Periostin Production Research Articles

Suppressive Activity of Boiogito, a Japanese Traditional Kampo Medicine, on Periostin Secretion in Human Fibroblast-Like Synoviocytes In Vitro.

Background Knee osteoarthritis (KOA) is a prevalent degenerative disease that affects the knee joints, particularly among individuals aged over 40 years. It leads to pain, stiffness, and reduced quality of life; affects approximately 300 million individuals worldwide; and is increasing, particularly in developed nations. Although treatments for KOA range from conservative measures to surgical interventions, such as total knee arthroplasty (TKA), the financial burden of TKA in many countries underscores the urgent need for effective conservative therapies. The pathophysiology of KOA involves articular cartilage degeneration, increased subchondral bone turnover, synovitis, and periarticular soft tissue contracture. Abnormal bone turnover, intensified by factors, such as weight gain and knee injury, precedes cartilage degeneration. Synovitis, characterized by inflammation in the synovial tissue, plays a crucial role in perpetuating the disease by triggering a cascade of catabolic and proinflammatory mediators, including cytokines, such as interleukin (IL)-1 beta, tumor necrosis factor-alpha, and IL-13. Periostin, an extracellular matrix protein, is implicated in KOA progression, with its levels increasing with disease severity. Materials & methods In this study, the preventive effect of boiogito (BOT), a traditional herbal medicine, on periostin secretion in human fibroblast-like synoviocytes (hFLS) stimulated by IL-13 was investigated. Synoviocyte Growth Medium and recombinant human IL-13 were used for cell culture and stimulation. BOT was dissolved in phosphate-buffered saline and applied to cell cultures. Periostin secretion and mRNA expression were measured using enzyme-linked immunosorbent assay and quantitative reverse transcription polymerase chain reaction, respectively. Cell viability was assessed using an MTT assay, and signal transducer and activator of transcription factor 6 (STAT6) phosphorylation was examined using Western blotting. Results IL-13 stimulation of hFLS significantly increased periostin secretion, with levels rising above 20 ng/mL after 72 h of stimulation. Pretreatment with BOT dose-dependently suppressed periostin secretion, with doses of 1,000μg/mL significantly reducing periostin levels. Furthermore, BOT inhibited periostin mRNA expression and STAT6 phosphorylation in IL-13-stimulated hFLS, suggesting its potential in modulating IL-13-mediated inflammatory pathways in KOA. Conclusion This study demonstrated the preventive effect of BOT on periostin secretion in IL-13-stimulated hFLS, highlighting its potential as a therapeutic agent for KOA. By inhibiting periostin production and downstream signaling pathways, BOT may offer a promising conservative treatment option for KOA, addressing the inflammatory cascade implicated in disease progression. Further research is warranted to elucidate the specific herbal components responsible for the therapeutic effects of BOT and to validate its efficacy in clinical settings.

IL-6 induces periostin production in human ACL remnants: a possible mechanism causing post-traumatic osteoarthritis

ObjectivePerostin (POSTN) and IL-6 consistently elevated after ACL injury, and ACL has been proposed as the major source of POSTN. However, there is a lack of evidence whether IL-6 induces ACL remnants to produce POSTN. This study aimed to investigate the effect of IL-6 on POSTN production in ACL fibroblasts, which may help us understand more about the mechanism of PTOA after ACL injury and ACL reconstruction.MethodsACL remnants were harvested from 27 patients undergoing ACL reconstruction. Quantitative real-time polymerase chain reaction (PCR) was performed to examine the POSTN gene expression of ACL fibroblasts after treatment of different concentrations of IL-6. The POSTN protein production of ACL fibroblasts was determined using western blot analysis. The blockers of possible signaling pathways, including PI3K/Akt, Ras/MAPK, and JAK/STAT pathways, were added to test whether the effect of IL-6 on ACL fibroblast could be attenuated. ACL fibroblast and chondrocyte co-culture was carried out to determine the influence of ACL and IL-6 on chondrocytes.ResultsQuantitative real-time PCR showed that IL-6 time-dependently and dose-dependently increased POSTN gene expression of ACL fibroblast. Western blot analysis also revealed that IL-6 dose-dependently induced POSTN protein production. Regarding the chronicity of ACL injury, the POSTN protein production was comparable between ACL remnants which were derived within 3 months of injury and at least 6 months after injury. PI3K/Akt blockers could attenuate the effect of IL-6 on ACL remnants, whereas Ras/MAPK and JAK/STAT did not decrease POSTN production. The coexistence of ACL and IL-6 induced more MMP-13 and ADAMTS-4 by chondrocytes.ConclusionsIL-6 induced ACL remnants to produce POSTN. This effect could be attenuated by the PI3K/Akt blocker. Coexistence of IL-6 and ACL remnants may accelerate post-traumatic arthritis.

