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Iron is an indispensable element for plant growth and defense and hence it is essential to improve the plant’s ability to accumulate iron. Besides, it is also an important aspect for human health. In view of this, we attempted to increase the iron content in banana cultivar Rasthali using MusaFer1 as a candidate gene. Initially, the expression of all five genes of the MusaFer family (MusaFer1-5) was quantified under iron-excess and -deficient conditions. The supplementation of 250 and 350 μM iron enhanced expression of all MusaFer genes; however, MusaFer1 was increased maximally by 2- and 4- fold in leaves and roots respectively. Under iron deficient condition, all five MusaFer genes were downregulated, indicating their iron dependent regulation. In MusaFer1 overexpressing lines, iron content was increased by 2- and 3-fold in leaves and roots respectively, as compared with that of untransformed lines. The increased iron was mainly localized in the epidermal regions of petiole. The analysis of MusaFer1 promoter indicated that it might control the expression of iron metabolism related genes and also other genes of MusaFer family. MusaFer1 overexpression led to downregulated expression of MusaFer3, MusaFer4 and MusaFer5 in transgenic leaves which might be associated with the plant’s compensatory mechanism in response to iron flux. Other iron metabolism genes like Ferric reductase (FRO), transporters (IRT, VIT and YSL) and chelators (NAS, DMAS and NAAT) were also differentially expressed in transgenic leaf and root, suggesting the multifaceted impact of MusaFer1 towards iron uptake and organ distribution. Additionally, MusaFer1 overexpression increased plant tolerance against methyl viologen and excess iron which was quantified in terms of photosynthetic efficiency and malondialdehyde content. Thus, the study not only broadens our understanding about iron metabolism but also highlights MusaFer1 as a suitable candidate gene for iron fortification in banana. [ABSTRACT FROM AUTHOR]

Degeneration of the intervertebral discs (IVD) is a leading cause of neck and low back pain. Degeneration begins in the central nucleus pulposus region, leading to loss of IVD osmotic properties. Regeneration approaches include administration of matrix-mimicking scaffolds, cells and/or therapeutic factors. Cell-targeting strategies are likely to improve delivery due to the low cell numbers in the IVD. Single-chain antibody fragments (scFvs) that bind IVD cells were isolated for potential delivery of therapeutics to degenerated IVD. The most cell-distal domain of neural cell adhesion molecule 1 (NCAM1) was cloned and expressed in Escherichia coli. Phage display technology was used to isolate a human scFv against the recombinant domain by panning a scFv library on the immobilised protein. The isolated scFv bound cultured rat astrocytes, as well as bovine nucleus pulposus and annulus fibrosus cells in immunocytochemical studies. The scFv also labelled cells in bovine spinal cord and six-month and two-year old bovine IVD sections by immunohistochemistry. Antibody fragments can provide cell-binding moieties at improved cost, time, yield and functionalisation potential over whole antibodies. The described scFv has potential application in delivery of therapeutics to NCAM1-expressing cells in degenerated IVD. [ABSTRACT FROM AUTHOR]

βA3/A1-crystallin is a lens structural protein that plays an important role in maintaining lens transparency via interactions with other crystallins. While the function of βA3/A1-crystallin in the retina is well studied, its functions in the lens, other than as a structural protein, remain unclear. In the current study, we generated the lens-specific βA3/A1-crystallin conditional knockout mouse (named βA3/A1ckO) and explored phenotypic changes and the function of the crystallin in the lens. The βA3/A1ckO mice showed congenital cataract at birth and exhibited truncation of lens proteins. Several truncated protein fragments were recovered as a pellet during a low-speed centrifugation (800 rpm, 70 x g) followed by a relatively higher speed centrifugation (5000 rpm, 2744 x g). Mass spectrometric analysis of pellets recovered following the two centrifugations showed that among the fragments with Mr < 20 kDa, the majority of these were from β-tubulin, and some from phakinin, αA-crystallin, and calpain-3. Further, we observed that in vitro activation of calpain-3 by calcium treatment of the wild-type-lens homogenate resulted in the degradation of calpain-3, αA-crystallin and β-tubulin and insolubilization of these proteins. Based on these results, it was concluded that the activation of calpain 3 resulted in proteolysis of β-tubulin, which disrupted cellular microtubular structure, and caused proteolysis of other lens proteins (αA-crystallin and phakinin). These proteolyzed protein fragments become insoluble, and together with the disruption of microtubular structure, and could be the causative factors in the development of congenital nuclear cataract in βA3/A1cKO mice. [ABSTRACT FROM AUTHOR]

Objective: Evaluate the efficacy of AYUSH 64, a standard polyherbal Ayurvedic drug in COVID-19. Methods: During the first pandemic wave, 140 consenting and eligible hospitalized adult participants with mild-moderate symptomatic disease (specific standard RT-PCR assay positive) were selected as per a convenience sample, and randomized (1:1 ratio) to an open-label (assessor blind) two-arm multicentric drug trial; standard of care (SOC as per Indian guidelines) versus AYUSH 64 combined with SOC (AYUSH plus). Participants were assessed daily and discharged once clinical recovery (CR, primary efficacy) was achieved which was based on a predetermined set of criteria (resolution of symptoms, normal peripheral oximetry, and negative specific RT-PCR assay). Each participant was followed using an indigenous software program(mobile phone) and completed a 12-week study period. The dose of AYUSH 64 was 2 tablets oral, 500 mg each, bid for 12 weeks (AYUSH plus only). Significant P was <0.05 (two-sided). On randomization, the groups were found well matched. Results: The mean interval time from randomization to CR was significantly superior in the AYUSH plus group [mean 6.45 days versus 8.26 days, 95% Confidence Interval of the difference -3.02 to -0.59 (P = 0.003, Student's 't test] as per-protocol analysis (134 participants); significant (P = 0.002) on an intention to treat analysis. 70% of the participants in AYUSH plus recovered during the first week (P = 0.046, Chi-square) and showed a significantly better change in physical health, fatigue, and quality of life measures. 48 adverse events, mostly mild and gut related, were reported by each group. There were 20 patient withdrawals (8 in AYUSH plus) but none due to an AE. There were no deaths. Daily assessment (hospitalization) and supervised drug intake ensured robust efficacy data. The open-label design was a concern (study outcome). Conclusions: AYUSH 64 in combination with SOC hastened recovery, reduced hospitalization, and improved health in COVID-19. It was considered safe and well-tolerated. Further clinical validation (Phase III) is required. Trial registration: CTRI/2020/06/025557. [ABSTRACT FROM AUTHOR]

Vascular smooth muscle cells (VSMC) from spontaneously hypertensive rats (SHR) exhibit decreased levels of nitric oxide (NO) that may be responsible for the overexpression of Giα proteins that has been shown as a contributing factor for the pathogenesis of hypertension in SHR. The present study was undertaken to investigate if increasing the intracellular levels of NO by NO donor S-Nitroso-N-acetyl-DL-penicillamine (SNAP) could attenuate the enhanced expression of Giα proteins in VSMC from SHR and explore the underlying mechanisms responsible for this response. The expression of Giα proteins and phosphorylation of ERK1/2, growth factor receptors and c-Src was determined by Western blotting using specific antibodies. Treatment of VSMC from SHR with SNAP for 24 hrs decreased the enhanced expression of Giα-2 and Giα-3 proteins and hyperproliferation that was not reversed by 1H (1, 2, 4) oxadiazole (4, 3-a) quinoxalin-1-one (ODQ), an inhibitor of soluble guanylyl cyclase, however, PD98059, a MEK inhibitor restored the SNAP-induced decreased expression of Giα proteins towards control levels. In addition, the increased production of superoxide anion, NAD(P)H oxidase activity, overexpression of AT1 receptor, Nox4, p22phox and p47phox proteins, enhanced levels of TBARS and protein carbonyl, increased phosphorylation of PDGF-R, EGF-R, c-Src and ERK1/2 in VSMC from SHR were all decreased to control levels by SNAP treatment. These results suggest that NO decreased the enhanced expression of Giα-2/3 proteins and hyperproliferation of VSMC from SHR by cGMP-independent mechanism and involves ROS and ROS-mediated transactivation of EGF-R/PDGF-R and MAP kinase signaling pathways. [ABSTRACT FROM AUTHOR]

The recent Zika viral (ZIKV) epidemic has been associated with severe neurological pathologies such as neonatal microcephaly and Guillain-Barre syndrome but unfortunately no vaccine or medication is effectively available yet. Zika NS2B-NS3pro is essential for the proteolysis of the viral polyprotein and thereby viral replication. Thus NS2B-NS3pro represents an attractive target for anti-Zika drug discovery/design. Here, we have characterized the solution conformations and catalytic parameters of both linked and unlinked Zika NS2B-NS3pro complexes and found that the unlinked complex manifested well-dispersed NMR spectra. Subsequently with selective isotope-labeling using NMR spectroscopy, we demonstrated that C-terminal residues (R73-K100) of NS2B is highly disordered without any stable tertiary and secondary structures in the Zika NS2B-NS3pro complex in the free state. Upon binding to the well-characterized serine protease inhibitor, bovine pancreatic trypsin inhibitor (BPTI), only the extreme C-terminal residues (L86-K100) remain disordered. Additionally, we have identified five flavonoids and one natural phenol rich in edible plants including fruits and vegetables, which inhibit Zika NS2B-NS3pro in a non-competitive mode, with Ki ranging from 770 nM for Myricetin to 34.02 μM for Apigenin. Molecular docking showed that they all bind to a pocket on the back of the active site and their structure-activity relationship was elucidated. Our study provides valuable insights into the solution conformation of Zika NS2B-NS3pro and further deciphers its susceptibility towards allosteric inhibition by natural products. As these natural product inhibitors fundamentally differ from the currently-known active site inhibitors in terms of both inhibitory mode and chemical scaffold, our finding might open a new avenue for development of better allosteric inhibitors to fight ZIKV infection. [ABSTRACT FROM AUTHOR]

