DNM2 levels normalization improves muscle phenotypes of a novel mouse model for moderate centronuclear myopathy.

Dynamin 2 (DNM2) is a ubiquitously expressed GTPase regulating membrane trafficking and cytoskeleton dynamics. Heterozygous dominant mutations in DNM2 cause centronuclear myopathy (CNM), associated with muscle weakness and atrophy and histopathological hallmarks as fiber hypotrophy and organelles mis-position. Different severities range from the severe neonatal onset form to the moderate form with childhood onset and to the mild adult onset form. No therapy is approved for CNM. Here we aimed to validate and rescue a mouse model for the moderate form of DNM2-CNM harboring the common DNM2 R369W missense mutation. Dnm2R369W/+ mice presented with increased DNM2 protein level in muscle and moderate CNM-like phenotypes with force deficit, muscle and fiber hypotrophy, impaired mTOR signaling, and progressive mitochondria and nuclei mis-position with age. Molecular analyses revealed a fiber type switch toward oxidative metabolism correlating with decreased force and alteration of mitophagy markers paralleling mitochondria structural defects. Normalization of DNM2 levels through intramuscular injection of AAV-shDnm2 targeting Dnm2 mRNA significantly improved histopathology and muscle and myofiber hypotrophy. These results showed that the Dnm2R369W/+ mouse is a faithful model for the moderate form of DNM2-CNM and revealed that DNM2 normalization after a short 4-week treatment is sufficient to improve the CNM phenotypes.

Growth Differentiation Factor 15 Ameliorates Anti-Glomerular Basement Membrane Glomerulonephritis in Mice.

Growth differentiation factor 15 (GDF15) is a member of the transforming growth factor-ß (TGF-ß) cytokine family and an inflammation-associated protein. Here, we investigated the role of GDF15 in murine anti-glomerular basement membrane (GBM) glomerulonephritis. Glomerulonephritis induction in mice induced systemic expression of GDF15. Moreover, we demonstrate the protective effects for GDF15, as GDF15-deficient mice exhibited increased proteinuria with an aggravated crescent formation and mesangial expansion in anti-GBM nephritis. Herein, GDF15 was required for the regulation of T-cell chemotactic chemokines in the kidney. In addition, we found the upregulation of the CXCR3 receptor in activated T-cells in GDF15-deficient mice. These data indicate that CXCL10/CXCR3-dependent-signaling promotes the infiltration of T cells into the organ during acute inflammation controlled by GDF15. Together, these results reveal a novel mechanism limiting the migration of lymphocytes to the site of inflammation during glomerulonephritis.

Glucagon-Like Peptide-1 Receptor Agonism Improves Nephrotoxic Serum Nephritis by Inhibiting T-Cell Proliferation.

Glucagon-like peptide (GLP)-1 analogs such as liraglutide improved albuminuria in patients with type 2 diabetes in large randomized controlled trials. One of the suspected mechanisms is the anti-inflammatory potential of GLP-1 receptor (Glp1r) agonism. Thus, the anti-inflammatory action of Glp1r agonism was tested in a nondiabetic, T-cell-mediated murine model of nephrotoxic serum nephritis (NTS). The role of Glp1r in NTS was evaluated by using Glp1r-/- mice or C57BL/6 mice treated with liraglutide. In vitro, murine T cells were stimulated in the presence of liraglutide or vehicle. Glp1r-/- mice displayed increased renal infiltration of neutrophils and T cells after induction of NTS. Splenocyte proliferation and TH1 cytokine transcription were increased in spleen and lymph nodes of Glp1r-/- mice. Liraglutide treatment significantly improved the renal outcome of NTS in C57BL/6 mice by decreasing renal infiltration and proliferation of T cells, which resulted in decreased macrophage infiltration. In vitro, T cells stimulated in the presence of liraglutide showed decreased proliferation of TH1 and TH17 cells. Liraglutide blocked glycolysis in T cells and decreased their Glut1 mRNA expression. Together, Glp1r agonism protects mice from a T-cell-dependent glomerulonephritis model by inhibition of T-cell proliferation, possibly by interacting with their metabolic program. This mechanism may explain in part the renoprotective effects of Glp1r agonism in diabetic nephropathy.

IFN Regulatory Factor 4 Controls Post-ischemic Inflammation and Prevents Chronic Kidney Disease.