Abnormal basement membrane results in increased keratinocyte-derived periostin expression in psoriasis similar to wound healing

The psoriatic skin resembles wound healing, and it shows abnormalities at the basement membrane (BM), also in the non-lesional skin. Fibroblast-derived dermal periostin has well-known functions in wound healing and Th2-mediated diseases, such as atopic dermatitis. Here we show that serum periostin level was elevated in psoriatic patients, remarkably in the systemically treated ones. Obvious periostin positivity was detected in basal keratinocytes of the non-lesional, lesional, and previously-lesional psoriatic vs. healthy skin. Ex vivo skin models were generated to examine how different skin injuries affect periostin expression during wound healing. Our newly developed cultured salt-split model demonstrated that BM-injury induced periostin expression in basal keratinocytes, and periostin levels in the supernatant were also increased upon healing. In wound healing models, β1-integrin expression was similarly induced. β1-integrin blocking caused reduced periostin expression in in vitro scratch assay, indicating that β1-integrin can mediate periostin production. In contrast to atopic dermatitis, psoriatic basal keratinocytes are in an activated state and show a stable wound healing-like phenotype with the overexpression of periostin. This abnormal BM-induced wound healing as a potential compensatory mechanism can be initiated already in the non-lesional skin present in the lesion and keratinocytes can remain activated in the healed skin.

Caesalpinia sappan Linn. Ameliorates Allergic Nasal Inflammation by Upregulating the Keap1/Nrf2/HO-1 Pathway in an Allergic Rhinitis Mouse Model and Nasal Epithelial Cells.

Allergic rhinitis (AR) is a common upper-airway inflammatory disease of the nasal mucosa caused by immunoglobulin (IgE)-mediated inflammation. AR causes various painful clinical symptoms of the nasal mucosa that worsen the quality of daily life, necessitating the urgent development of therapeutic agents. Herein, we investigated the effects of Caesalpinia sappan Linn. heartwood water extract (CSLW), which has anti-inflammatory and antioxidant properties, on AR-related inflammatory responses. We examined the anti-inflammatory and anti-allergic effects of CSLW in ovalbumin (OVA)-induced AR mice and in primary human nasal epithelial cells (HNEpCs). Administration of CSLW mitigated allergic nasal symptoms in AR mice, decreased total immune cell and eosinophil counts in nasal lavage fluid, and significantly reduced serum levels of OVA-specific IgE, histamine, and Th2 inflammation-related cytokines. CSLW also inhibited the infiltration of several inflammatory and goblet cells, thereby ameliorating OVA-induced thickening of the nasal mucosa tissue. We found that CSLW treatment significantly reduced infiltration of eosinophils and production of periostin, MUC5AC, and intracellular reactive oxygen species through the Keap1/Nrf2/HO-1 pathway in HNEpCs. Thus, our findings strongly indicate that CSLW is a potent therapeutic agent for AR and can improve the daily life of patients by controlling the allergic inflammatory reaction of the nasal epithelium.

IL13 and periostin in active fibrogenic areas of the extrahepatic bile ducts in biliary atresia patients.

The leading pathology of biliary atresia (BA) is inflammatory and fibrous obstruction of extrahepatic bile duct, but the pathogenesis remains unclear. IL13 is a cytokine associated with allergies and inflammatory fibrosis, and periostin induces fibrogenesis by stimulation with IL13. We analyzed the involvement of IL13 and periostin in inflammatory fibrosis in the extrahepatic bile duct of BA patients. Surgically resected tissues from the hepatic hilar area of BA patients were immunostained with CD45, α-SMA, IL13 and periostin and statistically analyzed. Fibroblasts from the resected tissue were cultured with recombinant IL13, and periostin production was analyzed by quantitative polymerase chain reaction and Western blotting. IL13 was stained in 93% of large and micro bile ducts, and 92.1% matched with the CD45 location (p = 0.006) around the large bile ducts. Periostin staining correlated with the localization of IL13 and αSMA (p < 0.001) around the large bile ducts. Periostin mRNA and protein were upregulated by IL13 stimulation in cultured fibroblasts. IL13 was associated with induced periostin expression by fibroblasts, playing a vital role in the pathogenesis of fibrogenesis around the extrahepatic bile duct in BA.

Matricellular Protein Periostin Promotes Pericyte Migration in Fibrotic Airways.

