CCR3-dependent eosinophil recruitment is regulated by sialyltransferase ST3Gal-IV.

Eosinophil recruitment is a pathological hallmark of many allergic and helminthic diseases. Here, we investigated chemokine receptor CCR3-induced eosinophil recruitment in sialyltransferase St3gal4-/- mice. We found a marked decrease in eosinophil extravasation into CCL11-stimulated cremaster muscles and into the inflamed peritoneal cavity of St3gal4-/- mice. Ex vivo flow chamber assays uncovered reduced adhesion of St3gal4-/- compared to wild type eosinophils. Using flow cytometry, we show reduced binding of CCL11 to St3gal4-/- eosinophils. Further, we noted reduced binding of CCL11 to its chemokine receptor CCR3 isolated from St3gal4-/- eosinophils. This was accompanied by almost absent CCR3 internalization of CCL11-stimulated St3gal4-/- eosinophils. Applying an ovalbumin-induced allergic airway disease model, we found a dramatic reduction in eosinophil numbers in bronchoalveolar lavage fluid following intratracheal challenge with ovalbumin in St3gal4-deficient mice. Finally, we also investigated tissue-resident eosinophils under homeostatic conditions and found reduced resident eosinophil numbers in the thymus and adipose tissue in the absence of ST3Gal-IV. Taken together, our results demonstrate an important role of ST3Gal-IV in CCR3-induced eosinophil recruitment in vivo rendering this enzyme an attractive target in reducing unwanted eosinophil infiltration in various disorders including allergic diseases.

Muscle eosinophilia is a hallmark of chronic disease in facioscapulohumeral muscular dystrophy.

Facioscapulohumeral muscular dystrophy (FSHD) is a progressive myopathy caused by the aberrant increased expression of the DUX4 retrogene in skeletal muscle cells. The DUX4 gene encodes a transcription factor that functions in zygotic genome activation and then is silenced in most adult somatic tissues. DUX4 expression in FSHD disrupts normal muscle cell function; however, the downstream pathogenic mechanisms are still unclear. Histologically, FSHD affected muscles show a characteristic dystrophic phenotype that is often accompanied by a pronounced immune cell infiltration, but the role of the immune system in FSHD is not understood. Previously, we used ACTA1;FLExDUX4 FSHD-like mouse models varying in severity as discovery tools to identify increased Interleukin 6 and microRNA-206 levels as serum biomarkers for FSHD disease severity. In this study, we use the ACTA1;FLExDUX4 chronic FSHD-like mouse model to provide insight into the immune response to DUX4 expression in skeletal muscles. We demonstrate that these FSHD-like muscles are enriched with the chemoattractant eotaxin and the cytotoxic eosinophil peroxidase, and exhibit muscle eosinophilia. We further identified muscle fibers with positive staining for eosinophil peroxidase in human FSHD muscle. Our data supports that skeletal muscle eosinophilia is a hallmark of FSHD pathology.

Profiling CCR3 target pathways for discovering novel antagonists from natural products using label-free cell phenotypic assays.

CC chemokine receptor 3 (CCR3) plays important roles in atopic dermatitis (AD) and other related allergic diseases. Activation of CCR3 receptor signaling pathways regulates the recruitment of eosinophils to related tissues, releasing inflammatory mediators and causing inflammatory responses. However, none of the known CCR3 antagonists exhibit promising efficacy in clinical trials. In this work, we sought new natural CCR3 antagonists for drug development. To construct a high-throughput screening model, we established a stably transfected CHO-K1-Gα15-CCR3 cell line, and receptor expression was demonstrated by real-time quantitative PCR, confocal detection and flow cytometry analysis. Then, we applied a label-free cell phenotyping technique to profile and deconvolute CCR3 target pathways in CHO-K1-Gα15-CCR3 cells and found that activation of CCR3 triggered the Gq-PLC-Ca2+ and MAPK-P38-ERK pathways. By in vitro and in silico experiments, we discovered a novel CCR3 antagonist emodin, with an IC50 value of 27.28 ± 1.71 µM out of 266 compounds that were identified in 15 traditional Chinese medicines used in the clinical treatment of skin diseases. Molecular docking graphically presented the binding mode of emodin on CCR3. This work reports a new approach for CCR3 antagonist screening and pathway detection and identifies a new antagonist that would benefit future drug development.

