Periostin Expression Research Articles

Kidney Injury Molecule-1 and Periostin Urinary Excretion and Tissue Expression Levels and Association with Glomerular Disease Outcomes

Introductions: Kidney injury molecule-1 (KIM-1) and periostin (POSTN) are proximal and distal tubule injury biomarkers. We tested whether baseline urine KIM-1/Cr (uKIM-1/Cr) and/or uPOSTN/Cr correlated with disease severity or improved a remission prediction model. Methods: Baseline uKIM-1/Cr and uPOSTN/Cr were measured on spot urine samples from immunosuppression-free patients enrolled in Nephrotic Syndrome Study Network until December 15, 2014. Urine protein/Cr (UPCR) and albumin/Cr (UACR) were measured at baseline, 4 months, and until last follow-up. Glomerular and tubulointerstitial (TI) expression arrays were analyzed from a baseline research renal biopsy core collected during a clinically indicated biopsy. Renal diagnoses were centrally confirmed, sections scanned, and measured morphometrically. Correlations between baseline uKIM-1/Cr and uPOSTN/Cr and UPCR, UACR, histopathologic features, glomerular and TI KIM-1 and POSTN expression levels, and renal outcomes were assessed. Results: Baseline uKIM-1/cr correlated with UPCR and UACR and were associated with complete remission (CR) after adjustment for proteinuria, histopathologic diagnosis, and treatment. Baseline uKIM-1/Cr also correlated with degree of foot process effacement and acute tubular injury. Glomerular and TI KIM-1 expression levels correlated with UPCR and UACR. Higher TI KIM-1 expression levels correlated with interstitial fibrosis, tubular atrophy, and global glomerulosclerosis, while glomerular KIM-1 expression correlated with time to remission. Findings for POSTN were of lesser statistical strength. Discussion/Conclusion: Lower baseline uKIM-1/Cr values were associated with more rapid time to CR after adjusting for proteinuria, histopathologic diagnosis, and treatment. Increased TI KIM-1 expression levels in proteinuric states were associated with chronic morphological injury; lower glomerular expression levels were associated with a greater potential for proteinuria reversibility.

Expression of Musashi-1 Increases in Bone Healing.

Musashi-1 (MSI1) is an RNA-binding protein that regulates progenitor cells in adult and developing organisms to maintain self-renewal capacities. The role of musashi-1 in the bone healing environment and its relation with other osteogenic factors is unknown. In the current study, we analyze the expression of MSI1 in an experimental model of rat femoral bone fractures. We also analyze the relation between MSI1 expression and the expression of two osteogenic markers: periostin (POSTN) and runt-related transcription factor 2 (RUNX2). We use histological, immunohistochemical, and qPCR techniques to evaluate bone healing and the expression of MSI1, POSTN, and RUNX2 over time (4, 7, and 14 days). We compare our findings with non-fractured controls. We find that in bone calluses, the number of cells expressing MSI1 and RUNX2 increase over time and the intensity of POSTN expression decreases over time. Within bone calluses, we find the presence of MSI1 expression in mesenchymal stromal cells, osteoblasts, and osteocytes but not in hypertrophic chondrocytes. After 14 days, the expression of MSI1, POSTN, and RUNX2 was significantly correlated. Thus, we conclude that musashi-1 potentially serves in the osteogenic differentiation of mesenchymal stromal cells and bone healing. Therefore, further studies are needed to determine the possibility of musashi-1′s role as a clinical biomarker of bone healing and therapeutic agent for bone regeneration.

Role of Periostin Expression in Canine Osteosarcoma Biology and Clinical Outcome.

Periostin is a matricellular protein important in regulating bone, tooth, and cardiac development. In pathologic conditions, periostin drives allergic and fibrotic inflammatory diseases and is also overexpressed in certain cancers. Periostin signaling in tumors has been shown to promote angiogenesis, metastasis, and cancer stem cell survival in rodent models, and its overexpression is associated with poor prognosis in human glioblastoma. However, the role of periostin in regulating tumorigenesis of canine cancers has not been evaluated. Given its role in bone development, we sought to evaluate mRNA and protein expression of periostin in canine osteosarcoma (OS) and assess its association with patient outcome. We validated an anti-human periostin antibody cross-reactive to canine periostin via western blot and immunohistochemistry and evaluated periostin expression in microarray data from 49 primary canine OS tumors and 8 normal bone samples. Periostin mRNA was upregulated greater than 40-fold in canine OS tumors compared to normal bone and was significantly correlated with periostin protein expression based on quantitative image analysis. However, neither periostin mRNA nor protein expression were associated with time to metastasis in this cohort. Gene Set Enrichment Analysis demonstrated significant enhancement of pro-tumorigenic pathways including canonical WNT signaling, epithelial-mesenchymal transition, and angiogenesis in periostin-high tumors, while periostin-low tumors demonstrated evidence of heightened antitumor immune responses. Overall, these data identify a novel antibody that can be used as a tool for evaluation of periostin expression in dogs and suggest that investigation of Wnt pathway-targeted drugs in periostin overexpressing canine OS may be a potential therapeutic target.

