Unambiguous diagnosis of the two main forms of inflammatory bowel diseases (IBD): Ulcerative colitis (UC) and Crohn’s disease (CD), represents a challenge in the early stages of the diseases. The diagnosis...Unambiguous diagnosis of the two main forms of inflammatory bowel diseases (IBD): Ulcerative colitis (UC) and Crohn’s disease (CD), represents a challenge in the early stages of the diseases. The diagnosis may be established several years after the debut of symptoms. Hence, protein biomarkers for early and accurate diagnostic could help clinicians improve treatment of the individual patients. Moreover, the biomarkers could aid physicians to predict disease courses and in this way, identify patients in need of intensive treatment. Patients with low risk of disease flares may avoid treatment with medications with the concomitant risk of adverse events. In addition, identification of disease and course specific biomarker profiles can be used to identify biological pathways involved in the disease development and treatment. Knowledge of disease mechanisms in general can lead to improved future development of preventive and treatment strategies. Thus, the clinical use of a panel of biomarkers represents a diagnostic and prognostic tool of potentially great value. The technological development in recent years within proteomic research (determination and quantification of the complete protein content) has made the discovery of novel biomarkers feasible. Several IBD-associated protein biomarkers are known, but none have been successfully implemented in daily use to distinguish CD and UC patients. The intestinal tissue remains an obvious place to search for novel biomarkers, which blood, urine or stool later can be screened for. When considering the protein complexity encountered in intestinal biopsy-samples and the recent development within the field of mass spectrometry driven quantitative proteomics, a more thorough and accurate biomarker discovery endeavor could today be performed than ever before. In this review, we report the current status of the proteomics IBD biomarkers and discuss various emerging proteomic strategies for identifying and characterizing novel biomarkers, as well as suggesting future targe展开更多
Excessive release of neutrophil extracellular traps(NETs)is associated with disease severity and contributes to tissue injury,followed by severe organ damage.Pharmacological or genetic inhibition of NET release reduce...Excessive release of neutrophil extracellular traps(NETs)is associated with disease severity and contributes to tissue injury,followed by severe organ damage.Pharmacological or genetic inhibition of NET release reduces pathology in multiple inflammatory disease models,indicating that NETs are potential therapeutic targets.Here,we demonstrate using a preclinical basket approach that our therapeutic anti-citrullinated protein antibody(tACPA)has broad therapeutic potential.Treatment with tACPA prevents disease symptoms in various mouse models with plausible NET-mediated pathology,including inflammatory arthritis(IA),pulmonary fibrosis,inflammatory bowel disease and sepsis.We show that citrulline residues in the N-termini of histones 2A and 4 are specific targets for therapeutic intervention,whereas antibodies against other N-terminal post-translational histone modifications have no therapeutic effects.Because citrullinated histones are generated during NET release,we investigated the ability of tACPA to inhibit NET formation.tACPA suppressed NET release from human neutrophils triggered with physiologically relevant human disease-related stimuli.Moreover,tACPA diminished NET release and potentially initiated NET uptake by macrophages in vivo,which was associated with reduced tissue damage in the joints of a chronic arthritis mouse model of IA.To our knowledge,we are the first to describe an antibody with NET-inhibiting properties and thereby propose tACPA as a drug candidate for NET-mediated inflammatory diseases,as it eliminates the noxious triggers that lead to continued inflammation and tissue damage in a multidimensional manner.展开更多
