Smad mediated regulation of inhibitor of DNA binding 2 and its role in phenotypic maintenance of human renal proximal tubule epithelial cells.

The basic-Helix-Loop-Helix family (bHLH) of transcriptional factors plays a major role in regulating cellular proliferation, differentiation and phenotype maintenance. The downregulation of one of the members of bHLH family protein, inhibitor of DNA binding 2 (Id2) has been shown to induce de-differentiation of epithelial cells. Opposing regulators of epithelial/mesenchymal phenotype in renal proximal tubule epithelial cells (PTEC), TGFβ1 and BMP7 also have counter-regulatory effects in models of renal fibrosis. We investigated the regulation of Id2 by these growth factors in human PTECs and its implication in the expression of markers of epithelial versus myofibroblastic phenotype. Cellular Id2 levels were reduced by TGFβ1 treatment; this was prevented by co-incubation with BMP7. BMP7 alone increased cellular levels of Id2. TGFβ1 and BMP7 regulated Id2 through Smad2/3 and Smad1/5 dependent mechanisms respectively. TGFβ1 mediated Id2 suppression was essential for α-SMA induction in PTECs. Although Id2 over-expression prevented α-SMA induction, it did not prevent E-cadherin loss under the influence of TGFβ1. This suggests that the loss of gate keeper function of E-cadherin alone may not necessarily result in complete EMT and further transcriptional re-programming is essential to attain mesenchymal phenotype. Although BMP7 abolished TGFβ1 mediated α-SMA expression by restoring Id2 levels, the loss of Id2 was not sufficient to induce α-SMA expression even in the context of reduced E-cadherin expression. Hence, a reduction in Id2 is critical for TGFβ1-induced α-SMA expression in this model of human PTECs but is not sufficient in it self to induce α-SMA even in the context of reduced E-cadherin

Systematic review and meta-analysis of COVID-19 and kidney transplant recipients, the South West London Kidney Transplant Network experience.

Introduction: There is paucity of literature comparing outcomes of kidney transplant patients with Covid-19 to that of dialysis and waitlisted patients. This report describes our data, provides comparative analysis, together with meta-analysis of published studies and describes our protocols to restart the transplant programme. Methods: Data were analysed on kidney transplant, dialysis and waitlisted patients tested positive for SARS-CoV-2 (naso-pharyngeal swab PCR) between March 1, 2020 and June 30, 2020 together with meta-analysis of 16 studies. Results: 23/1494 kidney transplant patients tested positive for SARS-CoV-2 compared to 123/1278 haemodialysis patients (1.5% vs 9.6%, p<0.001), 12/253 waitlisted patients (1.5% vs 4.7%, p=0.002). 19 required hospital admission, 6 died and 13 developed AKI. Overall case fatality ratio was 26.1% compared to patients on haemodialysis (27.6%, p=0.99) and waitlisted patients (8.3%, p=0.38). Within our entire cohort, 0.4% of transplant patients died compared to 0.4% of waitlisted patients and 2.7% of haemodialysis patients. Patients who died were older [Alive (median 71years) vs. Dead (median 59years), p=0.01]. In meta-analysis of 16 studies, including ours, pooled case fatality ratio was 24% [95%CI (19%, 28%)]; AKI proportion in 10 studies was 50% [95%CI (45%, 56%)], with some evidence against no heterogeneity between studies (p=0.02). Conclusions: From our cohort of transplant patients, a significantly lower proportion of patients contracted COVID-19 compared to waitlisted and dialysis patients. The case fatality ratio was comparable to that of dialysis cohort and pooled case fatality ratio from meta-analysis of 16 studies. The pooled AKI ratio in the meta-analysis was similar to our experience

The characteristics, dynamics and risk of death in COVID-19 positive dialysis patients in London, UK

Background: Dialysis patients, with frequent co-morbidities, advanced age and frailty, visiting treatment facilities frequently are perhaps more prone to SARS-Cov-2 infection and related death - the risk-factors and dynamics of which are unknown. The aim of this study was to investigate the hospital outcomes in SARS-CoV-2 infected dialysis patients. Methods: Data on 224 hemodialysis patients between 02/29/2020 and 05/15/2020 with confirmed SARS-CoV-2 were analyzed for outcomes and potential risk factors for death, using competing risk regression model assessed by sub-distribution hazards ratio (SHR). Results: Crude data analyses suggest an overall case fatality ratio of 22.7(95%CI(17.3-28.3)%) overall but that varies across age groups from 11.4(95%CI(0.9-9.2)) in 80 years; with 60% of deaths occurring in the first 15 days and 80% within 21 days indicating a rapid deterioration towards death after admission. Almost 90% of surviving patients were discharged within 28 days. Death was more likely than hospital discharge in more frail (WHO performance status 3-4) [SHR=2.16(1.25-3.74);p=0.006)], ischemic heart disease [SHR=2.28(1.32-3.94),p=0.003], cerebrovascular disease [SHR=2.11(1.20-3.72),p=0.010], smoking history [SHR=2.69(1.33-5.45),p=0.006], and (completely or partially) hospitalized patients [SHR=10.26.(3.10-33.94),p<.001]; and in patients with high CRP [SHR=1.35(1.10-1.67)] and high neutrophil:lymphocyte ratio [SHR=1.03(1.01-1.04),p<0.001]. Our data did not support differences in the risk of death associated with gender, ethnicity, dialysis vintage or other comorbidities. However, comparison with the entire dialysis population attending these hospitals, and 12.9% being affected, revealed that non-Caucasians (62% vs. 52% in all patients, p=0.001) and diabetic patients (54% vs. 22%, p<0.001) were disproportionately affected. Conclusion: This report discusses the outcomes of a large cohort of dialysis patients with SARS-CoV-2, infection affecting more diabetics and non-Caucasians; with a high case fatality ratio, which increased significantly with age, frailty, smoking, increasing CRP and neutrophil:lymphocyte ratio at presentation

