Sphingosine 1-phosphate receptor 1 is required for MMP-2 function in bone marrow mesenchymal stromal cells: implications for cytoskeleton assembly and proliferation

Bone marrow-derived mesenchymal stromal cell- (BM-MSC-) based therapy is a promising option for regenerative medicine. An important role in the control of the processes influencing the BM-MSC therapeutic efficacy, namely, extracellular matrix remodelling and proliferation and secretion ability, is played by matrix metalloproteinase- (MMP-) 2. Therefore, the identification of paracrine/autocrine regulators of MMP-2 function may be of great relevance for improving BM-MSC therapeutic potential. We recently reported that BM-MSCs release the bioactive lipid sphingosine 1-phosphate (S1P) and, here, we demonstrated an impairment of MMP-2 expression/release when the S1P receptor subtype S1PR1 is blocked. Notably, active S1PR1/MMP-2 signalling is required for F-actin structure assembly (lamellipodia, microspikes, and stress fibers) and, in turn, cell proliferation. Moreover, in experimental conditions resembling the damaged/regenerating tissue microenvironment (hypoxia), S1P/S1PR1 system is also required for HIF-1α expression and vinculin reduction. Our findings demonstrate for the first time the trophic role of S1P/S1PR1 signalling in maintaining BM-MSCs' ability to modulate MMP-2 function, necessary for cytoskeleton reorganization and cell proliferation in both normoxia and hypoxia. Altogether, these data provide new perspectives for considering S1P/S1PR1 signalling a pharmacological target to preserve BM-MSC properties and to potentiate their beneficial potential in tissue repair

Mesenchymal Stromal Cell Secreted Sphingosine 1-Phosphate (S1P) Exerts a Stimulatory Effect on Skeletal Myoblast Proliferation.

Bone-marrow-derived mesenchymal stromal cells (MSCs) have the potential to significantly contribute to skeletal muscle healing through the secretion of paracrine factors that support proliferation and enhance participation of the endogenous muscle stem cells in the process of repair/regeneration. However, MSC-derived trophic molecules have been poorly characterized. The aim of this study was to investigate paracrine signaling effects of MSCs on skeletal myoblasts. It was found, using a biochemical and morphological approach that sphingosine 1-phosphate (S1P), a natural bioactive lipid exerting a broad range of muscle cell responses, is secreted by MSCs and represents an important factor by which these cells exert their stimulatory effects on C2C12 myoblast and satellite cell proliferation. Indeed, exposure to conditioned medium obtained from MSCs cultured in the presence of the selective sphingosine kinase inhibitor (iSK), blocked increased cell proliferation caused by the conditioned medium from untreated MSCs, and the addition of exogenous S1P in the conditioned medium from MSCs pre-treated with iSK further increased myoblast proliferation. Finally, we also demonstrated that the myoblast response to MSC-secreted vascular endothelial growth factor (VEGF) involves the release of S1P from C2C12 cells. Our data may have important implications in the optimization of cell-based strategies to promote skeletal muscle regeneration

The Efficacy of Umbelliferone, Arbutin, and N-Acetylcysteine to Prevent Microbial Colonization and Biofilm Development on Urinary Catheter Surface: Results from a Preliminary Study

We evaluated, in a preliminary study, the efficacy of umbelliferone, arbutin, and N-acetylcysteine to inhibit biofilm formation on urinary catheter. We used 20 urinary catheters: 5 catheters were incubated with Enterococcus faecalis (control group); 5 catheters were incubated with E. faecalis in presence of umbelliferone (150 mg), arbutin (60 mg), and N-acetylcysteine (150 mg) (group 1); 5 catheters were incubated with E. faecalis in presence of umbelliferone (150 mg), arbutin (60 mg), and N-acetylcysteine (400 mg) (group 2); and 5 catheters were incubated with E. faecalis in presence of umbelliferone (300 mg), arbutin (60 mg), and N-acetylcysteine (150 mg) (group 3). After 72 hours, planktonic microbial growth and microorganisms on catheter surface were assessed. In the control group, we found a planktonic load of ≥105 CFU/mL in the inoculation medium and retrieved 3.69 × 106 CFU/cm from the sessile cells adherent to the catheter surface. A significantly lower amount in planktonic (p < 0.001) and sessile (p = 0.004) bacterial load was found in group 3, showing <100 CFU/mL and 0.12 × 106 CFU/cm in the incubation medium and on the catheter surface, respectively. In groups 1 and 2, 1.67 × 106 CFU/cm and 1.77 × 106 CFU/cm were found on catheter surface. Our results document that umbelliferone, arbutin, and N-acetylcysteine are able to reduce E. faecalis biofilm development on the surface of urinary catheters

