Finding the Needle in the Haystack: Serological and Urinary Biomarkers in Behçet's Disease: A Systematic Review.

Urinary and serological markers play an essential role in the diagnostic process of autoimmune diseases. However, to date, specific and reliable biomarkers for diagnosing Behçet's disease (BD) are still lacking, negatively affecting the management of these patients. To analyze the currently available literature on serological and urinary BD biomarkers investigated in the last 25 years, we performed a systematic literature review using the Population, Intervention, Comparison, and Outcomes (PICO) strategy. One hundred eleven studies met the eligibility criteria (6301 BD patients, 5163 controls). Most of them were retrospective, while five (5%) were prospective. One hundred ten studies (99%) investigated serological biomarkers and only two (2%) focused on urinary biomarkers. One hundred three studies (93%) explored the diagnostic potential of the biomolecules, whereas sixty-two (56%) tested their effect on disease activity monitoring. Most articles reported an increase in inflammatory markers and pro-oxidant molecules, with a decrease in antioxidants. Promising results have been shown by the omics sciences, offering a more holistic approach. Despite the vast number of investigated markers, existing evidence indicates a persistent gap in BD diagnostic/prognostic indices. While new steps have been taken in the direction of pathogenesis and disease monitoring, international efforts for the search of a diagnostic marker for BD are still needed.

Extracellular Vesicles Derived from Endothelial Progenitor Cells Protect Human Glomerular Endothelial Cells and Podocytes from Complement- and Cytokine-Mediated Injury.

Glomerulonephritis are renal inflammatory processes characterized by increased permeability of the Glomerular Filtration Barrier (GFB) with consequent hematuria and proteinuria. Glomerular endothelial cells (GEC) and podocytes are part of the GFB and contribute to the maintenance of its structural and functional integrity through the release of paracrine mediators. Activation of the complement cascade and pro-inflammatory cytokines (CK) such as Tumor Necrosis Factor α (TNF-α) and Interleukin-6 (IL-6) can alter GFB function, causing acute glomerular injury and progression toward chronic kidney disease. Endothelial Progenitor Cells (EPC) are bone-marrow-derived hematopoietic stem cells circulating in peripheral blood and able to induce angiogenesis and to repair injured endothelium by releasing paracrine mediators including Extracellular Vesicles (EVs), microparticles involved in intercellular communication by transferring proteins, lipids, and genetic material (mRNA, microRNA, lncRNA) to target cells. We have previously demonstrated that EPC-derived EVs activate an angiogenic program in quiescent endothelial cells and renoprotection in different experimental models. The aim of the present study was to evaluate in vitro the protective effect of EPC-derived EVs on GECs and podocytes cultured in detrimental conditions with CKs (TNF-α/IL-6) and the complement protein C5a. EVs were internalized in both GECs and podocytes mainly through a L-selectin-based mechanism. In GECs, EVs enhanced the formation of capillary-like structures and cell migration by modulating gene expression and inducing the release of growth factors such as VEGF-A and HGF. In the presence of CKs, and C5a, EPC-derived EVs protected GECs from apoptosis by decreasing oxidative stress and prevented leukocyte adhesion by inhibiting the expression of adhesion molecules (ICAM-1, VCAM-1, E-selectin). On podocytes, EVs inhibited apoptosis and prevented nephrin shedding induced by CKs and C5a. In a co-culture model of GECs/podocytes that mimicked GFB, EPC-derived EVs protected cell function and permeselectivity from inflammatory-mediated damage. Moreover, RNase pre-treatment of EVs abrogated their protective effects, suggesting the crucial role of RNA transfer from EVs to damaged glomerular cells. In conclusion, EPC-derived EVs preserved GFB integrity from complement- and cytokine-induced damage, suggesting their potential role as therapeutic agents for drug-resistant glomerulonephritis.

PMMA-Based Continuous Hemofiltration Modulated Complement Activation and Renal Dysfunction in LPS-Induced Acute Kidney Injury.

