Montserrat Estruch1, Jose Luis Sánchez-Quesada2, Jordi Ordóñez-Llanos3, Sonia Benítez4. 1. Biomedical Research Institute Sant Pau (IIB-Sant Pau), Hospital de la Santa Creu i Sant Pau, Barcelona, Spain; Molecular Biology and Biochemistry Department, Universitat Autònoma de Barcelona (UAB), Barcelona, Spain. Electronic address: mestruch@santpau.cat. 2. Biomedical Research Institute Sant Pau (IIB-Sant Pau), Hospital de la Santa Creu i Sant Pau, Barcelona, Spain; Molecular Biology and Biochemistry Department, Universitat Autònoma de Barcelona (UAB), Barcelona, Spain. 3. Biochemistry Department, Hospital de la Santa Creu i Sant Pau, Barcelona, Spain; Molecular Biology and Biochemistry Department, Universitat Autònoma de Barcelona (UAB), Barcelona, Spain. 4. Biomedical Research Institute Sant Pau (IIB-Sant Pau), Hospital de la Santa Creu i Sant Pau, Barcelona, Spain; Molecular Biology and Biochemistry Department, Universitat Autònoma de Barcelona (UAB), Barcelona, Spain. Electronic address: sbenitez@santpau.cat.
Abstract
INTRODUCTION: In vitro ceramide-enriched LDL (CER-LDL) reproduces most of the properties of electronegative LDL (LDL(-)), a heterogeneous subfraction of LDL found in plasma. LDL(-) comprises several modifications of LDL and has an increased content in ceramide (CER). It promotes cytokine release in monocytes through CD14 and TLR4. CER-LDL also induces cytokine release in these cells but the mechanism is unknown. AIM: To evaluate TLR4 and CD14 as the putative receptors involved in cytokine release induced by CER-LDL. METHODS: CER-LDL was obtained by incubating native LDL with CER-enriched liposomes. CER content in CER-LDL was assessed by thin layer chromatography of lipid extracts. CER-LDL and LDL(-) were incubated for 20 h with human monocytes in the presence or absence of a TLR4 signaling inhibitor. Both LDLs were also incubated with two human monocytic cell lines, normal and THP1 overexpressing CD14 (THP1-CD14) cells. The release of IL-6, IL-10 and MCP-1 was evaluated by ELISA in culture medium. RESULTS: The release of IL-6, IL-10 and MCP-1 induced by CER-LDL in monocytes was inhibited by VIPER (90% inhibition), a specific TLR4 inhibitor. The cytokine release induced by CER-LDL was negligible in THP1, cells presenting a very low CD14 expression. In contrast, the induction of cytokine release in THP1-CD14 was high and dependent on the CER content in LDL. CONCLUSION: CER-LDL induces IL-6, IL-10 and MCP-1 release through the activation of CD14 and TLR4 in monocytes, reproducing the behavior of LDL(-). The increased content of CER in LDL(-) is then related to the inflammatory action of LDL(-).
INTRODUCTION: In vitro ceramide-enriched LDL (CER-LDL) reproduces most of the properties of electronegative LDL (LDL(-)), a heterogeneous subfraction of LDL found in plasma. LDL(-) comprises several modifications of LDL and has an increased content in ceramide (CER). It promotes cytokine release in monocytes through CD14 and TLR4. CER-LDL also induces cytokine release in these cells but the mechanism is unknown. AIM: To evaluate TLR4 and CD14 as the putative receptors involved in cytokine release induced by CER-LDL. METHODS:CER-LDL was obtained by incubating native LDL with CER-enriched liposomes. CER content in CER-LDL was assessed by thin layer chromatography of lipid extracts. CER-LDL and LDL(-) were incubated for 20 h with human monocytes in the presence or absence of a TLR4 signaling inhibitor. Both LDLs were also incubated with two human monocytic cell lines, normal and THP1 overexpressing CD14 (THP1-CD14) cells. The release of IL-6, IL-10 and MCP-1 was evaluated by ELISA in culture medium. RESULTS: The release of IL-6, IL-10 and MCP-1 induced by CER-LDL in monocytes was inhibited by VIPER (90% inhibition), a specific TLR4 inhibitor. The cytokine release induced by CER-LDL was negligible in THP1, cells presenting a very low CD14 expression. In contrast, the induction of cytokine release in THP1-CD14 was high and dependent on the CER content in LDL. CONCLUSION:CER-LDL induces IL-6, IL-10 and MCP-1 release through the activation of CD14 and TLR4 in monocytes, reproducing the behavior of LDL(-). The increased content of CER in LDL(-) is then related to the inflammatory action of LDL(-).
Authors: Núria Puig; Lara Montolio; Pol Camps-Renom; Laia Navarra; Francesc Jiménez-Altayó; Elena Jiménez-Xarrié; Jose Luis Sánchez-Quesada; Sonia Benitez Journal: Cells Date: 2020-03-01 Impact factor: 6.600