Pub Date : 2023-05-01DOI: 10.4049/jimmunol.210.supp.79.17
S. Miaw, W. Chin
Macrophage plays a crucial role in the front line of host defense against pathogens. Classically activated macrophages (M1), induced by IFN-g and LPS, highly express inflammatory cytokines and contribute to inflammatory processes. By contrast, alternatively activated macrophages (M2) are induced by IL-4/IL-13, produce IL-10, and display anti-inflammatory activity. Adenylate kinase 4 (Ak4), an enzyme that transfers phosphate group among ATP/GTP, AMP, and ADP, is a key modulator of ATP. Ak4 is involved in maintaining the homeostasis of cellular nucleotides which is essential for cellular function. We observed Ak4 is preferentially expressed in M1 macrophages compared to M2 macrophages. Whether Ak4 is critical for M1 macrophage function remains elusive. Here we demonstrated that Ak4 maintained ATP homeostasis, and was critical for ROS production, glycolysis, and bactericidal ability in M1 macrophages. Moreover, Ak4 promoted the expression of inflammatory genes, including Il1b, Il6, Tnfa, Nos2, Nox2and Hif1a, in M1 macrophages via Hif1a and AMPK. However, Ak4 deficiency did not affect the development of murine immune cells. Taken together, our data depict a potential mechanism linking nucleotide homeostasis and the function of M1 macrophage. Taiwan Ministry of Science and Technology (111-2320-B-002-068-MY3)
{"title":"Adenylate kinase 4 is critical to the function of classically activated macrophage","authors":"S. Miaw, W. Chin","doi":"10.4049/jimmunol.210.supp.79.17","DOIUrl":"https://doi.org/10.4049/jimmunol.210.supp.79.17","url":null,"abstract":"\u0000 Macrophage plays a crucial role in the front line of host defense against pathogens. Classically activated macrophages (M1), induced by IFN-g and LPS, highly express inflammatory cytokines and contribute to inflammatory processes. By contrast, alternatively activated macrophages (M2) are induced by IL-4/IL-13, produce IL-10, and display anti-inflammatory activity. Adenylate kinase 4 (Ak4), an enzyme that transfers phosphate group among ATP/GTP, AMP, and ADP, is a key modulator of ATP. Ak4 is involved in maintaining the homeostasis of cellular nucleotides which is essential for cellular function. We observed Ak4 is preferentially expressed in M1 macrophages compared to M2 macrophages. Whether Ak4 is critical for M1 macrophage function remains elusive. Here we demonstrated that Ak4 maintained ATP homeostasis, and was critical for ROS production, glycolysis, and bactericidal ability in M1 macrophages. Moreover, Ak4 promoted the expression of inflammatory genes, including Il1b, Il6, Tnfa, Nos2, Nox2and Hif1a, in M1 macrophages via Hif1a and AMPK. However, Ak4 deficiency did not affect the development of murine immune cells. Taken together, our data depict a potential mechanism linking nucleotide homeostasis and the function of M1 macrophage.\u0000 Taiwan Ministry of Science and Technology (111-2320-B-002-068-MY3)","PeriodicalId":22698,"journal":{"name":"The Journal of Immunology","volume":"17 1","pages":""},"PeriodicalIF":0.0,"publicationDate":"2023-05-01","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"81733179","PeriodicalName":null,"FirstCategoryId":null,"ListUrlMain":null,"RegionNum":0,"RegionCategory":"","ArticlePicture":[],"TitleCN":null,"AbstractTextCN":null,"PMCID":"","EPubDate":null,"PubModel":null,"JCR":null,"JCRName":null,"Score":null,"Total":0}
Pub Date : 2023-05-01DOI: 10.4049/jimmunol.210.supp.221.26
Y. Poluektov, M. Delcommenne
The use of MHC tetramers to identify antigen specific T cells has become widespread among immunologists since the late 90s. Tetramers have become so ubiquitous that many companies and research facilities can ship and deliver some of the most commonly used tetramers within a matter of days. But when it comes to the uncommon tetramers or tetramers with antigenic peptides that have only recently been identified, it may be quite a challenge to find the needed tetramer. In this case, most researchers are left with trying to produce their own tetramer or having another facility do it for them, which can become a lengthy process. In an effort to help those researchers, we have developed the QuickSwitch ™MHC tetramer. This MHC tetramer comes loaded with a place-holder peptide (or exiting peptide) that can be exchanged for most well-binding peptides of a given MHC haplotype. This way, researchers are able to make a tetramer with any peptide they have available in a matter of hours. More importantly, our MHC tetramer platform comes with a means to monitor how much of the available peptide has exchanged with the exiting peptide and quantify the MHC occupancy. This allows the user to not only determine if the peptide in question can make a viable MHC tetramer, but also identify the strong and weak binding peptides for that MHC haplotype. This piece of information can be invaluable in future studies, in particular for vaccine development. MBL International