Abiotic stresses adversely affect cellular homeostasis, impairing overall growth and development of plants. These initial stress signals activate downstream signalling processes, which, subsequently, activate stress-responsive mechanisms to re-establish homeostasis. Dehydrins (DHNs) play an important role in combating dehydration stress. Rice (Oryza sativa L.), which is a paddy crop, is susceptible to drought stress. As drought survival in rice might be viewed as a trait with strong evolutionary selection pressure, we observed DHNs in the light of domestication during the course of evolution. Overall, 65 DHNs were identified by a genome-wide survey of 11 rice species, and 3 DHNs were found to be highly conserved. The correlation of a conserved pattern of DHNs with domestication and diversification of wild to cultivated rice was validated by synonymous substitution rates, indicating that Oryza rufipogon and Oryza sativa ssp. japonica follow an adaptive evolutionary pattern; whereas Oryza nivara and Oryza sativa ssp. indica demonstrate a conserved evolutionary pattern. A comprehensive analysis of tissue-specific expression of DHN genes in japonica and their expression profiles in normal and PEG (poly ethylene glycol)-induced dehydration stress exhibited a spatiotemporal expression pattern. Their interaction network reflects the cross-talk between gene expression and the physiological processes mediating adaptation to dehydration stress. The results obtained strongly indicated the importance of DHNs, as they are conserved during the course of domestication. [ABSTRACT FROM AUTHOR]

A putative novel rhabdovirus (SfRV) was previously identified in a Spodoptera frugiperda cell line (Sf9 cells [ATCC CRL-1711 lot 58078522]) by next generation sequencing and extensive bioinformatic analysis. We performed an extensive analysis of our Sf9 cell bank (ATCC CRL-1711 lot 5814 [Sf9L5814]) to determine whether this virus was already present in cells obtained from ATCC in 1987. Inverse PCR of DNA isolated from Sf9 L5814 cellular DNA revealed integration of SfRV sequences in the cellular genome. RT-PCR of total RNA showed a deletion of 320 nucleotides in the SfRV RNA that includes the transcriptional motifs for genes X and L. Concentrated cell culture supernatant was analyzed by sucrose density gradient centrifugation and revealed a single band at a density of 1.14 g/ml. This fraction was further analysed by electron microscopy and showed amorphous and particulate debris that did not resemble a rhabdovirus in morphology or size. SDS-PAGE analysis confirmed that the protein composition did not contain the typical five rhabdovirus structural proteins and LC-MS/MS analysis revealed primarily of exosomal marker proteins, the SfRV N protein, and truncated forms of SfRV N, P, and G proteins. The SfRV L gene fragment RNA sequence was recovered from the supernatant after ultracentrifugation of the 1.14 g/ml fraction treated with diethyl ether suggesting that the SfRV L gene fragment sequence is not associated with a diethyl ether resistant nucleocapsid. Interestingly, the 1.14 g/ml fraction was able to transfer baculovirus DNA into Sf9L5814 cells, consistent with the presence of functional exosomes. Our results demonstrate the absence of viral particles in ATCC CRL-1711 lot 5814 Sf9 cells in contrast to a previous study that suggested the presence of infectious rhabdoviral particles in Sf9 cells from a different lot. This study highlights how cell lines with different lineages may present different virosomes and therefore no general conclusions can be drawn across Sf9 cells from different laboratories. [ABSTRACT FROM AUTHOR]

Introduction: Duct ectasia (DE) and periductal mastitis (PDM) are the most common benign breast conditions seen in women. The etiopathogenesis of these entities is still not clear and most of the theories regarding the causation are based on the histological features as seen on light microscopy. The ultramicroscopic features associated with these conditions that may give more insight to the etiopathogenesis are unknown. Aim: To study the ultrastructural changes occurring in mammary duct cones in patients with DE and PDM using Transmission Electron Microscopic (TEM). Method: Major ducts removed by radical duct excision from 21 patients with final histopathological diagnosis of DE and PDM were subjected to TEM study with 2 normal duct samples as controls. Results: The TEM features of DE were denudation of the epithelial cells with focal loss of microvilli, widening of the inter-epithelial junctions with focal disruption of the T bars, periductal collagenisation without inflammation, and features suggestive of Epithelial Mesenchymal Transition (EMT). PDM features are intact epithelial lining with proliferative epithelium and periductal collagenisation with inflammation. Conclusions: Based on the TEM findings, we suggest that DE and PDM are two different entities. EMT a novel finding observed in DE in this study. [ABSTRACT FROM AUTHOR]

Rhodococcus are gram-positive bacteria, which can exist in two different shapes rod and cocci. A number of studies have been done in the past on replication and stability of small plasmids in this bacterium; however, there are no reports on spatial localization and segregation of these plasmids. In the present study, a low copy number plasmid pDS3 containing pRC4 replicon was visualized in growing cells of Rhodococcus erythropolis PR4 (NBRC100887) using P1 parS-ParB-GFP system. Cells were initially cocci and then became rod shaped in exponential phase. Cocci cells were found to be non-replicating as evident by the presence of single fluorescence focus corresponding to the plasmid and diffuse fluorescence of DnaB-GFP. Rod shaped cells contained plasmid either present as one fluorescent focus observed at the cell center or two foci localized at quarter positions. The results suggest that the plasmid is replicated at the cell center and then it goes to quarter position. In order to observe the localization of plasmid with respect to nucleoid, plasmid segregation was also studied in filaments where it was found to be replicated at the cell center in a nucleoid free region. To the best of our knowledge, this is the first report on segregation of small plasmids in R. erythropolis. [ABSTRACT FROM AUTHOR]

Two broad classes of RNase P trim the 5' leader of precursor tRNAs (pre-tRNAs): ribonucleoprotein (RNP)- and proteinaceous (PRORP)-variants. These two RNase P types, which use different scaffolds for catalysis, reflect independent evolutionary paths. While the catalytic RNA-based RNP form is present in all three domains of life, the PRORP family is restricted to eukaryotes. To obtain insights on substrate recognition by PRORPs, we examined the 5' processing ability of recombinant Arabidopsis thaliana PRORP1 (AtPRORP1) using a panel of pre-tRNASer variants and model hairpin-loop derivatives (pATSer type) that consist of the acceptor-T-stem stack and the T-/D-loop. Our data indicate the importance of the identity of N-1 (the residue immediately 5' to the cleavage site) and the N-1:N+73 base pair for cleavage rate and site selection of pre-tRNASer and pATSer. The nucleobase preferences that we observed mirror the frequency of occurrence in the complete suite of organellar pre-tRNAs in eight algae/plants that we analyzed. The importance of the T-/D-loop in pre-tRNASer for tight binding to AtPRORP1 is indicated by the 200-fold weaker binding of pATSer compared to pre-tRNASer, while the essentiality of the T-loop for cleavage is reflected by the near-complete loss of activity when a GAAA-tetraloop replaced the T-loop in pATSer. Substituting the 2'-OH at N-1 with 2'-H also resulted in no detectable cleavage, hinting at the possible role of this 2'-OH in coordinating Mg2+ ions critical for catalysis. Collectively, our results indicate similarities but also key differences in substrate recognition by the bacterial RNase P RNP and AtPRORP1: while both forms exploit the acceptor-T-stem stack and the elbow region in the pre-tRNA, the RNP form appears to require more recognition determinants for cleavage-site selection. [ABSTRACT FROM AUTHOR]

Tillage and crop residue management play an imperative role in soil physico-chemical properties that eventually affects crop productivity. The objective of the study to find out a compatible combination of tillage and crop residue management for achieving sustainable food production by improving soil properties, providing favorable environment to crop plants. Secondly, managing crop residues effectively to reduce environmental pollution arising due to crop residue burning. With this aim, a field experiment was conducted on six years continued running experiment under conservation agricultural practices during rabi season of 2019–20 on chickpea. The experiment was comprised of five tillage operations with or without crop residue in main plot and three levels of nutrients in sub plots laid out in split plot design with three replications. Reduced Tillage with 60cm residue height (RT60) was recorded higher growth and yield attributes over conventional tillage practice that attributed to economic yield enhancement. The percent yield increment under NT and RT with 30 and 60cm height residue retention varied from 6.91% to 9.67% over conventional tillage. Maximum grain (2380 kg ha-1) and biological output (5762 kg ha-1) was recorded under RT60 (T4), which ascribed to higher net return (Rs 60551 ha-1) and benefit-cost ratio (2.97). The augmentation in net monetary benefit among tillage systems was lies between 24.32% to 37.78% over conventional tillage. The seed protein content ranged between 20.38 to 21.69% among the treatments. Moreover, total N uptake was maximum under RT60, while total P and K uptake was higher in No Tillage with 30cm residue height (T1). No-Tillage with 60cm residue height (NT60) recorded relatively higher soil moisture content (SMC) (22.71 and 15.40%). Treatment NT30 accrued relatively higher value of soil bulk density (1.42 Mg m-3) followed by NT60 and RT60 in comparison to conventional tillage (1.34 Mg m-3). In conclusion, NT and RT with 60cm residue height along with STCR (N3) nutrient dose was found effective for sustainable food production. [ABSTRACT FROM AUTHOR]

