Anti-E. coli Beta Galactosidase Polyclonal antibody

Background: Ageing is associated with progressive endothelial senescence and dysfunction, and cardiovascular risk. Circulating endothelial microvesicles (MVs) are pro-senescent and pro-inflammatory endothelial effectors in acute coronary syndrome. Omega-3 PUFA intake was claimed beneficial in cardiovascular prevention. Purpose: To investigate whether the intake of the omega-3 formulation EPA:DHA 6:1 by middle-aged and old rats reduces the shedding of pro-senescent microvesicles from cultured spleen leukocytes (SMVs) and clarify the underlying mechanisms in target coronary primary endothelial cells (ECs). Methods: Middle-aged male Wistar rats (M, 48-week old) received 500 mg/kg/d of either EPA:DHA 6:1, EPA:DHA 1:1, or vehicle (CTL) for 7 days, old rats (72-week old) for 14 days. Spleen-derived leukocytes were prepared and cultured for 24 h and MVs collected from supernatants (SMVs). Cultured ECs were prepared from freshly isolated porcine coronary arteries. Senescence-associated beta-galactosidase activity (SA-beta-gal) was assessed by C12FDG, protein expression by Western blot analysis, oxidative stress by dihydroethidium using confocal microscopy, and procoagulant MVs by prothrombinase assay. The pro-senescent potential of SMVs from middle-aged rats (M-SMVs) was analyzed by comparison with young (Y, 12-week) and old (O) rats. Results: The shedding of SMVs significantly increased with age and was inhibited by EPA:DHA 6:1 intake that also prevented ROS accumulation in spleen. Incubation of ECs with 10 nM SMVs from middle-aged and old but not those from young rats induced premature senescence after 48 h. The pro-senescent effect of M-SMVs was prevented by Losartan and associated with endothelial oxidative stress. M-SMVs induced an up-regulation of senescence markers (p16, p21, p53), pro-atherothrombotic (VCAM-1, ICAM-1, tissue factor) and pro-inflammatory markers (pNF-kappa B, COX2) and proteins of the angiotensin system (ACE, AT1-R). Conversely, endothelial NO synthase was down-regulated. Intake of EPA:DHA 1:1 and 6:1 by middle-aged rats decreased SMV shedding by 14% and 24%, respectively. Only EPA:DHA 6:1 intake abolished the M-SMVs-induced endothelial senescence and reduced the pro-senescent action of O-SMVs by 45%. Protection of ECs was not observed in response to SMVs from EPA:DHA 1:1 treated rats. Conclusion: Ingestion of EPA:DHA 6:1 by middle-aged or old rats, respectively abolished or limited both the shedding of SMVs and their pro-senescent, pro-thrombotic and pro-inflammatory effects in ECs, most likely by triggering the local angiotensin system. EPA:DHA 6:1 may help to delay ageing-related endothelial dysfunction.

Objective. To preliminary explore the senescent dynamic changes of the bone marrow mesenchymal stem cells (BMMSCs) by human ageing and its possible mechanism. Methods. The bone marrows were harvested from healthy volunteers, and according to volunteers' age, these were divided into group A (<= 25 years), group B (26-45 years), group C (46-65 years), and group D (>65 years). Totally, the bone marrows were extracted from the posterior superior iliac spine from volunteers under aseptic conditions. Diluted with isovolumic PBS, followed by centrifugation at 1x10(5)/cm(2), cells were cultured in a 5% CO2 incubator at 37 degrees C. After three passages, surface marker identification of hBMMSCs was tested by flow cytometry (FCM), oil red O staining was used to observe the ability of osteogenic differentiation, alkaline phosphatase (ALP) staining and the levels of osteocalcin (OST) in the supernatants were used to observe the ability of adipogenic differentiation, senescence-associated beta-galactosidase (SA-beta-Gal) staining was used to detect the senescent BMSCs, the ability of BMSC proliferation was detected by cell counting kit-8 (CCK-8), the distribution of the cell cycle was analyzed by flow cytometry (FCM), and malondialdehyde (MDA) content, total glutathione peroxidase, total antioxidant capacity, and total superoxide dismutase (SOD) activity was analyzed using enzymatic assay. Results. The BMSCs highly expressed CD73 and CD90, but lowly expressed CD34 and CD19/CD14. With age, osteogenic differentiation was markedly increased and audiogenic differentiation was significantly decreased. The number of SA-beta-gal-positive cells was significantly increased, the proliferation ability of hBMMSCs declined, the BMSCs were held in the G1 phase, the MDA level of BMSCs was significantly increased, and total glutathione peroxidase, total antioxidant capacity, and SOD activity significantly declined. Conclusions. With age, the aging BMSCs were intensified; the mechanism may be related to oxidative damage mediated aging-related pathways.