An in vitro anti-inflammatory effect of Thai propolis in human dental pulp cells.

OBJECTIVE: To explore the potential for development of Thai propolis extract as a pulp capping agent to suppress pulpal inflammation from dental pulp infections. This study aimed to examine the anti-inflammatory effect of the propolis extract on the arachidonic acid pathway, activated by interleukin (IL)-1ß, in cultured human dental pulp cells. METHODOLOGY: Dental pulp cells, isolated from three freshly extracted third molars, were first characterized for their mesenchymal origin and treated with 10 ng/ml of IL-1ß in the presence or absence of non-toxic concentrations of the extract from 0.08 to 1.25 mg/ml, as determined by the PrestoBlue cytotoxic assay. Total RNA was harvested and analyzed for mRNA expressions of 5-lipoxygenase (5-LOX) and cyclooxygenase-2 (COX-2). Western blot hybridization was performed to investigate COX-2 protein expression. Culture supernatants were assayed for released prostaglandin E2 levels. Immunofluorescence was conducted to determine involvement of nuclear factor-kappaB (NF-kB) in the inhibitory effect of the extract. RESULTS: Stimulation of the pulp cells with IL-1ß resulted in the activation of arachidonic acid metabolism via COX-2, but not 5-LOX. Incubation with various non-toxic concentrations of the propolis extract significantly inhibited upregulated COX-2 mRNA and protein expressions upon treatment with IL-1ß (p<0.05), resulting in a significant decrease in elevated PGE2 levels (p<0.05). Nuclear translocation of the p50 and the p65 subunits of NF-kB upon treatment with IL-1ß was also blocked by incubation with the extract. CONCLUSIONS: Upregulated COX-2 expression and enhanced PGE2 synthesis upon treatment with IL-1ß in human dental pulp cells were suppressed by incubation with non-toxic doses of Thai propolis extract via involvement of the NF-kB activation. This extract could be therapeutically used as a pulp capping material due to its anti-inflammatory properties.

Anti-Inflammatory Comparison of Melatonin and Its Bromobenzoylamide Derivatives in Lipopolysaccharide (LPS)-Induced RAW 264.7 Cells and Croton Oil-Induced Mice Ear Edema.

The pineal gland is a neuroendocrine organ that plays an important role in anti-inflammation through the hormone melatonin. The anti-inflammatory effects of melatonin and its derivatives have been reported in both in vitro and in vivo models. Our previous study reported the potent antioxidant and neuroprotective activities of bromobenzoylamide substituted melatonin. In silico analysis successfully predicted that melatonin bromobenzoylamid derivatives were protected from metabolism by CYP2A1, which is a key enzyme of the melatonin metabolism process. Therefore, the anti-inflammatory activities of melatonin and its bromobenzoylamide derivatives BBM and EBM were investigated in LPS-induced RAW 264.7 macrophages and croton oil-induced ear edema in mice. The experiments showed that BBM and EBM significantly reduced production of the inflammatory mediators interleukin-6 (IL-6), prostaglandin E2 (PGE2), and nitric oxide (NO) in a dose-dependent manner, but only slightly affected TNF-α in LPS-induced RAW 264.7 macrophages. This suggests that modifying melatonin at either the N1-position or the N-acetyl side chain affected production of NO, PGE2 and IL-6 in in vitro model. In the croton oil-induced mouse ear edema model, BBM, significantly decreased ear edema thickness at 2-4 h. It leads to conclude that bromobenzoylamide derivatives of melatonin may be one of the potential candidates for a new type of anti-inflammatory agent.

Inhibitory Effects of Erythrosine/Curcumin Derivatives/Nano-Titanium Dioxide-Mediated Photodynamic Therapy on Candida albicans.

This study focuses on the role of photosensitizers in photodynamic therapy. The photosensitizers were prepared in combinations of 110/220 µM erythrosine and/or 10/20 µM demethoxy/bisdemethoxy curcumin with/without 10% (w/w) nano-titanium dioxide. Irradiation was performed with a dental blue light in the 395-480 nm wavelength range, with a power density of 3200 mW/cm2 and yield of 72 J/cm2. The production of ROS and hydroxyl radical was investigated using an electron paramagnetic resonance spectrometer for each individual photosensitizer or in photosensitizer combinations. Subsequently, a PrestoBlue® toxicity test of the gingival fibroblast cells was performed at 6 and 24 h on the eight highest ROS-generating photosensitizers containing curcumin derivatives and erythrosine 220 µM. Finally, the antifungal ability of 22 test photosensitizers, Candida albicans (ATCC 10231), were cultured in biofilm form at 37 °C for 48 h, then the colonies were counted in colony-forming units (CFU/mL) via the drop plate technique, and then the log reduction was calculated. The results showed that at 48 h the test photosensitizers could simultaneously produce both ROS types. All test photosensitizers demonstrated no toxicity on the fibroblast cells. In total, 18 test photosensitizers were able to inhibit Candida albicans similarly to nystatin. Conclusively, 20 µM bisdemethoxy curcumin + 220 µM erythrosine + 10% (w/w) nano-titanium dioxide exerted the highest inhibitory effect on Candida albicans.

Preservation of the viability and gene expression of human periodontal ligament cells by Thai propolis extract.

