UNASSIGNED

A prominent pathological feature of neuromyelitis optica spectrum disorders (NMOSD) is markedly greater eosinophilic infiltration than that seen in other demyelinating diseases, like multiple sclerosis (MS). Eosinophils express the chemokine receptor CCR3, which is activated by eotaxins (CCL11/eotaxin-1, CCL24/eotaxin-2, CCL26/eotaxin-3) and CCL13 [monocyte chemoattractant protein (MCP)-4]. Moreover, CCL13 is part of the chemokine set that activates CCR2. The present study aimed to evaluate plasma levels of eotaxins (CCL11, CCL24, and CCL26) and MCPs (CCL13, CCL2, CCL8, and CCL7) in patients with NMOSD during remission.

UNASSIGNED

Healthy controls (HC; n = 30) and patients with MS (n = 47) and NMOSD (n = 58) in remission were consecutively enrolled in this study between January 2016 and August 2017. Plasma CCL11, CCL24, CCL26, CCL2, CCL8, CCL7, CCL13, tumor necrosis factor (TNF)-α, and interleukin (IL)-1β levels were detected using the human cytokine multiplex assay.

UNASSIGNED

Plasma CCL13, CCL11, and CCL26 levels were all significantly higher in patients with NMOSD than in HC and patients with MS. No significant differences were found in the CCL13, CCL11, or CCL26 levels between patients with NMOSD receiving and not receiving immunosuppressive therapy. The plasma levels of TNF-α and IL-1β, which stimulate the above chemokines, were higher in patients with NMOSD than in HC. There was no difference in CCL24 levels among the three groups. In most cases, the CCL7 levels were below the threshold value of the human cytokine multiplex assay, which is in line with other studies. Adjusted multiple regression analyses showed a positive association of CCL13 levels with the number of relapses after controlling gender, age, body mass index, and disease duration in patients with NMOSD.

UNASSIGNED

The study indicates that in NMOSD, the overproduction of cytokines such as IL-1β and TNF-α during remission stimulates eosinophilic chemoattractants such as CCL13, CCL11, and CCL26, which in turn bind to their receptor (CCR3); this could lead to eosinophil hypersensitivity. These findings suggest that the elevated secretion of CCL13, CCL11, and CCL26 may be a critical step in eosinophil recruitment during NMOSD remission.

Bioprinting technology merges engineering and biological fields and together, they possess a great translational potential, which can tremendously impact the future of regenerative medicine and drug discovery. However, the molecular effects elicited by thermal inkjet bioprinting in breast cancer cells remains elusive. Previous studies have suggested that bioprinting can be used to model tissues for drug discovery and pharmacology. We report viability, apoptosis, phosphorylation, and RNA sequence analysis of bioprinted MCF7 breast cancer cells at separate timepoints post-bioprinting. An Annexin A5-FITC apoptosis stain was used in combination with flow cytometry at 2 and 24 h post-bioprinting. Antibody arrays using a Human phospho-MAPK array kit was performed 24 h post-bioprinting. RNA sequence analysis was conducted in samples collected at 2, 7, and 24 h post-bioprinting. The post-bioprinting cell viability averages were 77 and 76% at 24 h and 48 h, with 31 and 64% apoptotic cells at 2 and 24 h after bioprinting. A total of 21 kinases were phosphorylated in the bioprinted cells and 9 were phosphorylated in the manually seeded controls. The RNA seq analysis in the bioprinted cells identified a total of 12,235 genes, of which 9.7% were significantly differentially expressed. Using a ±2-fold change as the cutoff, 266 upregulated and 206 downregulated genes were observed in the bioprinted cells, with the following 5 genes uniquely expressed NRN1L, LUCAT1, IL6, CCL26, and LOC401585. This suggests that thermal inkjet bioprinting is stimulating large scale gene alterations that could potentially be utilized for drug discovery. Moreover, bioprinting activates key pathways implicated in drug resistance, cell motility, proliferation, survival, and differentiation.

Marek's disease virus (MDV), an alpha herpes virus, causes a lymphoproliferative state in chickens known as Marek's disease (MD), resulting in severe monetary losses to the poultry industry. Because lymphocytes of bursa of Fabricius and spleen are prime targets of MDV replication during the early cytolytic phase of infection, the immune response in bursa and spleen should be the foundation of late immunity induced by MDV. However, the mechanism of the MDV-mediated host immune response in lymphocytes in the early stage is poorly understood. The present study is primarily aimed at identifying the crucial genes and significant pathways involved in the immune response of chickens infected with MDV CVI988 and the very virulent RB1B (vvRB1B) strains. Using the RNA sequencing approach, we analyzed the generated transcriptomes from lymphocytes isolated from chicken bursa and spleen. Our findings validated the expression of previously characterized genes; however, they also revealed the expression of novel genes during the MDV-mediated immune response. The results showed that after challenge with CVI988 or vvRB1B strains, 634 and 313 differentially expressed genes (DEGs) were identified in splenic lymphocytes, respectively. However, 58 and 47 DEGs were observed in bursal lymphocytes infected with CVI988 and vvRB1B strains, respectively. Following MDV CVI988 or vvRB1B challenge, the bursal lymphocytes displayed changes in IL-6 and IL-4 gene expression. Surprisingly, splenic lymphocytes exhibited an overwhelming alteration in the expression of cytokines and cytokine receptors involved in immune response signaling. On the other hand, there was no distinct trend between infection with CVI988 and vvRB1B and the expression of cytokines and chemokines, such as IL-10, IFN-γ, STAT1, IRF1, CCL19, and CCL26. However, the expression profiles of IL-1β, IL-6, IL8L1, CCL4 (GGCL1), and CCL5 were significantly upregulated in splenic lymphocytes from chickens infected with CVI988 compared with those of chickens infected with vvRB1B. Because these cytokines and chemokines are considered to be associated with B cell activation and antigenic signal transduction to T cells, they may indicate differences of immune responses initiated by vaccinal and virulent strains during the early phase of infection. Collectively, our study provides valuable data on the transcriptional landscape using high-throughput sequencing to understand the different mechanism between vaccine-mediated protection and pathogenesis of virulent MDV in vivo.