{"title":"QuickSwitch ™: an MHC tetramer platform for making fast and reliable custom tetramers for T cell staining","authors":"Y. Poluektov, M. Delcommenne","doi":"10.4049/jimmunol.210.supp.221.26","DOIUrl":"https://doi.org/10.4049/jimmunol.210.supp.221.26","url":null,"abstract":"\u0000 The use of MHC tetramers to identify antigen specific T cells has become widespread among immunologists since the late 90s. Tetramers have become so ubiquitous that many companies and research facilities can ship and deliver some of the most commonly used tetramers within a matter of days. But when it comes to the uncommon tetramers or tetramers with antigenic peptides that have only recently been identified, it may be quite a challenge to find the needed tetramer. In this case, most researchers are left with trying to produce their own tetramer or having another facility do it for them, which can become a lengthy process. In an effort to help those researchers, we have developed the QuickSwitch ™MHC tetramer. This MHC tetramer comes loaded with a place-holder peptide (or exiting peptide) that can be exchanged for most well-binding peptides of a given MHC haplotype. This way, researchers are able to make a tetramer with any peptide they have available in a matter of hours. More importantly, our MHC tetramer platform comes with a means to monitor how much of the available peptide has exchanged with the exiting peptide and quantify the MHC occupancy. This allows the user to not only determine if the peptide in question can make a viable MHC tetramer, but also identify the strong and weak binding peptides for that MHC haplotype. This piece of information can be invaluable in future studies, in particular for vaccine development.\u0000 MBL International","PeriodicalId":22698,"journal":{"name":"The Journal of Immunology","volume":"30 1","pages":""},"PeriodicalIF":0.0,"publicationDate":"2023-05-01","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"81870610","PeriodicalName":null,"FirstCategoryId":null,"ListUrlMain":null,"RegionNum":0,"RegionCategory":"","ArticlePicture":[],"TitleCN":null,"AbstractTextCN":null,"PMCID":"","EPubDate":null,"PubModel":null,"JCR":null,"JCRName":null,"Score":null,"Total":0}
Pub Date : 2023-05-01DOI: 10.4049/jimmunol.210.supp.173.10
Young S. Lee, V. Saxena, Wenji Piao, Lushen Li, Long Wu, Marina Willsonshirkey, Allison Kensiski, Samuel J. Gavzy, Bing Ma, J. Bromberg
Regulatory T cell (Treg) lymphatic migration and maintenance of transcription factor Foxp3 expression are required for suppressor function and allograft protection, and migrated Tregs express ectonucleotidase CD39 hi. Purine metabolism is implicated in Treg phenotype but the precise functions of purine metabolites on Treg function and stability have remained unclear. Sorted Foxp3+ Tregs were used to test Treg stability, migration, and cellular viability in a transendothelial migration (TEM) in vitro model. Intracellular metabolome of Tregs was analyzed by mass spectrometry, and differentially expressed metabolites between Tregs and exTregs were identified and tested. Foxp3 hiTregs displayed higher suppressive function and migration across LECs compared to Foxp3 loexTregs, but cellular viability was similar. Tregs showed differential metabolic profiles compared to exTregs, and the top 16 most differentially expressed metabolites involved the tricarboxylic acid cycle, polyamines, and purine metabolism. Ten metabolites including nicotinamide and inosine monophosphate (IMP) were downregulated, while six metabolites including putrescine, cadaverine, and N-acetylglycine were upregulated in Tregs compared to exTregs. A purine ectonucleotidase CD73 inhibitor decreased Treg conversion to exTregs and adenosine increased Treg migration and suppressive function concomitant with decreased exTreg conversion. Polyamine metabolites showed only subtle effects on exTreg conversion and migration. These results suggest that purine metabolism is a potent regulator that maintains Treg Foxp3 expression and suppressive function during Treg TEM and thus, could be a promising target for therapeutic interventions. NIH grant RO1 A1062765
{"title":"Purine metabolic pathway regulates regulatory T cell stability and function","authors":"Young S. Lee, V. Saxena, Wenji Piao, Lushen Li, Long Wu, Marina Willsonshirkey, Allison Kensiski, Samuel J. Gavzy, Bing Ma, J. Bromberg","doi":"10.4049/jimmunol.210.supp.173.10","DOIUrl":"https://doi.org/10.4049/jimmunol.210.supp.173.10","url":null,"abstract":"\u0000 Regulatory T cell (Treg) lymphatic migration and maintenance of transcription factor Foxp3 expression are required for suppressor function and allograft protection, and migrated Tregs express ectonucleotidase CD39 hi. Purine metabolism is implicated in Treg phenotype but the precise functions of purine metabolites on Treg function and stability have remained unclear.\u0000 Sorted Foxp3+ Tregs were used to test Treg stability, migration, and cellular viability in a transendothelial migration (TEM) in vitro model. Intracellular metabolome of Tregs was analyzed by mass spectrometry, and differentially expressed metabolites between Tregs and exTregs were identified and tested.\u0000 Foxp3 hiTregs displayed higher suppressive function and migration across LECs compared to Foxp3 loexTregs, but cellular viability was similar. Tregs showed differential metabolic profiles compared to exTregs, and the top 16 most differentially expressed metabolites involved the tricarboxylic acid cycle, polyamines, and purine metabolism. Ten metabolites including nicotinamide and inosine monophosphate (IMP) were downregulated, while six metabolites including putrescine, cadaverine, and N-acetylglycine were upregulated in Tregs compared to exTregs. A purine ectonucleotidase CD73 inhibitor decreased Treg conversion to exTregs and adenosine increased Treg migration and suppressive function concomitant with decreased exTreg conversion. Polyamine metabolites showed only subtle effects on exTreg conversion and migration.\u0000 These results suggest that purine metabolism is a potent regulator that maintains Treg Foxp3 expression and suppressive function during Treg TEM and thus, could be a promising target for therapeutic interventions.\u0000 NIH grant RO1 A1062765","PeriodicalId":22698,"journal":{"name":"The Journal of Immunology","volume":"35 1","pages":""},"PeriodicalIF":0.0,"publicationDate":"2023-05-01","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"84420079","PeriodicalName":null,"FirstCategoryId":null,"ListUrlMain":null,"RegionNum":0,"RegionCategory":"","ArticlePicture":[],"TitleCN":null,"AbstractTextCN":null,"PMCID":"","EPubDate":null,"PubModel":null,"JCR":null,"JCRName":null,"Score":null,"Total":0}