Sepsis-induced acute kidney injury (AKI) is a frequent complication in critically ill patients, refractory to conventional treatments. Aberrant activation of innate immune system may affect organ damage with poor prognosis for septic patients. Here, we investigated the efficacy of polymethyl methacrylate membrane (PMMA)-based continuous hemofiltration (CVVH) in modulating systemic and tissue immune activation in a swine model of LPS-induced AKI. After 3 h from LPS infusion, animals underwent to PMMA-CVVH or polysulfone (PS)-CVVH. Renal deposition of terminal complement mediator C5b-9 and of Pentraxin-3 (PTX3) deposits were evaluated on biopsies whereas systemic Complement activation was assessed by ELISA assay. Gene expression profile was performed from isolated peripheral blood mononuclear cells (PBMC) by microarrays and the results validated by Real-time PCR. Endotoxemic pigs presented oliguric AKI with increased tubulo-interstitial infiltrate, extensive collagen deposition, and glomerular thrombi; local PTX-3 and C5b-9 renal deposits and increased serum activation of classical and alternative Complement pathways were found in endotoxemic animals. PMMA-CVVH treatment significantly reduced tissue and systemic Complement activation limiting renal damage and fibrosis. By microarray analysis, we identified 711 and 913 differentially expressed genes with a fold change >2 and a false discovery rate <0.05 in endotoxemic pigs and PMMA-CVVH treated-animals, respectively. The most modulated genes were Granzyme B, Complement Factor B, Complement Component 4 Binding Protein Alpha, IL-12, and SERPINB-1 that were closely related to sepsis-induced immunological process. Our data suggest that PMMA-based CVVH can efficiently modulate immunological dysfunction in LPS-induced AKI.

Targeted and untargeted quantification of quorum sensing signalling molecules in bacterial cultures and biological samples via HPLC-TQ MS techniques.

Quorum sensing (QS) is the ability of some bacteria to detect and to respond to population density through signalling molecules. QS molecules are involved in motility and cell aggregation mechanisms in diseases such as sepsis. Few biomarkers are currently available to diagnose sepsis, especially in high-risk conditions. The aim of this study was the development of new analytical methods based on liquid chromatography-mass spectrometry for the detection and quantification of QS signalling molecules, including N-acyl homoserine lactones (AHL) and hydroxyquinolones (HQ), in biofluids. Biological samples used in the study were Pseudomonas aeruginosa bacterial cultures and plasma from patients with sepsis. We developed two MS analytical methods, based on neutral loss (NL) and product ion (PI) experiments, to identify and characterize unknown AHL and HQ molecules. We then established a multiple-reaction-monitoring (MRM) method to quantify specific QS compounds. We validated the HPLC-MS-based approaches (MRM-NL-PI), and data were in accord with the validation guidelines. With the NL and PI MS-based methods, we identified and characterized 3 and 13 unknown AHL and HQ compounds, respectively, in biological samples. One of the newly found AHL molecules was C12-AHL, first quantified in Pseudomonas aeruginosa bacterial cultures. The MRM quantitation of analytes in plasma from patients with sepsis confirmed the analytical ability of MRM for the quantification of virulence factors during sepsis. Graphical abstract.

Regenerative Role of Stem Cell-Derived Extracellular Vesicles in Acute Kidney Injury.

Acute kidney injury (AKI) is a frequent complication of hospital admission and worsens short- and long-term patients' prognosis. Currently, AKI treatment remains supportive and no therapy has proven significant benefit in clinical trials. Stem cells (SCs) are a promising therapeutic option, but their translation to the clinical setting is limited by the risk of rejection or aberrant differentiation. Numerous studies have shown how SC effects are mediated by paracrine factors such as extracellular vesicles (EVs). In this review, we describe the preclinical evidence about EV efficacy in acute tubular and glomerular injury and the recently generated clinical data.

Citrate anion improves chronic dialysis efficacy, reduces systemic inflammation and prevents Chemerin-mediated microvascular injury.

Systemic inflammation and uremic toxins (UT) determine the increased cardiovascular mortality observed in chronic hemodialysis (HD) patients. Among UT, the adipokine Chemerin induces vascular dysfunction by targeting both endothelial and vascular smooth muscular cells (EC and VSMC). As Citrate anion modulates oxidative metabolism, systemic inflammation and vascular function, we evaluated whether citrate-buffered dialysis improves HD efficiency, inflammatory parameters and chemerin-mediated microvascular injury. 45 patients were treated in sequence with acetate, citrate and, again, acetate-buffered dialysis solution (3 months per interval). At study admission and after each treatment switch, we evaluated dialysis efficacy and circulating levels of chemerin and different inflammatory biomarkers. In vitro, we stimulated EC and VSMC with patients' plasma and we investigated the role of chemerin as UT. Citrate dialysis increased HD efficacy and reduced plasma levels of CRP, fibrinogen, IL6 and chemerin. In vitro, patients' plasma induced EC and VSMC dysfunction. These effects were reduced by citrate-buffered solutions and paralleled by the decrease of chemerin levels. Consistently, chemerin receptor knockdown reduced EC and VSMC dysfunction. In conclusion, Switching from acetate to citrate improved dialysis efficacy and inflammatory parameters; in vitro, chemerin-induced EC and VSMC injury were decreased by using citrate as dialysis buffer.

