knotting: a rare case report with review of literature†.

Ileosigmoid knotting (ISK) is a rare cause of intestinal obstruction in which loops of ileum and sigmoid colon wrap around each other. It is very uncommon in western world when compared with the African and Asian region. It is rapidly a progressive, fatal disease. Early diagnosis and intervention is the key of better outcome. We are reporting a case of 51-year-old male who presented with shock within 24 h of onset of symptoms. Exploratory laparotomy revealed ISK causing gangrene of ileum and sigmoid colon. In view of haemodynamic unstability, end ileostomy was done after excising gangrenous segments. The patient expired after 2 weeks due to complications of short bowl syndrome. We are also tabulating all cases of ISK reported in the literature till date.

Synthesis and antitubercular screening of some novel 4H-1,4- benzothiazines and their sulfones under environment benign solvent free conditions as future anti-tubercular agents.

Some novel analogs of 4H-1,4-benzothiazines were synthesized under environmentally benign solvent free conditions by one pot oxidative cyclocondensation of substituted 2- aminobenzenthiols with compounds having active methylene group and then converted in to sulfones. These compounds were examined as antitubercular agents against Mycobacterium tuberculosis H37Rv using REMA plate method. The best results have MICs from 6.4 to 8.8 µg/mL, comparable to phenothiazines. IR, NMR and mass spectral investigations are included for structural elucidation.

Synthesis, spectral characterization, and pharmacological importance of new 4H-1,4-benzothiazines, their sulfone analogues, and ribofuranosides.

The present article describes the synthesis of new 4H-1,4-benzothiazines via condensation and oxidative cyclization of substituted 2-aminobenzenethiols with compounds containing active methylene groups. It is believed that the reaction proceeds via intermediary of the enaminoketone system. The sulfone derivatives were synthesized by oxidation of 4H-1,4-benzothiazines using 30% hydrogen peroxide in glacial acetic acid. Benzothiazines were used as bases to prepare ribofuranosides by treatment with a sugar derivative (ß-D-ribofuranosyl-1-acetate-2,3,5-tribenzoate). The pharmacological importance of the synthesized compounds was evaluated by their, antimicrobial properties against various bacterial strains and fungal species. The structures of the compounds have been confirmed by spectral and chemical analysis.

Muscarinic acetylcholine receptors: novel opportunities for drug development.

The muscarinic acetylcholine receptors are a subfamily of G protein-coupled receptors that regulate numerous fundamental functions of the central and peripheral nervous system. The past few years have witnessed unprecedented new insights into muscarinic receptor physiology, pharmacology and structure. These advances include the first structural views of muscarinic receptors in both inactive and active conformations, as well as a better understanding of the molecular underpinnings of muscarinic receptor regulation by allosteric modulators. These recent findings should facilitate the development of new muscarinic receptor subtype-selective ligands that could prove to be useful for the treatment of many severe pathophysiological conditions.

A novel experimental strategy to assess the metabolic effects of selective activation of a G(q)-coupled receptor in hepatocytes in vivo.

Increased hepatic glucose production is a key pathophysiological feature of type 2 diabetes. Like all other cell types, hepatocytes express many G protein-coupled receptors (GPCRs) that are linked to different functional classes of heterotrimeric G proteins. The important physiological functions mediated by G(s)-coupled hepatic glucagon receptors are well-documented. In contrast, little is known about the in vivo physiological roles of hepatocyte GPCRs that are linked to G proteins of the G(q) family. To address this issue, we established a transgenic mouse line (Hep-Rq mice) that expressed a G(q)-linked designer receptor (Rq) in a hepatocyte-selective fashion. Importantly, Rq could no longer bind endogenous ligands but could be selectively activated by a synthetic drug, clozapine-N-oxide. Clozapine-N-oxide treatment of Hep-Rq mice enabled us to determine the metabolic consequences caused by selective activation of a G(q)-coupled GPCR in hepatocytes in vivo. We found that acute Rq activation in vivo led to pronounced increases in blood glucose levels, resulting from increased rates of glycogen breakdown and gluconeogenesis. We also demonstrated that the expression of the V(1b) vasopressin receptor, a G(q)-coupled receptor expressed by hepatocytes, was drastically increased in livers of ob/ob mice, a mouse model of diabetes. Strikingly, treatment of ob/ob mice with a selective V(1b) receptor antagonist led to reduced glucose excursions in a pyruvate challenge test. Taken together, these findings underscore the importance of G(q)-coupled receptors in regulating hepatic glucose fluxes and suggest novel receptor targets for the treatment of type 2 diabetes.

