Small Molecule Optical Probes for Detection of H2S in Water Samples: A Review.

Hydrogen sulfide (H2S) is closely linked to not only environmental hazards, but also it affects human health due to its toxic nature and the exposure risks associated with several occupational settings. Therefore, detection of this pollutant in water sources has garnered immense importance in the analytical research arena. Several research groups have devoted great efforts to explore the selective as well as sensitive methods to detect H2S concentrations in water. Recent studies describe different strategies for sensing this ubiquitous gas in real-life water samples. Though many of the designed and developed H2S detection approaches based on the use of organic small molecules facilitate qualitative/quantitative detection of the toxic contaminant in water, optical detection has been acknowledged as one of the best, attributed to the simple, highly sensitive, selective, and good repeatability features of the technique. Therefore, this review is an attempt to offer a general perspective of easy-to-use and fast response optical detection techniques for H2S, fluorimetry and colorimetry, over a wide variety of other instrumental platforms. The review affords a concise summary of the various design strategies adopted by various researchers in constructing small organic molecules as H2S sensors and offers insight into their mechanistic pathways. Moreover, it collates the salient aspects of optical detection techniques and highlights the future scope for prospective exploration in this field based on the limitations of the existing H2S probes.

A study of sexually transmitted infections and reproductive tract infections among the bar girls in Mumbai, India.

Introduction: There is a continuous and increasing trend in sexually transmitted infections (STIs) worldwide. There are severe consequences of STI especially among women practicing high-risk behavior. Control of STI among women practicing high-risk behavior is an important strategy to reduce HIV transmission. In the present study, STIs and reproductive tract infections (RTI) were assessed among bar girls in an urban metro city. Methods: The cross-sectional study was conducted among bar girls residing in an urban settlement from May 2018 to December 2018. 195 bar girls randomly selected from among the registered bar girls for HIV prevention services with a local NGO and willing to participate were enrolled after obtaining informed consent. They were interviewed in person and clinical examination was conducted for assessing symptoms and signs of STI and RTI. Results: The median age of women was 26 years. 45% women reported sexual activity 3-6 times/week. 49.7% women reported symptom of vaginal discharge in the preceding 3 months, 52.3% women reported pain in lower abdomen, 10.2% women complained of genital ulcer disease, 67.2% women reported itching in the genital area, and 36.4% women had complaint of dyspareunia. 90.2% women in the present study had at least one symptom of STI/RTI. Conclusion: High-risk sexual behavior of bar girls with their regular clients puts them at a higher risk of acquiring HIV/AIDS. Focused efforts need to be put in to encourage reporting of symptoms for early diagnosis and management of STI/RTIs.

Replication Protein A Enhances Kinetics of Uracil DNA Glycosylase on ssDNA and Across DNA Junctions: Explored with a DNA Repair Complex Produced with SpyCatcher/SpyTag Ligation.

DNA repair proteins participate in extensive protein-protein interactions that promote the formation of DNA repair complexes. To understand how complex formation affects protein function during base excision repair, we used SpyCatcher/SpyTag ligation to produce a covalent complex between human uracil DNA glycosylase (UNG2) and replication protein A (RPA). Our covalent "RPA-Spy-UNG2" complex could identify and excise uracil bases in duplex areas next to ssDNA-dsDNA junctions slightly faster than the wild-type proteins, but this was highly dependent on DNA structure, as the turnover of the RPA-Spy-UNG2 complex slowed at DNA junctions where RPA tightly engaged long ssDNA sections. Conversely, the enzymes preferred uracil sites in ssDNA where RPA strongly enhanced uracil excision by UNG2 regardless of ssDNA length. Finally, RPA was found to promote UNG2 excision of two uracil sites positioned across a ssDNA-dsDNA junction, and dissociation of UNG2 from RPA enhanced this process. Our approach of ligating together RPA and UNG2 to reveal how complex formation affects enzyme function could be applied to examine other assemblies of DNA repair proteins.

