Matrix metalloproteinase 9 Contributes to Glomerulosclerosis by Causing Profibrotic Changes in Podocytes and Glomerular Endothelial Cells

Background: Glomerulosclerosis is characterized by progressive (myo) fibroblast accumulation and collagen deposition involving profibrotic changes of podocytes and endothelial cells. A profibrotic role of MMP-9 in interstitial fibrosis has been reported. Whether MMP-9 plays a role in glomerulosclerosis is not clear yet. Methods: Mouse glomerulosclerosis model [Adriamycin Nephropathy (AN) model] was induced by a single adriamycin injection (10.2mg/kg, with physiological saline for controls) through tail vein in MMP-9-/- and wild-type control mice of BALB/c background. All animals were sacrificed at 4 weeks after injection. Albuminuria (albumin to creatinine ratio) and calculated GFR were measured. Gomori Trichrome (GT) and Sirius Red (SR) staining were used for assessment of glomerular fibrosis. Profibrotic changes of podocytes or glomerular endothelial cells were examined by confocal microscopy using immunofluorescence staining (IF) of desmin or a-SMA with P-cadherin or VEcadherin. Results: Albuminuria was reduced while GFR was increased in MMP-9-/- AN mice compared with those of wild-type mice. Confocal microscopy showed a significant decrease in podocytes double-stained with P-cadherin and desmin, demonstrating that MMP9-/- AN mice were protected from profibrotic changes in podocytes and glomerular endothelial cells. Glomerulosclerosis was significantly reduced in MMP9-/- AN mice compared to that of WT, as demonstrated by GT and SR staining. Conclusions: MMP-9 contributes to glomerulosclerosis at least in part by causing profibrotic changes in podocytes and glomerular endothelial cells.

Synaptopodin deficiency exacerbates kidney disease in a mouse model of Alport Syndrome

Synaptopodin (Synpo) is an actin-associated protein in podocyte foot processes. By generating mice that completely lack Synpo, we previously showed that Synpo is dispensable for normal kidney function. However, the lack of Synpo worsened Adriamycin nephropathy, indicating a protective role for Synpo in injured podocytes. Here we investigated whether the lack of Synpo directly impacts a genetic disease, Alport syndrome (AS), because Synpo is reduced in the podocytes of affected humans and mice; whether this is merely an association or pathogenic is unknown. We used Col4a5 mutant mice that model X-linked AS, showing glomerular basement membrane (GBM) abnormalities, eventual foot process effacement, and progression to ESKD. We intercrossed mice carrying mutations in Synpo and Col4a5 to produce doubly mutant mice. Urine and tissue were taken at select time points to evaluate albuminuria, histopathology, and glomerular capillary wall composition and ultrastructure. The lack of Synpo in Col4a5-/Y, Col4a5-/-, or Col4a5+/- Alport mice led to acceleration of disease progression, including more severe proteinuria and glomerulosclerosis. The absence of Synpo attenuated the shift of myosin IIA from the podocyte cell body and major processes to the actin cables near the GBM in the areas of effacement. We speculate that this is mechanistically associated with enhanced loss of podocytes due to easier detachment from the GBM. We conclude that Synpo deletion exacerbates the disease phenotype in Alport mice, revealing the podocyte actin cytoskeleton as a target for therapy in patients with AS.

Artemether ameliorates kidney injury by restoring redox imbalance and improving mitochondrial function in Adriamycin nephropathy in mice

The kidney is a high-energy demand organ rich in mitochondria especially renal tubular cells. Emerging evidence suggests that mitochondrial dysfunction, redox imbalance and kidney injury are interconnected. Artemether has biological effects by targeting mitochondria and exhibits potential therapeutic value for kidney disease. However, the underlying molecular mechanisms have not been fully elucidated. This study was performed to determine the effects of artemether on Adriamycin-induced nephropathy and the potential mechanisms were also investigated. In vivo, an Adriamycin nephropathy mouse model was established, and mice were treated with or without artemether for 2 weeks. In vitro, NRK-52E cells were stimulated with TGF-β1 and treated with or without artemether for 24 h. Then renal damage and cell changes were evaluated. The results demonstrated that artemether reduced urinary protein excretion, recovered podocyte alterations, attenuated pathological changes and alleviated renal tubular injury. Artemether also downregulated TGF-β1 mRNA expression levels, inhibited tubular proliferation, restored tubular cell phenotypes and suppressed proliferation-related signalling pathways. In addition, artemether restored renal redox imbalance, increased mtDNA copy number and improved mitochondrial function. In summary, we provided initial evidence that artemether ameliorates kidney injury by restoring redox imbalance and improving mitochondrial function in Adriamycin nephropathy in mice. Artemether may be a promising agent for the treatment kidney disease.