Myotubularin-related protein 6 is an ion channel-associated pro-leishmanial phosphatase.

Residing obligatorily as amastigotes within the mammalian macrophages, the parasite Leishmania donovani inflicts the potentially fatal, globally re-emerging disease visceral leishmaniasis (VL) by altering intracellular signaling through kinases and phosphatases. Because the phosphatases that modulate the VL outcome in humans remained unknown, we screened a human phosphatase siRNA-library for anti-leishmanial functions in THP-1, a human macrophage-like cell line. Of the 251 phosphatases, the screen identified the Ca++-activated K+-channel-associated phosphatase myotubularin-related protein-6 (MTMR6) as the only phosphatase whose silencing reduced parasite load and IL-10 production in human macrophages. Virulent, but not avirulent, L. donovani infection increased MTMR6 expression in macrophages. As virulent L. donovani parasites expressed higher lipophosphoglycan, a TLR2-ligand, we tested the effect of TLR2 stimulation or blockade on MTMR6 expression. TLR1/TLR2-ligand Pam3CSK4 enhanced, but TLR2 blockade reduced, MTMR6 expression. L. donovani infection of macrophages ex vivo increased, but miltefosine treatment reduced, MTMR6 expression. Corroboratively, compared to endemic controls, untreated VL patients had higher, but miltefosine-treated VL patients had reduced, MTMR6 expression. The phosphatase siRNA-library screening thus identified MTMR6 as the first TLR2-modulated ion channel-associated phosphatase with significant implications in VL patients and anti-leishmanial functions.

Copula-based risk aggregation with trapped ion quantum computers.

Copulas are mathematical tools for modeling joint probability distributions. In the past 60 years they have become an essential analysis tool on classical computers in various fields. The recent finding that copulas can be expressed as maximally entangled quantum states has revealed a promising approach to practical quantum advantages: performing tasks faster, requiring less memory, or, as we show, yielding better predictions. Studying the scalability of this quantum approach as both the precision and the number of modeled variables increase is crucial for its adoption in real-world applications. In this paper, we successfully apply a Quantum Circuit Born Machine (QCBM) based approach to modeling 3- and 4-variable copulas on trapped ion quantum computers. We study the training of QCBMs with different levels of precision and circuit design on a simulator and a state-of-the-art trapped ion quantum computer. We observe decreased training efficacy due to the increased complexity in parameter optimization as the models scale up. To address this challenge, we introduce an annealing-inspired strategy that dramatically improves the training results. In our end-to-end tests, various configurations of the quantum models make a comparable or better prediction in risk aggregation tasks than the standard classical models.

Human IL-17A protein production is controlled through a PIP5K1α-dependent translational checkpoint.

The cytokine interleukin-17 (IL-17) is secreted by T helper 17 (TH17) cells and is beneficial for microbial control; however, it also causes inflammation and pathological tissue remodeling in autoimmunity. Hence, TH17 cell differentiation and IL-17 production must be tightly regulated, but, to date, this has been defined only in terms of transcriptional control. Phosphatidylinositols are second messengers produced during T cell activation that transduce signals from the T cell receptor (TCR) and costimulatory receptors at the plasma membrane. Here, we found that phosphatidylinositol 4,5-bisphosphate (PIP2) was enriched in the nuclei of human TH17 cells, which depended on the kinase PIP5K1α, and that inhibition of PIP5K1α impaired IL-17A production. In contrast, nuclear PIP2 enrichment was not observed in TH1 or TH2 cells, and these cells did not require PIP5K1α for cytokine production. In T cells from people with multiple sclerosis, IL-17 production elicited by myelin basic protein was blocked by PIP5K1α inhibition. IL-17 protein was affected without altering either the abundance or stability of IL17A mRNA in TH17 cells. Instead, analysis of PIP5K1α-associating proteins revealed that PIP5K1α interacted with ARS2, a nuclear cap-binding complex scaffold protein, to facilitate its binding to IL17A mRNA and subsequent IL-17A protein production. These findings highlight a transcription-independent, translation-dependent mechanism for regulating IL-17A protein production that might be relevant to other cytokines.

Comparison of SARS-CoV-2 diagnosis by rapid antigen detection kit with RT-qPCR in a tertiary care setup in North Eastern India.

