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Summary: Ubiquitination of mitochondrial proteins provides a basis for the downstream recruitment of mitophagy machinery, yet whether ubiquitination of the machinery itself contributes to mitophagy is unknown. Here, we show that K63-linked polyubiquitination of the key mitophagy regulator TBK1 is essential for its mitophagy functions. This modification is catalyzed by the ubiquitin ligase TRIM5α and is required for TBK1 to interact with and activate a set of ubiquitin-binding autophagy adaptors including NDP52, p62/SQSTM1, and NBR1. Autophagy adaptors, along with TRIM27, enable TRIM5α to engage with TBK1 following mitochondrial damage. TRIM5α’s ubiquitin ligase activity is required for the accumulation of active TBK1 on damaged mitochondria in Parkin-dependent and Parkin-independent mitophagy pathways. Our data support a model in which TRIM5α provides a mitochondria-localized, ubiquitin-based, self-amplifying assembly platform for TBK1 and mitophagy adaptors that is ultimately necessary for the recruitment of the core autophagy machinery.

IntroductionNon-Hodgkin Lymphoma (NHL) is a heterogeneous lymphoproliferative malignancy with B cell origin. Combinatorial treatment of rituximab, cyclophsphamide, hydroxydaunorubicin, oncovin, prednisone (R-CHOP) is the standard treatment regimen for NHL, yielding a complete remission (CR) rate of 40-50%. Unfortunately, considerable patients undergo relapse after CR or initial treatment, resulting in poor clinical implications. Patient’s response to chemotherapy varies widely from static disease to cancer recurrence and later is primarily associated with the development of multi-drug resistance (MDR). The immunosuppressive cells within the tumor microenvironment (TME) have become a crucial target for improving the therapy efficacy. However, a better understanding of their involvement is needed for distinctive response of NHL patients after receiving chemotherapy to design more effective front-line treatment algorithms based on reliable predictive biomarkers.MethodsPeripheral blood from 61 CD20+ NHL patients before and after chemotherapy was utilized for immunophenotyping by flow-cytometry at different phases of treatment. In-vivo and in-vitro doxorubicin (Dox) resistance models were developed with murine Dalton’s lymphoma and Jurkat/Raji cell-lines respectively and impact of responsible immune cells on generation of drug resistance was studied by RT-PCR, flow-cytometry and colorimetric assays. Gene silencing, ChIP and western blot were performed to explore the involved signaling pathways.ResultsWe observed a strong positive correlation between elevated level of CD33+CD11b+CD14+CD15- monocytic MDSCs (M-MDSC) and MDR in NHL relapse cohorts. We executed the role of M-MDSCs in fostering drug resistance phenomenon in doxorubicin-resistant cancer cells in both in-vitro, in-vivo models. Moreover, in-vitro supplementation of MDSCs in murine and human lymphoma culture augments early expression of MDR phenotypes than culture without MDSCs, correlated well with in-vitro drug efflux and tumor progression. We found that MDSC secreted cytokines IL-6, IL-10, IL-1β are the dominant factors elevating MDR expression in cancer cells, neutralization of MDSC secreted IL-6, IL-10, IL-1β reversed the MDR trait. Moreover, we identified MDSC secreted IL-6/IL-10/IL-1β induced STAT1/STAT3/NF-κβ signaling axis as a targeted cascade to promote early drug resistance in cancer cells.ConclusionOur data suggests that screening patients for high titre of M-MDSCs might be considered as a new potential biomarker and treatment modality in overcoming chemo-resistance in NHL patients.