OBJECTIVE

The aim of the article is to describe an immunological reaction to shunt infection in children with hydrocephalus. The main cause of shunt infection involves methicillin resistant Staphylococcus epidermidis (Bhatia et al. Indian J Med Microbiol 35:120-123, 2017; Hayhurst et al. Childs Nerv Syst 24:557-562, 2008; Martínez-Lage et al. Childs Nerv Syst 26: 1795-1798, 2010; Simon et al. PLoS One, 2014; Snowden et al. PLoS One 8:e84089, 2013; Turgut et al. Pediatr Neurosurg 41:131-136, 2005), a bacterial strain which is responsible for the formation of biofilm on contaminated catheters (Snowden et al. PLoS One 8:e84089, 2013; Stevens et al. Br J of Neurosurg 26: 792-797, 2012).

METHODS

The study group involved 30 children with congenital hydrocephalus after shunt system implantation, whose procedures were complicated by S. epidermidis implant infection. Thirty children with congenital hydrocephalus awaiting their first-time shunt implantation formed the control group. The level of eosinophils in peripheral blood was assessed in both groups. Cerebrospinal fluid (CSF) was examined for protein level, pleocytosis, interleukins, CCL26/Eotaxin-3, IL-5, IL-6, CCL11/Eotaxin-1, CCL3/MIP-1a, and MBP. Three measurements were performed in the study group. The first measurement was obtained at the time of shunt infection diagnosis, the second one at the time of the first sterile shunt, and the third one at the time of shunt reimplantation. In the control group, blood and CSF samples were taken once, at the time of shunt implantation.

RESULTS

In the clinical material, the highest values of eosinophils in peripheral blood and CSF pleocytosis were observed in the second measurement. It was accompanied by an increase in the majority of analyzed CSF interleukins.

CONCLUSIONS

CSF pleocytosis observed in the study group shortly after CSF sterilization is presumably related to an allergic reaction to Staphylococcus epidermidis, the causative agent of ventriculoperitoneal shunt infection.

As manuka honey (MH) exhibits immunoregulatory and anti-staphylococcal activities, we aimed to investigate if it could be effective in the treatment of atopic dermatitis (AD).

Adult volunteers with bilateral AD lesions were asked to apply MH on one site overnight for seven consecutive days and leave the contralateral site untreated as possible. Three Item Severity score was used to evaluate the response. Skin swabs were obtained from both sites before and after treatment to investigate the presence of staphylococci and enterotoxin production. In addition, the ability of MH and its methanolic and hexane extracts to down regulate IL4-induced CCL26 protein release from HaCaT cells was evaluated by enzyme linked immunosorbent assay. Also, the ability of MH to modulate calcium ionophore-induced mast cell degranulation was assessed by enzyme immunoassay.

In 14 patients, AD lesions significantly improved post MH treatment versus pre-treatment as compared to control lesions. No significant changes in the skin staphylococci were observed after day 7, irrespective of honey treatment. Consistent with the clinical observation, MH significantly down regulated IL4-induced CCL26 release from HaCaT cells in a dose-dependent manner. This effect was partially lost, though remained significant, when methanolic and hexane extracts of MH were utilized. In addition, mast cell degranulation was significantly inhibited following treatment with MH.

MH is potentially effective in the treatment of AD lesions based on both clinical and cellular studies through different mechanisms. This needs to be confirmed by randomized and controlled clinical trials.

Background: Numerous tissue-derived factors have been postulated to be involved in tissue migration of circulating monocytes. The aim of this study was to evaluate whether a defined hypoxic gradient can induce directed migration of naïve human monocytes and to identify responsible autocrine/paracrine factors.

Methods: Monocytes were isolated from peripheral blood mononuclear cells, transferred into chemotaxis chambers and subjected to a defined oxygen gradient with or without the addition of CCL26. Cell migration was recorded and secretome analyses were performed.

Results: Cell migration recordings revealed directed migration of monocytes towards the source of hypoxia. Analysis of the monocyte secretome demonstrated a reduced secretion of 70% (19/27) of the analyzed cytokines under hypoxic conditions. The most down-regulated factors were CCL26 (- 99%), CCL1 (- 95%), CX3CL1 (- 95%), CCL17 (- 85%) and XCL1 (- 83%). Administration of recombinant CCL26 abolished the hypoxia-induced directed migration of human monocytes, while the addition of CCL26 under normoxic conditions resulted in a repulsion of monocytes from the source of CCL26.

