Abstract
Ulcerative colitis (UC) is a chronic, recurrent inflammatory bowel disease. UC confronts with severe challenges including the unclear pathogenesis and lack of specific diagnostic markers, demanding for identifying predictive biomarkers for UC diagnosis and treatment. We perform immune infiltration and weighted gene co-expression network analysis on gene expression profiles of active UC, inactive UC, and normal controls to identify UC related immune cell and hub genes. Neutrophils, M1 macrophages, activated dendritic cells, and activated mast cells are significantly enriched in active UC. MMP-9, CHI3L1, CXCL9, CXCL10, CXCR2 and S100A9 are identified as hub genes in active UC. Specifically, S100A9 is significantly overexpressed in mice with colitis. The receiver operating characteristic curve demonstrates the excellent performance of S100A9 expression in diagnosing active UC. Inhibition of S100A9 expression reduces DSS-induced colonic inflammation. These identified biomarkers associated with activity in UC patients enlighten the new insights of UC diagnosis and treatment.
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Introduction
Ulcerative colitis (UC) is a chronic and recurrent inflammatory bowel disease (IBD) with increasing incidence worldwide1. UC confronts with severe challenges including the unclear pathogenesis and lack of specific diagnostic markers, demanding for identifying predictive biomarkers for UC diagnosis and treatment.
Immune disorders play a crucial role in the etiology of UC among the complex pathogenic mechanisms2,3. Infiltrating immune cells in the intestinal mucosa of UC patients leads to mucosal inflammation. Early evidence suggests that UC is driven by Th2-polarized T cells in the lamina propria of the colon4. Additionally, neutrophils are key mediators of epithelial cytotoxicity and barrier dysfunction in UC5. Recently, targeting immune cells to inhibit inflammation has become a research hotspot. For example, 260 Itaconate modified cysteine sites were found in the macrophage proteome, and Itaconate could covalently modify the cysteine of macrophages to play an anti-inflammatory role6. However, the proportion and composition of infiltrating immune cells in the intestinal mucosal tissue of active UC remain unclear. Therefore, revealing the immune cells closely related to the pathogenesis of UC may provide a new direction for its treatment.
Limited achievements have been made in the field of diagnosis and surveillance of UC. The diagnosis of UC mainly relies on invasive tests including colonoscopy and histopathology. Besides, colonoscopy is highly costed and associated with complications such as perforation7,8. Although identified markers such as fecal calprotectin, C-reactive protein, and Intercellular adhesion molecule 1 were associated with disease activity, they were inaccurately correlated with endoscopic status9,10. Thus, it is emergently demanded for newly noninvasive and economic strategies for UC diagnosis.
Microrray technology have provided important insights into the pathophysiology mechanisms of disease at the genetic level11,12. Bioinformatics analysis has been widely used to search for differentially expression genes (DEGs), miRNA, and functional pathways involved in the development and progression of UC. For example, Wu et al. constructed a complete lncRNA-miRNA-mRNA network through bioinformatics to determine the specific immune infiltration characteristics of UC13. Moreover, protein–protein interaction (PPI) can provide information about direct and indirect protein interactions for screening hub genes14,15.
This study explored the difference of immune cell infiltration of UC in normal intestinal mucosal tissues through biological information. The hub genes (CXCL9, CXCL10, MMP-9, CHI3L1, CXCR2 and S100A9) of UC were identified by DEGs, weighted gene co-expression network analysis (WGCNA) and protein–protein interaction (PPI) network analysis. Then, we verified the results of bioinformatics analysis using external cohort and in vitro experiments, and explored the mechanism of hub genes in UC. Comprehensive analysis of immune infiltrating cells and hub genes will provide new biomarkers for the diagnosis and treatment of UC.
Materials and methods
Microarray data
The gene expression profile of GSE87466 was obtained from the GEO database (http://www.ncbi.nlm.nih.gov/geo/). A total of 108 mucosal biopsy samples were obtained from 87 active UC patients and 21 control subjects for subsequent analysis. The platform for GSE107499 was based on the GPL15207 Affymetrix Human Gene Expression Array, containing 75 active and 44 inactive UC mucosal biopsy samples.
Data preprocessing and DEGs analysis
All raw data were normalized and standardized by using the R software package. Gene differential expression analysis was conducted through the “limma” packages in the Bioconductor package (available online: http://www.bioconductor.org/). p value < 0.05 and |log2FC|> 2 were set as cut-off standards and considered to indicate statistical significance.