Peptidyl arginine deiminase 4(PAD4)plays a pivotal role in infection and inflammatory diseases by facilitating the formation of neutrophil extracellular traps(NETs).However,the substrates of PAD4 and its exact role in...Peptidyl arginine deiminase 4(PAD4)plays a pivotal role in infection and inflammatory diseases by facilitating the formation of neutrophil extracellular traps(NETs).However,the substrates of PAD4 and its exact role in inflammatory bowel disease(IBD)remain unclear.In this study,we employed single-cell RNA sequencing(scRNA-seq)and substrate citrullination mapping to decipher the role of PAD4 in intestinal inflammation associated with IBD.Our results demonstrated that PAD4 deficiency alleviated colonic inflammation and restored intestinal barrier function in a dextran sulfate sodium(DSS)-induced colitis mouse model.scRNA-seq analysis revealed significant alterations in intestinal cell populations,with reduced neutrophil numbers and changes in epithelial subsets upon PAD4 deletion.Gene expression analysis highlighted pathways related to inflammation and epithelial cell function.Furthermore,we found that neutrophil-derived extracellular vesicles(EVs)carrying PAD4 were secreted into intestinal epithelial cells(IECs).Within IECs,PAD4 citrullinates mitochondrial creatine kinase 1(CKMT1)at the R242 site,leading to reduced CKMT1 protein stability via the autophagy pathway.This action compromises mitochondrial homeostasis,impairs intestinal barrier integrity,and induces IECs apoptosis.IEC-specific depletion of CKMT1 exacerbated intestinal inflammation and apoptosis in mice with colitis.Clinical analysis of IBD patients revealed elevated levels of PAD4,increased CKMT1 citrullination,and decreased CKMT1 expression.In summary,our findings highlight the crucial role of PAD4 in IBD,where it modulates IECs plasticity via CKMT1 citrullination,suggesting that PAD4 may be a potential therapeutic target for IBD.展开更多
Rheumatoid arthritis(RA)is an inflammatory autoimmune disease triggered by antigenic peptides with environmental and genetic risk factors.It has been shown that antigen-specific targeting could be a promising therapeu...Rheumatoid arthritis(RA)is an inflammatory autoimmune disease triggered by antigenic peptides with environmental and genetic risk factors.It has been shown that antigen-specific targeting could be a promising therapeutical strategy for RA by restoring immune tolerance to self-antigens without compromising normal immunity.Citrullination of antigens enhances antigenic properties and induces autoimmune responses.Here,we showed that citrullinated antigenic(citAg)vaccine ameliorated collageninduced arthritis with decreased T-helper 1(Th1)and Th17 cells,downregulated proinflammatory cytokines including interlukin-6 and tumor necrosis factor-a,and inhibited antigen recall responses.B cell receptor sequencing further revealed that citAg vaccine could dampen the dysregulated V(D)J recombination,restoring the immune repertoire.Taken together,the results demonstrated that citAg vaccine might have a therapeutic effect on RA.展开更多
Histone citrullination,an important post-translational modification mediated by peptidyl arginine deiminases,is essential for many physiological processes and epigenetic regulation.However,the causal relationship betw...Histone citrullination,an important post-translational modification mediated by peptidyl arginine deiminases,is essential for many physiological processes and epigenetic regulation.However,the causal relationship between histone citrullination and specific gene regulation remains unresolved.In this study,we develop a programmable epigenetic editor by fusing the peptidyl arginine deiminase(PAD)PPAD from Porphyromonas gingivalis with d Cas9.With the assistance of g RNA,PPAD-d Cas9 can recruit PPADs to specific genomic loci,enabling direct manipulation of the epigenetic landscape and regulation of gene expression.Our citrullination editor allows for the site-specific manipulation of histone H3R2,8,17 and H3R26 at target human gene loci,resulting in the activation or suppression of different genes in a locus-specific manner.Moreover,the epigenetic effects of the citrullination editor are specific and sustained.This epigenetic editor offers an accurate and efficient tool for exploring gene regulation of histone citrullination.展开更多
Protein citrullination,including histone H1 and H3 citrullination,is important for transcriptional regulation,DNA damage response,and pluripotency of embryonic stem cells(ESCs).Tripartite motif containing 28(Trim28),a...Protein citrullination,including histone H1 and H3 citrullination,is important for transcriptional regulation,DNA damage response,and pluripotency of embryonic stem cells(ESCs).Tripartite motif containing 28(Trim28),an embryonic development regulator involved in ESC self-renewal,has recently been identified as a novel substrate for citrullination by Padi4.However,the physiological functions of Trim28 citrullination and its role in regulating the chromatin structure and gene transcription of ESCs remain unknown.In this paper,we show that Trim28 is specifically citrullinated in mouse ESCs(m