Acute Tubular Injury is Associated With Severe Traumatic Brain Injury: in Vitro Study on Human Tubular Epithelial Cells.

Acute kidney injury following traumatic brain injury is associated with poor outcome. We investigated in vitro the effects of plasma of brain injured patients with acute tubular kidney injury on kidney tubular epithelial cell function. we performed a prospective observational clinical study in ICU in a trauma centre of the University hospital in Italy including twenty-three ICU patients with traumatic brain injury consecutively enrolled. Demographic data were recorded on admission: age 39 ± 19, Glasgow Coma Score 5 (3-8). Neutrophil Gelatinase-Associated Lipocalin and inflammatory mediators were measured in plasma on admission and after 24, 48 and 72 hours; urine were collected for immunoelectrophoresis having healthy volunteers as controls. Human renal proximal tubular epithelial cells were stimulated with patients or controls plasma. Adhesion of freshly isolated human neutrophils and trans-epithelial electrical resistance were assessed; cell viability (XTT assay), apoptosis (TUNEL staining), Neutrophil Gelatinase-Associated Lipocalin and Megalin expression (quantitative real-time PCR) were measured. All patients with normal serum creatinine showed increased plasmatic Neutrophil Gelatinase-Associated Lipocalin and increased urinary Retinol Binding Protein and α1-microglobulin. Neutrophil Gelatinase-Associated Lipocalin was significantly correlated with both inflammatory mediators and markers of tubular damage. Patient' plasma incubated with tubular cells significantly increased adhesion of neutrophils, reduced trans-epithelial electrical resistance, exerted a cytotoxic effect and triggered apoptosis and down-regulated the endocytic receptor Megalin compared to control. Plasma of brain injured patients with increased markers of subclinical acute kidney induced a pro-inflammatory phenotype, cellular dysfunction and apoptotic death in tubular epithelial cells.

Online Hemodiafiltration Inhibits Inflammation-Related Endothelial Dysfunction and Vascular Calcification of Uremic Patients Modulating miR-223 Expression in Plasma Extracellular Vesicles.

Decreased inflammation and cardiovascular mortality are evident in patients with end-stage chronic kidney disease treated by online hemodiafiltration. Extracellular vesicles (EV) are mediators of cell-to-cell communication and contain different RNA types. This study investigated whether mixed online hemodiafiltration (mOL-HDF) beneficial effects associate with changes in the RNA content of plasma EV in chronic kidney disease patients. Thirty bicarbonate hemodialysis (BHD) patients were randomized 1:1 to continue BHD or switch to mOL-HDF. Concentration, size, and microRNA content of plasma EV were evaluated for 9 mo; we then studied EV effects on inflammation, angiogenesis, and apoptosis of endothelial cells (HUVEC) and on osteoblast mineralization of vascular smooth muscle cells (VSMC). mOL-HDF treatment reduced different inflammatory markers, including circulating CRP, IL-6, and NGAL. All hemodialysis patients showed higher plasma levels of endothelial-derived EV than healthy subjects, with no significant differences between BHD and mOL-HDF. However, BHD-derived EV had an increased expression of the proatherogenic miR-223 with respect to healthy subjects or mOL-HDF. Compared with EV from healthy subjects, those from hemodialysis patients reduced angiogenesis and increased HUVEC apoptosis and VSMC calcification; however, all these detrimental effects were reduced with mOL-HDF with respect to BHD. Cell transfection with miR-223 mimic or antagomiR proved the role of this microRNA in EV-induced HUVEC and VSMC dysfunction. The switch from BHD to mOL-HDF significantly reduced systemic inflammation and miR-223 expression in plasma EV, thus improving HUVEC angiogenesis and reducing VSMC calcification.

Perfluorocarbon solutions limit tubular epithelial cell injury and promote CD133+ kidney progenitor differentiation: potential use in renal assist devices for sepsis-associated acute kidney injury and multiple organ failure.