Pub Date : 2023-05-01DOI: 10.4049/jimmunol.210.supp.88.12
Brandon Lee Maniaci, D. J. Friedman, S. Crotts, Matthew J. Rajcula, Brady Hammer, Elissa Mai, V. Shapiro
The lifetime risk for colorectal cancer in the United States is approximately 4%. Individuals with Inflammatory Bowel Disease, including Ulcerative Colitis and Crohn’s Disease, have a substantially increased risk of developing colorectal cancer. The mechanisms for accelerated tumorigenesis due to enhanced inflammation are not fully characterized. Siglecs (sialic acid immunoglobulin lectin-like proteins) are a family of inhibitory receptors that are negative regulators of the immune response. Siglec-E is an inhibitory receptor that is expressed by innate immune cells, including monocytes, macrophages, neutrophils and dendritic cells. Interestingly, mice deficient in Siglec-E have accelerated development of tumors and reduced survival in a spontaneous mouse model of colorectal cancer (TS4-cre LSL-KRas G12DAPClox 468/wt). While tumors develop at approximately six months in mice with Siglec-E, tumors develop at approximately two months in Siglec-E knockout mice. Initial results indicate that Siglec-E knockout mice also have accelerated gut inflammation using the DSS colitis model as compared to WT mice. Current studies are examining inflammation that develops during tumorigenesis in Siglec-E knockout mice. Center for Biomedical Discovery NIH R01 CA243545-01A1 to VSS Mayo Clinic Graduate School of Biomedical Sciences Deans fellowship: Initiative for maximizing student development (IMSD) R25 GM055252-26 Doyon Foundation
{"title":"Accelerated tumorigenesis in a colorectal cancer model in Siglec-E knockout mice","authors":"Brandon Lee Maniaci, D. J. Friedman, S. Crotts, Matthew J. Rajcula, Brady Hammer, Elissa Mai, V. Shapiro","doi":"10.4049/jimmunol.210.supp.88.12","DOIUrl":"https://doi.org/10.4049/jimmunol.210.supp.88.12","url":null,"abstract":"\u0000 The lifetime risk for colorectal cancer in the United States is approximately 4%. Individuals with Inflammatory Bowel Disease, including Ulcerative Colitis and Crohn’s Disease, have a substantially increased risk of developing colorectal cancer. The mechanisms for accelerated tumorigenesis due to enhanced inflammation are not fully characterized. Siglecs (sialic acid immunoglobulin lectin-like proteins) are a family of inhibitory receptors that are negative regulators of the immune response. Siglec-E is an inhibitory receptor that is expressed by innate immune cells, including monocytes, macrophages, neutrophils and dendritic cells. Interestingly, mice deficient in Siglec-E have accelerated development of tumors and reduced survival in a spontaneous mouse model of colorectal cancer (TS4-cre LSL-KRas G12DAPClox 468/wt). While tumors develop at approximately six months in mice with Siglec-E, tumors develop at approximately two months in Siglec-E knockout mice. Initial results indicate that Siglec-E knockout mice also have accelerated gut inflammation using the DSS colitis model as compared to WT mice. Current studies are examining inflammation that develops during tumorigenesis in Siglec-E knockout mice.\u0000 Center for Biomedical Discovery NIH R01 CA243545-01A1 to VSS Mayo Clinic Graduate School of Biomedical Sciences Deans fellowship: Initiative for maximizing student development (IMSD) R25 GM055252-26 Doyon Foundation","PeriodicalId":22698,"journal":{"name":"The Journal of Immunology","volume":"59 1","pages":""},"PeriodicalIF":0.0,"publicationDate":"2023-05-01","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"84472140","PeriodicalName":null,"FirstCategoryId":null,"ListUrlMain":null,"RegionNum":0,"RegionCategory":"","ArticlePicture":[],"TitleCN":null,"AbstractTextCN":null,"PMCID":"","EPubDate":null,"PubModel":null,"JCR":null,"JCRName":null,"Score":null,"Total":0}
Pub Date : 2023-05-01DOI: 10.4049/jimmunol.210.supp.141.08
Pinghan Huang, Hsiao-Han Tsai, Vivian Y Tat, Jason C. Hsu, Drelich Aleksandra, Hui-Wen Chen, C. J. Hu, C. Tseng
After the outbreak of COVID-19, cellular immunity has been shown to provide protection especially in breakthrough infection and B cell-deficient patients. Furthermore, durable cellular immunity targets conserved epitopes among SARS-CoV-2 variants or different coronavirus then inspiring an attractive strategy for vaccine development. However, how to simultaneously induce balanced T cell responses against multiple epitopes still needs to be elucidated. In this study, we sought to design a T cell-based vaccine that can adjust valency against different T cell epitopes. We selected seven SARS-CoV-2 specific short peptides as antigens, including I-Ab/H-2Kb/H-2Db restricted epitopes. Furthermore, we used PLGA nanoparticle as vaccine platform to achieve the precise loading of the indicated peptides. Mice were immunized with different combinations of peptides and splenocytes were used to access epitope specific-IFN gamma expressing CD8+ or CD4+ T cell responses. First, we confirmed the robust immunogenicity of each peptide. We then combined an equal amount of indicated epitope peptides and accessed the immunogenicity of each epitope in the combination. The results showed peptides compete with each other to induce T cell responses regardless of MHC allele and MHC class. In addition, dominant epitopes significantly suppressed the immunogenicity of the subdominant epitopes. Finally, by optimizing the antigen dose, nanoparticle vaccine induced more balanced T responses against all seven epitopes. Taking advantage of nanoparticle, our results reveal a viable strategy to induce multivalent T cell response. Furthermore, the result suggests that competition of T cell epitopes is one of the factors controlling cellular immunity.
