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1.
World J Microbiol Biotechnol ; 40(7): 204, 2024 May 17.
Article in English | MEDLINE | ID: mdl-38755413

ABSTRACT

Globally colorectal cancer ranks as the third most widespread disease and the third leading cause of cancer-associated mortality. Immunotherapy treatments like PD-L1 blockade have been used to inhibit the PD-L1 legend, which boosts the activity of cytotoxic T lymphocytes. Recently, studies suggest that some probiotics could potentially enhance the effectiveness of immunotherapy treatments for cancer patients. We found that in Caco-2 and HT-29 cells, the live Leuconostoc mesenteroides treatment resulted an increase in the PD-L1 expression and this treatment stimulated interferon-gamma (IFN-γ) production in Jurkat T-cells. Due to the well-established ability of IFN-γ to enhance PD-L1 expression, the combination of IFN-γ and L. mesenteroides was used in colon cancer cell lines and a resulting remarkable increase of over tenfold in PD-L1 expression was obtained. Interestingly, when L. mesenteroides and IFN-γ are present, the blockage of PD-L1 using PD-L1 antibodies not only improved the viability of Jurkat T-cells but also significantly boosted the levels of IFN-γ and IL-2, the T-cells activation marker cytokines. In addition to upregulating PD-L1, L. mesenteroides also activated Toll-like receptors (TLRs) and NOD-like receptors (NODs) pathways, specifically through TLR2 and NOD2, while also exerting a suppressive effect on autophagy in colon cancer cell lines. In conclusion, our findings demonstrate a significant upregulation of PD-L1 expression in colon cancer cells upon co-culturing with L. mesenteroides. Moreover, the presence of PD-L1 antibodies during co-culturing activates Jurkat T cells. The observed enhancement in PD-L1 expression may be attributed to the inhibition of the Autophagy pathway or activation of the hippo pathway. KEY POINTS: Co-culturing L. mesenteroides increases PD-L1 gene and protein transaction in colon cancer. L. mesenteroides existing enhances T cells viability and activity. GPCR41/42 is a possible link between L. mesenteroides, YAP-1 and PD-L1.


Subject(s)
B7-H1 Antigen , Colonic Neoplasms , Interferon-gamma , Leuconostoc mesenteroides , Up-Regulation , Humans , B7-H1 Antigen/metabolism , B7-H1 Antigen/genetics , Interferon-gamma/metabolism , Colonic Neoplasms/immunology , HT29 Cells , Jurkat Cells , Caco-2 Cells , Leuconostoc mesenteroides/metabolism , Leuconostoc mesenteroides/genetics , Interleukin-2/metabolism , Lymphocyte Activation , T-Lymphocytes/immunology , T-Lymphocytes/metabolism , Probiotics/pharmacology , Cell Line, Tumor , Nod2 Signaling Adaptor Protein/metabolism , Nod2 Signaling Adaptor Protein/genetics , Toll-Like Receptor 2/metabolism , Toll-Like Receptor 2/genetics
2.
Eur J Med Chem ; 271: 116439, 2024 May 05.
Article in English | MEDLINE | ID: mdl-38691886

ABSTRACT

Nucleotide-binding oligomerization domain 2 (NOD2) is a receptor of the innate immune system that is capable of perceiving bacterial and viral infections. Muramyl dipeptide (MDP, N-acetyl muramyl L-alanyl-d-isoglutamine), identified as the minimal immunologically active component of bacterial cell wall peptidoglycan (PGN) is recognized by NOD2. In terms of biological activities, MDP demonstrated vaccine adjuvant activity and stimulated non-specific protection against bacterial, viral, and parasitic infections and cancer. However, MDP has certain drawbacks including pyrogenicity, rapid elimination, and lack of oral bioavailability. Several detailed structure-activity relationship (SAR) studies around MDP scaffolds are being carried out to identify better NOD2 ligands. The present review elaborates a comprehensive SAR summarizing structural aspects of MDP derivatives in relation to NOD2 agonistic activity.


Subject(s)
Acetylmuramyl-Alanyl-Isoglutamine , Nod2 Signaling Adaptor Protein , Nod2 Signaling Adaptor Protein/metabolism , Nod2 Signaling Adaptor Protein/agonists , Acetylmuramyl-Alanyl-Isoglutamine/pharmacology , Acetylmuramyl-Alanyl-Isoglutamine/chemistry , Structure-Activity Relationship , Humans , Animals , Molecular Structure
3.
Int J Mol Sci ; 25(10)2024 May 13.
Article in English | MEDLINE | ID: mdl-38791357