Gene set enrichment analysis (GSEA) and gene set variation analysis (GSVA)
In the training sets GSE87466 and GSE107499, a chip expression profile file and a sample data file (UC vs control, active vs inactive), respectively, were created and imported into the GSEA software16. Hallmark gene sets were selected to obtain pathway enrichment results for total gene expression levels. NOM p-value < 0.05 and FDR q-value < 25% were considered significantly enriched. To explore the biological function of S100A9 in UC, GSVA was performed on S100A9 high expression group and S100A9 low expression group17.
Immune cell infiltration
Gene expression datasets for GSE87466 and GSE107499 were uploaded to the CIBERSORT portal (http://cibersort.stanford.edu/) in the accepted CIBERSORT format. The original CIBERSORT gene signature file LM22, which defines 22 immune cell subtypes, was used to analyze immune cell infiltration in UC tissues and normal tissues, active UC tissues and inactive UC tissues18. The samples were screened according to P value < 0.05.
WGCNA analysis
The top 25% of genes with the largest variance in the gene expression dataset of GSE87466 were extracted to perform WGCNA. The R package “WGCNA” was applied to find clinical traits-related modules. In order to ensure the reliable results of network construction, one outlier sample was removed. Here, soft-thresholding power was set to 12 to convert the similarity matrix of gene expression into an adjacency matrix. The fitting degree of scale-free topological model was 0.80. Then, the adjacency relationship was transformed into a topological overlap matrix. The dynamic tree cutting method was used for module clustering, merging closer modules into new modules with a height of 0.2. Each gene network module sets a minimum number of bases of 50.
Gene ontology and KEGG pathway enrichment analysis
DAVID (https://david.ncifcrf.gov) is an online bioinformatics tool designed to identify a large number of gene or protein functions19. DAVID software was used to perform GO (including Biological process, cellular component and molecular function) and KEGG pathway analysis20. p value < 0.05 was considered to indicate statistical significance.
Protein–protein interaction network and hub gene definition
To predict protein–protein interactions, the online databases STRING21 and GENEMANIA22 were utilized in the PPI network analysis. The integrated regulatory networks were then visualized by cytoscape. The degree of protein nodes was calculated by using the cytoscape plugin, cytohubba, to find top10 hub genes.
Data validation
To verify the robustness of hub genes, the microarray data of GSE59071 was obtained from GPL6244 [HuGene-1_0-st] Affymetrix Human Gene 1.0 ST Array and included 97 colonic mucosal tissues from patients with active UC and 11 healthy colon mucosal tissues. The microarray data of GSE126124 [HuGene-1_0-st] Affymetrix Human Gene 1.0 ST Array [transcript (gene) version], which included 57 peripheral blood samples (18 UC samples and 39 control samples), were downloaded from the GEO database.
Animals and experimental design
SPF grade healthy female C57BL/6 mice aged 6–8 weeks were purchased from Beijing SiPeiFu Bio-Technology. Acute colitis was induced in mice by continuous feeding with 3% dextran sulfate sodium (DSS) (MPbio, USA) for 8 days. Mice were anesthetized by intraperitoneal injection of pentobarbital sodium. During the experiment, the body weight, fecal characteristics and blood in the stool of each mouse were recorded every day. According to the instructions, the mice in the treatment group were intraperitoneally injected with paquinimod (MedChemExpress, ABR 25757) at 5 mg/kg every 2 days. The mice were sacrificed (cervical disloaction) humanely and the colon length of each group of mice was measured after the experiment. Body weight, colon length and hematoxylin–eosin (H&E) staining were used to evaluate the severity of colitis in each group of mice. All animal procedures were performed in accordance with the Three Gorges University Institutional Animal Care and Use Committee.
Quantitative real-time PCR
According to the manufacturer’s instruction, total RNA from UC groups and controls colon tissues was extracted using a Trizol reagent kit (Takara, Dalian, China). qRT-PCR was then performed as described previously23. All primers were synthesized by TSINGKE (Shanghai, China) (Table S1). β-Actin was used as an internal control.