Abstract PO051: Fibroblast lineage tracing in metastatic and pre-metastatic microenvironments

Abstract Metastatic breast cancer is one of the leading causes of cancer-related mortality among women. However, the complex process of tumor cell dissemination and colonization of distant tissues is incompletely understood and largely incurable using existing therapies. Metastatic disease is not solely driven by tumor cell-intrinsic properties but is instead a consequence of dynamic interactions between cancer cells and other cell types in the tumor microenvironment (TME) including cancer-associated fibroblasts (CAFs). However, CAFs are known to display paradoxical functions across different cancer types, suggesting that heterogeneous CAF subpopulations play diverse or opposing roles in the TME. It was recently shown that periostin, a TGFβ-induced matricellular protein important for collagen cross-linking and extracellular matrix stiffening, can distinguish CAF subtypes in breast cancer. This specific CAF subpopulation is not well characterized, and their functional properties and contributions to disease progression have not been well-studied in vivo. Here, we have used in vivo genetic labelling of periostin+ subpopulations (Postn-Cre:ZSGreenl/s/l) and their progeny to assess their expansion and function during mammary tumor growth and metastasis. We find that ZSGreen-labelled periostin-expressing CAFs arise from mesenchymal-lineage cells and are more abundant in primary tumors, metastatic, and pre-metastatic sites of high-metastatic tumors versus their low-metastatic counterparts. We also find differential collagen organization in high- versus low-metastatic tumors: high-metastatic tumors display longer, aligned collagen fibers whereas low-metastatic tumors display shorter, disorganized fibers. Knocking down periostin in primary human breast CAFs inhibits tumor cell invasion through collagen in a CAF/tumor cell co-culture spheroid assay, indicating that periostin-expressing CAFs may remodel the extracellular matrix via collagen cross-linking to facilitate cell invasion. Finally, we show that periostin/ZSGreen+ cells in highly metastatic tumors are larger and more spindle-shaped, and that periostin knockdown in primary human breast CAFs reduces their size and ability to migrate, suggesting that periostin expression associates with CAF biomechanical properties such as spreading and motility. Our results indicate that highly-metastatic cancer cells mobilize periostin-expressing CAFs in the TME, which may promote collagen remodeling and collective migration of CAFs and cancer cells during tissue invasion. Citation Format: Jamie L. Null, Dae Joong Kim, Yu Zhang, James V. McCann, Thomas H. Barker, Andrew C. Dudley. Fibroblast lineage tracing in metastatic and pre-metastatic microenvironments [abstract]. In: Proceedings of the AACR Virtual Special Conference on the Evolving Tumor Microenvironment in Cancer Progression: Mechanisms and Emerging Therapeutic Opportunities; in association with the Tumor Microenvironment (TME) Working Group; 2021 Jan 11-12. Philadelphia (PA): AACR; Cancer Res 2021;81(5 Suppl):Abstract nr PO051.

Over-expression of stromal periostin correlates with poor prognosis of cutaneous squamous cell carcinomas.

Periostin, an extracellular matrix macromolecule implicated in tumorigenesis, serves as a prognostic marker for many cancer types. However, there are no data on periostin expression in cutaneous squamous cell carcinoma (cSCC). This study examined periostin expression in patients with cSCC and explored its clincopathological relationship and prognosis. Using immunohistochemistry and ImageJ analysis, we compared periostin expression in 95 cSCCs across a spectrum of cSCC aggressiveness: cSCC in situ (SCCIS) (n=25), low-risk cSCC (LR-cSCC) (n=26), high-risk cSCC (HR-cSCC) (n=38), and cSCC in recessive dystrophic epidermolysis bullosa patients (RDEB cSCC) (n=6). Immunohistochemistry demonstrated periostin expression within the intra-tumoral stroma but not within tumor cells. Periostin levels significantly (P<0.001) increased from SCCIS, LR-cSCC, HR-cSCC to RDEB SCC. The stroma of most of the cSCCs we evaluated contained cancer-associated fibroblasts with a myofibroblastic (α -SMA-positive) phenotype. Co-localization of periostin with α-SMA, evidence of fibroblast periostin expression, and absence of keratinocyte or tumor cell periostin expression suggest that, in cSCC, periostin is a product of the peritumoral microenvironment and not the tumor cells themselves. Our data indicate that fibroblast periostin expression is highly correlated with the aggressiveness of cSCC, and may thereby provide a molecular marker that will be useful for subtyping and diagnosing cSCCs according to their biological nature.

