Pre-TCR Signaling and Inactivation of p53 Induces Crucial Cell Survival Pathways in Pre-T Cells

AbstractSignaling through the pre-TCR is essential for early T cell development and is severely impaired in mice lacking the CD3γ chain of the pre-TCR. We here address the molecular mechanisms underlying this defect. Impaired pre-TCR signaling is shown to be associated with a profound increase in the number of apoptotic CD4−CD8− (DN) thymocytes. Introduction of p53 deficiency into CD3γ-deficient mice completely reverses the cell survival defect in CD3γ-deficient DN thymocytes and rescues the block in pre-T cell differentiation. In addition, the CD4+CD8+ (DP) compartment is expanded to its normal size. These findings suggest that the pre-TCR regulates progression through the DNA-damage checkpoint of the DN to DP transition by inactivating p53

Whole-blood transcriptomic signatures induced during immunization by chloroquine prophylaxis and Plasmodium falciparum sporozoites

A highly effective vaccine that confers sterile protection to malaria is urgently needed. Immunization under chemoprophylaxis with sporozoites (CPS) consistently confers high levels of protection in the Controlled Human Malaria infection (CHMI) model. To provide a broad, unbiased assessment of the composition and kinetics of direct ex vivo human immune responses to CPS, we profiled whole-blood transcriptomes by RNA-seq before and during CPS immunization and following CHMI challenge. Differential expression of genes enriched in modules related to T cells, NK cells, protein synthesis, and mitochondrial processes were detected in fully protected individuals four weeks after the first immunization. Non-protected individuals demonstrated transcriptomic changes after the third immunization and the day of treatment, with upregulation of interferon and innate inflammatory genes and downregulation of B-cell signatures. Protected individuals demonstrated more significant interactions between blood transcription modules compared to non-protected individuals several weeks after the second and third immunizations. These data provide insight into the molecular and cellular basis of CPS-induced immune protection from P. falciparum infection

Gene expression profiles classifying clinical stages of tuberculosis and monitoring treatment responses in Ethiopian HIV-negative and HIV-positive cohorts.

BACKGROUND: Validation of previously identified candidate biomarkers and identification of additional candidate gene expression profiles to facilitate diagnosis of tuberculosis (TB) disease and monitoring treatment responses in the Ethiopian context is vital for improving TB control in the future. METHODS: Expression levels of 105 immune-related genes were determined in the blood of 80 HIV-negative study participants composed of 40 active TB cases, 20 latent TB infected individuals with positive tuberculin skin test (TST+), and 20 healthy controls with no Mycobacterium tuberculosis (Mtb) infection (TST-), using focused gene expression profiling by dual-color Reverse-Transcription Multiplex Ligation-dependent Probe Amplification assay. Gene expression levels were also measured six months after anti-TB treatment (ATT) and follow-up in 38 TB patients. RESULTS: The expression of 15 host genes in TB patients could accurately discriminate between TB cases versus both TST+ and TST- controls at baseline and thus holds promise as biomarker signature to classify active TB disease versus latent TB infection in an Ethiopian setting. Interestingly, the expression levels of most genes that markedly discriminated between TB cases versus TST+ or TST- controls did not normalize following completion of ATT therapy at 6 months (except for PTPRCv1, FCGR1A, GZMB, CASP8 and GNLY) but had only fully normalized at the 18 months follow-up time point. Of note, network analysis comparing TB-associated host genes identified in the current HIV-negative TB cohort to TB-associated genes identified in our previously published Ethiopian HIV-positive TB cohort, revealed an over-representation of pattern recognition receptors including TLR2 and TLR4 in the HIV-positive cohort which was not seen in the HIV-negative cohort. Moreover, using ROC cutoff ≥ 0.80, FCGR1A was the only marker with classifying potential between TB infection and TB disease regardless of HIV status. CONCLUSIONS: Our data indicate that complex gene expression signatures are required to measure blood transcriptomic responses during and after successful ATT to fully diagnose TB disease and characterise drug-induced relapse-free cure, combining genes which resolve completely during the 6-months treatment phase of therapy with genes that only fully return to normal levels during the post-treatment resolution phase