Conclusions: Hypoxia induces directed migration of human monocytes in-vitro. Autocrine/paracrine released CCL26 is involved in the hypoxia-mediated monocyte migration and may represent a target molecule for the modulation of monocyte migration in-vivo.

Keywords: CCL26; Cytokines; Hypoxia; Migration; Monocytes.

Purpose: Inflammation plays a crucial role in epileptogenesis. We analyzed inflammatory cytokines in plasma and saliva from children with seizures and healthy controls and measured their associations with HHV6 and EBV infection.

Methods: We analyzed plasma from 36 children within 24 h of seizures (cases) and 43 healthy controls and saliva from 44 cases and 44 controls with a multiplex immunoassay. Saliva from all controls and 65 cases and blood from 26 controls and 35 cases were also analyzed by PCR for viral DNA. Primary outcome was cytokine levels in cases vs. controls. Secondary outcomes included detection of HHV-6 and EBV viral DNA in cases vs. controls and viral loads in cases vs. controls. Statistical analysis included the Wilcoxon Rank Sum test, Fisher's exact test, ANOVA, and Spearman correlation.

Results: Compared to controls, patients had higher levels of CCL11 (p = 0.0018), CCL26 (p<0.001), IL10 (p = 0.044), IL6 (p<0.001), IL8 (p = 0.018), and MIP1β (p = 0.0012). CCL11 was higher with 3 or more seizures (p = 0.01), seizures longer than 10 min (p = 0.001), and when EEG showed focal slowing (p = 0.02). In saliva, febrile seizures had higher levels of IL-1β (n = 7, p = 0.04) and new onset seizures had higher IL-6 (n = 15, p = 0.02). Plasma and saliva cytokine levels did not show a correlation. The frequency of HHV-6 and EBV detection was similar across groups and not different than controls. We found no correlation between viral load and cytokine levels.

Conclusions: We showed differential activation of neuroinflammatory pathways in plasma from different seizure etiologies compared to controls, unrelated to viral infection.

Keywords: CCL11; Children; Cytokines; Epilepsy; HHV6; Inflammation; Seizures.

Introduction: Dysregulation of NLRP3 inflammasome complex formation can promote chronic inflammation by increased release of IL-1β. However, the effect of NLRP3 complex formation on tumor progression remains controversial. Therefore, we sought to determine the effect of NLRP3 modulation on the growth of the different types of cancer cells, derived from lung, breast, and prostate cancers as well as neuroblastoma and glioblastoma in-vitro.

Method: The effect of Caspase 1 inhibitor (VX765) and combination of LPS/Nigericin on NLRP3 inflammasome activity was analyzed in A549 (lung cancer), MCF-7 (breast cancer), PC3 (prostate cancer), SH-SY5Y (neuroblastoma), and U138MG (glioblastoma) cells. Human fibroblasts were used as control cells. The effect of VX765 and LPS/Nigericin on NLRP3 expression was analyzed using western blot, while IL-1β and IL-18 secretion was detected by ELISA. Tumor cell viability and progression were determined using Annexin V, cell proliferation assay, LDH assay, sphere formation assay, transmission electron microscopy, and a multiplex cytokine assay. Also, angiogenesis was investigated by a tube formation assay. VEGF and MMPs secretion were detected by ELISA and a multiplex assay, respectively. Statistical analysis was done using one-way ANOVA with Tukey's analyses and Kruskal-Wallis one-way analysis of variance.

Results: LPS/Nigericin increased NRLP3 protein expression as well as IL-1β and IL-18 secretion in PC3 and U138MG cells compared to A549, MCF7, SH-SY5Y cells, and fibroblasts. In contrast, MIF expression was commonly found upregulated in A549, PC3, SH-SY5Y, and U138MG cells and fibroblasts after Nigericin treatment. Nigericin and a combination of LPS/Nigericin decreased the cell viability and proliferation. Also, LPS/Nigericin significantly increased tumorsphere size in PC3 and U138MG cells. In contrast, the sphere size was reduced in MCF7 and SH-SY5Y cells treated with LPS/Nigericin, while no effect was detected in A549 cells. VX765 increased secretion of CCL24 in A549, MCF7, PC3, and fibroblasts as well as CCL11 and CCL26 in SH-SY5Y cells. Also, VX765 significantly increased the production of VEGF and MMPs and stimulated angiogenesis in all tumor cell lines.

Discussion: Our data suggest that NLRP3 activation using Nigericin could be a novel therapeutic approach to control the growth of tumors producing a low level of IL-1β and IL-18.

Keywords: IL-1β; NLRP3; cancer; inflammasome; nigericin.