Introduction: Periostin is a matricellular protein that is currently used as a biomarker for asthma. However, its contribution to tissue remodeling in allergic asthma is currently unknown. We have previously demonstrated that tissue-resident mesenchymal stem cells known as pericytes are a key cell type involved in airway remodeling. This is thought to be caused the uncoupling of pericytes from the microvasculature supporting the large airways, facilitated by inflammatory growth factors and cytokines. It is hypothesized that periostin may be produced by profibrotic pericytes and contribute to the remodeling observed in allergic asthma.Methods: Lung sections from mice with allergic airway disease driven by exposure to house dust mite (HDM) were stained using an anti-periostin antibody to explore its involvement in fibrotic lung disease. Human pericytes were cultured in vitro and stained for periostin to assess periostin expression. Migration assays were performed using human pericytes that were pretreated with TGF-β or periostin. ELISAs were also carried out to assess periostin expression levels in bronchoalveolar lavage fluid as well as the induction of periostin production by IL-13.Results: Immunostaining indicated that pericytes robustly express periostin, with increased expression following treatment with TGF-β. Migration assays demonstrated that pericytes treated with periostin were more migratory. Periostin production was also increased in HDM exposed mice as well as in cultured pericytes treated with IL-13.Conclusion: Periostin is produced by pericytes in response to TGF-β or IL-13, and periostin plays a key role in inducing pericyte migration. The increase in periostin expression in TGF-β or IL-13 treated pericytes suggests that IL-13 may trigger periostin production in pericytes whilst TGF-β modulates periostin expression to promote pericyte migration in the context of tissue fibrosis.

Anti-allergic effects of Asarum heterotropoides on an ovalbumin-induced allergic rhinitis murine model

Allergic rhinitis (AR) is a common chronic respiratory disease. Asarum heterotropoides (AH) is predicted to be a treatment for allergic diseases, but its therapeutic effect is unclear. We aimed to determine the anti-allergic effects of AH in mice with ovalbumin (OVA)-induced AR. OVA-induced AR mouse model was constructed, and AH was orally administered for a week; next, nasal clinical symptoms were evaluated. The levels of serum histamine, OVA-specific IgE, and IL-13 were measured by ELISA. Inflammatory cells, including leukocytes, neutrophils, eosinophils, and macrophages were counted in the nasal lavage fluid (NALF). Histopathological examinations of the nasal tissues were performed using H&E, Giemsa, and PAS staining. The production of periostin and eotaxin-3 from AH-treated human nasal epithelial cells (HNEpCs) in vitro, was measured using ELISA. Oral administration of AH alleviated allergic symptoms in mice with AR; significantly decreased levels of allergic mediators, such as serum histamine and OVA-specific IgE. The decrease in allergic symptoms positively correlated with the decrease in serum allergic mediators. The NALF of AH-treated AR mice demonstrated lower number of eosinophils. AH demonstrated a capacity to reduce the infiltration of mast cells, eosinophils, and goblet cells, thereby resulting in thinner nasal tissues. Moreover, treatment of HNEpCs with AH demonstrated suppressed production of periostin and eotaxin-3. AH exerts a therapeutic effect in modulating AR through multi-target and multi-function influence on regulating B cells, mast cells, eosinophils, goblet cells, and epithelial cells.

POS0429 INTERLEUKIN-4 ACTIVATES EOSINOPHILS AND CCR3-POSITIVE T HELPER CELLS MIGRATION TO FASCIA AND PROMOTES FIBROSIS IN EOSINOPHILIC FASCIITIS

Background:Eosinophilic fasciitis (EF) is a rare disease that causes inflammation and fibrosis mainly in the fascia of the extremities with eosinophilia. It has been reported that the hypertrophied fascia in EF shows inflammatory cell infiltration by the lymphocytes and eosinophils and increased expression of fibrosis-related cytokines genes in fibroblast [1]. However, its pathophysiology in the fascia remains unresolved.Objectives:Therefore, we focused on fascial fibroblasts and aimed to determine the role of interleukin-4 (IL-4) in eosinophil and helper T cell infiltration and fibrosis in fascial fibroblast in EF.Methods:Fascial fibroblasts were obtained from fascia biopsy of a patient with EF, and were stimulated with pre- and post-treatment serum of a patient with EF and healthy control, followed by microarray to analyze gene expression. Fascial fibroblasts were stimulated with IL-4 10 ng/mL, and gene expression of IL-4 receptor and CCR3 ligands, CCL7 and CCL11 were measured by qPCR. Transforming growth factor (TGF) -β and periostin in the pre- and post-treatment serum of a patient with EF and conditioned medium of fascial fibroblasts stimulated with IL-4 were measured by ELISA. To examine the role of IL-4 in proliferation, we performed in proliferation assays using fascial fibroblasts treated with IL-4. CCR3-positive T cells in the fascial tissue of EF, dermatomyositis, and polymyositis patients were evaluated by immunostaining.Results:By microarray analysis, CCL7 and CCL11 expression of fascial fibroblasts stimulated with pre-treatment EF serum was higher than that in post-treatment EF serum and control serum. CCL7 and CCL11 mRNA in IL-4 stimulated facial fibroblasts were increased by 5.1-fold and 7.3-fold, respectively. TGF-β and periostin in IL-4 stimulated facial fibroblast conditioned medium were also increased. In addition, TGF-β and periostin in EF serum were gradually decreased by treatment for 4 and 10 weeks, compared to before treatment. Finally, fascial fibroblast proliferation was significantly increased by stimulation with IL-4. Furthermore, infiltration of CCR3-positive T cells was specific to the fascial tissue of EF.Conclusion:In EF, IL-4 enhances the production of CCR3 ligands, TGF-β, and periostin from fascial fibroblasts. As a result, it promotes the migration of eosinophils and CCR3-positive T helper cells to the fascia and fibrosis. These results suggest that inhibition of IL-4 pathway could be a novel strategy for eosinophilic fasciitis.