Metformin attenuates hyperlipidaemia-associated vascular calcification through anti-ferroptotic effects

Ferroptosis is a form of regulated cell death that involves metabolic dysfunction resulting from iron-dependent excessive lipid peroxidation. Elevated plasma levels of free fatty acids are tightly associated with cardiometabolic risk factors in patients with obesity, diabetes mellitus, and metabolic syndrome. Metformin (Met) is an antidiabetic drug with beneficial cardiovascular disease effects. The aim of this study was to determine the effects of Met on ferroptosis induced by lipid overload and the effects of these changes on vascular smooth muscle cells (VSMCs) calcification. We developed a hyperlipidaemia-related vascular calcification in vivo model with rats fed a high-fat diet combined with vitamin D3 plus nicotine, and palmitic acid (PA), the most abundant long-chain saturated fatty acid in plasma, was used to induce lipid overload and develop an oxidative stress-related calcification model in vitro. The results showed that Met inhibits hyperlipidaemia-associated calcium deposition in the rat aortic tissue. In vitro, treatment of VSMCs with PA stimulates ferroptosis concomitant with increased calcium deposition in VSMCs, while pretreatment with Met attenuates these effects. Furthermore, PA also promotes the protein expression of the extracellular matrix protein periostin (POSTN) and its secretion into the extracellular environment. More importantly, upregulation of POSTN increased the sensitivity of cells to ferroptosis. Mechanistically, upregulation of POSTN suppresses SLC7A11 expression through the inhibition of p53 in VSMCs, which contributes to a decrease in glutathione synthesis and therefore triggers ferroptosis. Interestingly, overexpression of p53 attenuates the inhibitory effect of POSTN on SLC7A11 expression, accompanied by increased Gpx4 expression. Furthermore, p53 knockdown suppresses Met-mediated anti-ferroptosis effects in PA-treated VSMCs, which may be related to the downregulation of SLC7A11 expression. In addition, supplementation of VSMCs with Met enhances the antioxidative capacity of VSMCs through Nrf2 signalling activation. Collectively, targeting POSTN in VSMCs may provide a new strategy for vascular calcification prevention or treatment.

Kcnk3 dysfunction exaggerates the development of pulmonary hypertension induced by left ventricular pressure overload.

Pulmonary hypertension (PH) is a common complication of left heart disease (LHD, Group 2 PH) leading to right ventricular (RV) failure and death. Several loss-of-function (LOF) mutations in KCNK3 were identified in pulmonary arterial hypertension (PAH, Group 1 PH). Additionally, we found that KCNK3 dysfunction is a hallmark of PAH at pulmonary vascular and RV levels. However, the role of KCNK3 in the pathobiology of PH due to LHD is unknown. We evaluated the role of KCNK3 on PH induced by ascending aortic constriction (AAC), in WT and Kcnk3-LOF-mutated rats, by echocardiography, RV catheterization, histology analyses, and molecular biology experiments. We found that Kcnk3-LOF-mutation had no consequence on the development of left ventricular (LV) compensated concentric hypertrophy in AAC, while left atrial emptying fraction was impaired in AAC-Kcnk3-mutated rats. AAC-animals (WT and Kcnk3-mutated rats) developed PH secondary to AAC and Kcnk3-mutated rats developed more severe PH than WT. AAC-Kcnk3-mutated rats developed RV and LV fibrosis in association with an increase of Col1a1 mRNA in right ventricle and left ventricle. AAC-Kcnk3-mutated rats developed severe pulmonary vascular (pulmonary artery as well as pulmonary veins) remodelling with intense peri-vascular and peri-bronchial inflammation, perivascular oedema, alveolar wall thickening, and exaggerated lung vascular cell proliferation compared to AAC-WT-rats. Finally, in lung, right ventricle, left ventricle, and left atrium of AAC-Kcnk3-mutated rats, we found a strong increased expression of Il-6 and periostin expression and a reduction of lung Ctnnd1 mRNA (coding for p120 catenin), contributing to the exaggerated pulmonary and heart remodelling and pulmonary vascular oedema in AAC-Kcnk3-mutated rats. Our results indicate that Kcnk3-LOF is a key event in the pathobiology of PH due to AAC, suggesting that Kcnk3 channel dysfunction could play a potential key role in the development of PH due to LHD.