ABSTRACT

The lung is prone to infections from respiratory viruses such as Severe Acute Respiratory Syndrome Coronavirus 2 (SARS-CoV-2). A challenge in combating these infections is the difficulty in targeting antiviral activity directly at the lung mucosal tract. Boosting the capability of the respiratory mucosa to trigger a potent immune response at the onset of infection could serve as a potential strategy for managing respiratory infections. This study focused on screening immunomodulators to enhance innate immune response in lung epithelial and immune cell models. Through testing various subfamilies and pathways of pattern recognition receptors (PRRs), the nucleotide-binding and oligomerization domain (NOD)-like receptor (NLR) family was found to selectively activate innate immunity in lung epithelial cells. Activation of NOD1 and dual NOD1/2 by the agonists TriDAP and M-TriDAP, respectively, increased the number of IL-8+ cells by engaging the NF-κB and interferon response pathways. Lung epithelial cells showed a stronger response to NOD1 and dual NOD1/2 agonists compared to control. Interestingly, a less-pronounced response to NOD1 agonists was noted in PBMCs, indicating a tissue-specific effect of NOD1 in lung epithelial cells without inducing widespread systemic activation. The specificity of the NOD agonist pathway was confirmed through gene silencing of NOD1 (siRNA) and selective NOD1 and dual NOD1/2 inhibitors in lung epithelial cells. Ultimately, activation induced by NOD1 and dual NOD1/2 agonists created an antiviral environment that hindered SARS-CoV-2 replication in vitro in lung epithelial cells.


Subject(s)
COVID-19 , Epithelial Cells , Immunity, Innate , Lung , Nod1 Signaling Adaptor Protein , Nod2 Signaling Adaptor Protein , SARS-CoV-2 , Humans , Nod1 Signaling Adaptor Protein/metabolism , Nod1 Signaling Adaptor Protein/agonists , Immunity, Innate/drug effects , SARS-CoV-2/physiology , SARS-CoV-2/immunology , COVID-19/immunology , COVID-19/virology , Epithelial Cells/virology , Epithelial Cells/metabolism , Epithelial Cells/drug effects , Epithelial Cells/immunology , Lung/immunology , Lung/virology , Lung/metabolism , Nod2 Signaling Adaptor Protein/agonists , Nod2 Signaling Adaptor Protein/metabolism , COVID-19 Drug Treatment , NF-kappa B/metabolism , Antiviral Agents/pharmacology , A549 Cells , Diaminopimelic Acid/analogs & derivatives , Diaminopimelic Acid/pharmacology , Signal Transduction/drug effects , Interleukin-8/metabolism
4.
J Mol Med (Berl) ; 102(6): 787-799, 2024 06.
Article in English | MEDLINE | ID: mdl-38740600

ABSTRACT

Nucleotide-binding oligomerization domain containing 2 (NOD2), located in the cell cytoplasm, is a pattern recognition receptor belonging to the innate immune receptor family. It mediates the innate immune response by identifying conserved sequences in bacterial peptide glycans and plays an essential role in maintaining immune system homeostasis. Gene mutations of NOD2 lead to the development of autoimmune diseases such as Crohn's disease and Blau syndrome. Recently, NOD2 has been shown to be associated with the pathogenesis of diabetes, cardiac-cerebral diseases, and cancers. However, the function of NOD2 in these non-communicable diseases (CNCDs) is not well summarized in reviews. Our report mainly discusses the primary function and molecular mechanism of NOD2 as well as its potential clinical significance in CNCDs.


Subject(s)
Nod2 Signaling Adaptor Protein , Noncommunicable Diseases , Humans , Nod2 Signaling Adaptor Protein/genetics , Nod2 Signaling Adaptor Protein/metabolism , Animals , Chronic Disease , Mutation , Genetic Predisposition to Disease , Immunity, Innate
5.
Front Immunol ; 15: 1374368, 2024.
Article in English | MEDLINE | ID: mdl-38715616

ABSTRACT

NOD1 and NOD2 as two representative members of nucleotide-binding oligomerization domain (NOD)-like receptor (NLR) family play important roles in antimicrobial immunity. However, transcription mechanism of nod1 and nod2 and their signal circle are less understood in teleost fish. In this study, with the cloning of card9 and ripk2 in Chinese perch, the interaction between NOD1, NOD2, and CARD9 and RIPK2 were revealed through coimmunoprecipitation and immunofluorescence assays. The overexpression of NOD1, NOD2, RIPK2 and CARD9 induced significantly the promoter activity of NF-κB, IFNh and IFNc. Furthermore, it was found that nod1 and nod2 were induced by poly(I:C), type I IFNs, RLR and even NOD1/NOD2 themselves through the ISRE site of their proximal promoters. It is thus indicated that nod1 and nod2 can be classified also as ISGs due to the presence of ISRE in their proximal promoter, and their expression can be mechanistically controlled through PRR pathway as well as through IFN signaling in antiviral immune response.