ESCs),and that the loss of Trim28 citrullination induces loss of pluripotency.Mechanistically,Trim28 citrullination enhances the interaction of Trim28with Smarcad1 and prevents chromatin condensation.Additionally,Trim28 citrullination regulates m ESC pluripotency by promoting transcription of Nanog and Klf4 which it does by increasing the enrichment of H3K27ac and H3K4me3 and decreasing the enrichment of H3K9me3 in the transcriptional regulatory region.Thus,our findings suggest that Trim28citrullination is the key for the epigenetic activation of pluripotency genes and pluripotency maintenance of ESCs.Together,these results uncover a role Trim28 citrullination plays in pluripotency regulation and provide novel insight into how citrullination of proteins other than histones regulates chromatin compaction.展开更多
Both cholinergic dysfunction and protein citrullination are the hallmarks of rheumatoid arthritis(RA),but the relationship between the two phenomena remains unclear.We explored whether and how cholinergic dysfunction ...Both cholinergic dysfunction and protein citrullination are the hallmarks of rheumatoid arthritis(RA),but the relationship between the two phenomena remains unclear.We explored whether and how cholinergic dysfunction accelerates protein citrullination and consequently drives the development of RA.Cholinergic function and protein citrullination levels in patients with RA and collageninduced arthritis(CIA)mice were collected.In both neuron-macrophage coculture system and CIA mice,the effect of cholinergic dysfunction on protein citrullination and expression of peptidylarginine deiminases(PADs)was assessed by immunofluorescence.The key transcription factors for PAD4 expression were predicted and validated.Cholinergic dysfunction in the patients with RA and CIA mice negatively correlated with the degree of protein citrullination in synovial tissues.The cholinergic or alpha7 nicotinic acetylcholine receptor(a7nAChR)deactivation and activation resulted in the promotion and reduction of protein citrullination in vitro and in vivo,respectively.Especially,the activation deficiency of a7nAChR induced the earlier onset and aggravation of CIA.Furthermore,deactivation of a7nAChR increased the expression of PAD4 and specificity protein-3(SP3)in vitro and in vivo.Our results suggest that cholinergic dysfunction-induced deficient a7nAChR activation,which induces the expression of SP3 and its downstream molecule PAD4,accelerating protein citrullination and the development of RA.展开更多
文摘Unambiguous diagnosis of the two main forms of inflammatory bowel diseases (IBD): Ulcerative colitis (UC) and Crohn’s disease (CD), represents a challenge in the early stages of the diseases. The diagnosis may be established several years after the debut of symptoms. Hence, protein biomarkers for early and accurate diagnostic could help clinicians improve treatment of the individual patients. Moreover, the biomarkers could aid physicians to predict disease courses and in this way, identify patients in need of intensive treatment. Patients with low risk of disease flares may avoid treatment with medications with the concomitant risk of adverse events. In addition, identification of disease and course specific biomarker profiles can be used to identify biological pathways involved in the disease development and treatment. Knowledge of disease mechanisms in general can lead to improved future development of preventive and treatment strategies. Thus, the clinical use of a panel of biomarkers represents a diagnostic and prognostic tool of potentially great value. The technological development in recent years within proteomic research (determination and quantification of the complete protein content) has made the discovery of novel biomarkers feasible. Several IBD-associated protein biomarkers are known, but none have been successfully implemented in daily use to distinguish CD and UC patients. The intestinal tissue remains an obvious place to search for novel biomarkers, which blood, urine or stool later can be screened for. When considering the protein complexity encountered in intestinal biopsy-samples and the recent development within the field of mass spectrometry driven quantitative proteomics, a more thorough and accurate biomarker discovery endeavor could today be performed than ever before. In this review, we report the current status of the proteomics IBD biomarkers and discuss various emerging proteomic strategies for identifying and characterizing novel biomarkers, as well as suggesting future targe
基金Feasibility grants from the Dutch government(MTVLA15144 and NITLS14050).
