Podocytes play a pivotal role in maintaining glomerular filtration function through their interdigitated foot processes. However, the mechanisms that govern the podocyte cytoskeletal rearrangement remain unclear. Through analyzing the transcriptional profile of renal biopsy specimens from patients with diabetic nephropathy (DN) and control donors, we identify SLIT-ROBO ρGTPase-activating protein 2a (SRGAP2a) as one of the main hub genes strongly associated with proteinuria and glomerular filtration in type 2 DN. Immunofluorescence staining and Western blot analysis revealed that human and mouse SRGAP2a is primarily localized at podocytes and largely colocalized with synaptopodin. Moreover, podocyte SRGAP2a is downregulated in patients with DN and db/db mice at both the mRNA and the protein level. SRGAP2a reduction is observed in cultured podocytes treated with tumor growth factor-β or high concentrations of glucose. Functional and mechanistic studies show that SRGAP2a suppresses podocyte motility through inactivating RhoA/Cdc42 but not Rac1. The protective role of SRGAP2a in podocyte function also is confirmed in zebrafish, in which knockdown of SRGAP2a, a SRGAP2 ortholog in zebrafish, recapitulates podocyte foot process effacement. Finally, increasing podocyte SRGAP2a levels in db/db mice through administration of adenovirus-expressing SRGAP2a significantly mitigates podocyte injury and proteinuria. The results demonstrate that SRGAP2a protects podocytes by suppressing podocyte migration.

Although the use of bortezomib alone and in combination with steroids has shown efficacy in AL amyloidosis, its role in combination with high-dose melphalan and autologous stem cell transplantation (HDM/SCT) is unknown. In this study, we evaluated bortezomib in combination with dexamethasone (BD) for induction chemotherapy prior to HDM/SCT. This was a single-center, prospective, randomized controlled trial comparing induction therapy consisting of two BD cycles followed by HDM/SCT (BD + HDM/SCT) with HDM/SCT alone in the treatment of patients with newly diagnosed AL amyloidosis. The hematological and organ responses of the patients were assessed every three months post HDM/SCT. Fifty-six patients newly diagnosed with renal (100%), cardiac (57.1%), liver (7.1%), or nervous system (8.9%) AL amyloidosis were enrolled in this study; 28 patients were assigned to each arm. Two patients died within 100 days of HDM/SCT (3.6% treatment-related mortality). The overall hematologic response rates in the BD + HDM/SCT arm and HDM/SCT arm at three, six and twelve months were 78.5% versus 50%, 82.1% versus 53.5% and 85.7% versus 53.5%, respectively. In the BD + HDM/SCT arm, 15 (53.5%) patients achieved a hematologic response after BD and before HDM/SCT. An intention-to-treat analysis revealed a higher rate of complete remission in the BD + HDM/SCT arm at both 12 and 24 months (67.9% and 70%, respectively) than with the HDM/SCT-only therapy (35.7% and 35%, respectively, P = 0.03). After a median follow-up of 28 months, the survival rates at 24 months post-treatment start were 95.0% in the BD + HDM/SCT group and 69.4% in the HDM/SCT alone group (P = 0.03). Our preliminary data suggest that the outcome of treating AL amyloidosis with BD induction and HDM/SCT was superior to the outcome of the HDM/SCT treatment alone. This trial has been registered at clinicaltrials.gov with the number NCT01998503 .

GADD45 gene has been implicated in cell cycle arrest, cell survival or apoptosis in a cell type specific and context-dependent manner. Members of GADD45 gene family have been found differentially expressed in several podocyte injury models, but their roles in podocytes are unclear. Using an in vivo zebrafish model of inducible podocyte injury that we have previously established, we found that zebrafish orthologs of gadd45b were induced upon the induction of podocyte injury. Podocyte-specific overexpression of zebrafish gadd45b exacerbated edema, proteinuria and foot-process effacement, whereas knockdown of gadd45b by morpholino-oligos in zebrafish larvae ameliorated podocyte injury. We then explored the role of GADD45B induction in podocyte injury using in vitro podocyte culture. We confirmed that GADD45B was significantly upregulated during the early phase of podocyte injury in cultured human podocytes and that podocyte apoptosis induced by TGF-β and puromycin aminonucleoside (PAN) was aggravated by GADD45B overexpression but ameliorated by shRNA-mediated GADD45B knockdown. We also showed that ROS inhibitor NAC suppressed PAN-induced GADD45B expression and subsequent activation of p38 MAPK pathway in podocytes and that inhibition of GADD45B diminished PAN-induced p38 MAPK activation. Taken together, our findings demonstrated that GADD45B has an important role in podocyte injury and may be a therapeutic target for the management of podocyte injury in glomerular diseases.

Nephrotic syndrome is a CKD defined by proteinuria with subsequent hypoalbuminemia, hyperlipidemia, and edema caused by impaired renal glomerular filtration barrier function. We previously identified mutations in epithelial membrane protein 2 (EMP2) as a monogenic cause of this disease. Here, we generated an emp2-knockout zebrafish model using transcription activator-like effector nuclease–based genome editing. We found that loss of emp2 in zebrafish upregulated caveolin-1 (cav1), a major component of caveolae, in embryos and adult mesonephric glomeruli and exacerbated podocyte injury. This phenotype was partially rescued by glucocorticoids. Furthermore, overexpression of cav1 in zebrafish podocytes was sufficient to induce the same phenotype observed in emp2 homozygous mutants, which was also treatable with glucocorticoids. Similarly, knockdown of EMP2 in cultured human podocytes resulted in increased CAV1 expression and decreased podocyte survival in the presence of puromycin aminonucleoside, whereas glucocorticoid treatment ameliorated this phenotype. Taken together, we have established excessive CAV1 as a mediator of the predisposition to podocyte injury because of loss of EMP2, suggesting CAV1 could be a novel therapeutic target in nephrotic syndrome and podocyte injury.

Podocyte injury is a primary contributor to proteinuria. Triptolide is a major active component of Tripterygium wilfordii Hook F that exhibits potent antiproteinuric effects. We used our previously developed in vivo zebrafish model of inducible podocyte-target injury and found that triptolide treatment effectively alleviated oedema, proteinuria and foot process effacement. Triptolide also inhibited podocyte apoptosis in our zebrafish model and in vitro. We also examined the mechanism of triptolide protection of podocyte. Whole-genome expression profiles of cultured podocytes demonstrated that triptolide treatment downregulated apoptosis pathway-related GADD45B expression. Specific overexpression of gadd45b in zebrafish podocytes abolished the protective effects of triptolide. GADD45B is a mediator of podocyte apoptosis that contains typical NF-κB binding sites in the promoter region, and NF-κB p65 primarily transactivates this gene. Triptolide inhibited NF-κB signalling activation and binding of NF-κB to the GADD45B promoter. Taken together, our findings demonstrated that triptolide attenuated proteinuria and podocyte apoptosis via inhibition of NF-κB/GADD45B signalling, which provides a new understanding of the antiproteinuric effects of triptolide in glomerular diseases.