PURPOSE: Real time reverse transcriptase polymerase chain reaction (RT-qPCR) is still considered a gold standard for the diagnosis of COVID-19. However, due to several limitations, use of RT-qPCR is limited in a resource poor setting like North East India. Rapid antigen detection testing kit has revolutionized the diagnosis and management of COVID-19 in India. However, conflicting reports exist regarding the efficacy of the kits for diagnosis of COVID-19. This study aims to highlight the performance of Standard Q COVID-19® Antigen detection kit (SD Biosensor) compared with RT-qPCR in the setup of North East India. METHODS: Nasopharyngeal and oropharyngeal swab samples were collected from consenting patients attending the flu clinic in the period from 1st July to December 31, 2020. Samples were transferred to Viral Research and Diagnostic Laboratory (VRDL) for RT-qPCR test. Antigen detection from the patient samples were undertaken using Standard Q ® COVID-19 antigen detection kit (SD Biosensor, Republic of Korea). Data were then analyzed for comparison between RT-qPCR and antigen kit results. RESULTS: During the study period, 189 samples were collected, out of which 119 were positive by RT-qPCR. Out of 119 positive samples, calculated sensitivity and specificity of the rapid antigen kit was 63% and 100% respectively. Sensitivity and diagnostic accuracy increases in symptomatic patients as compared to asymptomatic patients. Cohen's Kappa coefficient showed a moderate association (0.6) between the kit and RT-qPCR test. The kit performed optimally at a CT value of ≤32.5 for N gene with a predicted sensitivity of 77.3% and specificity of 93.3%. CONCLUSION: The study shows an overall acceptable sensitivity and specificity of the testing kit, with a better performance in symptomatic patients. The association of the kit result is moderate with the results obtained in RT-qPCR. In this study, the rapid antigen test kit performed optimally at N gene qRT PCR cut off value of ≤32.5.

Arid5a Mediates an IL-17-Dependent Pathway That Drives Autoimmunity but Not Antifungal Host Defense.

IL-17 contributes to the pathogenesis of certain autoimmune diseases, but conversely is essential for host defense against fungi. Ab-based biologic drugs that neutralize IL-17 are effective in autoimmunity but can be accompanied by adverse side effects. Candida albicans is a commensal fungus that is the primary causative agent of oropharyngeal and disseminated candidiasis. Defects in IL-17 signaling cause susceptibility to candidiasis in mice and humans. A key facet of IL-17 receptor signaling involves RNA-binding proteins, which orchestrate the fate of target mRNA transcripts. In tissue culture models we showed that the RNA-binding protein AT-rich interaction domain 5A (Arid5a) promotes the stability and/or translation of multiple IL-17-dependent mRNAs. Moreover, during oropharyngeal candidiasis, Arid5a is elevated within the oral mucosa in an IL-17-dependent manner. However, the contribution of Arid5a to IL-17-driven events in vivo is poorly defined. In this study, we used CRISPR-Cas9 to generate mice lacking Arid5a. Arid5a -/- mice were fully resistant to experimental autoimmune encephalomyelitis, an autoimmune setting in which IL-17 signaling drives pathology. Surprisingly, Arid5a -/- mice were resistant to oropharyngeal candidiasis and systemic candidiasis, similar to immunocompetent wild-type mice and contrasting with mice defective in IL-17 signaling. Therefore, Arid5a-dependent signals mediate pathology in autoimmunity and yet are not required for immunity to candidiasis, indicating that selective targeting of IL-17 signaling pathway components may be a viable strategy for development of therapeutics that spare IL-17-driven host defense.

In situ process quality monitoring and defect detection for direct metal laser melting.

Quality control and quality assurance are challenges in direct metal laser melting (DMLM). Intermittent machine diagnostics and downstream part inspections catch problems after undue cost has been incurred processing defective parts. In this paper we demonstrate two methodologies for in-process fault detection and part quality prediction that leverage existing commercial DMLM systems with minimal hardware modification. Novel features were derived from the time series of common photodiode sensors along with standard machine control signals. In one methodology, a Bayesian approach attributes measurements to one of multiple process states as a means of classifying process deviations. In a second approach, a least squares regression model predicts severity of certain material defects.