文摘Excessive release of neutrophil extracellular traps(NETs)is associated with disease severity and contributes to tissue injury,followed by severe organ damage.Pharmacological or genetic inhibition of NET release reduces pathology in multiple inflammatory disease models,indicating that NETs are potential therapeutic targets.Here,we demonstrate using a preclinical basket approach that our therapeutic anti-citrullinated protein antibody(tACPA)has broad therapeutic potential.Treatment with tACPA prevents disease symptoms in various mouse models with plausible NET-mediated pathology,including inflammatory arthritis(IA),pulmonary fibrosis,inflammatory bowel disease and sepsis.We show that citrulline residues in the N-termini of histones 2A and 4 are specific targets for therapeutic intervention,whereas antibodies against other N-terminal post-translational histone modifications have no therapeutic effects.Because citrullinated histones are generated during NET release,we investigated the ability of tACPA to inhibit NET formation.tACPA suppressed NET release from human neutrophils triggered with physiologically relevant human disease-related stimuli.Moreover,tACPA diminished NET release and potentially initiated NET uptake by macrophages in vivo,which was associated with reduced tissue damage in the joints of a chronic arthritis mouse model of IA.To our knowledge,we are the first to describe an antibody with NET-inhibiting properties and thereby propose tACPA as a drug candidate for NET-mediated inflammatory diseases,as it eliminates the noxious triggers that lead to continued inflammation and tissue damage in a multidimensional manner.
基金National Natural Science Foundation of China(No.82100587,No.82170567)National Key R&D Program of China(No.2023YFC2413801,China)+6 种基金Shanghai Sailing Program(No.21YF1458700)China National Postdoctoral Program for Innovative Talents(No.BX20220288)China Postdoctoral Science Foundation(No.2022M720138)Program of Shanghai Academic Research Leader(No.22XD1425000)Deep Blue Project of Naval Medical University(Pilot Talent Plan)Basic Medical Research Project of the First Affiliated Hospital of Naval Medical University(No.2023PY06)“Changying”Talent Program of Changhai Hospital of Naval Medical University,and the“Changjian”Talent Program of Changhai Hospital of Naval Medical University.
文摘Peptidyl arginine deiminase 4(PAD4)plays a pivotal role in infection and inflammatory diseases by facilitating the formation of neutrophil extracellular traps(NETs).However,the substrates of PAD4 and its exact role in inflammatory bowel disease(IBD)remain unclear.In this study,we employed single-cell RNA sequencing(scRNA-seq)and substrate citrullination mapping to decipher the role of PAD4 in intestinal inflammation associated with IBD.Our results demonstrated that PAD4 deficiency alleviated colonic inflammation and restored intestinal barrier function in a dextran sulfate sodium(DSS)-induced colitis mouse model.scRNA-seq analysis revealed significant alterations in intestinal cell populations,with reduced neutrophil numbers and changes in epithelial subsets upon PAD4 deletion.Gene expression analysis highlighted pathways related to inflammation and epithelial cell function.Furthermore,we found that neutrophil-derived extracellular vesicles(EVs)carrying PAD4 were secreted into intestinal epithelial cells(IECs).Within IECs,PAD4 citrullinates mitochondrial creatine kinase 1(CKMT1)at the R242 site,leading to reduced CKMT1 protein stability via the autophagy pathway.This action compromises mitochondrial homeostasis,impairs intestinal barrier integrity,and induces IECs apoptosis.IEC-specific depletion of CKMT1 exacerbated intestinal inflammation and apoptosis in mice with colitis.Clinical analysis of IBD patients revealed elevated levels of PAD4,increased CKMT1 citrullination,and decreased CKMT1 expression.In summary,our findings highlight the crucial role of PAD4 in IBD,where it modulates IECs plasticity via CKMT1 citrullination,suggesting that PAD4 may be a potential therapeutic target for IBD.
基金supported by the Guangdong Basic and Applied Basic Research Foundation(2020B1515130005)the National Natural Science Foundation of China(82371807,82171773,92374202,and 32141004)+1 种基金the National Key R&D Program of China(2022YFC3602000)the Beijing Nova Program(Z181100006218044 and Z211100002121163)。
文摘Rheumatoid arthritis(RA)is an inflammatory autoimmune disease triggered by antigenic peptides with environmental and genetic risk factors.It has been shown that antigen-specific targeting could be a promising therapeutical strategy for RA by restoring immune tolerance to self-antigens without compromising normal immunity.Citrullination of antigens enhances antigenic properties and induces autoimmune responses.Here,we showed that citrullinated antigenic(citAg)vaccine ameliorated collageninduced arthritis with decreased T-helper 1(Th1)and Th17 cells,downregulated proinflammatory cytokines including interlukin-6 and tumor necrosis factor-a,and inhibited antigen recall responses.B cell receptor sequencing further revealed that citAg vaccine could dampen the dysregulated V(D)J recombination,restoring the immune repertoire.Taken together,the results demonstrated that citAg vaccine might have a therapeutic effect on RA.
