Human pluripotent stem cell-derived kidney organoids for personalized congenital and idiopathic nephrotic syndrome modeling.

Nephrotic syndrome (NS) is characterized by severe proteinuria as a consequence of kidney glomerular injury due to podocyte damage. In vitro models mimicking in vivo podocyte characteristics are a prerequisite to resolve NS pathogenesis. The detailed characterization of organoid podocytes resulting from a hybrid culture protocol showed a podocyte population that resembles adult podocytes and was superior compared with 2D counterparts, based on single-cell RNA sequencing, super-resolution imaging and electron microscopy. In this study, these next-generation podocytes in kidney organoids enabled personalized idiopathic nephrotic syndrome modeling, as shown by activated slit diaphragm signaling and podocyte injury following protamine sulfate, puromycin aminonucleoside treatment and exposure to NS plasma containing pathogenic permeability factors. Organoids cultured from cells of a patient with heterozygous NPHS2 mutations showed poor NPHS2 expression and aberrant NPHS1 localization, which was reversible after genetic correction. Repaired organoids displayed increased VEGFA pathway activity and transcription factor activity known to be essential for podocyte physiology, as shown by RNA sequencing. This study shows that organoids are the preferred model of choice to study idiopathic and congenital podocytopathies.

Human scattered tubular cells represent a heterogeneous population of glycolytic dedifferentiated proximal tubule cells.

Scattered tubular cells (STCs) are a phenotypically distinct cell population in the proximal tubule that increase in number after acute kidney injury. We aimed to characterize the human STC population. Three-dimensional human tissue analysis revealed that STCs are preferentially located within inner bends of the tubule and are barely present in young kidney tissue (<2 years), and their number increases with age. Increased STC numbers were associated with acute tubular injury (kidney injury molecule 1) and interstitial fibrosis (alpha smooth muscle actin). Isolated CD13+ CD24- CD133- proximal tubule epithelial cells (PTECs) and CD13+ CD24+ and CD13+ CD133+ STCs were analyzed using RNA sequencing. Transcriptome analysis revealed an upregulation of nuclear factor κB, tumor necrosis factor alpha, and inflammatory pathways in STCs, whereas metabolism, especially the tricarboxylic acid cycle and oxidative phosphorylation, was downregulated, without showing signs of cellular senescence. Using immunostaining and a publicly available single-cell sequencing database of human kidneys, we demonstrate that STCs represent a heterogeneous population in a transient state. In conclusion, STCs are dedifferentiated PTECs showing a metabolic shift toward glycolysis, which could facilitate cellular survival after kidney injury. © 2022 The Authors. The Journal of Pathology published by John Wiley & Sons Ltd on behalf of The Pathological Society of Great Britain and Ireland.

Establishment and characterization of a novel conditionally immortalized human parietal epithelial cell line.

Parietal epithelial cells (PECs) are epithelial cells in the kidney, surrounding Bowman's space. When activated, PECs increase in cell volume, proliferate, migrate to the glomerular tuft and excrete extracellular matrix. Activated PECs are crucially involved in the formation of sclerotic lesions, seen in focal segmental glomerulosclerosis (FSGS). In FSGS, a number of glomeruli show segmental sclerotic lesions. Further disease progression will lead to increasing number of involved glomeruli and gradual destruction of the affected glomeruli. Although the involvement of PECs in FSGS has been acknowledged, little is known about the molecular processes driving PEC activation. To get more insights in this process, accurate in vivo and in vitro models are needed. Here, we describe the development and characterization of a novel conditionally immortalized human PEC (ciPEC) line. We demonstrated that ciPECs are differentiated when grown under growth-restrictive conditions and express important PEC-specific markers, while lacking podocyte and endothelial markers. In addition, ciPECs showed PEC-like morphology and responded to IL-1ß treatment. We therefore conclude that we have successfully generated a novel PEC line, which can be used for future studies on the role of PECs in FSGS.

Understanding the pathobiology in patent ductus arteriosus in prematurity-beyond prostaglandins and oxygen.

The ductus arteriosus (DA) is probably the most intriguing vessel in postnatal hemodynamic transition. DA patency in utero is an active state, in which prostaglandin E2 (PGE2) and nitric monoxide (NO), play an important role. Since the DA gets programmed for postnatal closure as gestation advances, in preterm infants the DA frequently remains patent (PDA). PGE2 exposure programs functional postnatal closure by inducing gene expression of ion channels and phosphodiesterases and anatomical closure by inducing intimal thickening. Postnatally, oxygen inhibits potassium and activates calcium channels, which ultimately leads to a rise in intracellular calcium concentration consequently inducing phosphorylation of the myosin light chain and thereby vasoconstriction of the DA. Since ion channel expression is lower in preterm infants, oxygen induced functional vasoconstriction is attenuated in comparison with full term newborns. Furthermore, the preterm DA is more sensitive to both PGE2 and NO compared to the term DA pushing the balance toward less constriction. In this review we explain the physiology of DA patency in utero and subsequent postnatal functional closure. We will focus on the pathobiology of PDA in preterm infants and the (un)intended effect of antenatal exposure to medication on both fetal and neonatal DA vascular tone.

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 profibrotic 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.