Protective effect and related mechanisms of curcumin in rat experimental periodontitis.

BACKGROUND: Curcumin exhibits anti-inflammatory effects and has been suggested as a treatment for inflammatory diseases. The aim of this study was to investigate the effects of curcumin on the lipopolysaccharide induced inflammatory response in rat gingival fibroblasts in vitro and ligation-induced experimental periodontitis in vivo, and to speculate the possible anti-inflammatory mechanism of curcumin. METHODS: The gingival fibroblasts were incubated with different concentrations of curcumin in the absence or presence of lipopolysaccharide (LPS). Concentrations of interleukin-1ß(IL-1ß), tumor necrosis factor-α (TNF-α), osteoprotegerin (OPG) and soluble receptor activator of nuclear factor kappa-B ligand (RANKL) culture supernatants of rat gingival fibroblasts were determined by enzyme linked immunosorbent assay. The nuclear fraction of rat gingival fibroblasts was extracted and nuclear factor kappa-B (NF-κB) activation was assessed by western blotting to elucidate related mechanisms. Curcumin was given every two days by oral gavage. The gingival inflammation and alveolar bone loss between the first and second molars were observed by hematoxylin and eosin staining. Collagen fibers were observed by picro-sirius red staining. Alveolar bone loss was assessed by micro-CT analysis. RESULTS: Curcumin attenuated the production of IL-1ß and TNF-α in rat gingival fibroblasts stimulated by LPS, and inhibited the LPS-induced decrease in OPG/sRANKL ratio and NF-κB activation. Curcumin significantly reduced gingival inflammation and modulated collagen fiber and alveolar bone loss in vivo. CONCLUSIONS: curcumin modulates inflammatory activity in rat periodontitis by inhibiting NF-κB activation and decreasing the OPG/sRANKL ratio induced by LPS.

Effect of hypoxia on the expression of RANKL/OPG in human periodontal ligament cells in vitro.

To investigate the impact of hypoxia on the expression of receptor activator of NF-kB ligand (RANKL) and osteoprotegerin (OPG) in human periodontal ligament cells (hPDLCs) in vitro. hPDLCs were incubated in a hypoxic atmosphere of 2% O2, 5% CO2, 94% N2 at 37°C for 6, 12, 24 and 48 h. After that, cell proliferation assay was determined using CCK-8 technique. SP immunocytochemistry method was performed to trace the expression of hypoxia-inducible factor 1 alpha (HIF-1α) in hPDLCs. The expression levels of RANKL and OPG were investigated using real-time PCR and ELISA. As a control, the cells were incubated at normoxic conditions of 20% O2, 5% CO2, 75% N2. All results were analyzed using one-way ANOVA at a significant level of P=0.05. OPG mRNA and protein levels were down-regulated meanwhile RANKL mRNA and soluble RANKL (sRANKL) protein levels were up-regulated after stimulated by hypoxia. The relative RANKL/OPG expression ratios were increased in both mRNA and protein levels. The expression of RANKL mRNA and sRANKL protein levels was enhanced significantly (P<0.05) under the hypoxia conditions at 12 h, 24 h and 48 h while OPG mRNA and protein were reduced significantly (P<0.05) at 12 h, 24 h and 48 h. Hypoxia can affect the expression of RANKL and OPG in hPDLCs, which constitute an important pathogenic event in the alveolar bone resorption. Lack of oxygen in periodontal tissue may accelerate the development of periodontitis.

[Proliferation, apoptosis and expression of tissue non-specific alkaline phosphatase in cells of stratum intermedium during amelogenesis in postnatal stage of BALB/c mouse].

PURPOSE: To study the apoptosis and expression of proliferating cell nuclear antigen(PCNA),tissue non-specific alkaline phosphatase(TNSALP) of stratum intermedium cell in postnatal stage of BALB/c mouse. METHODS: Fifty six BALB/c mice in different developmental days were sacrificed and their bilateral mandibular first molar germs with surrounding alveolar bone were taken out, then the tissues were fixed with 4% paraformaldehyde at 4degrees centigrade overnight, dehydrated, embedded in paraffin and serially sectioned at 5mum.Tdt-mediated Dutp nick end labeling (TUNEL) and immunohistochemical assay were adopted to determine the tissue distribution and cellular localization of bcl-2, PCNA and TNSALP,respectively. Image-pro plus 6.0 software was used to evaluate the histological sections and the data was analyzed by Students't test,one-way ANOVA and SNK-q test with SPSS 13.0 software package. RESULTS: The expression of PCNA in stratum intermedium was higher than ameloblast at postnatal day 1, but it gradually decreased and was negative at postnatal day 7. The apoptosis of stratum intermedium cells increased during enamel development.The expression of TNSALP appeared at postnatal day 3 and was most obvious at postnatal day 5 in stratum intermedium, then gradually decreased. However, the expression of TNSALP appeared at postnatal day 7 in ameloblasts, then gradually increased. CONCLUSIONS: There is a significant linear correlation between stratum intermedium cell and ameloblast in apoptosis and expression of PCNA and TNSALP.It is suggested that the stratum intermedium participate in the differentiation of ameloblast and formation of enamel.