{"title":"Induction of multivalent and balanced T cell responses by modular nanoshell vaccine technology","authors":"Pinghan Huang, Hsiao-Han Tsai, Vivian Y Tat, Jason C. Hsu, Drelich Aleksandra, Hui-Wen Chen, C. J. Hu, C. Tseng","doi":"10.4049/jimmunol.210.supp.141.08","DOIUrl":"https://doi.org/10.4049/jimmunol.210.supp.141.08","url":null,"abstract":"\u0000 After the outbreak of COVID-19, cellular immunity has been shown to provide protection especially in breakthrough infection and B cell-deficient patients. Furthermore, durable cellular immunity targets conserved epitopes among SARS-CoV-2 variants or different coronavirus then inspiring an attractive strategy for vaccine development. However, how to simultaneously induce balanced T cell responses against multiple epitopes still needs to be elucidated. In this study, we sought to design a T cell-based vaccine that can adjust valency against different T cell epitopes. We selected seven SARS-CoV-2 specific short peptides as antigens, including I-Ab/H-2Kb/H-2Db restricted epitopes. Furthermore, we used PLGA nanoparticle as vaccine platform to achieve the precise loading of the indicated peptides. Mice were immunized with different combinations of peptides and splenocytes were used to access epitope specific-IFN gamma expressing CD8+ or CD4+ T cell responses. First, we confirmed the robust immunogenicity of each peptide. We then combined an equal amount of indicated epitope peptides and accessed the immunogenicity of each epitope in the combination. The results showed peptides compete with each other to induce T cell responses regardless of MHC allele and MHC class. In addition, dominant epitopes significantly suppressed the immunogenicity of the subdominant epitopes. Finally, by optimizing the antigen dose, nanoparticle vaccine induced more balanced T responses against all seven epitopes. Taking advantage of nanoparticle, our results reveal a viable strategy to induce multivalent T cell response. Furthermore, the result suggests that competition of T cell epitopes is one of the factors controlling cellular immunity.","PeriodicalId":22698,"journal":{"name":"The Journal of Immunology","volume":"48 1","pages":""},"PeriodicalIF":0.0,"publicationDate":"2023-05-01","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"84486616","PeriodicalName":null,"FirstCategoryId":null,"ListUrlMain":null,"RegionNum":0,"RegionCategory":"","ArticlePicture":[],"TitleCN":null,"AbstractTextCN":null,"PMCID":"","EPubDate":null,"PubModel":null,"JCR":null,"JCRName":null,"Score":null,"Total":0}
Pub Date : 2023-05-01DOI: 10.4049/jimmunol.210.supp.67.16
Kun He, Zhongli Xu, W. MacDonald, A. Ray, Wei Chen, B. Lambrecht, Amanda C. Poholek
Allergic asthma remains a significant health burden for both children and adults. CD4 +Th2 cells are critical drivers of disease, yet the mechanisms that support initiation of the Th2 cell response to environmental allergens are not well understood. We demonstrated a distinct requirement for the transcriptional repressor Blimp-1 to promote Th2 cells in the lung to inhaled but not systemically or subcutaneously delivered allergens. Using temporal, spatial and single cell transcriptomic tracking of house dust mite (HDM) specific T cell responses, we demonstrate that inhalation of HDM drove early Blimp-1 expression necessary for GATA3 upregulation and subsequent Th2 differentiation in the lung that coincides with IL2Rα expression. Blimp-1 expression remains confined to HDM-specific Treg and Th2 cells that traffic to the lung but importantly is dispensable for Th2 cell maintenance. We found that inhaled allergens induce a pattern of STAT activation with transient pSTAT5 concomitant with sustained pSTAT3 within GATA3 +allergen-specific T cells, which is critical for the induction of Blimp-1 during the earliest phase of T cell priming in the lymph node. IL2Rα cooperates with IL10Rα signaling acting directly on allergen specific T cells to drive Th2 cell differentiation. Furthermore, IL-10 derived from allergen specific T cells was sufficient to induce Th2 cells suggesting an autocrine or paracrine loop of IL-10 supports Blimp-1 to regulate GATA3 upregulation at the T-B border and subsequent Th2 differentiation. These data shed light on the steps initiating Th2 responses to inhaled allergens and identify an unexpected pro-inflammatory requirement for IL-10 and Blimp-1 driving inflammatory T cell responses to environmental antigens.
