Expression and regulation of the CXCL9-11 chemokines and CXCR3 receptor in Atlantic salmon (Salmo salar).

Chemokines are cytokines that mediate leukocyte traffic between the lymphoid organs, the bloodstream, and the site of tissue damage, which is essential for an efficient immune response. In particular, the gamma interferon (IFN- γ) inducible chemokines CXCL9, CXCL10, and CXCL11, and their receptor CXCR3, are involved in T cell and macrophage recruitment to the site of infection. The nature and function of these chemokines and their receptor are well-known in mammals, but further research is needed to achieve a similar level of understanding in fish immunity. Thus, in this study, we seek to identify the genes encoding the components of the Atlantic salmon (Salmo salar) CXCL9, CXCL10, CXCL11/CXCR3 axis (CXCL9-11/CXCR3), predict the protein structure from the amino acid sequence, and explore the regulation of gene expression as well as the response of these chemokines and their receptor to viral infections. The cxcl9, cxcl10, cxcl11, and cxcr3 gene sequences were retrieved from the databases, and the phylogenetic analysis was conducted to determine the evolutionary relationships. The study revealed an interesting pattern of clustering and conservation among fish and mammalian species. The salmon chemokine sequences clustered with orthologs from other fish species, while the mammalian sequences formed separate clades. This indicates a divergent evolution of chemokines between mammals and fish, possibly due to different evolutionary pressures. While the structural analysis of the chemokines and the CXCR3 receptor showed the conservation of critical motifs and domains, suggesting preserved functions and stability throughout evolution. Regarding the regulation of gene expression, some components of the CXCL9-11/CXCR3 axis are induced by recombinant gamma interferon (rIFN-γ) and by Infectious pancreatic necrosis virus (IPNV) infection in Atlantic salmon cells. Further studies are needed to explore the role of Atlantic salmon CXCL9-11 chemokines in regulating immune cell migration and endothelial activation, as seen in mammals. To the best of our knowledge, there have been no functional studies of chemokines to understand these effects in Atlantic salmon.

Oxaliplatin inhibits colorectal cancer progression by inhibiting CXCL11 secreted by cancer-associated fibroblasts and the CXCR3/PI3K/AKT pathway.

PURPOSE: Colorectal cancer (CRC) is a malignant tumor. Oxaliplatin (OXA) can inhibit cancer-associated fibroblasts (CAFs)-induced cancer progression. This study sought to explore the mechanism of OXA in CAFs-induced CRC development. METHODS: CRC cell lines (Caco-2, SW620), normal fibroblasts (NFs), and CAFs were treated with OXA. NFs and CAFs were cultured. CAFs were treated with/without OXA (0.4 mM), and the supernatant was extracted as the conditioned medium (CM) to culture CRC cells. Cell malignant episodes, E-cadherin and Vimentin levels, CXCL1, CXCL2, CXCL3, CXCL8, and CXCL11 mRNA levels, CXCL11 protein level, and extracellular release were assessed. CAFs were transfected with interfering RNA sh-CXCL11 to silence CXCL11 or transfected with CXCL11 overexpression plasmids and treated with OXA to explore the role of CXCL11 in OXA-mediated CRC cells through CAFs. CXCL11 receptor CXCR3 levels in CRC cells and the PI3K/AKT pathway changes were examined. The xenogeneic tumor was transplanted in nude mice. CXCL11 and CXCR3 levels in tumor tissues, tumor volume, shape, size, weight, and Ki67 positive expressions were assessed. RESULTS: CRC cell growths and epithelial-mesenchymal transformation were stimulated after culture with CAFs-CM, while OXA averted these trends. CXCL11 mRNA level was elevated most significantly, and its protein and extracellular secretion levels were raised, while OXA diminished the levels. CXCL11 silencing weakened the effects of CAFs-CM on promoting CRC proliferation and malignant episodes and CXCL11 overexpression averted OXA property on inhibiting CAFs-promoted CRC cell growth. CXCR3 and PI3K and AKT1 phosphorylation levels were raised in the CAFs-CM group but diminished by OXA. CXCL11 overexpression in CAFs averted OXA property on inhibiting CAFs-activated CXCR3/PI3K/AKT in CRC cells. OXA also inhibited the progression of xenograft tumors by limiting CAFs-secreted CXCL11. CONCLUSIONS: OXA repressed CRC progression by inhibiting CAFs-secreted CXCL11 and the CXCR3/PI3K/AKT pathway.

Study of CXCL9-11 gene polymorphisms in liver fibrosis among patients with chronic hepatitis C.