[Hertwig's epithelial root sheath, bone matrix proteins, and initiation of cementogenesis in mouse teeth].

PURPOSE: The aim of this study was to analyze the association between Hertwig's epithelial root sheath (HERS), noncollagenous matrix proteins, and cementogenesis. METHODS: Eighteen BALB/c postnatal mice were divided into five groups according to the developing stages. The mouse teeth were examined at the onset of root development by light and transmission electron microscopy. PV two-step immunohistochemical method was used to detect the expression of proliferating cell nuclear antigen(PCNA), OPN and BSP during the root formation. RESULTS: Hertwig's epithelial root sheath, which derived from the apical extensions of inner and outer enamel epithelium, was formed at 5-day mouse teeth and expressed PCNA positively. During the fuse of inner and outer enamel epithelium, some cells, which displayed the cytologic features of protein synthesis and secretion, could be found between the two epithelium cells. Desmosomal cell junctions were observed. The positive expressions of OPN were observed at the beginning of HERS. An accumulation of OPN could also be observed at the dentin-cementum junction during cementogenesis, but no positive BSP signals were seen at the onset of HERS. CONCLUSION: At onset of HERS, the inner enamel epithelium, outer enamel epithelium embrace several cells with developed cytoplasmic organelles, which may derive from stratum intermedium or stellate reticulum, and form a structure like enamel organ, which may be associated with initial cementogenesis. OPN, the non collagenous matrix proteins of cementum, may also take part in the formation of the dentin-cementum junction.

[Reaction of epithelial cell rests of malassez to tooth emergence and occlusal function reaction of epithelial cell rests of Malassez to tooth emergence and occlusal function].

UNLABELLED: OBJECTIVE To observe the morphology and proliferation of epithelial cell rests of Malassez (ECRM) during tooth emergence and occlusal function, and to evaluate its roles. METHODS: Cytokeratin 14 (CK14) was applied as special marker of ECRM cells. The morphology and distribution of ECRM were examined by light and transmission electron microscopy. PV two-step immunohistochemical method was used to detect the expression of CK14 and proliferating cell nuclear antigen (PCNA) in ECRM. RESULTS: ECRM experienced instinct morphological changes during tooth emergence and occlusal function. They were observed as network of epithelial cells labeled by CK14, especially in furcation level regions of mouse molars and active cell proliferation during occlusion found period. Cell apoptosis was observed in many ECRM by transmission electron microscopy during late stage of the progess. CONCLUSION: ECRM may not only an accidental left-over of early embryonic development but rather play significant roles in occlusion found period.

[Influence of smoking on apoptosis in human gingival epithelium].

PURPOSE: Cigarette smoking was a major risk factor for oral hygiene and periodontal status. The study was to investigate the relationship between different exposure to cigarette smoke and apoptosis in stratum spinosum and stratum basale cells of human gingival epithelium. METHODS: 47 smoking patients with crown lengthening surgery or extraction of the impacted teeth were enrolled. It comprised 41 males and 6 females. Their age ranged from 18 to 29 years old (average age: 21.4). According to smoke history (SH), average cigarettes per day (ACD) and total smoke number (TSM), 47 smokers were divided into group I (SH<5a or ACD<10 and TSM<15 thousand, n=19), group II (5a< or = SH or 10< or = ACD and 15 thousand < or =TSM, n=28). Another 10 non-smoking patients were chosen as control. Apoptosis was identified by the terminal deoxy-transferase (TdT)-mediated dUTP-biotin nick end labeling (TUNEL) method. Differences between the two groups were analyzed using Student-Newman-Keuls (SNK).The criteria for statistical significance was accepted at the probability level P<0.05. RESULTS: There was increased apoptosis in stratum spinosumin and stratum basale cells of smokers compared to that of non-smokers. Apoptosis in stratum spinosumin cells was significantly different between smokers and non-smokers (P<0.01), between group I and group II (P<0.01). CONCLUSION: It is conclude that exposure to cigarette smoke can increase apoptosis in stratum spinosumin cells of human gingival epithelium, which may influence its normal metabolism and protein secretion.