Background: The identification of endometrial stem/progenitor cells in a high turnover rate tissue suggests that a well-orchestrated underlying network controls the behaviour of these stem cells. The thickness of the endometrium can grow from 0.5-1 mm to 5-7 mm within a week indicating the need of stem cells for self-renewal and differentiation during this period. The cyclical regeneration of the endometrium suggests specific signals can activate the stem cells during or shortly after menstruation.

Methods: Endometrial mesenchymal stem-like cells (eMSCs) were cocultured with endometrial epithelial or stromal cells from different phases of the menstrual cycle; the clonogenicity and the phenotypic expression of eMSC markers (CD140b and CD146) were assessed. The functional role of WNT/β-catenin signalling on eMSC was determined by western blot analysis, immunofluorescent staining, flow cytometry, quantitative real-time PCR and small interfering RNA. The cytokine levels in the conditioned medium of epithelial or stromal cells cocultured with eMSCs were evaluated by enzyme-linked immunosorbent assays.

Results: Coculture of endometrial cells (epithelial or stromal) from the menstrual phase enhanced the clonogenicity and self-renewal activities of eMSCs. Such phenomenon was not observed in niche cells from the proliferative phase. Coculture with endometrial cells from the menstrual phase confirmed an increase in expression of active β-catenin in the eMSCs. Treatment with IWP-2, a WNT inhibitor, suppressed the observed effects. Anti-R-spondin-1 antibody reduced the stimulatory action of endometrial niche cells on WNT/β-catenin activation in the T cell factor/lymphoid enhancer-binding factor luciferase reporter assay. Moreover, the mRNA level and protein immunoreactivities of leucine-rich repeat-containing G-protein coupled receptor 5 were higher in eMSCs than unfractionated stromal cells. Conditioned media of endometrial niche cells cocultured with eMSCs contained increased levels of C-X-C motif ligand 1 (CXCL1), CXCL5 and interleukin 6. Treatment with these cytokines increased the clonogenic activity and phenotypic expression of eMSCs.

Conclusions: Our findings indicate a role of WNT/β-catenin signalling in regulating activities of endometrial stem/progenitor cells during menstruation. Certain cytokines at menstruation can stimulate the proliferation and self-renewal activities of eMSCs. Understanding the mechanism in the regulation of eMSCs may contribute to treatments of endometrial proliferative disorders such as Asherman's syndrome.

Keywords: Cytokines; Endometrium; LGR5; Menstruation; RSPO1; Stem cells; WNT signalling.

The use of in vitro systems that allow efficient selection of probiotic candidates with immunomodulatory properties could significantly minimize the use of experimental animals. In this work, we generated an in vitro immunoassay system based on porcine intestinal epithelial (PIE) cells and dextran sodium sulfate (DSS) administration that could be useful for the selection and characterization of potential probiotic strains to be used in inflammatory bowel disease (IBD) patients. Our strategy was based on two fundamental pillars: on the one hand, the capacity of PIE cells to create a monolayer by attaching to neighboring cells and efficiently mount inflammatory responses and, on the other hand, the use of two probiotic bifidobacteria strains that have been characterized in terms of their immunomodulatory capacities, particularly in mouse IBD models and patients. Our results demonstrated that DSS administration can alter the epithelial barrier created in vitro by PIE cells and induce a potent inflammatory response, characterized by increases in the expression levels of several inflammatory factors including TNF-α, IL-1α, CCL4, CCL8, CCL11, CXCL5, CXCL9, CXCL10, SELL, SELE, EPCAM, VCAM, NCF2, and SAA2. In addition, we demonstrated that Bifidobacterium breve M-16V and B. longum BB536 are able to regulate the C-jun N-terminal kinase (JNK) intracellular signalling pathway, reducing the DSS-induced alterations of the in vitro epithelial barrier and differentially regulating the inflammatory response in a strain-dependent fashion. The good correlation between our in vitro findings in PIE cells and previous studies in animal models and IBD patients shows the potential value of our system to select new probiotic candidates in an efficient way.

Keywords: Bifidobacteria; Epithelial barrier; IBD; In vitro immunoassay; Inflammation; Probiotics.

