We show for the first time that sustained activity of histone deacetylase 6 (HDAC6) in wounds of diabetic mice contributes to delayed wound healing. Topical application of HDAC6 inhibitor; Tubastatin A (TSA) gel promoted the wound healing in diabetic mice compared to blank gel treated mice. TSA topical application reduced the infiltrating neutrophils, macrophages and T cells in the early phase of wounds. Similarly TSA topical application promoted the wound healing by inducing collagen deposition, angiogenesis, re-epithelization and fibrotic factors in the late stages of diabetic wounds. Protein analysis of the diabetic wounds treated with TSA showed increased acetylation of α-tubulin with no effect on the expression of pro-IL1β, pro-caspase-1 and active caspase-1 indicating no influence of TSA treatment on inflammasome activation. Macrophages are crucial for sustained inflammation activation; hence we have explored the effect of TSA on inflammatory factors (IL-1β and IL-10) expression using raw 264.7 cells. Macrophages exhibited upregulation of HDAC6, IL-1β and down regulation of IL-10 upon stimulation with high glucose and LPS. Selective inhibition of HDAC6 with TSA inhibited the IL-1β and promoted IL-10. Detailed probing to determine the effect on IL-1β resulted that TSA inhibit IL-1β release by inhibiting exocytosis while not showing any effect on its maturation. Similarly, inhibition of HDAC6 in macrophages stimulated with high glucose and LPS promoted the acetylation of tubulin. For further confirmation we have used nocodazole (known acetylation inhibitor) and found that nocodazole reversed the tubulin acetylation in high glucose conditions. Our findings indicate that sustained HDAC6 expression in diabetic wounds contributes to impaired early healing responses and HDAC6 may represent a new therapeutic target for diabetic wounds.
Hybrid implants combine both Titanium (Ti) and Magnesium (Mg) are prevalent nowadays. The long-term implications of Ti and Mg implants within the human body are not yet fully understood. Many implant failure cases due to inflammation, allergic responses, and aspect loosening have been reported frequently. Particles generated through daily wear and tear of implants may worsen the situation by causing acute complications. An in-depth understanding of the behavior of metal particles with human osteoblasts is necessary. In this study, a novel and systematic attempt was made to understand the effects of different concentrations of Ti and Mg particles to the osteoblastic SAOS2 cell: toxicity, alterations to mitochondria, and changes to the specific gene and protein expression. Ti particles were found toxic to SAOS2 cells at different dosages, while Mg particles at lower concentrations could improve cell viability. To understand this phenomenon better, we have measured cellular reactive oxygen species (ROS) production and cell apoptosis & necrosis percentage. We also have checked the mitochondrial structure with transmission electron microscope (TEM), and mitochondrial function using Tetramethyl rhodamine, ethyl ester staining (TMRE). NDUFB6, SDHC, and ATP5F1 were the essential mitochondrial genes involved in the ROS production and ATP production. Immunocytochemistry (ICC) and realtime polymerase chain reaction (qPCR) were implemented to check the regulations of these related genes.
: Biodentine™ is a tricalcium silicate-based cement material that has a great impact on different biological processes of dental stem cells, compared to other biomaterials. Therefore, we aimed to investigate the optimum biocompatible concentration of Biodentine™ with stem cells derived from periodontal ligament (hPDLSCs) by determining cell proliferation, cytotoxicity, migration, adhesion and mineralization potential. hPDLSCs were treated with Biodentine™ extract at different concentrations; 20, 2, 0.2 and 0.02 mg/mL. Cells cultured without Biodentine™ were used as a blank control. The proliferation potential of hPDLSCs was evaluated by MTT viability analysis for 6 days. Cytotoxicity assay was performed after 3 days by using AnnexinV/7AAD. Migration potential was investigated by wound healing and transwell migration assays at both cellular and molecular levels. The expression levels of chemokines CXCR4, MCP-1 and adhesion molecules FGF-2, FN, VCAM and ICAM-1 were measured by qPCR. The communication potentials of these cells were determined by adhesion assay. In addition, mineralization potential was evaluated by measuring the expression levels of osteogenic markers; ALP, OCN, OPN and Collagen type1 by qPCR. Our results showed significant increase in the proliferation of hPDLSCs at low concentrations of Biodentine™ (2, 0.2 and 0.02 mg/mL) while higher concentration (20 mg/mL) exhibited cytotoxic effect on the cells. Moreover, 2 mg/mL Biodentine™ showed a significant increase in the migration, adhesion and mineralization potentials of the derived cells among all concentrations and when compared to the blank control. Our findings suggest that 2 mg/mL of Biodentine™ is the most biocompatible concentration with hPDLSCs, showing a high stimulatory effect on the biological processes.
Metal particles or debris can be generated by wear and tear from bone implants. Reports mentioned that debris can circulate in blood and trigger inflammation, aseptic loosening, and other complications. The mechanism of these phenomena remains unclear. This research is to investigate the toxicity of titanium and magnesium in bone implants due to these two are the most commonly used biomaterials based on excellent biocompatibility, low elastic modulus, and good mechanical strength. During the cytotoxicity test, the metal particles were added to the petri dish. Ti particles showed toxic to osteoblast at different dosages and time, while Mg particles can reduce the Ti induced metal toxicity to the cells and boost cell proliferation. Mg particles can be toxic to osteoblast at a higher dosage as well. To better understand this phenomenon, human osteoblast cell line SAOS2 were exposed to different concentration of Ti/Mg/Ti-Mg particles. Cell proliferation was measured at 48/72/120 hours. The flow cytometry analysis results showed that Ti induced cell toxicity was through an increase in Reactive Oxygen Species production and induced cell apoptosis and necrosis. Fluorescence microscopy was implemented to observe the cell damage in mitochondria. Quantitative real-time PCR analysis for relative mRNA expression of SAOS2 cells was studied to understand the process better. Through this study, a proper protocol was established to measure the cytotoxicity of metal particles. However, how to quantify the debris from wear and tear inside the human body, and the comprehensive mechanism of cellular interaction among particles, cells, and tissues require further investigation. Biography: Niyou is currently a Ph.D. candidate and research engineer at the National University of Singapore. He is working on a collaboration project between Mechanical Engineering and Anatomy. He has a rich mechanical background, strong biological knowledge, and hands-on experience. He has published one paper in Materials Science and Engineering C, and two papers are currently under review. Presenting author details Full name: Niyou Wang Contact number: 90088491 Email: e0222912@u.nus.edu LinkedIn account: Niyou
Tissue designing is a biomedical designing discipline that utilizes a blend of cells, designing, materials techniques, and reasonable biochemical and physicochemical variables to reestablish, keep up with, improve, or supplant various sorts of natural tissues. Tissue designing frequently includes the utilization of cells put on tissue platforms in the arrangement of new suitable tissue for a clinical reason however isn't restricted to applications including cells and tissue frameworks. While it was once ordered as a sub-field of biomaterials, having filled in extension and significance it very well may be considered as a field in its own.