Eotaxin-1/CCL11 promotes cellular senescence in human-derived fibroblasts through pro-oxidant and pro-inflammatory pathways.

Introduction: Eotaxin-1/CCL11 is a pivotal chemokine crucial for eosinophil homing to the lungs of asthmatic patients. Recent studies also suggest that CCL11 is involved in the aging process, as it is upregulated in elderly, and correlated with shorter telomere length in leukocytes from asthmatic children. Despite its potential pro-aging effects, the precise contribution of CCL11 and the underlying mechanisms involved in the promotion of cellular senescence remains unclear. Therefore, the primary goal of this study was to explore the role of CCL11 on senescence development and the signaling pathways activated by this chemokine in lung fibroblasts. Methods: To investigate the targets potentially modulated by CCL11, we performed an in silico analysis using PseudoCell. We validated in vitro the activation of these targets in the human lung fibroblast cell line MRC-5 following rhCCL11 exposure. Finally, we performed differential gene expression analysis in human airway epithelial cells of asthmatic patients to assess CCL11 signaling and activation of additional senescent markers. Results: Our study revealed that eotaxin-1/CCL11 promote reactive oxygen secretion (ROS) production in lung fibroblasts, accompanied by increased activation of the DNA damage response (DDR) and p-TP53 and γH2AX. These modifications were accompanied by cellular senescence promotion and increased secretion of senescence-associated secretory phenotype inflammatory cytokines IL-6 and IL-8. Furthermore, our data show that airway epithelial lung cells from atopic asthmatic patients overexpress CCL11 along with aging markers such as CDKN2A (p16INK4a) and SERPINE1. Discussion: These findings provide new insights into the mechanisms underlying the pro-aging effects of CCL11 in the lungs of asthmatic patients. Understanding the role of CCL11 on senescence development may have important implications for the treatment of age-related lung diseases, such as asthma.

Young Astrocytic Mitochondria Attenuate the Elevated Level of CCL11 in the Aged Mice, Contributing to Cognitive Function Improvement.

Aging drives cognitive decline, and mitochondrial dysfunction is a hallmark of age-induced neurodegeneration. Recently, we demonstrated that astrocytes secrete functional mitochondria (Mt), which help adjacent cells to resist damage and promote repair after neurological injuries. However, the relationship between age-dependent changes in astrocytic Mt function and cognitive decline remains poorly understood. Here, we established that aged astrocytes secret less functional Mt compared to young astrocytes. We found the aging factor C-C motif chemokine 11 (CCL11) is elevated in the hippocampus of aged mice, and that its level is reduced upon systemic administration of young Mt, in vivo. Aged mice receiving young Mt, but not aged Mt improved cognitive function and hippocampal integrity. Using a CCL11-induced aging-like model in vitro, we found that astrocytic Mt protect hippocampal neurons and enhance a regenerative environment through upregulating synaptogenesis-related gene expression and anti-oxidants that were suppressed by CCL11. Moreover, the inhibition of CCL11-specific receptor C-C chemokine receptor 3 (CCR3) boosted the expression of synaptogenesis-related genes in the cultured hippocampal neurons and restored the neurite outgrowth. This study suggests that young astrocytic Mt can preserve cognitive function in the CCL11-mediated aging brain by promoting neuronal survival and neuroplasticity in the hippocampus.

Adeno-Associated Virus-Mediated Interleukin-12 Gene Expression Alleviates Lung Inflammation and Type 2 T-Helper-Responses in Ovalbumin-Sensitized Asthmatic Mice.