Abstract Background Multiple sclerosis (MS) is characterized by neuroinflammation and demyelination orchestrated by activated neuroglial cells, CNS infiltrating leukocytes, and their reciprocal interactions through inflammatory signals. An inflammatory stimulus triggers inducible nitric oxide synthase (NOS2), a pro-inflammatory marker of microglia/macrophages (MG/Mφ) to catalyze sustained nitric oxide production. NOS2 during neuroinflammation, has been associated with MS disease pathology; however, studies dissecting its role in demyelination are limited. We studied the role of NOS2 in a recombinant β-coronavirus-MHV-RSA59 induced neuroinflammation, an experimental animal model mimicking the pathological hallmarks of MS: neuroinflammatory demyelination and axonal degeneration. Objective Understanding the role of NOS2 in murine-β-coronavirus-MHV-RSA59 demyelination. Methods Brain and spinal cords from mock and RSA59 infected 4–5-week-old MHV-free C57BL/6 mice (WT) and NOS2-/- mice were harvested at different disease phases post infection (p.i.) (day 5/6-acute, day 9/10-acute-adaptive and day 30-chronic phase) and compared for pathological outcomes. Results NOS2 was upregulated at the acute phase of RSA59-induced disease in WT mice and its deficiency resulted in severe disease and reduced survival at the acute-adaptive transition phase. Low survival in NOS2-/- mice was attributed to (i) high neuroinflammation resulting from increased accumulation of macrophages and neutrophils and (ii) Iba1 + phagocytic MG/Mφ mediated-early demyelination as observed at this phase. The phagocytic phenotype of CNS MG/Mφ was confirmed by significantly higher mRNA transcripts of phagocyte markers-CD206, TREM2, and Arg1 and double immunolabelling of Iba1 with MBP and PLP. Further, NOS2 deficiency led to exacerbated demyelination at the chronic phase as well. Conclusion Taken together the results imply that the immune system failed to control the disease progression in the absence of NOS2. Thus, our observations highlight a protective role of NOS2 in murine-β-coronavirus induced demyelination.

Background: Bone remodeling is a balanced, dynamic function of two counteractive processes: osteoblasts-mediated bone formation and osteoclasts-dependent bone resorption. An imbalance between these two processes causes bone diseases. Herein, the effects and molecular modus operandi of plumbagin- a bioactive phytometabolite from Plumbago sp. have been investigated on osteoblast and osteoclast differentiation. Material and Methods: Cell viability assay and Alizarin red-S staining assessed the plumbagin effects on pre-osteoblastic MC3T3-E1 cell viability and differentiation. Alkaline phosphatase activity, the osteoblast-specific genes, and protein expression were investigated by colorimetry, quantitative-PCR and immunoblotting, respectively. TRAP assay and osteoclast-specific proteins' expression assessed plumbagin's effect on osteoclastogenesis. During plumbagin-driven osteoclastogenesis, we used paired-end RNA-sequencing data to identify the key up-/down-regulated DEGs, validation by qPCR, and bioinformatics-based pathway analysis. Results: Plumbagin at a very low dose (0.25 µM) significantly promoted osteoblast differentiation, as evident from alkaline phosphatase, matrix mineralization, and enhanced expression of osteoblast-specific markers, including Runx2 and its target genes. The reduced TRAP activity, osteoclast-specific protein expression, and RNA sequencing suggested RANKL-induced osteoclastogenesis inhibition by plumbagin at 2 µM.. Conclusion: Plumbagin significantly enhances osteoblast differentiation by enhanced matrix mineralization and osteoblast-specific gene expression but suppresses the RANKL-induced osteoclastogenesis by regulating key osteoclast-specific DEGs and protein expression. Plumbagin can thus significantly influence bone biology and affect bone diseases.

IntroductionCurrent anti-leukemic chemotherapies with multiple targets suffer from side effects. Synthetic drugs with huge off-target effects are detrimental to leukemic patients. Therefore, natural plant-based products are being increasingly tested for new anti-leukemic therapy with fewer or no side effects. Herein, we report the effect of ethanolic olive leaves extract (EOLE) on the K562 cell line and on the bone marrow (BM) of N-ethyl-N-nitrosourea (ENU)-induced leukemic mice.MethodsUsing standard methodologies, we assessed viability, chromatin condensation, and induction of apoptosis in EOLE-treated K562 cells in-vitro. The anti-leukemic activity of EOLE was assayed by measuring ROS, levels of various cytokines, expression of iNOS and COX-2 gene, and changes in the level of important apoptosis regulatory and cell signaling proteins in-vivo. ResultK562 cells underwent apoptotic induction after exposure to EOLE. In the BM of leukemic mice, EOLE therapy decreased the number of blast cells, ROS generation, and expression of NF-κB and ERK1/2. IL-6, IL-1β, TNF-α, iNOS, and COX-2 were among the inflammatory molecules that were down-regulated by EOLE therapy. Additionally, it decreased the expression of anti-apoptotic proteins BCL2A1, BCL-xL, and MCL-1 in the BM of leukemic mice.DiscussionChronic inflammation and anomalous apoptotic mechanism both critically contribute to the malignant transformation of cells. Inflammation in the tumor microenvironment promotes the growth, survival, and migration of cancer cells, accelerating the disease. The current investigation showed that EOLE treatment reduces inflammation and alters the expression of apoptosis regulatory protein in the BM of leukemic mice, which may halt the progression of the disease.