Assay design for analysis of human uracil DNA glycosylase.

Human uracil DNA glycosylase (UNG2) is an enzyme whose primary function is to remove uracil bases from genomic DNA. UNG2 activity is critical when uracil bases are elevated in DNA during class switch recombination and somatic hypermutation, and additionally, UNG2 affects the efficacy of thymidylate synthase inhibitors that increase genomic uracil levels. Here, we summarize the enzymatic properties of UNG2 and its mitochondrial analog UNG1. To facilitate studies on the activity of these highly conserved proteins, we discuss three fluorescence-based enzyme assays that have informed much of our understanding on UNG2 function. The assays use synthetic DNA oligonucleotide substrates with uracil bases incorporated in the DNA, and the substrates can be single-stranded, double-stranded, or form other structures such as DNA hairpins or junctions. The fluorescence signal reporting uracil base excision by UNG2 is detected in different ways: (1) Excision of uracil from end-labeled oligonucleotides is measured by visualizing UNG2 reaction products with denaturing PAGE; (2) Uracil excision from dsDNA substrates is detected in solution by base pairing uracil with 2-aminopurine, whose intrinsic fluorescence is enhanced upon uracil excision; or (3) UNG2 excision of uracil from a hairpin molecular beacon substrate changes the structure of the substrate and turns on fluorescence by relieving a fluorescence quench. In addition to their utility in characterizing UNG2 properties, these assays are being adapted to discover inhibitors of the enzyme and to determine how protein-protein interactions affect UNG2 function.

KAT2A complexes ATAC and SAGA play unique roles in cell maintenance and identity in hematopoiesis and leukemia.

Epigenetic histone modifiers are key regulators of cell fate decisions in normal and malignant hematopoiesis. Their enzymatic activities are of particular significance as putative therapeutic targets in leukemia. In contrast, less is known about the contextual role in which those enzymatic activities are exercised and specifically how different macromolecular complexes configure the same enzymatic activity with distinct molecular and cellular consequences. We focus on KAT2A, a lysine acetyltransferase responsible for histone H3 lysine 9 acetylation, which we recently identified as a dependence in acute myeloid leukemia stem cells and that participates in 2 distinct macromolecular complexes: Ada two-A-containing (ATAC) and Spt-Ada-Gcn5-Acetyltransferase (SAGA). Through analysis of human cord blood hematopoietic stem cells and progenitors, and of myeloid leukemia cells, we identify unique respective contributions of the ATAC complex to regulation of biosynthetic activity in undifferentiated self-renewing cells and of the SAGA complex to stabilization or correct progression of cell type-specific programs with putative preservation of cell identity. Cell type and stage-specific dependencies on ATAC and SAGA-regulated programs explain multilevel KAT2A requirements in leukemia and in erythroid lineage specification and development. Importantly, they set a paradigm against which lineage specification and identity can be explored across developmental stem cell systems.

Diabetes Care in India: A Descriptive Study.

CONTEXT: Diabetes is a chronic illness that requires continuing medical care and patient self-management education to prevent and reduce the risk of long-term complications. It requires an array of investigations to provide an accurate picture of the condition and its management accordingly by a qualified doctor. AIMS: This study was conducted to understand the treatment received by type 2 diabetes (T2DM) patients from various categories of health care professionals and awareness about diabetes reversal by lifestyle modification and prevention of complications. SETTINGS AND DESIGN: This was a community-based cross-sectional study. SUBJECTS AND METHODS: The link of the semi-structured questionnaire in Google form with e-consent was sent to all members in the selected groups of "World free of obesity and diabetes" campaign on their personal WhatsApp account. STATISTICAL ANALYSIS USED: A total of 3082 participants were included, and the data obtained were analyzed using SPSS v26. RESULTS: The mean age of the participants was 50.26 ± 9.78 years ranging from 18 to 81 years. A total of 35.8% of the study population was diabetic for 1-5 years. A total of 54.9% were started with antidiabetic medication on the same day of diagnosis. Only 1.5% of the patients had complete investigation profile for T2DM, 50.2% of the patients were briefed about hypoglycemia, and only 15.8% of the patients were checked for retinopathy. CONCLUSIONS: Most doctors, qualified as well as nonqualified, did not follow the standard guidelines for diagnosis, treatment, and patient education regarding T2DM; therefore, it is necessary to train all medical practitioners regarding these guidelines. Diabetes reversal by lifestyle modification must be prescribed as the first line of treatment in patients with T2DM.

