Efficacy of double antibiotics in hydroxypropyl methylcellulose for bactericidal activity against Enterococcus faecalis and Streptococcus gordonii in biofilm.

OBJECTIVE: This study aimed to investigate the efficacy of double antibiotics, including ciprofloxacin and metronidazole, in a new vehicle, hydroxypropyl methylcellulose (HPMC), against Enterococcus faecalis and Streptococcus gordonii grown in biofilm. DESIGN: Human mandibular premolars were prepared and divided into four groups: (i) negative control, (ii) positive control, (iii) infected with E. faecalis and S. gordonii for 21 days and intracanally medicated with double antibiotics in HPMC, and (iv) infected with E. faecalis and S. gordonii for 21 days and intracanally medicated with calcium hydroxide (UltraCal™). The efficacy of medication for 14 or 28 days was determined by bacterial cultures and RT-qPCR for absolute quantities of E. faecalis and S. gordonii cDNA and for relative mRNA expressions of pbp5 and gtfG genes. RESULTS: There were significant decreases in the mean colony forming units and mean cDNA amounts of E. faecalis and S. gordonii in group (iii) on days 14 and 28 compared to those in group (ii) (p < 0.01). However, the mean cDNA amounts of E. faecalis and S. gordonii in group (iv) were found to be significantly increased on day 28 (p < 0.05). The mRNA expression of gtfG was significantly decreased in groups (iii) and (iv) on days 14 and 28, whereas that of pbp5 was significantly increased in group (iv) on days 14 and 28 (p < 0.01). CONCLUSION: Double antibiotics in HPMC gel showed an in vitro efficacy against E. faecalis and S. gordonii grown in biofilm, suggesting its clinical application as an intracanal medicament for both primary and persistent infections.

A biological active artificial saliva formulation containing flower mucilage from Ceylon Spinach (Basella alba Linn.).

Ceylon Spinach (Basella albe) is an edible perennial vine found in tropical Asia and Africa, known as vegetables containing mucilage. Its mucilage from flowers was extracted by microwaving and precipitated with 95% ethanol. Five artificial saliva formulations composing of mucilage from Ceylon Spinach, calcium chloride (CaCl2), potassium chloride (KCl) and sodium fluoride (NF) were developed. The best formulation No.5 containing 0.61% of the mucilage with the non-Newtonian pseudoplastic flow (8.9 ± 0.2 cP) and the wetting time (12.50 ± 2.24 min) similar to the normal human saliva was selected. This artificial saliva formulation exhibited biological activities including an antioxidative activity by DPPH free radical scavenging with the SC50 of 14.26 ± 2.00 mg/ml (0.05 folds of ascorbic acid), and the adhesion inhibition of S. mutans on hydroxyapatite beads at 17.01 ± 7.75%, while the natural human saliva exhibited an increase bacterial adhesion of 33.10 ± 9.70%. The safety of this formulation which gave no cytotoxicity on normal human gingival fibroblasts at 99.20 ± 21.09% cell viability was also demonstrated. The results from this study have indicated high biological activity and safety of the developed formulation containing mucilage from Ceylon Spinach which is potential to be used as artificial saliva for xerostomia patients.

Effects of prostaglandin E2 on clonogenicity, proliferation and expression of pluripotent markers in human periodontal ligament cells.

BACKGROUND AND OBJECTIVE: Based on our earlier work on the response of periodontal ligament (PDL) cells to mechanical stress by induction of cyclooxygenase expression and production of prostaglandin PGE2 that could regulate mineralization of PDL cells, it was hypothesized that PGE2 had potential effects on PDL stemness. In this study, we aimed to investigate clonogenicity, proliferation and expression of certain pluripotent markers, considered to be characteristics of PDL stemness, in response to treatment with exogenously-added PGE2. MATERIAL AND METHODS: Human PDL cells were cultured and treated with various doses of PGE2, and the aforementioned characteristics of PDL stemness were analyzed. RESULTS: The clonogenicity and proliferation were significantly enhanced by PGE2 at low concentrations (0.01, 0.1 and 1ng/ml; P<0.05), but only the proliferation was significantly diminished by PGE2 at a high concentration (100ng/ml; P<0.05). Expression of NANOG and OCT4 mRNA and protein was increased by PGE2 treatment at 0.1 and 1ng/ml. Consistently, expression of stage-specific embryonic antigen 4, a putative stem cell marker, was significantly augmented by PGE2 treatment at 1ng/ml (P<0.05). CONCLUSION: Our findings suggest that although a high dose of PGE2 (100ng/ml) inhibits proliferation of PDL cells, PGE2 at low doses appears to play a role in the maintenance of PDL stemness.