Development of an engineered peptide antagonist against periostin to overcome doxorubicin resistance in breast cancer

BackgroundChemoresistance is one of the main problems in treatment of cancer. Periostin (PN) is a stromal protein which is mostly secreted from cancer associated fibroblasts in the tumor microenvironment and can promote cancer progression including cell survival, metastasis, and chemoresistance. The main objective of this study was to develop an anti-PN peptide from the bacteriophage library to overcome PN effects in breast cancer (BCA) cells.MethodsA twelve amino acids bacteriophage display library was used for biopanning against the PN active site. A selected clone was sequenced and analyzed for peptide primary structure. A peptide was synthesized and tested for the binding affinity to PN. PN effects including a proliferation, migration and a drug sensitivity test were performed using PN overexpression BCA cells or PN treatment and inhibited by an anti-PN peptide. An intracellular signaling mechanism of inhibition was studied by western blot analysis. Lastly, PN expressions in BCA patients were analyzed along with clinical data.ResultsThe results showed that a candidate anti-PN peptide was synthesized and showed affinity binding to PN. PN could increase proliferation and migration of BCA cells and these effects could be inhibited by an anti-PN peptide. There was significant resistance to doxorubicin in PN-overexpressed BCA cells and this effect could be reversed by an anti-PN peptide in associations with phosphorylation of AKT and expression of survivin. In BCA patients, serum PN showed a correlation with tissue PN expression but there was no significant correlation with clinical data.ConclusionsThis finding supports that anti-PN peptide is expected to be used in the development of peptide therapy to reduce PN-induced chemoresistance in BCA.

Chemokine C-C motif ligand 8 in periodontal ligament during orthodontic tooth movement

ObjectivesTo investigate the roles of chemokine (C-C motif) ligand 8 (CCL8) in periodontal ligament during orthodontic tooth movement (OTM). MethodsBioinformatics analyzed 100 genes in human periodontal ligament cells that were most upregulated after 48 hours of mechanical stress, and these genes were classified through GO and KEGG databases. Nickel-titanium closed-coil springs were placed between right first molar and incisors to produce 20 cN of orthodontic force in eight-week-old male SD rats for 1 and 2 days, followed by immunohistochemical staining of CCL8. Human periodontal ligament fibroblasts (hPDLFs) were stimulated by 14% cyclic tension force (Flexcell FX-5000 T Tension System) or hypoxia conditions to mimic OTM for 1 and 2 days, then the resulting CCL8 were examined through ELISA. Scratching assay was performed by treating hPDLFs with different concentrations of CCL8 (1 ng/ml, 10 ng/ml, 100 ng/ml). The migration, proliferation, and adhesion abilities of 100 ng/ml CCL8-treated hPDLFs were also examined. qRT-PCR and western blot detected matrix metalloproteinase 3, periostin, and osteoprotegrin expressions of hPDLFs under 100 ng/ml CCL8. ResultsBioinformatic analysis demonstrated that CCL8 was upregulated after applying mechanical stress for 48 hours. CCL8 secretion showed upregulation after 24 hours of OTM applicationsin vivo and in vitro. CCL8-treated hPDLFs showed significant positive effects on cell proliferation and matrix metalloproteinase 3. It also inhibited periostin and osteoprotegrin expressions. ConclusionsCCL8 was upregulated in periodontal ligament during initial stage of OTM. Although CCL8 in human periodontal ligaments showed no significant effects on cell migration ability, it did enhance cell proliferation and osteoclastogenesis.

Mouse models of atopic dermatitis: a critical reappraisal.

Mouse models for atopic dermatitis (AD) are an indispensable preclinical research tool for testing new candidate AD therapeutics and for interrogating AD pathobiology in vivo. In this Viewpoint, we delineate why, unfortunately, none of the currently available so-called "AD" mouse models satisfactorily reflect the clinical complexity of human AD, but imitate more "allergic" or "irriant" contact dermatitis conditions. This limits the predictive value of AD models for clinical outcomes of new tested candidate AD therapeutics and the instructiveness of mouse models for human AD pathophysiology research. Here, we propose to initiate a rational debate on the minimal criteria that a mouse model should meet in order to be considered relevant for human AD. We suggest that valid AD models should at least meet the following criteria: (a) an AD-like epidermal barrier defect with reduced filaggrin expression along with hyperproliferation, hyperplasia; (b) increased epidermal expression of thymic stromal lymphopoietin (TSLP), periostin and/or chemokines such as TARC (CCL17); (c) a characteristic dermal immune cell infiltrate with overexpression of some key cytokines such as IL-4, IL-13, IL-31 and IL-33; (d) distinctive "neurodermatitis" features (sensory skin hyperinnervation, defective beta-adrenergic signalling, neurogenic skin inflammation and triggering or aggravation of AD-like skin lesions by perceived stress); and (e) response of experimentally induced skin lesions to standard AD therapy. Finally, we delineate why humanized AD mouse models (human skin xenotransplants on SCID mice) offer a particularly promising preclinical research alternative to the currently available "AD" mouse models.