{"title":"Autocrine pro-inflammatory IL-10 initiates lung-specific Th2 responses to inhaled allergen to induce allergic asthma","authors":"Kun He, Zhongli Xu, W. MacDonald, A. Ray, Wei Chen, B. Lambrecht, Amanda C. Poholek","doi":"10.4049/jimmunol.210.supp.67.16","DOIUrl":"https://doi.org/10.4049/jimmunol.210.supp.67.16","url":null,"abstract":"\u0000 Allergic asthma remains a significant health burden for both children and adults. CD4 +Th2 cells are critical drivers of disease, yet the mechanisms that support initiation of the Th2 cell response to environmental allergens are not well understood. We demonstrated a distinct requirement for the transcriptional repressor Blimp-1 to promote Th2 cells in the lung to inhaled but not systemically or subcutaneously delivered allergens. Using temporal, spatial and single cell transcriptomic tracking of house dust mite (HDM) specific T cell responses, we demonstrate that inhalation of HDM drove early Blimp-1 expression necessary for GATA3 upregulation and subsequent Th2 differentiation in the lung that coincides with IL2Rα expression. Blimp-1 expression remains confined to HDM-specific Treg and Th2 cells that traffic to the lung but importantly is dispensable for Th2 cell maintenance. We found that inhaled allergens induce a pattern of STAT activation with transient pSTAT5 concomitant with sustained pSTAT3 within GATA3 +allergen-specific T cells, which is critical for the induction of Blimp-1 during the earliest phase of T cell priming in the lymph node. IL2Rα cooperates with IL10Rα signaling acting directly on allergen specific T cells to drive Th2 cell differentiation. Furthermore, IL-10 derived from allergen specific T cells was sufficient to induce Th2 cells suggesting an autocrine or paracrine loop of IL-10 supports Blimp-1 to regulate GATA3 upregulation at the T-B border and subsequent Th2 differentiation. These data shed light on the steps initiating Th2 responses to inhaled allergens and identify an unexpected pro-inflammatory requirement for IL-10 and Blimp-1 driving inflammatory T cell responses to environmental antigens.","PeriodicalId":22698,"journal":{"name":"The Journal of Immunology","volume":"59 1","pages":""},"PeriodicalIF":0.0,"publicationDate":"2023-05-01","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"84873071","PeriodicalName":null,"FirstCategoryId":null,"ListUrlMain":null,"RegionNum":0,"RegionCategory":"","ArticlePicture":[],"TitleCN":null,"AbstractTextCN":null,"PMCID":"","EPubDate":null,"PubModel":null,"JCR":null,"JCRName":null,"Score":null,"Total":0}
Pub Date : 2023-05-01DOI: 10.4049/jimmunol.210.supp.249.08
Lisa R. Volpatti, Joseph W. Reda, Gustavo Borjas, Zhengjie Zhou, Yun Fang, J. Hubbell
Chronic inflammatory diseases are often treated with corticosteroids or TNF blockers to suppress overactive immune responses. However, these immunosuppressants also dampen healthy immune responses to opportunistic pathogens (e.g., respiratory viruses) and are associated with adverse effects. Targeted immunotherapies are needed to provide local immunosuppression without systemic effects. In the context of atherosclerosis, the anti-inflammatory cytokine IL-10 has been shown to suppress vascular inflammation, but its poor pharmacokinetic profile and pleiotropic effects have limited its therapeutic potential. To overcome these challenges, we engineered IL-10 to specifically target atherosclerotic plaques. We accomplished this goal by constructing fusion proteins in which one side is IL-10 and the other side is an antibody fragment (Fab) that binds to protein epitopes of low-density lipoprotein (LDL). In murine models of atherosclerosis, we show that systemically administered Fab-IL-10 constructs bind circulating LDL and hitchhike a ride to atherosclerotic plaques. In a biodistribution study, we observe elevated levels of IL-10 in the aorta but not the lungs of hyperlipidemic mice, indicative of targeted delivery. The targeted Fab-IL-10 constructs significantly reduce aortic immune cell infiltration to levels comparable to healthy mice, whereas non-targeted IL-10 has no therapeutic effect. Mechanistically, we demonstrate that Fab-IL-10 constructs are preferentially taken up by macrophages to exert an anti-inflammatory effect. This platform technology can be applied to a variety of cytokines and shows promise as a potential targeted anti-inflammatory therapy. Supported by the Chicago Immunoengineering Innovation Center, the NIH National Heart, Lung, and Blood Institute, and the American Heart Association.