Introduction: Ursolic acid (UA) and oleanolic acid (OA) are triterpenoids. They are used to treat numerous diseases, including tuberculosis. Combinations of these drugs provide new insight into the management of tuberculosis. The major obstacle is the effective delivery of these drugs to the lungs, which are mainly affected due to M. tuberculosis. A metered-dose inhaler (MDI) was developed to address this issue containing UA and OA, followed by in-vitro and in-vivo evaluation. Methods: In the present study, MDI formulation was prepared by incorporating UA and OA at the dose level of 120 μg/ml in each actuation. In-vitro evaluation of this MDI formulation was performed to ensure its suitability to deliver UA and OA preciously. With prior approval of IAEC, a pharmacokinetic and acute inhalation toxicity study was conducted using MDI on Wistar rats. Results: The pharmacokinetic study showed an increased biological half-life of UA (9.23±0.104 h) and OA (8.93±0.166 h) in combination therapy. In-vivo toxicity study demonstrated no adverse effects on body weight and vital organs in the treatment group compared with the control group. Histopathology examination of these essential organs showed no abnormalities. Mild alternation in the biochemical and hematological parameters was observed. However, these alterations did not affect the overall health of the animals. Conclusion: The present study documents a detailed study for the safety and pharmacokinetics of UA and OA in-vivo for their advanced application in tuberculosis disease. [ABSTRACT FROM AUTHOR]

Dimethyl fumarate (DMF) is a first-line prodrug for the treatment of relapsing-remitting multiple sclerosis (RRMS) that is completely metabolized to monomethyl fumarate (MMF), the active metabolite, before reaching the systemic circulation. Its metabolism has been proposed to be due to ubiquitous esterases in the intestines and other tissues, but the specific enzymes involved are unknown. We hypothesized based on its structure and extensive presystemic metabolism that DMF would be a carboxylesterase substrate subject to interaction with alcohol. We sought to determine the enzymes(s) responsible for the extensive presystemic metabolism of DMF to MMF and the effect of alcohol on its disposition by conducting metabolic incubation studies in human recombinant carboxylesterase-1 (CES1), carboxylesterase-2 (CES2) and human intestinal microsomes (HIM), and by performing a follow-up study in an in vivo mouse model. The in vitro incubation studies demonstrated that DMF was only metabolized to MMF by CES1. Consistent with the incubation studies, the mouse pharmacokinetic study demonstrated that alcohol decreased the maximum concentration and area-under-the-curve of MMF in the plasma and the brain after dosing with DMF. We conclude that alcohol may markedly decrease exposure to the active MMF metabolite in the plasma and brain potentially decreasing the effectiveness of DMF in the treatment of RRMS. [ABSTRACT FROM AUTHOR]

Remote sensing can be used to map tillage practices at large spatial and temporal scales. However, detecting such management practices in smallholder systems is challenging given that the size of fields is smaller than historical readily-available satellite imagery. In this study we used newer, higher-resolution satellite data from Sentinel-1, Sentinel-2, and Planet to map tillage practices in the Eastern Indo-Gangetic Plains in India. We specifically tested the classification performance of single sensor and multiple sensor random forest models, and the impact of spatial, temporal, or spectral resolution on classification accuracy. We found that when considering a single sensor, the model that used Planet imagery (3 m) had the highest classification accuracy (86.55%) while the model that used Sentinel-1 data (10 m) had the lowest classification accuracy (62.28%). When considering sensor combinations, the model that used data from all three sensors achieved the highest classification accuracy (87.71%), though this model was not statistically different from the Planet only model when considering 95% confidence intervals from bootstrap analyses. We also found that high levels of accuracy could be achieved by only using imagery from the sowing period. Considering the impact of spatial, temporal, and spectral resolution on classification accuracy, we found that improved spatial resolution from Planet contributed the most to improved classification accuracy. Overall, it is possible to use readily-available, high spatial resolution satellite data to map tillage practices of smallholder farms, even in heterogeneous systems with small field sizes. [ABSTRACT FROM AUTHOR]

Background: In 2017, accidents and other acts of violence were the fifth most common cause of death in Switzerland. Moreover, there are increasing numbers of refugees and asylum seekers (AS), who often exhibit distinct disease profiles from those of the natives of the host country. If these differences could be clearly identified, this might help to develop and implement strategies to prevent injuries in health care programs for refugees and asylum seekers. The aim of this study was to examine the types and characteristics of physical trauma profiles in patients from the two largest groups of AS in Switzerland–from Eastern Africa (EA) and the Middle East (ME)–who consulted a Swiss Emergency Department (ED) in 2017/2018. Furthermore, the physical trauma profiles of Swiss national (SN) patients were examined in order to explore potential differences. Methods: Descriptive retrospective study of adult trauma patients consulting the ED of a Swiss University Hospital between 01/2017 and 12/2018. The study included 157 asylum seeking trauma patients from EA and ME were included in the study. These were matched by gender and age to 157 Swiss trauma patients consulting the ED in the study period. Results: There were significant differences between the groups with respect to type of admission, level of severity, localization and mechanisms of injury. While SN had higher levels of injuries related to road traffic or work, AS had higher levels of injuries related to attempted suicide or to assault. Conclusions: There were differences between AS and the local population with respect to the characteristics and patterns of injury, so that strategies for preventing injuries and promoting health must be tailored to the target population. Moreover, the observed high rates of outpatient treatment for both groups underline the increasing role of EDs as primary care providers for the population served. [ABSTRACT FROM AUTHOR]

Introduction: Real-world data on COVID-19 vaccine effectiveness are needed to validate evidence from randomized clinical trials. Accordingly, this study aims to evaluate, in a real-world setting in Brazil, the effectiveness of Pfizer-BioNTech BNT162b2 against symptomatic COVID-19 and COVID-19-related complications across diverse populations. Materials and methods: A test-negative case-control study with follow-up of cases is currently being conducted in Toledo, a city in southern Brazil, following a mass COVID-19 vaccination campaign with BNT162b2. The study is being conducted among patients aged 12 years or older seeking care in the public health system with acute respiratory symptoms and tested for SARS-CoV-2 on reverse transcription polymerase chain reaction (RT-PCR). Cases are RT-PCR positive and controls RT-PCR negative. Test-positive cases are prospectively followed through structured telephone interviews performed at 15 days post-enrollment, and at 1, 3, 6, 9 and 12 months. Baseline demographic, clinical, and vaccination data are being collected by means of structured interviews and medical registry records reviews at the time of enrollment. All RT-PCR-positive samples are screened for mutations to identify SARS-CoV-2 variants. Ethics and dissemination: The study protocol has been approved by the research ethics committee of all participant sites. Study findings will be disseminated through peer-reviewed publications and conference presentations. Trail registration: Clinicatrials.gov:NCT05052307. [ABSTRACT FROM AUTHOR]

Background: The Stanford Youth Diabetes Coaches Program (SYDCP) is a school based health program in which Family Medicine residents train healthy at-risk adolescents to become diabetes self-management coaches for family members with diabetes. This study evaluates the impact of the SYDCP when disseminated to remote sites. Additionally, this study aims to assess perceived benefit of enhanced curriculum. Methods: From 2012–2015, 10 high schools and one summer camp in the US and Canada and five residency programs were selected to participate. Physicians and other health providers implemented the SYDCP with racial/ethnic-minority students from low-income communities. Student coaches completed pre- and posttest surveys which included knowledge, health behavior, and psychosocial asset questions (i.e., worth and resilience), as well as open-ended feedback questions. T-test pre-post comparisons were used to determine differences in knowledge and psychosocial assets, and open and axial coding methods were used to analyze qualitative data. Results: A total of 216 participating high school students completed both pre-and posttests, and 96 nonparticipating students also completed pre- and posttests. Student coaches improved from pre- to posttest significantly on knowledge (p<0.005 in 2012–13, 2014 camp, and 2014–15); worth (p<0.1 in 2014–15); problem solving (p<0.005 in 2014 camp and p<0.1 in 2014–15); and self-efficacy (p<0.05 in 2014 camp). Eighty-two percent of student coaches reported that they considered making a behavior change to improve their own health as a result of program participation. Qualitative feedback themes included acknowledgment of usefulness and relevance of the program, appreciation for physician instructors, knowledge gain, pride in helping family members, improved relationships and connectedness with family members, and lifestyle improvements. Conclusion: Overall, when disseminated, this program can increase health knowledge and some psychosocial assets of at-risk youth and holds promise to empower these youth with health literacy and encourage them to adopt healthy behaviors. [ABSTRACT FROM AUTHOR]