Western blotting
Tissues protein extraction was carried out on ice using a RIPA buffer (Sigma-Aldrich, Darmstadt, Germany) containing proteinase and phosphatase inhibitors. Western blotting was performed as described previously23. Primary antibodies against S100A9 (Abcam, ab242945, Cambridge, UK, 1:1000 dilution), and β-actin (Proteintech, 66009-1-Ig, Wuhan, China, 1:5000 dilution). β-actin was used to normalize the protein level.
Immunohistochemistry
Immunohistochemical analysis of mouse intestinal tissue was performed using anti-S100A9 (Abcam, ab242945, Cambridge, UK, 1:200 dilution), anti-MPO (Abcam, ab208670, Cambridge, UK, 1:1000 dilution), or anti-F4/80 (CST, #70076, Danvers, USA, 1:1000 dilution) antibodies and incubated overnight at 4 °C. Immunohistochemical was performed as described previously23.
Statistical analysis
Data was analyzed using the R software. GraphPad prism 8.00 software was used to calculate the area under the curve. Statistical significance between the two groups was calculated by Student's t-test. ****P < 0.0001; ***P < 0.001; **P < 0.01; *P < 0.05; ns, not significant.
Ethical approval and informed consent
The China Three Gorges University Ethics Committee gave its approval to the animal experimentation methodology. The study is reported in accordance with ARRIVE guidelines.
Results
Immune infiltration in active UC
To understand the whole-gene enrichment annotation of UC and consider the potential role of genes with smaller differences in expression in UC and normal tissues, GSEA was conducted to search KEGG pathways enriched in the active UC. GSEA results showed that active UC was mainly enriched in nod-like receptors, cytokine-cytokine receptor interactions, Toll-like receptors, ECM receptor interactions, cell adhesion molecules, chemokines, leukocyte transendothelial migration and other inflammatory signaling pathways (Fig. 1a, S1a). To further elucidate the immune infiltration of active UC and normal tissues, CIBERSORT algorithm was used to calculate the proportion of 22 immune cell subsets between 87 UC and 21 normal tissues (Fig. S1b), 75 active UC and 44 inactive UC samples (Fig. S1c). M0 macrophages, M1 macrophages, activated DCs, activated mast cells, neutrophils, activated CD4+ memory T‐cells and gamma delta T‐cells were upregulated in active UC. Regulatory T cells, M2 macrophages, resting DCs, resting mast cells were downregulated in active UC (Fig. 1b, S1d). Correlation analysis suggests that the functions of activated mast cells, M1 macrophages, and follicle-assisted T cells may promote each other, while M2 macrophages may antagonize M1 macrophages and activated mast cells (Fig. S2a,b).
According to the corresponding steps of WGCNA described in the materials and methods, 15 modules were obtained after merging similar genes (Fig. 1c, S3a–c). According to the correlation diagram between modules and clinical information, the correlation between black modules and active UC was the highest, with a correlation coefficient of 0.87 (p = 5e−34). Followed by the red module (r = 0.67, p = 2e−15). Meanwhile, the black module and the red module are positively correlated with the expression of neutrophils, M1 macrophages, and negatively correlated with the M2 macrophages (Fig. 1d). Therefore, the black and red modules were considered the most study-worthy.
Identification of DEGs and functional enrichment
In order to explore the differential gene expression between active UC and normal intestinal mucosal tissues, “Limma” package was used to identify DEGs in GSE87466. A total of 111 DEGs were identified between the UC group and the normal control group, including 37 downregulated genes and 74 upregulated genes (Fig. 2a). The Venn diagram showed the overlap between the DEGs and the black and red modules, respectively (Fig. 2b). Additionally, in related to UC activity, the black and red modules were assessed for further functional enrichment, consisting of GO enrichment and KEGG pathway analysis of module genes of interest. KEGG pathway analysis of DEGs in black and red module demonstrate that the most significant pathways are cytokine-cytokine receptor interaction, chemokine signaling pathway and TNF signaling pathway (Fig. 2c,f). The results of the GO analysis of DEGs in black and red module were shown in Fig. 2d and e. The DEGs were particularly enriched in inflammatory response, extracellular space and chemokine activity.