{"title":"Engineered cytokines target atherosclerotic plaques and locally suppress inflammation","authors":"Lisa R. Volpatti, Joseph W. Reda, Gustavo Borjas, Zhengjie Zhou, Yun Fang, J. Hubbell","doi":"10.4049/jimmunol.210.supp.249.08","DOIUrl":"https://doi.org/10.4049/jimmunol.210.supp.249.08","url":null,"abstract":"\u0000 Chronic inflammatory diseases are often treated with corticosteroids or TNF blockers to suppress overactive immune responses. However, these immunosuppressants also dampen healthy immune responses to opportunistic pathogens (e.g., respiratory viruses) and are associated with adverse effects. Targeted immunotherapies are needed to provide local immunosuppression without systemic effects. In the context of atherosclerosis, the anti-inflammatory cytokine IL-10 has been shown to suppress vascular inflammation, but its poor pharmacokinetic profile and pleiotropic effects have limited its therapeutic potential. To overcome these challenges, we engineered IL-10 to specifically target atherosclerotic plaques. We accomplished this goal by constructing fusion proteins in which one side is IL-10 and the other side is an antibody fragment (Fab) that binds to protein epitopes of low-density lipoprotein (LDL). In murine models of atherosclerosis, we show that systemically administered Fab-IL-10 constructs bind circulating LDL and hitchhike a ride to atherosclerotic plaques. In a biodistribution study, we observe elevated levels of IL-10 in the aorta but not the lungs of hyperlipidemic mice, indicative of targeted delivery. The targeted Fab-IL-10 constructs significantly reduce aortic immune cell infiltration to levels comparable to healthy mice, whereas non-targeted IL-10 has no therapeutic effect. Mechanistically, we demonstrate that Fab-IL-10 constructs are preferentially taken up by macrophages to exert an anti-inflammatory effect. This platform technology can be applied to a variety of cytokines and shows promise as a potential targeted anti-inflammatory therapy.\u0000 Supported by the Chicago Immunoengineering Innovation Center, the NIH National Heart, Lung, and Blood Institute, and the American Heart Association.","PeriodicalId":22698,"journal":{"name":"The Journal of Immunology","volume":"1 1","pages":""},"PeriodicalIF":0.0,"publicationDate":"2023-05-01","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"84887505","PeriodicalName":null,"FirstCategoryId":null,"ListUrlMain":null,"RegionNum":0,"RegionCategory":"","ArticlePicture":[],"TitleCN":null,"AbstractTextCN":null,"PMCID":"","EPubDate":null,"PubModel":null,"JCR":null,"JCRName":null,"Score":null,"Total":0}
Pub Date : 2023-05-01DOI: 10.4049/jimmunol.210.supp.153.04
M. Pandey, Vyoma Snehal Trivedi, A. Magnusen, R. Rani, Christopher Woods, B. Dipasquale
Sandhoff disease (SD) is an ultra-rare lysosomal storage disorder (LSD), which affects ~ 1/1000, 000 live birth. SD is caused by genetic deficiency of beta (β) Hexosaminidase and resulting excess central nervous system (CNS) synthesis of GM2 ganglioside (GM2) and its impact on neuron death. The exact mechanisms underlying such GM2-driven neuron death are unknown in SD. Glucosylceramide (GC) induced complement 5a (C5a) and its C5aR1 activation causes tissue inflammation in experimental and clinical Gaucher disease. Additionally, C5a-C5aR1 axis was found to sparks CNS inflammation and neurodegeneration in several brain diseases including, intracerebral hemorrhage, traumatic brain injury, myasthenia gravis, amyotrophic lateral sclerosis, neuromyelitis optica spectrum, Alzheimer, and Huntington’s diseases. Here, we determined the increased brain levels of C5a and C5aR1 in a beta (β) hexosaminidase inhibitor (HABI)-induced experimental mouse model of SD, when compared to vehicle (PBS) injected WT control mice. Also, HABI-induced experimental mouse model of SD showed increased microglial cells activation, massive brain generation of pro-inflammatory cytokines and loss of neurons, when compared to vehicle injected WT mice. To assess the relevance of C5a-C5aR1 axis activation for brain inflammation in SD, we targeted β hexosaminidase with HAB-I in WT and C5aR1 −/−mice. Strikingly, as compared to HABI-injected WT mice, HABI-injected C5aR1 −/−mice caused marked reduction in microglial cell activation, brain production of pro-inflammatory cytokines, and the loss of neurons. These data suggest that the C5a-C5aR1 axis is a critical driver of neuroinflammation and neurodegeneration in SD. No Funding