Background: Large-scale war-related migration to Switzerland and other European countries is currently challenging European health systems. Little is known about recent patterns and trends in Emergency Department (ED) consultations by Asylum Seekers (AS). Methods: A retrospective single-centre analysis was performed of the data from all adult patients with the official status of “Asylum Seeker” or “Refugee” who consulted the ED of Bern University Hospital, Switzerland, between June 2012 and June 2015. Patient characteristics and clinical information, such as triage category, type of referral and discharge, violence-related injury and diagnostic group on discharge, were extracted from the computerised database or determined from the medical reports. Changes in categorical variables between the three studied years were described. Results: A total of 1,653 eligible adult patients were identified in the 3-year period. Between the first (06/12–06/13) and third periods (06/14–06/15), the number of presentations per year increased by about 45%. The AS came from 62 different nations, the most common countries being Eritrea (13%), Somalia (13%) and Syria (11%). The mean age was 33.3 years (SD 12.3) and two thirds (65.7%) were male. The proportion of women increased over time. Moreover the relative proportions shifted from patients between 20 and 50 years to patients of under 20 or over 60 years. Nearly two thirds of the patients were walk-in emergencies and this proportion increased over time. The mean triage score was 2.9 (SD 0.7), with more than 90% presenting as “urgent consultation”. About half of the patients were treated for trauma (17.2%), infections (16.8%) or psychiatric problems (14.2%). Trauma was seen in a higher proportion of male than female patients. About 25% of the patients were admitted for in-hospital treatment. Conclusions: The recent rise in AS in the population has lead to an increase in AS presenting to EDs. This changes the composition of ED patients and should raise awareness that changes in procedures may be needed. Infectious diseases and psychiatric problems remain a heavy burden for AS presenting in the ED. A trend towards an increasing proportion of walk-in patients to the ED could not be explained by this study. Further studies and surveillance are needed to investigate this trend. [ABSTRACT FROM AUTHOR]

Organisms quickly learn about their surroundings and display synaptic plasticity which is thought to be critical for their survival. For example, fruit flies Drosophila melanogaster exposed to highly enriched social environment are found to show increased synaptic connections and a corresponding increase in sleep. Here we asked if social environment comprising a pair of same-sex individuals could enhance sleep in the participating individuals. To study this, we maintained individuals of D. melanogaster in same-sex pairs for a period of 1 to 4 days, and after separation, monitored sleep of the previously socialized and solitary individuals under similar conditions. Males maintained in pairs for 3 or more days were found to sleep significantly more during daytime and showed a tendency to fall asleep sooner as compared to solitary controls (both measures together are henceforth referred to as “sleep-enhancement”). This sleep phenotype is not strain-specific as it is observed in males from three different “wild type” strains of D. melanogaster. Previous studies on social interaction mediated sleep-enhancement presumed ‘waking experience’ during the interaction to be the primary underlying cause; however, we found sleep-enhancement to occur without any significant increase in wakefulness. Furthermore, while sleep-enhancement due to group-wise social interaction requires Pigment Dispersing Factor (PDF) positive neurons; PDF positive and CRYPTOCHROME (CRY) positive circadian clock neurons and the core circadian clock genes are not required for sleep-enhancement to occur when males interact in pairs. Pair-wise social interaction mediated sleep-enhancement requires dopamine and olfactory signaling, while visual and gustatory signaling systems seem to be dispensable. These results suggest that socialization alone (without any change in wakefulness) is sufficient to cause sleep-enhancement in fruit fly D. melanogaster males, and that its neuronal control is context-specific. [ABSTRACT FROM AUTHOR]

βA3/A1-crystallin is an abundant structural protein of the lens that is very critical for lens function. Many different genetic mutations have been shown to associate with different types of cataracts in humans and in animal models. βA3/A1-crystallin has four Greek key-motifs that organize into two crystallin domains. It shown to bind calcium with moderate affinity and has putative calcium-binding site. Other than in the lens, βA3/A1 is also expressed in retinal astrocytes, retinal pigment epithelial (RPE) cells, and retinal ganglion cells. The function of βA3/A1-crystallin in the retinal cell types is well studied; however, a clear understanding of the function of this protein in the lens has not yet been established. In the current study, we generated the βA3/A1-crystallin knockout (KO) mouse and explored the function of βA3/A1-crystallin in lens development. Our results showed that βA3-KO mice develop congenital nuclear cataract and exhibit persistent fetal vasculature condition. At the cellular level KO lenses show defective lysosomal clearance and accumulation of nuclei, mitochondria, and autophagic cargo in the outer cortical region of the lens. In addition, the calcium level and the expression and activity of calpain-3 were increased in KO lenses. Taken together, these results suggest the lack of βA3-crystallin function in lenses, alters calcium homeostasis which in turn causes lysosomal defects and calpain activation. These defects are responsible for the development of nuclear cataract in KO lenses. [ABSTRACT FROM AUTHOR]

Purpose: Cisplatin based concomitant chemoradiation (CRT) is the standard treatment for locally advanced cervical cancer (CC). Glutathione S-transferase (GST), a phase II antioxidant enzyme is induced by oxidative stress generated by drugs and reactive oxidants. The present study was undertaken to evaluate the association of GSTM1, T1 and P1 polymorphisms with the outcome of CRT treatment in CC patients. Methods: A total of 227 cervical cancer patients with stages IIB-IIIB treated with the same chemoradiotherapy regimen were enrolled and genotyped for GSTM1, T1 and P1 gene polymorphisms by multiplex polymerase chain reaction (mPCR) and PCR-restriction fragment length polymorphism (PCR-RFLP). Overall survival was evaluated using Kaplan-Meier survival function and Cox proportional hazards model. All data were analyzed using SPSS (version 21.0). Results: Stratified analysis showed that GSTM1 null (M1-) genotype was associated with a significantly better survival among patients with stage IIB cervical cancer (log-rank P = 0.004) than cases with stage IIIA/IIIB. Death and recurrence were significantly higher in patients with GSTM1 present genotype (M1+) (P = 0.037 and P = 0.003 respectively) and those with M1- showed reduced hazard of death with an adjusted hazard ratio ‘HR’ of 0.47 (95% CI, 0.269–0.802, P = 0.006). Women with M1- genotype as well as in combination with GSTT1 null (T1-), GSTP1 (AG+GG) and GSTT1 null/GSTP1 (AG+GG) showed better survival and also reduced risk of death (HR = 0.31, P = 0.016; HR = 0.45, P = 0.013; HR = 0.31, P = 0.02 respectively). Conclusions: To the best of our knowledge, this is the first study to correlate the association of GSTM1, T1 and P1 gene polymorphisms with treatment outcome of CRT treated CC patients. Our results suggested that individuals with GSTM1 null genotype and in combination with GSTT1 null and GSTP1 (AG+GG) had a survival advantage. Such genetic studies may provide prognostic information in CRT treated CC patients. [ABSTRACT FROM AUTHOR]

The correct name is: David Jean Simon. Reference 1 Patel R, Marbaniang SP, Srivastava S, Kumar P, Chauhan S, Simon DJ (2021) Gender differential in low psychological health and low subjective well-being among older adults in India: With special focus on childless older adults. [Extracted from the article]

Ants often interact with other invertebrates as predators or mutualists. Epiphytic bromeliads provide nesting sites for ants, and could increase ant abundances in the tree canopy. We surveyed ants in the foliage of orange trees that either hosted bromeliads or did not. To determine if observed associations between bromeliads and tree ants were causal, we removed bromeliads from half of the trees, and resurveyed ants six weeks later. Our results show that bromeliad presence is correlated with higher ant abundances and different species of ants on orange trees during the dry season. This increase in ant abundance was driven primarily by Solenopsis ants, which were both numerous and found to facultatively nest in bromeliads. Bromeliad removal did not affect either ant abundance or composition, potentially because this manipulation coincided with the transition from dry to wet season. Other ant species were never encountered nesting in bromeliads, and the abundances of such ants on tree leaves were unaffected by bromeliad presence or removal. Considering the importance of ants in herbivore regulation, our findings suggest that bromeliads–through their association with ants–could indirectly be associated with biological control in agricultural systems. [ABSTRACT FROM AUTHOR]

Introduction: Two pharmacological possibilities exist for an acute ischemic stroke (AIS): recanalization of the occluded artery and neuroprotection from ischaemic injury, the latter's efficacy being debatable. We sought to determine whether administration of Citicoline immediately after recanalization therapy for AIS would improve clinical and radiological outcome at three months compared to standard treatment alone. Patients and methods: CAISR was a single centre, randomized, placebo-controlled, parallel-group trial with blinded endpoint assessment. It was approved by the All India Institute of Medical Sciences Institutional ethics committee and registered at the Clinical Trial Registry of India (CTRI/2018/011900). We recruited participants with AIS undergoing recanalization therapy and randomly assigned them to receive either Citicoline or placebo in 1:1 ratio. Citicoline arm patients received Citicoline 1gm BD intravenously for three days, followed by oral citicoline 1gm BD for 39 days. Placebo arm patients received 100ml intravenous normal saline for three days, followed by multivitamin tablet BD for 39 days. All patients received standard of care. Outcome: Blinded assessors did the follow-up assessment at six weeks (MRI Brain-stroke volume) and three months (NIHSS 0–2, mRS 0–2 and Barthel index> = 95). Results: The infarct volume decreased from week 1 to week 6 by 2.6 cm3 on placebo versus 4.2 cm3 on Citicoline (p-0.483). The OR for achieving NIHSS 0–2, mRS 0–2 and Barthel index> = 95 with Citicoline was found to be 0.96(95%CI 0.39–2.40), 0.92(95%CI 0.40–2.05) and 0.87(95%CI 0.22–2.98) respectively. Conclusion: CAISR was the first to evaluate the role of Citicoline, when used immediately after recanalization therapy, when the penumbral tissue is the most susceptible either to be protected from injury or become ischemic. We did not find any significant difference between the Citicoline or placebo arms with respect to either our primary or secondary outcomes. [ABSTRACT FROM AUTHOR]