POSTN promotes proliferation and epithelial-mesenchymal transition in renal cell carcinoma through ILK/AKT/mTOR pathway.

Periostin (POSTN) is an extracellular matrix (ECM) protein, involved in various diseases. This research focused on the detailed mechanisms study of periostin (POSTN) overexpression in renal cell carcinoma (RCC) invasion and migration. Western blot and RT-PCR were performed to explore POSTN expression in various RCC cells. Cells were transfected with siRNAs or lentivirus to regulate the expression of POSTN. The effects of POSTN on cell viability, apoptosis, migration, invasion and epithelial-to-mesenchymal transition (EMT) of RCC cells were determined by CCK-8, flow cytometry, migration and invasion assay and Western blot analysis. POSTN expression was significantly enhanced in RCC cells compared with renal tubular epithelial cells. In vitro experiments showed that POSTN knockdown could dramatically inhibit RCC cell proliferation, migration and invasion, while overexpression of POSTN could promote these biological behaviors. We further demonstrated that POSTN knockdown suppressed epithelial-mesenchymal transition (EMT), which was mediated via upregulation of E-cadherin and downregulation of N-cadherin and vimentin, through IKL/AKT/mTOR pathway. In contrast, overexpression of POSTN could promote EMT in RCC cells via the activation of IKL /AKT/mTOR pathway. Next, we demonstrated that higher POSTN expression promoted angiogenesis in vivo in an RCC xenograft tumor via activating IKL /AKT/mTOR pathway. Our study showed that POSTN could promote EMT through ILK/AKT/mTOR pathway and might be an alternative therapeutic strategy for RCC treatment.

Expression of “periostin” in leukoplakia and oral submucous fibrosis: An immunohistochemical study

Aim: Periostin is an extracellular matrix protein secreted in the tumor microenvironment. It is frequently overexpressed in various human cancers, including oral squamous cell carcinoma (OSCC). The role of periostin in carcinogenesis of OSCC is yet to be established. The present study was aimed at determining “Periostin” expression in leukoplakia and the oral submucous fibrosis (OSMF) to relate with its role in malignant transformation of these disorders to OSCC. Materials and Methods: A total of 63 cases were categorized into three groups, namely Group I: OSCC (n = 21), Group II: Leukoplakia (n = 21), and Group III: Oral submucous fibrosis (n = 21); these groups were subjected to immunohistochemical analysis by using the periostin antibody. Kruskal–Wallis test and Mann–Whitney U tests were used to assess the significant differences in the periostin expression between groups (SPSS software version 23.0). Results: Results indicated that there was significant difference in the periostin levels between the groups (P < 0.005), indicating a fairly strong relationship between periostin expression among groups. A significant difference between the different grades of leukoplakia was observed, whereas there were no significant differences between subgroups of the OSCC and OSMF. Conclusion: The present study justifies periostin as a key stromal element in tumor progression of OSCC. The potential role of periostin in malignant transformation of oral potentially malignant disorders (OPMD) has been suggested.

An exosomal-carried short periostin isoform induces cardiomyocyte proliferation.

