ABSTRACT
Type 2 diabetes mellitus (DM2) is a widespread metabolic disorder that results in podocyte damage and diabetic nephropathy. Previous studies demonstrated that TRPC6 channels play a pivotal role in podocyte function and their dysregulation is associated with development of different kidney diseases including nephropathy. Here, using single channel patch clamp technique, we demonstrated that non-selective cationic TRPC6 channels are sensitive to the Ca2+ store depletion in human podocyte cell line Ab8/13 and in freshly isolated rat glomerular podocytes. Ca2+ imaging indicated the involvement of ORAI and sodium-calcium exchanger in Ca2+ entry induced upon store depletion. In male rats fed a high-fat diet combined with a low-dose streptozotocin injection, which leads to DM2 development, we observed the reduction of a store-operated Ca2+ entry (SOCE) in rat glomerular podocytes. This was accompanied by a reorganization of store-operated Ca2+ influx such that TRPC6 channels lost their sensitivity to Ca2+ store depletion and ORAI-mediated Ca2+ entry was suppressed in TRPC6-independent manner. Altogether our data provide new insights into the mechanism of SOCE organization in podocytes in the norm and in pathology, which should be taken into account when developing pharmacological treatment of the early stages of diabetic nephropathy.
Subject(s)
Diabetes Mellitus, Experimental , Diabetes Mellitus, Type 2 , Diabetic Nephropathies , Podocytes , Humans , Rats , Male , Animals , TRPC6 Cation Channel/metabolism , Podocytes/metabolism , Calcium Channels/metabolism , Diabetic Nephropathies/metabolism , Calcium/metabolism , Diabetes Mellitus, Type 2/metabolism , Diabetes Mellitus, Experimental/metabolism , TRPC Cation Channels/metabolismABSTRACT
BACKGROUND/AIMS: Mutations of desmosomal genes are known to cause arrhythmogenic cardiomyopathy characterized by arrhythmias and sudden cardiac death. Previously, we described a novel genetic variant H1684R in desmoplakin gene (DSP), associated with a progressive cardiac conduction disease (PCCD). In the present study, we aimed to investigate an effect of the DSP-H1684R genetic variant on the activity of ion channels. METHODS: We used cardiomyocytes derived from induced pluripotent stem cells (iPSC cardiomyocytes) from a patient with DSP-H1684R genetic variant and from two healthy donors. Immunofluorescent staining and western blot analyses were used to characterize patient-specific cardiomyocytes. By the whole-cell voltage-clamp technique we estimated the activity of voltage-gated sodium, calcium, and potassium channels that are responsible for action potential generation and its shape. Action potentials' parameters were measured using whole-cell current-clamp technique. RESULTS: In patient-specific cardiomyocytes we observed both lower amplitudes of currents through sodium Nav1.5 channels and L-type calcium channels, but higher amplitude of current through transient-outward potassium channels in comparison to donor cardiomyocytes. Current-clamp measurements revealed shortening of action-potential in DSP-H1684R-carrying iPSC cardiomyocytes. Therefore, observed alterations in the channels activity might have a great impact on the properties of action potential and development of PCCD. CONCLUSION: Our results show that desmoplakin genetic variants, besides conduction slowing caused by structural heart remodeling, could affect multiple ion channel activity aggravating arrhythmia manifestation in PCCD.