Alternaria brassicicola causes black spot disease of Brassica species. The functional importance of pectin digestion enzymes and unidentified phytotoxins in fungal pathogenesis has been suspected but not verified in A. brassicicola. The fungal transcription factor AbPf2 is essential for pathogenicity and induces 106 genes during early pathogenesis, including the pectate lyase-coding gene, PL1332. The aim of this study was to test the importance and roles of PL1332 in pathogenesis. We generated deletion strains of the PL1332 gene, produced heterologous PL1332 proteins, and evaluated their association with virulence. Deletion strains of the PL1332 gene were approximately 30% less virulent than wild-type A. brassicicola, without showing differences in colony expansion on solid media and mycelial growth in nutrient-rich liquid media or minimal media with pectins as a major carbon source. Heterologous PL1332 expressed as fusion proteins digested polygalacturons in vitro. When the fusion proteins were injected into the apoplast between leaf veins of host plants the tissues turned dark brown and soft, resembling necrotic leaf tissue. The PL1332 gene was the first example identified as a general toxin-coding gene and virulence factor among the 106 genes regulated by the transcription factor, AbPf2. It was also the first gene to have its functions investigated among the 19 pectate lyase genes and several hundred putative cell-wall degrading enzymes in A. brassicicola. These results further support the importance of the AbPf2 gene as a key pathogenesis regulator and possible target for agrochemical development. [ABSTRACT FROM AUTHOR]

Seabuckthorn is an economically important dioecious plant in which mechanism of sex determination is unknown. The study was conducted to identify seabuckthorn homologous genes involved in floral development which may have role in sex determination. Forty four putative Genes involved in sex determination (GISD) reported in model plants were shortlisted from literature survey, and twenty nine seabuckthorn homologous sequences were identified from available seabuckthorn genomic resources. Of these, 21 genes were found to differentially express in either male or female flower bud stages. HrCRY2 was significantly expressed in female flower buds only while HrCO had significant expression in male flowers only. Among the three male and female floral development stages (FDS), male stage II had significant expression of most of the GISD. Information on these sex-specific expressed genes will help in elucidating sex determination mechanism in seabuckthorn. [ABSTRACT FROM AUTHOR]

There is limited data on the clinical, cellular and molecular changes in relapsed acute promyeloytic leukemia (RAPL) in comparison with newly diagnosed cases (NAPL). We undertook a prospective study to compare NAPL and RAPL patients treated with arsenic trioxide (ATO) based regimens. 98 NAPL and 28 RAPL were enrolled in this study. RAPL patients had a significantly lower WBC count and higher platelet count at diagnosis. IC bleeds was significantly lower in RAPL cases (P=0.022). The ability of malignant promyelocytes to concentrate ATO intracellularly and their in-vitro IC50 to ATO was not significantly different between the two groups. Targeted NGS revealed PML B2 domain mutations in 4 (15.38%) of the RAPL subset and none were associated with secondary resistance to ATO. A microarray GEP revealed 1744 genes were 2 fold and above differentially expressed between the two groups. The most prominent differentially regulated pathways were cell adhesion (n=92), cell survival (n=50), immune regulation (n=74) and stem cell regulation (n=51). Consistent with the GEP data, immunophenotyping revealed significantly increased CD34 expression (P=0.001) in RAPL cases and there was in-vitro evidence of significant microenvironment mediated innate resistance (EM-DR) to ATO. Resistance and relapse following treatment with ATO is probably multi-factorial, mutations in PML B2 domain while seen only in RAPL may not be the major clinically relevant cause of subsequent relapses. In RAPL additional factors such as expansion of the leukemia initiating compartment along with EM-DR may contribute significantly to relapse following treatment with ATO based regimens. [ABSTRACT FROM AUTHOR]

Hypoxia is a hallmark of solid tumors including glioblastoma (GBM). Its synergism with Notch signaling promotes progression in different cancers. However, Notch signaling exhibits pleiotropic roles and the existing literature lacks a comprehensive understanding of its perturbations under hypoxia in GBM with respect to all components of the pathway. We identified the key molecular cluster(s) characteristic of the Notch pathway response in hypoxic GBM tumors and gliomaspheres. Expression of Notch and hypoxia genes was evaluated in primary human GBM tissues by q-PCR. Clustering and statistical analyses were applied to identify the combination of hypoxia markers correlated with upregulated Notch pathway components. We found well-segregated tumor—clusters representing high and low HIF-1α/PGK1-expressors which accounted for differential expression of Notch signaling genes. In combination, a five-hypoxia marker set (HIF-1α/PGK1/VEGF/CA9/OPN) was determined as the best predictor for induction of Notch1/Dll1/Hes1/Hes6/Hey1/Hey2. Similar Notch-axis genes were activated in gliomaspheres, but not monolayer cultures, under moderate/severe hypoxia (2%/0.2% O2). Preliminary evidence suggested inverse correlation between patient survival and increased expression of constituents of the hypoxia-Notch gene signature. Together, our findings delineated the Notch-axis maximally associated with hypoxia in resected GBM, which might be prognostically relevant. Its upregulation in hypoxia-exposed gliomaspheres signify them as a better in-vitro model for studying hypoxia-Notch interactions than monolayer cultures. [ABSTRACT FROM AUTHOR]

As part of our drug discovery program for anti-filarial agents from Indian medicinal plants, leaves of Eucalyptus tereticornis were chemically investigated, which resulted in the isolation and characterization of an anti-filarial agent, ursolic acid (UA) as a major constituent. Antifilarial activity of UA against the human lymphatic filarial parasite Brugia malayi using in vitro and in vivo assays, and in silico docking search on glutathione-s-transferase (GST) parasitic enzyme were carried out. The UA was lethal to microfilariae (mf; LC100: 50; IC50: 8.84 µM) and female adult worms (LC100: 100; IC50: 35.36 µM) as observed by motility assay; it exerted 86% inhibition in MTT reduction potential of the adult parasites. The selectivity index (SI) of UA for the parasites was found safe. This was supported by the molecular docking studies, which showed adequate docking (LibDock) scores for UA (−8.6) with respect to the standard antifilarial drugs, ivermectin (IVM −8.4) and diethylcarbamazine (DEC-C −4.6) on glutathione-s-transferase enzyme. Further, in silico pharmacokinetic and drug-likeness studies showed that UA possesses drug-like properties. Furthermore, UA was evaluated in vivo in B. malayi-M. coucha model (natural infection), which showed 54% macrofilaricidal activity, 56% female worm sterility and almost unchanged microfilaraemia maintained throughout observation period with no adverse effect on the host. Thus, in conclusion in vitro, in silico and in vivo results indicate that UA is a promising, inexpensive, widely available natural lead, which can be designed and developed into a macrofilaricidal drug. To the best of our knowledge this is the first ever report on the anti-filarial potential of UA from E. tereticornis, which is in full agreement with the Thomson Reuter's ‘Metadrug’ tool screening predictions. [ABSTRACT FROM AUTHOR]

Mesangial cells are glomerular cells of stromal origin. During immune complex mediated crescentic glomerulonephritis (Crgn), infiltrating and proliferating pro-inflammatory macrophages lead to crescent formation. Here we have hypothesised that mesangial cells, given their mesenchymal stromal origin, show similar immunomodulatory properties as mesenchymal stem cells (MSCs), by regulating macrophage function associated with glomerular crescent formation. We show that rat mesangial cells suppress conA-stimulated splenocyte proliferation in vitro, as previously shown for MSCs. We then investigated mesangial cell-macrophage interaction by using mesangial cells isolated from nephrotoxic nephritis (NTN)-susceptible Wistar Kyoto (WKY) and NTN-resistant Lewis (LEW) rats. We first determined the mesangial cell transcriptome in WKY and LEW rats and showed that this is under marked genetic control. Supernatant transfer results show that WKY mesangial cells shift bone marrow derived macrophage (BMDM) phenotype to M1 or M2 according to the genetic background (WKY or LEW) of the BMDMs. Interestingly, these effects were different when compared to those of MSCs suggesting that mesangial cells can have unique immunomodulatory effects in the kidney. These results demonstrate the importance of the genetic background in the immunosuppressive effects of cells of stromal origin and specifically of mesangial cell-macrophage interactions in the pathophysiology of crescentic glomerulonephritis. [ABSTRACT FROM AUTHOR]

Biosurfactants are amphipathic molecules produced from microorganisms. There are relatively few species known where the detailed chemical characterization of biosurfactant has been reported. Here, we report isolation and chemical characterization of the biosurfactant produced by a biodesulfurizing bacterium Gordonia sp. IITR100. Biosurfactant production was determined by performing oil spreading, drop-collapse, Emulsion index (E24), and Bacterial adhesion to hydrocarbons (BATH) assay. The biosurfactant was identified as a glycolipid by LCMS and GCMS analysis. The chemical structure was further confirmed by performing FTIR and NMR of the extracted biosurfactant. The emulsion formed by the biosurfactant was found to be stable between temperatures of 4°C to 30°C, pH of 6 to 10 and salt concentrations up to 2%. It was successful in reducing the surface tension of the aqueous media from 61.06 mN/m to 36.82 mN/m. The biosurfactant produced can be used in petroleum, detergents, soaps, the food and beverage industry and the healthcare industry. [ABSTRACT FROM AUTHOR]