Background: The renal assist device (RAD) is a blood purification system containing viable renal tubular epithelial cells (TECs) that has been proposed for the treatment of acute kidney injury (AKI) and multiple organ failure. Perfluorocarbons (PFCs) are oxygen carriers used for organ preservation in transplantation. The aim of this study was to investigate the effect of PFCs on hypoxia- and sepsis-induced TEC injury and on renal CD133+ progenitor differentiation in a microenvironment similar to the RAD. Methods: TECs were seeded in a polysulphone hollow fibre under hypoxia or cultured with plasma from 10 patients with sepsis-associated AKI in the presence or absence of PFCs and were tested for cytotoxicity (XTT assay), apoptosis (terminal deoxynucleotidyl transferase dUTP nick end labeling assay, caspases, enzyme-linked immunosorbent assay, Fas/Fas Ligand pathway activation), mitochondrial activity, cell polarity [transepithelial electrical resistance (TEER)] and adenosine triphosphate production. The effect of PFCs on proliferation and differentiation of human CD133+ progenitors was also studied. Results: In the presence of PFCs, TECs seeded into the polysulphone hollow fibre showed increased viability and expression of insulin-like growth factor 1, hepatocyte growth factor and macrophage-stimulating protein. Plasma from septic patients induced TEC apoptosis, disruption of oxidative metabolism, alteration of cell polarity and albumin uptake, down-regulation of the tight junction protein ZO-1 and the endocytic receptor megalin on the TEC surface. These detrimental effects were significantly reduced by PFCs. Moreover, PFCs induced CD133+ renal progenitor cell proliferation and differentiation towards an epithelial/tubular-like phenotype. Conclusions: PFCs improved the viability and metabolic function of TECs seeded within a polysulphone hollow fibre and subjected to plasma from septic AKI patients. Additionally, PFCs promoted differentiation towards a tubular/epithelial phenotype of CD133+ renal progenitor cells.

The exciting "bench to bedside" journey of cell therapies for acute kidney injury and renal transplantation.

Acute kidney injury (AKI) is characterized by an increasing incidence and poor outcomes in both developed and undeveloped countries. AKI is also acquiring importance in the setting of kidney transplantation (KT): besides all the classical forms of AKI that KT patients may undergo, several transplant-specific injuries can also lead to the loss of graft function. The mechanisms of tissue damage in native and grafted kidneys share several common pathogenic elements. Since appropriate therapeutic treatments are still lacking-probably due to the disease complexity-clinicians are forced to provide only supportive care. In this composite scenario, cell therapies represent an evolving frontier for AKI treatment in native and transplanted kidneys: ex-vivo manipulated stem or immune cells are able to counteract renal dysfunction by a wide range of biological mechanisms. In this review, we will discuss the potential applications of cell therapies in AKI and KT by analyzing the available clinical data and the most promising experimental prospects from a "bench to bedside" perspective.

Neutrophil Gelatinase Associated Lipocalin Is an Early and Accurate Biomarker of Graft Function and Tissue Regeneration in Kidney Transplantation from Extended Criteria Donors.

BACKGROUND: Delayed graft function (DGF) is an early complication of kidney transplantation (KT) associated with increased risk of early loss of graft function. DGF increases using kidneys from extended criteria donors (ECD). NGAL is a 25KDa protein proposed as biomarker of acute kidney injury. The aim of this study was to investigate the role of NGAL as an early and accurate indicator of DGF and Tacrolimus (Tac) toxicity and as a mediator of tissue regeneration in KT from ECD. METHODS: We evaluated plasma levels of NGAL in 50 KT patients from ECD in the first 4 days after surgery or after Tac introduction. RESULTS: Plasma levels of NGAL at day 1 were significantly higher in DGF group. In the non DGF group, NGAL discriminated between slow or immediate graft function and decreased more rapidly than serum creatinine. NGAL increased after Tac introduction, suggesting a role as marker of drug toxicity. In vitro, hypoxia and Tac induced NGAL release from tubular epithelial cells (TEC) favoring an autocrine loop that sustains proliferation and inhibits apoptosis (decrease of caspases and Bax/Bcl-2 ratio). CONCLUSIONS: NGAL is an early and accurate biomarker of graft function in KT from ECD favoring TEC regeneration after ischemic and nephrotoxic injury.

