cis-Dihydroxylation by Synthetic Iron(III)-Peroxo Intermediates and Rieske Dioxygenases: Experimental and Theoretical Approaches Reveal the Key O-O Bond Activation Step.

Dioxygen (O2) activation by iron-containing enzymes and biomimetic compounds generates iron-oxygen intermediates, such as iron-superoxo, -peroxo, -hydroperoxo, and -oxo, that mediate oxidative reactions in biological and abiological systems. Among the iron-oxygen intermediates, iron(III)-peroxo species are less frequently implicated as active intermediates in oxidation reactions. In this study, we present the combined experimental and theoretical investigations on cis-dihydroxylation reactions mediated by synthetic mononuclear nonheme iron-peroxo intermediates, demonstrating the importance of supporting ligands and metal centers in activating the peroxo ligand toward the O-O bond homolysis for the cis-dihydroxylation reactions. We found a significant ring size effect of the TMC ligand in [FeIII(O2)(n-TMC)]+ (TMC = tetramethylated tetraazacycloalkane; n = 12, 13, and 14) on the cis-dihydroxylation reactivity order: [FeIII(O2)(12-TMC)]+ > [FeIII(O2)(13-TMC)]+ > [FeIII(O2)(14-TMC)]+. Additionally, we found that only [FeIII(O2)(n-TMC)]+, but not other metal-peroxo complexes such as [MIII(O2)(n-TMC)]+ (M = Mn, Co, and Ni), is reactive for the cis-dihydroxylation of olefins. Using density functional theory (DFT) calculations, we revealed that electron transfer from the Fe dxz orbital to the peroxo σ*(O-O) orbital facilitates the O-O bond homolysis, with the O-O bond cleavage barrier well correlated with the energy gap between the frontier molecular orbitals of dxz and σ*(O-O). Further computational studies showed that the reactivity of the synthetic [FeIII(O2)(12-TMC)]+ complex is comparable to that of Rieske dioxygenases in cis-dihydroxylation, providing compelling evidence of the potential involvement of Fe(III)-peroxo species in Rieske dioxygenases. Thus, the present results significantly advance our understanding of the cis-dihydroxylation mechanisms by Rieske dioxygenases and synthetic nonheme iron-peroxo models.

Electronic Structure and Reactivity of Mononuclear Nonheme Iron-Peroxo Complexes as a Biomimetic Model of Rieske Oxygenases: Ring Size Effects of Macrocyclic Ligands.

We report the macrocyclic ring size-electronic structure-electrophilic reactivity correlation of mononuclear nonheme iron(III)-peroxo complexes bearing N-tetramethylated cyclam analogues (n-TMC), [FeIII(O2)(12-TMC)]+ (1), [FeIII(O2)(13-TMC)]+ (2), and [FeIII(O2)(14-TMC)]+ (3), as a model study of Rieske oxygenases. The Fe(III)-peroxo complexes show the same δ and pseudo-σ bonds between iron and the peroxo ligand. However, the strength of these interactions varies depending on the ring size of the n-TMC ligands; the overall Fe-O bond strength and the strength of the Fe-O2 δ bond increase gradually as the ring size of the n-TMC ligands becomes smaller, such as from 14-TMC to 13-TMC to 12-TMC. MCD spectroscopy plays a key role in assigning the characteristic low-energy δ → δ* LMCT band, which provides direct insight into the strength of the Fe-O2 δ bond and which, in turn, is correlated with the superoxo character of the iron-peroxo group. In oxidation reactions, reactivities of 1-3 toward hydrocarbon C-H bond activation are compared, revealing the reactivity order of 1 > 2 > 3; the [FeIII(O2)(n-TMC)]+ complex with a smaller n-TMC ring size, 12-TMC, is much more reactive than that with a larger n-TMC ring size, 14-TMC. DFT analysis shows that the Fe(III)-peroxo complex is not reactive toward C-H bonds, but it is the end-on Fe(II)-superoxo valence tautomer that is responsible for the observed reactivity. The hydrogen atom abstraction (HAA) reactivity of these intermediates is correlated with the overall donicity of the n-TMC ligand, which modulates the energy of the singly occupied π* superoxo frontier orbital that serves as the electron acceptor in the HAA reaction. The implications of these results for the mechanism of Rieske oxygenases are further discussed.