Loss of Kat2a enhances transcriptional noise and depletes acute myeloid leukemia stem-like cells.

Acute Myeloid Leukemia (AML) is an aggressive hematological malignancy with abnormal progenitor self-renewal and defective white blood cell differentiation. Its pathogenesis comprises subversion of transcriptional regulation, through mutation and by hijacking normal chromatin regulation. Kat2a is a histone acetyltransferase central to promoter activity, that we recently associated with stability of pluripotency networks, and identified as a genetic vulnerability in AML. Through combined chromatin profiling and single-cell transcriptomics of a conditional knockout mouse, we demonstrate that Kat2a contributes to leukemia propagation through preservation of leukemia stem-like cells. Kat2a loss impacts transcription factor binding and reduces transcriptional burst frequency in a subset of gene promoters, generating enhanced variability of transcript levels. Destabilization of target programs shifts leukemia cell fate out of self-renewal into differentiation. We propose that control of transcriptional variability is central to leukemia stem-like cell propagation, and establish a paradigm exploitable in different tumors and distinct stages of cancer evolution.

Less than 30% of patients with acute myeloid leukaemia ­ an aggressive cancer of the white blood cells ­ survive five years post-diagnosis. This disease disrupts the maturation of white blood cells, resulting in the accumulation of immature cells that multiply and survive but are incapable of completing their maturation process. Amongst these, a group of cancer cells known as leukemic stem cells is responsible for continually replenishing the leukaemia, thus perpetuating its growth. Cancers develop when cells in the body acquire changes or mutations to their genetic makeup. The mutations that lead to acute myeloid leukaemia often affect the activity of genes known as epigenetic regulators. These genes regulate which proteins and other molecules cells make by controlling the way in which cells 'read' their genetic instructions. The epigenetic regulator Kat2a is thought to 'tune' the frequency at which cells read their genetic instructions. This tuning mechanism decreases random fluctuations in the execution of the instructions cells receive to make proteins and other molecules. In turn, this helps to ensure that individual cells of the same type behave in a similar way, for example by keeping leukaemia cells in an immature state. Here, Domingues, Kulkarni et al. investigated whether interfering with Kat2a can make acute myeloid leukaemia less aggressive by allowing the immature white blood cells to mature. Domingues, Kulkarni et al. genetically engineered mice to remove Kat2a from blood cells on demand and then inserted a mutation that causes acute myeloid leukaemia. The experiments showed that the loss of Kat2a delayed the development of leukaemia in the mice and progressively depleted leukaemia stem cells, causing the disease to become less aggressive. The results also showed that loss of Kat2a caused more fluctuations in how the white blood cells read their genetic code, which resulted in more variability in the molecules they produced and increased the tendency of the cells to mature. These findings establish that loss of Kat2a causes leukaemia stem cells to mature and stop multiplying by untuning the frequency at which the cells read their genetic instructions. In the future, it may be possible to develop drugs that target human KAT2A to treat acute myeloid leukaemia.

miRNA-dependent regulation of STIM1 expression in breast cancer.