Caffeic acid, a phenol found in white wine, modulates endothelial nitric oxide production and protects from oxidative stress-associated endothelial cell injury.

INTRODUCTION: Several studies demonstrated that endothelium dependent vasodilatation is impaired in cardiovascular and chronic kidney diseases because of oxidant stress-induced nitric oxide availability reduction. The Mediterranean diet, which is characterized by food containing phenols, was correlated with a reduced incidence of cardiovascular diseases and delayed progression toward end stage chronic renal failure. Previous studies demonstrated that both red and white wine exert cardioprotective effects. In particular, wine contains Caffeic acid (CAF), an active component with known antioxidant activities. AIM OF THE STUDY: The aim of the present study was to investigate the protective effect of low doses of CAF on oxidative stress-induced endothelial injury. RESULTS: CAF increased basal as well as acetylcholine-induced NO release by a mechanism independent from eNOS expression and phosphorylation. In addition, low doses of CAF (100 nM and 1 µM) increased proliferation and angiogenesis and inhibited leukocyte adhesion and endothelial cell apoptosis induced by hypoxia or by the uremic toxins ADMA, p-cresyl sulfate and indoxyl sulfate. The biological effects exerted by CAF on endothelial cells may be at least in part ascribed to modulation of NO release and by decreased ROS production. In an experimental model of kidney ischemia-reperfusion injury in mice, CAF significantly decreased tubular cell apoptosis, intraluminal cast deposition and leukocyte infiltration. CONCLUSION: The results of the present study suggest that CAF, at very low dosages similar to those observed after moderate white wine consumption, may exert a protective effect on endothelial cell function by modulating NO release independently from eNOS expression and phosphorylation. CAF-induced NO modulation may limit cardiovascular and kidney disease progression associated with oxidative stress-mediated endothelial injury.

Endothelial progenitor cell-derived extracellular vesicles protect from complement-mediated mesangial injury in experimental anti-Thy1.1 glomerulonephritis.

BACKGROUND: Endothelial progenitor cells (EPCs) are known to induce tissue repair by paracrine mechanisms including the release of growth factors and extracellular vesicles (EVs), nanoparticles able to carry proteins and genetic information to target cells. The aim of this study was to evaluate whether EVs derived from EPCs may protect from complement-mediated mesangial injury in experimental anti-Thy1.1 glomerulonephritis. METHODS: EVs were isolated by serial ultracentrifugation from supernatants of cultured human EPCs and characterized for their protein and RNA content. In vivo, EVs were injected i.v. in the experimental rat model of mesangiolytic anti-Thy1.1 glomerulonephritis evaluating renal function, proteinuria, complement activity and histological lesions. In vitro, the biological effects of EPC-derived EVs were studied in cultured rat mesangial cells incubated with anti-Thy1.1 antibody and rat or human serum as complement source. RESULTS: After i.v. injection in Thy1.1-treated rats, EVs localized within injured glomeruli and inhibited mesangial cell activation, leucocyte infiltration and apoptosis, decreased proteinuria, increased serum complement haemolytic activity (CH50) and ameliorated renal function. EV treatment decreased intraglomerular deposition of the membrane attack complex (MAC or C5b-9) and expression of smooth muscle cell actin and preserved the endothelial antigen RECA-1 and the podocyte marker synaptopodin. The protective effect of EVs was significantly reduced by pre-treatment with a high dose of RNase (1 U/mL), suggesting a key role for EV-carried RNAs in these mechanisms. Indeed, EPC-derived EVs contained different mRNAs coding for several anti-apoptotic molecules and for the complement inhibitors Factor H, CD55 and CD59 and the related proteins. The in vitro experiments aimed to investigate the mechanisms of EV protection indicated that EVs transferred to mesangial cell mRNAs coding for Factor H, CD55 and CD59 and inhibited anti-Thy1.1 antibody/complement-induced apoptosis and C5b-9/C3 mesangial cell deposition. CONCLUSIONS: EVs derived from EPCs exert a protective effect in Thy1.1 glomerulonephritis by inhibition of antibody- and complement-mediated injury of mesangial cells.

[New mechanisms and recent insights in the pathogenesis of acute kidney injury (AKI)]. / Nuovi meccanismi e recenti acquisizioni nella patogenesi del danno renale acuto.