Background: Immune checkpoint blockers (ICBs) are increasingly applied to treat patients with advanced HCC. However, the overall survival (OS) of HCC patients is still unsatisfactory, and there is no confirmed immune-related and prognostic gene to identify patients who could clinically benefit from this treatment. The tumor microenvironment (TME) is known to be closely related to immunotherapy and plays a pivotal role in the recurrence and progression of HCC. Our aim is to explore TME-related genes and identify the prognostic value in HCC patients.

Methods: ESTIMATE, immune, and stromal scores were calculated for HCC patients based on RNA expression data from The Cancer Genome Atlas database. Differential expression analysis was performed to screen the differentially expressed genes (DEGs). A protein-protein interaction (PPI) network was constructed to identify the key DEGs. Univariate and multivariate Cox analyses were adopted to validate hub DEGs associated with clinical prognosis, and a single-sample gene set enrichment analysis (ssGSEA) algorithm was used to dissect the landscape of tumor-infiltrating cells (TIC) in HCC. Finally, the relationship between hub immune-related genes and TIC was explored through difference and correlation analyses.

Results: ESTIMATE, immune and stromal scores were all found to be associated with the OS of patients (P<0.05). A total of 1,112 DEGs were identified by comparing low and high score groups of immune and stromal scores. Most of DEGs were enriched in immune-related gene sets by Gene Ontology (GO) and Kyoto Encyclopedia of Genes and Genomes (KEGG) analyses. Additionally, the top 34 genes were included in the protein-protein interaction (PPI) network, and univariate Cox analysis focus on a novel prognosis-related gene cluster CXCL5/CXCL8 (P<0.001). Regarding the immune landscape of HCC, univariable Cox regression analysis showed six immune cells to be associated with OS. Finally, 21 immune cells were commonly determined between high and low expression of CXCL5/CXCL8, suggesting there is a close relationship between expression of CXCL5 and CXCL8 .

Conclusions: Our study has revealed that the immune-related gene cluster of CXCL5 /CXCL8 could be a promising prognostic indicator for HCC and a potential novel biomarker to guide the selection of HCC patients for ICB immunotherapy.

Keywords: CXCL5; CXCL8; Hepatocellular carcinoma (HCC); tumor microenvironment (TME).

Pathologic findings showed that neutrophils played an important role in the pathogenesis of NMO. This study aims to investigate the CSF levels of neutrophil-related chemokines in NMO. CXCL1, CXCL5, and CXCL7 were measured in 95 patients with NMO, 15 patients with MS, 18 patients with GFAP astrocytopathy, and 16 controls. The CSF level of CXCL1, CXCL5, and CXCL7 was significantly elevated in the NMO group but not correlated with the patient clinical severity. Besides, the CSF CXCL1, CXCL5, and CXCL7 could act as biomarkers to distinguish NMO from MS with good reliability, especially the CXCL7.

Keywords: CXCL1; CXCL5; CXCL7; neuromyelitis optica; neutrophil-related chemokines.

A deficiency in cystic fibrosis transmembrane conductance regulator (CFTR) function in CF leads to chronic lung disease. CF is associated with abnormalities in fatty acids, ceramides, and cholesterol, their relationship with CF lung pathology is not completely understood. Therefore, we examined the impact of CFTR deficiency on lipid metabolism and pro-inflammatory signaling in airway epithelium using mass spectrometric, protein array. We observed a striking imbalance in fatty acid and ceramide metabolism, associated with chronic oxidative stress under basal conditions in CF mouse lung and well-differentiated bronchial epithelial cell cultures of CFTR knock out pig and CF patients. Cell-autonomous features of all three CF models included high ratios of ω-6- to ω-3-polyunsaturated fatty acids and of long- to very long-chain ceramide species (LCC/VLCC), reduced levels of total ceramides and ceramide precursors. In addition to the retinoic acid analog fenretinide, the anti-oxidants glutathione (GSH) and deferoxamine partially corrected the lipid profile indicating that oxidative stress may promote the lipid abnormalities. CFTR-targeted modulators reduced the lipid imbalance and oxidative stress, confirming the CFTR dependence of lipid ratios. However, despite functional correction of CF cells up to 60% of non-CF in Ussing chamber experiments, a 72-h triple compound treatment (elexacaftor/tezacaftor/ivacaftor surrogate) did not completely normalize lipid imbalance or oxidative stress. Protein array analysis revealed differential expression and shedding of cytokines and growth factors from CF epithelial cells compared to non-CF cells, consistent with sterile inflammation and tissue remodeling under basal conditions, including enhanced secretion of the neutrophil activator CXCL5, and the T-cell activator CCL17. However, treatment with antioxidants or CFTR modulators that mimic the approved combination therapies, ivacaftor/lumacaftor and ivacaftor/tezacaftor/elexacaftor, did not effectively suppress the inflammatory phenotype. We propose that CFTR deficiency causes oxidative stress in CF airway epithelium, affecting multiple bioactive lipid metabolic pathways, which likely play a role in CF lung disease progression. A combination of anti-oxidant, anti-inflammatory and CFTR targeted therapeutics may be required for full correction of the CF phenotype.