Although a small number of cardiomyocytes may reenter the cell cycle after injury, the adult mammalian heart is incapable of a robust cardiomyocyte proliferation. Periostin, a secreted extracellular matrix protein, has been implicated as a regulator of cardiomyocyte proliferation; however, this role remains controversial. Alternative splicing of the human periostin gene results in 6 isoforms lacking sequences between exons 17 and 21, in addition to full-length periostin. We previously showed that exosomes (Exo) secreted by human cardiac explant-derived progenitor cells (CPC) carried periostin. Here, we aimed to investigate their cell cycle activity.Methods: CPC were derived as the cellular outgrowth of ex vivo cultured cardiac atrial explants. Exo were purified from CPC conditioned medium using size exclusion chromatography. Exosomal periostin was analyzed by Western blotting using a pair of antibodies (one raised against aa 537-836, and one raised against amino acids mapping at exon 17 of human periostin), by ELISA, and by cryo-EM with immune-gold labeling. Cell cycle activity was assessed in neonatal rat cardiomyocytes, in human induced pluripotent stem cell (iPS)-derived cardiomyocytes, and in adult rat cardiomyocytes after myocardial infarction. The role of periostin in cell cycle activity was investigated by transfecting donor CPC with a siRNA against this protein.Results: Periostin expression in CPC-secreted exosomes was detected using the antibody raised against aa 537-836 of the human protein, but not with the exon 17-specific antibody, consistent with an isoform lacking exon 17. Periostin was visualized on vesicle surfaces by cryo-EM and immune-gold labeling. CPC-derived exosomes induced cell proliferation in neonatal rat cardiomyocytes both in vitro and in vivo, in human iPS-derived cardiomyocytes, and in adult rat cardiomyocytes after myocardial infarction. Exo promoted phosphorylation of focal adhesion kinase (FAK), actin polymerization, and nuclear translocation of Yes-associated protein (YAP) in cardiomyocytes. Knocking down of periostin or YAP, or blocking FAK phosphorylation with PF-573228 nullified Exo-induced proliferation. A truncated human periostin peptide (aa 22-669), but not recombinant human full-length periostin, mimicked the pro-proliferative activity of exosomes.Conclusions: Our results show, for the first time, that CPC-secreted exosomes promote cardiomyocyte cell cycle-reentry via a short periostin isoform expressed on their surfaces, whereas recombinant full-length periostin does not. These findings highlight isoform-specific roles of periostin in cardiomyocyte proliferation.

Periostin is Oncogenic and Promotes EMT in Gliomas via Inhibition of RIN1-Mediated Endocytosis of EGFR

Background: About 1/3 of all brain tumors are gliomas. Previous glioma studies have reported increased expression of Periostin (POSTN) in these cancerous tissues, but its role and mechanism in gliomas development remain unknown. Methods: Nanoscale liquid chromatography coupled to tandem mass spectrometry (Nano LC-MS/MS) and RNA sequencing were used to identify differential protein and mRNA expression in glioma samples from the clinic. Quantitative real-time PCR (qRT-PCR) was used to detect the expression of POSTN in tissues and cells. The effects of POSTN on glioma cell migration and invasion were examined using wound-healing, transwell, and three-dimensional spheroid assays in vitro, and a xenograft nude-mouse model in vivo. POSTN’s effect on the stability, endocytosis, and degradation of EGFR was examined by immunoblotting and immunofluorescence. Truncated mutation analysis was performed to interrogate any direct interactions between POSTN and EGFR. Immunohistochemical staining was carried out to confirm the clinical significance of POSTN. Findings: In this study, POSTN was upregulated in gliomas and was associated with poor prognosis. POSTN depletion inhibited glioma epithelial-to-mesenchymal transition (EMT) both in vitro and in vivo. Furthermore, POSTN downregulation dampened EGFR signaling via promotion of EGFR endocytosis and degradation. In addition, POSTN was found to bind to EGFR and RIN1, inhibiting EGFR endocytosis and degradation, thus promoting the activation of the PI3K-AKT signaling pathway. Interpretation: These findings indicate the POSTN/EGFR/RIN1 axis inhibits EGFR endocytosis and degradation. Targeting POSTN/EGFR/RIN1 interactions may prove beneficial the treatment of gliomas. Funding Information: This work was supported by grants from National Natural Science Foundation of China (81772682), and the Priority Academic Program Development of Jiangsu Higher Education Institutions (PAPD). Declaration of Interests: The authors have declared that no competing interest exists. Ethics Approval Statement: Our study protocol was approved by the Research Ethics Committee of Nanjing Medical University (Nanjing, Jiangsu, China).

Knockdown of POSTN Inhibits Osteogenic Differentiation of Mesenchymal Stem Cells From Patients With Steroid-Induced Osteonecrosis.

