Response to acute hyperglycemia and high fructose in cultured tenocytes.

High monosaccharide levels are intimately associated with diabetes and impact tendon cells through inflammation and impairment in metabolic homeostasis. Experiments were designed to understand the responses elicited by cultured tenocytes under monosaccharide stress induced by hyperglycemia and hyperfructosemia. We simulated hyperglycemia and hyperfructosemia in vitro by treating tenocytes with media containing sublethal concentrations of glucose and fructose, respectively. Exposure of tenocytes to high glucose and high fructose altered the levels of IL-1ß, IL-2, IL-6, IL10 and IL-17A. AMPK expression was increased in high-glucose and decreased in high-fructose groups. High fructose increased the level of IRS-1 compared with the control. Increased mitochondrial superoxide levels and compromised mitochondrial membrane integrity were exhibited by both the groups. The findings from the network analysis revealed many altered genes that are related to pathways for enzyme-linked receptor protein signaling, positive regulation of metabolic processes, transmembrane receptor tyrosine kinase pathway, insulin receptor signaling and regulation of cytokine production. Overall, the data suggest that the tenocytes under high monosaccharide levels exhibit survival responses by altering the expression status of cytokines and metabolic mediators that are involved in the underlying pathogenesis of tendinopathy.

Ischemia challenged epicardial adipose tissue stem cells-derived extracellular vesicles alter the gene expression of cardiac fibroblasts to cardiomyocyte like phenotype.

The present study hypothesizes that the ischemic insults activate epicardial adipose tissue-derived stem cells (EATDS) to secrete extracellular vesicles (EVs) packed with regenerative mediators to alter the gene expression in cardiac fibroblasts (CF). EATDS and CF were isolated from hyperlipidemic microswine and EVs were harvested from control, simulated ischemia (ISC) and ischemia-reperfusion (ISC/R) groups. The in vitro interaction between ISC-EVs and CF resulted in the upregulation of cardiomyocyte-specific transcription factors including GATA4, Nkx2.5, IRX4, and TBX5 in CF and the healing marker αSMA and the downregulation of fibroblast biomarkers such as vimentin, FSP1, and podoplanin and the cardiac biomarkers such as troponin-I and connexin-43. These results suggest a cardiomyocyte-like phenotype as confirmed by immunostaining and Western blot. The LC-MS/MS analysis of ISC-EVs LGALS1, PRDX2, and CCL2 to be the potent protein mediators which are intimately involved in versatile regenerative processes and connected with a diverse array of regenerative genes. Moreover, the LGALS1+, PRDX2+, and CCL2+ EATDS phenotypes were deciphered at single cell resolution revealing corresponding sub-populations with superior healing potential. Overall, the findings unveiled the healing potential of EATDS-derived EVs and sub-populations of regenerative EATDS promising novel translational opportunities in improved cardiac healing following ischemic injury.

Expression of specific HLA class II alleles is associated with an increased risk for active tuberculosis and a distinct gene expression profile.

Several HLA allelic variants have been associated with protection from or susceptibility to infectious and autoimmune diseases. Here, we examined whether specific HLA alleles would be associated with different Mycobacterium tuberculosis (Mtb) infection outcomes. The HLA alleles present at the -A, -B, -C, -DPA1, -DPB1, -DQA1, -DQB1, -DRB1, and -DRB3/4/5 loci were determined in a cohort of 636 individuals with known Mtb infection outcomes from South Africa and the United States. Among these individuals, 203 were QuantiFERON (QFT) negative, and 433 were QFT positive, indicating Mtb exposure. Of these, 99 QFT positive participants either had active tuberculosis (TB) upon enrollment or were diagnosed in the past. We found that DQA1*03:01, DPB1*04:02, and DRB4*01:01 were significantly more frequent in individuals with active TB (susceptibility alleles), as judged by Odds Ratios and associated p-values, while DPB1*105:01 was associated with protection from active TB. Peripheral blood mononuclear cells (PMBCs) from a subset of individuals were stimulated with Mtb antigens, revealing individuals who express any of the three susceptibility alleles were associated with lower magnitude of responses. Furthermore, we defined a gene signature associated with individuals expressing the susceptibility alleles that was characterized by lower expression of APC-related genes. In summary, we have identified specific HLA alleles associated with susceptibility to active TB and found that the expression of these alleles was associated with a decreased Mtb-specific T cell response and a specific gene expression signature. These results will help understand individual risk factors in progressing to active TB.

T-cell deficiency and hyperinflammatory monocyte responses associate with Mycobacterium avium complex lung disease.

Immunological mechanisms of susceptibility to nontuberculous mycobacterial (NTM) disease are poorly understood. To understand NTM pathogenesis, we evaluated innate and antigen-specific adaptive immune responses to Mycobacterium avium complex (MAC) in asymptomatic individuals with a previous history of MAC lung disease (MACDZ). We hypothesized that Mav-specific immune responses are associated with susceptibility to MAC lung disease. We measured MAC-, NTM-, or MAC/Mtb-specific T-cell responses by cytokine production, expression of surface markers, and analysis of global gene expression in 27 MACDZ individuals and 32 healthy controls. We also analyzed global gene expression in Mycobacterium avium-infected and uninfected peripheral blood monocytes from 17 MACDZ and 17 healthy controls. We were unable to detect increased T-cell responses against MAC-specific reagents in MACDZ compared to controls, while the responses to non-mycobacteria derived antigens were preserved. MACDZ individuals had a lower frequency of Th1 and Th1* T-cell populations. In addition, MACDZ subjects had lower transcriptional responses in PBMCs stimulated with a mycobacterial peptide pool (MTB300). By contrast, global gene expression analysis demonstrated upregulation of proinflammatory pathways in uninfected and M. avium-infected monocytes, i.e. a hyperinflammatory in vitro response, derived from MACDZ subjects compared to controls. Together, these data suggest a novel immunologic defect which underlies MAC pathogenesis and includes concurrent innate and adaptive dysregulation which persists years after completion of treatment.