Identification of hub genes
To identify candidate biomarkers related to UC activity, the STRING and GENEMANIA database were applied to identify the interactions between DEGs in black and red module (Fig. 3a,c, Fig. S4a,b). Cytoscape was then used to construct a network of PPI for DEGs in the black and red modules, respectively (Fig. 3a,c). Top 10 hub genes in the black and red module were identified in PPI network respectively (Fig. 3b,d). “Corrplot” R package was used to further calculate the relationship between hub genes and three types of immune cells (M1 macrophages, M2 macrophages and neutrophils). The results showed that MMP-9 and CHI3L1 had the most negative correlation with M2 macrophages, CXCL9 and CXCL10 exhibited the most positive correlation with M1 macrophages, S100A9 and CXCR2 were most positively correlated with neutrophils (Fig. 3e).
Data validation
To verify the reliability and robustness of the above six genes, two independent datasets were used for external validation. Compared with healthy and inactive UC patients, the expression of MMP-9, CHI3L1, CXCL9, CXCL10, and S100A9 were significantly increased in active UC patients (Fig. 4a,b). Transcriptome expression profiles of whole blood samples from UC and paired patients were used to identify potential molecules for UC diagnosis. The diagnostic accuracy of MMP-9 and S100A9 was 0.883 and 0.812, respectively (Fig. 4c), which was significantly higher than that of CRP, a commonly used clinical indicator.
S100a9 is highly expressed in intestinal inflammatory mucosal tissue
In order to verify whether in vivo experiments are consistent with our bioinformatics analysis results, the colitis model was carried out. Compared with the control group, the body weight and colon length of mice in DSS treatment group were significantly reduced (Fig. 5a–c). Histological lesions and inflammatory cytokine (Il-6, Tnf-α) levels indicate that the colitis model is successfully established (Fig. 5d,e). The mRNA expression levels of Cxcr2, Chi3l1 and S100a9 in colitis mice were significantly higher than those in control group, especially S100a9, which was consistent with our bioinformatics analysis results (Fig. 5e, S5a). Finally, S100a9 was selected from the six hub genes for further analysis. Consistent with the results of the qPCR experiment, the S100a9 protein was overexpressed in the intestinal tissue of colitis mice (Fig. 5f, S7). Histochemical results showed that the expression of S100a9 in intestinal tissue of colitis mice was increased, and it was positively correlated with neutrophils and macrophages (Fig. 5g). Furthermore, the gene set variation analysis results demonstrate that IL‐2/STAT5, IL‐6/JAK/STAT3 signalling, apoptosis, angiogenesis and epithelial‐mesenchymal transition (EMT) are highly enriched in active UC with high S100a9 expression (Fig. S5b).
Blockade of S100a9 alleviates DSS-induced colitis in mice
To determine the role of S100a9 in the pathogenesis of colonic inflammation, the S100a9-selective inhibitor paquinimod (Paq, 5 mg/kg) was administered intraperitoneally every two days. Paq had no obvious effect on mouse growth (Fig. 6a,c). Compared with DSS treated mice, Paq-treated mice had less symptoms of colitis/ weight loss/ intestinal mucosal destruction and less shortening colon (Fig. 6a–d, S6). Meanwhile, the intestinal tissue of the Paq-treated mice showed less immune cell (neutrophils and macrophages) infiltration (Fig. 6e,f). Taken together, our data suggest that S100a9 may serve as a potential therapeutic target for UC.
Discussion
In this study, we performed immune infiltration analysis on UC (active UC and inactive UC) patients and healthy volunteers, and find that a variety of immune cells such as neutrophils, macrophages and activated DCs were highly enriched in active UC. Among these immune cells, neutrophils, M1 macrophages and DCs can participate in the proinflammatory response in a variety of ways, such as producing granular lyase and neutrophils extracellular traps, and secreting pro-inflammatory cytokines including IL-6, IL-1β and TNF-α24,25,26,27. In contrast, M2 macrophages produces a large number of anti-inflammatory cytokines (IL-10) and anti-inflammatory chemokines (CCL-17, CCL-24) that are involved in suppressing immune responses and tissue healing28. To date, there are few reports on the involvement of mast cells in UC. Here we showed that activated mast cells increased significantly in UC patients, while resting mast cells decreased, suggesting that mast cells may be involved in the pathogenesis of UC. In addition, no significant changes in NK cells were observed in our study, suggesting that NK cells may not be involved in the pathogenesis of UC. Regulatory cells (Tregs) play an important role in suppressing the immune response and maintaining peripheral tolerance29. We found that the number of Tregs increased in UC patients compared with healthy individuals, which is inconsistent with the results of Yao et al.30. One possible reason is that the expression level of Tregs are related to the disease progression of UC. Also, we observed a significant increase of memory B cells in the intestinal tissues of patients with active UC, but the function of memory B cells in the pathogenesis of UC remains to be studied. It has been reported that circulating memory B cells are associated with serum immunoglobulin level in patients with ulcerative colitis and may be involved in the pathogenesis of UC31.
