Understanding the critical role of microRNAs in HIV-1 pathogenesis

MicroRNAs (miRNAs) are ~22 nucleotide small RNAs which are critical regulators of mRNA translation and have key roles in HIV-1 infection. We characterized miRNA changes in CD4+ T cells from 7 patients with chronic HIV-1 infection (CHI), 7 long term non progressors (LTNPs) and 8 healthy controls (HC). There were 111 up- and 52 down-regulated miRNAs in CHI relative to HC, and 70 up- and 27 down-regulated miRNAs in LTNPs relative to HC. Let-7 miRNAs were found to be highly expressed in CD4+ T cells from HC whilst significantly and differentially decreased in CHI and LTNPs. We modulated let-7 levels in an in-vitro model and found no direct effect on HIV-1 replication kinetics. We therefore investigated whether miRNAs played a role in the altered cytokine milieu associated with chronic infection. As several in silico algorithms consistently suggested that let-7 miRNAs target the 3’ untranslated (3’UTR) region of IL10, we focused on this as a target. IL-10 was significantly increased in plasma from CHI compared to HC and also in the supernatant of HIV-1infected HUT78 cells compared to uninfected cultures. Using HUT78 cells infected with HIV-1, we noted let-7b was down-regulated within 72 hours and let-7c within 24 hours. We over-expressed let-7 miRNAs and showed decreases in IL-10 for let-7b, let-7f and let-7c. Using anti-miRNAs we showed increased IL-10 levels for let-7c, let-7f and let-7b. We confirmed that let-7 was binding to the IL10 3’UTR using a reporter construct. We performed a microarray in CD4+ T cell subsets revealing that Blimp-1 was preferentially expressed in activated regulatory T cells. Blimp-1 is a zinc finger transcriptional repressor known to play a role in IL-2 expression. We found that miR-9 played a pivotal role in regulating Blimp-1, that Blimp-1 levels were markedly elevated in CHI compared to LTNPs and HC and its expression in CHI may explain the dysfunctional endogenous IL-2 secretion noted in CHI. We investigated the expression of miRNAs in monocytes and found only two miRNAs which were differentially expressed between patient groups. By understanding the interaction between miRNAs and HIV-1, this work has resulted in unique insights into HIV-1 pathogenesis

The conundrum of neuropsychiatric systemic lupus erythematosus: Current and novel approaches to diagnosis

Recognising neuropsychiatric involvement by systemic lupus erythematosus (SLE) is of growing importance, however many barriers to this exist at multiple levels of our currently available diagnostic algorithms that may ultimately delay its diagnosis and subsequent treatment. The heterogeneous and non-specific clinical syndromes, serological and cerebrospinal fluid (CSF) markers and neuroimaging findings that often do not mirror disease activity, highlight important research gaps in the diagnosis of neuropsychiatric SLE (NPSLE). Formal neuropsychological assessments or the more accessible screening metrics may also help improve objective recognition of cognitive or mood disorders. Novel serum and CSF markers, including autoantibodies, cytokines and chemokines have also shown increasing utility as part of diagnosis and monitoring, as well as in distinguishing NPSLE from SLE patients without SLE-related neuropsychiatric manifestations. Novel neuroimaging studies also expand upon our existing strategy by quantifying parameters that indicate microarchitectural integrity or provide an assessment of neuronal function. Some of these novel markers have shown associations with specific neuropsychiatric syndromes, suggesting that future research move away from considering NPSLE as a single entity but rather into its individually recognized neuropsychiatric manifestations. Nevertheless, it is likely that a composite panel of these investigations will be needed to better address the gaps impeding recognition of neuropsychiatric involvement by SLE

The interaction of multiple sclerosis risk loci with Epstein-Barr virus phenotypes implicates the virus in pathogenesis

