MUSCLE RELAXANT DERIVED FROM HUMAN GINGIVAL MICROVASCULAR ENDOTHELIAL CELLS

点击翻译按钮获取中文摘要

The purpose of this investigation was to detect the smooth muscle relaxant (Nitric oxide, No) released from human gingival microvascular endothelial cells (HGECs). The effect of mechanical stimulation on NO-release from HGECs was also compared with that from human umbilical vein and artery endothelial cells. Afer collagenase treatment, human gingival samples were compressed to release endothelial cells. Centrifuged cells were resuspended in the growth medium. The cells suspension was introduced into every well of a 96-well plate. Morphologically homogeneous cells with a cobblestone appearance were harvested. The isolated cells were identified as HGECs by the presence of von Willebrand factor, the uptake of acetylated LDL,and the formation of capillary-like structure on Matrigel. Fluorometric assay was used to detect the NO.The sample was reacted with DAN reagent to form 1- (H) -naphthotiazol. After 10-min incubation at 20 ﾟC,the reaction was terminated with NaOH.The end product, 2,3-diamino-naphthotriazol, was measured using a fluorescent plate reader with excitation at 360 nm and emission read at 460nm. At baseline, none of the cells tested released the NO.The NO was detected in the culture medium of the umbilical vein and artery endothelial cells after 3-day-and 5-day-mechanical stimulation by Flexercell. However, the HGECs did not release the NO during the whole experimental period. The cultivable HGECs were passage 4, which were thought not to be enough for detecting the NO.The further study will focus on more efficient method for cultivating the HGECs.