Esophageal adenocarcinoma (EAC) is an aggressive cancer with poor prognosis and incidence is increasing rapidly in the Western world. Multi-modal treatment has improved survival outcomes but only for a minority of patients. Currently no markers have been identified to predict treatment response. This study investigated the association between clinical outcomes and pre-treatment levels of 54 serum proteins in n = 80 patients with EAC. Low tumor regression grade (TRG), corresponding to a favorable treatment response, was linked to prolonged overall survival (OS). CCL4 was higher in patients with a favorable treatment response, while Tie2 and CRP were higher in poor responders. Elevated CCL22 and CCL26 was associated with improved OS, while elevated IL-10 showed a negative association. CCL3, CCL4, IL-1α and IL-12/IL23p40 were highest in individuals with no adverse features of tumor biology, whereas levels of Tie2 and VEGF were lowest in this cohort. CCL4 was also elevated in patients with high tumor lymphocyte infiltration. Comparison of matched pre- and post-treatment serum (n = 28) showed a large reduction in VEGFC, and a concomitant increase in other cytokines, including CCL4. These data link several serum markers with clinical outcomes, highlighting an important role for immune cell trafficking in the EAC antitumor immune response.

Keywords: chemokines; clinical outcome; cytokines; esophageal adenocarcinoma; prognostic markers; serum markers; survival; treatment response.

Allergic asthma is a chronic inflammatory airway disease characterized by dysregulated type 2 immune responses, including degranulating airway eosinophils that induce tissue damage and airway hyperresponsiveness (AHR). The type 2 cytokines interleukin 5 (IL-5) and IL-13 and the eosinophil-specific chemokine CCL11/CCL24/CCL26 axis recruit, activate, and regulate eosinophils in the airways. In this issue of the JCI, Karcz et al. identified a mechanism involving the nucleotide sugar UDP-glucose (UDP-G) and the purinergic receptor P2Y14R in amplifying eosinophil accumulation in the lung. During type 2 inflammation, UDP-G activates P2Y14R on eosinophils, inducing the cells to move and migrate into the lung. Pharmacologically or genetically inhibiting P2Y14R on eosinophils attenuated eosinophil infiltration and AHR. Future experiments, including identifying additional type 2 factors regulating P2Y14R expression on lung eosinophils, are necessary to ascertain the impact of targeting P2Y14R as an alternative or adjunctive therapy to current type 2 biologics for the treatment of asthma.

Background: Atopic dermatitis/AD is a common disease, with particularly high prevalence seen in Africa. It is increasingly recognized that AD patients of different ethnic backgrounds have unique molecular signatures in skin, potentially accounting for treatment response variations. However, the skin profile of AD patients from Africa is unknown, hindering development of new treatments targeted to this patient population.

Objective: To characterize the skin profile of AD patients from Africa.

Methods: Gene-expression studies including RNA-sequencing (using threshold of foldchange/FCH>2 and false discovery rate/FDR<0.05) and real-time PCR were performed on skin biopsies of Tanzanian moderate-to-severe AD patients and controls.

Results: Tanzanian AD skin presented robust up-regulations of multiple key mediators of both Th2 (IL-13, IL10, IL-4R, CCL13/CCL17/CCL18/CCL26) and Th22 (IL22, S100As) pathways. Markers related to Th17/IL-23 (IL-17A, IL-23p19/IL-23A, IL-23p40/IL-12, PI3, DEFB4B) and Th1 (IFNγ, CXCL9/CXCL10/CXCL11) were also significantly overexpressed in AD tissues, albeit to a lesser extent. IL-36 isoforms showed significant upregulations in African skin. The barrier fingerprint of Tanzanian AD showed no suppression of hallmark epidermal barrier differentiation genes, such as filaggrin/FLG, loricrin/LOR, and periplakin/PPL, with robust attenuation of lipid metabolism genes (i.e AWAT1).

Conclusion: The skin phenotype of Tanzanian AD patients is consistent with that of African-Americans, exhibiting dominant Th2/Th22-skewing, minimal dysregulation of terminal differentiation, and even broader attenuation of lipid metabolism-related products. These data highlight the unique characteristic of AD in black individuals, as well as the need to develop unique treatments targeting AD patients of these under-represented populations.

Keywords: African patients; Atopic dermatitis; T cell; Th2; barrier; biomarkers; clinical immunology; dermatology.

Type-2 (T2) inflammation is a characteristic feature of asthma. Biological therapies have been developed to target T2-inflammation in asthma. IL-13 is a key component of T2-inflammation in asthma, driving mucus hypersecretion, IgE-induction, and smooth muscle contraction. Early phase clinical trials for treatments that target T2-inflammation require biomarkers to assess pharmacological effects. The aim of this study was to examine levels of IL-13 inducible biomarkers in the airway epithelium of patients with mild asthma compared to healthy controls. Ten patients with mild asthma with high blood eosinophil and high fractional exhaled nitric oxide (FeNO) were recruited, and six healthy subjects. Blood eosinophil and FeNO reproducibility was assessed prior to bronchoscopy. Epithelial brushings were collected and assessed for IL-13 inducible gene expression. Blood eosinophil and FeNO levels remained consistent in both patients with asthma and healthy subjects. Of the 11 genes assessed, expression levels of 15LOX1, POSTN, CLCA1, SERPINB2, CCL26, and NOS2 were significantly higher in patients with asthma compared to healthy controls. These six genes, present in patients with mild asthma with T2 inflammation, have the potential to be used in translational early phase asthma clinical trials of novel therapies as bronchial epithelial biomarkers.