COVID-19 has led to unprecedented challenges and requires local and global efforts for its mitigation. Poor and marginalized populations are more vulnerable to the health, social and economic effects of the pandemic. The objective of this study was to know about the knowledge, attitude and practices towards COVID-19 among poor and marginalized communities in central India and the factors associated with them so that effective risk communication messages can be designed and community engagement needs and strategies can be identified. A cross-sectional survey was conducted using an Interactive Voice Response System as part of the NISHTHA-Swasthya Vani intervention, which is a platform for dissemination of key messages related to COVID-19, social welfare schemes, national health programs and other important information. A total of 1673 respondents participated in the survey. The mean knowledge, attitude and practice scores of the respondents was 4.06 (SD = 1.67) out of 8, 2.46 (SD = 1.18) out of 4 and 3.65 (SD = 0.73) out of 4 respectively. More than 50% respondents exhibited stigma towards recovered COVID-19 patients(n = 347) and towards health workers(n = 384) catering to COVID-19 patients. The factors associated with higher KAP scores were education, occupation, age and primary source of information on COVID-19. There was a positive correlation between knowledge and attitude (co-efficient: 0.32) and a negative correlation between knowledge and stigma (co-efficient: -0.28). The knowledge, and attitude scores related to COVID-19 were low among the poor and marginalized communities, while the prevalence of stigma was high. Therefore, there is a need for effective risk communication for these communities through alternate channels. [ABSTRACT FROM AUTHOR]

The periodic breakdowns of stripe rust resistance due to emergence of new virulent and more aggressive pathotypes of Puccinia striiformis f. sp. tritici have resulted in severe epidemics in India. This necessitates the search for new and more durable resistance sources against stripe rust. The three bread wheat cultivars PBW 343 (carries Yr9 and Yr27), PBW 621 (carries Yr17) and HD 2967 (gene not known) were highly popular among the farmers after their release in 2011. But presently all three cultivars are highly susceptible to stripe rust at seedling as well as at adult plant stages as their resistance has been broken down due to emergence of new pathotypes of the pathogen (110S119, 238S119). In previous study, the crosses of PBW 621 with PBW 343 and HD 2967 and evaluation of further generations (up to F4) against pathotype 78S84 resulted in resistant segregants. In the present study, the F5 and F6 RIL populations have been evaluated against new pathotypes of Pst. The RILs categorized based on the disease severity on the P (Penultimate leaf) and F (flag) leaf into three categories i.e., high, moderate and low level of APR (adult plant resistance) having 1–200, 201–400 and >400 values of AUDPC, respectively, upon infection with stripe rust. The various APR components (latent period, lesion growth rate, spore production and uredial density) were studied on each category, i.e., resistant, moderately resistant and susceptible. The values of APR parameters decreased as the level of resistance increased. Based on molecular analysis, the lines (representing different categories of cross PBW 621 X PBW 343) containing the genes Yr9 and Yr17 due to their interactive effect provide resistance. Based on BSA using 35k SNPs and KASP markers association with phenotypic data of the RIL population (PBW 621 X HD 2967) showed the presence of two QTLs (Q.Pst.pau-6B, Q.Pst.pau-5B) responsible for the residual resistance and two SNPs AX-94891670 and AX-94454107 were found to be associated with the trait of interest on chromosome 6B and 5B respectively. The present study concludes that in the population of both the crosses (PBW 621 X PBW 343 and PBW 621 X HD 2967) major defeated gene contributed towards residual resistance by interacting with minor gene/QTLs. [ABSTRACT FROM AUTHOR]

The portal venous circulation provides a conduit for pancreatic ductal adenocarcinoma (PDAC) tumor cells to the liver parenchyma sinusoids, a frequent site of metastasis. Turbulent flow in the portal circulation promotes retention of PDAC shed circulating tumor cells (CTC) and myeloid-derived immunosuppressor cells (MDSC). Excessive colony stimulating factor-1 receptor (CSF1R) signaling can induce myeloid differentiation to MDSC and transformation of MDSC to myeloid-derived fibroblasts (M-FB). Interactions between PDAC CTC and M-FB in the portal blood promotes the formation of immunoresistant clusters that enhance CTC proliferation, migration, and survival. Analysis of portal and peripheral blood samples collected intraoperatively from 30 PDAC patients undergoing pancreatico-duodenectomy showed that PDAC patient plasma contained high levels of macrophage colony stimulating factor (M-CSF/CSF1), granulocyte-macrophage colony stimulating factor (GM-CSF/CSF2), interleukin-8 (IL-8), and interleukin-34 (IL-34) compared to healthy control levels. Moreover, the level of M-CSF in portal blood was significantly higher than that detected in the peripheral blood of PDAC patients. PDAC CTC aseptically isolated by fluorescence activated cell sorting (FACS) out of freshly collected patient portal blood mononuclear cells (PortalBMC) had elevated RNA expression of IL34 (IL-34 gene) and CSF1 (M-CSF/CSF1 gene) which both signal through CSF1R. PDAC CTC also had high levels of RNA expression for CXCL8, the gene encoding chemokine interleukin-8 (IL-8) which can attract myeloid cells through their CXCR2 receptors. FACS-isolated portal PDAC CTC and M-FB co-cultured ex vivo had increased CTC proliferation, motility, and cluster formation compared to CTC cultured alone. CSF1R and CXCR2 cell surface expression were found on PDAC portal blood CTC and M-FB, suggesting that both cell types may respond to M-CSF, IL-34, and IL-8-mediated signaling. Portal PDAC CTC displayed enhanced RNA expression of CSF1 and IL34, while CTC+M-FB+ clusters formed in vivo had increased RNA expression of CSF2 and IL34. Portal M-FB were found to have high CSF1R RNA expression. CTC isolated from ex vivo 7-day cultures of PDAC patient portal blood mononuclear cells (PortalBMC) expressed elevated CSF1, IL34, and IL8 RNA, and CSF1 expression was elevated in M-FB. Treatment with rabbit anti-CSF1R antibodies decreased CTC proliferation. Treatment of PortalBMC cultures with humanized anti-CSF1R, humanized anti-IL-8, or anti-IL-34 antibodies disrupted CTC cluster formation and increased CTC apoptosis. U937 myeloid precursor cell line cultures treated with conditioned media from PortalBMC ex vivo cultures without treatment or treated with anti-IL-8 and/or anti-CSF1R did not prevent myeloid differentiation in the myeloid precursor cell line U937 to macrophage, dendritic cell, MDSC, and M-FB phenotypes; whereas, U937 cultures treated with conditioned media from PortalBMC ex vivo cultures exposed to anti-IL-34 were significantly inhibited in their myeloid differentiation to all but the M-FB phenotype. PDAC patient T cells that were found phenotypically anergic (CD3+CD25+CTLA4+PD1L1+) in PortalBMC could be re-activated (CD3+CD25+CTLA4-PD1L1-), and displayed increased interferon gamma (IFNγ) production when PortalBMC ex vivo cultures were treated with anti-CSF1R, anti-IL-8, and anti-IL-34 antibodies alone or in combination. These findings suggest that PDAC CTC have the potential to influence myeloid differentiation and/or antigen presenting cell activation in the PDAC portal blood microenvironment, and that disruption of CTC/M-FB interactions may be potential targets for reversing the immunosuppression supporting CTC survival in the portal blood. [ABSTRACT FROM AUTHOR]

The variability of clinical course and prognosis of COVID-19 highlights the necessity of patient sub-group risk stratification based on clinical data. In this study, clinical data from a cohort of Indian COVID-19 hospitalized patients is used to develop risk stratification and mortality prediction models. We analyzed a set of 70 clinical parameters including physiological and hematological for developing machine learning models to identify biomarkers. We also compared the Indian and Wuhan cohort, and analyzed the role of steroids. A bootstrap averaged ensemble of Bayesian networks was also learned to construct an explainable model for discovering actionable influences on mortality and days to outcome. We discovered blood parameters, diabetes, co-morbidity and SpO2 levels as important risk stratification features, whereas mortality prediction is dependent only on blood parameters. XGboost and logistic regression model yielded the best performance on risk stratification and mortality prediction, respectively (AUC score 0.83, AUC score 0.92). Blood coagulation parameters (ferritin, D-Dimer and INR), immune and inflammation parameters IL6, LDH and Neutrophil (%) are common features for both risk and mortality prediction. Compared with Wuhan patients, Indian patients with extreme blood parameters indicated higher survival rate. Analyses of medications suggest that a higher proportion of survivors and mild patients who were administered steroids had extreme neutrophil and lymphocyte percentages. The ensemble averaged Bayesian network structure revealed serum ferritin to be the most important predictor for mortality and Vitamin D to influence severity independent of days to outcome. The findings are important for effective triage during strains on healthcare infrastructure. [ABSTRACT FROM AUTHOR]

All stage resistance to stripe rust races prevalent in India was investigated in the European winter wheat cultivar 'Acienda'. In order to dissect the genetic basis of the resistance, a backcross population was developed between 'Acienda' and the stripe rust susceptible Indian spring wheat cultivar 'HD 2967'. Inheritance studies revealed segregation for a dominant resistant gene. High density SNP genotyping was used to map stripe rust resistance and marker regression analysis located stripe rust resistance to the distal end of wheat chromosome 1A. Interval mapping located this region between the SNP markers AX-95162217 and AX-94540853, at a LOD score of 15.83 with a phenotypic contribution of 60%. This major stripe rust resistance locus from 'Acienda' has been temporarily designated as Yraci. A candidate gene search in the 2.76 Mb region carrying Yraci on chromosome 1A identified 18 NBS-LRR genes based on wheat RefSeqv1.0 annotations. Our results indicate that as there is no major gene reported in the Yraci chromosome region, it is likely to be a novel stripe rust resistance locus and offers potential for deployment, using the identified markers, to confer all stage stripe rust resistance. [ABSTRACT FROM AUTHOR]