Contributions of the T Cell Receptor–associated CD3γ–ITAM to Thymocyte Selection

The immunoreceptor tyrosine-based activation motifs (ITAMs) in the CD3 chains associated with the T cell receptor (TCR) are crucial for TCR signaling. To probe the role of the CD3γ–ITAM in T cell development, we created knock-in mice in which the CD3γ chain of the TCR complex is replaced by a mutant signaling-deficient CD3γ chain, lacking the CD3γ–ITAM. This mutation results in considerable impairment in positive selection in the polyclonal TCR repertoire. When CD3γ–ΔITAM mice are crossed to mice expressing transgenic F5 TCRs, their thymocytes are completely unable to perform positive selection in vivo in response to intrathymic ligands. Also, the in vitro positive selection response of double-positive (DP) thymocytes with F5–CD3γ–ΔITAM mutant receptors to their agonist ligand and many of its variants is severely impaired or abrogated. Yet, the binding and dissociation constants of agonist ligands for the F5 receptor are not affected by the CD3γ–ΔITAM mutation. Furthermore, DP thymocytes with mutant receptors can respond to agonist ligand with normal antigen sensitivity and to normal levels, as shown by their ability to induce CD69 up-regulation, TCR down-regulation, negative selection, and ZAP70 and c-Jun NH2-terminal kinase activation. In sharp contrast, induction of extracellular signal-regulated kinase (ERK) activation and linker for activation of T cells (LAT) phosphorylation are severely impaired in these cells. Together, these findings underscore that intrinsic properties of the TCR–CD3 complex regulate selection at the DP checkpoint. More importantly, this analysis provides the first direct genetic evidence for a role of the CD3γ–ITAM in TCR-driven thymocyte selection

Novel transcriptional signatures for sputum-independent diagnostics of tuberculosis in children

Pediatric tuberculosis (TB) is challenging to diagnose, confirmed by growth of Mycobacterium tuberculosis at best in 40% of cases. The WHO has assigned high priority to the development of non-sputum diagnostic tools. We therefore sought to identify transcriptional signatures in whole blood of Indian children, capable of discriminating intra-thoracic TB disease from other symptomatic illnesses. We investigated the expression of 198 genes in a training set, comprising 47 TB cases (19 definite/28 probable) and 36 asymptomatic household controls, and identified a 7- and a 10-transcript signature, both including NOD2, GBP5, IFITM1/3, KIF1B and TNIP1. The discriminatory abilities of the signatures were evaluated in a test set comprising 24 TB cases (17 definite/7 probable) and 26 symptomatic non-TB cases. In separating TB-cases from symptomatic non-TB cases, both signatures provided an AUC of 0.94 (95%CI, 0.88–1.00), a sensitivity of 91.7% (95%CI, 71.5–98.5) regardless of culture status, and 100% sensitivity for definite TB. The 7-transcript signature provided a specificity of 80.8% (95%CI, 60.0–92.7), and the 10-transcript signature a specificity of 88.5% (95%CI, 68.7–96.9%). Although warranting exploration and validation in other populations, our findings are promising and potentially relevant for future non-sputum based POC diagnostic tools for pediatric TB.publishedVersio

BLR1 and FCGR1A transcripts in peripheral blood associate with the extent of intrathoracic tuberculosis in children and predict treatment outcome