Several factors are associated with the progression of chronic hepatitis C: comorbidities, lifestyle, and pathogenic factors, including immune response, apoptosis and heredity. Single nucleotide polymorphisms (SNPs) in the PNPLA3 and TM6SF2 genes are more widely studied genetic risk factors, while CXCL9-11 chemokines produced by hepatocytes in the process of infection are less well studied. Our aim was to evaluate the influence of CXCL9 rs10336, CXCL10 rs3921 and CXCL11 rs4619915 in liver fibrosis when analysed together with PNPLA3 rs738409 and TM6SF2 rs58542926. The study included 219 patients with chronic hepatitis C. SNP genotyping was performed by real-time PCR. Univariate and multivariate analyses were used to detect the association between SNPs and advanced fibrosis in a recessive genetic model. All SNPs had a minimum allele frequency >5%, and CXCL9 rs10336, CXCL10 rs3921 and CXCL11 rs4619915 were in high linkage disequilibrium (D' ≥ 0.84). In the multivariate analysis, we observed that male gender (P = 0.000), older age (P = 0.025), moderate to intense inflammatory activity (P = 0.002), moderate to accentuated hepatic steatosis (P = 0.026) and the CT genotype of the TM6SF2 rs58542926 SNP (P = 0.014) presented significant associations with advanced fibrosis. Overall, the CXCL9 rs10336, CXCL10 rs3921, CXCL11 rs4619915 and PNPLA3 rs738409 SNPs did not influence liver fibrosis among patients with chronic hepatitis C.

Computational Study of C-X-C Chemokine Receptor (CXCR)3 Binding with Its Natural Agonists Chemokine (C-X-C Motif) Ligand (CXCL)9, 10 and 11 and with Synthetic Antagonists: Insights of Receptor Activation towards Drug Design for Vitiligo.

Vitiligo is a hypopigmentary skin pathology resulting from the death of melanocytes due to the activity of CD8+ cytotoxic lymphocytes and overexpression of chemokines. These include CXCL9, CXCL10, and CXCL11 and its receptor CXCR3, both in peripheral cells of the immune system and in the skin of patients diagnosed with vitiligo. The three-dimensional structure of CXCR3 and CXCL9 has not been reported experimentally; thus, homology modeling and molecular dynamics could be useful for the study of this chemotaxis-promoter axis. In this work, a homology model of CXCR3 and CXCL9 and the structure of the CXCR3/Gαi/0ßγ complex with post-translational modifications of CXCR3 are reported for the study of the interaction of chemokines with CXCR3 through all-atom (AA-MD) and coarse-grained molecular dynamics (CG-MD) simulations. AA-MD and CG-MD simulations showed the first activation step of the CXCR3 receptor with all chemokines and the second activation step in the CXCR3-CXCL10 complex through a decrease in the distance between the chemokine and the transmembrane region of CXCR3 and the separation of the ßγ complex from the α subunit in the G-protein. Additionally, a general protein-ligand interaction model was calculated, based on known antagonists binding to CXCR3. These results contribute to understanding the activation mechanism of CXCR3 and the design of new molecules that inhibit chemokine binding or antagonize the receptor, provoking a decrease of chemotaxis caused by the CXCR3/chemokines axis.

Immunosuppressive therapies after intestinal transplant modulate the expression of Th1 signature genes during acute cellular rejection. Implications in the search for rejection biomarkers.

BACKGROUND AND AIMS: Acute cellular rejection (ACR) and infections are leading causes of graft loss and death in intestinal transplant patients. Our aim was to evaluate the impact of maintenance immunosuppressive therapies on the expression of pro-inflammatory mediators in small bowel at ACR diagnosis. MATERIALS AND METHODS: We analyzed expression levels of Th1-associated genes, IFNG, CXCL10, and CXCL11 by qPCR in 46 selected graft biopsies unequivocally assigned to mild ACR (n = 14) or normal histopathology and clinical condition (n = 32) from 15 patients receiving two different immunosuppressive (IS) schemes. Double treatment: corticosteroids and tacrolimus (n = 17) and triple treatment: sirolimus or mycophenolate mofetil in addition to the basal therapy (n = 29). RESULTS: IFNG, CXCL10, and CXCL11 were induced during rejection (p < 0.05; p < 0.005, and p < 0.05, respectively). However, when rejection and control groups were classified according to immunosuppressive treatment, in the rejection group, significant differences of IFNG, CXCL10, and CXCL11 expression (p < 0.001; p < 0.005, and 0.01, respectively) were detected, whereas no differences were observed in the control group. CONCLUSION: Gene expression of Th1 response mediators is higher during ACR. Triple IS group showed significantly lower expression of pro-inflammatory Th1 mediators during mild ACR indicating that use of these markers to monitor rejection can be affected by the IS treatment used.

Role of CXCR3/CXCL10 axis in immune cell recruitment into the small intestine in celiac disease.