In This Issue

HomeCirculation ResearchVol. 127, No. 9In This Issue Free AccessResearch ArticlePDF/EPUBAboutView PDFView EPUBSections ToolsAdd to favoritesDownload citationsTrack citationsPermissions ShareShare onFacebookTwitterLinked InMendeleyRedditDiggEmail Jump toFree AccessResearch ArticlePDF/EPUBIn This Issue Ruth Williams Ruth WilliamsRuth Williams Search for more papers by this author Originally published8 Oct 2020https://doi.org/10.1161/RES.0000000000000443Circulation Research. 2020;127:1119is related toEpigenomic and Transcriptomic Dynamics During Human Heart OrganogenesisEnhancement of ALK1 Signaling as a HHT Therapy (p 1122)Boosting activity of kinase ALK1 could be a treatment for hereditary hemorrhagic telangiectasia, say Kim et al.Download figureDownload PowerPointHereditary hemorrhagic telangiectasia (HHT) is a genetic disease characterized by vascular abnormalities called arteriovenous malformations (AVMs) in which arteries are directly connected to veins. Without intervening capillaries, blood pressure in the veins becomes excessive and can cause aneurysms and ruptures. Indeed, HHT patients often endure frequent bleeds and anemia, and sometimes suffer more severe events such as brain or pulmonary hemorrhages. HHT is caused by mutations in one of three genes—ALK1, ENG, or SMAD4—all of which encode components of a TGFβ-related growth factor signaling pathway. Because ninety percent of HHT cases are driven by ALK1 or ENG mutations, Kim et al focused on these two factors, examining how they might interact and compensate for each other. Using mice that conditionally lacked either ALK1 or ENG and had AVMs as a result, the team found that genetically increasing ALK1 production could prevent AVMs in the ENG- and ALK-lacking mice, but that increasing ENG production could only compensate for deficiency of ENG, not ALK1. Together these results suggest that ALK1 acts downstream of ENG in their shared signaling pathway and that augmenting ALK1 production could be a therapeutic approach for treating HHT.Periostin Is a Target to Treat PH (p 1138)Nie et al suggest periostin as a novel therapeutic target for treating pulmonary hypertension.Download figureDownload PowerPointPulmonary hypertension (PH) is a life threatening disease in which excess proliferation of vascular smooth muscle cells (VSMCs) and deposition of extracellular matrix (ECM) thickens the walls of the lung vasculature causing an increase in pulmonary blood pressure. Vasodilatory medications are used to treat the symptoms of the disease, but there are no available treatments that target the underlying pathologic remodeling. Nie and colleagues now suggest that drugs targeting the secreted ECM protein periostin might work. Periostin is unusually abundant in the arteries of PH patients’ lungs and is thought to be involved in cell adhesion and wound healing mechanisms such as proliferation and migration of VSMCs. The team has now confirmed the increased production of periostin in patient lungs and shown the same is true for mice with induced PH. Moreover they showed that genetic deletion of periostin attenuated PH in mice, while suppression of periostin via RNA inhibition could even reverse pathologic vessel thickening and right ventricle hypertrophy. The team went on to identify factors—HIF-1A and TrkB—that mediate periostin’s effects in cultured arterial cells, and suggest that blocking either these factors or periostin itself could be novel strategies for treating PH.Gene Regulatory Dynamics of Developing Human Heart (p e184)VanOudenhove et al examine the epigenomics and transcriptomics of the developing human heart.Download figureDownload PowerPointCongenital heart defects (CHDs) are a common form of birth abnormality. While some genes have been linked to CHDs, the majority—nearly sixty percent—have unknown etiologies. It’s thought that multiple genetic and environmental factors contribute to CHDs, one of which could be variations in regulatory regions of the genome. To find such heart-specific regulatory regions, VanOudenhove and colleagues examined heart tissue from human embryos obtained 4 to 8 weeks after conception. They performed chromatin immunoprecipitation (ChIP) experiments to scour the heart genomes for histone modifications associated with increased or decreased gene transcription. They also performed transcriptome analysis to see whether the genomic regions identified by ChIP corresponded with the activity status of nearby genes. In total, the team found more than 12 thousand previously unknown enhancers that were enriched for binding sites for heart-specific transcription factors (including GATA, MEF2 and NKX) and that tended to be close to genes activated in the heart. Many of the regions also contained sequence variations previously associated with atrial fibrillation. These newly identified sites are potential CHD candidate loci, say the authors, who have made their data freely available for study. Previous Back to top Next FiguresReferencesRelatedDetailsRelated articlesEpigenomic and Transcriptomic Dynamics During Human Heart OrganogenesisJennifer VanOudenhove, et al. Circulation Research. 2020;127:e184-e209 October 9, 2020Vol 127, Issue 9Article InformationMetrics Download: 270 © 2020 American Heart Association, Inc.https://doi.org/10.1161/RES.0000000000000443 Originally publishedOctober 8, 2020 PDF download