Subject(s)
Fish Proteins , Nod1 Signaling Adaptor Protein , Nod2 Signaling Adaptor Protein , Receptor-Interacting Protein Serine-Threonine Kinase 2 , Signal Transduction , Animals , Nod1 Signaling Adaptor Protein/genetics , Nod1 Signaling Adaptor Protein/metabolism , Receptor-Interacting Protein Serine-Threonine Kinase 2/metabolism , Receptor-Interacting Protein Serine-Threonine Kinase 2/genetics , Nod2 Signaling Adaptor Protein/genetics , Nod2 Signaling Adaptor Protein/metabolism , Fish Proteins/genetics , Fish Proteins/metabolism , Fish Proteins/immunology , Perches/genetics , Perches/immunology , Perches/metabolism , Interferons/metabolism , Interferons/genetics , Promoter Regions, Genetic , Transcription, Genetic , Immunity, Innate/genetics , Protein Binding
6.
J Immunol ; 212(11): 1791-1806, 2024 Jun 01.
Article in English | MEDLINE | ID: mdl-38629918

ABSTRACT

RIG-I-like receptors and NOD-like receptors play pivotal roles in recognizing microbe-associated molecular patterns and initiating immune responses. The LGP2 and NOD2 proteins are important members of the RIG-I-like receptor and NOD-like receptor families, recognizing viral RNA and bacterial peptidoglycan (PGN), respectively. However, in some instances bacterial infections can induce LPG2 expression via a mechanism that remains largely unknown. In the current study, we found that LGP2 can compete with NOD2 for PGN binding and inhibit antibacterial immunity by suppressing the NOD2-RIP2 axis. Recombinant CiLGP2 (Ctenopharyngodon idella LGP2) produced using either prokaryotic or eukaryotic expression platform can bind PGN and bacteria in pull-down and ELISA assays. Comparative protein structure models and intermolecular interaction prediction calculations as well as pull-down and colocalization experiments indicated that CiLGP2 binds PGN via its EEK motif with species and structural specificity. EEK deletion abolished PGN binding of CiLGP2, but insertion of the CiLGP2 EEK motif into zebrafish and mouse LGP2 did not confer PGN binding activity. CiLGP2 also facilitates bacterial replication by interacting with CiNOD2 to suppress expression of NOD2-RIP2 pathway genes. Sequence analysis and experimental verification demonstrated that LGP2 having EEK motif that can negatively regulate antibacterial immune function is present in Cyprinidae and Xenocyprididae families. These results show that LGP2 containing EEK motif competes with NOD2 for PGN binding and suppresses antibacterial immunity by inhibiting the NOD2-RIP2 axis, indicating that LGP2 plays a crucial negative role in antibacterial response beyond its classical regulatory function in antiviral immunity.


Subject(s)
Nod2 Signaling Adaptor Protein , Peptidoglycan , Animals , Nod2 Signaling Adaptor Protein/metabolism , Nod2 Signaling Adaptor Protein/immunology , Nod2 Signaling Adaptor Protein/genetics , Peptidoglycan/metabolism , Peptidoglycan/immunology , Fish Proteins/immunology , Fish Proteins/genetics , Fish Proteins/metabolism , Receptor-Interacting Protein Serine-Threonine Kinase 2/metabolism , Carps/immunology , Mice , Protein Binding , Signal Transduction/immunology , Humans , Amino Acid Motifs , Zebrafish/immunology
7.
Infect Immun ; 92(5): e0000424, 2024 May 07.
Article in English | MEDLINE | ID: mdl-38563734

ABSTRACT

Neisseria gonorrhoeae, a human restricted pathogen, releases inflammatory peptidoglycan (PG) fragments that contribute to the pathophysiology of pelvic inflammatory disease. The genus Neisseria is also home to multiple species of human- or animal-associated Neisseria that form part of the normal microbiota. Here we characterized PG release from the human-associated nonpathogenic species Neisseria lactamica and Neisseria mucosa and animal-associated Neisseria from macaques and wild mice. An N. mucosa strain and an N. lactamica strain were found to release limited amounts of the proinflammatory monomeric PG fragments. However, a single amino acid difference in the PG fragment permease AmpG resulted in increased PG fragment release in a second N. lactamica strain examined. Neisseria isolated from macaques also showed substantial release of PG monomers. The mouse colonizer Neisseria musculi exhibited PG fragment release similar to that seen in N. gonorrhoeae with PG monomers being the predominant fragments released. All the human-associated species were able to stimulate NOD1 and NOD2 responses. N. musculi was a poor inducer of mouse NOD1, but ldcA mutation increased this response. The ability to genetically manipulate N. musculi and examine effects of different PG fragments or differing amounts of PG fragments during mouse colonization will lead to a better understanding of the roles of PG in Neisseria infections. Overall, we found that only some nonpathogenic Neisseria have diminished release of proinflammatory PG fragments, and there are differences even within a species as to types and amounts of PG fragments released.