Diagnosis of Indigenous Non-Malarial Vector-Borne Infections from Malaria Negative Samples from Community and Rural Hospital Surveillance in Dhalai District, Tripura, North-East India.

The aetiology of non-malaria vector-borne diseases in malaria-endemic, forested, rural, and tribal-dominated areas of Dhalai, Tripura, in north-east India, was studied for the first time in the samples collected from malaria Rapid Diagnostic Kit negative febrile patients by door-to-door visits in the villages and primary health centres. Two hundred and sixty serum samples were tested for the Dengue NS1 antigen and the IgM antibodies of Dengue, Chikungunya, Scrub Typhus (ST), and Japanese Encephalitis (JE) during April 2019-March 2020. Fifteen Dengue, six JE, twelve Chikungunya, nine ST and three Leptospirosis, and mixed infections of three JE + Chikungunya, four Dengue + Chikungunya, three Dengue + JE + Chikungunya, one Dengue + Chikungunya + ST, and one Dengue + ST were found positive by IgM ELISA tests, and four for the Dengue NS1 antigen, all without any travel history. True prevalence values estimated for infections detected by Dengue IgM were 0.134 (95% CI: 0.08-0.2), Chikungunya were 0.084 (95% CI: 0.05-0.13), Scrub were 0.043 (95% CI: 0.01-0.09), and Japanese Encephalitis were 0.045 (95% CI: 0.02-0.09). Dengue and Chikungunya were associated significantly more with a younger age. There was a lack of a defined set of symptoms for any of the Dengue, Chikungunya, JE or ST infections, as indicated by the k-modes cluster analysis. Interestingly, most of these symptoms have an overlapping set with malaria; thereby, it becomes imperative that malaria and these non-malaria vector-borne disease diagnoses are made in a coordinated manner. Findings from this study call for advances in routine diagnostic procedures and the development of a protocol that can accommodate, currently, in practicing the rapid diagnosis of malaria and other vector-borne diseases, which is doable even in the resource-poor settings of rural hospitals and during community fever surveillance.

The RNA-binding protein IMP2 drives a stromal-Th17 cell circuit in autoimmune neuroinflammation.

Stromal cells are emerging as key drivers of autoimmunity, partially because they produce inflammatory chemokines that orchestrate inflammation. Chemokine expression is regulated transcriptionally but also through posttranscriptional mechanisms, the specific drivers of which are still incompletely defined. CCL2 (MCP1) is a multifunctional chemokine that drives myeloid cell recruitment. During experimental autoimmune encephalomyelitis (EAE), an IL-17-driven model of multiple sclerosis, CCL2 produced by lymph node (LN) stromal cells was essential for immunopathology. Here, we showed that Ccl2 mRNA upregulation in human stromal fibroblasts in response to IL-17 required the RNA-binding protein IGF-2 mRNA-binding protein 2 (IGF2BP2, IMP2), which is expressed almost exclusively in nonhematopoietic cells. IMP2 binds directly to CCL2 mRNA, markedly extending its transcript half-life, and is thus required for efficient CCL2 secretion. Consistent with this, Imp2-/- mice showed reduced CCL2 production in LNs during EAE, causing impairments in monocyte recruitment and Th17 cell polarization. Imp2-/- mice were fully protected from CNS inflammation. Moreover, deletion of IMP2 after EAE onset was sufficient to mitigate disease severity. These data showed that posttranscriptional control of Ccl2 in stromal cells by IMP2 was required to permit IL-17-driven progression of EAE pathogenesis.

IL-17-dependent fibroblastic reticular cell training boosts tissue protective mucosal immunity through IL-10-producing B cells.