We screened the genes most related to UC through WGCNA analysis, and obtained two modules, which were also significantly correlated with the differentially expressed immune cells (neutrophils, macrophages and activated DCs). In addition to inflammatory responses and innate immune responses, we also found that differential genes in the modules were particularly enriched in multiple metabolic pathways, such as extracellular space, serine endopeptidase activity, and zinc ion binding. There is increasing evidence showing that the extracellular space is important for UC initiation32. According to free radical induction theory, excessive hydrogen peroxide can be produced by abnormal metabolism of colonic epithelial cells, which can cause extensive oxidative damage to the intestinal barrier33. Recently, serine endopeptidase activity was reported to be most prominent among several protein functions associated with UC34. Besides, overexpression of zinc-binding protein may lead to the inactivation of p53 tumor suppressor gene and promote UC-associated colorectal cancer progression35. In short, functional annotation and pathway enrichment analysis may provide new directions for the pathogenesis of UC.
In this study, six potential hub genes (CXCL9, CXCL10, MMP-9, CHI3L1, CXCR2 and S100A9) in UC were identified. CXCL9 and CXCL10 genes also play important roles in tumors, such as melanoma and colorectal cancer36,37. As the expression pattern in tumor, the CXCL9, -10, -11/CXCR3 axis impacts TAMs polarization. However, immune cells, show anti-tumor effect against cancer cells through paracrine CXCL9, -10, -11/CXCR3 axis, the autocrine CXCL9, -10, -11/CXCR3 signaling in cancer cells increases cancer cell proliferation, angiogenesis, and metastasis38. Studies have proved that the expression levels of CXCL10 and CXCL9 in tumor may be correlated with a poor prognosis of overall survival39. However, some studies also show that CXCL10 and CXCL9 may promote colonic tumorigenesis via promotes the cytokine-mediated mucosal injury and inflammation response40. In UC, CXCL10 and CXCL9 can recruit leukocytes to the site of inflammation and promote the occurrence and development of inflammation through CXCL9, CXCL10, CXCL11/CXCR3 axis, which works primarily for immune cell migration, differentiation, and activation. CXCL10 and CXCL9 drive increased transcription of T-bet and RORγ, leading to Th1 polarization. After polarization, Th1 cells induce activation of CTLs, NK cells, and NKT cells through IFN-γ41.
In the analysis of the whole blood samples from UC patients, strikingly, the sensitivity of MMP-9 and S100A9 in the diagnosis of active UC was significantly higher than that of CRP, a commonly used clinical diagnostic molecule. This suggests that MMP-9 and S100A9 can be used as specific, sensitive and less invasive biomarkers of active UC, which is consistent with other reports42,43. A large number of studies have shown that MMP-9 is highly expressed in the intestinal tissues of patients with UC and actively participated in the pathophysiological process of UC44,45,46. MMP-9 can affect the tight connection between mucosal cells, increase intestinal mucosal permeability and aggravate mucosal barrier function47. Studies on MMP-9 deficient mice also suggest that MMP-9 is associated with mucosal damage in the early stages of colitis48. However, it has also been reported that MMP-9 restricts the accumulation of reactive oxygen species and DNA damage in colon, and thus inhibits the occurrence colitis-associated cancer49. Therefore, understanding the role of MMP-9 in UC and colitis-associated cancer is of great importance for exploring the treatment of UC and colitis-associated cancer.
S100A9 is a calcium-binding protein mainly expressed by neutrophils, monocytes, and macrophages, and play a key role in the pathophysiology of various inflammatory diseases50. In rheumatoid arthritis, S100A9-mediated neutrophil migration and secretion of pro-inflammatory cytokines from monocytes lead to joint inflammation and joint destruction51. In gout, it`s a key factor that S100A9 drives the production of sodium urate crystals and further induces the secretion of IL-1β in pathogenesis52. In contrast, in streptococcal pneumonia, blocking S100A9 significantly inhibited the migration of neutrophils and macrophages to alveoli53. However, it remains unknown about the biological function of S100A9 in intestinal inflammation. Our analysis indicated that the IL-2/STAT5, IL6/JAK/STAT3 and TNF‐α/NF‐κB pathway were significantly activated in UC with higher expression of S100A9. This partly reveals the specific mechanism of S100A9 in UC inflammatory response. In addition, angiogenesis and EMT pathways were also significantly activated in patients with high S100A9 expression, suggesting that S100A9 may be involved in UC-related tumor development.
