Wissam H Faour1, Hussein Fayyad-Kazan2, Nabil El Zein3. 1. School of Medicine, Lebanese American University, P.O. Box 36, Byblos, Lebanon. 2. Faculty of Sciences, Branch 1, Lebanese University, Rafic Hariri University Campus, Hadath, Lebanon. 3. Faculty of Sciences, Branch 1, Lebanese University, Rafic Hariri University Campus, Hadath, Lebanon. nabilzein@hotmail.fr.
Abstract
OBJECTIVE AND DESIGN: The objective of this study is to uncover the signal transduction pathways of N-formyl methionyl-leucyl-phenylalanine (fMLP) in monocyte. MATERIALS OR SUBJECTS: Freshly isolated human peripheral blood monocytes (PBMC) were used for in vitro assessment of signal transduction pathways activated by fMLP. TREATMENT: Time-course and dose-response experiments were used to evaluate the effect of fMLP along with the specific inhibitors/stimulators on the activation of downstream signaling kinases. METHODS: Freshly isolated human PBMC were stimulated with fMLP for the desired time. Western blot and siRNA analysis were used to evaluate the activated intracellular signaling kinases, and flow analysis was performed to assess the levels of CD11b. Furthermore, luminescence spectrometry was performed to measure the levels of released hydrogen peroxide in the media. RESULTS: fMLP strongly stimulated the activation of AKT and ERK1/2 through a RhoA-GTPase-dependent manner and also induced H2O2 release by monocytes. Furthermore, fMLP mediated its effects through restricted activation of formylpeptide receptor-like 1 (FPRL1/FPR2), but independently of either EGFR transactivation or intracellular calcium release. In addition, NAC reversed fMLP- and H2O2-induced activation of Akt and RhoA-GTPase. CONCLUSION: Collectively, these data suggested that fMLP-activated ERK1/2 and Akt pathways through specific activation of the FPRL1/ROS/RoA-GTPase pathway.
OBJECTIVE AND DESIGN: The objective of this study is to uncover the signal transduction pathways of N-formyl methionyl-leucyl-phenylalanine (fMLP) in monocyte. MATERIALS OR SUBJECTS: Freshly isolated human peripheral blood monocytes (PBMC) were used for in vitro assessment of signal transduction pathways activated by fMLP. TREATMENT: Time-course and dose-response experiments were used to evaluate the effect of fMLP along with the specific inhibitors/stimulators on the activation of downstream signaling kinases. METHODS: Freshly isolated human PBMC were stimulated with fMLP for the desired time. Western blot and siRNA analysis were used to evaluate the activated intracellular signaling kinases, and flow analysis was performed to assess the levels of CD11b. Furthermore, luminescence spectrometry was performed to measure the levels of released hydrogen peroxide in the media. RESULTS:fMLP strongly stimulated the activation of AKT and ERK1/2 through a RhoA-GTPase-dependent manner and also induced H2O2 release by monocytes. Furthermore, fMLP mediated its effects through restricted activation of formylpeptide receptor-like 1 (FPRL1/FPR2), but independently of either EGFR transactivation or intracellular calcium release. In addition, NAC reversed fMLP- and H2O2-induced activation of Akt and RhoA-GTPase. CONCLUSION: Collectively, these data suggested that fMLP-activated ERK1/2 and Akt pathways through specific activation of the FPRL1/ROS/RoA-GTPase pathway.
Authors: B Q Li; M A Wetzel; J A Mikovits; E E Henderson; T J Rogers; W Gong; Y Le; F W Ruscetti; J M Wang Journal: Blood Date: 2001-05-15 Impact factor: 22.113