Synthesis of indol-3-yl-benzofurans and carbazoles via Cu(OTf)2-catalyzed [3 + 2] and [4 + 2] cycloaddition.

An efficient Cu(OTf)2-catalyzed [3 + 2] cycloaddition of indole-3-acrylate with p-benzoquinone has been developed to construct two distinct indole-tethered benzofuran scaffolds, offering the first-ever selective access to these scaffolds. Moreover, the [4 + 2] cycloaddition reaction of indole-3-acrylate with vinyl ketone derivatives was used to synthesize carbazoles in a one-pot manner. The disclosed strategies provided a series of selective transformations under low-catalyst loading, with a broad substrate scope featuring diverse applicability and practical simplicity of the developed protocol with easily available substrates.

Seeing the cis-Dihydroxylating Intermediate: A Mononuclear Nonheme Iron-Peroxo Complex in cis-Dihydroxylation Reactions Modeling Rieske Dioxygenases.

The nature of reactive intermediates and the mechanism of the cis-dihydroxylation of arenes and olefins by Rieske dioxygenases and synthetic nonheme iron catalysts have been the topic of intense research over the past several decades. In this study, we report that a spectroscopically well characterized mononuclear nonheme iron(III)-peroxo complex reacts with olefins and naphthalene derivatives, yielding iron(III) cycloadducts that are isolated and characterized structurally and spectroscopically. Kinetics and product analysis reveal that the nonheme iron(III)-peroxo complex is a nucleophile that reacts with olefins and naphthalenes to yield cis-diol products. The present study reports the first example of the cis-dihydroxylation of substrates by a nonheme iron(III)-peroxo complex that yields cis-diol products.

Surface-Modified Lyotropic Crystalline Nanoconstructs Bearing Doxorubicin and Buparvaquone Target Sigma Receptors through pH-Sensitive Charge Conversion to Improve Breast Cancer Therapy.

In the current study, we aimed to develop lyotropic crystalline nanoconstructs (LCNs) based on poly(l-glutamic acid) (PLG) with a two-tier strategy. The first objective was to confer pH-responsive charge conversion properties to facilitate the delivery of both doxorubicin (DOX) and buparvaquone (BPQ) in combination (B + D@LCNs) to harness their synergistic effects. The second goal was to achieve targeted delivery to sigma receptors within the tumor tissues. To achieve this, we designed a pH-responsive charge conversion system using a polymer consisting of poly(ethylenimine), poly(l-lysine), and poly(l-glutamic acid) (PLG), which was then covalently coupled with methoxybenzamide (MBA) for potential sigma receptor targeting. The resulting B + D@LCNs were further modified by surface functionalization with PLG-MBA to confer both sigma receptor targeting and pH-responsive charge conversion properties. Our observations indicated that at physiological pH 7.4, P/B + D-MBA@LCNs exhibited a negative charge, while under acidic conditions (pH 5.5, characteristic of the tumor microenvironment), they acquired a positive charge. The particle size of P/B + D-MBA@LCNs was determined to be 168.23 ± 2.66 nm at pH 7.4 and 201.23 ± 1.46 nm at pH 5.5. The crystalline structure of the LCNs was confirmed through small-angle X-ray scattering (SAXS) diffraction patterns. Receptor-mediated endocytosis, facilitated by P/B + D-MBA@LCNs, was confirmed using confocal laser scanning microscopy and flow cytometry. The P/B + D-MBA@LCNs formulation demonstrated a higher rate of G2/M phase arrest (55.20%) compared to free B + D (37.50%) and induced mitochondrial depolarization (59.39%) to a greater extent than P/B + D@LCNs (45.66%). Pharmacokinetic analysis revealed significantly improved area under the curve (AUC) values for both DOX and BPQ when administered as P/B + D-MBA@LCNs, along with enhanced tumor localization. Tumor regression studies exhibited a substantial reduction in tumor size, with P/B + D-MBA@LCNs leading to 3.2- and 1.27-fold reductions compared to B + D and nontargeted P/B + D@LCNs groups, respectively. In summary, this two-tier strategy demonstrates substantial promise for the delivery of a drug combination through the prototype formulation. It offers a potential chemotherapeutic option by minimizing toxic effects on healthy cells while maximizing therapeutic efficacy.