Steroid-induced osteonecrosis of femoral head (SONFH) is a common and serious complication caused by long-term and/or excessive use of glucocorticoids (GCs). The decreased activity and abnormal differentiation of bone marrow mesenchymal stem cells (BMSCs) are considered to be one of the major reasons for the onset and progression of this disease. Periostin (POSTN) is a matricellular protein which plays an important role in regulating osteoblast function and bone formation. Sclerostin (SOST) is a secreted antagonist of Wnt signaling that is mainly expressed in osteocytes to inhibit bone formation. However, the exact role of POSTN and SOST in SONFH has not been reported yet. Therefore, we detected the differential expression of POSTN and SOST in BMSCs of SONFH Group patients, and Control Group was patients with traumatic ONFH (TONFH) and developmental dysplasia of the hip (DDH). Furthermore, we used lentiviral transfection to knockdown POSTN expression in BMSCs of patients with SONFH to study the effect of POSTN knockdown on the SOST expression and osteogenic differentiation of BMSCs. The results indicated that the endogenous expression of POSTN and SOST in BMSCs of SONFH Group was upregulated, compared with Control Group. POSTN was upregulated gradually while SOST was downregulated gradually at days 0, 3, and 7 of osteogenic differentiation of BMSCs in Control Group. Contrarily, POSTN was gradually downregulated while SOST was gradually upregulated during osteogenic differentiation of BMSCs in SONFH Group. This could be due to increased expression of SOST in BMSCs, which was caused by excessive GCs. In turn, the increased expression of POSTN in BMSCs may play a role in antagonizing the continuous rising of SOST during the osteogenic differentiation of BMSCs in patients with SONFH. POSTN knockdown significantly attenuated osteo-specific gene expression, alkaline phosphatase activity, and calcium nodule formation in vitro; thus inhibiting the osteogenic differentiation of BMSCs in patients with SONFH. Besides, POSTN knockdown upregulated SOST expression, increased GSK-3β activity, and downregulated β-catenin. These findings suggest that POSTN have an essential role in regulating the expression of SOST and osteogenic differentiation of BMSCs in patients with SONFH, and POSTN knockdown suppresses osteogenic differentiation by upregulating SOST and partially inactivating Wnt/β-catenin signaling pathway. Therefore, targeting POSTN and SOST may serve as a promising therapeutic target for the prevention and treatment of SONFH.

Preservation of the viability and gene expression of human periodontal ligament cells by Thai propolis extract.

Success of tooth replantation depends on the quality and quantity of periodontal ligament (PDL) cells. The aims of this study were to evaluate Thai propolis extract as a storage medium for maintaining PDL cell viability and preserving gene expressions in PDL tissues. PDL cells from human premolars were tested for cytotoxicity of the extract by PrestoBlue assay to determine a non-toxic concentration. Subsequently, 96 freshly extracted premolars were allocated into different treatment groups. Control groups were freshly extracted premolars or they had been stored dry for 12hours. Experimental avulsed teeth were created by leaving them air-dried for 30minutes immediately after extraction, then they were immersed in Thai propolis extract, HBSS or milk for 3, 6 and 12hours. After tooth storage, the remaining PDL cells were determined for their cell viability. RNA isolated from PDL tissues of three premolars treated similarly was analysed for periostin and S100A4 expressions using RT-qPCR. Thai propolis extract at 0.625mgmL-1 promoted the greatest PDL cell viability. Tooth storage in 0.625mgmL-1 Thai propolis extract, HBSS or milk showed no difference in maintaining cell viability. Periostin mRNA level was preserved by Thai propolis extract. Expression of S100A4 mRNA in PDL tissues stored in all tested media was dampened. PDL cells from mock avulsed teeth stored in 0.625mgmL-1 Thai propolis extract for 3, 6 and 12hours remained viable and the expression of periostin was preserved. This study suggests this extract as an alternative for a tooth storage medium for up to 12hours. However, transporting an avulsed tooth in a storage medium for extended extra-oral time might affect the PDL cell phenotypes.

Periostin as a key molecule defining desmoplastic environment in colorectal cancer.

Categorizing desmoplastic reaction (DR) based on the histological findings of cancer-associated fibroblasts is shown to be a promising novel method to predict prognosis of patients with colorectal cancer (CRC). Periostin (POSTN) in cancer-associated stroma is reportedly associated with poor clinical outcomes. Immunohistochemical staining with an anti-POSTN antibody was performed in 73 patients with pStage III CRC (cohort 1). In addition, to evaluate mRNA and protein expression levels of POSTN, we analyzed paired normal and invasive cancer frozen specimens by quantitative real-time polymerase chain reaction and western blot analysis in 41 patients (cohort 2). In cohort 1, according to the DR categorization, 18, 22, and 33 patients were classified as immature, intermediate, and mature, respectively. High immunoreactivity of POSTN was observed 100%, 68.2%, and 27.3%, respectively (p < 0.0001). The 5-year relapse-free survival rates were 56.8% and 82.7% in high and low POSTN expression subgroups, respectively (p = 0.015). In cohort 2, the POSTN mRNA and protein levels were significantly higher in the immature stroma as compared to the stroma characterized as other DR patterns. POSTN expression was closely associated with DR categorization. POSTN may be a key molecule that contributes to the malignant potential of CRC.

Enhanced 15-Lipoxygenase 1 Production is Related to Periostin Expression and Eosinophil Recruitment in Eosinophilic Chronic Rhinosinusitis.

