Defective Mitochondrial Quality Control during Dengue Infection Contributes to Disease Pathogenesis.

Mitochondrial fitness is governed by mitochondrial quality control pathways comprising mitochondrial dynamics and mitochondrial-selective autophagy (mitophagy). Disruption of these processes has been implicated in many human diseases, including viral infections. Here, we report a comprehensive analysis of the effect of dengue infection on host mitochondrial homeostasis and its significance in dengue disease pathogenesis. Despite severe mitochondrial stress and injury, we observed that the pathways of mitochondrial quality control and mitochondrial biogenesis are paradoxically downregulated in dengue-infected human liver cells. This leads to the disruption of mitochondrial homeostasis and the onset of cellular injury and necrotic death in the infected cells. Interestingly, dengue promotes global autophagy but selectively disrupts mitochondrial-selective autophagy (mitophagy). Dengue downregulates the expression of PINK1 and Parkin, the two major proteins involved in tagging the damaged mitochondria for elimination through mitophagy. Mitophagy flux assays also suggest that Parkin-independent pathways of mitophagy are also inactive during dengue infection. Dengue infection also disrupts mitochondrial biogenesis by downregulating the master regulators PPARγ and PGC1α. Dengue-infected cells release mitochondrial damage-associated molecular patterns (mtDAMPs) such as mitochondrial DNA into the cytosol and extracellular milieu. Furthermore, the challenge of naive immune cells with culture supernatants from dengue-infected liver cells was sufficient to trigger proinflammatory signaling. In correlation with our in vitro observations, dengue patients have high levels of cell-free mitochondrial DNA in their blood in proportion to the degree of thrombocytopenia. Overall, our study shows how defective mitochondrial homeostasis in dengue-infected liver cells can drive dengue disease pathogenesis. IMPORTANCE Many viruses target host cell mitochondria to create a microenvironment conducive to viral dissemination. Dengue virus also exploits host cell mitochondria to facilitate its viral life cycle. Dengue infection of liver cells leads to severe mitochondrial injury and inhibition of proteins that regulate mitochondrial quality control and biogenesis, thereby disrupting mitochondrial homeostasis. A defect in mitochondrial quality control leads to the accumulation of damaged mitochondria and promotes cellular injury. This leads to the release of mitochondrial damage-associated molecular patterns (mt-DAMPs) into the cell cytoplasm and extracellular milieu. These mt-DAMPs activate the naive immune cells and trigger proinflammatory signaling, leading to the release of cytokines and chemokines, which may trigger systemic inflammation and contribute to dengue disease pathogenesis. In correlation with this, we observed high levels of cell-free mitochondrial DNA in dengue patient blood. This study provides insight into how the disruption of mitochondrial quality control in dengue-infected cells can trigger inflammation and drive dengue disease pathogenesis.

Perturbations in spike-specific peripheral T follicular helper cells in SARS-CoV2 breakthrough convalescent individuals immunized by BBV152 vaccine.

Severe acute respiratory syndrome coronavirus 2 (SARS-Cov2) infection has caused an increase in mortality and morbidity, but with vaccination, the disease severity has significantly reduced. With the emergence of various variants of concern (VOCs), the vaccine breakthrough infection has also increased. Here we studied circulating spike-specific T follicular response (cTfh) in infection-naïve vaccinees and convalescent vaccinees (individuals who got the Delta breakthrough infection after two doses of BBV152 vaccine) to understand their response as they are the most crucial cells that are involved in vaccine-mediated protection by helping in B-cell maturation. Our results indicated that cTfh cells in both the groups recognized the wild-type and Delta spike protein but memory response to the wild-type spike was superior in infection-naïve than in the convalescent group. The cytokine response, particularly interleukin-21 (IL-21) from cTfh, was also higher in infection-naïve than in convalescent vaccinees, indicating a dampened cTfh response in convalescent vaccinees after breakthrough infection. Also, there was a positive correlation between IL-21 from cTfh cells and neutralizing antibodies of infection-naïve vaccinees. Multiple cytokine analysis also revealed higher inflammation in convalescent vaccinees. Our data indicated that the necessity of a third booster dose may be individual-specific depending on the steady-state functional phenotype of immune cells.

IL-21, Inflammatory Cytokines and Hyperpolarized CD8+ T Cells Are Central Players in Lupus Immune Pathology.