Prior experience of pathogen-associated stimuli reduces morbidity and mortality to newly encountered infections through innate immune training, which can be enhanced by childhood vaccination. Fibroblastic reticular cells (FRCs) are stromal cells in lymphoid organs that support lymphocyte localization and survival and modulate adaptive immune responses. IL-17 signaling is important for FRC metabolism and proliferation during inflammatory responses. Here, we show that FRC-intrinsic IL-17 signaling was required for protective antibody-mediated immunity to the gut bacterial pathogen Citrobacter rodentium. We asked whether prior activation of FRC through nonspecific inflammatory "training" of the gut would alter subsequent immune response to C. rodentium. Inflammatory training increased the number of activated FRC in mesenteric LN (MLN) and enhanced the antibody response to C. rodentium in an IL-17­dependent manner. FRC demonstrated cardinal features of innate immune training, including increased epigenetic markers of activation and increased metabolic response to infection. Enhanced responses were still evident 6 weeks after training. The kinetics of bacterial infection were not changed by inflammatory training, but colon inflammation was paradoxically reduced. Mechanistically, IL-10 production by activated B cells was required for colon protective effects of inflammatory training. Enhancing tissue protective B cell responses thus led to increased production of antibody and IL-10, allowing clearance of infection with reduced tissue inflammation. These data identify a new mode of immune training through FRC to modulate future adaptive responses and better preserve host health.

IL-17 in the Pathogenesis of Disease: Good Intentions Gone Awry.

The IL-17 family is an evolutionarily old cytokine family consisting of six members (IL-17A through IL-17F). IL-17 family cytokines signal through heterodimeric receptors that include the shared IL-17RA subunit, which is widely expressed throughout the body on both hematopoietic and nonhematopoietic cells. The founding family member, IL-17A, is usually referred to as IL-17 and has received the most attention for proinflammatory roles in autoimmune diseases like psoriasis. However, IL-17 is associated with a wide array of diseases with perhaps surprisingly variable pathologies. This review focuses on recent advances in the roles of IL-17 during health and in disease pathogenesis. To decipher the functions of IL-17 in diverse disease processes it is useful to first consider the physiological functions that IL-17 contributes to health. We then discuss how these beneficial functions can be diverted toward pathogenic amplification of deleterious pathways driving chronic disease.

Noncanonical STAT3 activity sustains pathogenic Th17 proliferation and cytokine response to antigen.

The STAT3 signaling pathway is required for early Th17 cell development, and therapies targeting this pathway are used for autoimmune disease. However, the role of STAT3 in maintaining inflammatory effector Th17 cell function has been unexplored. Th17ΔSTAT3 mice, which delete STAT3 in effector Th17 cells, were resistant to experimental autoimmune encephalomyelitis (EAE), a murine model of MS. Th17 cell numbers declined after STAT3 deletion, corresponding to reduced cell cycle. Th17ΔSTAT3 cells had increased IL-6-mediated phosphorylation of STAT1, known to have antiproliferative functions. Th17ΔSTAT3 cells also had reduced mitochondrial membrane potential, which can regulate intracellular Ca2+. Accordingly, Th17ΔSTAT3 cells had reduced production of proinflammatory cytokines when stimulated with myelin antigen but normal production of cytokines when TCR-induced Ca2+ flux was bypassed with ionomycin. Thus, early transcriptional roles of STAT3 in developing Th17 cells are later complimented by noncanonical STAT3 functions that sustain pathogenic Th17 cell proliferation and cytokine production.

IL-17 metabolically reprograms activated fibroblastic reticular cells for proliferation and survival.

Lymph-node (LN) stromal cell populations expand during the inflammation that accompanies T cell activation. Interleukin-17 (IL-17)-producing helper T cells (TH17 cells) promote inflammation through the induction of cytokines and chemokines in peripheral tissues. We demonstrate a critical requirement for IL-17 in the proliferation of LN and splenic stromal cells, particularly fibroblastic reticular cells (FRCs), during experimental autoimmune encephalomyelitis and colitis. Without signaling via the IL-17 receptor, activated FRCs underwent cell cycle arrest and apoptosis, accompanied by signs of nutrient stress in vivo. IL-17 signaling in FRCs was not required for the development of TH17 cells, but failed FRC proliferation impaired germinal center formation and antigen-specific antibody production. Induction of the transcriptional co-activator IκBζ via IL-17 signaling mediated increased glucose uptake and expression of the gene Cpt1a, encoding CPT1A, a rate-limiting enzyme of mitochondrial fatty acid oxidation. Hence, IL-17 produced by locally differentiating TH17 cells is an important driver of the activation of inflamed LN stromal cells, through metabolic reprogramming required to support proliferation and survival.