In this study, we determined the cytotoxic effects of piperine, a major constituent of black and long pepper in melanoma cells. Piperine treatment inhibited the growth of SK MEL 28 and B16 F0 cells in a dose and time-dependent manner. The growth inhibitory effects of piperine were mediated by cell cycle arrest of both the cell lines in G1 phase. The G1 arrest by piperine correlated with the down-regulation of cyclin D1 and induction of p21. Furthermore, this growth arrest by piperine treatment was associated with DNA damage as indicated by phosphorylation of H2AX at Ser139, activation of ataxia telangiectasia and rad3-related protein (ATR) and checkpoint kinase 1 (Chk1). Pretreatment with AZD 7762, a Chk1 inhibitor not only abrogated the activation of Chk1 but also piperine mediated G1 arrest. Similarly, transfection of cells with Chk1 siRNA completely protected the cells from G1 arrest induced by piperine. Piperine treatment caused down-regulation of E2F1 and phosphorylation of retinoblastoma protein (Rb). Apoptosis induced by piperine was associated with down-regulation of XIAP, Bid (full length) and cleavage of Caspase-3 and PARP. Furthermore, our results showed that piperine treatment generated ROS in melanoma cells. Blocking ROS by tiron protected the cells from piperine mediated cell cycle arrest and apoptosis. These results suggest that piperine mediated ROS played a critical role in inducing DNA damage and activation of Chk1 leading to G1 cell cycle arrest and apoptosis. [ABSTRACT FROM AUTHOR]

Vascular smooth muscle cells (VSMC) from spontaneously hypertensive rats (SHR) exhibit hyperproliferation and overexpression of cell cycle proteins. We earlier showed that small peptide fragments of cytoplasmic domain of natriuretic receptor-C (NPR-C) attenuate vasoactive peptide-induced hyperproliferation of VSMC. The present study investigated if C-ANP4–23, a specific agonist of NPR-C, could attanuate the hyperproliferation of VSMC from SHR by inhibiting the overexpression of cell cycle proteins and examine the underlying signaling pathways contributing to this inhibition. The proliferation of VSMC was determined by [3H] thymidine incorporation and the expression of proteins was determined by Western blotting. The hyperproliferation of VSMC from SHR and overexpression of cyclin D1,cyclin A, cyclin E, cyclin-dependent kinase 2 (cdk2), phosphorylated retinoblastoma protein (pRb), Giα proteins and enhanced phosphorylation of ERK1/2 and AKT exhibited by VSMC from SHR were attenuated by C-ANP4–23 to control levels. In addition, in vivo treatment of SHR with C-ANP4–23 also attenuated the enhanced proliferation of VSMC. Furthemore, PD98059, wortmannin and pertussis toxin, the inhibitors of MAP kinase, PI3kinase and Giα proteins respectively, also attenuated the hyperproliferation of VSMC from SHR and overexpression of cell cycle proteins to control levels. These results indicate that NPR-C activation by C-ANP4–23 attenuates the enhanced levels of cell cycle proteins through the inhibition of enhanced expression of Giα proteins and enhanced activation of MAPkinase/PI3kinase and results in the attenuation of hyperproliferation of VSMC from SHR. It may be suggested that C-ANP4–23 could be used as a therapeutic agent in the treatment of vascular complications associated with hypertension, atherosclerosis and restenosis. [ABSTRACT FROM AUTHOR]

The role of CD4+ T cells in the generation of therapeutic primary and memory immune responses in cancer diverse immunotherapy settings remains ambiguous. We herein investigated this issue using two vaccine formulations containing a novel costimulatory molecule, SA-4-1BBL, as adjuvant and HPV E7 or survivin (SVN) as tumor associated antigens (TAAs) in two mouse transplantable tumor models; the TC-1 cervical cancer expressing xenogeneic HPV E7 and 3LL lung carcinoma overexpressing autologous SVN. Single vaccination with optimized SA-4-1BBL/TAA formulations resulted in the eradication of 6-day established TC-1 and 3LL tumors in >70% of mice in both models. The in vivo depletion of CD4+ T cells one day before tumor challenge resulted in compromised vaccine efficacy in both TC-1 (25%) and 3LL (12.5%) tumor models. In marked contrast, depletion of CD4+ T cells 5 days post-tumor challenge and one day prior to vaccination did not significantly alter the therapeutic efficacy of these vaccines. However, long-term immunological memory was compromised in the 3LL, but not in TC-1 model as a significant number (85.7%) of tumor free-mice succumbed to tumor growth when rechallenged with 3LL cells 60 days after the initial tumor inoculation. Collectively, these results demonstrate the indispensable role CD4+ T cells play in the generation of therapeutic primary immune responses elicited by SA-4-1BBL/TAA-based vaccines irrespective of the nature of TAAs and establish the importance of CD4+ T cells for long-term immune memory against 3LL tumor expressing self-antigen SVN, but not TC-1 expressing xenogeneic viral antigen E7. [ABSTRACT FROM AUTHOR]

Background: The rising drug resistance in pathogenic bacteria and inefficiency of current antibiotics to meet clinical requirements has augmented the need to establish new and innovative approaches for antibacterial drug discovery involving identification of novel antibacterial targets and inhibitors. Being obligatory for bacterial growth, essential gene products are considered vital as drug targets. The bacterial protein YidC is highly conserved among pathogens and is essential for membrane protein insertion due to which it holds immense potential as a promising target for antibacterial therapy. Methods/Principal Findings: The aim of this study was to explore the feasibility and efficacy of expressed antisense-mediated gene silencing for specific downregulation of yidC in Escherichia coli. We induced RNA silencing of yidC which resulted in impaired growth of the host cells. This was followed by a search for antibacterial compounds sensitizing the YidC depleted cells as they may act as inhibitors of the essential protein or its products. The present findings affirm that reduction of YidC synthesis results in bacterial growth retardation, which warrants the use of this enzyme as a viable target in search of novel antibacterial agents. Moreover, yidC antisense expression in E. coli resulted in sensitization to antibacterial essential oils eugenol and carvacrol. Fractional Inhibitory Concentration Indices (FICIs) point towards high level of synergy between yidC silencing and eugenol/carvacrol treatment. Finally, as there are no known YidC inhibitors, the RNA silencing approach applied in this study put forward rapid means to screen novel potential YidC inhibitors. Conclusions/Significance: The present results suggest that YidC is a promising candidate target for screening antibacterial agents. High level of synergy reported here between yidC silencing and eugenol/carvacrol treatment is indicative of a potential antibacterial therapy. This is the first report indicating that the essential gene yidC is a therapeutic target of the antibacterial essential oils eugenol and carvacrol in E. coli. [ABSTRACT FROM AUTHOR]

Epidemiological evidence supports that maternal infection during gestation are notable risk factors for developmental mental illnesses including schizophrenia and autism. In prenatal lipopolysaccharide (LPS) model of immune activation in rats, the offspring exhibit significant impairments in behaviors mediated by central dopamine (DA) system. This study aimed to examine the temporal and regional pattern of postnatal DA development in the male offspring of pregnant Sprague-Dawley rats administered with 100 µg/kg LPS or saline at gestational days 15/16. Using ligand autoradiography, D1 and D2 dopamine receptors (D1R, D2R) and dopamine transporter (DAT) binding levels were measured in the prefrontal cortex (PFC) and sub cortical regions (dorsal striatum and nucleus accumbens core and shell) at pre pubertal (P35) and post pubertal ages (P60). We found a significant decrease in D2R ligand [(3)H] YM-90151-2 binding in the medial PFC (mPFC) in prenatal LPS-treated animals at P35 and P60 compared to respective saline groups. The decrease in D2R levels was not observed in the striatum or accumbens of maternal LPS-treated animals. No significant changes were observed in [(3)H] SCH23390 binding to D1R. However, the level of [(125)I] RTI-121 binding to DAT was selectively reduced in the nucleus accumbens core and shell at P35 in the prenatal LPS group. Immunohistochemical analysis showed that number of D2R immunopositive cells in infralimbic/prelimbic (IL/PL) part of mPFC was significantly reduced in the LPS group at P60. Prenatal LPS treatment did not significantly affect either the total number of mature neurons or parvalbumin (PV)-immunopositive interneurons in this region. However the number of PV and D2R co-labeled neurons was significantly reduced in the IL/PL subregion of PFC of LPS treated animals. Our data suggests D2R deficit in the PFC and PV interneurons may be relevant to understanding mechanisms of cortical dysfunctions described in prenatal infection animal models as well as schizophrenia. [ABSTRACT FROM AUTHOR]