High levels of allergen exposure increase the prevalence of asthma in developed countries. The asthmatic type 2 T-helper (Th2) response is characterized with high eosinophil infiltration, elevated Th2 cytokines, and immunoglobulin (Ig) E secretion resulting in local or systemic inflammation. However, the treatment with palliative Th2 inhibitor drugs cannot completely control asthma and that is why the development of novel approaches is still important. Based on type 1 T-helper (Th1) and Th2 immune homeostasis, the enhanced Th1 immune response has high potential to alleviate Th2 immune response. Thus, we aimed to overexpress single chain IL-12 (scIL-12) through recombinant adeno-associated virus (rAAV) vector (as rAAV-IL-12) and test the efficacy in an ovalbumin (OVA)-induced asthmatic murine model. We firstly demonstrated the bioactivity of exogenous scIL-12. The expression of exogenous scIL-12 was also detected in the lungs of rAAV-IL-12 transduced mice. The data demonstrated that overexpression of exogenous scIL-12 significantly suppressed total number of cells and eosinophil infiltration, as well as the mucus secretion in rAAV-IL-12-treated mice. The decreased OVA-specific IgE in bronchoalveolar lavage fluid and gene expression of Th2-cytokines or C-C motif ligand (CCL) 11 (also eotaxin-1) in lung were observed. In addition, the production of cytokines in the supernatants of OVA-stimulated splenocytes were suppressed with rAAV-IL-12 treatment. Thus, scIL-12 expression by rAAV vector was able to modulate Th2 activity and has the potential to be developed as a feasible strategy in modulating allergic diseases.

Eotaxin-1 (CCL11) in neuroinflammatory disorders and possible role in COVID-19 neurologic complications.

The related neurologic complications of SARS-CoV-2 infection in COVID-19 patients and survivors comprise symptoms including depression, anxiety, muscle pain, dizziness, headaches, fatigue, and anosmia/hyposmia that may continue for months. Recent studies have been demonstrated that chemokines have brain-specific attraction and effects such as chemotaxis, cell adhesion, modulation of neuroendocrine functions, and neuroinflammation. CCL11 is a member of the eotaxin family that is chemotactic agents for eosinophils and participate in innate immunity. Eotaxins may exert physiological and pathological functions in the central nerve system, and CCL11 may induce neuronal cytotoxicity effects by inducing the production of reactive oxygen species (ROS) in microglia cells. Plasma levels of CCL11 elevated in neuroinflammation and neurodegenerative disorders. COVID-19 patients display elevations in CCL11 levels. As CCL11 plays roles in physiosomatic and neuroinflammation, analyzing the level of this chemokine in COVID-19 patients during hospitalization and to predicting post-COVID-19-related neurologic complications may be worthwhile. Moreover, using chemokine modulators may be helpful in lessening the neurologic complications in such patients.

Trans-activation of eotaxin-1 by Brg1 contributes to liver regeneration.

Infiltration of eosinophils is associated with and contributes to liver regeneration. Chemotaxis of eosinophils is orchestrated by the eotaxin family of chemoattractants. We report here that expression of eotaxin-1 (referred to as eotaxin hereafter), but not that of either eotaxin-2 or eotaxin-3, were elevated, as measured by quantitative PCR and ELISA, in the proliferating murine livers compared to the quiescent livers. Similarly, exposure of primary murine hepatocytes to hepatocyte growth factor (HGF) stimulated eotaxin expression. Liver specific deletion of Brahma-related gene 1 (Brg1), a chromatin remodeling protein, attenuated eosinophil infiltration and down-regulated eotaxin expression in mice. Brg1 deficiency also blocked HGF-induced eotaxin expression in cultured hepatocytes. Further analysis revealed that Brg1 could directly bind to the proximal eotaxin promoter to activate its transcription. Mechanistically, Brg1 interacted with nuclear factor kappa B (NF-κB)/RelA to activate eotaxin transcription. NF-κB knockdown or pharmaceutical inhibition disrupted Brg1 recruitment to the eotaxin promoter and blocked eotaxin induction in hepatocytes. Adenoviral mediated over-expression of eotaxin overcame Brg1 deficiency caused delay in liver regeneration in mice. On the contrary, eotaxin depletion with RNAi or neutralizing antibodies retarded liver regeneration in mice. More important, Brg1 expression was detected to be correlated with eotaxin expression and eosinophil infiltration in human liver specimens. In conclusion, our data unveil a novel role of Brg1 as a regulator of eosinophil trafficking by activating eotaxin transcription.

Chemoattraction of Neoplastic Glial Cells with CXCL10, CCL2 and CCL11 as a Paradigm for a Promising Therapeutic Approach for Primary Brain Tumors.