Background: COVID-19 pandemic induced lockdown and the social restrictions had a profound impact on the circadian rhythm driven sleep-wake schedule of the home confined population. Aims and Objective: Our study explored the effect of COVID-19 pandemic induced lockdown on sleep wake pattern and personal well-being of medical students of West Bengal Materials and Methods: An observational, questionnaire-based online survey was conducted using the URL linked Google form. The online survey was conducted through social media platforms as per CHERRIES checklist guideline. Information on demographic profile, before and after lockdown sleep-wake pattern, social wellbeing and general lifestyle was obtained. Any student with any sleeping disorder or on drugs was excluded. Mean (SE), median, range, proportion was calculated as per attributes. Fisher’s exact chi square test and paired t test was done to test association and to compare means of the attributes. Results: The mean age of students was 20.26 years. The average daily time spent on electronic media got doubled during lockdown. Sleep disorder was reported by 53.0% male and 47.0% students. More than half of respondents reported increased napping during daytime, irregular timing for meal intake and low mood. Anxiety was present in every two out of three participants. Females gained more weight during lockdown. The gender difference in sleep duration became significant. Lockdown effect on the average daily media time was significant. The sleep disturbance at night and physical inactivity was significantly more among female students. Conclusion: COVID-19 lockdown leads to delayed sleep-wake cycle, irregular meal timings and excessive digital exposure among medical students with gender based differential impact.

In the era of antibiotic resistance, in silico prediction of bacterial resistome profiles, likely to be associated with inactivation of new potential antibiotics is of utmost importance. Despite this, to the best of our knowledge, no tool exists for such prediction. Therefore, under the rationale that drugs with similar structures have similar resistome profiles, we developed two models, a deterministic model and a stochastic model, to predict the bacterial resistome likely to neutralize uncharacterized but potential chemical structures. The current version of the tool involves the prediction of a resistome for Escherichia coli and Pseudomonas aeruginosa. The deterministic model on omitting two diverse but relatively less characterized drug classes, polyketides and polypeptides showed an accuracy of 87%, a sensitivity of 85%, and a precision of 89%, whereas the stochastic model predicted antibiotic classes of the test set compounds with an accuracy of 72%, a sensitivity of 75%, and a precision of 83%. The models have been implemented in both a standalone package and an online server, uCAREChemSuiteCLI and uCARE Chem Suite, respectively. In addition to resistome prediction, the online version of the suite enables the user to visualize the chemical structure, classify compounds in 19 predefined drug classes, perform pairwise alignment, and cluster with database compounds using a graphical user interface. Availability: uCARE Chem Suite can be browsed at: https://sauravsaha.shinyapps.io/ucarechemsuite2/, and uCAREChemSuiteCLI can be installed from:1. CRAN (https://cran.r-project.org/package=uCAREChemSuiteCLI) and2. GitHub (https://github.com/sauravbsaha/uCAREChemSuiteCLI).

As effector innate immune cells and as a host to the protozoan parasite Leishmania, macrophages play a dual role in antileishmanial immunoregulation. The 2 key players in this immunoregulation are the macrophage-expressed microRNAs (miRNAs) and the macrophage-secreted cytokines. miRNAs, as small noncoding RNAs, play vital roles in macrophage functions including cytokines and chemokines production. In the reverse direction, Leishmania-regulated cytokines alter miRNAs expression to regulate the antileishmanial functions of macrophages. The miRNA patterns vary with the time and stage of infection. The cytokine-regulated macrophage miRNAs not only help parasite elimination or persistence but also regulate cytokine production from macrophages. Based on these observations, we propose a novel immunoregulatory framework as a scientific rationale for antileishmanial therapy.