Keywords: bronchial epithelial cell; ceramide species; cystic fibrosis; cystic fibrosis transmembrane conductance regulator corrector therapy; cytokine array; lipidomics; oxidative stress; polyunsaturated (essential) fatty acids.

Lewisite is a strong vesicating and chemical warfare agent. Because of the rapid transdermal absorption, cutaneous exposure to lewisite can also elicit severe systemic injury. Lewisite (2.5, 5.0, and 7.5 mg/kg) was applied to the skin of Ptch1^+/- /SKH-1 mice and acute lung injury (ALI) was assessed after 24 hours. Arterial blood gas measurements showed hypercapnia and hypoxemia in the lewisite-exposed group. Histological evaluation of lung tissue revealed increased levels of proinflammatory neutrophils and a dose-dependent increase in structural changes indicative of injury. Increased inflammation was also confirmed by altered expression of cytokines, including increased IL-33, and a dose-dependent elevation of CXCL1, CXCL5, and GCSF was observed in the lung tissue. In the bronchoalveolar lavage fluid of lewisite-exposed animals, there was a significant increase in HMGB1, a damage-associated molecular pattern molecule, as well as elevated CXCL1 and CXCL5, which coincided with an influx of neutrophils to the lungs. Complete blood cell analysis revealed eosinophilia and altered neutrophil-lymphocyte ratios as a consequence of lewisite exposure. Mean platelet volume and RBC distribution width, which are predictors of lung injury, were also increased in the lewisite group. These data demonstrate that cutaneous lewisite exposure causes ALI and may contribute to mortality in exposed populations.

Background and aims Recent research indicates a previously unknown low-grade systemic or neurogenic inflammation in groups of chronic pain (CP) patients. Low-grade inflammation may have an important role in symptoms that have previously not been well depicted: widespread pain, tiredness and cognitive dysfunctions frequently seen in severely impaired CP patients. This study aimed to investigate the plasma inflammatory profile in a group of very complex CP patients at baseline and at a 1-year follow-up after participation in a cognitive behavior therapy (CBT)-based multimodal pain rehabilitation program (PRP). Methods Blood samples were collected from 52 well-characterized CP patients. Age- and sex-matched healthy blood donors served as controls. The samples were analyzed with a multiple Proximal Extension Analysis allowing a simultaneous analysis of 92 inflammation-related proteins consisting mainly of cytokines, chemokines and growth-factors. At follow-up, 1-year after participation in the RPR samples from 28 patients were analyzed. The results were confirmed by a multi-array technology that allows quantitative estimation. Results Clear signs of increased inflammatory activity were detected in the CP patients. Accepting a false discovery rate (FDR) of 5%, there were significant differences in 43/92 inflammatory biomarkers compared with the controls. In three biomarkers (CXCL5, SIRT2, AXIN1) the expression levels were elevated more than eight times. One year after the PRP, with the patients serving as their own controls, a significant decrease in overall inflammatory activity was found. Conclusions Our results indicate that the most impaired CP patients suffer from low-grade chronic systemic inflammation not described earlier with this level of detail. The results may have implications for a better understanding of the cluster of co-morbid symptoms described as the "sickness-syndrome" and the wide-spread pain seen in this group of patients. The decrease in inflammatory biomarkers noted at the follow-up after participation in the PRP may reflect the positive effects obtained on somatic and psycho-social mechanisms involved in the inflammatory process by a rehabilitation program. Besides the PRP, no major changes in medication or lifestyle factors were implemented during the same period. To our knowledge, this is the first study reporting that a PRP may induce inflammatory-reducing effects. Further studies are needed to verify the objective findings in CP patients and address the question of causality that remains to be solved. Implications The findings offer a new insight into the complicated biological processes underlying CP. It may have implications for the understanding of symptoms collectively described as the "sickness-syndrome" - frequently seen in this group of patients. The lowering of cytokines after the participation in a PRP indicate a new way to evaluate this treatment; by measuring inflammatory biomarkers.