Translating the findings of genome wide association studies (GWAS) to new therapies requires identification of the relevant immunological contexts to interrogate for genetic effects. In one of the largest GWAS, more than 200 risk loci have been identified for Multiple Sclerosis (MS) susceptibility. Infection with Epstein-Barr virus (EBV) appears to be necessary for the development of Multiple Sclerosis (MS). Many MS risk loci are associated with altered gene expression in EBV infected B cells (LCLs). We have interrogated this immunological context to identify interaction between MS risk loci and EBV DNA copy number, intrinsic growth rate and EBV encoded miRNA expression. The EBV DNA copy number was associated with significantly more risk alleles for MS than for other diseases or traits. EBV miRNAs BART4-3p and BART3-5p were highly associated with EBV DNA copy number and MS risk loci. The poliovirus receptor (PVR) risk SNP was associated with EBV DNA copy number, PVR and miRNA expression. Targeting EBV miRNAs BART4-3p and BART3-5p, and the gene PVR, may provide therapeutic benefit in MS. This study also indicates how immunological context and risk loci interactions can be exploited to validate and develop novel therapeutic approaches. © 2020, The Author(s)

The interaction of Epstein-Barr virus encoded transcription factor EBNA2 with multiple sclerosis risk loci is dependent on the risk genotype

Background: Epstein-Barr virus (EBV) infection may be necessary for the development of Multiple sclerosis (MS). Earlier we had identified six MS risk loci that are co-located with binding sites for the EBV transcription factor Epstein-Barr Nuclear Antigen 2 (EBNA2) in EBV-infected B cells (lymphoblastoid cell lines – LCLs). Methods: We used an allele-specific chromatin immunoprecipitation PCR assay to assess EBNA2 allelic preference. We treated LCLs with a peptide inhibitor of EBNA2 (EBNA2-TAT), reasoning that inhibiting EBNA2 function would alter gene expression at these loci if it was mediated by EBNA2. Findings: We found that EBNA2 binding was dependent on the risk allele for five of these six MS risk loci (p < 0·05). Treatment with EBNA2-TAT significantly altered the expression of TRAF3 (p < 0·05), CD40 (p < 0·001), CLECL1 (p <0·0001), TNFAIP8 (p < 0·001) and TNFRSF1A (p < 0·001). Interpretation: These data suggest that EBNA2 can enhance or reduce expression of the gene depending on the risk allele, likely promoting EBV infection. This is consistent with the concept that these MS risk loci affect MS risk through altering the response to EBNA2. Together with the extensive data indicating a pathogenic role for EBV in MS, this study supports targeting EBV and EBNA2 to reduce their effect on MS pathogenesis

Transcribed B lymphocyte genes and multiple sclerosis risk genes are underrepresented in Epstein–Barr Virus hypomethylated regions

Epstein–Barr Virus (EBV) infection appears to be necessary for the development of Multiple Sclerosis (MS), although the specific mechanisms are unknown. More than 200 single-nucleotide polymorphisms (SNPs) are known to be associated with the risk of developing MS. About a quarter of these are also highly associated with proximal gene expression in B cells infected with EBV (lymphoblastoid cell lines—LCLs). The DNA of LCLs is hypomethylated compared with both uninfected and activated B cells. Since methylation can affect gene expression, and so cell differentiation and immune evasion, we hypothesised that EBV-driven hypomethylation may affect the interaction between EBV infection and MS. We interrogated an existing dataset comprising three individuals with whole-genome bisulfite sequencing data from EBV transformed B cells and CD40L-activated B cells. DNA methylation surrounding MS risk SNPs associated with gene expression in LCLs (LCLeQTL) was less likely to be hypomethylated than randomly selected chromosomal regions. Differential methylation was independent of genomic features such as promoter regions, but genes preferentially expressed in EBV-infected B cells, including the LCLeQTL genes, were underrepresented in the hypomethylated regions. Our data does not indicate MS genetic risk is affected by EBV hypomethylation

The interaction of human and Epstein–Barr virus miRNAs with Multiple Sclerosis risk loci