Improvement of Legionnaires' disease diagnosis using real-time PCR assay: a retrospective analysis, Italy, 2010 to 2015

AimTo evaluate real-time PCR as a diagnostic method for Legionnaires' disease (LD). Detection of Legionella DNA is among the laboratory criteria of a probable LD case, according to the European Centre for Disease Prevention and Control, although the utility and advantages, as compared to culture, are widely recognised.MethodsTwo independent laboratories, one using an in-house and the other a commercial real-time PCR assay, analysed 354 respiratory samples from 311 patients hospitalised with pneumonia between 2010-15. The real-time PCR reliability was compared with that of culture and urinary antigen tests (UAT). Concordance, specificity, sensitivity and positive and negative predictive values (PPV and NPV, respectively) were calculated.ResultsOverall PCR detected eight additional LD cases, six of which were due to Legionella pneumophila (Lp) non-serogroup 1. The two real-time PCR assays were concordant in 99.4% of the samples. Considering in-house real-time PCR as the reference method, specificity of culture and UAT was 100% and 97.9% (95% CI: 96.2-99.6), while the sensitivity was 63.6% (95%CI: 58.6-68.6) and 77.8% (95% CI: 72.9-82.7). PPV and NPV for culture were 100% and 93.7% (95% CI: 91.2-96.3). PPV and NPV for UAT were 87.5% (95% CI: 83.6-91.4) and 95.8% (95% CI: 93.5-98.2).ConclusionRegardless of the real-time PCR assay used, it was possible to diagnose LD cases with higher sensitivity than using culture or UAT. These data encourage the adoption of PCR as routine laboratory testing to diagnose LD and such methods should be eligible to define a confirmed LD case

Modulation of MMP-2 function in bone marrow mesenchymal stromal cells requires sphingosine 1-phopsphate receptor 1 mediated signaling: implications for cytoskeletal assembly and proliferation

Bone-marrow-derived mesenchymal stromal cells (BM-MSCs)–based therapy represents a promising option in the field of regenerative medicine. Their therapeutic potential is mainly dependent on paracrine secretion, proliferation and ECM remodeling abilities whose modulation involves Matrix Metalloproteinase (MMP)-2 functionality. Thus, the identification of paracrine/autocrine factors regulating MMP-2 expression/activity may be of great biological relevance for potentiating BM-MSC theraputic efficacy. Our research group has demonstrated that BM-MSCs release the bioactive lipid sphingosine-1-phosphate (S1P). Here we demonstrated : i) the requirement for BM-MSC of S1P production to synthesize functional gelatinases; ii) an impairment of gelatinolytic activity and MMP-2 expression/release when the S1P receptor subtype 1 (S1PR1) is blocked. Notably, in these experimental conditions BM-MSCs did not exhibit the formation of plasmamembrane-associated F-actin structures (lamellipodia, filopodia, microspikes) and, in turn, showed a reduction of the proliferation rate. Moreover, S1P1-mediated signaling is required for HIF-1alpha expression and MMP-2 expression/activity, reduction of vinculin expression and stress fiber formation and proliferation in hypoxia, an experimental condition mimicking the injured/regenerating tissue microenvironment. In conclusion, our findings, demonstrating the trophic role exerted by the autocrine S1P/S1PR1 signaling in maintaining BM-MSC ability to modulate MMP-2 function, required for ECM remodeling, cytoskeleton assembly and cell proliferation may provide perspectives for considering S1P/S1PR1 as a pharmacological target to preserve BM-MSCs properties and improve their efficacy in tissue repair

Inflammation and urothelial bladder cancer. What we need to know?

The association between inflammation and bladder cancer has been debated in several studies, highlighting that inflammation may be a crucial component both in tumor development or progression. On the other hand, several authors suggest that the presence of an inflammatory cell infiltrate within the urothelial bladder cancer is a good prognostic predictor in terms of recurrence-free survival time. The question is: What is the prognostic role of inflammation in patients affected by urothelial bladder cancer? On one hand, chronic inflammation should be considered a risk factor in developing bladder cancer, as demonstrated by Schistosoma haematobium infection and, on the other hand, the inflammation induced by the Bacillus Calmette-Guérin intravesical therapy has a protective effect on cancer recurrence. Recently, some authors highlight that the presence of an inflammatory cell infiltrate within the urothelial bladder cancer is a good prognostic predictor in terms of recurrence-free survival time, due to the host generating angiogenic stimulation of a local inflammatory reaction against cancer. This is probably due to the angiogenetic stimulation of a local inflammatory reaction generated by the host against superficial bladder cancer. However, the debate is still open. This review will summarize recent data regarding inflammation and urothelial cell carcinoma, with special emphasis on the role that the inflammatory response is likely to have on recurrence risk and progression in superficial bladder cancer patients