Decreased levels of matrix metalloproteinase-2 in root-canal exudates during root canal treatment.

OBJECTIVE: To determine the matrix metalloproteinase-2 (MMP-2) levels in root-canal exudates from teeth undergoing root-canal treatment. MATERIAL AND METHODS: The root-canal exudates from six teeth with normal pulp and periradicular tissues that required intentional root canal treatment for prosthodontic reasons and from twelve teeth with pulp necrosis and asymptomatic apical periodontitis (AAP) were sampled with paper points for bacterial culture and aspirated for the detection of proMMP-2 and active MMP-2 by gelatin zymography and the quantification of MMP-2 levels by ELISA. RESULTS: By gelatin zymography, both proMMP-2 and active MMP-2 were detected in the first collection of root-canal exudates from teeth with pulp necrosis and AAP, but not from teeth with normal pulp, and their levels gradually decreased and disappeared at the last collection. Consistently, ELISA demonstrated a significant decrease in MMP-2 levels in the root-canal exudates of teeth with pulp necrosis and AAP following root canal procedures (p<0.05). Furthermore, the MMP-2 levels were significantly lower in the negative bacterial culture than those in the positive bacterial culture (p<0.001). CONCLUSIONS: The levels of MMP-2 in root-canal exudates from teeth with pulp necrosis and AAP were gradually reduced during root canal procedures. Future studies are required to determine if MMP-2 levels may be used as a biomolecule for the healing of apical lesions, similar to the clinical application of MMP-8 as a biomarker.

Physicochemical properties and biological activities of Thai plant mucilages for artificial saliva preparation.

CONTEXT: Plant mucilages can be found in various parts of several Thai plants, which can be used as thickening, moisturizing, and lubricating agents in artificial saliva formulations. OBJECTIVE: The objective of this study was to evaluate the physicochemical properties, biological activity, and cytotoxicity of Thai plant mucilages. MATERIALS AND METHODS: The mucilages from Thai plants were extracted by various processes (temperature and pH variation, microwave oven, steam, and Tris-HCl buffer extraction). The viscosity and the rheology were evaluated using viscometer. Antioxidative activities including DPPH radical scavenging and metal chelating activities were investigated. The mucilages were determined for cytotoxicity on normal human gingival fibroblasts and anti-adherent activity of Streptococcus mutans. RESULTS: Mucilages from Ocimum citriodorum Vis. (Lamiaceae), Artocarpus heterophyllus Lam. (Moraceae), Abelmoschus esculentus (Linn.) Moench. (Malvaceae), and Basella alba Linn. (Basellaceae) exhibited pseudoplastic non-Newtonian rheology. The highest DPPH radical-scavenging and metal-chelating activities were observed in the mucilages from B. alba (microwave, 3 min) and A. esculentus (microwave, 1 min) with the SC50 and MC50 values (50% of scavenging activity and 50% of metal chelating activity, respectively) of 0.71 ± 0.32 and 1.11 ± 0.52 mg/ml, respectively. Most mucilages exhibited no cytotoxicity to normal human gingival fibroblasts. The mucilage from A. esculentus (microwave, 5 min) gave the shortest wetting time of 2.75 ± 0.51 min. The highest S. mutans adhesion inhibition was observed in A. esculentus (pH 11) of 5.39 ± 9.70%. DISCUSSION AND CONCLUSION: This study has indicated the suitable physicochemical and biological properties and the potential application of mucilages from Thai plants for artificial saliva preparation.

Potent anti-proliferative effects against oral and cervical cancers of Thai medicinal plants selected from the Thai/Lanna medicinal plant recipe database "MANOSROI III".