Design, Synthesis, and Applications of a Vanadium Complex: An Effective Catalyst for the Direct Conversion of Alcohols and Aldehydes to Esters.

A novel bench-stable V-catalyst [(L2)VIVO](ClO4) was synthesized and characterized by X-ray diffraction (XRD) analysis and FT-IR, UV-visible, and EPR spectroscopies, which confirmed its excellent catalytic activity. In application, aldehydes are rapidly converted into their corresponding esters without additives in a one-pot manner using a newly developed catalyst [(L2)VIVO](ClO4) and H2O2 as a green oxidant. The developed method is compatible with a broad range of densely substituted aldehydes and allows for the facile preparation of aliphatic, aromatic, and heterocyclic esters, including esters derived from CD3OD, methanol, ethanol, iso-propanol, n-butanol, sec-butyl alcohol, and propargylic alcohol. Gratifyingly, numerous alcohols also directly converted to their corresponding esters in a one-pot manner. We disclose herein the direct conversion of two different functionalities (alcohols and aldehydes) into esters (33 examples) with satisfactory yields, showing the potential of the developed catalyst toward varied oxidative organic transformations in a one-pot manner.

The rapid construction and biological evaluation of densely substituted pyrrolo[1,2-a]indoles via a BF3·OEt2-assisted cascade approach.

Lewis-acid cascade reactions promoted by BF3·OEt2 are reported for the synthesis of highly substituted pyrrolo[1,2-a]indoles and congeners of benzofuro[2,3-b]indoles. These reactions are highly regio- and diastereoselective towards generating up to five contiguous stereogenic centers, including two vicinal quaternary centers. Furthermore, an established cascade approach and the mechanism proposed herein are well supported by quantum chemistry calculations. In addition, a self-dimerization intermediate was trapped and isolated to establish a strategy for potential access to both pyrrolo and benzo indole derivatives, leaving sufficient freedom for broadening. Furthermore, in-silico molecular docking and all atomistic molecular dynamic (MD) simulation analysis suggests that the synthesized pyrrolo[1,2-a]indole derivatives stably bind at the active site of the mycobacterial secreted tyrosine phosphatase B (MptpB) enzyme, an emerging anti-mycobacterial drug target. Deep learning-based affinity predictions and MMPBGBSA-based energy calculations of the docked poses are presented herein.

Delayed Compression Paralysis Following an Iliopsoas Hematoma 30 Days After Saw-Scaled Viper (Echis carinatus sochureki) Envenoming: A Case Report.

Snakebite envenoming is a neglected tropical disease disproportionately affecting the rural and marginalized population in low-middle-income countries. The saw-scaled viper (Echis carinatus) is a clinically important snake that causes serious morbidity and mortality in the Indian subcontinent. Even though it is within the so-called big-four snakes against which polyvalent antivenom is available throughout India, reports of antivenom ineffectiveness are emerging in saw-scaled viper envenoming, especially around Jodhpur, Rajasthan, India. This case report highlights a patient with saw-scaled viper envenoming with an ineffective antivenom response complicated by acute kidney injury as well as local and systemic bleeding complications, which subsequently resulted in a pelvic hematoma that compressed the lumbosacral nerves, causing lower-limb weakness and sensory deficits. He was successfully managed with hematoma aspiration and supportive care. This case brings into focus the challenges of managing saw-scaled viper envenoming in this region with antivenom ineffectiveness, resulting in delayed and significant coagulopathy and its complications leading to prolonged hospital stay and morbidity. Our report spotlights less emphasized aspects of long-term morbidity in snakebite survivors, such as loss of working days and productivity. We also highlight the need for an organized system of long-term follow-up of snakebite survivors to screen for possible complications and manage them early.

Investigating impact of CORDEX-based predicted climatic and LCM-based LULC scenarios on hydrologic response of a semi-gauged Indian catchment.