Store-operated Ca2+ entry (SOCE) has been shown to be important for breast cancer metastasis in xenograft mouse models. The ER Ca2+ sensor STIM1 and Orai plasma membrane Ca2+ channels molecularly mediate SOCE. Here we investigate the role of the microRNA machinery in regulating STIM1 expression. We show that STIM1 expression is regulated post-transcriptionally by the miRNA machinery and identify miR-223 and miR-150 as regulators of STIM1 expression in the luminal non-aggressive MCF7 breast cancer cell line. In contrast, STIM1 expression in the more aggressive basal triple-negative MDA-MB-231 cell line is not significantly modulated by a single miRNA species but is rather upregulated due to inhibition of the miRNA machinery through downregulation of Ago2. Consistently, overexpression of Ago2 results in decreased STIM1 protein levels in MDA-MB-231 cells. Clinically, STIM1 and Ago2 expression levels do not correlate with breast cancer progression, however in the basal subtype high STIM1 expression is associated with poorer survival. Our findings show that STIM1 expression is differentially regulated by the miRNA machinery in different cell types and argue for a role for this regulation in breast cancer.

Remodeling of ER-plasma membrane contact sites but not STIM1 phosphorylation inhibits Ca2+ influx in mitosis.

Store-operated Ca2+ entry (SOCE), mediated by the endoplasmic reticulum (ER) Ca2+ sensor stromal interaction molecule 1 (STIM1) and the plasma membrane (PM) channel Orai1, is inhibited during mitosis. STIM1 phosphorylation has been suggested to mediate this inhibition, but it is unclear whether additional pathways are involved. Here, we demonstrate using various approaches, including a nonphosphorylatable STIM1 knock-in mouse, that STIM1 phosphorylation is not required for SOCE inhibition in mitosis. Rather, multiple pathways converge to inhibit Ca2+ influx in mitosis. STIM1 interacts with the cochaperone BAG3 and localizes to autophagosomes in mitosis, and STIM1 protein levels are reduced. The density of ER-PM contact sites (CSs) is also dramatically reduced in mitosis, thus physically preventing STIM1 and Orai1 from interacting to activate SOCE. Our findings provide insights into ER-PM CS remodeling during mitosis and a mechanistic explanation of the inhibition of Ca2+ influx that is required for cell cycle progression.

Histone Acetyltransferase KAT2A Stabilizes Pluripotency with Control of Transcriptional Heterogeneity.

Cell fate transitions in mammalian stem cell systems have often been associated with transcriptional heterogeneity; however, existing data have failed to establish a functional or mechanistic link between the two phenomena. Experiments in unicellular organisms support the notion that transcriptional heterogeneity can be used to facilitate adaptability to environmental changes and have identified conserved chromatin-associated factors that modulate levels of transcriptional noise. Herein, we show destabilization of pluripotency-associated gene regulatory networks through increased transcriptional heterogeneity of mouse embryonic stem cells in which paradigmatic histone acetyl-transferase, and candidate noise modulator, Kat2a (yeast orthologue Gcn5), have been inhibited. Functionally, network destabilization associates with reduced pluripotency and accelerated mesendodermal differentiation, with increased probability of transitions into lineage commitment. Thus, we show evidence of a relationship between transcriptional heterogeneity and cell fate transitions through manipulation of the histone acetylation landscape of mouse embryonic stem cells, suggesting a general principle that could be exploited in other normal and malignant stem cell fate transitions. Stem Cells 2018;36:1828-11.

Fatal Iron Toxicity in an Adult: Clinical Profile and Review.

Acute iron toxicity is usually seen in children with accidental ingestion of iron-containing syrups. However, the literature on acute iron toxicity with suicidal intent in adults is scant. We report an instance wherein an adult committed suicide by ingestion of multiple iron tablets. Delay in treatment was there due to misdiagnosis of the intoxicating agent. She developed fulminant hepatic failure with rapid clinical deterioration. Despite aggressive supportive management, the patient succumbed to the toxic doses of iron. Clinical course and postmortem features are discussed with a review of the literature.

Effects of Hyperglycemia on Vascular Smooth Muscle Ca2+ Signaling.