Background: Social distancing is important preventive strategy to prevent spreading of SARSCOV-2. The medical faculties are continuing the teaching-learning session using different online platform. Aims and Objective: This online survey aims at exploring the different aspect of online teaching effects among medical students. Materials and Methods: Forty-four validated questions in the Google survey form were distributed among the MBBS students of different medical colleges of West Bengal in this cross-sectional study, and analysed. Results: The responses from 416 participants show that internet-based teaching has been adopted by 96.4% of colleges. Practical class and classes regarding clinical exposure are the two most neglected section of the MBBS curriculum during this pandemic. Total teaching hours per day also drastically reduced from mean of 5.56 h/ day in pre-pandemic time to 1.95 h/ day during this pandemic. Out of 69.7% of the participants who faced difficulty in following online classes, majority (56.25%) was due to technical problems including issues related to internet connectivity and software. Participants were of opinion that current session should be extended on an average of 3 months as the practical and clinical classes are way behind in syllabus completion. According to 72.35% of participants, online classes are less effective than the offline classes. Conclusions: In nutshell, online teaching activities cannot replace the classical classroom teaching, though it is a cost-effective and viable method during this pandemic period.

Abstract Artemisinin is the frontline fast-acting anti-malarial against P. falciparum. Emergence and spread of resistant parasite in eastern-India poses a threat to national malaria control programs. Therefore, the objective of our study is to evaluate the artesunate-sulfadoxine-pyrimethamine efficacy in Central India. 180 monoclonal P. falciparum-infected patients received standard ASSP therapy during August 2015–January 2017, soon after diagnosis and monitored over next 42-days. Artemisinin-resistance was assessed through in-vivo parasite clearance half-life (PC1/2), ex-vivo ring-stage survivability (RSA), and genome analysis of kelch13 and other candidate gene (pfcrt, pfmdr1, pfatpase 6, pfdhfr and pfdhps). Of 180 P. falciparum positive patients, 9.5% showed increased PC1/2 (> 5.5 h), among them eleven isolates (6.1%) showed reduced sensitivity to RSA. In 4.4% of cases, parasites were not cleared by 72 h and showed prolonged PC1/2(5.6 h) (P

In malaria, anti-parasite immune response of the host may lead to dysregulated inflammation causing severe neuropathology arising from extensive damage to the Blood Brain Barrier (BBB). Use of anti-malarial drugs alone can control parasitemia and reduce inflammation but it cannot reduce pathology if chronic inflammation has already set in. In the present study, we have tested the efficacy of a new oral artemsinin based combination therapy (ACT) regimen using a combination of anti-malarial compounds like nanoartemisinin and nanoallylated-chalcone9 [{1-(4-Chlorophenyl)-3-[3-methoxy-4-(prop-2-en-1-yloxy) phenyl]-prop-2-en-1-one}]given together with anti-inflammatory-cum- anti-malarial compounds like nanoandrographolide and nanocurcumin to C57BL/6 mice infected with P. berghei ANKA. Untreated infected mice developed Experimental Cerebral Malaria (ECM) and died between 10 to 12 days after infection from severe BBB damage. We observed that oral treatments with nanoartemisinin or nano allylated chalcone 9 or nanoandrographolide alone, for 4 days after the onset of ECM, delayed the development of severe neurolopathology but could not prevent it. Nanocurcumin treatment for 4 days on the other hand, prevented damage to the BBB but the mice died because of hyperparasitemia. A single time oral administration of our ACT controlled blood parasitemia and prevented damage to the BBB, but recrudescence occurred due to persistence of parasites in the spleen. However the recrudescent parasites failed to induce ECM and BBB damage, leading to prolonged survival of the animals. A second time treatment at the start of recrudescence led to complete parasite clearance and survival of mice without pathology or parasitemia for 90 days. FACS analysis of spleen cells and gene expression profile in brain and spleen as well as quantitation of serum cytokine by ELISA showed that P. berghei ANKA infection in C57Bl/6 mice leads to a Th1-skewed immune response that result in severe inflammation and early death from ECM. Oral treatment with our ACT prevented a heightened pro-inflammatory response by modulating the Th1, Th2 and Treg immune responses and prevented ECM and death.

