A Podocyte view on RhoGTPases and actin cytoskeleton regulation

Proteinuria is a hallmark symptom of chronic kidney disease, that if left to persist constitutes a risk for progression of disease. Symptomatic treatment aiming at decreasing proteinuria is therefore standard practice. Curative treatments for the underlying cause of disease are however lacking and treatments currently in use to induce disease remission are associated with unfavorable side effects. Dysregulation of the podocyte actin cytoskeleton underlies the pathological process called foot process effacement (FPE), which is one of the leading causes of proteinuria. The studies included in this thesis have focused on podocyte actin cytoskeleton regulation and a group of proteins called RhoGTPases, known to be involved in actin cytoskeleton regulation in podocytes. In the first study, glomerular microarray analysis showed an increase in the expression of the melanocortin 1-receptor (MC1R) in renal diseases focal segmental glomerulosclerosis and membranous nephropathy. Subsequent mass spectrometry analysis in combination with pathway and biochemical analysis revealed the podocyte protective effects of MC1R stimulation in vitro. Activation of MC1R proved to be stabilizing the podocyte actin cytoskeleton through inhibition of the epidermal growth factor receptor (EGFR) and maintenance of the actin associated protein synaptopodin. In the second study, the depletion of the prenylation enzyme Geranylgeranyl transferase type I (GGTase-I) in podocytes led to the development of proteinuria and FPE in mice due to an imbalanced RhoGTPase activity and disruption of the actin cytoskeleton. These findings suggest that GGTase-I activity is essential for podocyte function. In the last study, a guanine nucleotide exchange factor (activator of RhoGTPases) named Bpix was identified to be modulated in podocytes following treatment with a renal stressor, using mass spectrometry analysis. Gene silencing of Bpix protected against actin cytoskeleton remodulation in a model of podocyte injury, demonstrating the importance of Bpix for podocyte actin cytoskeleton regulation. In conclusion, the results in this thesis confirm the importance of actin cytoskeleton regulation for podocyte integrity. Further on, the results provide new information on actin cytoskeleton regulatory pathways involving RhoGTPases in podocytes, which can be of importance for future attempts in finding targeted treatments of proteinuria and chronic kidney disease

Orellanine specifically targets renal clear cell carcinoma

Renal cell carcinoma (RCC), arising from the proximal tubule in the kidney, accounts for approximately 85% of kidney cancers and causes over 140,000 annual deaths worldwide. In the last decade, several new therapies have been identified for treatment of metastatic RCC. Although these therapies increase survival time compared to standard care, none of them has curative properties. The nephrotoxin orellanine specifically targets proximal tubular epithelial cells, leaving other organs unaffected. We therefore hypothesized that the selective toxicity of orellanine extends to clear cell RCC (ccRCC) cells since they emanate from proximal tubular cells. Orellanine would thus target both primary and metastatic ccRCC in vitro and in vivo. We found that orellanine induces dose-dependent cell death in proximal tubular cells and in all ccRCC cells tested, both primary and cell lines, with no toxicity detected in control cells. The toxic action of orellanine involve decreased protein synthesis, disrupted cell metabolism and induction of apoptosis. In nude rats carrying human ccRCC xenografts, brief orellanine treatment eliminated more than 90% of viable tumor mass compared to control rats. This identifies orellanine as a potential treatment concept for ccRCC patients on dialysis, due to its unique selective toxicity towards ccRCC