The present study aims at documenting the impact of different climate and land use change scenarios on runoff in the Kangsabati River basin. While the study relies on India Meteorological Department (IMD), National Oceanic and Atmospheric Administration's Physical Sciences Laboratory (NOAA-PSL), and a multi-model ensemble of six driving models from Coordinated Regional Downscaling Experiment-Regional Climate Models (CORDEX RCM) for climate data input, it depends on IDRISI Selva's Land Change Modeller (LCM) and Soil and Water Assessment Tool (SWAT) model to generate projected land use land change maps and simulate its streamflow response, respectively. A total of four land use and land cover (LULC) scenarios, representing four projected land use change, were modelled across three climatic scenarios, called Representative Concentration Pathways (RCPs). With runoff being predominantly impacted more by climate change than LULC, volumetric runoff is expected to be 12-46% higher than the baseline period of 1982-2017. Conversely, while surface runoff is expected to decrease by 4-28% in lower parts of the basin, it will increase by 2-39% in the rest of it, depending on the subtle alterations in land use and climatic variability.

Fut9 Deficiency Causes Abnormal Neural Development in the Mouse Cerebral Cortex and Retina.

α1,3-Fucosyltransferase 9 (Fut9) is responsible for the synthesis of Lewis X [LeX, Galß1-4(Fucα1-3)GlcNAc] carbohydrate epitope, a marker for pluripotent or multipotent tissue-specific stem cells. Although Fut9-deficient mice show anxiety-related behaviors, structural and cellular abnormalities in the brain remain to be investigated. In this study, using in situ hybridization and immunohistochemical techniques in combination, we clarified the spatiotemporal expression of Fut9, together with LeX, in the brain and retina. We found that Fut9-expressing cells are positive for Ctip2, a marker of neurons residing in layer V/VI, and TLE4, a marker of corticothalamic projection neurons (CThPNs) in layer VI, of the cortex. A birthdating analysis using 5-ethynyl-2'-deoxyuridine at embryonic day (E)11.5, 5-bromo-2'-deoxyuridine at E12.5, and in utero electroporation of a GFP expression plasmid at E14.5 revealed a reduction in the percentage of neurons produced at E11.5 in layer VI/subplate of the cortex and in the ganglion cell layer of the retina in P0 Fut9-/- mice. Furthermore, this reduction in layer VI/subplate neurons persisted into adulthood, leading to a reduction in the number of Ctip2strong/Satb2- excitatory neurons in layer V/VI of the adult Fut9-/- cortex. These results suggest that Fut9 plays significant roles in the differentiation, migration, and maturation of neural precursor cells in the cortex and retina.

CRISPR-Cas system: from diagnostic tool to potential antiviral treatment.

This mini review focuses on the diagnosis and treatment of virus diseases using Crisper-Cas technology. The present paper describes various strategies involved in diagnosing diseases using Crispr-Cas-based assays. Additionally, CRISPR-Cas systems offer great potential as new therapeutic tools for treating viral infections including HIV, Influenza, and SARS-CoV-2. There are several major challenges to be overcome before this technology can be applied routinely in clinical settings, such as finding a suitable delivery tool, toxicity, and immunogenicity, as well as off-target effects. This review also discusses ways to deal with the challenges associated with Crisper-Cas technology. KEY POINTS: • Crisper technology is being applied to diagnose infectious and non-infectious diseases. • A new generation of CRISPR-Cas-based assays has been developed which detect pathogens within minutes, providing rapid diagnosis of diseases. • Crispr-Cas tools can be used to combat viral infections, specifically HIV, influenza, and SARS-CoV-2.

Autochthonous buffalo-gut origin Pediococcus pentosaceus RM119 decreased diarrhea and enhanced gut health, immunity in neonatal Murrah calves.

This study assessed the effect of autochthonous probiotic Pediococcus pentosaceus RM119 on gut health, growth, and nutrient utilization in calves. Twelve buffalo calves (<15 d) were divided into two groups, control without probiotics, and probiotic group with P. pentosaceus RM119 @ 108 CFU/calf/d. The probiotic group showed a reduction (p < 0.05) in fecal score, diarrhea episodes and duration of diarrhea. The fecal pH, fecal ammonia was lower, whereas lactate was higher in probiotic group than control. There was a significant increase (p < 0.01) in the concentration of fecal acetate, propionate and butyrate levels in the probiotic supplemented group. The fecal lactobacilli and bifidobacterium were higher (p < 0.01), whereas, fecal coliform and clostridial count were lower (p < 0.01) in P. pentosaceus RM119 supplemented group. There was an improvement in reduced glutathione anti oxidant. Overall, buffalo-gut origin P. pentosaceus RM119 reduced the frequency and severity of diarrhea in neonatal buffalo calves and improved the gut health.