IL-13-induced periostin production from human bronchial epithelial cells and human nasal epithelial cells

IL-13-induced periostin production from human bronchial epithelial cells and human nasal epithelial cells

Serum periostin levels in asthma patients in relation to omalizumab therapy and presence of chronic rhinosinusitis with nasal polyps.

IntroductionThe serum periostin level is a promising biomarker of type 2- high inflammation pattern of bronchial asthma. It has been proven that serum periostin levels decrease in response to systemic and inhaled corticosteroid (ICS) therapy. However, we have only limited knowledge about changes in serum periostin levels reflecting omalizumab (OMA) treatment and other variables, such as chronic rhinosinusitis with nasal polyps (CRSwNP).AimTo critically appraise clinically relevant parameters influencing periostin levels in asthma patients.Material and methodsA pilot, cross-sectional, observational study to assess serum periostin levels of 48 asthma patients (38 treated by conventional therapy comprising ICS and 10 treated by ICS and OMA as an add-on therapy) with respect to asthma clinical traits, comorbidities and to other biomarkers of type 2-high asthma phenotype (total IgE, absolute and relative eosinophil count, eosinophilic cationic protein (ECP) and a fraction of exhaled NO (FeNO)).ResultsSerum periostin correlates with total IgE levels (Spearman rho = 0.364, p = 0.025) in a subgroup of conventionally treated patients, and with eosinophil count (Spearman rho = 0.401, p = 0.021) in a subgroup of patients with concurrent CRSwNP. Serum periostin levels were decreased in omalizumab-treated patients in comparison to conventionally treated patients (p = 0.025). This effect was remarkably apparent only if CRSwNP was not present (p = 0.005). Conversely, we measured elevated periostin levels in OMA-treated patients with concurrent CRSwNP (p = 0.017).ConclusionsSerum periostin production is significantly associated with treatment modality (omalizumab vs. conventional) and presence of CRSwNP. These variables need to be taken into account to interpret periostin levels accurately.

Periostin Promotes Cell Proliferation and Macrophage Polarization to Drive Repair after AKI.

The matricellular protein periostin has been associated with CKD progression in animal models and human biopsy specimens. Periostin functions by interacting with extracellular matrix components to drive collagen fibrillogenesis and remodeling or by signaling through cell-surface integrin receptors to promote cell adhesion, migration, and proliferation. However, its role in AKI is unknown. We used mice with conditional tubule-specific overexpression of periostin or knockout mice lacking periostin expression in the renal ischemia-reperfusion injury model, and primary cultures of isolated tubular cells in a hypoxia-reoxygenation model. Tubular epithelial cells showed strong production of periostin during the repair phase of ischemia reperfusion. Periostin overexpression protected mice from renal injury compared with controls, whereas knockout mice showed increased tubular injury and deteriorated renal function. Periostin interacted with its receptor, integrin-β1, to inhibit tubular cell cycle arrest and apoptosis in in vivo and in vitro models. After ischemia-reperfusion injury, periostin-overexpressing mice exhibited diminished expression of proinflammatory molecules and had more F4/80+ macrophages compared with knockout mice. Macrophages from periostin-overexpressing mice showed increased proliferation and expression of proregenerative factors after ischemia-reperfusion injury, whereas knockout mice exhibited the opposite. Coculturing a macrophage cell line with hypoxia-treated primary tubules overexpressing periostin, or treating such macrophages with recombinant periostin, directly induced macrophage proliferation and expression of proregenerative molecules. In contrast to the detrimental role of periostin in CKD, we discovered a protective role of periostin in AKI. Our findings suggest periostin may be a novel and important mediator of mechanisms controlling renal repair after AKI.

