Human umbilical vein endothelial cells (HUVECs) were cultured on microcarrier beads to accommodate different experiment apparatus such as rotating wall vessel. In this study, fluid operating apparatus (FPA) was used. However, the effect of inflammation and endothelial activation biomarkers in HUVECs cultured on different culture surface and containers are not well established. The effects of temperature changes on these biomarkers in HUVECs grown in FPA, a spaceflight hardware, are still unclear. The objective of this study was to compare the protein and gene expression of inflammation and endothelial activation biomarkers in (i) HUVECs cultured on microcarrier beads in conventional culture flask (CCFMC) vs. conventional culture flask (CCF) (ii) HUVECs cultured on microcarrier in FPA (FPAMC) vs. CCFMC and (iii) HUVEC cultured in FPAMC with ideal temperature (37°C) (FPAMC) vs. simulated space travel temperature(25-37°C), (FPAMC-ST). sICAM-1 and sVCAM-1protein expression in HUVECs grown in CCFMC were higher than CCF. FPAMC had higher IL-6, TNF-α, ICAM-1, VCAM-1, e-selectin, NFκB and eNOS gene expression than in CCFMC. FPAMC-ST had higher ICAM-1 and e-selectin protein expression than FPAMC- in ideal temperature. HUVECs are cultured onto microcarrier in simulated space flight temperature compared with ideal temperature had higher protein expression of sICAM-1 and e-selectin but the protein and gene expression of other biomarkers of inflammation and endothelial activation are comparable. This suggests that differences in culture surface and container are have an impact on the expression of inflammation and adhesion molecule by HUVECs.