Characterisation of feline renal cortical fibroblast cultures and their transcriptional response to transforming growth factor beta 1

Chronic kidney disease (CKD) is common in geriatric cats, and the most prevalent pathology is chronic tubulointerstitial inflammation and fibrosis. The cell type predominantly responsible for the production of extra-cellular matrix in renal fibrosis is the myofibroblast, and fibroblast to myofibroblast differentiation is probably a crucial event. The cytokine TGF-β1 is reportedly the most important regulator of myofibroblastic differentiation in other species. The aim of this study was to isolate and characterise renal fibroblasts from cadaverous kidney tissue of cats with and without CKD, and to investigate the transcriptional response to TGF-β1

A Prokaryotic S1P Lyase Degrades Extracellular S1P In Vitro and In Vivo: Implication for Treating Hyperproliferative Disorders

Sphingosine-1-phosphate (S1P) regulates a broad spectrum of fundamental cellular processes like proliferation, death, migration and cytokine production. Therefore, elevated levels of S1P may be causal to various pathologic conditions including cancer, fibrosis, inflammation, autoimmune diseases and aberrant angiogenesis. Here we report that S1P lyase from the prokaryote Symbiobacterium thermophilum (StSPL) degrades extracellular S1P in vitro and in blood. Moreover, we investigated its effect on cellular responses typical of fibrosis, cancer and aberrant angiogenesis using renal mesangial cells, endothelial cells, breast (MCF-7) and colon (HCT 116) carcinoma cells as disease models. In all cell types, wild-type StSPL, but not an inactive mutant, disrupted MAPK phosphorylation stimulated by exogenous S1P. Functionally, disruption of S1P receptor signaling by S1P depletion inhibited proliferation and expression of connective tissue growth factor in mesangial cells, proliferation, migration and VEGF expression in carcinoma cells, and proliferation and migration of endothelial cells. Upon intravenous injection of StSPL in mice, plasma S1P levels rapidly declined by 70% within 1 h and then recovered to normal 6 h after injection. Using the chicken chorioallantoic membrane model we further demonstrate that also under in vivo conditions StSPL, but not the inactive mutant, inhibited tumor cell-induced angiogenesis as an S1P-dependent process. Our data demonstrate that recombinant StSPL is active under extracellular conditions and holds promise as a new enzyme therapeutic for diseases associated with increased levels of S1P and S1P receptor signaling

Tubulointerstitial injury and the progression of chronic kidney disease

In chronic kidney disease (CKD), once injury from any number of disease processes reaches a threshold, there follows an apparently irreversible course toward decline in kidney function. The tubulointerstitium may play a key role in this common progression pathway. Direct injury, high metabolic demands, or stimuli from various other forms of renal dysfunction activate tubular cells. These, in turn, interact with interstitial tissue elements and inflammatory cells, causing further pathologic changes in the renal parenchyma. The tissue response to these changes thus generates a feed-forward loop of kidney injury and progressive loss of function. This article reviews the mechanisms of this negative cycle mediating CKD

Roadmap on Electronic Structure Codes in the Exascale Era

Electronic structure calculations have been instrumental in providing many important insights into a range of physical and chemical properties of various molecular and solid-state systems. Their importance to various fields, including materials science, chemical sciences, computational chemistry and device physics, is underscored by the large fraction of available public supercomputing resources devoted to these calculations. As we enter the exascale era, exciting new opportunities to increase simulation numbers, sizes, and accuracies present themselves. In order to realize these promises, the community of electronic structure software developers will however first have to tackle a number of challenges pertaining to the efficient use of new architectures that will rely heavily on massive parallelism and hardware accelerators. This roadmap provides a broad overview of the state-of-the-art in electronic structure calculations and of the various new directions being pursued by the community. It covers 14 electronic structure codes, presenting their current status, their development priorities over the next five years, and their plans towards tackling the challenges and leveraging the opportunities presented by the advent of exascale computing.Comment: Submitted as a roadmap article to Modelling and Simulation in Materials Science and Engineering; Address any correspondence to Vikram Gavini ([email protected]) and Danny Perez ([email protected]

Recombinant Expression, Purification, and Functional Characterisation of Connective Tissue Growth Factor and Nephroblastoma-Overexpressed Protein

The CCN family of proteins, especially its prominent member, the Connective tissue growth factor (CTGF/CCN2) has been identified as a possible biomarker for the diagnosis of fibrotic diseases. As a downstream mediator of TGF-β1 signalling, it is involved in tissue scarring, stimulates interstitial deposition of extracellular matrix proteins, and promotes proliferation of several cell types. Another member of this family, the Nephroblastoma-Overexpressed protein (NOV/CCN3), has growth-inhibiting properties. First reports further suggest that these two CCN family members act opposite to each other in regulating extracellular matrix protein expression and reciprocally influence their own expression when over-expressed. We have established stable HEK and Flp-In-293 clones as productive sources for recombinant human CCN2/CTGF. In addition, we generated an adenoviral vector for recombinant expression of rat NOV and established protocols to purify large quantities of these CCN proteins. The identity of purified human CCN2/CTGF and rat CCN3/NOV was proven by In-gel digest followed by ESI-TOF/MS mass spectrometry. The biological activity of purified proteins was demonstrated using a Smad3-sensitive reporter gene and BrdU proliferation assay in permanent cell line EA•hy 926 cells. We further demonstrate for the first time that both recombinant CCN proteins are N-glycosylated