099 Skin atrophy due to topical glucocorticoids is less common in patients with atopic dermatitis than psoriasis

Atopic dermatitis (AD) and psoriasis are the most common chronic inflammatory dermatitis treated with topical glucocorticoids (GC). Although long-term use of topical GC may induce skin atrophy including striae distensa (SD), AD patients appear to cause less skin atrophy compared to psoriasis. Chronic AD lesions are characterized by eosinophil infiltration and dermal fibrosis. Periostin, encoded by POSTN gene, augments TGF-β production in eosinophils and inversely activated eosinophils can induce periostin production in fibroblasts. Periostin involves in tissue remodeling such as fibrosis and lichenification of chronic AD lesion. This study was designed to reveal that AD patients have less skin atrophy due to skin fibrosis related with eosinophils and periostin compared to psoriasis. For the skin atrophy including SD, we enrolled AD and psoriasis patients with similar ages and treatment durations. For the skin fibrosis and eosinophils, we compared lesional and non-lesional skin from the patients with AD, psoriasis, and allergic contact dermatitis (ACD). Also, we compared mRNA expression in the dermis from the mouse model of AD or psoriasis. AD patients have less SD than psoriasis. In AD patients, the ratio of the dermal fibrous tissues was significantly higher than psoriasis. Tissue and blood eosinophils were significantly higher in AD patients than ACD as well as psoriasis. Dermal thickness and fibrosis were significantly higher in AD mice than psoriasis even after the treatment with topical GC. There was a significant positive correlation between dermal fibrosis and tissue eosinophils counts. The mRNAs of POSTN, TGFB1, COL11A1, MMP1, and IL1R2 were significantly up-regulated in AD mice than psoriasis. We found that AD patients had less skin atrophy than psoriasis after the long-term use of topical GC. It might be resulted from the skin fibrosis caused by upregulation of tissue eosinophils and periostin.

Proteomic analysis of synovial fluid identifies periostin as a biomarker for anterior cruciate ligament injury

Emerging evidence suggests that injury to the anterior cruciate ligament (ACL) typically initiates biological changes that contribute to the development of osteoarthritis (OA). The molecular biomarkers or mediators of these biological events remain unknown. The goal of this exploratory study was to identify novel synovial fluid biomarkers associated with early biological changes following ACL injury distinct from findings in end-stage OA. Synovial fluid was aspirated from patients with acute (≤30 days) and subacute (31-90 days) ACL tears and from patients with advanced OA and probed via tandem mass spectrometry for biomarkers to distinguish OA from ACL injury. Periostin (POSTN) was identified as a potential candidate. Further analyses of POSTN were performed in synovial fluid, OA cartilage, torn ACL remnants, and cultured cells and media by Western blot, PCR, immunostaining and ELISA. Synovial fluid analysis revealed that POSTN exhibited higher expression in subacute ACL injury than OA. POSTN expression was relatively low in cartilage/chondrocytes suggesting it is also produced by other intra-articular tissues. Conversely, high and time-dependent expression of POSTN in ACL tear remnants and isolated cells was consistent with the synovial fluid results. Elevated POSTN may provide a synovial fluid biomarker of subacute ACL injury setting separate from OA. Increased expression of POSTN in ACL suggests that the injured ACL may play a pivotal role in POSTN production, which is sensitive to time from injury. Previous studies have shown potential catabolic effects of POSTN, raising the possibility that POSTN contributes to the initiation of joint degeneration and may offer a window of opportunity to intervene in the early stages of post-traumatic OA.

Matrisome Properties of Scaffolds Direct Fibroblasts in Idiopathic Pulmonary Fibrosis.

In idiopathic pulmonary fibrosis (IPF) structural properties of the extracellular matrix (ECM) are altered and influence cellular responses through cell-matrix interactions. Scaffolds (decellularized tissue) derived from subpleural healthy and IPF lungs were examined regarding biomechanical properties and ECM composition of proteins (the matrisome). Scaffolds were repopulated with healthy fibroblasts cultured under static stretch with heavy isotope amino acids (SILAC), to examine newly synthesized proteins over time. IPF scaffolds were characterized by increased tissue density, stiffness, ultimate force, and differential expressions of matrisome proteins compared to healthy scaffolds. Collagens, proteoglycans, and ECM glycoproteins were increased in IPF scaffolds, however while specific basement membrane (BM) proteins such as laminins and collagen IV were decreased, nidogen-2 was also increased. Findings were confirmed with histology, clearly showing a disorganized BM. Fibroblasts produced scaffold-specific proteins mimicking preexisting scaffold composition, where 11 out of 20 BM proteins were differentially expressed, along with increased periostin and proteoglycans production. We demonstrate how matrisome changes affect fibroblast activity using novel approaches to study temporal differences, where IPF scaffolds support a disorganized BM and upregulation of disease-associated proteins. These matrix-directed cellular responses emphasize the IPF matrisome and specifically the BM components as important factors for disease progression.