Senescent thymic stromal cells (TSCs) producing senescence-associated secretory phenotype (SASP) may play a role at later phases of thymic involution. However, the etiology and mechanisms responsible for TSC senescence remain to be elucidated. In the present study, the effects of oxidative stress on TSCs and role of miRNA-146a-5p in stress-induced premature senescence (SIPS) were identified. D-galactose (D-gal) induced oxidative stress in primary TSCs and a limited cumulative oxidative stress induced premature senescence but not apoptosis of TSCs. miRNA-146a-5p overexpression can mitigate the SIPS by targeting tumor necrosis factor receptor-associated factor 6 (TRAF6) instead of increasing autophagy clearance. Furthermore, exogenous miRNA-146a-5p reversed the upregulation of chemokines including Cxcl5, pro-inflammatory cytokines, and antimicrobial peptides in TSCs with SIPS. In conclusion, the accumulated oxidative stress may be partially responsible for senescence in TSCs and modulation of miRNA-146a-5p may attenuate this process.

Keywords: D-galactose; Oxidative stress; Premature senescence; TSCs; miRNA-146a-5p.

Background. Despite considerable interest in the search for a spinal cord injury (SCI) therapy, there is a critical need to develop a panel of diagnostic biomarkers to determine injury severity. In this regard, there is a requirement for continuing research into the fundamental processes of neuroinflammatory and autoimmune reactions in SCI, identifying changes in the expression of cytokines. Methods. In this pilot study, an extended multiplex analysis of the cytokine profiles in the serum of patients at 2 weeks post-SCI (n = 28) was carried out, together with an additional assessment of neuron-specific enolase (NSE) and vascular endothelial growth factor (VEGF) levels by enzyme-linked immunosorbent assay. A total of 16 uninjured subjects were enrolled as controls. Results. The data obtained showed a large elevation of IFNγ (>52 fold), CCL27 (>13 fold), and CCL26 (>8 fold) 2 weeks after SCI. The levels of cytokines CXCL5, CCL11, CXCL11, IL10, TNFα, and MIF were different between patients with baseline American Spinal Injury Association Impairment Scale (AIS) grades of A or B, whilst IL2 (>2 fold) and MIP-3a (>6 fold) were significantly expressed in the cervical and thoracic regions. There was a trend towards increasing levels of NSE. However, the difference in NSE was lost when the patient set was segregated based on AIS group. Conclusions. Our pilot research demonstrates that serum concentrations of cytokines can be used as an affordable and rapid detection tool to accurately stratify SCI severity in patients.

Keywords: blood serum; clinical trial; cytokine profile; inflammation; traumatic spinal cord injury.

One of the most prominent features of glioblastoma (GBM) is hyper-vascularization. Bone marrow-derived macrophages are actively recruited to the tumor and referred to as glioma-associated macrophages (GAMs) which are thought to provide a critical role in tumor neo-vascularization. However, the mechanisms by which GAMs regulate endothelial cells (ECs) in the process of tumor vascularization and response to anti-angiogenic therapy (AATx) is not well-understood. Here we show that GBM cells secrete IL-8 and CCL2 which stimulate GAMs to produce TNFα. Subsequently, TNFα induces a distinct gene expression signature of activated ECs including VCAM-1, ICAM-1, CXCL5, and CXCL10. Inhibition of TNFα blocks GAM-induced EC activation both in vitro and in vivo and improve survival in mouse glioma models. Importantly we show that high TNFα expression predicts worse response to Bevacizumab in GBM patients. We further demonstrated in mouse model that treatment with B20.4.1.1, the mouse analog of Bevacizumab, increased macrophage recruitment to the tumor area and correlated with upregulated TNFα expression in GAMs and increased EC activation, which may be responsible for the failure of AATx in GBMs. These results suggest TNFα is a novel therapeutic that may reverse resistance to AATx. Future clinical studies should be aimed at inhibiting TNFα as a concurrent therapy in GBMs.

Keywords: Anti-angiogenic therapy; Endothelial cells activation; Glioblastoma; Glioma-associated macrophages; TNFα.