Critical metabolic roles of ß-cell M3 muscarinic acetylcholine receptors.

Muscarinic acetylcholine (ACh) receptors (mAChRs; M(1)-M(5)) regulate the activity of an extraordinarily large number of important physiological processes. We and others previously demonstrated that pancreatic ß-cells are endowed with M(3) mAChRs which are linked to G proteins of the G(q) family. The activation of these receptors by ACh or other muscarinic agonists leads to the augmentation of glucose-induced insulin release via multiple mechanisms. Interestingly, in humans, ACh acting on human ß-cell mAChRs is released from adjacent α-cells which express both choline acetyltransferase (ChAT) and the vesicular acetylcholine transporter (vAChT), indicative of the presence of a non-neuronal cholinergic system in human pancreatic islets. In order to shed light on the physiological roles of ß-cell M(3) receptors, we recently generated and analyzed various mutant mouse models. Specifically, we carried out studies with mice which overexpressed M(3) receptors or mutant M(3) receptors in pancreatic ß-cells or which selectively lacked M(3) receptors or M(3)-receptor-associated proteins in pancreatic ß-cells. Our findings indicate that ß-cell M(3) receptors play a key role in maintaining proper insulin release and whole body glucose homeostasis and that strategies aimed at enhancing signaling through ß-cell M(3) receptors may prove useful to improve ß-cell function for the treatment of type 2 diabetes (T2D).

Novel insights into the function of ß-cell M3 muscarinic acetylcholine receptors: therapeutic implications.

Impaired function of pancreatic ß-cells is one of the hallmarks of type 2 diabetes. ß-cell function is regulated by the activity of many hormones and neurotransmitters, which bind to specific cell surface receptors. The M(3) muscarinic acetylcholine receptor (M3R) belongs to the superfamily of G protein-coupled receptors and, following ligand dependent activation, selectively activates G proteins of the G(q/11) family. Recent studies with M3R mutant mice strongly suggest that ß-cell M3Rs play a central role in promoting insulin release and maintaining correct glucose homeostasis. In this review, we highlight recent studies indicating that ß-cell M3Rs and components of downstream signaling pathways might represent promising new targets for the treatment of type 2 diabetes.

Beneficial metabolic effects caused by persistent activation of beta-cell M3 muscarinic acetylcholine receptors in transgenic mice.

Previous studies have shown that ß-cell M(3) muscarinic acetylcholine receptors (M3Rs) play a key role in maintaining blood glucose homeostasis by enhancing glucose-dependent insulin release. In this study, we tested the hypothesis that long-term, persistent activation of ß-cell M3Rs can improve glucose tolerance and ameliorate the metabolic deficits associated with the consumption of a high-fat diet. To achieve the selective and persistent activation of ß-cell M3Rs in vivo, we generated transgenic mice that expressed the Q490L mutant M3R in their pancreatic ß-cells (ß-M3-Q490L Tg mice). The Q490L point mutation is known to render the M3R constitutively active. The metabolic phenotypes of the transgenic mice were examined in several in vitro and in vivo metabolic tests. In the presence of 15 mm glucose and the absence of M3R ligands, isolated perifused islets prepared from ß-M3-Q490L Tg mice released considerably more insulin than wild-type control islets. This effect could be completely blocked by incubation of the transgenic islets with atropine (10 µm), an inverse muscarinic agonist, indicating that the Q490L mutant M3R exhibited ligand-independent signaling (constitutive activity) in mouse ß-cells. In vivo studies showed that ß-M3-Q490L Tg mice displayed greatly improved glucose tolerance and increased serum insulin levels as well as resistance to diet-induced glucose intolerance and hyperglycemia. These results suggest that chronic activation of ß-cell M3Rs may represent a useful approach to boost insulin output in the long-term treatment of type 2 diabetes.

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.