Periostin is induced by IL-4/IL-13 in dermal fibroblasts and promotes RhoA/ROCK pathway-mediated TGF-β1 secretion in abnormal scar formation

Excess scar formation can occur after skin injurふy and lead to abnormal scar formation, such as keloids and hypertrophic scars, which are characterised by substantial deposition of extracellular matrix in the dermis. Periostin, an extracellular matrix protein that plays a crucial role in skin development and maintaining homeostasis, is also involved in skin disorders such as systemic/limited scleroderma, wound closure, and abnormal scar formation. However, the mechanism of periostin involvement in abnormal scar formation is not yet fully understood. In this study, we investigated the mechanism by which periostin is involved in abnormal scar formation. Treatment of human dermal fibroblasts (HDFs) with IL-4 and IL-13, which are cytokines of Th2 type immune responses that are up-regulated in abnormal scars, dramatically elevated the levels of periostin mRNA and protein, and also promoted the secretion of periostin by HDFs. Transforming growth factor-β1 (TGF-β1) had the same effect on HDFs as IL-4 and IL-13. Stimulation of HDFs with periostin promoted RhoA/ROCK pathway-mediated TGF-β1 secretion from HDFs. Our results suggest that IL-4 and IL-13 induce periostin expression and secretion, and in turn, secreted periostin induces RhoA/ROCK pathway-mediated TGF-β1 secretion. Secreted TGF-β1 then induces further periostin production and secretion, thereby promoting abnormal scar formation.

P116 PERIOSTIN PROMOTES ENDOPLASMIC RETICULUM STRESS-MEDIATED FIBROSIS IN CROHN’S DISEASE

Fibrostenosis complicates Crohn’s disease in the 30-50% of patients with a Montreal B2 phenotype. Current medical therapies can delay but no prevent critical fibrostenosis and the need for surgery. Transforming growth factor-β1 (TGF-β1)-induced excess extracellular matrix (ECM) protein production in ileal subepithelial myofibroblasts (SEMF) are a major structural component of fibrosis. The role of endoplasmic reticulum (ER) stress in the pathogenesis of Crohn’s disease has been delineated in epithelial and lamina propria mononuclear cells and more recently we have delineated the role of ER stress in SEMF of fibrostenotic ileum. ER stress marker and chaperone GRP78 can bind to type I collagen in a stimulated extracellular environment, which suggests the potential link between ER stress and fibrosis formation. The secreted matricellular protein periostin, typically promotes fibrosis in a TGF-β1-dependent fashion and interacts with plasma membrane integrin family members, including αVβ3. We have previously shown αVβ3 integrin is crucial for LAP-TGF-β1 activation in ileal mesenchymal cells, SEMF and smooth muscle cells. Our RNAseq data identified TGFB1, ITGAV, ITGB3 COL1A1 and all as significantly upregulated (25-200 fold) in SEMF of strictured ileum. However, the role of periostin in SEMF during the ER stress-mediated component of intestinal fibrosis in Crohn’s disease remains unknown. To determine the role of a periostin during the development of intestinal fibrosis in fibrostenotic Crohn’s disease, SEMF were isolated from the patients with each Crohn’s disease phenotype and place them into primary culture or 3D culture. Periostin expression was determined by qRT-PCR and immunoblot analysis. Binding of periostin to αvβ3 integrin was determined by proximity ligation hybridization assay. Knockdown of periostin in cells was accomplished with siRNA. Active and total TGF-β1 protein levels were measured by ELISA. Our data show that periostin expression and production are increased in SEMF from strictured ileum compared to normal ileum in the same patient. IL-6 promotes the interaction between secreted periostin and integrin αvβ3 on the SEMF cell membrane. This was associated with increased levels of active TGF-β1 and increased the resulting production of collagen 1α1. Induction of ER stress in SEMF from normal ileum with tunicamycin elicited increased expression of periostin and αVβ3. The ER stress-induced increase in αVβ3, however, was inhibited by siRNA-mediated knockdown of periostin. The increase in active TGF-β1 seen following ER stress was also decreased in the SEMF in which periostin was knocked down. Our data suggest ER stress-induced periostin links increased αVβ3 integrin and αVβ3-dependent activation of latent TGF-β1 and the resulting increase in collagen production (Figure 1).

