Y-box protein-1 is actively secreted through a non-classical pathway and acts as an extracellular mitogen

Y-box protein (YB)-1 of the cold-shock protein family functions in gene transcription and RNA processing. Extracellular functions have not been reported, but the YB-1 staining pattern in inflammatory glomerular diseases, without adherence to cell boundaries, suggests an extracellular occurrence. Here, we show the secretion of YB-1 by mesangial and monocytic cells after inflammatory challenges. It should be noted that YB-1 was secreted through a non-classical mode resembling that of the macrophage migration inhibitory factor. YB-1 release requires ATP-binding cassette transporters, and microvesicles protect YB-1 from protease degradation. Two lysine residues in the YB-1 carboxy-terminal domain are crucial for its release, probably because of post-translational modifications. The addition of purified recombinant YB-1 protein to different cell types results in increased DNA synthesis, cell proliferation and migration. Thus, the non-classically secreted YB-1 has extracellular functions and exerts mitogenic as well as promigratory effects in inflammation

SARS-CoV-2 infects the human kidney and drives fibrosis in kidney organoids

Kidney failure is frequently observed during and after COVID-19, but it remains elusive whether this is a direct effect of the virus. Here, we report that SARS-CoV-2 directly infects kidney cells and is associated with increased tubule-interstitial kidney fibrosis in patient autopsy samples. To study direct effects of the virus on the kidney independent of systemic effects of COVID-19, we infected human induced pluripotent stem cell-derived kidney organoids with SARS-CoV-2. Single cell RNA-sequencing indicated injury and dedifferentiation of infected cells with activation of pro-fibrotic signaling pathways. Importantly, SARS-CoV-2 infection also led to increased collagen 1 protein expression in organoids. A SARS-CoV-2 protease inhibitor was able to ameliorate the infection of kidney cells by SARS-CoV-2. Our results suggest that SARS-CoV-2 can directly infect kidney cells and induce cell injury with subsequent fibrosis. These data could explain both acute kidney injury in COVID-19 patients and the development of chronic kidney disease in Long-COVID

Untersuchungen zur Rolle von Notch-3 Rezeptoren in experimentellen Schädigungsmodellen der Niere

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

3D printed tubulointerstitium chip as an in vitro testing platform

Chronic kidney disease (CKD) ranks as the twelfth leading cause of death worldwide and represents a major global health problem with still rather limited treatment options. The development of new in vitro models replicating defined segments of the kidney functional units, i.e., the nephrons, in a physiologically relevant and reproducible manner could facilitate drug testing and development. We aimed at producing an in vitro organ-on-a-chip platform with extrusion-based 3D printing approach with sacrificial components. We optimized the manufacturing of the tubular platform produced by printing sacrificial fibers in varying diameters, which provide a suitable structure for cell adhesion and proliferation. The manufactured chip platform was seeded with primary murine tubular epithelial cells and human umbilical vein endothelial cells. The effect of channel geometry, its reproducibility, coatings for optimal cell adhesion, and specific cell markers were investigated. The developed chip presents single and dual channels, mimicking segments of a renal tubule and the capillary network. Together with an extracellular matrix gel analogue placed in the middle of the two channels we aimed at mimicking the renal tubulointerstitium in vitro. The proposed 3D printed platform enables perfusable circular cross-section channels with fully automated, rapid and reproducible manufacturing processes at low costs. In conclusion, this kidney tubulointerstitium on-a-chip provides the first step toward the production of more complex in vitro models for drug testing and identification. This article is protected by copyright. All rights reserved