Evaluation and management of "low" anorectal malformation in male children: an observational study.

PURPOSE: ARM with perineal fistula has been traditionally defined as low ARM (LARM). This study was conducted to evaluate LARM in male patients with an emphasis on the role of various factors on the outcome and follow-up of them. MATERIALS AND METHODS: It was a retrospective cohort study. The clinical presentation, associated anomalies, and complications were assessed. The operative procedures included cutback anoplasty and others. The patients were followed in the outpatient department. The complications were assessed and managed accordingly. RESULTS: During the study period of 8 years, 301 patients were admitted. The complaints included absent or abnormal anal opening, abdominal distension, constipation, and peritonitis. Most of the children (n = 214) presented in the neonatal period. The most common clinical presentation was the perineal fistula. The most common associated anomaly was urologic. Fourteen patients were referred from other centers after complications. The most common problem in follow-up was constipation. CONCLUSION: LARM in male patients may have a diverse presentation. The associated anomalies need proper assessment. Awareness may avoid delayed presentation and unwanted complications. When managed by an expert, the condition can be effectively managed. Regular follow-up is important.

Effects of Spirulina (Arthrospira platensis) as a drinking water supplement during cyclical chronic heat stress on broiler chickens: Assessing algal composition, production, stress, health and immune-biochemical indices.

Spirulina, the blue green algae is considered to exhibit multifaceted benefits on both human health and animal production. Three hundred sixty day old unsexed broiler chicks of CARIBROVISHAL strain were assigned to five treatment groups each comprising nine replicates of 8 chicks. The experiment was carried out during the hot humid summer season (Mid-April to May) under deep litter rearing system with uniform managemental conditions. Birds were administered orally with Spirulina through drinking water in the morning (06:00-12:00 PM) on daily basis throughout the experimental period at 5, 10, 15 and 20 gL-1 concentration. Spirulina supplementation neither improved nor compromised production performance of broilers reared during hot climatic condition. Results based on one way analysis of variance indicated a significant effect on haemoglobin and total red blood cell count. Serum lipid content and transaminases were reduced, while serum protein concentration was higher (P < 0.01) in the groups administered with 15 and 20 gL-1 of Spirulina. The extent of imparting shank pigmentation was improved in all the supplemented groups. Cell mediated and humoral immunity against Phytoheamagglutunin-P and Newcastle disease vaccination respectively were maximized (P < 0.05) at 20 gL-1. These findings provide direct evidence of dose-related modulation of production, physiological and immunological attributes by Spirulina engendering its further investigation as a potential source of drinking water supplement for stress alleviation in broilers. From the results, it may concluded that Spirulina can be incorporated at 15 or 20 gL-1 for achieving optimal improvement of health and welfare attributes in broilers reared during hot summer without compromising production.

Effectiveness of early awake self proning strategy in non-intubated patients with COVID-19 hypoxemia: an open-labelled randomized clinical trial from Jodhpur, India.

Awake self-proning is being used widely as respiratory support in COVID-19 hypoxemia, in resource-limited settings. We aimed to investigate the effectiveness of early awake self-proning in preventing mortality and the need for intubation in adults with moderate COVID-19 hypoxemia. In this randomized clinical trial with inten-tion-to-treat analysis, we enrolled eligible adults with COVID-19 hypoxemia (SpO2 <94%), requiring supplemental oxygen via nasal prongs or facemask from a tertiary-care setting in Jodhpur, India between June 15 to December 24, 2020. Awake proning comprised of 4-hour cycles with prone position maintained 2 h per cycle. The control group did not maintain any specific position. All participants received standard care. The primary outcomes were 30-day mortal-ity and requirement for mechanical ventilation. Of 502 participants included, mean (SD) age was 59.7 (12.7) years with 124 women (24.6%); 257 were randomized to awake-proning, 245 to control group and all 502 were included for follow-up mortality analysis. Mortality at follow-up was 16.3% in the awake-prone and 15.1% in the control group [OR:1.10 (0.68-1.78), p=0.703). The requirement of mechanical ventilation was 10% in both groups (p=0.974). Survival time (in days) was not significantly different between the groups [Log-rank test, HR: 1.08 (95% CI, 0.70-1.68), p=0.726]. Likewise, time to intubation was comparable (Log-rank test, HR: 0.93 (95% CI, 0.56-1.70), p=0.974). Hence, awake self-proning did not improve survival or requirement of mechanical-ventilation in non-intubated patients with mild to moderate COVID-19 hypox-emia. Trial Registration: Clinical trial registry of India, ID: CTRI/2020/06/025804.   The trial is accessible from WHO's International Clinical Trials Registry Platform (ICTRP) at https://trialsearch.who.int ***************************************************************   *Appendix Authors list  Deepak Kumar1, Gopal Krishna Bohra1, Nishant Kumar Chauhan2, Nikhil Kothari3, Vijaya Lakshmi Nag4 Sanjeev Misra5  1Department of Internal Medicine; 2Department of Pulmonary Medicine; 3Department of Anaesthesiology and Critical Care; 4Department of Microbiology; 5Department of Surgical Oncology, All India Institute of Medical Sciences, Jodhpur, India.