Chemoattraction is a normal and essential process, but it can also be involved in tumorigenesis. This phenomenon plays a key role in glioblastoma (GBM). The GBM tumor cells are extremely difficult to eradicate, due to their strong capacity to migrate into the brain parenchyma. Consequently, a complete resection of the tumor is rarely a possibility, and recurrence is inevitable. To overcome this problem, we proposed to exploit this behavior by using three chemoattractants: CXCL10, CCL2 and CCL11, released by a biodegradable hydrogel (GlioGel) to produce a migration of tumor cells toward a therapeutic trap. To investigate this hypothesis, the agarose drop assay was used to test the chemoattraction capacity of these three chemokines on murine F98 and human U87MG cell lines. We then studied the potency of this approach in vivo in the well-established syngeneic F98-Fischer glioma-bearing rat model using GlioGel containing different mixtures of the chemoattractants. In vitro assays resulted in an invasive cell rate 2-fold higher when chemokines were present in the environment. In vivo experiments demonstrated the capacity of these specific chemoattractants to strongly attract neoplastic glioblastoma cells. The use of this strong locomotion ability to our end is a promising avenue in the establishment of a new therapeutic approach in the treatment of primary brain tumors.

Eosinophils are dispensable for development of MOG35-55-induced experimental autoimmune encephalomyelitis in mice.

Experimental autoimmune encephalomyelitis (EAE) represents the mouse model of multiple sclerosis, a devastating neurological disorder. EAE development and progression involves the infiltration of different immune cells into the brain and spinal cord. However, less is known about a potential role of eosinophil granulocytes for EAE disease pathogenesis. In the present study, we found enhanced eosinophil abundance accompanied by increased concentration of the eosinophil chemoattractant eotaxin-1 in the spinal cord in the course of EAE induced in C57BL/6 mice by immunization with MOG35-55 peptide. However, the absence of eosinophils did not affect neuroinflammation, demyelination and clinical development or severity of EAE, as assessed in ∆dblGATA1 eosinophil-deficient mice. Taken together, despite their enhanced abundance in the inflamed spinal cord during disease progression, eosinophils were dispensable for EAE development.

Repetitive mild traumatic brain injury affects inflammation and excitotoxic mRNA expression at acute and chronic time-points.

The cumulative effect of mild traumatic brain injuries (mTBI) can result in chronic neurological damage, however the molecular mechanisms underpinning this detriment require further investigation. A closed head weight drop model that replicates the biomechanics and head acceleration forces of human mTBI was used to provide an exploration of the acute and chronic outcomes following single and repeated impacts. Adult male C57BL/6J mice were randomly assigned into one of four impact groups (control; one, five and 15 impacts) which were delivered over 23 days. Outcomes were assessed 48 hours and 3 months following the final mTBI. Hippocampal spatial learning and memory assessment revealed impaired performance in the 15-impact group compared with control in the acute phase that persisted at chronic measurement. mRNA analyses were performed on brain tissue samples of the cortex and hippocampus using quantitative RT-PCR. Eight genes were assessed, namely MAPT, GFAP, AIF1, GRIA1, CCL11, TARDBP, TNF, and NEFL, with expression changes observed based on location and follow-up duration. The cortex and hippocampus showed vulnerability to insult, displaying upregulation of key excitotoxicity and inflammation genes. Serum samples showed no difference between groups for proteins phosphorylated tau and GFAP. These data suggest that the cumulative effect of the impacts was sufficient to induce mTBI pathophysiology and clinical features. The genes investigated in this study provide opportunity for further investigation of mTBI-related neuropathology and may provide targets in the development of therapies that help mitigate the effects of mTBI.

Increased Thrombin-Activatable Fibrinolysis Inhibitor in Response to Sublingual Immunotherapy for Allergic Rhinitis.

OBJECTIVES/HYPOTHESIS: The objective of this study was to determine the role of thrombin-activatable fibrinolysis inhibitor (TAFI) as a candidate biomarker for therapeutic efficacy of sublingual immunotherapy (SLIT) and to identify the role of TAFI in the pathogenesis of allergic rhinitis (AR). STUDY DESIGN: Retrospective cohort study and laboratory study. METHODS: Serum was collected from patients with allergies to Japanese cedar pollen before, during, and after treatment with SLIT. We measured the levels of immunoreactive TAFI, C3a, and C5a in serum by enzyme-linked immunosorbent assay (ELISA) and assessed their relative impact on a combined symptom-medication score. We also examined the impact of TAFI on mast cells and fibroblasts in experiments performed in vitro. RESULTS: Serum levels of TAFI increased significantly in response to SLIT. By contrast, serum C3a levels decreased significantly over time; we observed a significant negative correlation between serum levels of TAFI versus C3a and symptom-medication score. Mast cell degranulation was inhibited in response to TAFI, as it was the expression of both CCL11 and CCL5 in cultured fibroblasts. CONCLUSIONS: High serum levels of TAFI may be induced by SLIT. TAFI may play a critical protective role in pathogenesis of AR by inactivating C3a and by inhibiting mast cell degranulation and chemokines expression in fibroblasts. LEVEL OF EVIDENCE: 4 Laryngoscope, 131:2413-2420, 2021.