The ischemia-reperfusion (I/R)-caused damage in human brain microvascular endothelial cells (BMECs) is associated with the disruption of blood-brain barrier (BBB). CXC chemokine ligand 5 (CXCL5) is reported upregulated in ischemia stroke. However, the detailed function of CXCL5 in this pathological process remains largely unclear. To further analyze the function of CXCL5 in ischemia stroke, we constructed oxygen-glucose deprivation (OGD) model on human BMECs to mimic ischemic stroke condition in vitro. Cell proliferation was analyzed using cell counting kit-8 (CCK-8) assay. qRT-PCR and western blot were utilized to determine gene expression. The barrier function of BMECs was assessed using fluorescently labeled dextran (FITC-Dextran) assay and Trans-epithelial/endothelial electrical resistance (TEER) technique. Our results indicated that CXCL5 antibody (anti-CXCL5) promoted the proliferation of model cells, whereas reduced its permeability. Moreover, the TEER value of model cells was enhanced in the presence of anti-CXCL5. Therefore, these findings demonstrated that CXCL5 silencing attenuated the ischemic/hypoxic-induced injury in human BMECs. Importantly, human recombinant protein CXCL5 (Re-CXCL5) deeply disrupted the function of BMECs in normoxic condition. Furthermore, p38 inhibitor SB203580 significantly abolished the function of CXCL5 in model cells. Much importantly, the similar results were also obtained in BMECs with normoxic conditions in the presence of Re-CXCL5. These results indicated CXCL5 might regulate the function of BMECs via mediating p38 pathway. This investigation not only enhanced the understanding into the biological effect of CXCL5 in human BMECs with ischemic/hypoxic conditions but also indicated its potential value as a therapeutic target for ischemic-induced brain disease. This article is protected by copyright. All rights reserved.

Keywords: BBB; BMECs; CXCL5; ischemic/hypoxic; p38.

Sex steroid hormones have been reported to induce inflammation causing dysregulation of cytokines in prostate cancer cells. However, the underlying epigenetic mechanism has not well been studied. The objective of this study was to evaluate the effect of sex steroid hormones on epigenetic DNA methylation changes in prostate cancer cells using a signature PCR methylation array panel that correspond to 96 genes with biological function in the human inflammatory and autoimmune signals in prostate cancer. Of the 96-gene panel, 32 genes showed at least 10% differentially methylation level in response to hormonal treatment when compared to untreated cells. Genes that were hypomethylated included CXCL12, CXCL5, CCL25, IL1F8, IL13RAI, STAT5A, CXCR4 and TLR5; and genes that were hypermethylated included ELA2, TOLLIP, LAG3, CD276 and MALT1. Quantitative RT-PCR analysis of select genes represented in a cytokine expression array panel showed inverse association between DNA methylation and gene expression for TOLLIP, CXCL5, CCL18 and IL5 genes and treatment of prostate cancer cells with 5'-aza-2'-deoxycytidine with or without trichostatin A induced up-regulation of TOLLIP expression. Further analysis of relative gene expression of matched prostate cancer tissues when compared to benign tissues from individual patients with prostate cancer showed increased and significant expression for CCL18 (2.6-fold; p<0.001), a modest yet significant increase in IL5 expression (1.17-fold; p=0.015), and a modest increase in CXCL5 expression (1.4-fold; p=0.25). In conclusion, our studies demonstrate that sex steroid hormones can induce aberrant gene expression via differential methylation changes in prostate carcinogenesis.

Background: RNF43 is an E3 ubiquitin ligase that is recurrently mutated in pancreatic ductal adenocarcinoma (PDAC) and precursor cystic neoplasms of the pancreas. The impact of RNF43 mutations on PDAC is poorly understood and autochthonous models have not been sufficiently characterized. In this study we describe a genetically engineered mouse model (GEMM) of PDAC with conditional expression of oncogenic Kras and deletion of the catalytic domain of Rnf43 in exocrine cells.

Methods: We generated Ptf1a-Cre;LSL-KrasG12D;Rnf43^flox/flox (KRC) and Ptf1a-Cre; LSL-KrasG12D (KC) mice and animal survival was assessed. KRC mice were sacrificed at 2 months, 4 months and at moribund status followed by analysis of pancreata by single cell RNA sequencing (scRNAseq). Comparative analyses between moribund KRC and a moribund Kras/Tp53 driven PDAC GEMM (KPC) was performed. Cell lines were isolated from KRC and KC tumors and interrogated by cytokine array analyses, ATAC-seq and in vitro drug assays. KRC GEMMs were also treated with an anti-CTLA4 neutralizing antibody with treatment response measured by magnetic response imaging.

