J P Gertler1, V H Ocasio. 1. Division of Vascular Surgery, Massachusetts General Hospital, Boston 02114.
Abstract
PURPOSE: The physiologic significance of endothelin remains incompletely defined. Procoagulant and antifibrinolytic activities are increased in hypoxic cultured human umbilical venous endothelial cells (HUVEC). We examined the effect of hypoxia on HUVEC endothelin-1 production in vitro to determine whether a correlation existed between the procoagulant and antifibrinolytic response to hypoxia previously observed and an increase in vasoconstrictor peptide secretion by hypoxic HUVEC. METHODS: Cultured HUVEC were rendered hypoxic (PO2 = 40 mm Hg) or control (PO2 = 120 mm Hg) for 24 hours. Media were either standard, 5 gm glucose/L (high glucose), or contained 500 units superoxide dismutase/ml (SOD). Endothelin-like immunoreactivity for endothelin-1 (ET-IR) in conditioned media was measured by radioimmunoassay and expressed as mean femtomoles per milliliter (+/- SD) per 100,000 cells. Viability of HUVEC was assessed by trypan blue exclusion. Significance was determined by use of Student's t test. RESULTS: Conditioned media from hypoxic cells contained 76% more ET-IR than was found in control counterparts (p < 0.004). The addition of high glucose or SOD did not diminish ET-IR; a trend to higher ET-IR was present in both these groups versus standard media (303% and 226%, respectively, p < 0.03). CONCLUSIONS: Thus 24 hours of hypoxia caused an increase in conditioned-media ET-IR in cultured HUVEC. Because SOD or greater substrate availability did not diminish endothelin presence in conditioned media, it seems that hypoxic induction of endothelin-1 production or secretion is signaled in a fashion unrelated to cell toxicity from the hypoxic period.
PURPOSE: The physiologic significance of endothelin remains incompletely defined. Procoagulant and antifibrinolytic activities are increased in hypoxic cultured human umbilical venous endothelial cells (HUVEC). We examined the effect of hypoxia on HUVEC endothelin-1 production in vitro to determine whether a correlation existed between the procoagulant and antifibrinolytic response to hypoxia previously observed and an increase in vasoconstrictor peptide secretion by hypoxic HUVEC. METHODS: Cultured HUVEC were rendered hypoxic (PO2 = 40 mm Hg) or control (PO2 = 120 mm Hg) for 24 hours. Media were either standard, 5 gm glucose/L (high glucose), or contained 500 units superoxide dismutase/ml (SOD). Endothelin-like immunoreactivity for endothelin-1 (ET-IR) in conditioned media was measured by radioimmunoassay and expressed as mean femtomoles per milliliter (+/- SD) per 100,000 cells. Viability of HUVEC was assessed by trypan blue exclusion. Significance was determined by use of Student's t test. RESULTS: Conditioned media from hypoxic cells contained 76% more ET-IR than was found in control counterparts (p < 0.004). The addition of high glucose or SOD did not diminish ET-IR; a trend to higher ET-IR was present in both these groups versus standard media (303% and 226%, respectively, p < 0.03). CONCLUSIONS: Thus 24 hours of hypoxia caused an increase in conditioned-media ET-IR in cultured HUVEC. Because SOD or greater substrate availability did not diminish endothelin presence in conditioned media, it seems that hypoxic induction of endothelin-1 production or secretion is signaled in a fashion unrelated to cell toxicity from the hypoxic period.