Influences of maternal factors on the estimate of genetic parameters for goat feed efficiency traits.

The objective of the present study was to estimate the genetic parameters for the feed efficiency traits in Barbari goats. The data records of 9332 progenies born to 413 sires and 2580 dams were collected with respect to the average daily weight gain (ADG), i.e., ADG1 (birth to weaning), ADG2 (weaning to 6 months), ADG3 (6 to 12 months), as well as derived trait Kleiber ratio (KR), i.e., KR1 (ADG1/3MW0.75), KR2 (ADG2/6MW0.75), and KR3 (ADG3/12MW0.75). The data were corrected for fixed covariates like period of kidding, the season of birth, sex, type of birth, and parity. Univariate and multivariate animal models with an average information function of restricted maximum likelihood (REML) were used to estimate genetic factors for these traits. The best model was evaluated based on the likelihood ratio test. The direct heritability estimates were 0.21 ± 0.03, 0.17 ± 0.03, 0.23 ± 0.04, 0.22 ± 0.04, 0.16 ± 0.04, and 0.26 ± 0.04 for ADG1, ADG2, ADG3, KR1, KR2, and KR3, respectively. However, they were inflated due to high and negative estimates of covariance between direct animal and maternal genetic effects. Moderate estimates of heritability augur the scope for improvement for feed efficiency traits. The maternal genetic effects (m2) significantly contributed to 3-12% of the total phenotypic variance. The realized heritability of mass selection, which takes into account direct and maternal genetic variance together, shows a low to moderate estimate of genetic variance for ADG and KR. The genetic correlation ranged from - 0.48 ± 0.11 (ADG1-KR3) to 0.95 ± 0.00 (ADG1-KR1), phenotypic correlation ranged from - 0.28 ± 0.01 (ADG2-KR3) to 0.94 ± 0.01 (ADG1-KR1), maternal genetic correlation ranged from - 0.22 (KR2-KR3) to 0.96 (ADG1-KR1) and - 0.69 (ADG1-KR3) to 0.95 (ADG1-KR1) for the maternal permanent environment, respectively. Kids can be indirectly chosen for higher feed efficiency since ADG and their associated KR have substantial genetic correlations. It is suggested that the KR should be used as a selection criterion for Barbari goats for improving feed efficiency.

Dendritic cell engineered cTXN as new vaccine prospect against L. donovani.

Dendritic cells (DCs), as antigen-presenting cells, can reportedly be infected withLeishmaniaparasites and hence provide a better option to trigger T-cell primary immune responses and immunological memory. We consistently primed DCs during culture with purified recombinant cytosolic tryparedoxin (rcTXN) and then evaluated the vaccine prospect of presentation of rcTXN against VL in BALB/c mice. We reported earlier the immunogenic properties of cTXN antigen derived fromL. donovani when anti-cTXN antibody was detected in the sera of kala-azar patients. It was observed that cTXN antigen, when used as an immunogen with murine DCs acting as a vehicle, was able to induce complete protection against VL in an infected group of immunized mice. This vaccination triggered splenic macrophages to produce more IL-12 and GM-CSF, and restricted IL-10 release to a minimum in an immunized group of infected animals. Concomitant changes in T-cell responses against cTXN antigen were also noticed, which increased the release of protective cytokine-like IFN-γ under the influence of NF-κß in the indicated vaccinated group of animals. All cTXN-DCs-vaccinated BALB/c mice survived during the experimental period of 120 days. The results obtained in our study suggest that DCs primed with cTXN can be used as a vaccine prospect for the control of visceral leishmaniasis.