Results: We demonstrate that KRC mice display a marked increase in incidence of high-grade cystic lesions of the pancreas and PDAC compared to KC. Importantly, KRC mice have a significantly decreased survival compared to KC mice. By use of scRNAseq we demonstrated that KRC tumor progression is accompanied by a decrease in macrophages, as well as an increase in T and B lymphocytes with evidence of increased immune checkpoint molecule expression and affinity maturation, respectively. This was in stark contrast to the tumor immune microenvironment observed in the KPC PDAC GEMM. Furthermore, expression of the chemokine, CXCL5, was found to be specifically decreased in KRC cancer cells by means of epigenetic regulation and emerged as a putative candidate for mediating the unique KRC immune landscape.

Conclusions: The KRC GEMM establishes RNF43 as a bona fide tumor suppressor gene in PDAC. This GEMM features a markedly different immune microenvironment compared to previously reported PDAC GEMMs and puts forth a rationale for an immunotherapy approach in this subset of PDAC cases.

Keywords: KRAS; genetically engineered mouse models; pancreatic cancer; single cell RNA sequencing; tumor suppressor gene.

OBJECTIVE

This study aimed to investigate whether plasma levels of four adipokines (chemerin, macrophage migratory inhibitory factor [MIF], visceral adipose tissue-derived serine protease inhibitor [vaspin] and chemokine CXCL5) are associated with the presence of obstructive sleep apnea syndrome (OSAS) in patients.

METHODS

A total of 58 male patients with OSAS and 16 healthy male subjects were enrolled in this study.

RESULTS

Four plasma adipokines (chemerin, MIF, vaspin and chemokine CXCL5) were significantly higher (P < 0.05) in severe OSAS patients than in the control group after polysomnography. Plasma levels of these four adipokines were higher (P < 0.05) after sleep than before sleep. These levels were also associated with anthropometric measurements for BMI, neck circumference, body fat percentage, sleep parameters including the apnea/hypopnea index (AHI) and minimum SaO2 %. Multiple regression analyses showed that BMI, AHI and mean SaO2 % were major factors affecting the four plasma adipokine levels in OSAS patients.

CONCLUSIONS

Plasma chemerin, MIF, vaspin and chemokine CXCL5 levels were severely elevated with OSAS, and were also connected with obesity.

Staphylococcus aureus is a skin commensal microorganism commonly colonizing healthy humans. Nevertheless, S. aureus can also be responsible for cutaneous infections and contribute to flare-up of inflammatory skin diseases such as atopic dermatitis (AD), which is characterized by dysbiosis of the skin microbiota with S. aureus as the predominant species. However, the role of major virulence factors of this pathogen such as phenol-soluble modulin (PSM) toxins in epidermal inflammation remains poorly understood. Stimulation of primary human keratinocytes with sublytic concentrations of synthetic and purified PSM α3 resulted in upregulation of a large panel of pro-inflammatory chemokine and cytokine gene expression, including CXCL1, CXCL2, CXCL3, CXCL5, CXCL8, CCL20, IL-1α, IL-1β, IL-6, IL-36γ and TNF-α, while inducing the release of CXCL8, CCL20, TNF-α and IL-6. In addition, using S. aureus culture supernatant from mutants deleted from genes encoding either α-type PSMs or all PSM production, PSMs were shown to be the main factors of S. aureus secretome responsible for pro-inflammatory mediator induction in human keratinocytes. On the other hand, α-type PSM-containing supernatant triggered an intense induction of pro-inflammatory mediator expression and secretion during both topical and basal layer stimulation of an ex vivo model of human skin explants, a physiologically relevant model of pluristratified epidermis. Taken together, the results of this study show that PSMs and more specifically α-type PSMs are major virulence factors of S. aureus inducing a potent inflammatory response during infection of the human epidermis and could thereby contribute to AD flare-up through exacerbation of skin inflammation.

Keywords: PSM α3; Staphylococcus aureus; atopic dermatitis; epidermis; inflammatory response; keratinocyte; phenol soluble modulins; toxins.