Neurotropic mouse hepatitis virus (MHV-A59/RSA59) infection in mice induces acute neuroinflammation due to direct neural cell dystrophy, which proceeds with demyelination with or without axonal loss, the pathological hallmarks of human neurological disease, Multiple sclerosis (MS). Recent studies in the RSA59-induced neuroinflammation model of MS showed a protective role of CNS-infiltrating CD4+ T cells compared to their pathogenic role in the autoimmune model. The current study further investigated the molecular nexus between CD4+ T cell-expressed CD40Ligand and microglia/macrophage-expressed CD40 using CD40L-/- mice. Results demonstrate CD40L expression in the CNS is modulated upon RSA59 infection. We show evidence that CD40L-/- mice are more susceptible to RSA59 induced disease due to reduced microglia/macrophage activation and significantly dampened effector CD4+ T recruitment to the CNS on day 10 p.i. Additionally, CD40L-/- mice exhibited severe demyelination mediated by phagocytic microglia/macrophages, axonal loss, and persistent poliomyelitis during chronic infection, indicating CD40-CD40L as host-protective against RSA59-induced demyelination. This suggests a novel target in designing prophylaxis for virus-induced demyelination and axonal degeneration, in contrast to immunosuppression which holds only for autoimmune mechanisms of inflammatory demyelination.

Abstract Background Ras are small cellular GTPases which regulate diverse cellular processes. It has three isoforms: H-Ras, K-Ras, and N-Ras. Owing to the N-terminus (1–165 residues) sequence homology these isoforms were thought to be functionally redundant. However, only K-Ras-deficient mice but not H-Ras- and N-Ras-deficient mice show embryonic lethality. Similarly, mutations in a given Ras isoform are associated with a particular type of cancer. Moreover, we have previously reported that Ras isoforms perform unique functions in Leishmania major infection. Thus, Ras isoforms are implicated to have signaling and functional specificity but the mechanism remains to be elucidated. Result Using CD40 as a model receptor, we showed that depending on the strength of signaling, specific Ras isoforms are activated. Weak CD40 signal activates N-Ras, whereas strong signal activates H-Ras and K-Ras. Additionally, we showed that suppression of N-Ras expression reduced CD40-induced extracellular signal–regulated kinase-1/2 (ERK-1/2) activation and Interleukin (IL)-10 production; whereas suppression of H-Ras or K-Ras reduced CD40-induced p38 mitogen-activated protein kinase (p38MAPK) activation and IL-12 production. Furthermore, we showed that Ras isoforms have activator (GEF) specificity as weak CD40 signal-activated N-Ras requires Sos-1/2 whereas strong CD40 signal-activated H-Ras/K-Ras requires Ras-GRP as the guanine-nucleotide exchange factor (GEF) inducing ERK-1/2- or p38MAPK-mediated IL-10 or IL-12 productions, respectively, in macrophages. Silencing of syk reduced CD40-induced N-Ras activation but silencing of lyn inhibited H-Ras and K-Ras activation. In CD40 signaling, Ras isoforms also showed effector specificity; while H-Ras and K-Ras showed specificity for phosphatidyl inositol-3 kinase activation at high dose of CD40 stimulation, N-Ras primarily associated with Raf-1 at low dose of CD40 stimulation. Moreover, fractal analysis showed that functional site surface roughness for H-Ras (SurfaceFD = 2.39) and K-Ras (SurfaceFD = 2.39) are similar but significantly different from N-Ras (SurfaceFD = 2.25). Conclusion The activator and effector specificities of Ras isoforms in CD40 signaling indicated their differential involvement in CD40 pathway and in maintaining the reciprocity. Our observations reveal Ras-regulated signaling outcome and its potential for developing Ras isoform-targeted immunotherapy and prophylaxis. Graphical abstract

