Three-dimensional cultures of gingival fibroblasts on fibrin-based scaffolds for gingival augmentation: A proof-of-concept study.

OBJECTIVE: Gingival tissue regeneration is associated with several challenges. Tissue engineering regenerates the different components of the tissues, providing three major elements: living cells, appropriate scaffolds, and tissue-inducing substances. This study aimed to regenerate the gingival connective tissue in vitro, using human gingival fibroblasts cultured in three-dimensional fibrin gel scaffolds. DESIGN: Human gingival fibroblasts were seeded in a novel three-dimensional fibrin gel scaffold and maintained in two media types: platelet lysate media (control) and collagen-stimulating media (test). Cellular viability and proliferation were assessed, and the production of collagen and other extracellular matrix components in these constructs was investigated and compared. RESULTS: Human gingival fibroblasts cultured in three-dimensional cultures were metabolically active and proliferated in both media. Furthermore, histologic sections, scanning electron microscopy, and quantitative polymerase chain reaction confirmed the production of higher levels of collagen and other extracellular matrix fibers in three-dimensional constructs cultured in collagen-stimulating media. CONCLUSIONS: Culturing human gingival fibroblasts in a novel three-dimensional fibrin gel scaffold containing collagen-stimulating media resulted in a tissue-equivalent construct that mimics human gingival connective tissue. The impact of these results should be considered for further investigations, which may help to develop a compatible scaffold for gingival soft tissue regeneration and treatment of mucogingival deformities.

Phytochemical Analysis and Anticancer Properties of Drimia maritima Bulb Extracts on Colorectal Cancer Cells.

Cancer is a worldwide health problem and is the second leading cause of death after heart disease. Due to the high cost and severe side effects associated with chemotherapy treatments, natural products with anticancer therapeutic potential may play a promising role in anticancer therapy. The purpose of this study was to investigate the cytotoxic and apoptotic characteristics of the aqueous Drimia maritima bulb extract on Caco-2 and COLO-205 colorectal cancer cells. In order to reach such a purpose, the chemical composition was examined using the GC-MS method, and the selective antiproliferative effect was determined in colon cancer cell lines in normal gingival fibroblasts. The intracellular ROS, mitochondrial membrane potential, and gene expression changes in selected genes (CASP8, TNF-α, and IL-6 genes) were assessed to determine the molecular mechanism of the antitumor effect of the extract. GC-MS results revealed the presence of fifty-seven compounds, and Proscillaridin A was the predominant secondary metabolite in the extract. The IC50 of D. maritima bulb extract on Caco-2, COLO-205, and the normal human gingival fibroblasts were obtained at 0.9 µg/mL, 2.3 µg/mL, and 13.1 µg/mL, respectively. The apoptotic effect assay indicated that the bulb extract induced apoptosis in both colon cancer cell lines. D. maritima bulb extract was only able to induce statistically significant ROS levels in COLO-205 cells in a dose-dependent manner. The mitochondrial membrane potential (MMP) revealed a significant decrease in the MMP of Caco-2 and COLO-205 to various concentrations of the bulb extract. At the molecular level, RT-qPCR was used to assess gene expression of CASP8, TNF-α, and IL-6 genes in Caco-2 and COLO-205 cancer cells. The results showed that the expression of pro-inflammatory genes TNF-α and IL-6 were upregulated. The apoptotic initiator gene CASP8 was also upregulated in the Caco-2 cell line and did not reach significance in COLO-205 cells. These results lead to the conclusion that D. maritima extract induced cell death in both cell lines and may have the potential to be used in CRC therapy in the future.

The Immunomodulatory and Regenerative Effect of Biodentine™ on Human THP-1 Cells and Dental Pulp Stem Cells: In Vitro Study.

Background: Pulp tissue affected by deep caries and trauma can be protected by vital pulp therapies in which pulp regeneration success depends on the degree of pulp inflammation and the presence of regenerative signals. Reparative dentinogenesis requires dental pulp stem cell (DPSC) activity which can be stimulated by many bioactive molecules to repair the dentine, mediating a balance between the inflammatory response and the reparative events. Therefore, this study was performed in order to investigate the immune-inflammatory effect of Biodentine capping material on DPSCs and macrophages. Method: THP-1, a human monocytic cell line, was differentiated to macrophages, and flow cytometry was used to analyze the expressions of specific macrophage markers. LPS-mediated infection was created for macrophages and DPSCs followed by treatment with Biodentine. CBA array was used to investigate the cytokine secretion followed by qPCR. Migration potential of treated DPSCs was also determined. Results: Our results showed that THP-1 cell line was successfully differentiated into macrophages as shown by surface marker expression. CBA array and qPCR results showed that Biodentine-treated DPSCs and macrophages upregulated anti-inflammatory cytokines and downregulated proinflammatory cytokines. Also, Biodentine enhances the migration potential of treated DPSCs. Conclusion: Biodentine capping material mediated the polarization of M1 to M2 macrophages suggestive of tissue repair properties of macrophages and enhanced the anti-inflammatory cytokines of DPSCs responsible for dentine-pulp regeneration.

Generation of an induced pluripotent stem cell (iPSC) line (JUCTCi017-A) from a patient with limb-girdle muscular dystrophy (LGMD) due to a homozygous p.Lue287Ser fs14* mutation in the SGCB gene.

Limb-girdle muscular dystrophies (LGMDs) are a large group of heterogenous genetic diseases characterized by muscle weakness. In this study, an induced pluripotent stem cell (iPSC) line was generated from LGMD patient's skin dermal fibroblasts, carrying a homozygous mutation in the Sarcoglycan Beta (SGCB) gene; chr4:52890221, c. 859 delC, p.Lue 287Ser fs14*. The reprogramming process was carried out using Sendai viruses encoding for Yamanaka factors. The resulting iPSCs showed normal morphology and karyotype, expressed pluripotency markers, demonstrated the potential to differentiate in vitro into three germ layers and retained the disease-causing SGCB mutation. This iPSC line represents an ideal source of cells for the investigation of LGMD disease mechanisms.