This study is primarily aimed at assessing serum changes on a large panel of proteins in patients with chronic back pain following spa therapy, as well as evaluating different spa therapy regimens as a preliminary exploratory clinical study. Sixty-six patients with chronic back pain secondary to osteoarthritis were randomly enrolled and treated with daily mud packs and bicarbonate-alkaline mineral water baths, or a thermal hydrotherapy rehabilitation scheme, the combination of the two regimens or usual medication only (control group), for two weeks. Clinical variables were evaluated at baseline, after 2 and 12 weeks. One thousand serum proteins were tested before and after a two-week mud bath therapy. All spa treatment groups showed clinical benefit as determined by improvements in VAS pain, Roland Morris disability questionnaire and neck disability index at both time points. The following serum proteins were found greatly increased (≥2.5 fold) after spa treatment: inhibin beta A subunit (INHBA), activin A receptor type 2B (ACVR2B), angiopoietin-1 (ANGPT1), beta-2-microglobulin (B2M), growth differentiation factor 10 (GDF10), C-X-C motif chemokine ligand 5 (CXCL5), fibroblast growth factor 2 (FGF2), fibroblast growth factor 12 (FGF12), oxidized low density lipoprotein receptor 1 (OLR1), matrix metallopeptidase 13 (MMP13). Three proteins were found greatly decreased (≤0.65 fold): apolipoprotein C-III (Apoc3), interleukin 23 alpha subunit p19 (IL23A) and syndecan-1 (SDC1). Spa therapy was confirmed as beneficial for chronic back pain and proved to induce changes in proteins involved in functions such as gene expression modulation, differentiation, angiogenesis, tissue repair, acute and chronic inflammatory response.

The C-X-C motif (CXC) chemokines play an important role in inflammatory processes and angiogenesis and are also associated with tumor development, progression and metastasis. They can be either promoting or inhibiting factors in colorectal cancers (CRC). The expression patterns and prognostic values of the CXC family still need further investigation. In this study, we investigated data related to transcription, translation, survival and tumor immune infiltration for CXC chemokines in patients with CRC from the ONCOMINE, GEPIA, cBioPortal, HPA and TIMER databases. We found that the expression levels of CXCL1-3, CXCL5, and CXCL8 were higher in CRC tissues than in colorectal tissues. Expression among stages significantly varied for CXCL1-3 and CXCL9-11. The survival analysis revealed that high transcriptional levels of CXCL4 and CXCL9-11 could serve as positive prognostic factors for patients with CRC. CXCL9-11 were highly associated with CD8+ T cells and natural killer (NK) cells in the tumor immune infiltration analysis, indicating their role in the antitumor immune response. This study implies that CXCL1-3, CXCL5, and CXCL8 are important factors during CRC oncogenesis and that CXCL9-11 could be new biomarkers for the prognosis of CRC.

Keywords: Chemokines; Colorectal adenocarcinoma; Prognosis; Tumor-infiltrating immune cells.

Monocytes are among the major myeloid cells that respond to Toxoplasma, a ubiquitous foodborne that infects ≥ 1 billion people worldwide, in human peripheral blood. As such, a molecular understanding of human monocyte-Toxoplasma interactions can expedite the development of novel human toxoplasmosis control strategies. Current molecular studies on monocyte-Toxoplasma interactions are based on average cell or parasite responses across bulk cell populations. Although informative, population-level averages of monocyte responses to Toxoplasma have sometimes produced contradictory results, such as whether CCL2 or IL12 define effective monocyte responses to the parasite. Here, we used single-cell dual RNA sequencing (scDual-Seq) to comprehensively define, for the first time, the monocyte and parasite transcriptional responses that underpin human monocyte-Toxoplasma encounters at the single cell level. We report extreme transcriptional variability between individual monocytes. Furthermore, we report that Toxoplasma-exposed and unexposed monocytes are transcriptionally distinguished by a reactive subset of CD14⁺CD16^- monocytes. Functional cytokine assays on sorted monocyte populations show that the infection-distinguishing monocytes secrete high levels of chemokines, such as CCL2 and CXCL5. These findings uncover the Toxoplasma-induced monocyte transcriptional heterogeneity and shed new light on the cell populations that largely define cytokine and chemokine secretion in human monocytes exposed to Toxoplasma.