Eotaxin-1/CCL11 is involved in cell migration in rheumatoid arthritis.

Eotaxin-1 (CCL11) induces the migration of different leukocyte types by interacting with CCR3. In rheumatoid arthritis (RA), fibroblast-like synoviocytes (FLS) are pathogenic effectors and a major CCR3-expressing cell. The aim of this study was to investigate the expression and function of CCL11 in RA FLS. The expression of CCL11 and CCR3 was evaluated by ELISA, immunofluorescence and quantitative PCR analysis. The CCL11 levels in serum and synovial fluids (SFs) from RA patients were significantly higher than those in serum from healthy controls and SFs from osteoarthritis patients. CCL11 and CCR3 were expressed in the RA synovial tissue lining layers. The secretion of CCL11 in RA FLS-conditioned medium and the mRNA expression of CCL11 and CCR3 were induced by TNF-α. Furthermore, CCL11 induced the mRNA expression of CCL11 and CCR3. Application of a CCR3 antagonist reduced TNF-α-induced CCL11 secretion from RA FLS. CCL11 induced the migration of RA FLS and monocytes. RA FLS migration was decreased by treatment with CCL11 siRNA. The migration of monocytes to medium conditioned with CCL11 siRNA-transfected and TNF-α-stimulated RA FLS was reduced. These data indicate that the self-amplification of CCL11 via CCR3 may play an important role in cell migration in RA.

Identifying the tumor-progressive gene expression profile in high-risk papillary thyroid cancer.

PURPOSE: Papillary thyroid cancer (PTC) is generally associated with a favorable prognosis. However, some patients have fatal disease, with locally infiltrating tumors or progressive distant metastases; yet few studies have investigated the characteristics of the tumor-progressive gene expression profile in advanced PTC. We conducted this study to clarify the gene expression status in advanced PTC and identify candidate molecules for prognostic biomarkers. METHODS: We analyzed 740 tumor-progressive gene expression levels from formalin-fixed paraffin-embedded blocks of samples from six patients with low-risk PTC and six patients with high-risk PTC, using the nCounter PanCancer Progression panel. Then, we investigated the association between the expression levels of focused genes and pathological factors in PTC patients in The Cancer Genome Atlas (TCGA) database. RESULTS: The expression levels of 14 genes in the high-risk PTC specimens were more than two-fold those in the low-risk PTC specimens. In the TCGA database, expression levels of four genes (CCL11, COL6A3, INHBA, and SRPX2) were significantly higher in patients with advanced PTC. Among the patients with advanced PTC, those with high SRPX2 expression levels had poor disease-free survival. Univariate and multivariate analyses revealed that high SRPX2 expression was an independent prognostic factor. CONCLUSION: Based on the findings of this study, CCL11, COL6A3, INHBA, and SRPX2 are potential biomarkers that indicate advanced PTC. SRPX2, in particular, is considered a prognostic biomarker.

High chemokine ligand 11 levels in nasal lavage fluid: A potential predictor of and therapeutic target for murine eosinophilic chronic rhinosinusitis.