A zinc Schiff base complex inhibits cancer progression both in vivo and in vitro by inducing apoptosis.

Cancer chemotherapy suffers from selectivity and undesired toxicity of the drugs. Since zinc is a biocompatible tracer element and cysteine derivatives are used in cancer chemoprevention, we intend to develop a complex of zinc and cysteine-derivatives as potent, non-toxic anticancer agents. Herein, we synthesized and characterized cysteine based ligand, 2-[(2-Hydroxy-3-methoxy-benzylidene)-amino]-3-mercapto-propionic acid and its Zn-complex, which are found to be non-toxic towards normal human PBMC. Data also revealed that only Zn-complex exhibited remarkable apoptosis in drug-sensitive CCRF-CEM and multidrug-resistant CEM/ADR5000 cancer cells as assessed by MTT, Cell cycle and AnnexinV binding assay. Moreover, Zn-complex altered ROS and GSH level of the respective cell lines. Finally, treatment of Zn-complex in Swiss albino mice did not show any systemic toxicity in preliminary trials in normal mice and remarkably increased the life-span of EAC bearing mice. In conclusion, the synthesized Zn-complex may be developed for efficacious, multidrug resistance reversal, non-toxic chemotherapeutic agents in future.

Modulation of cell death in human colorectal and breast cancer cells through a manganese chelate by involving GSH with intracellular p53 status.

Chemotherapy is central to current treatment modality especially for advanced and metastatic colorectal and breast cancers. Targeting the key molecular events of the neoplastic cells may open a possibility to treat cancer. Although some improvements in understanding of colorectal and breast cancer treatment have been recorded, the involvement of glutathione (GSH) and dependency of p53 status on the modulation of GSH-mediated treatment efficacy have been largely overlooked. Herein, we tried to decipher the underlying mechanism of the action of Mn-N-(2-hydroxyacetophenone) glycinate (MnNG) against differential p53 status bearing Hct116, MCF-7, and MDA-MB-468 cells on the backdrop of intracellular GSH level and reveal the role of p53 status in modulating GSH-dependant abrogation of MnNG-induced apoptosis in these cancer cells. Present study discloses that MnNG targets specifically wild-type-p53 expressing Hct116 and MCF-7 cells by significantly depleting both cytosolic, mitochondrial GSH, and modulating nuclear GSH through Glutathione reductase and Glutamate-cysteine ligase depletion that may in turn induce p53-mediated intrinsic apoptosis in them. Thus GSH addition abrogates p53-mediated apoptosis in wild-type-p53 expressing cells. GSH addition also overrides MnNG-induced modulation of phase II detoxifying parameters in them. However, GSH addition partially replenishes the down-regulated or modulated GSH pool in cytosol, mitochondria, and nucleus, and relatively abrogates MnNG-induced intrinsic apoptosis in p53-mutated MDA-MB-468 cells. On the contrary, although MnNG induces significant cell death in p53-null Hct116 cells, GSH addition fails to negate MnNG-induced cell death. Thus p53 status with intracellular GSH is critical for the modulation of MnNG-induced apoptosis.

Leishmania donovani-Induced Prostaglandin E2 Generation Is Critically Dependent on Host Toll-Like Receptor 2-Cytosolic Phospholipase A2 Signaling.

