LPS-induced effects on angiotensin I-converting enzyme expression and shedding in human pulmonary microvascular endothelial cells.

Angiotensin I-converting enzyme (kininase II, ACE, and CD143) availability is a determinant of local angiotensin and kinin concentrations and their physiological actions. Until now, it is unclear whether the decrease of pulmonary ACE activity in sepsis-described in clinical studies-is due to an enzyme compensatory downregulation (reduced ACE-mRNA expression) to shedding of ACE or endothelial damage. To address these questions, ACE distribution under septic conditions was studied in vitro by treating pulmonary microvascular endothelial cells (HPMEC) and human umbilical vein endothelial cells (HUVEC) with lipopolysaccharide from Escherichia coli (LPS). Primary isolated HUVEC and HPMEC were compared by detecting ACE activity, membrane-bound ACE, as well as shedding and mRNA production of ACE with and without LPS (1 ng/ml-1 μg/ml). ACE mRNA expression was detected by real-time PCR, and shedded ACE was measured in cell culture supernatant by ELISA. Additionally, membrane-bound protein expression was investigated by immunohistochemistry in situ. In septic ARDS, the distribution of ACE protein was significantly reduced in all lung endothelial cells (p<0.001). After stimulation with LPS, cultivated HPMEC showed more markedly than HUVEC, a concentration-dependent reduction of ACE protein expression compared to the respective untreated controls. Real-time PCR demonstrated a reduced ACE mRNA expression after LPS stimulation, predominantly in HPMEC. Specifically, in HPMEC, a concentration-dependent increase of shedded ACE was shown 24 h after LPS treatment. HPMEC cultures are an apt model for the investigation of pulmonary ACE expression in sepsis. This study suggests that reduced pulmonary microvascular endothelial ACE expression in septic ARDS is caused by two processes: (initial) increased shedding of ACE accompanied by a compensatory downregulation of ACE-mRNA and membrane-bound protein expression.