Oral Soft Tissue Regeneration Using Nano Controlled System Inducing Sequential Release of Trichloroacetic Acid and Epidermal Growth Factor

Background@#The effect of nano controlled sequential release of trichloroacetic acid (TCA) and epidermal growth factor (EGF) on the oral soft tissue regeneration was determined. @*Methods@#Hydrophobically modified glycol chitosan (HGC) nano controlled system was developed for the sequential release of TCA and EGF, and the release pattern was identified. The HGC-based nano controlled release system was injected into the critical-sized defects created in beagles’ palatal soft tissues. The palatal impression and its scanned body was obtained on various time points post-injection, and the volumetric amount of soft tissue regeneration was compared among the three groups: CON (natural regeneration control group), EXP1 (TCA-loaded nano controlled release system group), EXP2 (TCA and EGF individually loaded nano controlled release system). DNA microarray analysis was performed and various soft tissue regeneration parameters in histopathological specimens were measured. @*Results@#TCA release was highest at Day 1 whereas EGF release was highest at Day 2 and remained high until Day 3. In the volumetric measurements of impression body scans, no significant difference in soft tissue regeneration between the three groups was shown in two-way ANOVA. However, in the one-way ANOVA at Day 14, EXP2 showed a significant increase in soft tissue regeneration compared to CON. High correlation was determined between the histopathological results of each group. DNA microarray showed up-regulation of various genes and related cell signaling pathways in EXP2 compared to CON. @*Conclusion@#HGC-based nano controlled release system for sequential release of TCA and EGF can promote regeneration of oral soft tissue defects.

Oral Soft Tissue Regeneration Using Nano Controlled System Inducing Sequential Release of Trichloroacetic Acid and Epidermal Growth Factor

Background@#The effect of nano controlled sequential release of trichloroacetic acid (TCA) and epidermal growth factor (EGF) on the oral soft tissue regeneration was determined. @*Methods@#Hydrophobically modified glycol chitosan (HGC) nano controlled system was developed for the sequential release of TCA and EGF, and the release pattern was identified. The HGC-based nano controlled release system was injected into the critical-sized defects created in beagles’ palatal soft tissues. The palatal impression and its scanned body was obtained on various time points post-injection, and the volumetric amount of soft tissue regeneration was compared among the three groups: CON (natural regeneration control group), EXP1 (TCA-loaded nano controlled release system group), EXP2 (TCA and EGF individually loaded nano controlled release system). DNA microarray analysis was performed and various soft tissue regeneration parameters in histopathological specimens were measured. @*Results@#TCA release was highest at Day 1 whereas EGF release was highest at Day 2 and remained high until Day 3. In the volumetric measurements of impression body scans, no significant difference in soft tissue regeneration between the three groups was shown in two-way ANOVA. However, in the one-way ANOVA at Day 14, EXP2 showed a significant increase in soft tissue regeneration compared to CON. High correlation was determined between the histopathological results of each group. DNA microarray showed up-regulation of various genes and related cell signaling pathways in EXP2 compared to CON. @*Conclusion@#HGC-based nano controlled release system for sequential release of TCA and EGF can promote regeneration of oral soft tissue defects.

Chemical Regeneration of Wound Defects: Relevance to the Canine Palatal Mucosa and Cell Cycle Up-Regulation in Human Gingival Fibroblasts