The clinical significance and comprehensive features of chemokines and their receptors in lung adenocarcinoma (LUAD) have not been clarified. We aimed to characterize the expression profiles of chemokine and chemokine receptor family members and construct a chemokine- and chemokine receptor-based prognosis signature. A total of 1511 patients with LUAD from seven independent cohorts were included in the study. The training set collected from The Cancer Genome Atlas (TCGA) database containing 468 cases. The validation was performed on the basis of six different cohorts downloaded from Gene Expression Omnibus (GEO) database. A five-chemokine- and chemokine receptor-(CXCL2, CXCL13, CCL26, CCL20, CX3CR1) based prognosis signature was constructed with TCGA dataset using LASSO Cox regression and Cox proportional hazards regression analysis. A multivariate analysis verified that this signature was an independent prognostic factor. The predictive value of this signature was further verified by other six independent cohorts and multiple clinical subtypes. We performed immune cell infiltration analysis and biological pathway analysis which provided more insight into this signature-related immune and inflammatory landscape and clarified the intrinsic molecular mechanism by which this signature could be used to predict clinical prognosis. Furthermore, we explored the close relationship between this signature and tumor mutation burden (TMB), neoantigen burden, PD-1, PD-L1, CTLA4, TIDE score, T cell-inflamed score. This signature provided a robust prognostic biomarker for LUAD and could serve as a predictor for immunotherapy response, which may be used as an important supplement to immunotherapy to achieve individualized tumor treatment by optimizing the prognostic management and immunotherapy for patients with LUAD.

Keywords: Chemokines and chemokine receptors; Neoantigens; TIDE score; TMB.

Background: Tumor protein p63 has been shown to be important for epithelial dysfunction, including epithelial barrier defects and mucosal inflammation, in the development of chronic rhinosinusitis with nasal polyps (CRSwNP). Basonuclin1 (BNC1), an epithelial-specific transcriptional factor, is a direct downstream target of p63 and thus might be involved in the pathogenesis of CRSwNP.

Objective: We sought to investigate whether BNC1 was associated with p63-mediated epithelial barrier defects and nasal mucosal inflammation in CRSwNP.

Methods: Nasal tissue biopsies were obtained from 91 patients to CRSwNP, 49 chronic rhinosinusitis without nasal polyps (CRSsNP) patients, and 28 control subjects. Immunohistochemistry and immunofluorescence staining were used to determine the distribution of BNC1 in tissues and localization in cells, respectively. Quantitative PCR was performed to detect the expression levels of BNC1, TP63, epithelial barrier proteins, and type-2 helper T-cell inflammation-related genes.

Results: BNC1 mRNA expression was significantly elevated in the tissues in CRSwNP patients compared with CRSsNP (1.96-fold, p = 0.0003) and control groups (2.40-fold, p < 0.0001). BNC1 staining was strongly positive in the nasal epithelium and co-localized with p63-positive epithelial cells. The expression of BNC1 mRNA was strongly correlated with TP63 mRNA level both in tissue biopsies (r = 0.78, p < 0.0001) and epithelial scrapings (r = 0.97, p < 0.0001). BNC1 expression was also positively correlated with epithelial barrier protein genes (CDH1, CLDN1, CLDN4, TJP1, and TJP2) and epithelial genes involved in TH2 inflammation (IL33, CCL26, CLC, and ALOX15).

Conclusions: Overexpression of BNC1 may be associated with increased expression of TP63, and possibly contribute to the epithelial barrier defects and TH2 inflammation in CRSwNP.

Keywords: Basonuclin1; Chronic rhinosinusitis with nasal polyps; Epithelial barrier defects; Type-2 helper T-cell inflammation; p63.

Background: Emerging evidence suggests that chronic rhinosinusitis with nasal polyps (CRSwNP) is a highly heterogeneous disease with disparate inflammatory characteristics between different racial groups and geographies. Little is known currently about possible distinguishing factors underlying these inflammatory differences.

Objective: To interrogate for differences between Caucasian and Japanese CRSwNP disease using whole transcriptome and single-cell RNA gene expression profiling of nasal polyps (NPs).

Methods: We performed whole transcriptome RNA sequencing (RNA-seq) with endotype stratification of NPs from 8 Caucasian (residing in USA) and 9 Japanese (residing in Japan) patients. Reproducibility was confirmed by qPCR in an independent validation set of 46 Caucasian and 31 Japanese patients. Single-cell RNA-seq stratified key cell types for contributory transcriptional signatures.

Results: Unsupervised clustering analysis identified two major endotypes present within both NP cohorts, which have previously been reported at the cytokine level: 1) type 2 endotype and 2) non-type 2 endotype. Importantly, there was a statistically significant difference in the proportion of these endotypes between these geographically distinct NP subgroups (p = 0.03). Droplet-based single-cell RNA sequencing further identified prominent type 2 inflammatory transcript expression: C-C motif chemokine ligand 13 (CCL13) and CCL18 in M2 macrophages, as well as cystatin SN (CST1) and CCL26 in basal, suprabasal, and secretory epithelial cells.