Biomarkers reflecting the extent of Mycobacterium tuberculosis-induced pathology and normalization during anti-tuberculosis treatment (ATT) would considerably facilitate trials of new treatment regimens and the identification of patients with treatment failure. Therefore, in a cohort of 99 Indian children with intrathoracic tuberculosis (TB), we performed blood transcriptome kinetic analysis during ATT to explore 1) the association between transcriptional biomarkers in whole blood (WB) and the extent of TB disease at diagnosis and treatment outcomes at 2 and 6 months, and 2) the potential of the biomarkers to predict treatment response at 2 and 6 months. We present the first data on the association between transcriptional biomarkers and the extent of TB disease as well as outcome of ATT in children: Expression of three genes down-regulated on ATT (FCGR1A, FPR1 and MMP9) exhibited a positive correlation with the extent of TB disease, whereas expression of eight up-regulated genes (BCL, BLR1, CASP8, CD3E, CD4, CD19, IL7R and TGFBR2) exhibited a negative correlation with the extent of disease. Baseline levels of these transcripts displayed an individual capacity >70% to predict the six-month treatment outcome. In particular, BLR1 and FCGR1A seem to have a potential in monitoring and perhaps tailoring future antituberculosis therapy

Host Gene Expression Kinetics During Treatment of Tuberculosis in HIV-Coinfected Individuals Is Independent of Highly Active Antiretroviral Therapy.

Background: Limitations in diagnostic tools to discriminate between active tuberculosis and latent Mycobacterium tuberculosis infection and for monitoring antituberculosis treatment responses are major challenges in tuberculosis control, especially in human immunodeficiency virus (HIV)-coinfected individuals. Methods: Expression levels of 105 immune-related genes were determined in 131 HIV-infected patients with active tuberculosis (n = 48), patients with latent M. tuberculosis infection (LTBI; n = 37), and controls with no M. tuberculosis infection (n = 46) in Addis Ababa, Ethiopia, using focused gene expression profiling with a dual-color reverse-transcription multiplex ligation-dependent probe amplification assay. Results: Within the cohort of HIV-positive subjects, the expression profiles of 7 genes at baseline (FCGR1A, RAB24, TLR1, TLR4, MMP9, NLRC4, and IL1B) could accurately discriminate between active tuberculosis and both latent and no M. tuberculosis infection, largely independently of (in)eligibility for highly active antiretroviral therapy (HAART). Six months after antituberculosis treatment, biomarker profiles of patients with tuberculosis became indistinguishable from those of patients with LTBI and controls. Importantly, host gene expression kinetics during antituberculosis treatment in HIV-coinfected individuals was found to be independent of HAART use. Conclusions: Blood transcriptomic profiles can potentially be used as biomarkers to discriminate the different clinical stages of tuberculosis in HIV-coinfected individuals and to monitor tuberculosis treatment responses in both HAART recipients and untreated individuals

Lawan penyakit demi PhD

Kuantan - Semangat yang tinggi untuk menamatkan pengajian di peringkat Doktor Falsafah (PhD) membuatkan pensyarah Bahasa Inggeris Universiti Tenaga Nasional (Uniten) Muadzam Shah, Dr. Umi Kalsom Masrom, 37, tabah melawan penyakit 'adhesion colic' yang dihidapinya dua tahun lalu

Transcriptomic evidence for modulation of host inflammatory responses during febrile Plasmodium falciparum malaria

Identifying molecular predictors and mechanisms of malaria disease is important for understanding how Plasmodium falciparum malaria is controlled. Transcriptomic studies in humans have so far been limited to retrospective analysis of blood samples from clinical cases. In this prospective, proof-of-principle study, we compared whole-blood RNA-seq profiles at pre-and post-infection time points from Malian adults who were either asymptomatic (n = 5) or febrile (n = 3) during their first seasonal PCR-positive P. falciparum infection with those from malaria-naïve Dutch adults after a single controlled human malaria infection (n = 5). Our data show a graded activation of pathways downstream of pro-inflammatory cytokines, with the highest activation in malaria-naïve Dutch individuals and significantly reduced activation in malaria-experienced Malians. Newly febrile and asymptomatic infections in Malians were statistically indistinguishable except for genes activated by pro-inflammatory cytokines. The combined data provide a molecular basis for the development of a pyrogenic threshold as individuals acquire immunity to clinical malaria