A humanized anti-Toll like receptor 4 antibody Fab fragment inhibits pro-inflammatory responses induced by lipopolysaccharide through TLR4 in vitro and in vivo
Introduction: Toll like receptor 4 (TLR4) and its co-receptor MD-2 recognize bacterial lipopolysaccharide (LPS), initiating responses to infections caused by Gram-negative bacteria. TLR4 also plays a role in various pathological processes, including viral infections and sterile inflammation. Howeve...
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The Journal of Infection in Developing Countries
2025-06-01
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| Series: | Journal of Infection in Developing Countries |
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| Online Access: | https://www.jidc.org/index.php/journal/article/view/20194 |
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| author | Wenkai Zheng Fang Xie Shuping Si Xi Xiong Jing Xu Chuanxia Yao Cong Li Jin Zhu Ping Li Binggang Cai Maorong Wang |
| author_facet | Wenkai Zheng Fang Xie Shuping Si Xi Xiong Jing Xu Chuanxia Yao Cong Li Jin Zhu Ping Li Binggang Cai Maorong Wang |
| author_sort | Wenkai Zheng |
| collection | DOAJ |
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Introduction: Toll like receptor 4 (TLR4) and its co-receptor MD-2 recognize bacterial lipopolysaccharide (LPS), initiating responses to infections caused by Gram-negative bacteria. TLR4 also plays a role in various pathological processes, including viral infections and sterile inflammation. However, effective methods to inhibit LPS/TLR4-mediated inflammation remain elusive. This study aimed to evaluate the inhibitory effects of a constructed hTLR4-Fab on LPS-induced inflammation in both in vitro and in vivo settings.
Methodology: In vitro, mouse dendritic cells (DCs), human macrophages, and human DCs were incubated with hTLR4-Fab and then stimulated with LPS. In vivo, mice were pre-treated with a humanized anti-TLR4 antibody Fab prior to LPS injection. We examined the activation of various signaling pathways to elucidate the molecular mechanism underlying the inhibition of LPS-induced inflammation by hTLR4-Fab.
Results: We observed that the binding affinity of hTLR4-Fab to TLR4 on mouse bone marrow-derived dendritic cells (DCs) was approximately 81.8%, while the binding affinity to human blood monocyte-derived macrophages and DCs exceeded 90%. Pretreatment with hTLR4-Fab significantly reduced both mRNA and protein levels of LPS-induced proinflammatory cytokines. In vivo, a significant suppression of serum cytokine expression was driven by hTLR4-Fab treatment.
Conclusions: The results demonstrated that the antibody Fab could impede the phosphorylation of downstream components, including the nuclear factor κB (NF-κB) signaling pathway, the mitogen-activated protein kinase (MAPK) signaling pathway, and IFN regulatory factor 3 (IRF-3), all of which are activated by TLR4. Consequently, our study demonstrates that our hTLR4-Fab is effective in mitigating LPS-induced inflammation, both in vitro and in vivo.
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| format | Article |
| id | doaj-art-baeacf5ee425412badb538cd6e297e38 |
| institution | Kabale University |
| issn | 1972-2680 |
| language | English |
| publishDate | 2025-06-01 |
| publisher | The Journal of Infection in Developing Countries |
| record_format | Article |
| series | Journal of Infection in Developing Countries |
| spelling | doaj-art-baeacf5ee425412badb538cd6e297e382025-08-20T03:29:14ZengThe Journal of Infection in Developing CountriesJournal of Infection in Developing Countries1972-26802025-06-01190610.3855/jidc.20194A humanized anti-Toll like receptor 4 antibody Fab fragment inhibits pro-inflammatory responses induced by lipopolysaccharide through TLR4 in vitro and in vivoWenkai Zheng0Fang Xie1Shuping Si2Xi Xiong3Jing Xu4Chuanxia Yao5Cong Li6Jin Zhu7Ping Li8Binggang Cai9Maorong Wang10Department of Liver Diseases of Qinhuai Medical District, Jinling Hospital, Medical School of Nanjing University, Nanjing, Jiangsu, ChinaDepartment of Liver Diseases of Qinhuai Medical District, Jinling Hospital, Medical School of Nanjing University, Nanjing, Jiangsu, ChinaDepartment of Gastroenterology, Wuxi People's Hospital, Wuxi Medical Center, Nanjing Medical University, Wuxi, Jiangsu, ChinaDepartment of Liver Diseases of Qinhuai Medical District, Jinling Hospital, Medical School of Nanjing University, Nanjing, Jiangsu, ChinaDepartment of Liver Diseases of Qinhuai Medical District, Jinling Hospital, Medical School of Nanjing