The primary function of apolipoprotein E (APOE) is to mediate the transport of cholesterol- and lipid-containing lipoprotein particles into cells by receptor-mediated endocytosis. APOE also has pro- and anti-inflammatory effects, which are both context- and concentration-dependent. For example, Apoe-/- mice have enhanced airway remodeling and hyperreactivity in experimental asthma, whereas increased APOE levels in lung epithelial lining fluid induce IL-1β secretion from human asthmatic alveolar macrophages. However, APOE-mediated airway epithelial cell inflammatory responses and signaling pathways have not been defined. Here, RNA-sequencing of human asthmatic bronchial brushing cells stimulated with APOE identified the increased expression of mRNA transcripts encoding multiple pro-inflammatory genes, including CXCL5 (C-X-C Motif Chemokine Ligand 5), an epithelial-derived chemokine that promotes neutrophil activation and chemotaxis. We subsequently characterized the APOE signaling pathway that induces CXCL5 secretion by human asthmatic small airway epithelial cells (SAECs). Neutralizing antibodies directed against TLR4, but not TLR2, attenuated APOE-mediated CXCL5 secretion by human asthmatic SAECs. Inhibition of TAK1, IκKβ, TPL2, and JNK, but not p38 MAPK nor MEK1/2, attenuated APOE-mediated CXCL5 secretion. The roles for TAK1, IκKβ, TPL2, and JNK in APOE-mediated CXCL5 secretion were verified by RNA-interference. Furthermore, RNA-interference showed that following APOE stimulation, both NF-κB p65 and TPL2 are downstream of TAK1 and IκKβ, while JNK is downstream of TPL2. In summary, elevated levels of APOE in the airway may activate a TLR4/TAK1/IκKβ/NF-κB/TPL2/JNK signaling pathway that induces CXCL5 secretion by human asthmatic SAECs. These findings identify new roles for TLR4 and TPL2 in APOE-mediated pro-inflammatory responses in asthma.

Corticotropin-releasing hormone (CRH) is believed to play a critical role in stress-induced synaptic formation and modification. In the current study, we explored the mechanisms underlying CRH modulation of synaptic formation in the hippocampus by using various models in vitro. In cultured hippocampal slices, CRH treatment decreased synapsin I and postsynaptic density protein 95 (PSD95) levels via CRH receptor type 1 (CRHR1). In isolated hippocampal neurons, however, it increased synapsin I-labeled presynaptic terminals and PSD95-labeled postsynaptic terminals via CRHR1. Interestingly, the inhibitory effect of CRH on synapsin I-labeled and PSD95-labeled terminals occurred in the model of neuron-glia cocultures. These effects were prevented by CRHR1 antagonist. Moreover, treatment of the neurons with the media of CRH-treated glia led to a decrease in synaptic terminal formation. The media collected from CRH-treated glial cells with CRHR1 knockdown did not show an inhibitory effect on synaptic terminals in hippocampal neurons. Unbiased cytokine array coupled with confirmatory enzyme-linked immunosorbent assay revealed that CRH suppressed C-X-C motif chemokine 5 (CXCL5) production in glia via CRHR1. Administration of CXCL5 reversed the inhibitory effects of CRH-treated glia culture media on synaptic formation. Our data suggest that CRH suppresses synapse formation through inhibition of CXCL5 secretion from glia in the hippocampus. Our study indicates that glia-neuron intercommunication is one of the mechanisms responsible for neuronal circuit remodeling during stress.

Cigarette smoking is the main risk factor for chronic obstructive pulmonary disease, and to date, existing pharmacologic interventions have been ineffective at controlling inflammatory processes associated with the disease. To address this issue, we used the Connectivity Map (cMap) database to identify drug candidates with the potential to attenuate cigarette smoke-induced inflammation. We queried cMap using three independent in-house cohorts of healthy nonsmokers and smokers. Potential drug candidates were validated against four publicly available human datasets, as well as six independent datasets from cigarette smoke-exposed mice. Overall, these analyses yielded two potential drug candidates: kaempferol and bethanechol. Subsequently, the efficacy of each drug was validated in vivo in a model of cigarette smoke-induced inflammation. BALB/c mice were exposed to room air or cigarette smoke and treated with each of the two candidate drugs either prophylactically or therapeutically. We found that kaempferol, but not bethanechol, was able to reduce cigarette smoke-induced neutrophilia, both when administered prophylactically and when administered therapeutically. Mechanistically, kaempferol decreased expression of IL-1α and CXCL5 concentrations in the lung. Our data suggest that cMap analyses may serve as a useful tool to identify novel drug candidates against cigarette smoke-induced inflammation.