{"title":"Complement Turning Violent in Ultra-Rare Genetic Disorder","authors":"M. Pandey, Vyoma Snehal Trivedi, A. Magnusen, R. Rani, Christopher Woods, B. Dipasquale","doi":"10.4049/jimmunol.210.supp.153.04","DOIUrl":"https://doi.org/10.4049/jimmunol.210.supp.153.04","url":null,"abstract":"\u0000 Sandhoff disease (SD) is an ultra-rare lysosomal storage disorder (LSD), which affects ~ 1/1000, 000 live birth. SD is caused by genetic deficiency of beta (β) Hexosaminidase and resulting excess central nervous system (CNS) synthesis of GM2 ganglioside (GM2) and its impact on neuron death. The exact mechanisms underlying such GM2-driven neuron death are unknown in SD. Glucosylceramide (GC) induced complement 5a (C5a) and its C5aR1 activation causes tissue inflammation in experimental and clinical Gaucher disease. Additionally, C5a-C5aR1 axis was found to sparks CNS inflammation and neurodegeneration in several brain diseases including, intracerebral hemorrhage, traumatic brain injury, myasthenia gravis, amyotrophic lateral sclerosis, neuromyelitis optica spectrum, Alzheimer, and Huntington’s diseases. Here, we determined the increased brain levels of C5a and C5aR1 in a beta (β) hexosaminidase inhibitor (HABI)-induced experimental mouse model of SD, when compared to vehicle (PBS) injected WT control mice. Also, HABI-induced experimental mouse model of SD showed increased microglial cells activation, massive brain generation of pro-inflammatory cytokines and loss of neurons, when compared to vehicle injected WT mice. To assess the relevance of C5a-C5aR1 axis activation for brain inflammation in SD, we targeted β hexosaminidase with HAB-I in WT and C5aR1 −/−mice. Strikingly, as compared to HABI-injected WT mice, HABI-injected C5aR1 −/−mice caused marked reduction in microglial cell activation, brain production of pro-inflammatory cytokines, and the loss of neurons. These data suggest that the C5a-C5aR1 axis is a critical driver of neuroinflammation and neurodegeneration in SD.\u0000 No Funding","PeriodicalId":22698,"journal":{"name":"The Journal of Immunology","volume":"3 1","pages":""},"PeriodicalIF":0.0,"publicationDate":"2023-05-01","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"84958647","PeriodicalName":null,"FirstCategoryId":null,"ListUrlMain":null,"RegionNum":0,"RegionCategory":"","ArticlePicture":[],"TitleCN":null,"AbstractTextCN":null,"PMCID":"","EPubDate":null,"PubModel":null,"JCR":null,"JCRName":null,"Score":null,"Total":0}
Pub Date : 2023-05-01DOI: 10.4049/jimmunol.210.supp.76.16
C. Elkins, Pulavendran Sivasami, Jennifer Bae, Chaoran Li
A unique population of regulatory T cells (Tregs), characterized by a clonally expanded TCR repertoire and distinct transcriptional profile, is highly enriched in the visceral adipose tissue (VAT) at steady state but is lost during obesity, which exacerbates VAT inflammation and promotes metabolic disease. Therefore, understanding the factors that control the accumulation of VAT Tregs, which might be dysregulated during obesity, is important for developing novel therapies for obesity-associated metabolic diseases. Here we show that cellular cholesterol homeostasis is particularly important for VAT Treg accumulation and function. First, bulk RNA sequencing analysis of VAT Tregs from mice fed long-term high fat diet (HFD) showed a reduction in gene expression involved in cholesterol homeostasis (CH), corresponding to loss of VAT Tregs in vivo. Second, we found that VAT Tregs showed increased cholesterol-associated gene expression, cholesterol levels, and cholesterol uptake compared to lymphoid Tregs at steady state, suggesting this pathway is particularly important for VAT Tregs. Third, CRISPR-Cas9-mediated ablation of Srebf2, the master regulator of cholesterol homeostasis, followed by adoptive Treg transfer or Treg-specific germline deletion of Srebf2(Foxp3cre Srebf2-flox) reduced Tregs in the VAT, but not those in other tissues. Deletion of Srebf2in Tregs also increased VAT inflammatory cytokine expression, inflammatory cell infiltration, and insulin resistance in mice following short-term HFD feeding. These data highlight an importance of cellular CH for VAT Treg accumulation and function and implicate this pathway as a potential therapeutic target for treatment of obesity-associated metabolic diseases. NIH R01 (5R01DK128061-02)
{"title":"Cellular cholesterol homeostasis supports visceral adipose tissue (VAT) regulatory T cell (Treg) accumulation and promotes metabolic health","authors":"C. Elkins, Pulavendran Sivasami, Jennifer Bae, Chaoran Li","doi":"10.4049/jimmunol.210.supp.76.16","DOIUrl":"https://doi.org/10.4049/jimmunol.210.supp.76.16","url":null,"abstract":"\u0000 A unique population of regulatory T cells (Tregs), characterized by a clonally expanded TCR repertoire and distinct transcriptional profile, is highly enriched in the visceral adipose tissue (VAT) at steady state but is lost during obesity, which exacerbates VAT inflammation and promotes metabolic disease. Therefore, understanding the factors that control the accumulation of VAT Tregs, which might be dysregulated during obesity, is important for developing novel therapies for obesity-associated metabolic diseases. Here we show that cellular cholesterol homeostasis is particularly important for VAT Treg accumulation and function. First, bulk RNA sequencing analysis of VAT Tregs from mice fed long-term high fat diet (HFD) showed a reduction in gene expression involved in cholesterol homeostasis (CH), corresponding to loss of VAT Tregs in vivo. Second, we found that VAT Tregs showed increased cholesterol-associated gene expression, cholesterol levels, and cholesterol uptake compared to lymphoid Tregs at steady state, suggesting this pathway is particularly important for VAT Tregs. Third, CRISPR-Cas9-mediated ablation of Srebf2, the master regulator of cholesterol homeostasis, followed by adoptive Treg transfer or Treg-specific germline deletion of Srebf2(Foxp3cre Srebf2-flox) reduced Tregs in the VAT, but not those in other tissues. Deletion of Srebf2in Tregs also increased VAT inflammatory cytokine expression, inflammatory cell infiltration, and insulin resistance in mice following short-term HFD feeding. These data highlight an importance of cellular CH for VAT Treg accumulation and function and implicate this pathway as a potential therapeutic target for treatment of obesity-associated metabolic diseases.