Rat Anti-Murine Vcam1 Heterohybridoma [N/L-2.0]

Human monocytes comprise three distinct subsets, defined by their relative expression of CD14 and CD16. These subsets appear to have different functional roles within homeostasis and inflammation, but little is known about the manner in which they interact with macro- and microvascular endothelial cells, a key enabling component for the fulfillment of their functional roles. In the present study, we examined the locomotory behavior of the three major human monocyte subsets over human endothelial monolayers subjected to physiologically relevant levels of shear flow in vitro. Each subset was shown to preferentially perform different types of locomotory behavior in a resting state. A long-range crawling behavior, similar to the "patrolling" behavior of murine Ly6C(-) monocytes, was observed in CD14(+)CD16(-) and CD14(dim) CD16(+) monocytes, but not in CD14(+)CD16(+) monocytes. CD14(dim)CD16(+) and CD14(+)CD16(-) monocytes showed a preference for adhering to microvascular over macrovascular endothelium, whereas CD14(+)CD16(+) monocytes showed the opposite. Transendothelial migration was not observed in CD14(dim)CD16(+) monocytes during the 30-min observation period. Long-range crawling behavior in CD14(dim)CD16(+) monocytes was abrogated by blockade of ICAM1, VCAM1, or CX(3)CL1, in contrast with CD14(+)CD16(-) monocytes, which only required ICAM1 for this behavior. These studies indicate the existence of subtype-specific human monocyte migratory behavior patterns with distinct adhesion molecule dependence, which may assist in elucidating their physiological function and relevance to disease.

Background: Obesity is associated with hyperleptinemia and endothelial dysfunction. Hyperleptinemia has been reported to induce both oxidative stress and inflammation by increasing reactive oxygen species production. Objective: The objective of this study was to determine the effects of 1,25-dihydroxycholecalciferol [1,25(OH)(2)D-3] against leptin-induced oxidative stress and inflammation in human endothelial cells. Methods: Small interfering RNA (siRNA) were used to knock down the expression of vitamin D receptor (VDR) in human umbilical vein endothelial cells (HUVECs). HUVECs were pretreated for 4 h with physiologic (10(-10) M) or supraphysiologic (10(-7) M) concentrations of 1,25(OH)(2)D-3 and exposed to leptin (10 ng/mL). Superoxide anion production and translocation of nuclear factor (erythroid-derived 2)-like 2 (NRF2) and nuclear transcription factor kappa B (NF-kappa B) subunit p65 to the nucleus and the activation of their target genes were quantified. Results: Pretreatment of HUVECs with 1,25(OH) 2D3 prevented the leptin-induced increase in superoxide anion production (P < 0.05). Pretreatment with 1,25(OH)(2)D-3 further increased NRF2 translocation to the nucleus (by 3-fold; P < 0.05) and increased mRNA expression of superoxide dismutase 2 (SOD2; by 2-fold), glutathione peroxidase (GPX; by 3-fold), NAD(P)H dehydrogenase (quinone) 1 (NQO1; by 4-fold), and heme oxygenase 1 (HMOX1; by 2-fold) (P < 0.05). Leptin doubled the translocation of NF-kappa B (P < 0.05) to the nucleus and increased (P < 0.05) the upregulation of vascular inflammatory mediators such as monocyte chemoattractant protein 1 (MCP1; by 1-fold), transforming growth factor beta (TGF beta; by 1-fold), and vascular cell adhesion molecule 1 (VCAM1; by 4-fold) (P < 0.05), which were prevented (P < 0.05) by pretreatment with 1,25(OH)(2)D-3. Protective effects of 1,25(OH)(2)D-3 were confirmed to be VDR dependent by using VDR siRNA. Conclusion: Pretreatment with 1,25(OH)(2)D-3 in the presence of a high concentration of leptin has a beneficial effect on HUVECs through the regulation of mediators of antioxidant activity and inflammation.