Subject(s)
Neisseria , Nod1 Signaling Adaptor Protein , Nod2 Signaling Adaptor Protein , Peptidoglycan , Animals , Humans , Mice , Bacterial Proteins/metabolism , Bacterial Proteins/genetics , Membrane Transport Proteins , Neisseria/genetics , Neisseria gonorrhoeae/immunology , Neisseria gonorrhoeae/genetics , Neisseria gonorrhoeae/metabolism , Nod1 Signaling Adaptor Protein/metabolism , Nod1 Signaling Adaptor Protein/genetics , Nod2 Signaling Adaptor Protein/metabolism , Nod2 Signaling Adaptor Protein/genetics , Peptidoglycan/metabolism
8.
J Dent Res ; 103(6): 672-682, 2024 Jun.
Article in English | MEDLINE | ID: mdl-38679731

ABSTRACT

Enterococcus faecalis is an important contributor to the persistence of chronic apical periodontitis. However, the mechanism by which E. faecalis infection in the root canals and dentinal tubules affects periapical tissue remains unclear. Bacterial extracellular vesicles (EVs) act as natural carriers of microbe-associated molecular patterns (MAMPs) and have recently attracted considerable attention. In this study, we investigated the role of EVs derived from E. faecalis in the pathogenesis of apical periodontitis. We observed that E. faecalis EVs can induce inflammatory bone destruction in the periapical areas of mice. Double-labeling immunofluorescence indicated that M1 macrophage infiltration was increased by E. faecalis EVs in apical lesions. Moreover, in vitro experiments demonstrated the internalization of E. faecalis EVs into macrophages. Macrophages tended to polarize toward the M1 profile after treatment with E. faecalis EVs. Pattern recognition receptors (PRRs) can recognize MAMPs of bacterial EVs and, in turn, trigger inflammatory responses. Thus, we performed further mechanistic exploration, which showed that E. faecalis EVs considerably increased the expression of NOD2, a cytoplasmic PRR, and that inhibition of NOD2 markedly reduced macrophage M1 polarization induced by E. faecalis EVs. RIPK2 ubiquitination is a major downstream of NOD2. We also observed increased RIPK2 ubiquitination in macrophages treated with E. faecalis EVs, and E. faecalis EV-induced macrophage M1 polarization was notably alleviated by the RIPK2 ubiquitination inhibitor. Our study revealed the potential for EVs to be considered a virulence factor of E. faecalis and found that E. faecalis EVs can promote macrophage M1 polarization via NOD2/RIPK2 signaling. To our knowledge, this is the first report to investigate apical periodontitis development from the perspective of bacterial vesicles and demonstrate the role and mechanism of E. faecalis EVs in macrophage polarization. This study expands our understanding of the pathogenic mechanism of E. faecalis and provides novel insights into the pathogenesis of apical periodontitis.


Subject(s)
Enterococcus faecalis , Extracellular Vesicles , Macrophages , Periapical Periodontitis , Periapical Periodontitis/microbiology , Periapical Periodontitis/metabolism , Animals , Mice , Macrophages/microbiology , Nod2 Signaling Adaptor Protein/metabolism , Mice, Inbred C57BL , Disease Models, Animal
9.
J Stroke Cerebrovasc Dis ; 33(6): 107689, 2024 Jun.
Article in English | MEDLINE | ID: mdl-38527567

ABSTRACT

OBJECTIVES: Microglia-mediated neuroinflammation plays a crucial role in the pathophysiological process of multiple neurological disorders such as ischemic stroke, which still lacks effective therapeutic agents. Shikonin possesses anti-inflammatory and neuroprotective properties. However, its underlying mechanism remains elusive. This study aimed to investigate whether Shikonin confers protection against cerebral ischemia/reperfusion (I/R) injury by modulating microglial polarization and elucidate the associated mechanisms. METHODS: This study employed an oxygen-glucose deprivation and reoxygenation (OGD/R) BV2 microglial cellular model and a middle cerebral artery occlusion/reperfusion (MCAO/R) animal model to investigate the protection and underlying mechanism of Shikonin against ischemic stroke. RESULTS: The results demonstrated that Shikonin treatment significantly reduced brain infarction volume and improved neurological function in MCAO/R rats. Simultaneously, Shikonin treatment significantly reduced microglial proinflammatory phenotype and levels of proinflammatory markers (inducible-NO synthase (iNOS), tumor necrosis factor-alpha (TNF-α), interleukin-1 beta (IL-1ß), and IL-6), increased microglial anti-inflammatory phenotype and levels of anti-inflammatory markers (Arginase-1 (Arg1), transforming growth factor-beta (TGF-ß), and IL-10), reversed the expression of Nucleotide-binding oligomerization domain 2 (NOD2) and phosphorylation receptor interacting protein 2 (p-RIP2), and suppressed nuclear factor kappa-B (NF-κB) signaling activation in the ischemic penumbra regions. These effects of Shikonin were further corroborated in OGD/R-treated BV2 cells. Furthermore, overexpression of NOD2 markedly attenuated the neuroprotective effects of Shikonin treatment in MCAO/R rats. NOD2 overexpression also attenuated the regulatory effects of Shikonin on neuroinflammation, microglial polarization, and NF-κB signaling activation. CONCLUSION: This study illustrates that Shikonin mitigates inflammation mediated by microglial proinflammatory polarization by inhibiting the NOD2/RIP2/NF-κB signaling pathway, thereby exerting a protective role. The findings uncover a potential molecular mechanism for Shikonin in treating ischemic stroke.