AIM: We aimed to discover whether group 2 innate lymphoid cells (ILC2s) and cytokines in nasal lavage fluid could be used to predict eosinophilic infiltration in mice with eosinophilic chronic rhinosinusitis (ECRS). METHODS: Ten mice were divided into two groups. The ECRS group received an intranasal challenge of Aspergillus oryzae protease (AP) and ovalbumin (OVA) to establish disease. A control group received intranasal phosphate-buffered saline. Histopathology of nasal cavities and paranasal sinuses, and cytokine and ILC2s levels in nasal lavage fluid were analyzed and compared between the ECRS and control mouse groups. KEY FINDINGS: ILC2s numbers were not significantly higher in the nasal lavage fluid of the ECRS group mice compared with those of the control group. Eotaxin/chemokine (CC motif) ligand 11 (CCL11) levels were significantly higher in the nasal lavage fluid of mice in the ECRS group compared with those in the control group. However, no statistical differences were seen in the classic proinflammatory cytokines, IL-33, IL-25, and thymic stromal thymopoietin (TSLP), or the classic type 2 cytokines, IL-4, IL-5, and IL-13 between groups. SIGNIFICANCE: Eotaxin/CCL11 levels in nasal lavage fluid rather than that of ILC2s and classic proinflammatory and type 2 cytokines were significantly higher in ECRS mice compared with control ones. Eotaxin/CCL11 showed diagnostic and therapeutic value; however, more studies are needed to test and verify its value.

Asthma-related inflammation promotes lung metastasis of breast cancer cells through CCL11-CCR3 pathway.

BACKGROUND: Mechanisms that preclude lung metastasis are still barely understood. The possible consequences of allergic airways inflammation on cancer dissemination were studied in a mouse model of breast cancer. METHODS: Balb/c mice were immunized and daily exposed to ovalbumin (OVA) from day 21. They were subcutaneously injected with 4T1 mammary tumor cells on day 45 and sacrificed on day 67. Lung metastases were measured by biophotonic imaging (IVIS® 200 Imaging System) and histological measurement of tumor area (Cytomine software). Effects of CCL11 were assessed in vivo by intratracheal instillations of recCCL11 and in vitro using Boyden chambers. CCR3 expression on cell surface was assessed by flow cytometry. RESULTS: The extent of tumor metastases was significantly higher in lungs of OVA-exposed mice and increased levels of CCL11 expression were measured after OVA exposure. Migration of 4T1 cells and neutrophils was stimulated in vitro and in vivo by recCCL11. 4T1 cells and neutrophils express CCR3 as shown by flow cytometry and a selective CCR3 antagonist (SB-297006) inhibited the induction of 4T1 cells migration and proliferation in response to recCCL11. CONCLUSIONS: Allergic inflammation generated by exposure to allergens triggers the implantation of metastatic cells from primary breast tumor into lung tissues plausibly in a CCL11-CCR3-dependent manner. This indicates that asthma related inflammation in lungs might be a risk factor for lung metastasis in breast cancer patients.

Effects of Platelet-Rich Plasma on Cellular Populations of the Central Nervous System: The Influence of Donor Age.

Platelet-rich plasma (PRP) is a biologic therapy that promotes healing responses across multiple medical fields, including the central nervous system (CNS). The efficacy of this therapy depends on several factors such as the donor's health status and age. This work aims to prove the effect of PRP on cellular models of the CNS, considering the differences between PRP from young and elderly donors. Two different PRP pools were prepared from donors 65‒85 and 20‒25 years old. The cellular and molecular composition of both PRPs were analyzed. Subsequently, the cellular response was evaluated in CNS in vitro models, studying proliferation, neurogenesis, synaptogenesis, and inflammation. While no differences in the cellular composition of PRPs were found, the molecular composition of the Young PRP showed lower levels of inflammatory molecules such as CCL-11, as well as the presence of other factors not found in Aged PRP (GDF-11). Although both PRPs had effects in terms of reducing neural progenitor cell apoptosis, stabilizing neuronal synapses, and decreasing inflammation in the microglia, the effect of the Young PRP was more pronounced. In conclusion, the molecular composition of the PRP, conditioned by the age of the donors, affects the magnitude of the biological response.

circARRDC3 contributes to interleukin­13­induced inflammatory cytokine and mucus production in nasal epithelial cells via the miR­375/KLF4 axis.