Visceral leishmaniasis (VL) is the second-largest parasitic killer disease after malaria. During VL, the protozoan Leishmania donovani induces prostaglandin E2 (PGE2) generation within host macrophages to aid parasite survival. PGE2 significantly influences leishmanial pathogenesis, as L. donovani proliferation is known to be attenuated in PGE2-inhibited macrophages. Here, we report for the first time that signaling via macrophage Toll-like receptor 2 (TLR2) plays an instrumental role in inducing PGE2 release from L. donovani-infected macrophages. This signaling cascade, mediated via the TLR2-phosphatidylinositol 3-kinase (PI3K)-phospholipase C (PLC) signaling pathway, was found to be indispensable for activation of two major enzymes required for PGE2 generation: cytosolic phospholipase A2 (cPLA2) and cyclooxygenase 2 (Cox2). Inhibition of cPLA2, but not secreted phospholipase A2 (sPLA2) or calcium-independent phospholipase A2 (iPLA2), arrested L. donovani infection. During infection, cPLA2 activity increased >7-fold in a calcium-dependent and extracellular signal-regulated kinase (ERK)-dependent manner, indicating that elevation of intracellular calcium and ERK-mediated phosphorylation was necessary for L. donovani-induced cPLA2 activation. For transcriptional upregulation of cyclooxygenase 2, activation of the calcium-calcineurin-nuclear factor of activated T cells (NFAT) signaling was required in addition to the TLR2-PI3K-PLC pathway. Detailed studies by site-directed mutagenesis of potential NFAT binding sites and chromatin immunoprecipitation (ChIP) analysis revealed that the binding of macrophage NFATc2, at the -73/-77 site on the cox2 promoter, induced L. donovani-driven cox2 transcriptional activation. Collectively, these findings highlight the contribution of TLR2 downstream signaling toward activation of cPLA2 and Cox2 and illustrate how the TLR2-PI3K-PLC pathway acts in a concerted manner with calcium-calcineurin-NFATc2 signaling to modulate PGE2 release from L. donovani-infected macrophages.

Inflammatory Th17 Cells Express Integrin αvß3 for Pathogenic Function.

Interleukin-23 (IL-23) is required for inflammatory Th17 cell function in experimental autoimmune encephalomyelitis (EAE), and IL-23 blockade reduces the number of effector Th17 cells in the CNS. We report that pro-inflammatory Th17 cells express high integrin ß3 that is IL-23 dependent. Integrin ß3 was not upregulated on all activated T cells; rather, integrin ß3 was upregulated along with its functional partner integrin αv on effector Th17 cells and "ex-Th17" cells, and αvß3(hi) RORγt(+) cells expanded during EAE. Integrin αvß3 inhibitors ameliorated clinical signs of EAE, and integrin ß3 deficiency on CD4(+) T cells alone was sufficient to block EAE induction. Furthermore, integrin-ß3-deficient Th17 cells, but not Th1 cells, were impaired in their ability to induce EAE. Integrin ß3(-/-) T cells induced smaller demyelinated lesions and showed reduced spread and accumulation within the CNS, corresponding with impaired extracellular-matrix-mediated migration. Hence, integrin ß3 is required for Th17 cell-mediated autoimmune CNS inflammation.

SLA-PGN-primed dendritic cell-based vaccination induces Th17-mediated protective immunity against experimental visceral leishmaniasis: a crucial role of PKCß.

Emergence of drug resistance during visceral leishmaniasis (VL) is a major obstacle imposed during successful therapy. An effective vaccine strategy against this disease is therefore necessary. Our present study exploited the SLA (soluble leishmanial antigen) and PGN (peptidoglycan) stimulated bone marrow-derived dendritic cells (DCs) as a suitable vaccine candidate during experimental VL. SLA-PGN-stimulated DCs showed a significant decrease in hepatic and splenic parasite burden, which were associated with increased production of nitric oxide and pro-inflammatory cytokines such as IL-12, IFN-γ and IL-17. Elevated level of IL-17 was accompanied with the generation of more Th17 cells. Further studies on DC provided the evidence that these SLA-PGN-stimulated DCs played an important role in providing necessary cytokines such as IL-6, IL-23 and TGF-ß for the generation of Th17 cells. Interestingly, inhibition of protein kinase C-ß (PKCß) in DCs led to decreased production of Th17 polarizing cytokines, causing reduction of the Th17 population size. Altogether, our finding highlighted the important role of DC-based PKCß in regulation of the function and generation of Th17 cells.

Immunomodulation of host-protective immune response by regulating Foxp3 expression and Treg function in Leishmania-infected BALB/c mice: critical role of IRF1.