Acute kidney injury (AKI) is a frequent complication in hospitalized patients often associated with multiple organ failure, increased mortality and progression toward chronic kidney disease. The identification of new cellular and molecular targets involved in AKI may lead to an improvement of diagnostic and therapeutic approaches. In recent years, the pathogenetic mechanisms of AKI have been fully elucidated: tubular epithelial cells and endothelial cells present in the microvasculature have been identified as the main targets of ischemia and of nephrotoxic drugs. Indeed, endothelial cell injury is associated with an extension phase of AKI, whereas tubular cells are subjected to an alteration of cell polarity, mislocalization of tight junction proteins and membrane transporters, and finally to the development of necrosis or apotosis. Apoptosis, or programmed cell death, is also a key component of sepsis-associated AKI in which the mechanisms of tissue damage are associated not only with hypoperfusion but also with a direct detrimental effect of bacterial products and inflammatory mediators on resident kidney cells. Endothelial and tubular epithelial cells also represent the main targets in the immunological mechanisms of AKI in kidney transplantation during cell-mediated and antibody-mediated rejection. Recent studies evidenced new molecules as early biomarkers of AKI. Among these molecules, NGAL and KIM-1 play a possible role in the progression toward chronic kidney disease. Lastly, the new frontier of AKI therapy is represented by the use of bone marrow-derived mesenchymal stem cells able to induce a regenerative program in the damaged kidney.

Microvesicles derived from endothelial progenitor cells protect the kidney from ischemia-reperfusion injury by microRNA-dependent reprogramming of resident renal cells.

Endothelial progenitor cells are known to reverse acute kidney injury by paracrine mechanisms. We previously found that microvesicles released from these progenitor cells activate an angiogenic program in endothelial cells by horizontal mRNA transfer. Here, we tested whether these microvesicles prevent acute kidney injury in a rat model of ischemia-reperfusion injury. The RNA content of microvesicles was enriched in microRNAs (miRNAs) that modulate proliferation, angiogenesis, and apoptosis. After intravenous injection following ischemia-reperfusion, the microvesicles were localized within peritubular capillaries and tubular cells. This conferred functional and morphologic protection from acute kidney injury by enhanced tubular cell proliferation, reduced apoptosis, and leukocyte infiltration. Microvesicles also protected against progression of chronic kidney damage by inhibiting capillary rarefaction, glomerulosclerosis, and tubulointerstitial fibrosis. The renoprotective effect of microvesicles was lost after treatment with RNase, nonspecific miRNA depletion of microvesicles by Dicer knock-down in the progenitor cells, or depletion of pro-angiogenic miR-126 and miR-296 by transfection with specific miR-antagomirs. Thus, microvesicles derived from endothelial progenitor cells protect the kidney from ischemic acute injury by delivering their RNA content, the miRNA cargo of which contributes to reprogramming hypoxic resident renal cells to a regenerative program.

Microvesicles derived from endothelial progenitor cells enhance neoangiogenesis of human pancreatic islets.

The efficacy of islet transplantation is limited by poor graft vascularization. We herein demonstrated that microvesicles (MVs) released from endothelial progenitor cells (EPCs) enhanced human islet vascularization. After incorporation into islet endothelium and ß-cells, EPC-derived MVs favored insulin secretion, survival, and revascularization of islets transplanted in SCID mice. MVs induced in vitro islet endothelial cell proliferation, migration, resistance to apoptosis, and organization in vessel-like structures. Moreover, MVs partially overcame the antiangiogenic effect of rapamycin and inhibited endothelial-leukocyte interaction via L-selectin and CD40. MVs were previously shown to contain defined patterns of mRNAs. Here we demonstrated that MVs carried the proangiogenic miR-126 and miR-296 microRNAs (miRNAs). MVs pretreated with RNase or derived from Dicer knocked-down EPCs showed a reduced angiogenic effect. In addition, MVs overcame the antiangiogenic effect of the specific antagomiRs of miR-126 and miR-296, suggesting a relevant contribution of miRNAs delivered by MVs to islet endothelium. Microarray analysis of MV-stimulated islet endothelium indicated the upregulation of mRNAs coding for factors involved in endothelial proliferation, differentiation, and angiogenesis. In addition, MVs induced the activation of the PI3K-Akt and eNOS signaling pathways in islet endothelium. These results suggest that MVs activate an angiogenic program in islet endothelium that may sustain revascularization and ß-cell function.