P-191 Poster of distinction

Isolation, characterization and pro-angiogenic effect of microvesicles (MVs) derived from purified human pancreatic islets

F. Figliolini¹, V. Cantaluppi², M. De Lena¹, S. Beltramo¹, D. Medica¹, L. Biancone², G.P. Segoloni², G. Camussi^2
1 Research Center for Experimental Medicine (CeRMS), University of Turin, Turin, Italy; ² Nephrology, Dialysis and Renal Transplantation Unit, University of Turin, Turin, Italy

Objective: Microvesicles (MVs) are small particles that play a key role in cell-to-cell communication through the transfer of proteins and RNAs. MVs are enriched of microRNAs (miRNAs), small non coding RNAs able to modulate protein transduction. The aims of this study were: 1) isolation and characterization of MVs from purified human islets; 2) evaluation of the biological effects of MVs on islet-derived endothelial cells (IECs).

Methods: MVs were isolated by ultracentrifugation from purified human islets and characterized by electronic microscopy, FACS, western blot, bioanalyzer, RT-PCR for specific islet-associated genes and miRNAs. We evaluated on isolated IECs MV-induced; a) transfer of insulin mRNA/protein; b) proliferation (BrdU); c) resistance to apoptosis (TUNEL); d) angiogenesis on Matrigel.

Results: Islet-derived MVs sized between 0.1-1 ?m and expressed different membrane molecules (integrins, CD44, classI-II HLA). MVs also expressed insulin and C-peptide and shuttled mRNAs involved in insulin secretion and signal transduction (insulin receptor, IRS2, AKT2, PDX1) and several miRNAs, including the beta-cell specific miR-7 and miR-375 and the pro-angiogenic miR-126 and miR-296. MVs were internalized into IECs inducing insulin mRNA/protein expression, cell proliferation, resistance to apoptosis and enhanced angiogenesis. All these effects were significantly decreased by pre-treatment of MVs with 1U/ml RNase, suggesting a key role of RNA transfer from MVs to IECs. In addition, MV-stimulated IECs showed the up-regulation of pro-angiogenic genes and the activation of the P-Akt/Akt pathway.

Conclusions: Human islets release biologically active MVs able to shuttle specific proteins, mRNAs and miRNAs into target cells. MVs exert a pro-angiogenic effect on IECs, suggesting a putative role for islet-derived MVs in beta cell-endothelium cross-talk and in the neoangiogenesis processes of transplanted islets.

P-191 Poster of distinction

Isolation, characterization and pro-angiogenic effect of microvesicles (MVs) derived from purified human pancreatic islets

F. Figliolini¹, V. Cantaluppi², M. De Lena¹, S. Beltramo¹, D. Medica¹, L. Biancone², G.P. Segoloni², G. Camussi^2
1 Research Center for Experimental Medicine (CeRMS), University of Turin, Turin, Italy; ² Nephrology, Dialysis and Renal Transplantation Unit, University of Turin, Turin, Italy

Objective: Microvesicles (MVs) are small particles that play a key role in cell-to-cell communication through the transfer of proteins and RNAs. MVs are enriched of microRNAs (miRNAs), small non coding RNAs able to modulate protein transduction. The aims of this study were: 1) isolation and characterization of MVs from purified human islets; 2) evaluation of the biological effects of MVs on islet-derived endothelial cells (IECs).

Methods: MVs were isolated by ultracentrifugation from purified human islets and characterized by electronic microscopy, FACS, western blot, bioanalyzer, RT-PCR for specific islet-associated genes and miRNAs. We evaluated on isolated IECs MV-induced; a) transfer of insulin mRNA/protein; b) proliferation (BrdU); c) resistance to apoptosis (TUNEL); d) angiogenesis on Matrigel.

Results: Islet-derived MVs sized between 0.1-1 ?m and expressed different membrane molecules (integrins, CD44, classI-II HLA). MVs also expressed insulin and C-peptide and shuttled mRNAs involved in insulin secretion and signal transduction (insulin receptor, IRS2, AKT2, PDX1) and several miRNAs, including the beta-cell specific miR-7 and miR-375 and the pro-angiogenic miR-126 and miR-296. MVs were internalized into IECs inducing insulin mRNA/protein expression, cell proliferation, resistance to apoptosis and enhanced angiogenesis. All these effects were significantly decreased by pre-treatment of MVs with 1U/ml RNase, suggesting a key role of RNA transfer from MVs to IECs. In addition, MV-stimulated IECs showed the up-regulation of pro-angiogenic genes and the activation of the P-Akt/Akt pathway.

Conclusions: Human islets release biologically active MVs able to shuttle specific proteins, mRNAs and miRNAs into target cells. MVs exert a pro-angiogenic effect on IECs, suggesting a putative role for islet-derived MVs in beta cell-endothelium cross-talk and in the neoangiogenesis processes of transplanted islets.