Hexachlorocyclohexane dehydrochlorinase (LinA) mediates dehydrochlorination of γ-HCH to 1, 3, 4, 6-tetrachloro-1,4-cyclohexadiene that constitutes first step of the aerobic degradation pathway. We report the 3.5 Å crystal structure of a thermostable LinA-type2 protein, obtained from a soil metagenome, in the hexagonal space group P6322 with unit cell parameters a = b = 162.5, c = 186.3 Å, respectively. The structure was solved by molecular replacement using the coordinates of LinA-type1 that exhibits mesophile-like properties. Structural comparison of LinA-type2 and -type1 proteins suggests that thermostability of LinA-type2 might partly arise due to presence of higher number of ionic interactions, along with 4% increase in the intersubunit buried surface area. Mutational analysis involving the differing residues between the - type1 and -type2 proteins, circular dichroism experiments and functional assays suggest that Q20 and G23 are determinants of stability for LinA-type2. It was earlier reported that LinA-type1 exhibits enantioselectivity for the (-) enantiomer of α-HCH. Contrastingly, we identified that -type2 protein prefers the (+) enantiomer of α-HCH. Structural analysis and molecular docking experiments suggest that changed residues K20Q, L96C and A131G, vicinal to the active site are probably responsible for the altered enantioselectivity of LinA-type2. Overall the study has identified features responsible for the thermostability and enantioselectivity of LinA-type2 that can be exploited for the design of variants for specific biotechnological applications. [ABSTRACT FROM AUTHOR]

Use of Env in HIV vaccine development has been disappointing. Here we show that, in the presence of a biologically active Tat subunit vaccine, a trimeric Env protein prevents in monkeys virus spread from the portal of entry to regional lymph nodes. This appears to be due to specific interactions between Tat and Env spikes that form a novel virus entry complex favoring R5 or X4 virus entry and productive infection of dendritic cells (DCs) via an integrin-mediated pathway. These Tat effects do not require Tat-transactivation activity and are blocked by anti-integrin antibodies (Abs). Productive DC infection promoted by Tat is associated with a highly efficient virus transmission to T cells. In the Tat/Env complex the cysteine-rich region of Tat engages the Env V3 loop, whereas the Tat RGD sequence remains free and directs the virus to integrins present on DCs. V2 loop deletion, which unshields the CCR5 binding region of Env, increases Tat/Env complex stability. Of note, binding of Tat to Env abolishes neutralization of Env entry or infection of DCs by anti-HIV sera lacking anti-Tat Abs, which are seldom present in natural infection. This is reversed, and neutralization further enhanced, by HIV sera containing anti-Tat Abs such as those from asymptomatic or Tat-vaccinated patients, or by sera from the Tat/Env vaccinated monkeys. Thus, both anti-Tat and anti-Env Abs are required for efficient HIV neutralization. These data suggest that the Tat/Env interaction increases HIV acquisition and spreading, as a mechanism evolved by the virus to escape anti-Env neutralizing Abs. This may explain the low effectiveness of Env-based vaccines, which are also unlikely to elicit Abs against new Env epitopes exposed by the Tat/Env interaction. As Tat also binds Envs from different clades, new vaccine strategies should exploit the Tat/Env interaction for both preventative and therapeutic interventions. [ABSTRACT FROM AUTHOR]

Tumor hypoxia is a known driver of angiogenesis that also facilitates tumor growth. Moreover, poorly oxygenated central tumor area remains relatively radio or chemo resistant. HBO therapy is known to elevate the levels of dissolved oxygen and eliminates tumor hypoxia. It has been one of the modalities in cancer treatment; therefore its optimization is important. In this experimental study, no cancer enhancing effect was seen during the course of HBO therapy; however, post therapy there was an accelerated growth and progression of tumor. HBO treated mice lived shorter and the response to therapy was dose & tumor volume dependent. HBO therapy probably exert its effect on the cancer proliferating cells through multiple pathways such as increased DNA damage, apoptosis & geno-toxicity leading to slow cancer progression while post therapy tumorigenic effect could be due to impaired DNA repair mechanism, mutagenic effect & aneuploidy as well as altered blood supply & nutrients. Tumor growth reached plateau with time and this finding validated theoretical model predicting tumor reaching an asymptotic limit. While, marked asymmetry observed in tumor volume progression or cancer cell proliferation rate in each of the experimental C3H mouse suggested a need for an alternate small animal pre-clinical cancer therapeutic model. [ABSTRACT FROM AUTHOR]

Extracellular amyloid-β (Aβ) plaques and intracellular neurofibrillary tangles constitute the major neuropathological hallmarks of Alzheimer's disease (AD). It is now apparent that parenchymal Aβ plaque deposition precedes behavioral signs of disease by several years. The development of agents that can target these plaques may be useful as diagnostic or therapeutic tools. In this study, we synthesized an Aβ-targeted lipid conjugate, incorporated it in stealth liposomal nanoparticles and tested their ability to bind amyloid plaque deposits in an AD mouse model. The results show that the particles maintain binding profiles to synthetic Aβ aggregates comparable to the free ligand, and selectively bind Aβ plaque deposits in brain tissue sections of an AD mouse model (APP/PSEN1 transgenic mice) with high efficiency. When administered intravenously, these long circulating nanoparticles appear to cross the blood-brain barrier and bind to Aβ plaque deposits, labeling parenchymal amyloid deposits and vascular amyloid characteristic of cerebral amyloid angiopathy. [ABSTRACT FROM AUTHOR]

Tumors convert conventional CD4+ T cells into induced CD4+CD25+FoxP3+ T regulatory (iTreg) cells that serve as an effective means of immune evasion. Therefore, the blockade of conventional CD4+ T cell conversion into iTreg cells represents an attractive target for improving the efficacy of various immunotherapeutic approaches. Using a novel form of 4-1BBL molecule, SA-4-1BBL, we previously demonstrated that costimulation via 4-1BB receptor renders both CD4+ and CD8+ T effector (Teff) cells refractory to inhibition by Treg cells and increased intratumoral Teff/Treg cell ratio that correlated with therapeutic efficacy in various preclinical tumor models. Building on these studies, we herein show for the first time, to our knowledge, that signaling through 4-1BB inhibits antigen- and TGF-β-driven conversion of naïve CD4+FoxP3- T cells into iTreg cells via stimulation of IFN-c production by CD4+FoxP3- T cells. Importantly, treatment with SA-4-1BBL blocked the conversion of CD4+FoxP32 T cells into Treg cells by EG.7 tumors. Taken together with our previous studies, these results show that 4-1BB signaling negatively modulate Treg cells by two distinct mechanisms: I) inhibiting the conversion of CD4+FoxP3- T cells into iTreg cells and ii) endowing Teff cells refractory to inhibition by Treg cells. Given the dominant role of Treg cells in tumor immune evasion mechanisms, 4-1BB signaling represents an attractive target for favorably tipping the Teff:Treg balance toward Teff cells with important implications for cancer immunotherapy. [ABSTRACT FROM AUTHOR]

Clostridium difficile is considered to be one of the most important causes of health care-associated infections worldwide. In order to understand more fully the adaptive response of the organism to stressful conditions, we examined transcriptional changes resulting from a clinically relevant heat stress (41°C versus 37°C) in C. difficile strain 630 and identified 341 differentially expressed genes encompassing multiple cellular functional categories. While the transcriptome was relatively resilient to the applied heat stress, we noted upregulation of classical heat shock genes including the groEL and dnaK operons in addition to other stress-responsive genes. Interestingly, the flagellin gene (fliC) was downregulated, yet genes encoding the cell-wall associated flagellar components were upregulated suggesting that while motility may be reduced, adherence - to mucus or epithelial cells - could be enhanced during infection. We also observed that a number of phage associated genes were downregulated, as were genes associated with the conjugative transposon Tn5397 including a group II intron, thus highlighting a potential decrease in retromobility during heat stress. These data suggest that maintenance of lysogeny and genome wide stabilisation of mobile elements could be a global response to heat stress in this pathogen. [ABSTRACT FROM AUTHOR]

An infection represents a highly dynamic process involving complex biological responses of the host at many levels. To describe such processes at a global level, we recorded gene expression changes in mouse lungs after a non-lethal infection with influenza A virus over a period of 60 days. Global analysis of the large data set identified distinct phases of the host response. The increase in interferon genes and up-regulation of a defined NK-specific gene set revealed the initiation of the early innate immune response phase. Subsequently, infiltration and activation of T and B cells could be observed by an augmentation of T and B cell specific signature gene expression. The changes in B cell gene expression and preceding chemokine subsets were associated with the formation of bronchus-associated lymphoid tissue. In addition, we compared the gene expression profiles from wild type mice with Rag2 mutant mice. This analysis readily demonstrated that the deficiency in the T and B cell responses in Rag2 mutants could be detected by changes in the global gene expression patterns of the whole lung. In conclusion, our comprehensive gene expression study describes for the first time the entire host response and its kinetics to an acute influenza A infection at the transcriptome level. [ABSTRACT FROM AUTHOR]

Background: β-Galactosidase is a vital enzyme with diverse application in molecular biology and industries. It was covalently attached onto functionalized graphene nano-sheets for various analytical applications based on lactose reduction. Methodology/Principal Findings: Response surface methodology based on Box-Behnken design of experiment was used for determination of optimal immobilization conditions, which resulted in 84.2% immobilization efficiency. Native and immobilized functionalized graphene was characterized with the help of transmission and scanning electron microscopy, followed by Fourier transform infrared (FTIR) spectroscopy. Functionalized graphene sheets decorated with islands of immobilized enzyme were evidently visualized under both transmission and scanning electron microscopy after immobilization. FTIR spectra provided insight on various chemical interactions and bonding, involved during and after immobilization. Optimum temperature and energy of activation (Ea) remains unchanged whereas optimum pH and Km were changed after immobilization. Increased thermal stability of enzyme was observed after conjugating the enzyme with functionalized graphene. Significance: Immobilized β-galactosidase showed excellent reusability with a retention of more than 92% enzymatic activity after 10 reuses and an ideal performance at broad ranges of industrial environment. [ABSTRACT FROM AUTHOR]