Although the causes of Multiple Sclerosis (MS) still remain largely unknown, multiple lines of evidence suggest that Epstein–Barr virus (EBV) infection may contribute to the development of MS. Here, we aimed to identify the potential contribution of EBV-encoded and host cellular miRNAs to MS pathogenesis. We identified differentially expressed host miRNAs in EBV infected B cells (LCLs) and putative host/EBV miRNA interactions with MS risk loci. We estimated the genotype effect of MS risk loci on the identified putative miRNA:mRNA interactions in silico. We found that the protective allele of MS risk SNP rs4808760 reduces the expression of hsa-mir-3188-3p. In addition, our analysis suggests that hsa-let-7b-5p may interact with ZC3HAV1 differently in LCLs compared to B cells. In vitro assays indicated that the protective allele of MS risk SNP rs10271373 increases ZC3HAV1 expression in LCLs, but not in B cells. The higher expression for the protective allele in LCLs is consistent with increased IFN response via ZC3HAV1 and so decreased immune evasion by EBV. Taken together, this provides evidence that EBV infection dysregulates the B cell miRNA machinery, including MS risk miRNAs, which may contribute to MS pathogenesis via interaction with MS risk genes either directly or indirectly

Politics and society between elections

The problems of India’s development and governance are routinely linked to the logic of India’s electoral democracy. As a result, a great deal is known about elections, but paradoxically our knowledge of politics and society between elections is relatively underdeveloped. As much as anything else, development and governance outcomes are shaped by how the government functions between elections; including how it relates to citizens on a regular basis, how it provides routine public services to them, and how public order is maintained. Further, governance processes are nested in the social and political relationships between citizens and government functionaries

Sputum conversion and treatment success among tuberculosis patients with diabetes treated under the Tuberculosis Control Programme in an Urban setting in South India

Background: Studies from India on sputum conversion and tuberculosis (TB) treatment outcomes among TB patients with diabetes are limited. Objective: The objective of this study is to estimate the proportion of sputum smear conversion and successful treatment outcomes among diabetic–TB patients treated under Revised National TB Control Programme (RNTCP). Methodology: Information on TB disease, diabetes, sputum conversion, and treatment outcomes were collected from treatment cards of adult TB patients (age >18 years) treated in the District TB Centre TB Unit, Tiruchirapalli, Tamil Nadu from July 1, 2014, to October 31, 2015. Results: Diabetes was documented in 163 (14%) of 1131 TB patients. Sputum conversion was in 107 (94%) of 114 smear positive-TB patients. Successful TB treatment outcome was in 116 (85%) of 136 patients and 107 (86%) of 124 new TB patients. Conclusion: Sputum conversion was as per RNTCP target while treatment success rate among the new TB patients with diabetes was suboptimal

Abrogation of Cbl–PI3K Interaction Increases Bone Formation and Osteoblast Proliferation

Cbl is an adaptor protein and E3 ligase that plays both positive and negative roles in several signaling pathways that affect various cellular functions. Tyrosine 737 is unique to Cbl and phosphorylated by Src family kinases. Phosphorylated CblY737 creates a binding site for the p85 regulatory subunit of phosphatidylinositol 3 kinase (PI3K) that also plays an important role in the regulation of bone homeostasis. To investigate the role of Cbl–PI3K interaction in bone homeostasis, we examined knock-in mice in which the PI3K binding site on Cbl was ablated due to the substitution of tyrosine 737 to phenylalanine (CblYF/YF, YF mice). We previously reported that bone volume in these mice is increased due to decreased osteoclast function (Adapala et al., J Biol Chem 285:36745–36758, 19). Here, we report that YF mice also have increased bone formation and osteoblast numbers. In ex vivo cultures bone marrow-derived YF osteoblasts showed increased Col1A expression and their proliferation was also significantly augmented. Moreover, proliferation of MC3T3-E1 cells was increased after treatment with conditioned medium generated by culturing YF bone marrow stromal cells. Expression of stromal derived factor-1 (SDF-1) was increased in YF bone marrow stromal cells compared to wild type. Increased immunostaining of SDF-1 and CXCR4 was observed in YF bone marrow stromal cells compared to wild type. Treatment of YF condition medium with neutralizing anti-SDF-1 and anti-CXCR4 antibodies attenuated MC3T3-E1 cell proliferation. Cumulatively, these results show that abrogation of Cbl–PI3K interaction perturbs bone homeostasis, affecting both osteoclast function and osteoblast proliferation