Ischemia reperfusion injury (IRI) of the kidney results in interferon regulatory factor 4 (IRF4)-mediated counter-regulation of the acute inflammatory response. Beyond that, IRF4 exerts important functions in controlling the cytokine milieu, T-cell differentiation, and macrophage polarization. The latter has been implicated in tissue remodeling. It therefore remains elusive what the role of IRF4 is in terms of long-term outcome following IRI. We hypothesized that an inability to resolve chronic inflammation in Irf4-/- mice would promote chronic kidney disease (CKD) progression. To evaluate the effects of IRF4 in chronic upon acute injury in vivo, a mouse model of chronic injury following acute IRI was employed. The expression of Irf4 increased within 10 days after IRI in renal tissue. Both mRNA and protein levels remained high up to 5 weeks upon IRI, suggesting a regulatory function in the chronic phase. Mice deficient in IRF4 display increased tubular cell loss and defective clearance of infiltrating macrophages. These phenomena were associated with increased expression of pro-inflammatory macrophage markers together with reduced expression of alternatively activated macrophage markers. In addition, IRF4-deficient mice showed defective development of alternatively activated macrophages. Hints of a residual M1 macrophage signature were further observed in human biopsy specimens of patients with hypertensive nephropathy vs. living donor specimens. Thus, IRF4 restricts CKD progression and kidney fibrosis following IRI, potentially by enabling M2 macrophage polarization and restricting a Th1 cytokine response. Deteriorated alternative macrophage subpopulations in Irf4-/- mice provoke chronic intrarenal inflammation, tubular epithelial cell loss, and renal fibrosis in the long course after IRI in mice. The clinical significance of these finding for human CKD remains uncertain at present and warrants further studies.

MicroRNA-142-3p improves vascular relaxation in uremia.

BACKGROUND AND AIMS: Chronic kidney disease (CKD) is strongly associated with a high burden of cardiovascular morbidity and mortality. Therefore, we aimed to characterize the putative role of microRNAs (miR)s in uremic vascular remodelling and endothelial dysfunction. METHODS: We investigated the expression pattern of miRs in two independent end-stage renal disease (ESRD) cohorts and in the animal model of uremic DBA/2 mice via quantitative RT-PCR. Moreover, DBA/2 mice were treated with intravenous injections of synthetic miR-142-3p mimic and were analysed for functional and morphological vascular changes by mass spectrometry and wire myography. RESULTS: The expression pattern of miRs was regulated in ESRD patients and was reversible after kidney transplantation. Out of tested miRs, only blood miR-142-3p was negatively associated with carotid-femoral pulse-wave velocity in CKD 5D patients. We validated these findings in a murine uremic model and found similar suppression of miR-142-3p as well as decreased acetylcholine-mediated vascular relaxation of the aorta. Therefore, we designed experiments to restore bioavailability of aortic miR-142-3p in vivo via intravenous injection of synthetic miR-142-3p mimic. This intervention restored acetylcholine-mediated vascular relaxation. CONCLUSIONS: Taken together, we provide compelling evidence, both in humans and in mice, that miR-142-3p constitutes a potential pharmacological agent to prevent endothelial dysfunction and increased arterial stiffness in ESRD.

Innate and adaptive immunity in experimental glomerulonephritis: a pathfinder tale.

The role of innate and adaptive immune cells in the experimental model of nephrotoxic serum nephritis (NTS) has been rigorously studied in recent years. The model is dependent on kidney-infiltrating T helper (TH) 17 and TH1 cells, which recruit neutrophils and macrophages, respectively, and cause sustained kidney inflammation. In a later phase of disease, regulatory T cells (Tregs) infiltrate the kidney in an attempt to limit disease activity. In the early stage of NTS, lymph node drainage plays an important role in disease initiation since dendritic cells present the antigen to T cells in the T cell zones of the draining lymph nodes. This results in the differentiation and proliferation of TH17 and TH1 cells. In this setting, immune regulatory cells (Tregs), namely, CCR7-expressing Tregs and mast cells (MCs), which are recruited by Tregs via the production of interleukin-9, exert their immunosuppressive capacity. Together, these two cell populations inhibit T cell differentiation and proliferation, thereby limiting disease activity by as yet unknown mechanisms. In contrast, the spleen plays no role in immune activation in NTS, but constitutes a place of extramedullary haematopoiesis. The complex interactions of immune cells in NTS are still under investigation and might ultimately lead to targeted therapies in glomerulonephritis.

A New Murine Model of Chronic Kidney Disease-Mineral and Bone Disorder.

Chronic kidney disease (CKD) is associated with mineral and bone disorder (MBD), which is the main cause of the extensively increased cardiovascular mortality in the CKD population. We now aimed to establish a new murine experimental CKD-MBD model. Dilute brown non-Agouti (DBA/2) mice were fed with high-phosphate diet for 4 (HPD4) or 7 (HPD7) days, then with standard chow diet (SCD) and subsequently followed until day 84. They were compared to DBA/2 mice maintained on SCD during the whole study period. Both 4 and 7 days HPD-fed mice developed phosphate nephropathy with tubular atrophy, interstitial fibrosis, decreased glomerular filtration rate, and increased serum urea levels. The abdominal aorta of HPD-treated mice showed signs of media calcification. Histomorphometric analysis of HPD-treated mice showed decreased bone volume/tissue volume, low mineral apposition rate, and low bone formation rate as compared to SCD-fed mice, despite increased parathyroid hormone levels. Overall, the observed phenotype was more pronounced in the HPD7 group. In summary, we established a new, noninvasive, and therefore easy to perform reproducible CKD-MBD model, which showed media calcification, secondary hyperparathyroidism, and low-turnover bone disease.