Visceral leishmaniasis (VL), caused by a protozoan parasite Leishmania donovani, is still a threat to mankind due to treatment failure, drug resistance and coinfection with HIV. The limitations of first-line drugs have led to the development of new strategies to combat this dreaded disease. Recently, we have shown the immunomodulatory property of Ara-LAM, a TLR2 ligand, against leishmanial pathogenesis. In this study, we have extended our study to the effect of Ara-LAM on regulatory T cells in a murine model of VL. We observed that Ara-LAM-treated infected BALB/c mice showed a strong host-protective Th1 immune response due to reduced IL-10 and TGF-ß production, along with marked decrease in CD4(+) CD25(+) Foxp3(+) GITR(+) CTLA4(+) regulatory T cell (Treg) generation and activation. The reduction in Foxp3 expression was due to effective modulation of TGF-ß-induced SMAD signaling in Treg cells by Ara-LAM. Moreover, we demonstrated that Ara-LAM-induced IRF1 expression in the Treg cells, which negatively regulated foxp3 gene transcription, resulting in the reduced immunosuppressive activity of Treg cells. Interestingly, irf1 gene knockdown completely abrogated the effect of Ara-LAM on Treg cells. Thus, these findings provide detailed mechanistic insight into Ara-LAM-mediated modulation of Treg cells, which might be helpful in combating VL.

The host-protective effect of arabinosylated lipoarabinomannan against Leishmania donovani infection is associated with restoration of IFN-γ responsiveness.

Visceral leishmaniasis (VL), which is endemic as a major infectious disease in the tropical and subtropical countries, is caused by a protozoan parasite Leishmania donovani. At present, restricted treatment options and lack of vaccines intensify the problem of controlling VL. Therefore, finding a novel immunoprophylactic or therapeutic principle is a pressing need. Here, we report that arabinosylated lipoarabinomannan (Ara-LAM), a TLR2-ligand isolated from Mycobacterium smegmatis, exhibits a strong immunomodulatory property that conferred protection against L. donovani infection. Although, Ara-LAM modulates TLR2 and MAPK signaling, it is not known whether Ara-LAM involves IFN-γ signaling for effective parasite clearance. Because, it is reported that IFN-γ signaling, a principle mediator of NO generation and macrophage and Tcell activation, is hampered during leishmanial pathogenesis. Ara-LAM increases IFN-γ receptor expression and potentiates IFN-γ receptor signaling through JAK-STAT pathway. Moreover, Ara-LAM reciprocally modulates IRF4 and IRF8 expression and reinstates anti-leishmanial Th1 response that eventuates in significantly reduced parasite load in spleen and liver of L. donovani-infected BALB/c mice. IFN-γRα silencing resulted in the suppression of these host-protective mechanisms affected by Ara-LAM. Thus, Ara-LAM-mediated restoration of IFN-γ responsiveness is a novel immuno-modulatory principle for protection against L. donovani susceptible host.

Co-administration of glycyrrhizic acid with the antileishmanial drug sodium antimony gluconate (SAG) cures SAG-resistant visceral leishmaniasis.

Since there are very few affordable antileishmanial drugs available, antimonial resistance has crippled antileishmanial therapy, thereby emphasising the need for development of novel therapeutic strategies. This study aimed to evaluate the antileishmanial role of combined therapy with sodium antimony gluconate (SAG) and the triterpenoid glycyrrhizic acid (GA) against infection with SAG-resistant Leishmania (GE1F8R). Combination therapy with GA and SAG successfully limited infection with SAG-resistant Leishmania in a synergistic manner (fractional inhibitory concentration index <1.0). At the same time, mice infected with SAG-resistant Leishmania and co-treated with GA and SAG exhibited a significant reduction in hepatic and splenic parasite burden. In probing the mechanism, it was observed that GA treatment suppressed the expression and efflux activity of P-glycoprotein (P-gp) and multidrug resistance-associated protein 1 (MRP1), two host ABC transporters responsible for antimony efflux from host cells infected with SAG-resistant parasites. This suppression correlated with greater intracellular antimony retention during SAG therapy both in vitro and in vivo, which was reflected in the reduced parasite load. Furthermore, co-administration of GA and SAG induced a shift in the cytokine balance towards a Th1 phenotype by augmenting pro-inflammatory cytokines (such as IL-12, IFNγ and TNFα) and inducing nitric oxide generation in GE1F8R-infected macrophages as well as GE1F8R-infected mice. This study aims to provide an affordable leishmanicidal alternative to expensive antileishmanial drugs such as miltefosine and amphotericin B. Furthermore, this report explores the role of GA as a resistance modulator in MRP1- and P-gp-overexpressing conditions.