TRIM5α is a key cross-species barrier to retroviral infection, with certain TRIM5 alleles conferring increased risk of HIV-1 infection in humans. TRIM5α is best known as a species-specific restriction factor that directly inhibits the viral life cycle. Additionally, it is also a pattern-recognition receptor (PRR) that activates inflammatory signaling. How TRIM5α carries out its multi-faceted actions in antiviral defense remains incompletely understood. Here, we show that proteins required for autophagy, a cellular self-digestion pathway, play an important role in TRIM5α's function as a PRR. Genetic depletion of proteins involved in all stages of the autophagy pathway prevented TRIM5α-driven expression of NF-κB and AP1 responsive genes. One of these genes is the preeminent antiviral cytokine interferon β (IFN-β), whose TRIM5-dependent expression was lost in cells lacking the autophagy proteins ATG7, BECN1, and ULK1. Moreover, we found that the ability of TRIM5α to stimulate IFN-β expression in response to recognition of a TRIM5α-restricted HIV-1 capsid mutant (P90A) was abrogated in cells lacking autophagy factors. Stimulation of human macrophage-like cells with the P90A virus protected them against subsequent infection with an otherwise resistant wild type HIV-1 in a manner requiring TRIM5α, BECN1, and ULK1. Mechanistically, TRIM5α was attenuated in its ability to activate the kinase TAK1 in autophagy deficient cells, and both BECN1 and ATG7 contributed to the assembly of TRIM5α-TAK1 complexes. These data demonstrate a non-canonical role for the autophagy machinery in assembling antiviral signaling complexes and in establishing a TRIM5α-dependent antiviral state.

Summary: CD40-Ligand (CD40L)-CD40 interaction regulates immune responses against pathogens, autoantigens, and tumor and transplantation antigens. Single amino acid mutations within the 115–155 amino acids stretch, which is responsible for CD40L functions, result in XIgM syndrome. We hypothesize that each of these amino acids of CD40L encodes specific message that, when decoded by CD40 signaling, induces a specific profile of functions. We observed that every single substitution in the XIgM-related amino acids in the 115–155 41-mer peptide in CD40L selectively altered CD40 signaling and effector functions—cytokine productions, HMGCoA reductase, ceramide synthase, inducible nitric oxide synthase and arginase expression, survival of B cells, and control of Leishmania infection and anti-leishmanial T cell response—suggesting residue-specific encoding of a distinct set of messages that collectively define CD40L pleiotropy, serve as a target for engineering the ligand to generate superagonists as immunotherapeutic, and implicate the evolutionary diversification of functions among the ligands in a protein superfamily.

IL-27 is a cytokine that exerts diverse effects on the cells of innate and adaptive immune systems. Chiefly expressed in macrophages and dendritic cells during the early phase of Leishmania infection, IL-27 contributes to the protection against L. major infection but suppresses the protective Th1 response against L. donovani, L. infantum, L. amazonensis and L. braziliensis infections, suggesting its functional duality. During the late stage of Leishmania infection, IL-27 limits the immunopathogenic reactions and tissue damages. Herein, we analyze the mechanism of the functional duality of IL-27 in the resistance or susceptibility to Leishmania infection, prompting IL-27 for anti-Leishmanial therapy.

Tumor progression in the host leads to severe impairment of intrathymic T-cell differentiation/maturation, leading to the paralysis of cellular anti-tumor immunity. Such suppression manifests the erosion of CD4+CD8+ double-positive (DP) immature thymocytes and a gradual increase in CD4−CD8− double negative (DN) early T-cell progenitors. The impact of such changes on the T-cell progenitor pool in the context of cancer remains poorly investigated. Here, we show that tumor progression blocks the transition of Lin−Thy1.2+CD25+CD44+c-KitlowDN2b to Lin−Thy1.2+CD25+CD44−c-Kit−DN3 in T-cell maturation, instead leading to DN2-T-cell differentiation into dendritic cells (DC). We observed that thymic IL-10 expression is upregulated, particularly at cortico-medullary junctions (CMJ), under conditions of progressive disease, resulting in the termination of IL-10Rhigh DN2-T-cell maturation due to dysregulated expression of Notch1 and its target, CCR7 (thus restricting these cells to the CMJ). Intrathymic differentiation of T-cell precursors in IL-10−/− mice and in vitro fetal thymic organ cultures revealed that IL-10 promotes the interaction between thymic stromal cells and Notch1low DN2-T cells, thus facilitating these DN2-T cells to differentiate toward CD45+CD11c+MHC-II+ thymic DCs as a consequence of activating the Ikaros/IRF8 signaling axis. We conclude that a novel function of thymically-expressed IL-10 in the tumor-bearing host diverts T-cell differentiation toward a DC pathway, thus limiting the protective adaptive immune repertoire.