CD4+CCR6+ T cells dominate the BCG-induced transcriptional signature.

BACKGROUND: The century-old Mycobacterium bovis Bacillus Calmette-Guerin (BCG) remains the only licensed vaccine against tuberculosis (TB). Despite this, there is still a lot to learn about the immune response induced by BCG, both in terms of phenotype and specificity. METHODS: We investigated immune responses in adult individuals pre and 8 months post BCG vaccination. We specifically determined changes in gene expression, cell subset composition, DNA methylome, and the TCR repertoire induced in PBMCs and CD4 memory T cells associated with antigen stimulation by either BCG or a Mycobacterium tuberculosis (Mtb)-derived peptide pool. FINDINGS: Following BCG vaccination, we observed increased frequencies of CCR6+ CD4 T cells, which includes both Th1* (CXCR3+CCR6+) and Th17 subsets, and mucosal associated invariant T cells (MAITs). A large number of immune response genes and pathways were upregulated post BCG vaccination with similar patterns observed in both PBMCs and memory CD4 T cells, thus suggesting a substantial role for CD4 T cells in the cellular response to BCG. These upregulated genes and associated pathways were also reflected in the DNA methylome. We described both qualitative and quantitative changes in the BCG-specific TCR repertoire post vaccination, and importantly found evidence for similar TCR repertoires across different subjects. INTERPRETATION: The immune signatures defined herein can be used to track and further characterize immune responses induced by BCG, and can serve as reference for benchmarking novel vaccination strategies.

Treg cells in atherosclerosis.

Atherosclerosis involves both innate and adaptive immunity. Here, we provide an overview of the role of regulatory T (Treg) cells in atherosclerotic diseases. Treg cells and their inhibitory cytokines, IL-10 and TGF-ß, have been identified in atherosclerotic lesions and to inhibit progression through lipoprotein metabolism modulation. Treg cells have also been found to convert to T follicular helper (Tfh) cells and promote atherosclerosis progression. Treg cell involvement in different stages of atherosclerotic progression and Treg cell-mediated modulation of plaque development occurs via inflammation suppression and atheroma formation has been focused. Moreover, existing knowledge suggests that Treg cells are likely involved in the pathology of other specific circumstances including in-stent restenosis, neointimal hyperplasia, vessel graft failure, and ischemic arterial injury; however, there remain gaps regarding their specific contribution. Hence, advancements in the knowledge regarding Treg cells in diverse aspects of atherosclerosis offer translational significance for the management of atherosclerosis and associated diseases.

Quantitative and Qualitative Perturbations of CD8+ MAITs in Healthy Mycobacterium tuberculosis-Infected Individuals.

CD8 T cells are considered important contributors to the immune response against Mycobacterium tuberculosis, yet limited information is currently known regarding their specific immune signature and phenotype. In this study, we applied a cell population transcriptomics strategy to define immune signatures of human latent tuberculosis infection (LTBI) in memory CD8 T cells. We found a 41-gene signature that discriminates between memory CD8 T cells from healthy LTBI subjects and uninfected controls. The gene signature was dominated by genes associated with mucosal-associated invariant T cells (MAITs) and reflected the lower frequency of MAITs observed in individuals with LTBI. There was no evidence for a conventional CD8 T cell-specific signature between the two cohorts. We, therefore, investigated MAITs in more detail based on Vα7.2 and CD161 expression and staining with an MHC-related protein 1 (MR1) tetramer. This revealed two distinct populations of CD8+Vα7.2+CD161+ MAITs: MR1 tetramer+ and MR1 tetramer-, which both had distinct gene expression compared with memory CD8 T cells. Transcriptomic analysis of LTBI versus noninfected individuals did not reveal significant differences for MR1 tetramer+ MAITs. However, gene expression of MR1 tetramer- MAITs showed large interindividual diversity and a tuberculosis-specific signature. This was further strengthened by a more diverse TCR-α and -ß repertoire of MR1 tetramer- cells as compared with MR1 tetramer+ Thus, circulating memory CD8 T cells in subjects with latent tuberculosis have a reduced number of conventional MR1 tetramer+ MAITs as well as a difference in phenotype in the rare population of MR1 tetramer- MAITs compared with uninfected controls.

α-Synuclein-specific T cell reactivity is associated with preclinical and early Parkinson's disease.

A diagnosis of motor Parkinson's disease (PD) is preceded by a prolonged premotor phase with accumulating neuronal damage. Here we examined the temporal relation between α-synuclein (α-syn) T cell reactivity and PD. A longitudinal case study revealed that elevated α-syn-specific T cell responses were detected prior to the diagnosis of motor PD, and declined after. The relationship between T cell reactivity and early PD in two independent cohorts showed that α-syn-specific T cell responses were highest shortly after diagnosis of motor PD and then decreased. Additional analysis revealed significant association of α-syn-specific T cell responses with age and lower levodopa equivalent dose. These results confirm the presence of α-syn-reactive T cells in PD and show that they are most abundant immediately after diagnosis of motor PD. These cells may be present years before the diagnosis of motor PD, suggesting avenues of investigation into PD pathogenesis and potential early diagnosis.