Subject(s)
Anti-Inflammatory Agents , Infarction, Middle Cerebral Artery , NF-kappa B , Naphthoquinones , Neuroprotective Agents , Nod2 Signaling Adaptor Protein , Receptor-Interacting Protein Serine-Threonine Kinase 2 , Reperfusion Injury , Animals , Male , Mice , Anti-Inflammatory Agents/pharmacology , Cell Line , Cytokines/metabolism , Disease Models, Animal , Infarction, Middle Cerebral Artery/pathology , Infarction, Middle Cerebral Artery/metabolism , Infarction, Middle Cerebral Artery/drug therapy , Inflammation Mediators/metabolism , Microglia/drug effects , Microglia/metabolism , Microglia/pathology , Naphthoquinones/pharmacology , Neuroinflammatory Diseases/drug therapy , Neuroinflammatory Diseases/metabolism , Neuroinflammatory Diseases/pathology , Neuroprotective Agents/pharmacology , NF-kappa B/metabolism , Nod2 Signaling Adaptor Protein/metabolism , Phenotype , Rats, Sprague-Dawley , Receptor-Interacting Protein Serine-Threonine Kinase 2/metabolism , Reperfusion Injury/metabolism , Reperfusion Injury/drug therapy , Reperfusion Injury/pathology , Signal Transduction/drug effects
10.
Chem Commun (Camb) ; 60(16): 2212-2215, 2024 Feb 20.
Article in English | MEDLINE | ID: mdl-38305731

ABSTRACT

To further understand the specificity of muramyl dipeptide (MDP) sensing by NOD2, we evaluated the compatibility of synthetic MDP analogues for cellular uptake and NAGK phosphorylation, the pre-requisite steps of intracellular NOD2 activation. Our results revealed that these two prior steps do not confer ligand stereoselectivity; yet NAGK strictly discriminates against the disaccharide NOD2 agonists for phosphorylation in vitro, despite it being indispensable for the cellular NOD2-stimulating effects of these analogues, implying potential glycosidase cleavage as a novel intermediate step for cellular activation of NOD2.


Subject(s)
Acetylmuramyl-Alanyl-Isoglutamine , Nod2 Signaling Adaptor Protein , Acetylmuramyl-Alanyl-Isoglutamine/pharmacology , Acetylmuramyl-Alanyl-Isoglutamine/metabolism , Ligands , Nod2 Signaling Adaptor Protein/metabolism
11.
Arthritis Res Ther ; 26(1): 58, 2024 02 23.
Article in English | MEDLINE | ID: mdl-38395960

ABSTRACT

BACKGROUND AND OBJECTIVES: The study investigated the pathogenesis of Yao syndrome (YAOS), a rare systemic autoinflammatory disease associated with the nucleotide-binding oligomerization domain containing 2 (NOD2) gene variants. METHODS: RNA sequencing analyses were used to detect transcriptomic profile changes. Immunoblot and immunohistochemistry were used to examine the NOD2-mediated inflammatory signaling pathways and ELISA was used to detect cytokines. RESULTS: Transcriptome analysis of YAOS revealed NOD-like receptor signaling pathway enrichment. Compared with HCs, P-RIP2, p-p65, p-p38, p-ERK, and p-JNK notably increased in PBMCs of a patient with YAOS. P-RIP2, p-p65, and p-p38 elevated in small intestinal mucosa tissues. P-p65 and p-p38 in synovial tissues from YAOS were higher than those in patients with rheumatoid arthritis (RA) and osteoarthritis (OA). Serum interleukin (IL)-6 level along with tumor necrosis factor (TNF)-α and IL-6 secreted from PBMCs were markedly higher in patients with YAOS in comparison to healthy controls (HCs). The supernatants of synovial cells from a patient with YAOS showed substantially higher IL-1ß and IL-6 levels than those of RA and OA. Canakinumab therapy of a Q902K heterozygous patient with YAOS resulted in notable clinical improvement. CONCLUSION: Overproduction of pro-inflammatory cytokines and the hyperactivation of NOD2-mediated signaling pathways were found in the NOD2 variant Q902K patient with YAOS. NOD2-RIP2-MAPK pathway might play a pivotal role in the pathogenesis of YAOS. These results provide new perspectives for targeted therapies in YAOS.


Subject(s)
Hereditary Autoinflammatory Diseases , Nod2 Signaling Adaptor Protein , Humans , Arthritis, Rheumatoid/metabolism , Cytokines/metabolism , Interleukin-6/metabolism , Nod2 Signaling Adaptor Protein/genetics , Nod2 Signaling Adaptor Protein/metabolism , Osteoarthritis/metabolism , Synovial Membrane/metabolism , Tumor Necrosis Factor-alpha/metabolism , Hereditary Autoinflammatory Diseases/genetics
12.
J Med Chem ; 67(4): 3004-3017, 2024 Feb 22.
Article in English | MEDLINE | ID: mdl-38301029

ABSTRACT

NOD1 and NOD2 are members of the pattern recognition receptors involved in the innate immune response. Overactivation of NOD1 is implicated in inflammatory disorders, multiple sclerosis, and cancer cell metastases. NOD1 antagonists would represent valuable pharmacological tools to gain further insight into protein roles, potentially leading to new therapeutic strategies. We herein report the expansion of the chemical space of NOD1 antagonists via a multicomponent synthetic approach affording a novel chemotype, namely, 2,3-diaminoindoles. These efforts resulted in compound 37, endowed with low micromolar affinity toward NOD1. Importantly, a proof-of-evidence of direct binding to NOD1 of Noditinib-1 and derivative 37 is provided here for the first time. Additionally, the combination of computational studies and NMR-based displacement assays enabled the characterization of the binding modality of 37 to NOD1, thus providing key unprecedented knowledge for the design of potent and selective NOD1 antagonists.