BACKGROUND: Trichloroacetic acid (TCA) is an agent widely applied in dermatology for skin regeneration. To test whether TCA can offer an advantage for the regeneration of oral soft tissue defects, the cellular events following TCA application were explored in vitro and its influence on the oral soft tissue wound healing was evaluated in a canine palate model.METHODS: The cytotoxicity and growth factor gene expression in human gingival fibroblasts were tested in vitro following the application of TCA at four concentrations (0.005%, 0.05%, 0.5% and 1%) with different time intervals (0, 3, 9 and 21 h). One concentration of TCA was selected to screen the genes differentially expressed using DNA microarray and the associated pathways were explored. TCA was injected in open wound defects of the palatal mucosa from beagle dogs (n = 3) to monitor their healing and regeneration up to day 16-post-administration.RESULTS: While the 0.5–1% concentration induced the cytoxicity, a significantly higher expression of growth factor genes was observed after 3 and 9 h following the 0.5% TCA application in comparison to other groups. DNA microarray analysis in 0.5% TCA group showed 417 genes with a significant 1.5-fold differential expression, involving pathways of cell cycle, FoxO signaling, p53 signaling, ubiquitin mediated proteolysis and cAMP signaling. In vivo results showed a faster reepithelialization of TCA-treated wounds as compared to spontaneous healingCONCLUSION: TCA promoted the healing and regeneration of oral soft tissue wound defects by up-regulating the cell cycle progression, cell growth, and cell viability, particularly at a concentration of 0.5%.

The Effect of Fibronectin-Immobilized Microgrooved Titanium Substrata on Cell Proliferation and Expression of Genes and Proteins in Human Gingival Fibroblasts

BACKGROUND: We aimed to determine the effect of fibronectin (FN)-immobilized microgrooved titanium (Ti) on human gingival fibroblast proliferation, gene expression and protein expression. METHODS: Photolithography was used to fabricate the microgrooved Ti, and amine funtionalization (silanization) was used for FN immobilization on titanium surfaces. Cell proliferation, gene expression and protein expression were analyzed, followed by multiple regression analysis for determining the influential factors on cell proliferation. RESULTS: FN-immobilized microgrooved Ti significantly enhanced the fibroblast proliferation in various timelines of culture, among which a burst of fivefold increase is induced at 96 h of culture compared to that on the control smooth Ti. We suggest a presence of the synergistic promotion effect of microgrooves and FN immobilization on fibroblast proliferation. Through a series of analyses on the expression of various genes and proteins involved in cell adhesion and proliferation, cyclin-dependent kinase 6, cyclin D1, integrin α5, oncogene c-Src, osteonectin, paxillin and talin-2 were determined as influential factors on promoting fibroblast proliferation induced by FN-immobilized microgrooved Ti. CONCLUSION: FN-immobilized microgrooved Ti can act as an effective surface for enhancing fibroblast proliferation, and can be used for promoting soft tissue response on the connective tissue attachment zone of biomaterial surfaces.

Effect of microgrooves and fibronectin conjugation on the osteoblast marker gene expression and differentiation

PURPOSE: To determine the effect of fibronectin (FN)-conjugated, microgrooved titanium (Ti) on osteoblast differentiation and gene expression in human bone marrow-derived mesenchymal stem cells (MSCs). MATERIALS AND METHODS: Photolithography was used to fabricate the microgrooved Ti, and amine functionalization (silanization) was used to immobilize fibronectin on the titanium surfaces. Osteoblast differentiation and osteoblast marker gene expression were analyzed by means of alkaline phosphatase activity assay, extracellular calcium deposition assay, and quantitative real-time PCR. RESULTS: The conjugation of fibronectin on Ti significantly increased osteoblast differentiation in MSCs compared with non-conjugated Ti substrates. On the extracellular calcium deposition assays of MSCs at 21 days, an approximately two-fold increase in calcium concentration was observed on the etched 60-microm-wide/10-microm-deep microgrooved surface with fibronectin (E60/10FN) compared with the same surface without fibronectin (E60/10), and a more than four-fold increase in calcium concentration was observed on E60/10FN compared with the non-etched control (NE0) and etched control (E0) surfaces. Through a series of analyses to determine the expression of osteoblast marker genes, a significant increase in all the marker genes except type I collagen alpha1 mRNA was seen with E60/10FN more than with any of the other groups, as compared with NE0. CONCLUSION: The FN-conjugated, microgrooved Ti substrate can provide an effective surface to promote osteoblast differentiation and osteoblast marker gene expression in MSCs.