Conclusion: NPs from both racial groups harbor the same two major endotypes, which we determine are present in differing ratios between each cohort with CRSwNP disease. Distinct inflammatory and epithelial cells contribute to the type 2 inflammatory profiles observed.

The specific changes linked to de novo development of postoperative PVR have remained elusive and were the object of the underlying study. Vitreous fluid (VF) was obtained at the beginning of vitrectomy from 65 eyes that underwent vitrectomy for primary rhegmatogenous retinal detachment (RRD) without preoperative PVR. Eyes developing postoperative PVR within 6 months after re-attachment surgery were compared to those which did not regarding the preoperative concentrations of 43 cytokines and chemokines in the VF, using multiplex beads analysis. For all comparisons Holm's correction was applied in order to control for multiple comparisons. Twelve out of 65 eyes (18.5%) developed PVR postoperatively. While 12 of the chemokines and cytokines presented concentration differences on a statistical level of p < 0.05 (CXCL5, CCL11, CCL24, CCL26, GM-CSF, IFN-γ, CCL8, CCL7, MIF, MIG/CXCL9, CCL19, and CCL25), CXCL5 was the only cytokine with sufficiently robust difference in its VF concentrations to achieve significance in eyes developing postoperative PVR compared to eyes without PVR. CXCL5 may represent a potent biomarker for the de novo development of postoperative PVR. In line with its pathophysiological role in the development of PVR, it might serve as a basis for the development of urgently needed preventive options.

Background: Integrated care pathways (ICPs) improve the management of patients with chronic rhinosinusitis with nasal polyps (CRSwNP). The application of ICPs requires development of endotype-based biomarkers to stratify patients. The value of cytokines and markers induced by cytokines for the management of CRSwNP is largely unknown.

Objectives: To determine the prognostic and pharmacologic value of type 2, non-type 2 cytokines, and markers associated with type 2 inflammation including CCL26, periostin, and cystatin SN, in nasal secretions for CRSwNP.

Methods: This retrospective study divided 151 patients with CRSwNP into discovery and validation phases. Concentrations of cytokines, CCL26, periostin, and cystatin SN in nasal secretions were determined using Luminex and ELISA. Predictive significance was assessed with receiver-operating characteristic curves. Survival analysis was performed using Kaplan-Meier curves and Cox regression models.

Results: Cystatin SN was an independent predictor of the uncontrolled status of CRSwNP over a 2-year follow-up after adjusting for other risk factors (hazard ratio [HR] of 1.168 and 1.132 in discovery and validation phases, respectively; both p < 0.001). Patients with high cystatin SN concentrations presented with a faster onset and higher rate of uncontrolled status than those with low levels (p < 0.001). Enhanced medical treatment for patients with high cystatin SN levels postponed the uncontrolled status in discovery (p = 0.016) and validation phases (p = 0.002) but did not completely abolish it by the end of the follow-up.

Conclusion: Cystatin SN levels in nasal secretions hold strong prognostic value and can facilitate medical instructions for managing CRSwNP.

Keywords: chronic rhinosinusitis with nasal polyps (CRSwNP); cystatin SN; cytokines; endotype; management.

Background: A growing body of evidence links various biomarkers to atopic dermatitis (AD). Still, little is known about the association of specific biomarkers to disease characteristics and severity in AD.

Objective: To explore the relationship between various immunological markers in the serum and disease severity in a hospital cohort of AD patients.

Methods: Outpatients with AD referred to the Department of Dermatology, Bispebjerg Hospital, Copenhagen, Denmark, were divided into groups based on disease severity (SCORAD). Serum levels of a preselected panel of immunoinflammatory biomarkers were tested for association with disease characteristics. Two machine learning models were developed to predict SCORAD from the measured biomarkers.

Results: A total of 160 patients with AD were included; 53 (33.1%) with mild, 73 (45.6%) with moderate, and 34 (21.3%) with severe disease. Mean age was 29.2 years (range 6-70 years) and 84 (52.5%) were females. Numerous biomarkers showed a statistically significant correlation with SCORAD, with the strongest correlations seen for CCL17/thymus and activation-regulated chemokine (chemokine ligand-17/TARC) and CCL27/cutaneous T cell-attracting-chemokine (CTACK; Spearman R of 0.50 and 0.43, respectively, p < 0.001). Extrinsic AD patients were more likely to have higher mean SCORAD (p < 0.001), CCL17 (p < 0.001), CCL26/eotaxin-3 (p < 0.001), and eosinophil count (p < 0.001) than intrinsic AD patients. Predictive models for SCORAD identified CCL17, CCL27, serum total IgE, IL-33, and IL-5 as the most important predictors for SCORAD, but with weaker associations than single cytokines.

Conclusions: Specific immunoinflammatory biomarkers in the serum, mainly of the Th2 pathway, are correlated with disease severity in patients with AD. Predictive models identified biomarkers associated with disease severity but this finding warrants further investigation.

Keywords: Atopic dermatitis; Biomarkers; Disease severity; Inflammation.

