One recent line of cancer research shows increasing interest for biological factor such as IL-2, TNF-alpha, and leptin, which have been found to participate in the development and progression of non-small cell lung cancer (NSCLC). The aim of this study was to measure IL-2, TNF-alpha, and leptin concentrations in the airways and in the systemic circle of patients with NSCLC, investigating the role of these factors in the lung tumors. We enrolled 32 patients (17 men, 71 +/- 7 years) with a histological diagnosis of NSCLC and 20 healthy ex-smoker controls, negative for computed tomography of the chest (14 men, 69 +/- 8 years). IL-2, TNF-alpha, and leptin levels were measured in the serum, the urine, the bronchoalveolar lavage, the induced sputum, and exhaled breath condensate (EBC) of patients enrolled by means of a specific enzyme immunoassay kit. Higher concentrations of IL-2, TNF-alpha and leptin were found in NSCLC patients than in controls (p < 0.0001). A statistically significant increase of IL-2, TNF-alpha, and leptin concentrations was observed in patients from stage I to stage III of NSCLC. These findings suggest that IL-2, TNF-alpha, and the leptin play an important role in the cancerogenesis of NSCLC. Their measure in the EBC could be proposed as noninvasive markers for an early detection of NSCLC and in the follow-up of this tumor.

No abstract

Background and Aims EVs (Extracellular vesicles) are circulating microparticles able to mediate cell-to cell communication by carrying proteins, DNA, RNA or antibodies on their surface. EV are emerging as pivotal in renal Antibody-mediated rejection (AMR), one of the major causes of transplant failure associated to a massive complement activation. AMR is characterized by endothelial to mesenchymal transition (EndMT) and the occurrence of tubular accelerated senescence process, known as graft “Inflammaging”. However, the potential role of EVs in accelerating the renal aging and graft fibrosis after AMR is not well understood. Method Plasma EVs were isolated from 10 Acute AMR (AAMR), 10 Chronic AMR (CAMR) patients, 5 stable graft transplanted patients and 5 healthy volunteers. EVs were isolated by ultracentrifugation at 100.000 g for 1h at 4°C, quantified by Nanoparticle Tracking Analysis (Nanosight, NTA EV/ml) and characterized by FACS (Attune Nxt). RPTEC and endothelial cell culture were incubated with EVs (5e+4 EVs/cells target for 24h. MTT test was performed to assess cell viability. Cellular senescence was investigated by qPCR for p21, p53, Klotho and CYP1B1 and SA-β-gal staining were performed. To assess EndMT analysis for CD31, VE-cadherin, Vimentin, Collagen I was performed by FACS. mRNA level of C3 and CFH were also measured by qPCR and C4d deposits were evaluated in endothelial cell colture by IF. Renal biopsies were then analyzed for inflammaging (p16INK4a) and EndMT markers (CD31/αSMA) by IHC and IF. Results The Nanosight analysis showed significant differences in the amount of-EVs count per ml of plasma in AMR patients compared to healthy subjects. EVs appeared to be significantly augmented in acute AMR, in a higher manner than chronic AMR (NTA, p=0.0154). By cytofluorimetric analysis, the endothelial markers CD31 and VE- cadherin appeared to be significantly increased compared of healthy control, indicating a predominant endothelial EV origin (p&lt;0.05). Furthermore, an increase for CD3, CD4, CD8, CD40 and CD40L lymphocyte/monocytes markers was found. In vitro, the exposure of RPTEC and endothelial cells to AMR-derived EVs did not induce the loss of cell proliferation. However, AMR-derived EVs induced senescence in RPTEC, as observed by increase in SA-β-gal positive cells, upregulation of p21, p53, CYP1B1 and downregulated KL gene expression (p&lt;0.05). EVs induced the EndMT as observed by FACS with the downregulation of CD31, VE-cadherin (CD144) and increase of Vimentin and Collagen I (p=0.025). To evaluate the contribution of EVs to local complement activation, C3 and CFH qPCR analysis were performed. EVs from AMR patients induced a significant increase in C3 gene expression with concomitant downregulation in CFH in RPTEC. EV exposition induced the classical and lectin pathway activation in endothelial culture medium, and C4d deposition. These data support the hypothesis that circulating EVs can amplify local complement activation in systemic endothelial and tubular cells during AMR, therefore leading to accelerated senescence and fibrosis as later effects. Finally, renal AMR biopsies showed significant tubular senescence as indicated by p16 expression; p16 was significantly upregulated in Chronic compared with Acute AMR biopsies (p&lt;0.05). The AMR biopsies showed positivity for EndMT, as indicated by CD31 decrease and interstitial αSMA upregulation (p&lt;0.05). Conclusion These results suggest a putative role for circulating AMR derived-EVs in inducing the tubular inflammaging by local complement activation and early fibrosis by EndMT. The EVs cargo characterization that is ongoing might highlight novel targets for therapeutic intervention.