In our animal model, S100a9 mRNA and protein levels were significantly elevated in mice with colitis. Moreover, the expression of S100a9 was significantly correlated with neutrophils and macrophages, which were closely related to the pathogenesis of UC. Inhibition of S100a9 expression significantly reduced intestinal immune cell infiltration and inflammatory response in mice with colitis. Our results suggest that blocking S100a9 inhibits inflammatory symptoms associated with UC and S100a9 may be a potential therapeutic target for UC. However, there were some limitations in this study such as small sample size and lack of own sequencing data. Thus, more animal and clinical studies are needed to validate the results of this study in order to develop new treatments for UC in the future.
Conclusions
In this study, through multiple bioinformatics approaches, immune cell infiltration characteristics were in active UC, and six genes (CXCL9, CXCL10, MMP-9, CHI3L1, CXCR2 and S100A9) were screened out and verified as potential key genes of UC. Furthermore, S100A9 serve as candidate diagnostic and therapeutic biomarkers for UC.
Data availability
The datasets used and analyzed during the current study are available from the corresponding author on reasonable request. The accession number for GEO are GSE87466, GSE107499, GSE59071 and GSE126124.
References
Ng, S. C. et al. Worldwide incidence and prevalence of inflammatory bowel disease in the 21st century: A systematic review of population-based studies. Lancet 390, 2769–2778 (2017).
Lee, S. H., Kwon, J. E. & Cho, M.-L. Immunological pathogenesis of inflammatory bowel disease. Intest. Res. 16, 26–42 (2018).
Xue, G. et al. Characteristics of immune cell infiltration and associated diagnostic biomarkers in ulcerative colitis: Results from bioinformatics analysis. Bioengineered 12, 252–265 (2021).
Manousou, P. et al. Increased expression of chemokine receptor CCR3 and its ligands in ulcerative colitis: The role of colonic epithelial cells in in vitro studies. Clin. Exp. Immunol. 162, 337–347 (2010).
Brazil, J. C., Louis, N. A. & Parkos, C. A. The role of polymorphonuclear leukocyte trafficking in the perpetuation of inflammation during inflammatory bowel disease. Inflamm. Bowel Dis. 19, 1556–1565 (2013).
Gao, G. et al. Brilliant glycans and glycosylation: Seq and ye shall find. Int. J. Biol. Macromol. 189, 279–291 (2021).
Arora, G. et al. Risk of perforation from a colonoscopy in adults: A large population-based study. Gastrointest Endosc. 69, 654–664 (2009).
Abraham, C. & Cho, J. H. Mechanisms of disease inflammatory bowel disease. N. Engl. J. Med. 361, 2066–2078 (2009).
Kourkoulis, P. et al. Novel potential biomarkers for the diagnosis and monitoring of patients with ulcerative colitis. Eur. J. Gastroenterol. Hepatol. 31, 1173–1183 (2019).
Vainer, B. Intercellular adhesion molecule-1 (ICAM-1) in ulcerative colitis: Presence, visualization, and significance. Inflamm. Res. 54, 313–327 (2005).
Hephzibah, C. R. et al. A review of bioinformatics tools and web servers in different microarray platforms used in cancer research. Adv. Protein Chem. Struct. Biol. 131, 85–164 (2022).
Udhaya, K. S. et al. A systemic approach to explore the mechanisms of drug resistance and altered signaling cascades in extensively drug-resistant tuberculosis. Adv. Protein Chem. Struct. Biol. 127, 343–364 (2021).
Dong, L. et al. Construction, bioinformatics analysis, and validation of competitive endogenous RNA networks in ulcerative colitis. Front. Genet. 13, 951243 (2022).
Balasundaram, A. et al. A computational model revealing the immune-related hub genes and key pathways involved in rheumatoid arthritis (RA). Adv. Protein Chem. Struct. Biol. 129, 247–273 (2022).