University, Nanjing, Jiangsu, ChinaDepartment of Liver Diseases of Qinhuai Medical District, Jinling Hospital, Medical School of Nanjing University, Nanjing, Jiangsu, ChinaDepartment of Liver Diseases of Qinhuai Medical District, Jinling Hospital, Medical School of Nanjing University, Nanjing, Jiangsu, ChinaResearch Institute for Medicine of Nanjing Command, Nanjing, Jiangsu, ChinaDepartment of Liver Diseases of Qinhuai Medical District, Jinling Hospital, Medical School of Nanjing University, Nanjing, Jiangsu, ChinaYancheng No.1 People's Hospital, Affiliated Hospital of Medical School, Nanjing University; The First People’s Hospital of Yancheng, Yancheng, Jiangsu, ChinaDepartment of Liver Diseases of Qinhuai Medical District, Jinling Hospital, Medical School of Nanjing University, Nanjing, Jiangsu, China Introduction: Toll like receptor 4 (TLR4) and its co-receptor MD-2 recognize bacterial lipopolysaccharide (LPS), initiating responses to infections caused by Gram-negative bacteria. TLR4 also plays a role in various pathological processes, including viral infections and sterile inflammation. However, effective methods to inhibit LPS/TLR4-mediated inflammation remain elusive. This study aimed to evaluate the inhibitory effects of a constructed hTLR4-Fab on LPS-induced inflammation in both in vitro and in vivo settings. Methodology: In vitro, mouse dendritic cells (DCs), human macrophages, and human DCs were incubated with hTLR4-Fab and then stimulated with LPS. In vivo, mice were pre-treated with a humanized anti-TLR4 antibody Fab prior to LPS injection. We examined the activation of various signaling pathways to elucidate the molecular mechanism underlying the inhibition of LPS-induced inflammation by hTLR4-Fab. Results: We observed that the binding affinity of hTLR4-Fab to TLR4 on mouse bone marrow-derived dendritic cells (DCs) was approximately 81.8%, while the binding affinity to human blood monocyte-derived macrophages and DCs exceeded 90%. Pretreatment with hTLR4-Fab significantly reduced both mRNA and protein levels of LPS-induced proinflammatory cytokines. In vivo, a significant suppression of serum cytokine expression was driven by hTLR4-Fab treatment. Conclusions: The results demonstrated that the antibody Fab could impede the phosphorylation of downstream components, including the nuclear factor κB (NF-κB) signaling pathway, the mitogen-activated protein kinase (MAPK) signaling pathway, and IFN regulatory factor 3 (IRF-3), all of which are activated by TLR4. Consequently, our study demonstrates that our hTLR4-Fab is effective in mitigating LPS-induced inflammation, both in vitro and in vivo. https://www.jidc.org/index.php/journal/article/view/20194TLR4 signalingsepsisanti-inflammatory therapy lipopolysaccharidetoll like receptor 4 signalinginflammation |
| spellingShingle | Wenkai Zheng Fang Xie Shuping Si Xi Xiong Jing Xu Chuanxia Yao Cong Li Jin Zhu Ping Li Binggang Cai Maorong Wang A humanized anti-Toll like receptor 4 antibody Fab fragment inhibits pro-inflammatory responses induced by lipopolysaccharide through TLR4 in vitro and in vivo Journal of Infection in Developing Countries TLR4 signaling sepsis anti-inflammatory therapy lipopolysaccharide toll like receptor 4 signaling inflammation |
| title | A humanized anti-Toll like receptor 4 antibody Fab fragment inhibits pro-inflammatory responses induced by lipopolysaccharide through TLR4 in vitro and in vivo |
| title_full | A humanized anti-Toll like receptor 4 antibody Fab fragment inhibits pro-inflammatory responses induced by lipopolysaccharide through TLR4 in vitro and in vivo |
| title_fullStr | A humanized anti-Toll like receptor 4 antibody Fab fragment inhibits pro-inflammatory responses induced by lipopolysaccharide through TLR4 in vitro and in vivo |
| title_full_unstemmed | A humanized anti-Toll like receptor 4 antibody Fab fragment inhibits pro-inflammatory responses induced by lipopolysaccharide through TLR4 in vitro and in vivo |
| title_short | A humanized anti-Toll like receptor 4 antibody Fab fragment inhibits pro-inflammatory responses induced by lipopolysaccharide through TLR4 in vitro and in vivo |
| title_sort | humanized anti toll like receptor 4 antibody fab fragment inhibits pro inflammatory responses induced by lipopolysaccharide through tlr4 in vitro and in vivo |
| topic | TLR4 signaling sepsis anti-inflammatory therapy lipopolysaccharide toll like receptor 4 signaling inflammation |
| url | https://www.jidc.org/index.php/journal/article/view/20194 |
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