Diabetes is a complex disease that is characterized with hyperglycemia, dyslipidemia, and insulin resistance. These pathologies are associated with significant cardiovascular implications that affect both the macro- and microvasculature. It is therefore important to understand the effects of various pathologies associated with diabetes on the vasculature. Here we directly test the effects of hyperglycemia on vascular smooth muscle (VSM) Ca2+ signaling in an isolated in vitro system using the A7r5 rat aortic cell line as a model. We find that prolonged exposure of A7r5 cells to hyperglycemia (weeks) is associated with changes to Ca2+ signaling, including most prominently an inhibition of the passive ER Ca2+ leak and the sarcoplasmic reticulum Ca2+-ATPase (SERCA). To translate these findings to the in vivo condition, we used primary VSM cells from normal and diabetic subjects and find that only the inhibition of the ER Ca2+ leaks replicates in cells from diabetic donors. These results show that prolonged hyperglycemia in isolation alters the Ca2+ signaling machinery in VSM cells. However, these alterations are not readily translatable to the whole organism situation where alterations to the Ca2+ signaling machinery are different.

Synthesis of Naked-eye Detectable Fluorescent 2H-chromen-2-One 2, 6-Dicyanoanilines: Effect of Substituents and pH on Its Luminous Behavior.

A variety of new coumarin derivatives containing C-4 bridged 2,6-dicyanoanilines (4a-4d) were synthesized via multicomponent one pot approach. These novel sensors were characterized by spectral analysis and a series of pH sensing fluorescence studies were performed, the results indicating that the sensors are highly selective and more effective at various pH. The fluorescence colour changes at different pH could be directly detected by naked eyes.

3,4-Dihydropyrimidinone-coumarin analogues as a new class of selective agent against S. aureus: Synthesis, biological evaluation and molecular modelling study.

Bacterial infections are increasingly difficult to combat as bacteria evolve resistance to antibiotic drugs and have severely compromised the arsenal of antibiotic drugs. On the other hand matrix metalloproteinases (MMPs) play a fundamental role in inflammation and extracellular matrix degradation in physiological and pathological conditions. In search of potent antibiotic, taking coumarin and dihydropyrimidinone as lead compound, a green, eco-friendly and efficient protocol has been developed and synthesized the dihydropyrimidin-2(1H)-one/thione derivatives of coumarin 3/4 from substituted 4-formylcoumarins 2 and ethylacetoacetate using urea/thiourea in the presence of catalytic amount of ceric ammonium nitrate is reported. All the synthesized compounds were evaluated for their antibacterial activity against four bacterial strains by broth dilution method. The tested compounds have exhibited promising in vitro potency with low MIC values against the drug susceptive S. aureus strain with low MIC values ranging from 0.2 to 6.25µg/mL. The in vivo anti-inflammatory potency of 3a-e and 4a-e by gelatin zymography is comparable to that of tetracycline. Molecular docking study performed for all the synthesized compounds with S. aureus DNA gyrase and results obtained were quite promising.

Greener synthesis of indolizine analogues using water as a base and solvent: study for larvicidal activity against Anopheles arabiensis.

Greener synthesis of a series of novel indolizine analogues have been achieved by the cyclization of aromatic cycloimmonium ylides with electron-deficient alkynes in the presence of water as the base and solvent at 80 °C. Yield of the title compounds was good and reactions performed were eco-friendly. The structures of these newly synthesized compounds have been confirmed by spectroscopic techniques such as FTIR, NMR, LC-MS, and elemental analysis. Characterized title compounds were evaluated for larvicidal activity against Anopheles arabiensis by standard WHO larvicidal assay using Temefos as standard at 4 µg/mL. Title compounds 2e, 2f, and 2g emerged as promising larvicidal agents.

Release from Xenopus oocyte prophase I meiotic arrest is independent of a decrease in cAMP levels or PKA activity.