NOTCH Activation at the Hematovascular Mesoderm Stage Facilitates Efficient Generation of T Cells with High Proliferation Potential from Human Pluripotent Stem Cells.

Human pluripotent stem cells (hPSCs) offer the potential to serve as a versatile and scalable source of T cells for immunotherapies, which could be coupled with genetic engineering technologies to meet specific clinical needs. To improve T cell production from hPSCs, it is essential to identify cell subsets that are highly enriched in T cell progenitors and those stages of development at which NOTCH activation induces the most potent T cells. In this study, we evaluated the efficacy of T cell production from cell populations isolated at different stages of hematopoietic differentiation, including mesoderm, hemogenic endothelium (HE), and multipotent hematopoietic progenitors. We demonstrate that KDRhiCD31- hematovascular mesodermal progenitors (HVMPs) with definitive hematopoietic potential produce the highest numbers of T cells when cultured on OP9-DLL4 as compared with downstream progenitors, including HE and multipotent hematopoietic progenitors. In addition, we found that T cells generated from HVMPs have the capacity to expand for 6-7 wk in vitro, in comparison with T cells generated from HE and hematopoietic progenitors, which could only be expanded for 4-5 wk. Demonstrating the critical need of NOTCH activation at the HVMP stage of hematopoietic development to establish robust T cell production from hPSCs may aid in establishing protocols for the efficient off-the-shelf production and expansion of T cells for treating hematologic malignancies.

Regulatory safety pharmacology and toxicity assessments of a standardized stem extract of Cassia occidentalis Linn. in rodents.

Cassia occidentalis Linn (CO) is an annual/perennial plant having traditional uses in the treatments of ringworm, gastrointestinal ailments and piles, bone fracture, and wound healing. Previously, we confirmed the medicinal use of the stem extract (ethanolic) of CO (henceforth CSE) in fracture healing at 250 mg/kg dose in rats and described an osteogenic mode of action of four phytochemicals present in CSE. Here we studied CSE's preclinical safety and toxicity. CSE prepared as per regulations of Current Good Manufacturing Practice for human pharmaceuticals/phytopharmaceuticals and all studies were performed in rodents in a GLP-accredited facility. In acute dose toxicity as per New Drug and Clinical Trial Rules, 2019 (prior name schedule Y), in rats and mice and ten-day dose range-finding study in rats, CSE showed no mortality and no gross abnormality at 2500 mg/kg dose. Safety Pharmacology showed no adverse effect on central nervous system, cardiovascular system, and respiratory system at 2500 mg/kg dose. CSE was not mutagenic in the Ames test and did not cause clastogenicity assessed by in vivo bone marrow genotoxicity assay. By a sub chronic (90 days) repeated dose (as per OECD, 408 guideline) study in rats, the no-observed-adverse-effect-level was found to be 2500 mg/kg assessed by clinico-biochemistry and all organs histopathology. We conclude that CSE is safe up to 10X the dose required for its osteogenic effect.

The wandering ventriculoperitoneal shunt and the scope of its salvage.

PURPOSE: Distal shunt tube migration following ventriculoperitoneal (VP) shunt placement in children is mostly managed by an initial shunt diversion/removal and subsequent replacement. Lately, shunt salvage is being used as an alternative in certain conditions. We have focused on the situations where one can consider or disregard shunt salvage in such cases. METHOD: A retrospective study of children treated for distal shunt migration following VP shunt placement between January 2013 and December 2019. RESULT: Seventeen children were managed for over 7 years. These included cutaneous extrusions (n = 4), hollow viscus perforation (n = 6), inguinal hernias (n = 5), and umbilical extrusion (n = 2). The surgical treatment varied from a cutaneous wound closure (with a tube in situ), temporary external shunt diversion, and laparotomy with shunt reposition into the peritoneal cavity. Shunt salvage was possible in three cases, whereas in 2 cases even though shunt salvage was possible, it was not feasible due to a short residual shunt length. CONCLUSION: VP shunt salvage is possible in certain cases of distal shunt migration with a functional uninfected shunt. Small cutaneous extrusions can be covered by a local skin flap. Also, one should consider the residual intraperitoneal shunt length before its salvage in small children.