Recently, the combined use of FOLFIRINOX (leucovorin and fluorouracil plus irinotecan and oxaliplatin) and gemcitabine plus nab-paclitaxel has significantly improved the prognosis of patients with pancreatic cancer. However, there is still a high proportion of patients who develop metastatic pancreatic cancer in the course of chemotherapy or within a short period after chemotherapy. Previous reports have shown that chemotherapy-driven cytokine storms or the direct effects of certain chemotherapeutics on stromal and/or immune cells collectively change the microenvironment of the primary tumor, thus indirectly promoting metastasis. However, the mechanism underlying chemotherapy-induced metastasis in the course of chemotherapy, and afterwards, remains elusive in pancreatic cancer. In the present study, we aimed to determine the expression of CCL26 in the pancreatic cancer-associated fibroblasts (CAFs) after nab-paclitaxel treatment and to explore the role of CCL26 in the pancreatic adenocarcinoma (PDAC) invasion. Our results showed that nab-paclitaxel increased CCL26 mRNA and protein expression levels in a dose- and time-dependent manner. Subsequently, PDAC cell lines were treated with recombinant CCL26 for 48 h. The transwell migration assay showed that recombinant CCL26 enhanced the invasion of PDAC cells. Western blot analysis showed that the protein expression levels of phospho-(p-)PI3K, p-AKT, and p-mTOR were increased by CCL26 in PDAC cells. CCL26 expressions in 95 PDAC tissues and adjacent normal tissues were evaluated using reverse transcription-quantitative polymerase chain reaction and immunohistochemistry. CCL26 was found to be overexpressed in PDAC samples, and upregulated CCL26 expression was significantly associated with advanced perineural invasion, lymph node metastasis, and poor differentiation. In summary, our results showed that nab-paclitaxel increased the expression of CCL26 in CAFs, and CCL26 enhanced the invasive potential of pancreatic cancer cells by activating the PI3K/AKT/mTOR axis. Thus, CCL26 may be a potential prognostic biomarker for pancreatic cancer.

Keywords: CCL26; PI3K/AKT/mTOR signaling pathway; cancer-associated fibroblasts; invasion; pancreatic cancer.

Cytokines are immune response components important in innate immunity and inflammatory response. They are harnessed as part of local immunological responses by animals to combat local infections and/or infestations. This study investigated expression of four selected cytokine genes, namely, interleukin 1 beta (IL-1β), chemokine C-C ligand 2 (CCL2), chemokine C-C ligand 26 (CCL26) and interleukin 8 (IL-8), at tick attachment and control sites in a South African indigenous sheep breed the Namaqua Afrikaner (NA) and two commercial breeds, the Dorper and South African Mutton Merino (SAMM). The NA was previously shown to be more resistant to infestation by ticks than the two commercial breeds. NA ewes expressed IL-1β more at tick attachment sites compared to Dorpers. The NA breed was also more likely to upregulate the expression of the CCL2, CCL26 and IL-8 genes at tick attachment sites compared to control sites than the other breeds. The results of this study gave an indication that cytokines are involved in immune responses to tick challenge and laid a foundation for further studies under controlled challenge conditions.

Keywords: Dorper; Immune response. Namaqua Afrikaner; South African Mutton Merino; Tick attachment.

Background: The purpose of this study was to investigate the role of eotaxin family members including C-C motif chemokine 11 (CCL11), C-C motif chemokine 24 (CCL24), and C-C motif chemokine 26 (CCL26) as the subgroups of CC-chemokine in patients affected with osteoporosis and osteopenia.

Methods: Overall, 19 osteoporotic patients, 18 osteopenic individuals, and 20 healthy subjects were recruited in this study. The bone mineral density (BMD) was then measured at the lumbar spine (L1-L4) and the hip (femoral neck and total hip) using dual-energy X-ray absorptiometry for diagnosis of bone density and related disorders. Additionally, enzyme-linked immunosorbent assay (ELISA) technique was employed to measure the serum levels of CCL11, CCL24, and CCL26.

Results: The circulating levels of CCL11, CCL24, and CCL26 had been increased in both groups of patients with osteopenia and osteoporosis compared to those in healthy subjects (P<0.05); while no significant difference was observed between serum levels of these chemokines in such patients.

Conclusion: Eotaxins can play a role in the pathogenesis of osteoporosis and osteopenia; however, further studies are needed to clarify various roles of eotaxins in the pathophysiology of osteoporosis and osteopenia.

Keywords: Eotaxins; Human; Osteopenia; Osteoporosis; Protein.

Background: Targeted approaches may not account for the complexity of inflammation involved in children with severe asthma (SA), highlighting the need to consider more global analyses. We aimed to identify sets of immune constituents that distinguish children with SA from disease-control subjects through a comprehensive analysis of cells and immune constituents measured in bronchoalveolar lavage (BAL) and blood.

Methods: Twenty children with SA and 10 age-matched control subjects with chronic respiratory disorders other than asthma were included. Paired blood and BAL samples were collected and analyzed for a large set of cellular (eosinophils, neutrophils, and subsets of lymphocytes and innate lymphoid cells) and soluble (chemokines, cytokines, and total antibodies) immune constituents. First, correlations of all immune constituents between BAL and blood and with demographic and clinical data were assessed (Spearman correlations). Then, all data were modelled using supervised multivariate analyses (partial least squares discriminant analysis, PLS-DA) to identify immune constituents that significantly discriminate between SA and control subjects. Univariate analyses were performed (Mann-Whitney tests) and then PLS-DA and univariate analyses were combined to identify the most discriminative and significant constituents.

