Mesenchymal Stromal Cell-Derived Extracellular Vesicles Pass through the Filtration Barrier and Protect Podocytes in a 3D Glomerular Model under Continuous Perfusion

Background: Dynamic cultures, characterized by continuous fluid reperfusion, elicit physiological responses from cultured cells. Mesenchymal stem cell-derived EVs (MSC-EVs) has been proposed as a novel approach in treating several renal diseases, including acute glomerular damage, by using traditional two-dimensional cell cultures and in vivo models. We here aimed to use a fluidic three-dimensional (3D) glomerular model to study the EV dynamics within the glomerular structure under perfusion. Methods: To this end, we set up a 3D glomerular model culturing human glomerular endothelial cells and podocytes inside a bioreactor on the opposite sides of a porous membrane coated with type IV collagen. The bioreactor was connected to a circuit that allowed fluid passage at the rate of 80 µl/min. To mimic glomerular damage, the system was subjected to doxorubicin administration in the presence of therapeutic MSC-EVs. Results: The integrity of the glomerular basal membrane in the 3D glomerulus was assessed by a permeability assay, demonstrating that the co-culture could limit the passage of albumin through the filtration barrier. In dynamic conditions, serum EVs engineered with cel-miR-39 passed through the glomerular barrier and transferred the exogenous microRNA to podocyte cell lines. Doxorubicin treatment increased podocyte apoptosis, whereas MSC-EV within the endothelial circuit protected podocytes from damage, decreasing cell death and albumin permeability. Conclusion: Using an innovative millifluidic model, able to mimic the human glomerular barrier, we were able to trace the EV passage and therapeutic effect in dynamic conditions.

Effect of a Gluten-Free Diet on Albuminuria in Children with Newly Diagnosed Celiac Disease

<b><i>Background and Objectives:</i></b> Altered gastrointestinal permeability in celiac disease (CD) is mediated by zonulin. The receptor for zonulin is expressed on podocytes. Therefore, we tested the effect of a gluten-free diet (GFD) on albuminuria in pediatric patients with newly diagnosed CD. <b><i>Methods:</i></b> We performed a cohort study comparing urinary albumin (μg): Cr (mg) ratio (ACR) in CD patients versus controls and in response to a GFD. <b><i>Results:</i></b> Children with CD (<i>n</i> = 46) had higher ACR than controls (<i>n</i> = 21), 20.2 ± 5.6 versus 8.4 ± 1.1 μg/mg, <i>p</i> = 0.16 and exceeded 30 μg/mg (microalbuminuria cutoff) in 7/46 cases. Seventeen patients had a follow-up assessment (interval 6.1 ± 0.7 months) on a GFD. Baseline ACR was 20.7 ± 5.2 that fell to 10.4 ± 1.5 μg/mg, <i>p</i> = 0.035. <b><i>Conclusion:</i></b> Children and adolescents with newly diagnosed CD have low-grade albuminuria that is numerically higher than controls and that declined after implementation of a GFD. CD may be associated with reversible defects in the glomerular barrier.

Soluble Urokinase-Type Plasminogen Activator Receptor (suPAR) is a Risk Indicator for eGFR Loss in Kidney Transplant Recipients

Background suPAR is a signaling protein of the Ly6/alpha-neurotoxin family. In the kidney, suPAR contributes to podocyte foot process effacement and glomerular barrier function disruption via activating αvβ3 integrin on the podocyte membrane. Its role in allograft function or transplant-specific outcomes needs clarification. Therefore, we prospectively investigated the prognostic relevance of suPAR in patients before and one year after kidney transplantation (KTx).Methods We included 100 patients who had received a kidney transplantation between 2013 and 2015. The plasma concentration of suPAR was measured using an uPAR ELISA assay.Results In patients who had received a living donation (LD), pre-transplant suPAR levels were significantly lower than those who had received a deceased donation (DD). After KTx, suPAR levels significantly declined in LD and DD recipients, without a detectable difference between LD and DD recipients. Higher suPAR levels in recipients one year after KTx were associated with a more severe eGFR loss in the following three years (n = 82, p = 0.021).Conclusions After KTx, suPAR levels drop significantly. Nevertheless, suPAR-levels above 6,212 pg/ml one year after KTx are independently associated with a nearly twice as fast loss of renal function > 30% (p < 0.001). Therefore, suPAR level at one year mark might be a risk indicator of increased eGFR loss.