Udhaya, K. S. et al. Analysis of differentially expressed genes and molecular pathways in familial hypercholesterolemia involved in atherosclerosis: A systematic and bioinformatics approach. Front. Genet. 11, 734 (2020).
Subramanian, A. et al. Gene set enrichment analysis: A knowledge-based approach for interpreting genome-wide expression profiles. Proc. Natl. Acad. Sci. U. S. A. 102, 15545–15550 (2005).
Hänzelmann, S., Castelo, R. & Guinney, J. GSVA: Gene set variation analysis for microarray and RNA-seq data. BMC Bioinform. 14, 7 (2013).
Newman, A. M. et al. Determining cell type abundance and expression from bulk tissues with digital cytometry. Nat. Biotechnol. 37, 773–782 (2019).
Sherman, B. T. et al. DAVID: A web server for functional enrichment analysis and functional annotation of gene lists (2021 update). Nucleic Acids Res. 50, W216–W221 (2022).
Kanehisa, M. et al. KEGG for taxonomy-based analysis of pathways and genomes. Nucleic Acids Res. 51(D1), D587–D592 (2023).
Milosavljevic, F. et al. Association of CYP2C19 and CYP2D6 poor and intermediate metabolizer status with antidepressant and antipsychotic exposure: A systematic review and meta-analysis. JAMA Psychiatry 78, 270–280 (2021).
Warde-Farley, D. et al. The GeneMANIA prediction server: Biological network integration for gene prioritization and predicting gene function. Nucleic Acids Res. 38, W214–W220 (2010).
Tan, L. et al. Interferon regulatory factor-1 suppresses DNA damage response and reverses chemotherapy resistance by downregulating the expression of RAD51 in gastric cancer. Am. J. Cancer Res. 10, 1255–1270 (2020).
Dinallo, V. et al. Neutrophil extracellular traps sustain inflammatory signals in ulcerative colitis. J. Crohns Colitis. 13, 772–784 (2019).
Zhang, J. et al. Macrophage-based nanotherapeutic strategies in ulcerative colitis. J. Control. Release. 320, 363–380 (2020).
Hart, A. L. et al. Characteristics of intestinal dendritic cells in inflammatory bowel diseases. Gastroenterology 129, 50–65 (2005).
Watanabe, S. et al. Correlation of dendritic cell infiltration with active crypt inflammation in ulcerative colitis. Clin. Immunol. 122, 288–297 (2007).
Kmiec, Z., Cyman, M. & Slebioda, T. J. Cells of the innate and adaptive immunity and their interactions in inflammatory bowel disease. Adv. Med. Sci. 62, 1–16 (2017).
Yang, W. Y. et al. Pathological conditions re-shape physiological Tregs into pathological Tregs. Burns Trauma 3, 1 (2015).
Yao, J. et al. Effect of resveratrol on Treg/Th17 signaling and ulcerative colitis treatment in mice. World J. Gastroenterol. 21, 6572–6581 (2015).
Wang, X. et al. Circulating memory B cells and plasmablasts are associated with the levels of serum immunoglobulin in patients with ulcerative colitis. J. Cell. Mol. Med. 20, 804–814 (2016).
Zhang, X. et al. Transcription factor paired related homeobox 1 (PRRX1) activates matrix metalloproteinases (MMP)13, which promotes the dextran sulfate sodium-induced inflammation and barrier dysfunction of NCM460 cells. Bioengineered 13, 645–654 (2022).
Pravda, J. Radical induction theory of ulcerative colitis. World J. Gastroenterol. 11, 2371–2384 (2005).
Thuy-Boun, P. S. et al. Quantitative metaproteomics and activity-based protein profiling of patient fecal microbiome identifies host and microbial serine-type endopeptidase activity associated with ulcerative colitis. Mol. Cell. Proteomics 21, 100197 (2022).
Bruewer, M. et al. Metallothionein: Early marker in the carcinogenesis of ulcerative colitis-associated colorectal carcinoma. World J. Surg. 26, 726–731 (2002).
Kawada, K. et al. Pivotal role of CXCR3 in melanoma cell metastasis to lymph nodes. Cancer Res. 64, 4010–4017 (2004).
Zipin-Roitman, A. et al. CXCL10 promotes invasion-related properties in human colorectal carcinoma cells. Cancer Res. 67, 3396–3405 (2007).