Results: Concentrations of soluble immune constituents poorly correlated between BAL and blood. Certain constituents correlated with age or body mass index and, in asthmatics, with clinical symptoms, such as the number of exacerbations in the previous year, asthma control test score, or forced expiratory volume. Multivariate supervised analysis allowed construction of a model capable of distinguishing children with SA from control subjects with 80% specificity and 100% sensitivity. All immune constituents contributed to the model but some, identified by variable-important-in-projection values > 1 and p < 0.1, contributed more strongly, including BAL Th1 and Th2 cells and eosinophilia, CCL26 (Eotaxin 3), IgA and IL-19 concentrations in blood. Blood concentrations of IL-26, CCL13, APRIL, and Pentraxin-3 may also help in the characterization of SA.

Conclusions: The analysis of a large set of immune constituents may allow the identification of a biological immune signature of SA. Such an approach may provide new leads for delineating the pathogenesis of SA in children and identifying new targets for its diagnosis, prediction, and personalized treatment.

Keywords: children; immune signature; pathogenesis; precision medicine; severe asthma.

The clinical significance and regulators of IL-13Rα2 in itch and atopic dermatitis (AD) remain unclear. To identify disease-driven regulatory circuits of IL-13Rα2, transcriptomic/pathological analysis was performed in skin from patients with AD, psoriasis, healthy subjects, and murine AD model. Functionality was investigated in sensory neurons, keratinocytes and animal model, by using knockdown (KD), calcium imaging, RNA-seq, cytokine arrays, pharmacological assays, and behavioural investigations. In our study, an upregulated IL-13Rα2 expression was revealed in skin of AD patients, but not psoriasis, in a disease activity-dependent manner. In cultured human keratinocytes, IL-13 increased IL-13Rα2 transcription levels, and this were downregulated by IL-13Rα1KD. IL-13Rα2KD reduced transcription levels of EDNRA, CCL20, CCL26. In contrast, sensory neuron-derived IL-13Rα2 was upregulated by TLR2 heterodimer agonists, Pam3CSK4 and FSL-1. In a mouse cheek model, pre-administration of Pam3CSK4 and FSL-1 enhanced IL-13-elicited scratching behaviour. Consistently, in cultured sensory neurons Pam3CSK4 enhanced IL-13-elicted calcium transients, increased number of responders, and orchestrated chemerin, CCL17 and CCL22 release. These release was inhibited by IL-13Rα2KD. Collectively, IL-13 regulates keratinocyte-derived IL-13Rα2 and TLR2 to modulate neuronal IL-13Rα2, thereby promoting neurogenic inflammation and exacerbating AD and itch. Thus, the cutaneous IL-13-IL-13Rα2 and neuronal TLR2-IL-13Rα2 pathway represent important targets to treat AD and itch.

Keywords: Atopic Dermatitis; IL-13, Cytokine; IL-13Rα1 and 2; Innate Immune; Toll-like Receptor.

Neuropathic pain is a serious clinical issue, and its treatment remains a challenge in contemporary medicine. Thus, dynamic development in the area of animal and clinical studies has been observed. The mechanisms of neuropathic pain are still not fully understood; therefore, studies investigating these mechanisms are extremely important. However, much evidence indicates that changes in the activation and infiltration of immune cells cause the release of pronociceptive cytokines and contribute to neuropathic pain development and maintenance. Moreover, these changes are associated with low efficacy of opioids used to treat neuropathy. To date, the role of CC chemokine receptor type 3 (CCR3) in nociception has not been studied. Similarly, little is known about its endogenous ligands (C-C motif ligand; CCL), namely, CCL5, CCL7, CCL11, CCL24, CCL26, and CCL28. Our research showed that the development of hypersensitivity in rats following chronic constriction injury (CCI) of the sciatic nerve is associated with upregulation of CCL7 and CCL11 in the spinal cord and dorsal root ganglia (DRG). Moreover, our results provide the first evidence that single and repeated intrathecal administration of the CCR3 antagonist SB328437 diminishes mechanical and thermal hypersensitivity. Additionally, repeated administration enhances the analgesic properties of morphine and buprenorphine following nerve injury. Simultaneously, the injection of SB328437 reduces the protein levels of some pronociceptive cytokines, such as IL-6, CCL7, and CCL11, in parallel with a reduction in the activation and influx of GFAP-, CD4- and MPO-positive cells in the spinal cord and/or DRG. Moreover, we have shown for the first time that an inhibitor of myeloperoxidase-4-aminobenzoic hydrazide may relieve pain and simultaneously enhance morphine and buprenorphine efficacy. The obtained results indicate the important role of CCR3 and its modulation in neuropathic pain treatment and suggest that it represents an interesting target for future investigations.

Keywords: 4-aminobenzoic hydrazide; CCR3; SB328437; buprenorphine; cytokines; morphine; myeloperoxidase; neutrophils.