Autoimmunity in Focal Segmental Glomerulosclerosis: A Long-Standing Yet Elusive Association

Focal segmental glomerulosclerosis (FSGS) is a histological term that describes a pathologic renal entity affecting both adults and children, with a wide array of possible underlying etiologies. Podocyte damage with scarring, the hallmark of this condition, leads to altered permeability of the glomerular barrier, which may result in massive proteinuria and relentless renal function deterioration. A definite cause of focal segmental glomerulosclerosis can be confirmed in a minority of cases, while most forms have been traditionally labeled as primary or idiopathic. Despite this definition, increasing evidence indicates that primary forms are a heterogenous group rather than a single disease entity: several circulating factors that may affect glomerular permeability have been proposed as potential culprits, and both humoral and cellular immunity have been implicated in the pathogenesis of the disease. Consistently, immunosuppressive drugs are considered as the cornerstone of treatment for primary focal segmental glomerulosclerosis, but response to these agents and long-term outcomes are highly variable. In this review we provide a summary of historical and recent advances on the pathogenesis of primary focal segmental glomerulosclerosis, focusing on implications for its differential diagnosis and treatment.

Risedronate Attenuates Podocyte Injury in Phosphate Transporter-Overexpressing Rats

Osteoporosis patients with chronic kidney disease (CKD) are becoming common in our superaging society. Renal dysfunction causes phosphorus accumulation in the circulating plasma and leads to the development of CKD-mineral bone disorder (MBD). We have previously reported that type III Pi transporter-overexpressing transgenic (Pit-1 TG) rats manifest phosphate (Pi)-dependent podocyte injury. In the present study, we explored the effect of risedronate on Pi-induced podocyte injury in vivo. Pit-1 TG rats and wild-type rats at 5 weeks old were divided into a risedronate-treated group and an untreated group. We subcutaneously administered 5 μg/kg body weight of risedronate or saline twice a week during the experimental period. Risedronate did not alter serum creatinine levels at 5, 8, and 12 weeks of age. However, electron microscopy images showed that thickening of the glomerular basement membrane was improved in the risedronate treatment group. Furthermore, immunostaining for podocyte injury markers revealed that both desmin- and connexin43-positive areas were smaller in the risedronate-treated group than in the untreated group, suggesting that bisphosphonates could rescue Pi-induced podocyte injury. In conclusion, our findings suggest that risedronate could maintain glomerular barrier function by rescuing Pi-induced podocyte injury.

A microphysiological system to investigate the pressure dependent filtration at an artificial glomerular kidney barrier

Chronic kidney disease (CKD) is a global health problem that affects around 11 to 13% of the world’s population and more than 18% of European citizens. Characteristic syndromes of CKD during all stages of the disease are proteinuria and ongoing glomerular dysfunction caused by cellular damages at the glomerular filtration barrier. While some rare cases of the disease are correlated to genetic depositions the majority of cases are caused by diabetes, glomerulosclerosis, high blood pressure and glomerulonephritis. Thus, recapitulating the interplay of high blood pressure and changes at the glomerular filtration barrier in vitro seems an adequate way to mimic CKD. Here we present a microphysiological system of the glomerular filter that is capable to simulate high blood pressure at the glomerular filtration barrier in vitro. It consists of a closed loop microfluidic circuit with an integrated pneumatically driven heart like micro pump that constantly circulates the cell culture media at the blood site of the glomerular barrier. The ThinCert™ insert could be reversibly integrated into a holder system that ensures the correct position of the insert within the microfluidic circuit. By using different modulations of the integrated pneumatic micro pump different physiological and pathophysiological conditions e.g. hypertonic stress, like in CKD, could be applied. The influence of hypertonic conditions on the filtration above the barrier was studied by changes of TEER values and measurement of the flux of fluorescent labelled albumin through the cellular barrier.