Abstract 481: Exosomes as a Potential Acellular Signaling Mechanism in the Kidney
Exosomes are small 50-90 nm vesicles containing mRNA, proteins, and other renal epithelial sub-cellular components and might serve as an intra-nephron signaling mechanism. Exosomes were isolated from cultured immortalized human renal proximal tubule cells (RPTC) by ultracentrifugation following agonist treatment. 24 hour stimulation with the dopaminergic agonist fenoldopam (1 μM) produced a 2.38±0.053 fold increase in exosome release (RFU Vehicle = 0.038±0.0072; n=15, p<0.04). Angiotensin II stimulation (1 nM, 24 hours) caused a 4.71±0.35 fold increase in exosome release (n=18, p<0.04). Agonist-induced responses persisted after removal of extracellular calcium, indicating that renal exosome release is not dependent upon a calcium influx into cells. We next showed that exosomes are able to transfer between cells by electroporating Lamp1-RFP or CD82-YFP (two proteins previously found to be contained in exosomes) into RPTC. We found that both GFP fusion proteins were secreted into the culture medium and taken up by non-transfected cells. We further hypothesized that sodium regulatory proteins would be secreted into exosomes. Proteomic analysis demonstrated the presence of the dopamine-1 receptor and the dopamine-1 receptor selective kinase GRK4, which are known to affect natriuresis, in exosomes derived from RPTC. Furthermore, GRK4 mRNA was detected in exosomes by RT-PCR. In conclusion, exosomes contain proteins and mRNA which could serve as an intra-nephron acellular signaling mechanism that may alter sodium homeostasis.
- © 2012 by American Heart Association, Inc.