Background: The pathological features of chronic rhinosinusitis (CRS) with nasal polyps (CRSwNP) tissues include an eosinophilic infiltration pattern (eosinophilic CRS (ECRS)) or a less eosinophilic pattern (non-ECRS). Recently, it has been suggested that 15-lipoxygenase 1 (15-LOX-1) may have significant roles in allergic disease; however, the significance of 15-LOX-1 in CRS is not well understood. The objective of this study was to demonstrate the expression of 15-LOX-1 in CRS. Methods: The mRNA expression levels of 15-LOX-1 and periostin in nasal tissues were measured by quantitative real-time polymerase chain reaction. We also performed an immunofluorescence study of nasal tissues. Cells of the Eol-1 eosinophilic leukemic cell line were stimulated with interleukin-33 to test the induction of 15-LOX-1. Results: The expression level of 15-LOX-1 mRNA in nasal polyps (NPs) was significantly higher in ECRS patients than in non-ECRS patients. The immunofluorescence study revealed that both airway epithelial cells and eosinophils in NPs expressed 15-LOX-1. A significant correlation was seen between the number of eosinophils and the mRNA expression levels of 15-LOX-1 and periostin in nasal polyps. Moreover, interleukin-33 enhanced 15-LOX-1 expression in Eol-1 cells. Conclusions: 15-LOX-1 was shown to be a significant molecule that facilitates eosinophilic inflammation in ECRS.

Periostin+ cancer-associated fibroblasts promote lymph node metastasis by impairing the lymphatic endothelial barriers in cervical squamous cell carcinoma.

Lymph node metastasis (LNM), a critical prognostic determinant in cancer patients, is critically influenced by the presence of numerous heterogeneous cancer-associated fibroblasts (CAFs) in the tumor microenvironment. However, the phenotypes and characteristics of the various pro-metastatic CAF subsets in cervical squamous cell carcinoma (CSCC) remain unknown. Here, we describe a CAF subpopulation with elevated periostin expression (periostin+ CAFs), located in the primary tumor sites and metastatic lymph nodes, that positively correlated with LNM and poor survival in CSCC patients. Mechanistically, periostin+ CAFs impaired lymphatic endothelial barriers by activating the integrin-FAK/Src-VE-cadherin signaling pathway in lymphatic endothelial cells and consequently enhanced metastatic dissemination. In contrast, inhibition of the FAK/Src signaling pathway alleviated periostin-induced lymphatic endothelial barrier dysfunction and its related effects. Notably, periostin- CAFs were incapable of impairing endothelial barrier integrity, which may explain the occurrence of CAF-enriched cases without LNM. In conclusion, we identified a specific periostin+ CAF subset that promotes LNM in CSCC, mainly by impairing the lymphatic endothelial barriers, thus providing the basis for potential stromal fibroblast-targeted interventions that block CAF-dependent metastasis.

Recruited CD68+CD206+ macrophages orchestrate graft immune tolerance to prompt xenogeneic-dentin matrix-based tooth root regeneration

Successful regenerative medicine strategies of xenogeneic extracellular matrix need a synergistic balance among inflammation, fibrosis, and remodeling process. Adaptive macrophage subsets have been identified to modulate inflammation and orchestrate the repair of neighboring parenchymal tissues. This study fabricated PPARγ-primed CD68+CD206+ M2 phenotype (M2γ), and firstly verified their anti-inflammatory and tissue-regenerating roles in xenogeneic bioengineered organ regeneration. Our results showed that Th1-type CD3+CD8+ T cell response to xenogeneic-dentin matrix-based bioengineered root complex (xeno-complex) was significantly inhibited by M2γ macrophage in vitro. PPARγ activation also timely recruited CD68+CD206+ tissue macrophage polarization to xeno-complex in vivo. These subsets alleviated proinflammatory cytokines (TNF-α, IFN-γ) at the inflammation site and decreased CD3+CD8+ T lymphocytes in the periphery system. When translated to an orthotopic nonhuman primate model, PPARγ-primed M2 macrophages immunosuppressed IL-1β, IL-6, TNF-α, MMPs to enable xeno-complex to effectively escape immune-mediated rejection and initiate graft-host synergistic integrity. These collective activities promoted the differentiation of odontoblast-like and periodontal-like cells to guide pulp-dentin and cementum-PDLs-bone regeneration and rescued partially injured odontogenesis such as DSPP and periostin expression. Finally, the regenerated root showed structure-biomechanical and functional equivalency to the native tooth. The timely conversion of M1-to-M2 macrophage mainly orchestrated odontogenesis, fibrogenesis, and osteogenesis, which represents a potential modulator for intact parenchymal-stromal tissue regeneration of targeted organs.