Probiotic Lisosan G preserves functional properties of human endo- thelial progenitor cells exposed to lipopolysaccharides

Background and aims: Endogenous signals (such as oxidized-LDL) and exogenous signals (such as lipopolysaccharides, LPS) derived by the gut microbiota orchestrate inflammatory responses and con- tribute to plaque evolution in obesity, metabolic syndrome and dia- betes. The involvement of LPS as a potentially important source of vascular inflammation in the induction of atherogenesis has been postulated. Several strategies have been developed to change gut microbiota. For instance, probiotics can modify gut microbiota. With this background, we have evaluated the effects of the probiotic Lisosan G (LG) on human endothelial progenitor cells (EPCs) exposed to LPS. Materials and methods: Early EPCs, obtained from peripheral blood mononuclear cell (PBMC) of healthy subjects, pre-treated with LG 0.7 mg/ml for 1 hour, and/or exposed to LPS 10 μg/ml for the next 23 hours, have been evaluated at 5-hour and 24-hour for: 1. cells viability; and 2. reactive oxigen species (ROS) production; at 24-hour for: 3. adhesion to fibronectin; and 4. expression of IL-6, COX-2, ICAM-1, ET-1, Casp-3, Casp-9, CHOP, SOD2, CAT, GPx1; at 5-hour and at 24-hour for: 5. the nuclear factor NF-kB translocation. Results: First, cells viability at 5-hour and 24-hour: no differences in LG-/ LPS- (the control experimental condition, C), vs. LG+/LPS-, LG-/LPS+ and LG+/LPS+. Adhesion at 24-hour: it was higher (p=0.021) in LG+/ LPS+ vs. C, marginally higher (p=0.09) vs. other experimental condi- tions. ROS production: differences have been observed both at 5h (p=0.0026) and 24h (p=0.051); reduced in LG+/LPS- vs. C (p=0.033 and p=0.119, at 5h and 24h, respectively); marginally increased in LG-/ LPS+ vs. C at 5h and 24h; reduced in LG+/LPS+ vs. LG-/LPS+ (p=0.0043 and p=0.067, at 5h and 24h, respectively). At 24h, as com- pared to C, LPS (LG-/LPS+) rises expression of IL-6, COX-2, ICAM-1 (p=0.0001 for all), ET-1 (p=0.0003), Casp-3 e Casp-9 (p=0.005), CHOP (p=0.0004), SOD2 (p=0.0001) and GPx1 (p=0.0073). LG+/LPS+ de- creases IL-6 and ICAM-1 (both p=0.0001), normalizes COX-2 (p=0.0001), ET-1 (p=0.0002), Casp-3 (p=0.0001), Casp-9 (p=0.0001) and CHOP (p=0.0001), further increases SOD2 (p=0.0017) and GPx1 (p=0.0048). Finally, LPS pauperises CAT (p=0.012), with LG normaliz- ing its expression. In C and in LG+/LPS-, NF-kB was mainly localized into the citosol, in LG-/LPS+ mainly within the nucleus; in LG+/LPS+ nuclear translocation of NF-kB was reduced. Conclusion: In human EPCs, LPS increases ROS, up-regulates pro-in- flammatory tone, pro-apoptotic factors and anti-oxidants. Lisosan G pro- tects human EPCs exposed to LPS reducing ROS, down-regulating pro- inflammatory and pro-apoptotic factors and strengthening anti-oxidant defenses through NF-kB translocation. Supported by: Regione Toscana, Grant n. D55E11002680005