House, I. G. et al. Macrophage-derived CXCL9 and CXCL10 are required for antitumor immune responses following immune checkpoint blockade. Clin. Cancer Res. 26, 487–504 (2020).
Jin, J. et al. Identification of CXCL10-relevant tumor microenvironment characterization and clinical outcome in ovarian cancer. Front Genet. 12, 678747 (2021).
Shukla, P. K. et al. Chronic ethanol feeding promotes azoxymethane and dextran sulfate sodium-induced colonic tumorigenesis potentially by enhancing mucosal inflammation. BMC Cancer 16, 189 (2016).
Hosomi, S. et al. Increased numbers of immature plasma cells in peripheral blood specifically overexpress chemokine receptor CXCR3 and CXCR4 in patients with ulcerative colitis. Clin. Exp. Immunol. 163, 215–224 (2011).
Shamseya, A. M. et al. Serum matrix metalloproteinase-9 concentration as a marker of disease activity in patients with inflammatory bowel disease. Eur. J. Gastroenterol. Hepatol. 33, e803–e809 (2021).
Kopi, T. A. et al. The value of mRNA expression of S100A8 and S100A9 as blood-based biomarkers of inflammatory bowel disease. Arab. J. Gastroenterol. 20, 135–140 (2019).
de Bruyn, M. et al. The molecular biology of matrix metalloproteinases and tissue inhibitors of metalloproteinases in inflammatory bowel diseases. Crit. Rev. Biochem. Mol. Biol. 51, 295–358 (2016).
Buisson, A. et al. Fecal matrix metalloprotease-9 and lipocalin-2 as biomarkers in detecting endoscopic activity in patients with inflammatory bowel diseases. J. Clin. Gastroenterol. 52, E53–E62 (2018).
Martinesi, M. et al. Down-regulation of adhesion molecules and matrix metalloproteinases by ZK 156979 in inflammatory bowel diseases. Clin. Immunol. 136, 51–60 (2010).
Bai, X. et al. Changes in MMP-2, MMP-9, inflammation, blood coagulation and intestinal mucosal permeability in patients with active ulcerative colitis. Exp. Ther. Med. 20, 269–274 (2020).
Santana, A. et al. Attenuation of dextran sodium sulphate induced colitis in matrix metalloproteinase-9 deficient mice. World J. Gastroenterol. 12, 6464–6472 (2006).
Walter, L. et al. Matrix metalloproteinase 9 (MMP9) limits reactive oxygen species (ROS) accumulation and DNA damage in colitis-associated cancer. Cell Death Dis. 11, 767 (2020).
Jukic, A. et al. Calprotectin: from biomarker to biological function. Gut 70, 1978–1988 (2021).
Cesaro, A. et al. An inflammation loop orchestrated by s100a9 and calprotectin is critical for development of arthritis. PLOS ONE 7, e45478 (2012).
Holzinger, D. et al. Myeloid-related proteins 8 and 14 contribute to monosodium urate monohydrate crystal-induced inflammation in gout. Arthritis Rheum. 66, 1327–1339 (2014).
Raquil, M.-A. et al. Blockade of antimicrobial proteins S100A8 and S100A9 inhibits phagocyte migration to the alveoli in streptococcal pneumonia. J. Immunol. 180, 3366–3374 (2008).
Funding
This work was supported by the Science and Technology Research Project of Education Department of Hubei Province (B2019026), and Medical and Health Research Project of Yichang (A23-1-107).
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L.T.: Data curation, Formal analysis, Validation, Visualization, Writing—original draft. X.L.: Methodology, Investigation, Validation, Formal analysis. H.Q.: Formal analysis, Methodology, Writing—review and editing. Q.Z.: Methodology, Software, J.W.: Methodology, investigation. T.C.: Writing—review and editing. C.Z.: Funding acquisition, Supervision. X.Z.: Conceptualization, writing—review and editing, supervision. Y.T.: Conceptualization, writing—review and editing, supervision, funding acquisition.
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Tan, L., Li, X., Qin, H. et al. Identified S100A9 as a target for diagnosis and treatment of ulcerative colitis by bioinformatics analysis. Sci Rep 14, 5517 (2024). https://doi.org/10.1038/s41598-024-55944-3
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DOI: https://doi.org/10.1038/s41598-024-55944-3
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