P116 PERIOSTIN PROMOTES ENDOPLASMIC RETICULUM STRESS-MEDIATED FIBROSIS IN CROHN’S DISEASE

Fibrostenosis complicates Crohn’s disease in the 30-50% of patients with a Montreal B2 phenotype. Current medical therapies can delay but no prevent critical fibrostenosis and the need for surgery. Transforming growth factor-β1 (TGF-β1)-induced excess extracellular matrix (ECM) protein production in ileal subepithelial myofibroblasts (SEMF) are a major structural component of fibrosis. The role of endoplasmic reticulum (ER) stress in the pathogenesis of Crohn’s disease has been delineated in epithelial and lamina propria mononuclear cells and more recently we have delineated the role of ER stress in SEMF of fibrostenotic ileum. ER stress marker and chaperone GRP78 can bind to type I collagen in a stimulated extracellular environment, which suggests the potential link between ER stress and fibrosis formation. The secreted matricellular protein periostin, typically promotes fibrosis in a TGF-β1-dependent fashion and interacts with plasma membrane integrin family members, including αVβ3. We have previously shown αVβ3 integrin is crucial for LAP-TGF-β1 activation in ileal mesenchymal cells, SEMF and smooth muscle cells. Our RNAseq data identified TGFB1, ITGAV, ITGB3 COL1A1 and all as significantly upregulated (25-200 fold) in SEMF of strictured ileum. However, the role of periostin in SEMF during the ER stress-mediated component of intestinal fibrosis in Crohn’s disease remains unknown. To determine the role of a periostin during the development of intestinal fibrosis in fibrostenotic Crohn’s disease, SEMF were isolated from the patients with each Crohn’s disease phenotype and place them into primary culture or 3D culture. Periostin expression was determined by qRT-PCR and immunoblot analysis. Binding of periostin to αvβ3 integrin was determined by proximity ligation hybridization assay. Knockdown of periostin in cells was accomplished with siRNA. Active and total TGF-β1 protein levels were measured by ELISA. Our data show that periostin expression and production are increased in SEMF from strictured ileum compared to normal ileum in the same patient. IL-6 promotes the interaction between secreted periostin and integrin αvβ3 on the SEMF cell membrane. This was associated with increased levels of active TGF-β1 and increased the resulting production of collagen 1α1. Induction of ER stress in SEMF from normal ileum with tunicamycin elicited increased expression of periostin and αVβ3. The ER stress-induced increase in αVβ3, however, was inhibited by siRNA-mediated knockdown of periostin. The increase in active TGF-β1 seen following ER stress was also decreased in the SEMF in which periostin was knocked down. Our data suggest ER stress-induced periostin links increased αVβ3 integrin and αVβ3-dependent activation of latent TGF-β1 and the resulting increase in collagen production (Figure 1).

Elevated levels of periostin and TGF-β1 in the bronchoalveolar lavage fluid of patients with idiopathic eosinophilic pneumonia.

Periostin is induced in bronchial epithelial cells and fibroblasts by various stimuli including interleukin (IL)13 and transforming growth factor (TGF)-β1, and is involved in allergic diseases such as asthma and atopic dermatitis, playing an important role in tissue remodeling and fibrosis. The role of periostin in the pathogenesis of eosinophilic lung diseases, however, is unclear. To examine the contribution of periostin to eosinophilic inflammation of the lung in humans, we evaluated periostin, IL-13, and TGF-β1 levels in the bronchoalveolar lavage fluid (BALF) of patients with eosinophilic pneumonia (EP). Periostin, IL-13, and TGF-β1 concentrations in the BALF were measured by enzyme-linked immunosorbent assay in patients with acute EP, chronic EP, idiopathic pulmonary fibrosis (IPF), and sarcoidosis. Further, we analyzed the relationship between periostin, IL-13, and TGFβ-1, levels and the number of inflammatory cells in the BALF. The absolute number of eosinophils, and the periostin, IL-13, and TGF-β1 levels in the BALF were significantly higher in patients with EP than in patients with IPF and sarcoidosis. Concentrations of periostin significantly correlated with the concentrations of TGF-β1, but not those of IL-13, in the BALF of patients with EP. Periostin levels also significantly correlated with the absolute number of eosinophils in the BALF of patients with IPF, but not EP. Our findings suggest that TGF-β1 might increase the production of periostin in the lungs of patients with EP. Periostin might contribute the pathogenesis of not only EP, but also IPF.

Year in review 2017: Chronic obstructive pulmonary disease and asthma.

Year in review 2017: Chronic obstructive pulmonary disease and asthma.