Subject(s)
Immunity, Innate , Nod1 Signaling Adaptor Protein , Nod2 Signaling Adaptor Protein/metabolism , Indoles/chemistry , Indoles/metabolism
13.
Atherosclerosis ; 389: 117436, 2024 Feb.
Article in English | MEDLINE | ID: mdl-38277990

ABSTRACT

BACKGROUNDS AND AIMS: The role of inflammation in driving atherosclerosis is well-established. It exerts systemic effects beyond the local site of plaque formation. In the context of coronary artery disease (CAD), the proteins that show altered levels in the plasma, are potentially important for understanding the key regulatory mechanism in the pathogenesis of atherosclerosis. A case-control study revealed that plasma soluble Peptidoglycan Recognition Protein 2 (PGLYRP2) primarily produced by the liver, is increased in subjects with CAD. Furthermore, the concentration of PGLYRP2 in the blood correlates with the severity of coronary artery disease. Thus, it raises interest in understanding the exact role of the protein in aortic inflammation and plaque progression. METHODS: We evaluated the plasma concentration of PGLYRP2 in three distinct groups: patients with CAD (N = 68), asymptomatic individuals (N = 34), and healthy volunteers (N = 20). Furthermore, we investigated the correlation between disease severity and PGLYRP2 levels in CAD patients. To identify potential binding partners of PGLYRP2, we employed computational analysis. We verified the PGLYRP2-NOD2 interaction in macrophage cells and elucidated the inflammatory pathways activated by PGLYRP2 within these cells. To assess the impact of PGLYRP2, we examined its effects in the atherosclerotic mice model (ApoE-/-). RESULTS: In this study, we report for the first time that Nucleotide-binding Oligomerization domain 2 (NOD2) which is expressed on the surface of macrophages, is a receptor of PGLYRP2. The N-terminal domain of PGLYRP2 directly binds to NOD2 and activates the NOD2-RIP2-NFκB cascade that promotes the secretion of proinflammatory cytokines like TNFα, IL1ß, and IL-8. In the atherosclerotic mice model (ApoE-/-) we demonstrate that elevated PGLYRP2 level is parallel with increased proinflammatory cytokines in the plasma when fed a High Cholesterol Diet (HCD). Immunohistochemical analysis reveals that PGLYRP2 is co-localized with NOD2 on the macrophages at the site of the lesion. CONCLUSIONS: Taken together, our data demonstrate that NOD2 acts as a receptor of PGLYRP2 on macrophages, which mediates the activation of the NOD2-RIP2-NFκB pathway and promotes inflammation, thus significantly contributing to the development and progression of atherosclerosis.


Subject(s)
Carrier Proteins , Coronary Artery Disease , N-Acetylmuramoyl-L-alanine Amidase , Animals , Humans , Mice , Apolipoproteins E/metabolism , Atherosclerosis/pathology , Carrier Proteins/metabolism , Case-Control Studies , Cytokines/metabolism , Inflammation/metabolism , Nod2 Signaling Adaptor Protein/genetics , Nod2 Signaling Adaptor Protein/metabolism , N-Acetylmuramoyl-L-alanine Amidase/metabolism
14.
J Med Chem ; 66(21): 14391-14410, 2023 11 09.
Article in English | MEDLINE | ID: mdl-37857324

ABSTRACT

Receptor interacting serine/threonine protein kinase 2 (RIPK2) is a downstream signaling molecule essential for the activation of several innate immune receptors, including the NOD-like receptors (NOD1 and NOD2). Recognition of pathogen-associated molecular pattern proteins by NOD1/2 leads to their interaction with RIPK2, which induces release of pro-inflammatory cytokines through the activation of NF-κB and MAPK pathways, among others. Thus, RIPK2 has emerged as a key mediator of intracellular signal transduction and represents a new potential therapeutic target for the treatment of various conditions, including inflammatory diseases and cancer. In this Perspective, first, an overview of the mechanisms that underlie RIPK2 function will be presented along with its role in several diseases. Then, the existing inhibitors that target RIPK2 and different therapeutic strategies will be reviewed, followed by a discussion on current challenges and outlook.