Lymphocytic infiltration in the lamina propria (LP), which is primarily composed of CD4(+) Th1 cells and plasma cells, and increased numbers of intraepithelial lymphocytes (IELs), is a characteristic finding in active celiac disease (CD). Signals for this selective cell recruitment have not been fully established. CXCR3 and its ligands, particularly CXCL10, have been suggested to be one of the most relevant pathways in the attraction of cells into inflamed tissues. In addition, CXCR3 is characteristically expressed by Th1 cells. The aim of this work was to investigate the participation of the chemokine CXCL10/CXCR3 axis in CD pathogenesis. A higher concentration of CXCL10 was found in the serum of untreated CD patients. The mRNA levels of CXCL10 and CXCL11 but not CXCL9 were significantly higher in duodenal biopsies from untreated CD patients compared with non-CD controls or treated patients. The results demonstrate that CXCL10 is abundantly produced in untreated CD and reduced in treated patients, and the expression of CXCL10 was found to be correlated with the IFNγ levels in the tissue. Plasma cells and enterocytes were identified as CXCL10-producing cells. Moreover, the CXCL10 expression in intestinal tissues was upregulated by poly I:C and IL-15. IELs, LP T lymphocytes, and plasma cells, which infiltrate the intestinal mucosa in untreated CD, express CXCR3. The CXCR3/CXCL10 signalling axis is overactivated in the small intestinal mucosa in untreated patients, and this finding explains the specific recruitment of the major cell populations that infiltrate the epithelium and the LP in CD.

A gene expression profile related to immune dampening in the tumor microenvironment is associated with poor prognosis in gastric adenocarcinoma.

BACKGROUND: The TNM Classification of Malignant Tumours (TNM) staging system is the primary means of determining a prognosis for gastric adenocarcinoma (GC). However, tumor behavior in the individual patient is unpredictable and in spite of treatment advances, a classification of 'advanced stage' still portends a poor prognosis. Thus, further insights from molecular analyses are needed for better prognostic stratification and determination of new therapeutic targets. METHODS: A total of fifty-one fresh frozen tumor samples from patients with histopathologically confirmed diagnoses of GC, submitted to surgery with curative intent, were included in the study. Total RNA was extracted from an initial group of fifteen samples matched for known prognostic factors, categorized into two subgroups, according to patient overall survival: poor (<24 months) or favorable (at or above 24 months), and hybridized to Affymetrix Genechip human genome U133 plus 2.0 for genes associated with prognosis selection. Thirteen genes were selected for qPCR validation using those initial fifteen samples plus additional thirty-six samples. RESULTS: A total of 108 genes were associated with poor prognosis, independent of tumor staging. Using systems biology, we suggest that this panel reflects the dampening of immune/inflammatory response in the tumor microenvironment level and a shift to Th2/M2 activity. A gene trio (OLR1, CXCL11 and ADAMDEC1) was identified as an independent marker of prognosis, being the last two markers validated in an independent patient cohort. CONCLUSIONS: We determined a panel of three genes with prognostic value in gastric cancer, which should be further investigated. A gene expression profile suggestive of a dysfunctional inflammatory response was associated with unfavorable prognosis.

Expression analysis of wound healing genes in human periapical granulomas of progressive and stable nature.

INTRODUCTION: Wound healing process involves the activation of extracellular matrix components, remodeling enzymes, cellular adhesion molecules, growth factors, cytokines and chemokines genes. However, the molecular patterns underlying the healing process at the periapical environment remain unclear. Here we hypothesized that endodontic infection might result in an imbalance in the expression of wound healing genes involved in the pathogenesis of periapical lesions. Furthermore, we suggest that differential expression of wound healing markers in active and latent granulomas could account for different clinical outcomes for such lesions. METHODS: Study samples consisted of 93 periapical granulomas collected after endodontic surgeries and 24 healthy periodontal ligament tissues collected from premolars extracted for orthodontic purposes as control samples. Of these, 10 periapical granulomas and 5 healthy periapical tissues were used for expression analysis of 84 wound healing genes by using a pathway-specific real-time polymerase chain reaction array. The remaining 83 granulomas and all 24 control specimens were used to validate the obtained array data by real-time polymerase chain reaction. Observed variations in expression of wound healing genes were analyzed according to the classification of periapical granulomas as active/progressive versus inactive/stable (as determined by receptor activator for nuclear factor kappa B ligand/osteoprotegerin expression ratio). RESULTS: We observed a marked increase of 5-fold or greater in SERPINE1, TIMP1, COL1A1, COL5A1, VTN, CTGF, FGF7, TGFB1, TNF, CXCL11, ITGA4, and ITGA5 genes in the periapical granulomas when compared with control samples. SERPINE1, TIMP1, COL1A1, TGFB1, and ITGA4 mRNA expression was significantly higher in inactive compared with active periapical granulomas (P < .001), whereas TNF and CXCL11 mRNA expression was higher in active lesions (P < .001). CONCLUSIONS: The identification of novel gene targets that curb the progression status of periapical lesions might contribute to a more accurate diagnosis and lead to treatment modalities more conducive to endodontic success.