基金funded by the Tianjin Synthetic Biotechnology Innovation Capacity Improvement Project(TSBICIP-CXRC-048)。
文摘Histone citrullination,an important post-translational modification mediated by peptidyl arginine deiminases,is essential for many physiological processes and epigenetic regulation.However,the causal relationship between histone citrullination and specific gene regulation remains unresolved.In this study,we develop a programmable epigenetic editor by fusing the peptidyl arginine deiminase(PAD)PPAD from Porphyromonas gingivalis with d Cas9.With the assistance of g RNA,PPAD-d Cas9 can recruit PPADs to specific genomic loci,enabling direct manipulation of the epigenetic landscape and regulation of gene expression.Our citrullination editor allows for the site-specific manipulation of histone H3R2,8,17 and H3R26 at target human gene loci,resulting in the activation or suppression of different genes in a locus-specific manner.Moreover,the epigenetic effects of the citrullination editor are specific and sustained.This epigenetic editor offers an accurate and efficient tool for exploring gene regulation of histone citrullination.
基金supported by the National Natural Science Foundation of China(31972884)the National Key Research and Development Program of China(2018YFC1312300)+3 种基金the National Clinical Research Center for Geriatrics(Z20201007)1·3·5 Project for Disciplines of Excellence,West China Hospital(ZYGD18003)the Postdoctoral Research Project,West China Hospital,Sichuan University(18HXBH068)the Natural Science Foundation of Sichuan,China(2022NSFSC1424)。
文摘Protein citrullination,including histone H1 and H3 citrullination,is important for transcriptional regulation,DNA damage response,and pluripotency of embryonic stem cells(ESCs).Tripartite motif containing 28(Trim28),an embryonic development regulator involved in ESC self-renewal,has recently been identified as a novel substrate for citrullination by Padi4.However,the physiological functions of Trim28 citrullination and its role in regulating the chromatin structure and gene transcription of ESCs remain unknown.In this paper,we show that Trim28 is specifically citrullinated in mouse ESCs(m ESCs),and that the loss of Trim28 citrullination induces loss of pluripotency.Mechanistically,Trim28 citrullination enhances the interaction of Trim28with Smarcad1 and prevents chromatin condensation.Additionally,Trim28 citrullination regulates m ESC pluripotency by promoting transcription of Nanog and Klf4 which it does by increasing the enrichment of H3K27ac and H3K4me3 and decreasing the enrichment of H3K9me3 in the transcriptional regulatory region.Thus,our findings suggest that Trim28citrullination is the key for the epigenetic activation of pluripotency genes and pluripotency maintenance of ESCs.Together,these results uncover a role Trim28 citrullination plays in pluripotency regulation and provide novel insight into how citrullination of proteins other than histones regulates chromatin compaction.
基金supported by the“Double First-Class”University Project(CPU2022QZ31,China)。
文摘Both cholinergic dysfunction and protein citrullination are the hallmarks of rheumatoid arthritis(RA),but the relationship between the two phenomena remains unclear.We explored whether and how cholinergic dysfunction accelerates protein citrullination and consequently drives the development of RA.Cholinergic function and protein citrullination levels in patients with RA and collageninduced arthritis(CIA)mice were collected.In both neuron-macrophage coculture system and CIA mice,the effect of cholinergic dysfunction on protein citrullination and expression of peptidylarginine deiminases(PADs)was assessed by immunofluorescence.The key transcription factors for PAD4 expression were predicted and validated.Cholinergic dysfunction in the patients with RA and CIA mice negatively correlated with the degree of protein citrullination in synovial tissues.The cholinergic or alpha7 nicotinic acetylcholine receptor(a7nAChR)deactivation and activation resulted in the promotion and reduction of protein citrullination in vitro and in vivo,respectively.Especially,the activation deficiency of a7nAChR induced the earlier onset and aggravation of CIA.Furthermore,deactivation of a7nAChR increased the expression of PAD4 and specificity protein-3(SP3)in vitro and in vivo.Our results suggest that cholinergic dysfunction-induced deficient a7nAChR activation,which induces the expression of SP3 and its downstream molecule PAD4,accelerating protein citrullination and the development of RA.