CONTEXT: Thai/Lanna medicinal plant recipes have been used for the treatment of several diseases including oral and cervical cancers. OBJECTIVE: To investigate anti-proliferative activity on human cervical (HeLa) and oral (KB) cancer cell lines of medicinal plants selected from Thai/Lanna medicinal plant recipe database "MANOSROI III". MATERIALS AND METHODS: Twenty-three methanolic plant crude extracts were tested for phytochemicals and anti-proliferative activity on HeLa and KB cell lines for 24 h by the sulforhodamine B (SRB) assay at the doses of 1 × 10(1)-1 × 10(-6 )mg/ml. The nine extracts with the concentrations giving 50% growth inhibition (GI50) lower than 100 µg/ml were further semi-purified by liquid/liquid partition in order to evaluate and enhance the anti-proliferative potency. RESULTS: All extracts contained steroids/triterpenoids, but not xanthones. The methanolic extracts of Gloriosa superba L. (Colchinaceae) root and Albizia chinensis (Osbeck) Merr. (Leguminosae-Mimosoideae) wood gave the highest anti-proliferative activity on HeLa and KB cell lines with the GI50 values of 0.91 (6.0- and 0.31-fold of cisplatin and doxorubicin) and 0.16 µg/ml (28.78- and 82.29-fold of cisplatin and doxorubicin), respectively. Hexane and methanol-water fractions of G. superba exhibited the highest anti-proliferative activity on HeLa and KB cell lines with the GI50 values of 0.15 (37- and 1.9-fold of cisplatin and doxorubicin) and 0.058 µg/ml (77.45- and 221.46-fold of cisplatin and doxorubicin), respectively. DISCUSSION AND CONCLUSION: This study has demonstrated the potential of plants selected from MANOSROI III database especially G. superba and A. chinensis for further development as anti-oral and cervical cancer agents.

Effects of plasma treatment on the shear bond strength between fiber-reinforced composite posts and resin composite for core build-up.

The aim of this study was to evaluate the effects of plasma treatment on adhesion between fiber-reinforced posts and a composite core material. Two types of posts, methacrylate-based (FRC Postec) and epoxy resin-based (DT Light-Post), were treated with oxygen plasma (O(2)), argon plasma (Ar), nitrogen plasma (N(2)), or helium mixed with nitrogen plasma (He+N(2)) using a radio-frequency generator before bonding to a methacrylate-based composite. Pull-out tests were performed using a universal testing machine. Surface roughness of each group was evaluated using a profilometer. On tensile-shear bond strength, statistical analysis revealed that the type of post, type of plasma treatment, and their interaction significantly influenced the results (p<0.05). Tukey's test revealed significant differences in tensile-shear bond strength between the control and other plasma treatment groups (p<0.05). On surface roughness, Tukey's test revealed significant differences between the control group and the Ar group (p<0.05) with DT Light Post. Plasma treatment appeared to increase the tensile-shear bond strength between post and composite.

Aberrant expression of Smad4, a TGF-beta signaling molecule, in oral squamous cell carcinoma.

Although carcinogenesis of oral squamous cell carcinoma (OSCC) has been studied by many investigators in the past decade, the available evidence about its molecular mechanism is inconclusive. The objective of the present study was to compare expression of Smad4, a signaling molecule of the transforming growth factor beta (TGF-beta) pathway, between OSCC and normal oral mucosa. We assayed expression of Smad4 in OSCC and normal oral mucosa by performing immunohistochemistry using paraffin-embedded tissue samples. We also compared expression of Smad4 protein between OSCC lines and normal oral keratinocytes, using Western blot analysis. Smad4 expression was observed in only 60% of OSCC tissue samples, whereas it was observed in 82% of normal oral mucosa samples. Reduced Smad4 expression was clearly observed in all OSCC lines, compared with normal oral keratinocytes. These findings suggest that aberration of the TGF-beta pathway, as indicated by a reduction or absence of Smad4 expression, promotes carcinogenesis of OSCC.

Actinobacillus actinomycetemcomitans lipopolysaccharide activates matrix metalloproteinase-2 and increases receptor activator of nuclear factor-kappaB ligand expression in human periodontal ligament cells.

