Xi Luo1, Xiaohong Zhang2, Tie Zhao1, Tiebing Zeng1, Wen Liu3, Meixia Deng3, Feijun Zhao4. 1. Institute of Pathogenic Biology, Key Laboratory of Special Pathogen Prevention and Control of Hunan Province, University of South China, Hengyang 421001, PR China; Collaborative Innovation Center for New Molecular Drug Research, Hengyang 421001, PR China. 2. Institute of Pathogenic Biology, Key Laboratory of Special Pathogen Prevention and Control of Hunan Province, University of South China, Hengyang 421001, PR China; Department of Histology and Embryology, School of Medicine, University of South China, Hengyang 421001, PR China. 3. Institute of Pathogenic Biology, Key Laboratory of Special Pathogen Prevention and Control of Hunan Province, University of South China, Hengyang 421001, PR China. 4. Institute of Pathogenic Biology, Key Laboratory of Special Pathogen Prevention and Control of Hunan Province, University of South China, Hengyang 421001, PR China; Collaborative Innovation Center for New Molecular Drug Research, Hengyang 421001, PR China. Electronic address: nhdxzhfj@163.com.
Abstract
AIMS: To determine proinflammatory mechanisms of Treponema pallidum outer membrane protein Tp92 in the early syphilis infection in human macrophages and HMEC-1 cells. METHODS: Recombinant Tp92 protein was used to stimulate target human macrophages and HMEC-1 cells. PDTC (Pyrrolidinedithiocarbamic acid), SB202190 and Z-YVAD-FMK were used to block the MyD88/NF-κB, MAPKs/p38 and NLRP3/Caspase-1 pathway, respectively. TNF-α, IL-1β, IL-6, IL-8,NLRP3, casepase-1 were detected by ELISA or Western blot. Lactate dehydrogenase (LDH) activity was measured. RESULTS: Tp92 protein could significantly induced the secretion of proinflammatory cytokines TNF-α, IL-1β, IL-6 and IL-8 in HMEC-1 cells, but not in macrophages except IL-8. When MyD88/NF-κB pathway was blocked, differences in the secretion of TNF-α, IL-6 and IL-1β levels and LDH enzyme activity between Tp92 group and Tp92 + PDTC group were not significant (P > 0.05) in HMEC-1 cells and macrophages except IL-8(P < 0.05). When MAPKs/p38 pathway was blocked, differences in the secretion of TNF-α, IL-1β, IL-6 and IL-8 and LDH enzyme activity both Tp92 group and Tp92 + SB2010190 group were not significant (P > 0.05) in HMEC-1 cells and macrophages. In contrast, when NLRP3/Caspase-1 pathway was blocked with Z-YVAD-FMK, TNF-α, IL-6 and IL-1β levels, LDH enzyme activity, and Caspase-1 and NLRP3 protein levels were significantly declined (P < 0.05) in HMEC-1 cells except IL-8(P > 0.05). The LDH enzyme activity in macrophages was decreased before and after Z-YVAD-FMK blocking (P < 0.05),however, differences in the secretion of TNF-α, IL-1β, IL-6 and IL-8 between Tp92 group and Tp92+Z-YVAD-FMK group in macrophages were not significant (P > 0.05). CONCLUSIONS: Tp92 protein may promote proinflammatory cytokines TNF-α, IL-1β, IL-6 secretion of HMEC-1 cells, but not in macrophages, and increase the LDH enzyme activity of HMEC-1 cells and macrophages through NLRP3/Caspase-1 pathway. However, Tp92 protein may promote IL-8 secretion of HMEC-1 cells and macrophages through MyD88/NF-κB pathway.
AIMS: To determine proinflammatory mechanisms of Treponema pallidum outer membrane protein Tp92 in the early syphilis infection in human macrophages and HMEC-1 cells. METHODS: Recombinant Tp92 protein was used to stimulate target human macrophages and HMEC-1 cells. PDTC (Pyrrolidinedithiocarbamic acid), SB202190 and Z-YVAD-FMK were used to block the MyD88/NF-κB, MAPKs/p38 and NLRP3/Caspase-1 pathway, respectively. TNF-α, IL-1β, IL-6, IL-8,NLRP3, casepase-1 were detected by ELISA or Western blot. Lactate dehydrogenase (LDH) activity was measured. RESULTS: Tp92 protein could significantly induced the secretion of proinflammatory cytokines TNF-α, IL-1β, IL-6 and IL-8 in HMEC-1 cells, but not in macrophages except IL-8. When MyD88/NF-κB pathway was blocked, differences in the secretion of TNF-α, IL-6 and IL-1β levels and LDH enzyme activity between Tp92 group and Tp92 + PDTC group were not significant (P > 0.05) in HMEC-1 cells and macrophages except IL-8(P < 0.05). When MAPKs/p38 pathway was blocked, differences in the secretion of TNF-α, IL-1β, IL-6 and IL-8 and LDH enzyme activity both Tp92 group and Tp92 + SB2010190 group were not significant (P > 0.05) in HMEC-1 cells and macrophages. In contrast, when NLRP3/Caspase-1 pathway was blocked with Z-YVAD-FMK, TNF-α, IL-6 and IL-1β levels, LDH enzyme activity, and Caspase-1 and NLRP3 protein levels were significantly declined (P < 0.05) in HMEC-1 cells except IL-8(P > 0.05). The LDH enzyme activity in macrophages was decreased before and after Z-YVAD-FMK blocking (P < 0.05),however, differences in the secretion of TNF-α, IL-1β, IL-6 and IL-8 between Tp92 group and Tp92+Z-YVAD-FMK group in macrophages were not significant (P > 0.05). CONCLUSIONS: Tp92 protein may promote proinflammatory cytokines TNF-α, IL-1β, IL-6 secretion of HMEC-1 cells, but not in macrophages, and increase the LDH enzyme activity of HMEC-1 cells and macrophages through NLRP3/Caspase-1 pathway. However, Tp92 protein may promote IL-8 secretion of HMEC-1 cells and macrophages through MyD88/NF-κB pathway.