Vertebrate oocytes arrest at prophase of meiosis I as a result of high levels of cyclic adenosine monophosphate (cAMP) and protein kinase A (PKA) activity. In Xenopus, progesterone is believed to release meiotic arrest by inhibiting adenylate cyclase, lowering cAMP levels and repressing PKA. However, the exact timing and extent of the cAMP decrease is unclear, with conflicting reports in the literature. Using various in vivo reporters for cAMP and PKA at the single-cell level in real time, we fail to detect any significant changes in cAMP or PKA in response to progesterone. More interestingly, there was no correlation between the levels of PKA inhibition and the release of meiotic arrest. Furthermore, we devised conditions whereby meiotic arrest could be released in the presence of sustained high levels of cAMP. Consistently, lowering endogenous cAMP levels by >65% for prolonged time periods failed to induce spontaneous maturation. These results argue that the release of oocyte meiotic arrest in Xenopus is independent of a reduction in either cAMP levels or PKA activity, but rather proceeds through a parallel cAMP/PKA-independent pathway.

Anti-inflammatory and antioxidant effect of ginger in tuberculosis.

BACKGROUND: Tuberculosis (TB) has reemerged to become the world's leading cause of death from a single infectious agent. Inflammatory cytokines play an important role during the course of the disease and may be responsible for tissue damage by lipid peroxidation. The study was aimed to explore the anti-inflammatory and antioxidant effect of ginger in pulmonary TB patients. METHODS: A total of 69 pulmonary TB patients participated in a randomized and placebo-controlled study. The intervention group received 3 g of ginger extract daily for 1 month and placebo group was supplemented with starch capsule. Participants of both groups were taking standard antitubercular treatment during the study. The concentrations of tumor necrosis factor (TNF) alpha, ferritin and malondialdehyde (MDA) in blood samples were analyzed before and after the intervention by using enzyme-linked immunosorbent assay for TNF alpha and ferritin and spectrophotometry for MDA. RESULTS: Ginger supplementation significantly reduced the levels of TNF alpha, ferritin and MDA in ginger supplemented group in comparison to baseline. Ginger supplementation with antitubercular treatment significantly lowered TNF alpha, ferritin and MDA concentrations in comparison to control group. CONCLUSIONS: Ginger was found to be effective as an anti-inflammatory and antioxidant supplement along with anti-TB therapy as it possesses strong free radical scavenging property.

Prospects of poisoning - a multi facet study.

Aim of the study is to find out demographic profile, clinical characteristics and analysis of poison in clinical set up. The study carried out in Sri Aurobindo Medical College and PG Institute Indore, Madhya Pradesh. Total 75 cases of poisoning were studied for demographic profile, vitals (BP, pulse, heart rate, pupils, etc.), clinical features (such as vomiting, salivation, consciousness, etc.), type of poison and its analysis. Results: Poisoning was more common in cases between 15 and 25 years of age, in males than in females and in Hindu religion. Poisoning cases were predominantly from rural areas and in married people. Majority of cases were discharged after proper treatment and counseling. Altered vitals and clinical features were found in most of the cases. Organophosphate and aluminum phosphide compound were evaluated in most of the cases. Conclusions: Preventive measures should be applied through educating people, proper counseling, promoting poison information centers, and introducing separate toxicological units in hospitals.

Role for endocytosis of a constitutively active GPCR (GPR185) in releasing vertebrate oocyte meiotic arrest.

Vertebrate oocytes are naturally arrested at prophase of meiosis I for sustained periods of time before resuming meiosis in a process called oocyte maturation that prepares the egg for fertilization. Members of the constitutively active GPR3/6/12 family of G-protein coupled receptors represent important mediators of meiotic arrest. In the frog oocyte the GPR3/12 homolog GPRx (renamed GPR185) has been shown to sustain meiotic arrest by increasing intracellular cAMP levels through GαSßγ. Here we show that GPRx is enriched at the cell membrane (~80%), recycles through an endosomal compartment at steady state, and loses its ability to signal once trapped intracellularly. Progesterone-mediated oocyte maturation is associated with significant internalization of both endogenous and overexpressed GPRx. Furthermore, a GPRx mutant that does not internalize in response to progesterone is significantly more efficient than wild-type GPRx at blocking oocyte maturation. Collectively our results argue that internalization of the constitutively active GPRx is important to release oocyte meiotic arrest.