Photodynamic Therapy has no Adverse Effects In Vitro on Human Gingival Fibroblasts and Osteoblasts.

BACKGROUND: Invasion of periodontal pockets with persistent microorganisms and subsequent development of a biofilm are the main cause of periodontal infections. In severe cases, additional use of antibiotics to the standard therapy of scaling and root planing (SRP), is necessary, but the use of antibiotics may lead to resistance. As an alternative, the combination of SRP and adjunct photodynamic therapy (PDT) is applied in the treatment of periodontal infections to improve periodontal therapy. The aim of this study was to determine possible side effects of PDT on human gingival fibroblasts (GF) and human osteoblasts (OB). METHODS: GF and OB were either untreated or treated with methylene blue (MB) only, with MB and subsequently irradiation with a soft laser (PDT) or irradiated with a soft laser only. All cells were analyzed for viability using the MTT test, migration capacity using Boyden chambers, and the scratch wound assay. RESULTS: Viability and migration capacity of GF and OB were not affected by PDT (for GF: p = 0.48, for OB: p = 0.08 compared to control group) whereas soft laser irradiation only improved cell viability and migration and MB treatment only reduced cell viability and migration. After 72 hours of incubation of both GF and OB, the gaps were almost closed. CONCLUSIONS: In vitro, PDT did not affect viability and migration capacity of GF and OB whereas soft laser treatment only had a positive effect on GF and OB. Therefore, PDT seems to be a safe method in the treatment of periodontal infections without significant side effects.

Three-dimensional histochemistry and imaging of human gingiva.

In the present study, 3D histochemistry and imaging methodology is described for human gingiva to analyze its vascular network. Fifteen human gingiva samples without signs of inflammation were cleared using a mixture of 2-parts benzyl benzoate and 1-part benzyl alcohol (BABB), after being immunofluorescently stained for CD31, marker of endothelial cells to visualize blood vessels in combination with fluorescent DNA dyes. Samples were imaged in 3D with the use of confocal microscopy and light-sheet microscopy and image processing. BABB clearing caused limited tissue shrinkage 13 ± 7% as surface area and 24 ± 1% as volume. Fluorescence remained intact in BABB-cleared gingiva samples and light-sheet microscopy was an excellent tool to image gingivae whereas confocal microscopy was not. Histochemistry on cryostat sections of gingiva samples after 3D imaging validated structures visualized in 3D. Three-dimensional images showed the vascular network in the stroma of gingiva with one capillary loop in each stromal papilla invading into the epithelium. The capillary loops were tortuous with structural irregularities that were not apparent in 2D images. It is concluded that 3D histochemistry and imaging methodology described here is a promising novel approach to study structural aspects of human gingiva in health and disease.

The angiogenic switch leads to a metabolic shift in human glioblastoma.

Background: Invasion and angiogenesis are major hallmarks of glioblastoma (GBM) growth. While invasive tumor cells grow adjacent to blood vessels in normal brain tissue, tumor cells within neovascularized regions exhibit hypoxic stress and promote angiogenesis. The distinct microenvironments likely differentially affect metabolic processes within the tumor cells. Methods: In the present study, we analyzed gene expression and metabolic changes in a human GBM xenograft model that displayed invasive and angiogenic phenotypes. In addition, we used glioma patient biopsies to confirm the results from the xenograft model. Results: We demonstrate that the angiogenic switch in our xenograft model is linked to a proneural-to-mesenchymal transition that is associated with upregulation of the transcription factors BHLHE40, CEBPB, and STAT3. Metabolic analyses revealed that angiogenic xenografts employed higher rates of glycolysis compared with invasive xenografts. Likewise, patient biopsies exhibited higher expression of the glycolytic enzyme lactate dehydrogenase A and glucose transporter 1 in hypoxic areas compared with the invasive edge and lower-grade tumors. Analysis of the mitochondrial respiratory chain showed reduction of complex I in angiogenic xenografts and hypoxic regions of GBM samples compared with invasive xenografts, nonhypoxic GBM regions, and lower-grade tumors. In vitro hypoxia experiments additionally revealed metabolic adaptation of invasive tumor cells, which increased lactate production under long-term hypoxia. Conclusions: The use of glycolysis versus mitochondrial respiration for energy production within human GBM tumors is highly dependent on the specific microenvironment. The metabolic adaptability of GBM cells highlights the difficulty of targeting one specific metabolic pathway for effective therapeutic intervention.