Introduction and Aims: Acute kidney injury (AKI) is emerging as a public health problem worldwide. Several pharmacologic therapies that can accelerate recovery and improve survival have been attempted and were efficacious in experimental models but failed to manifest any substantial beneficial effect in clinical practice. Recent studies showed that adult renal progenitor cells (ARPCs) can participate in kidney repair processes and might be potentially used in the clinic to improve regeneration in acute and progressive kidney disease. Our aim was to study the influence of ARPCs on the regenerative process of cisplatin-injured renal proximal tubular epithelial cells (RPTECs) and validate some the ARPC-secreted molecules able to induce regenerative processes. Methods: CD133-positive ARPCs were isolated by magnetic sorting, starting from healthy sections of kidney removed for renal carcinoma. An in vitro model of cisplatin induced cell toxicity, in which RPTECs were co-cultured with ARPCs, was used. Caspase 3 expressions was studied to evaluate apoptosis. Cell proliferation induced by ARPCs was evaluated by BrdU proliferation assay. ELISA assays were performed to evaluate inhibin-A (INHB-A), decorin (DCN) and FGF2 chemokines. A natural polymer-based nanosystem for efficacious delivery of molecules was developed. INHB-A-loaded Polysaccharides Synthetic Vesicles (INHB-A-PSSV) and DCN-loaded Polysaccharides Synthetic Vesicles (DCN-PSSV) were synthesized by two steps methods: ionotropic pre-gelation of alginate core, followed by chitosan polyelectrolyte complexation. A microfluidic device was appositely fabricated in order to optimize the INHB-A and DCN-PSSV at interface-assembly process, in terms of polymers and INHB-A and DCN amount as well as vesicles size distribution. Cellular uptake and INHB-A-PSSV and DCN-PSSV effectiveness were tested. Results: We showed that the induction of tubular cell regeneration process was specific of tARPCs and occured only after that they detect the damage. On the contrary, glomerular ARPCs could not induce RPTEC regeneration. tARPCs protected RPTEC and HK2 cells from cisplatin toxicity by preventing cisplatin-induced apoptosis and enhancing proliferation of survived cells. Regenerative effect was completely abrogated blocking the Toll-like receptor 2 (TLR2), or using tARPC not expressing the TLR2. We found that INHB-A, DCN and FGF2 were secreted by ARPCs following the damage of RPTEC and that they were involved in the RPTEC regeneration. We showed that addition of INHB-A and DCN PSSV to cisplatin-treated RPTECs led to a substantial increase in cell number and viability after 3 days of culture. Remarkably, a very low dosage of functional loaded proteins (8 ng/25 ul) was sufficient to induce cell regeneration and the percentage of viable cells was similar to that of RPTECs without damage. Conclusions: We demonstrated that tARPCs have a regenerative effect on damaged RPTEC, by both preventing apoptosis and enhancing proliferation of surviving cells. They act by means of their secretoma, t...

No abstract