Summary: Hypoxia-inducible factor-1 alpha (HIF-1α) is considered a global regulator of cellular metabolism and innate immune cell functions. Intracellular pathogens such as Leishmania have been reported to manipulate host cell metabolism. Herein, we demonstrate that myeloid cells from myeloid-restricted HIF-1α-deficient mice and individuals with loss-of-function HIF1A gene polymorphisms are more susceptible to L. donovani infection through increased lipogenesis. Absence of HIF-1α leads to a defect in BNIP3 expression, resulting in the activation of mTOR and nuclear translocation of SREBP-1c. We observed the induction of lipogenic gene transcripts, such as FASN, and lipid accumulation in infected HIF-1α−/− macrophages. L. donovani-infected HIF-1α-deficient mice develop hypertriglyceridemia and lipid accumulation in splenic and hepatic myeloid cells. Most importantly, our data demonstrate that manipulating FASN or SREBP-1c using pharmacological inhibitors significantly reduced parasite burden. As such, genetic deficiency of HIF-1α is associated with increased lipid accumulation, which results in impaired host-protective anti-leishmanial functions of myeloid cells. : Mesquita et al. show that genetic deficiency of HIF-1α in the myeloid compartment promotes de novo lipogenesis through the BNIP3/mTOR/SREBP-1c axis. This is associated with higher susceptibility to Leishmania donovani infection, which is reduced upon pharmacological inhibition of fatty acid synthesis. Keywords: HIF-1α, visceral leishmaniasis, macrophages, lipogenesis, SREBP-1c, FASN, acetate, myeloid cells

BackgroundThe disturbance of host metabolic pathways by Leishmania parasites has crucial consequences for the activation status of immune cells and the outcome of infection. Glutamine has been described as an immunomodulatory amino acid, yet its role during Leishmania infection is still unknown.MethodsWe performed transcriptomics in uninfected and L. donovani-infected macrophages 6 hours post-infection. Glutamine quantification by HPLC was assessed in the supernatant of macrophages throughout the infection course. For experimental L. donovani infections, mice were infected with 1.0 x 108 stationary L. donovani promastigotes. Glutaminase (GLS) chemical inhibition was performed using BPTES and glutamine was administered throughout infection. For combined therapy experiment, a daily administration of miltefosine and glutamine was performed by oral gavage. Parasite burden was determined using a Taqman-based assay. Immune cell phenotyping and cytotoxicity were performed in splenic cells using flow cytometry.FindingsWe show that glutamine is essential for the control of L. donovani infection. Transcriptomic analysis of L. donovani-infected macrophages demonstrated an upregulation of genes involved in glutamine metabolism. Pharmacological inhibition of glutaminolysis significantly increased the susceptibility to infection, accompanied by an increased recruitment of anti-inflammatory myeloid cells and impaired T cell responses. Remarkably, the supplementation of glutamine to mice infected with L. donovani during miltefosine treatment potentiates parasite clearance through the development of a more effective anti-Leishmania adaptive immune response.ConclusionsOur data indicates that dietary glutamine supplementation may act as a promising adjuvant for the treatment of visceral leishmaniasis.

Autophagy is a cellular degradative process that has multiple important actions in cancer. Autophagy modulation is under consideration as a promising new approach to cancer therapy. However, complete autophagy dysregulation is likely to have substantial undesirable side effects. Thus, more targeted approaches to autophagy modulation may prove clinically beneficial. One potential avenue to achieving this goal is to focus on the actions of tripartite motif-containing protein family members (TRIMs). TRIMs have key roles in an array of cellular processes, and their dysregulation has been extensively linked to cancer risk and prognosis. As detailed here, emerging data shows that TRIMs can play important yet context-dependent roles in controlling autophagy and in the selective targeting of autophagic substrates. This review covers how the autophagy-related actions of TRIM proteins contribute to cancer and the possibility of targeting TRIM-directed autophagy in cancer therapy.