Subject(s)
Receptor-Interacting Protein Serine-Threonine Kinase 2 , Signal Transduction , Receptor-Interacting Protein Serine-Threonine Kinase 2/metabolism , NF-kappa B/metabolism , Cytokines/metabolism , Nod1 Signaling Adaptor Protein , Nod2 Signaling Adaptor Protein/metabolism
15.
Front Immunol ; 14: 1242659, 2023.
Article in English | MEDLINE | ID: mdl-37869013

ABSTRACT

Nucleotide-binding oligomerization domain-containing proteins, NOD1 and NOD2, are cytosolic receptors that recognize dipeptides and tripeptides derived from the bacterial cell wall component peptidoglycan (PGN). During the past two decades, studies have revealed several roles for NODs beyond detecting PGN fragments, including activation of an innate immune anti-viral response, NOD-mediated autophagy, and ER stress induced inflammation. Recent studies have also clarified the dynamic regulation of NODs at cellular membranes to generate specific and balanced immune responses. This review will describe how NOD1 and NOD2 detect microbes and cellular stress and detail the molecular mechanisms that regulate activation and signaling while highlighting new evidence and the impact on inflammatory disease pathogenesis.


Subject(s)
Nod Signaling Adaptor Proteins , Nod1 Signaling Adaptor Protein , Humans , Nod Signaling Adaptor Proteins/metabolism , Nod2 Signaling Adaptor Protein/metabolism , Inflammation , Nucleotides/metabolism
16.
Innate Immun ; 29(8): 186-200, 2023 Nov.
Article in English | MEDLINE | ID: mdl-37828863

ABSTRACT

NOD1 and NOD2 sense small bacterial peptidoglycan fragments, often called muropeptides, that access the cytosol. These muropeptides include iE-DAP and MDP, the minimal agonists for NOD1 and NOD2, respectively. Here, we synthesized and validated alkyne-modified muropeptides, iE-DAP-Alk and MDP-Alk, for use in click-chemistry reactions. While it has long been known that many cell types respond to extracellular exposure to muropeptides, it is unclear how these innate immune activators access their cytosolic innate immune receptors, NOD1 and NOD2. The subcellular trafficking and transport mechanisms by which muropeptides access these cytosolic innate immune receptors are a major gap in our understanding of these critical host responses. The click-chemistry-enabled agonists developed here will be particularly powerful to decipher the underlying cell biology and biochemistry of NOD1 and NOD2 innate immune sensing.


Subject(s)
Nod1 Signaling Adaptor Protein , Receptor Protein-Tyrosine Kinases , Diaminopimelic Acid/metabolism , Receptor Protein-Tyrosine Kinases/metabolism , Nod1 Signaling Adaptor Protein/metabolism , Nod2 Signaling Adaptor Protein/genetics , Nod2 Signaling Adaptor Protein/metabolism
17.
J Exp Clin Cancer Res ; 42(1): 236, 2023 Sep 09.
Article in English | MEDLINE | ID: mdl-37684625

ABSTRACT

BACKGROUND: Fusobacterium nucleatum (Fn) acts as a procarcinogenic bacterium in colorectal carcinoma (CRC) by regulating the inflammatory tumor microenvironment (TME). Neutrophil extracellular traps (NETs), which can be generated by persistent inflammation, have been recently considered to be significant contributors in promoting cancer progression. However, whether NETs are implicated in Fn-related carcinogenesis is still poorly characterized. Here, we explored the role of NETs in Fn-related CRC as well as their potential clinical significance. METHODS: Fn was measured in tissue specimens and feces samples from CRC patients. The expression of NET markers were also detected in tissue specimens, freshly isolated neutrophils and blood serum from CRC patients, and the correlation of circulating NETs levels with Fn was evaluated. Cell-based experiments were conducted to investigate the mechanism by which Fn modulates NETs formation. In addition, we clarified the functional mechanism of Fn-induced NETs on the growth and metastasis of CRC in vitro and in vivo experiments. RESULTS: Tissue and blood samples from CRC patients, particularly those from Fn-infected CRC patients, exhibited greater neutrophil infiltration and higher NETs levels. Fn infection induced abundant NETs production in in vitro studies. Subsequently, we demonstrated that Fn-induced NETs indirectly accelerated malignant tumor growth through angiopoiesis, and facilitated tumor metastasis, as manifested by epithelial-mesenchymal transition (EMT)-related cell migration, matrix metalloproteinase (MMP)-mediated basement membrane protein degradation, and trapping of CRC cells. Mechanistically, the Toll-like receptor (TLR4)-reactive oxygen species (ROS) signaling pathway and NOD-like receptor (NOD1/2)-dependent signaling were responsible for Fn-stimulated NETs formation. More importantly, circulating NETs combined with carcinoembryonic antigen (CEA) could predict CRC occurrence and metastasis, with areas under the ROC curves (AUCs) of 0.92 and 0.85, respectively. CONCLUSIONS: Our findings indicated that Fn-induced NETs abundance by activating TLR4-ROS and NOD1/2 signalings in neutrophils facilitated CRC progression. The combination of circulating NETs and CEA was identified as a novel screening strategy for predicting CRC occurrence and metastasis.