Systemic lupus erythematous (SLE) is a chronic autoimmune disorder, broadly characterized by systemic inflammation along with heterogeneous clinical manifestations, severe morbidity, moribund organ failure and eventual mortality. In our study, SLE patients displayed a higher percentage of activated, inflamed and hyper-polarized CD8+ T cells, dysregulated CD8+ T cell differentiation, significantly elevated serum inflammatory cytokines and higher accumulation of cellular ROS when compared to healthy controls. Importantly, these hyper-inflammatory/hyper-polarized CD8+ T cells responded better to an antioxidant than to an oxidant. Terminally differentiated Tc1 cells also showed plasticity upon oxidant/antioxidant treatment, but that was in contrast to the SLE CD8+ T cell response. Our studies suggest that the differential phenotype and redox response of SLE CD8+ T cells and Tc1 cells could be attributed to their cytokine environs during their respective differentiation and eventual activation environs. The polarization of Tc1 cells with IL-21 drove hyper-cytotoxicity without hyper-polarisation suggesting that the SLE inflammatory cytokine environment could drive the extreme aberrancy in SLE CD8+ T cells.

IL-21/23 axis modulates inflammatory cytokines and RANKL expression in RA CD4+ T cells via p-Akt1 signaling.

Introduction: CD4+ T cells are critically involved in the pathogenesis of Rheumatoid Arthritis; an autoimmune disorder characterized by joint inflammation and bone degeneration. In this study, we focused on the critical role of cytokines, IL-21 and IL-23 in facilitating the aberrant status of RA Th17-like cells and report their significant contribution(s) in modulating the expression of inflammatory cytokines and RANKL. Methods: Blood and synovial fluid collected from a total of 167 RA patients and 25 healthy volunteers were assessed for various inflammatory markers and RANKL expression in plasma and CD4+ T cells. Subsequent ex vivo studies examined the role of specific cytokines, IL-21 and IL-23 in mediating inflammation and RANKL upregulation by blocking their expression with neutralizing antibodies in RA CD4+ T cells and terminally differentiated human Th17 cells. Further, the role of p-Akt1 as a signalling target downstream of IL-21 and IL-23 was evinced with IL-21 and IL-23 inhibition and phospho Akt-1/2 kinase inhibitor. Results: Our observations highlighted the augmented inflammatory cytokine levels in plasma and an aberrant CD4+ T cell phenotype expressing exaggerated inflammatory cytokines and membrane RANKL expression in RA as opposed to healthy controls. Neutralization of either IL-21 or IL-23 (p19 and p40) or both, resulted in downregulation of the cytokines, TNF-α, IFN-γ and IL-17 and RANKL expression in these cells, signifying the critical role of IL-21/23 axis in modulating inflammation and RANKL. Subsequent dissection of the signaling pathway found p-Akt1 as the key phosphoprotein downstream of both IL-21 and IL-23, capable of increasing inflammatory cytokines and RANKL production. Discussion: Our findings unequivocally identify IL-21/23 axis in RA CD4+ T cells as a key regulator dictating two critical processes i.e. exaggerated inflammation and higher RANKL expression and provide critical targets in their downstream signalling for therapeutic approaches.

Underlying Co-Morbidity Reveals Unique Immune Signatures in Type II Diabetes Patients Infected With SARS-CoV2.

Background: SARS-CoV2 infection in patients with comorbidities, particularly T2DM, has been a major challenge globally and has been shown to be associated with high morbidity and mortality. Here, we did whole blood immunophenotyping along with plasma cytokine, chemokine, antibody isotyping, and viral load from oropharyngeal swab to understand the immune pathology in the T2DM patients infected with SARS-CoV2. Methods: Blood samples from 25 Covid-19 positive patients having T2DM, 10 Covid-19 positive patients not having T2DM, and 10 Covid-19 negative, non-diabetic healthy controls were assessed for various immune cells by analyzing for their signature surface proteins in mass cytometry. Circulating cytokines, chemokines, and antibody isotypes were determined from plasma while viral copy number was determined from oropharyngeal swabs. All our representative data corroborated with laboratory findings. Results: Our observations encompass T2DM patients having elevated levels of both type I and type II cytokines and higher levels of circulating IgA, IgM, IgG1, and IgG2 as compared to NDM and healthy volunteers. They also displayed higher percentages of granulocytes, mDCs, plasmablasts, Th2-like cells, CD4+ EM cells, and CD8+ TE cells as compared to healthy volunteers. T2DM patients also displayed lower percentages of pDCs, lymphocytes, CD8+ TE cells, CD4+, and CD8+ EM. Conclusion: Our study demonstrated that patients with T2DM displayed higher inflammatory markers and a dysregulated anti-viral and anti-inflammatory response when compared to NDM and healthy controls and the dysregulated immune response may be attributed to meta inflammation.