BACKGROUND: The lipopolysaccharide (LPS) of A. actinomycetemcomitans is one of the major pathogenic factors in periodontal disease. It induces secretion of proinflammatory cytokines and is involved in alveolar bone destruction. We hypothesized that the LPS of A. actinomycetemcomitans could affect the activation of matrix metalloproteinase (MMP)-2 and the expression of receptor activator of nuclear factor kappa B ligand (RANKL) and osteoprotegerin in human periodontal ligament (HPDL) cells leading to the destruction of periodontium. METHODS: HPDL cells were cultured in serum-free medium with or without the LPS of A. actinomycetemcomitans for 36 hours. The activation of MMP-2 was analyzed by zymography. Changes of the expression of RANKL and osteoprotegerin (OPG) were examined by reverse transcription-polymerase chain reaction and supported by Western blot analysis. RESULTS: The activation of MMP-2 could be induced by the LPS of A. actinomycetemcomitans in HPDL cells and could be inhibited by a serine protease inhibitor. This result suggested that the LPS might activate MMP-2 through a serine protease-dependent pathway. This activation was also blocked by NF-kappaB inhibitor, which indicated the involvement of NF-kappaB. The upregulation of RANKL but not OPG by the LPS was found in both transcription and translation and could be reduced by indomethacin. In addition, serine protease inhibitor also inhibited the upregulation of RANKL, suggesting the activity of serine protease. CONCLUSIONS: The effect of the LPS of A. actinomycetemcomitans on HPDL cells is serum-independent and the induction of the activation of MMP-2 and the expression of RANKL are serine protease-dependent pathways. The results suggest the role of HPDL cells in the pathogenesis of periodontitis.

The synergistic effect of TGF-beta and 1,25-dihydroxyvitamin D3 on SPARC synthesis and alkaline phosphatase activity in human pulp fibroblasts.

1,25(OH)2D3 and TGF-beta can influence the function and differentiation of dental pulp fibroblasts. In this study, we examined the effect of 1,25(OH)2D3 and TGF-beta on the synthesis of SPARC and ALP activity in human pulp fibroblasts. Two isoforms of SPARC, the 43 and 38 kDa, were detected in this cell type. TGF-beta increased the synthesis of SPARC about 2.5-fold after 3 days of treatment but had no effect on the ALP activity. On the contrary, 1,25(OH)2D3 increased ALP activity 2-fold but had no effect on SPARC. The combination of TGF-beta and 1,25(OH)2D3 significantly induced SPARC synthesis and ALP activity by 5 and 9 folds, respectively (P<0.05). This finding suggested the synergistic effect between TGF-beta and 1,25(OH)2D3 in dental pulp fibroblasts on the synthesis of SPARC and ALP activity. This interaction could influence the function and differentiation of dental pulp fibroblasts.

Activation of MMP-2 by Porphyromonas gingivalis in human periodontal ligament cells.

It has been reported that matrix metalloproteinase (MMP) produced by host cells plays a major role in periodontal tissue destruction. In addition, secreted virulence factors from Porphyromonas gingivalis can alter MMP secretion and cause activation in host cells that lead to the tissue degradation. In this study, we examine the effects of P. gingivalis supernatant on matrix metalloproteinase-2 (MMP-2) activation in human periodontal ligament (HPDL) cells. Cultures of HPDL cells were treated with P. gingivalis supernatant for 48 h and the level of MMP-2 activation was monitored by gelatin zymography. The profound activation of MMP-2 was seen only in the treated group. The activation of MMP-2 was inhibited by MMP inhibitors phenanthroline and EDTA, but not serine protease or cysteine protease inhibitors. To study the correlation between the expression of membrane-type-1 matrix metalloproteinase (MT1-MMP) and the activation of MMP-2, the level of MT1-MMP was analyzed. The results from reverse-transcription polymerase chain reaction (RT-PCR) and Western analysis indicated that P. gingivalis supernatant up-regulated the expression of MT1-MMP in both transcription and translation levels within 48 h. These results suggest that P. gingivalis supernatant can activate MMP-2 in HPDL cells and the mechanism of activation may involve the increased amount of MT1-MMP. It is possible that the activation of MMP-2 by P. gingivalis plays a role in the process of chronic periodontitis.