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Severe COVID-19 is linked to both dysfunctional immune response and unrestrained immunopathology, and it remains unclear whether T cells contribute to disease pathology. Here, we combined single-cell transcriptomics and single-cell proteomics with mechanistic studies to assess pathogenic T cell functions and inducing signals. We identified highly activated, CD16+ T cells with increased cytotoxic functions in severe COVID-19. CD16 expression enabled immune complex-mediated, T cell receptor-independent degranulation and cytotoxicity not found in other diseases. CD16+ T cells from COVID-19 patients promoted microvascular endothelial cell injury and release of neutrophil and monocyte chemoattractants. CD16+ T cell clones persisted beyond acute disease maintaining their cytotoxic phenotype. Increased generation of C3a in severe COVID-19 induced activated CD16+ cytotoxic T cells. Proportions of activated CD16+ T cells and plasma levels of complement proteins upstream of C3a were associated with fatal outcome of COVID-19, supporting a pathological role of exacerbated cytotoxicity and complement activation in COVID-19., Generation of the C3a complement protein fragment by SARS-CoV-2 infection drives differentiation of a CD16-expressing T cell population that associates with severe COVID-19 disease outcomes.

Journal of cellular and molecular medicine :JCMM 21(12), 3494-3505 (2017). doi:10.1111/jcmm.13260, Published by Wiley-Blackwell, Hoboken, NJ

Kidney disease imparts a vast burden on affected individuals and the overall health care system. Progressive tubulointerstitial fibrosis represents a common final pathway observed in all forms of chronic kidney diseases, characterized by deposition of extracellular matrix, immune cell infiltration, fibroblast proliferation and activation, injury to the tubular epithelium and rarefaction of the peritubular microvasculature. In the recent past, Notch receptors, which are involved in the nephron development, attributed increasingly a significant role in the development of various kidney diseases, and also in the tubulointerstitial fibrosis. In the present work, the involvement of the receptor Notch-3 in the immune cell response and kidney fibrosis was examined. First evidence for an involvement of Notch-3 receptor on the fibrogenesis was shown by in vitro studies in cultured kidney cells. Stimulation with the central profibrotic mediator TGF-beta1 resulted in an increased expression of the receptor Notch-1 and Notch-3 and so on both transcription and protein level. This has been confirmed in vivo in two different animal models of renal damage, the nephrotoxic nephritis (NTN) and the unilateral ureteral obstruction (UUO). The NTN is a primary inflammatory, glomerular and thus proteinuric model whose profile corresponds to a rapidly progressive glomerulonephritis. In addition to the glomerular injury it results in tubulointerstitial immune cell infiltration and fibrosis. The murine UUO model primarily caused damage to the tubular structures and also reflects the hallmarks of tubulointerstitial fibrosis. In both models an activation of the Notch-3 receptor, and increased expression of Notch ligands and target genes was detectable. Striking is, that the increased expression of Notch-3 occurred cell-specific and so after ureteral obstruction detected only in isolated tubular and in some tubulointerstitial. cells. This suggests that the Notch-3-mediated regulatory events are controlled only by subpopulations of cells. We hypothesized that Notch-3 plays a functional relevance in the chemotactic response. Therefore, we studied animals with genetic depletion of Notch-3 receptor after UUO and NTN induction. We could show that Notch-3 knockout (Notch-3 KO) mice were protected from injury and tubular cell loss. The infiltration of inflammatory cells was significantly reduced and delayed, presumably due to a reduced chemokine synthesis and release. The Notch-3 knockout mice also exhibited a very significant reduction in the matrix accumulation that is due to reduced proliferation of interstitial cells. To investigate a direct influence of Notch-3 receptor on the cell proliferation, a cell model was established, which simulates an increased Notch-3 expression/ activation. Overexpression of constitutively active intracellular Notch-3 domain increased proliferation rate of renal tubular cells in vitro. This result was confirmed by incubating the cells with the Notch ligand Jagged-1. In addition to the in vivo and in vitro studies in a third approach, we correlated the expression of members of the Notch receptor family with human renal diseases. Human biopsies of patients with tubular and glomerular kidney diseases have been studied in terms of a gene array where mRNA expression of Notch signal pathways components were measured and hence a heat map was created. It was found that Notch-3 was expressed strongest by all of the Notch family receptors. In summary, our results show that the Notch-3 receptor plays an important role in the orchestration of tubulointerstitial, inflammatory response and in particular the monocytic cell infiltration is affected by Notch-3. This makes the Notch pathway an attractive target for future therapeutic interventions in which an excessive matrix deposition and the consequent destruction of the tissue architecture is to be countered.

International audience; Kidney diseases impart a vast burden on affected individuals and the overall health care system. Progressive loss of renal parenchymal cells and functional decline following injury are often observed. Notch-1 and -2 receptors are crucially involved in nephron development and contribute to inflammatory kidney diseases. We specifically determined the participation of receptor Notch-3 following tubulointerstitial injury and in inflammatory responses. Here we show by heat map analyses that Notch-3 transcripts are up-regulated in human kidney diseases. A similar response was corroborated with kidney cells following TGF-β exposure in vitro. The murine unilateral ureteral obstruction (UUO) model mirrors hallmarks of tubulointerstitial injury and damage. A subset of tubular and interstitial cells demonstrated up-regulated Notch-3 receptor expression in diseased animals. We hypothesized a relevance of Notch-3 receptors for the chemotactic response. To address this question, animals with genetic ablation of receptor Notch-3 were analysed following UUO. As a result, we found that Notch-3-deficient animals are protected from tubular injury and cell loss with significantly reduced interstitial collagen deposition. Monocytic cell infiltration was significantly reduced and retarded, likely due to abrogated chemokine synthesis. A cell model was set up that mimics enhanced receptor Notch-3 expression and activation. Here a pro-mitogenic response was seen with activated signalling in tubular cells and fibroblasts. In conclusion, Notch-3 receptor fulfils non-redundant roles in the inflamed kidney that may not be replaced by other Notch receptor family members. Thus, specific blockade of this receptor may be suitable as therapeutic option to delay progression of kidney disease.

EMBO molecular medicine (2020). doi:10.15252/emmm.201911021, Published by Wiley-VCH, Weinheim