BACKGROUND/AIM: Success of tooth replantation depends on the quality and quantity of periodontal ligament (PDL) cells. The aims of this study were to evaluate Thai propolis extract as a storage medium for maintaining PDL cell viability and preserving gene expressions in PDL tissues. MATERIALS AND METHODS: PDL cells from human premolars were tested for cytotoxicity of the extract by PrestoBlue assay to determine a non-toxic concentration. Subsequently, 96 freshly extracted premolars were allocated into different treatment groups. Control groups were freshly extracted premolars or they had been stored dry for 12 hours. Experimental avulsed teeth were created by leaving them air-dried for 30 minutes immediately after extraction, then they were immersed in Thai propolis extract, HBSS or milk for 3, 6 and 12 hours. After tooth storage, the remaining PDL cells were determined for their cell viability. RNA isolated from PDL tissues of three premolars treated similarly was analysed for periostin and S100A4 expressions using RT-qPCR. RESULTS: Thai propolis extract at 0.625 mg mL-1 promoted the greatest PDL cell viability. Tooth storage in 0.625 mg mL-1 Thai propolis extract, HBSS or milk showed no difference in maintaining cell viability. Periostin mRNA level was preserved by Thai propolis extract. Expression of S100A4 mRNA in PDL tissues stored in all tested media was dampened. CONCLUSIONS: PDL cells from mock avulsed teeth stored in 0.625 mg mL-1 Thai propolis extract for 3, 6 and 12 hours remained viable and the expression of periostin was preserved. This study suggests this extract as an alternative for a tooth storage medium for up to 12 hours. However, transporting an avulsed tooth in a storage medium for extended extra-oral time might affect the PDL cell phenotypes.

Anti-inflammatory effect of photodynamic therapy using guaiazulene and red lasers on peripheral blood mononuclear cells.

INTRODUCTION: Photodynamic therapy improves oral mucositis treatment. The reactive oxygen species (ROS) generated from this reaction could contribute to an anti-inflammatory effect by suppressing inflammatory cells. OBJECTIVE: To evaluate the anti-inflammatory effect of photodynamic therapy using guaiazulene and a red laser in peripheral blood mononuclear cells (PBMCs). METHODS: Guaiazulene solutions (1, 2, 5, 25, 35, and 100 µM in 99.8 % methanol) were irradiated with red laser light (625 nm, 146.2 mW/cm2) in continuous mode at 0, 4, and 8 J/cm2 in black 96-well plates. ROS were measured using spin trapping technique with electron spin resonance (ESR) spectroscopy and fluorescence. The two highest concentrations were tested using cell viability (PrestoBlue®) and anti-inflammation (RANTES and PGE2 ELISA) assay kits. Kruskal-Wallis and Dunn Bonferroni tests were used for statistical analyses with significant differences at p-value < 0.05. RESULTS: Guaiazulene solutions between 2 and 5 µM exposed to red laser light at 4-8 J/cm2 generated significantly more singlet oxygen compared to the no guaiazulene group (p < 0.01) and reduced RANTES and PGE2 levels in TNF-α-inflamed peripheral blood mononuclear cells without affecting cell viability. CONCLUSION: Photodynamic activation of guaiazulene generated singlet oxygen and suppressed inflammatory markers in PBMCs.

Topical oral wound healing potential of anthocyanin complex: animal and clinical studies.

AIM: An anthocyanin complex (AC), combined Zea mays and Clitoria ternatea extracts, was evaluated for topical oral wound healing in rats and a clinical trial in orthodontic patients. METHODS/RESULTS: AC enhanced anthocyanin permeation in vitro. In rats, 10% w/w of AC in a mucoadhesive gel (AG) reduced erythema and sizes of oral wounds after topical applications at higher extent than its placebo gel. Acute orthodontic wounds in 68 volunteers were randomly assigned to topically receive either AG or placebo gel and double-blind assessed. Wound size reduction and wound closure enhancement were obvious in AG-treated group on day 3 (p < 0.05). CONCLUSION: At 10% w/w, AC promoted wound closure and possessed a potential in healing stimulation of acute oral wounds.

Topical Niosome Gel Containing an Anthocyanin Complex: a Potential Oral Wound Healing in Rats.

Anthocyanins from dietary sources showing potential benefits as anti-inflammatory in oral lesions were developed as an anthocyanin complex (AC), comprised of extracts of Zea mays (CC) and Clitoria ternatea (CT), and formulated into a niosome gel to prove its topical oral wound healing in vitro and in vivo investigations. The AC formed nano-sized clusters of crystalline-like aggregates, occurring through both intra- and inter-molecular interactions, resulting in delivery depots of anthocyanins, following encapsulation in niosomes and incorporation into a mucoadhesive gel. In vitro permeation of anthocyanins was improved by complexation and further enhanced by encapsulation in niosomes. Collagen production in human gingival fibroblasts was promoted by AC and AC niosomes, but not CC or CT. The in vivo wound healing properties of AC gel (1 and 10%), AC niosome gel (1 and 10%), fluocinolone acetonide gel, and placebo gel were investigated for incisional wounds in the buccal cavities of Wistar rats. AC gel and AC niosome gel both reduced wound sizes after 3 days. AC niosome gel (10%) gave the highest reduction in wound sizes after day 3 (compared to fluocinolone acetonide gel, p < 0.05), and resulted in 100% wound healing by day 5. Histological observations of cross-sectioned wound tissues revealed the adverse effects of fluocinolone gel and wound healing potential of AC niosome gel. Topical application of AC niosome gel exhibited an anti-inflammatory effect and promoted oral wound closure in rats, possibly due to the improved mucosal permeability and presence of delivery depots of AC in the niosome gel.