Subject(s)
Colorectal Neoplasms , Extracellular Traps , Fusobacterium nucleatum , Neutrophils , Colorectal Neoplasms/microbiology , Colorectal Neoplasms/pathology , Disease Progression , Humans , Tumor Microenvironment , Inflammation , Signal Transduction , Reactive Oxygen Species/metabolism , Toll-Like Receptor 4/metabolism , Nod1 Signaling Adaptor Protein/metabolism , Nod2 Signaling Adaptor Protein/metabolism , Carcinoembryonic Antigen/blood , Male , Female , Middle Aged , Cell Line, Tumor , Mice, Inbred BALB C , Animals , Mice , Neoplasm Metastasis
18.
J Biochem Mol Toxicol ; 37(12): e23510, 2023 Dec.
Article in English | MEDLINE | ID: mdl-37700718

ABSTRACT

Recent years have witnessed increasing studies on the effect of epigenetic silencing of genes in the progression of chronic lymphocytic leukemia (CLL). This study investigates whether the nucleotide binding oligomerization domain containing 2 (NOD2) participates in the cell apoptosis and drug resistance of CLL cells. Cells were treated with adriamycin (ADR), etoposide, aclacinomycin and daunorubicin. After treatment, drug resistance and cell proliferation were examined to detect the inhibitory effect of ADR on cell proliferation; flow cytometry to identify ADR accumulation, the cell cycle distribution and apoptosis after transfection, and rhodamine 123 accumulation and efflux tests to assess P-glycoprotein (P-gp) function. NOD2 silencing or inhibition of the nuclear factor kappa-B (NF-κB) signaling pathway suppressed the multidrug resistance level in CLL, the inhibition rate, and cell proliferation caused by ADR at concentrations of approximately 0.25-1.5 µmol/L. Greater accumulation of ADR was observed in the CLL-AAT cell line than in the CLL-AAT/A02 cell line, but NOD2 silencing or inhibition of the NF-κB signaling pathway further increased the accumulation of ADR drugs in the CLL-AAT cell line and inhibited the drug efflux pump function of P-gp. Additionally, NOD2 silencing or NF-κB signaling pathway inhibition increased the apoptotic rate. The results of this study indicate that NOD2 promotes cell apoptosis and reduces the drug resistance of CLL by inhibiting the NF-κB signaling pathway.


Subject(s)
Leukemia, Lymphocytic, Chronic, B-Cell , NF-kappa B , Humans , NF-kappa B/metabolism , Leukemia, Lymphocytic, Chronic, B-Cell/drug therapy , Leukemia, Lymphocytic, Chronic, B-Cell/genetics , Leukemia, Lymphocytic, Chronic, B-Cell/metabolism , Drug Resistance, Neoplasm , Signal Transduction , Doxorubicin/pharmacology , Doxorubicin/therapeutic use , Apoptosis , ATP Binding Cassette Transporter, Subfamily B, Member 1 , ATP Binding Cassette Transporter, Subfamily B/metabolism , Nod2 Signaling Adaptor Protein/genetics , Nod2 Signaling Adaptor Protein/metabolism , Nod2 Signaling Adaptor Protein/pharmacology
19.
Gut Microbes ; 15(2): 2249960, 2023 12.
Article in English | MEDLINE | ID: mdl-37655966

ABSTRACT

Over 90% of epidemic non-bacterial gastroenteritis are caused by human noroviruses (NoVs), which persist in a substantial subset of people allowing their spread worldwide. This has led to a significant number of endemic cases and up to 70,000 children deaths in developing countries. NoVs are primarily transmitted through the fecal-oral route. To date, studies have focused on the influence of the gut microbiota on enteric viral clearance by mucosal immunity. In this study, the use of mouse norovirus S99 (MNoV_S99) and CR6 (MNoV_CR6), two persistent strains, allowed us to provide evidence that the norovirus-induced exacerbation of colitis severity relied on bacterial sensing by nucleotide-binding oligomerization domain 2 (Nod2). Consequently, Nod2-deficient mice showed reduced levels of gravity of Dextran sodium sulfate (DSS)-induced colitis with both viral strains. And MNoV_CR6 viremia was heightened in Nod2-/- mice in comparison with animals hypomorphic for Atg16l1, which are prone to aggravated inflammation under DSS. Accordingly, the infection of macrophages derived from WT mice promoted the phosphorylation of Signal Transducer and Activator of Transcription 1 (STAT1) and NOD2's expression levels. Higher secretion of Tumor Necrosis Factor alpha (TNFα) following NOD2 activation and better viral clearance were measured in these cells. By contrast, reduced levels of pSTAT1 and blunted downstream secretion of TNFα were found in Nod2-deficient macrophages infected by MNoV_S99. Hence, our results uncover a previously unidentified virus-host-bacterial interplay that may represent a novel therapeutic target for treating noroviral origin gastroenteritis that may be linked with susceptibility to several common illnesses such as Crohn's disease.


Subject(s)
Caliciviridae Infections , Colitis , Gastroenteritis , Gastrointestinal Microbiome , Nod2 Signaling Adaptor Protein , Animals , Mice , Caliciviridae Infections/immunology , Colitis/chemically induced , Colitis/virology , Gastroenteritis/immunology , Gastroenteritis/virology , Nod2 Signaling Adaptor Protein/metabolism
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