\u0000 NIH R01 (5R01DK128061-02)","PeriodicalId":22698,"journal":{"name":"The Journal of Immunology","volume":"26 1","pages":""},"PeriodicalIF":0.0,"publicationDate":"2023-05-01","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"84989747","PeriodicalName":null,"FirstCategoryId":null,"ListUrlMain":null,"RegionNum":0,"RegionCategory":"","ArticlePicture":[],"TitleCN":null,"AbstractTextCN":null,"PMCID":"","EPubDate":null,"PubModel":null,"JCR":null,"JCRName":null,"Score":null,"Total":0}
Pub Date : 2023-05-01DOI: 10.4049/jimmunol.210.supp.141.06
Ravindi Dissanayake, S. Nada, Radhika Thanvi, C. O. Sebilleau, E. Prestwich, S. Sucheck, K. Wall
Pseudomonas aeruginosa is an opportunistic pathogen associated with an increased risk of morbidity and mortality among cystic fibrosis patients and immunocompromised individuals. Current antibiotics in the market are not successful against P. aeruginosa infections due to the pathogen’s ability to cause antibiotic resistance. Therefore, our prime objective is to formulate a liposomal-based anti-Pseudomonas vaccine that can produce long-lasting IgG antibodies and T cell memory. The vaccine we introduced accommodates several B cell epitopes of the outer-membrane protein F (OprF) of P. aeruginosa. The recombinant OprF antigen was conjugated with a Toll-like receptor 2/1 (TH1/TH2) agonist, Pam 3CysSK 4, and formulated into dipalmitoylphosphatidylcholine (DPPC)/cholesterol (Chol) liposomes. Our results show that the resulting vaccine conjugate can successfully produce high anti-OprF antibody titers in mice. These antibodies can also recognize intact P. aeruginosa bacteria and kill them with the support of rabbit complement and murine macrophage RAW264.7 cells. To further enhance the vaccine efficacy, the liposome was optimized with 2-dimyristoyl-sn-glycerol-3-phosphocholine (DMPC), 1,2-dimyristoyl-sn-glycerol-3-phospho-(1′-rac-glycerol) (DMPG), Chol, Pam 3CysSK 4and Quillaja Saponaria-derived saponin adjuvant QS21. The mice immunized with the modified vaccine elicit higher antibody titers and increased IgG 2a antibodies. Overall, our liposomal vaccines were found to be highly effective at generating a balanced and robust TH1/TH2 response. Supported by grants from NIH (R01AI148570)
{"title":"Testing of a novel liposomal anti-Pseudomonasvaccine based on outer-membrane protein F (OprF) of Pseudomonas aeruginosa","authors":"Ravindi Dissanayake, S. Nada, Radhika Thanvi, C. O. Sebilleau, E. Prestwich, S. Sucheck, K. Wall","doi":"10.4049/jimmunol.210.supp.141.06","DOIUrl":"https://doi.org/10.4049/jimmunol.210.supp.141.06","url":null,"abstract":"\u0000 Pseudomonas aeruginosa is an opportunistic pathogen associated with an increased risk of morbidity and mortality among cystic fibrosis patients and immunocompromised individuals. Current antibiotics in the market are not successful against P. aeruginosa infections due to the pathogen’s ability to cause antibiotic resistance. Therefore, our prime objective is to formulate a liposomal-based anti-Pseudomonas vaccine that can produce long-lasting IgG antibodies and T cell memory. The vaccine we introduced accommodates several B cell epitopes of the outer-membrane protein F (OprF) of P. aeruginosa. The recombinant OprF antigen was conjugated with a Toll-like receptor 2/1 (TH1/TH2) agonist, Pam 3CysSK 4, and formulated into dipalmitoylphosphatidylcholine (DPPC)/cholesterol (Chol) liposomes. Our results show that the resulting vaccine conjugate can successfully produce high anti-OprF antibody titers in mice. These antibodies can also recognize intact P. aeruginosa bacteria and kill them with the support of rabbit complement and murine macrophage RAW264.7 cells. To further enhance the vaccine efficacy, the liposome was optimized with 2-dimyristoyl-sn-glycerol-3-phosphocholine (DMPC), 1,2-dimyristoyl-sn-glycerol-3-phospho-(1′-rac-glycerol) (DMPG), Chol, Pam 3CysSK 4and Quillaja Saponaria-derived saponin adjuvant QS21. The mice immunized with the modified vaccine elicit higher antibody titers and increased IgG 2a antibodies. Overall, our liposomal vaccines were found to be highly effective at generating a balanced and robust TH1/TH2 response.\u0000 Supported by grants from NIH (R01AI148570)","PeriodicalId":22698,"journal":{"name":"The Journal of Immunology","volume":"94 1","pages":""},"PeriodicalIF":0.0,"publicationDate":"2023-05-01","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"84997713","PeriodicalName":null,"FirstCategoryId":null,"ListUrlMain":null,"RegionNum":0,"RegionCategory":"","ArticlePicture":[],"TitleCN":null,"AbstractTextCN":null,"PMCID":"","EPubDate":null,"PubModel":null,"JCR":null,"JCRName":null,"Score":null,"Total":0}
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