A bi-layered asymmetric membrane loaded with demineralized dentin matrix for guided bone regeneration.

OBJECTIVES: Guided bone regeneration (GBR) is a well-established method for repairing hard tissue deficiency in reconstructive dentistry. The aim of this study was to investigate the barrier function, osteogenic activity and immunomodulatory ability of a novel bi-layered asymmetric membrane loaded with demineralized dentin matrix (DDM). METHODS: DDM particles were harvested from healthy, caries-free permanent teeth. Electrospinning technique was utilized to prepare bi-layered DDM-loaded poly(lactic-co-glycolic acid) (PLGA)/poly(lactic acid) (PLA) membranes (abbreviated as DPP bilayer membranes). We analyzed the membranes' surface properties, cytocompatibility and barrier function, and evaluated their osteogenic activity in vitro. In addition, its effects on the osteogenic immune microenvironment were also investigated. RESULTS: Synthetic DPP bilayer membranes presented suitable surface characteristics and satisfactory cytocompatibility. Transwell assays showed significant fewer migrated cells by the DPP bilayer membranes compared with blank control, with or without in vitro degradation (all P < 0.001). In vitro experiments indicated that our product elevated messenger ribonucleic acid (mRNA) expression levels of osteogenic genes alkaline phosphatase (ALP), osteopontin (OPN), osteocalcin (OCN) and runt-related transcription factor 2 (Runx2). Among all groups, 20% DPP bilayer membrane displayed highest ALP activity (P < 0.001). Furthermore, DPP bilayer membranes enhanced the mRNA expression of M2 macrophage markers and increased the proportion of CD206+ M2 macrophages by 100% (20% DPP: P < 0.001; 30% DPP: P < 0.001; 40% DPP: P < 0.05), thus exerting an inflammation suppressive effect. CONCLUSIONS: DPP bilayer membranes exhibited notable biological safety and osteogenic activity in vitro, and have potential as a prospective candidate for GBR approach in the future.

CC chemokine receptor 7 promotes macrophage recruitment and induces M2-polarization through CC chemokine ligand 19&21 in oral squamous cell carcinoma.

PURPOSE: This study aimed to investigate the impact of CC chemokine receptor 7 (CCR7) on the recruitment and polarization of tumor-associated macrophages (TAMs) in oral squamous cell carcinoma (OSCC). METHODS: We analyzed CCR7 expression pattern, clinicopathological significance, and its association with M2 macrophage infiltration in OSCC by bioinformatic methods. Small interfering RNA (siRNA) was utilized to silence CCR7 in OSCC cells. Conditioned media (CM) was harvested from transfected OSCC cells to establish a co-culture model of THP-1 derived macrophages and OSCC cells. Transwell assay and cell adhesion assay were performed to examine the effect of CCR7 on macrophages recruitment and adhesion. Cytoskeleton was labelled by phalloidin to observe macrophage morphological changes. Moreover, phenotypic alteration of macrophages was measured using quantitative real-time PCR (qRT-PCR), flow cytometry, and immunofluorescence (IF) staining. Ultimately, recombinant human CCL19 and CCL21 were added into the medium of THP-1 derived macrophages to explore their effects on polarization in vitro. RESULTS: In OSCC patients, the overexpression of CCR7 positively correlated with lymph node metastasis and M2 macrophage infiltration. Macrophage not only exhibited enhanced migration, invasion and adhesion abilities, but also appeared more spindle and branched in vitro when treated with CM from OSCC cells. However, these phenomena were abrogated with knockdown of CCR7. We also discovered that inhibition of CCR7 in OSCC cells suppressed TAMs polarization to an M2 phenotype. In addition, recombinant human CCL19 and CCL21 promoted macrophage M2-polarization in vitro. CONCLUSION: CCR7 in OSCC cells promoted recruitment and M2-polarization of THP-1 derived macrophages in vitro by regulating production of CCL19 and CCL21.

The Overexpression of Fibronectin 1 Promotes Cancer Progression and Associated with M2 Macrophages Polarization in Head and Neck Squamous Cell Carcinoma Patients.

Purpose: This study aimed to investigate the biological roles of fibronectin 1 (FN1) in head and neck squamous cell carcinoma (HNSCC) and its effects on macrophage M2 polarization. Methods: We analyzed FN1 expression pattern and examined its clinical relevance in HNSCC progression by bioinformatic analysis. Small interfering RNA (siRNA) was utilized to silence FN1 in HNSCC cells. Cell counting kit-8 (CCK-8) assay, colony formation assay, Transwell assay and wound healing assay were performed to reveal the effect of FN1 on malignant behaviors of HNSCC cells. Moreover, a co-culture model of macrophages and HNSCC cells was established to investigate whether FN1 induce macrophage M2 polarization. Finally, we used bioinformatic methods to explore the possible FN1-related pathways in HNSCC. Results: FN1 is significantly overexpressed in HNSCC patients and has been obviously correlated with higher pathological stage and poor prognosis. Downregulation of FN1 suppressed the proliferation, migration and invasion of HNSCC cells, and inhibited macrophage M2 polarization in vitro. In addition, "PI3K-Akt" and "MAPK" signaling pathways may be involved in the malignant process of FN1 in HNSCC. Conclusion: The overexpression of FN1 promotes HNSCC progression and induces macrophages M2 polarization. FN1 may serve as a promising prognostic biomarker and therapeutic target in HNSCC.