Allergic rhinitis (AR) is a common inflammatory disorder of the nasal mucosa. It is a major risk factor for asthma development, and uncontrolled AR can lead to the worsening of asthma symptoms, which affects the quality of life and productivity of patients. Circular RNAs (circRNA) were reported to be involved in the pathogenesis of AR. The aim of the present study was to investigate the functional role of circRNA arrestin domain­containing 3 (circARRDC3) in AR progression. circARRDC3 knockdown suppressed the levels of granulocyte­macrophage colony­stimulating factor (GM­CSF) and eotaxin and mucin 5AC (MUC5AC) in IL­13­induced nasal epithelial cells. Moreover, circARRDC3 silencing promoted viability and suppressed apoptosis in IL­13­induced NECs. circARRDC3 targeted microRNA (miR)­375 and negatively regulated its expression. miR­375 inhibition reversed the effects of circARRDC3 knockdown on GM­CSF, eotaxin and MUC5AC expression levels, cell viability and cell apoptosis. In addition, miR­375 inhibited krueppel­like factor 4 (KLF4) expression through direct interaction, and miR­375 overexpression inhibited GM­CSF, eotaxin and MUC5AC expression levels, and cell apoptosis, which was abolished following KLF4 overexpression. In addition, circARRDC3, miR­375 and KLF4 were all dysregulated in the nasal mucosa of patients with AR. miR­375 expression was negatively correlated with circARRDC3 and KLF4 expression, and circARRDC3 expression was positively correlated with KLF4 expression. In conclusion, circARRDC3 contributed to the development of AR by regulating the miR­375/KLF4 axis. These findings may provide novel insights into the pathogenesis of AR.

IL-10, IL-13, Eotaxin and IL-10/IL-6 ratio distinguish breast implant-associated anaplastic large-cell lymphoma from all types of benign late seromas.

Breast implant-associated anaplastic large-cell lymphoma (BI-ALCL) is an uncommon peripheral T cell lymphoma usually presenting as a delayed peri-implant effusion. Chronic inflammation elicited by the implant has been implicated in its pathogenesis. Infection or implant rupture may also be responsible for late seromas. Cytomorphological examination coupled with CD30 immunostaining and eventual T-cell clonality assessment are essential for BI-ALCL diagnosis. However, some benign effusions may also contain an oligo/monoclonal expansion of CD30 + cells that can make the diagnosis challenging. Since cytokines are key mediators of inflammation, we applied a multiplexed immuno-based assay to BI-ALCL seromas and to different types of reactive seromas to look for a potential diagnostic BI-ALCL-associated cytokine profile. We found that BI-ALCL is characterized by a Th2-type cytokine milieu associated with significant high levels of IL-10, IL-13 and Eotaxin which discriminate BI-ALCL from all types of reactive seroma. Moreover, we found a cutoff of IL10/IL-6 ratio of 0.104 is associated with specificity of 100% and sensitivity of 83% in recognizing BI-ALCL effusions. This study identifies promising biomarkers for initial screening of late seromas that can facilitate early diagnosis of BI-ALCL.

Expression of mitochondrial TSPO and FAM173B is associated with inflammation and symptoms in patients with painful knee osteoarthritis.

OBJECTIVES: To characterize the expression profiles of two nuclear-encoded mitochondrial genes previously associated with chronic pain, the translocator protein (TSPO) and family with sequence similarity 173B (FAM173B), in different knee compartments from patients with painful knee OA. Also, to examine their association with the joint expression of inflammatory cytokines/chemokines and clinical symptoms. METHODS: The study was performed on 40 knee OA patients and 19 postmortem (PM) controls from which we collected the knee tissues: articular cartilage (AC), synovial membrane (SM) and subchondral bone (SB). Quantitative real-time polymerase chain reaction was used to determine the relative mRNA levels of TSPO, FAM173B, and inflammatory mediators IL6, IL8, IL10, IL12, MCP1, CCL11 and CCL17. OA patients rated their pain intensity (visual analogue scale), severity of knee-related outcomes (KOOS) and pain sensitivity assessed by pressure algometry. RESULTS: The gene expression of TSPO in SM was elevated in OA patients compared with control subjects while there were no group differences in AC and SB. Expression of FAM173B was reduced in SM but elevated in SB in OA patients compared with controls. The expression of TSPO and FAM173B in SM and SB was associated with the expression of inflammatory substances, but not in AC. Synovial expression of TSPO correlated with lower pain intensity and FAM173B with increased pressure pain sensitivity in OA. CONCLUSION: Our results suggest that altered expression of TSPO and FAM173B is associated with joint expression of inflammatory mediators and with clinical symptoms indicating the relevance for the pathophysiology of knee OA.