Leishmaniasis is the second largest parasitic killer disease caused by the protozoan parasite Leishmania, transmitted by the bite of sand flies. It's endemic in the eastern India with 165.4 million populations at risk with the current drug regimen. Three forms of leishmaniasis exist in which cutaneous is the most common form caused by Leishmania major. Trypanothione Reductase (TryR), a flavoprotein oxidoreductase, unique to thiol redox system, is considered as a potential target for chemotherapy for trypanosomatids infection. It is involved in the NADPH dependent reduction of Trypanothione disulphide to Trypanothione. Similarly, is Tryparedoxin Peroxidase (Txnpx), for detoxification of peroxides, an event pivotal for survival of Leishmania in two disparate biological environment. Fe-S plays a major role in regulating redox balance. To check for the closeness between human homologs of these proteins, we have carried the molecular clock analysis followed by molecular modeling of 3D structure of this protein, enabling us to design and test the novel drug like molecules. Molecular clock analysis suggests that human homologs of TryR i.e. Glutathione Reductase and Txnpx respectively are highly diverged in phylogenetic tree, thus, they serve as good candidates for chemotherapy of leishmaniasis. Furthermore, we have done the homology modeling of TryR using template of same protein from Leishmania infantum (PDB ID: 2JK6). This was done using Modeller 9.18 and the resultant models were validated. To inhibit this target, molecular docking was done with various screened inhibitors in which we found Taxifolin acts as common inhibitors for both TryR and Txnpx. We constructed the protein-protein interaction network for the proteins that are involved in the redox metabolism from various Interaction databases and the network was statistically analysed. Keywords: Trypanothione Reductase, Tryparedoxin Peroxidase, L.major, Homology modeling, Molecular clock analysis, Network analysis

Abstract The disease tuberculosis is fatal if untreated. It is caused by the acid‐fast bacilli Mycobacterium tuberculosis. Mycobacterium resides and replicates within the alveolar macrophages, causing inflammation and granuloma, wherein macrophage‐T cell interactions enhance the inflammation‐causing pulmonary caseous lesions. The first interactions between Mycobacterium and the receptors on macrophages decide the fate of Mycobacterium because of phagolysosomal impairments and the expression of several miRNAs, which may regulate CD40 expression on macrophages. While the altered phagolysosomal functions impede antigen presentation to the T cell‐expressed antigen receptor, the interactions between the macrophage‐expressed CD40 and the T cell‐expressed CD40‐ligand (CD40L or CD154) provide signals to T cells and Mycobacterium‐infected macrophages. These two functions significantly influence the resolution or persistence of Mycobacterium infection. CD40 controls T‐cell polarisation and host‐protective immunity by eliciting interleukin‐12p40, nitric oxide, reactive oxygen species and IFN‐γ production. Indeed, CD40‐deficient mice succumb to low‐dose aerosol infection with Mycobacterium because of deficient interleukin (IL)‐12 production leading to impaired IFN‐γ‐secreting T‐cell response. In contrast, despite generating fewer granulomas, the CD40L‐deficient mice developed anti‐mycobacterial T‐cell responses to the levels observed in the wild‐type mice. These host‐protective responses are significantly subdued by the Mycobacterium‐infected macrophage produced TGF‐β and IL‐10, which promote pro‐mycobacterial T‐cell responses. The CD40‐CD40L‐induced counteractive immune responses against Mycobacterium thus present a conundrum that we explain here with a reconciliatory hypothesis. Experimental validation of the hypothesis will provide a rationale for designing anti‐tubercular immunotherapy.

Leishmaniasis is a neglected protozoan parasitic disease that occurs in 88 countries but a vaccine is unavailable. Vaccination with live, killed, attenuated (physically or genetically) Leishmania have met with limited success, while peptide-, protein-, or DNA-based vaccines showed promise only in animal models. Here, we critically assess several technical issues in vaccination and expectation of a host-protective immune response. Several studies showed that antigen presentation during priming and triggering of the same cells in infected condition are not comparable. Altered proteolytic processing, antigen presentation, protease-susceptible sites, and intracellular expression of pathogenic proteins during Leishmania infection may vary dominant epitope selection, MHC-II/peptide affinity, and may deter the reactivation of desired antigen-specific T cells generated during priming. The robustness of the memory T cells and their functions remains a concern. Presentation of the antigens by Leishmania-infected macrophages to antigen-specific memory T cells may lead to change in the T cells’ functional phenotype or anergy or apoptosis. Although cells may be activated, the peptides generated during infection may be different and cross-reactive to the priming peptides. Such altered peptide ligands may lead to suppression of otherwise active antigen-specific T cells. We critically assess these different immunological issues that led to the non-availability of a vaccine for human use.