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PURPOSE: To investigate the effect of hypoxia on the expression of HIF-lα and VEGF in human periodontal ligament cells (hPDLCs) in vitro. METHODS: HPDLCs were cultured in Dulbecco's modified Eagle's medium(DMEM) and subcultured at confluence. Experiments were carried out with the third passage of hPDLCs populations. In the normoxic control group, cells were incubated under normoxic conditions of 20%O2, 5%CO2, 75%N2 for 24 hours and 48 hours, respectively. In the hypoxic group, cells were incubated in a humidified atmosphere of 1%O2, 5%CO2, 94%N2 for 24 hours and 48 hours, respectively. The expression of HIF-1α and VEGF in hPDLCs in vitro was measured using reverse transcription-polymerase chain reaction. SP immunohistochemistry method was performed to localize the distribution of HIF-1α and VEGF in hPDLCs. The data was analyzed by Student's t test, one-way ANOVA and LSD test with SPSS 13.0 software package. RESULTS: There was no significant difference between the hypoxic group and the normoxic control group in the expression of HIF-1α mRNA in hPDLCs. The expression of VEGF mRNA in the hypoxic group of 24 hours and 48 hours was both statistically higher than in the control group (P<0.05). The expression of HIF-1α protein was significantly increased with hypoxia time while the staining of HIF-1α was negative under normoxic condition. The expression of VEGF protein in hypoxic groups was significantly higher than in normoxic control groups. The staining of VEGF gradually enhanced in a time-dependent manner and was weakly positive after 48 hours under normoxic condition. CONCLUSION: Hypoxia could change the metabolic pathway of human periodontal ligament cells by upregulating the expression of HIF-1α and other relevant growth factors. [ABSTRACT FROM AUTHOR]

PURPOSE : To study the effects of epidermal growth factor (EGF) on the secretion of monocyte chemotactic protein-one (MCP-1) in rat dental follicle cells. METHODS : Dental follicle was harvested from 4-6 days postnatal Wistar rats and cultured in α-MEM with 15% FBS. Cells in the third passage were incubated with EGF at different concentrations to choose the best dosage. MTT assays were used to evaluate cell proliferation. Using the suitable concentration, 10ng/ml EGF cells were cultured for 0, 0.5, 1, 3 and 6 hours respectively, and then collected for RT-PCR analysis. The results were analyzed with SPSS13.0 software package. RESULTS: EGF(ng/ml) at 5-10 ng/ml increased the cell proliferation (P<0.05), which reached the peak at 10ng/ml according to MTT assay (P<0.01). 10ng/ml and 3 hours EGF exposure up-regulated the cellular MCP-1 expression dramatically (P<0.01). CONCLUSION: Proper concentration of EGF may promote dental follicle cell proliferation, EGF may up-regulate MCP-1 transcription in the rat dental follicle cells. [ABSTRACT FROM AUTHOR]

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 clays 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 4°C overnight, dehydrated, embedded in paraffin and serially sectioned at 5µm.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 clay 1, but it gradually decreased and was negative at postnatal clay 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. [ABSTRACT FROM AUTHOR]

Purpose: To investigate the effect of hypoxia on the expression of HIF-1alpha and VEGF in human periodontal ligament cells (hPDLCs) in vitro., Methods: HPDLCs were cultured in Dulbecco's modified Eagle's medium (DMEM) and subcultured at confluence. Experiments were carried out with the third passage of hPDLCs populations. In the normoxic control group, cells were incubated under normoxic conditions of 20% O2, 5% CO2, 75% N2 for 24 hours and 48 hours, respectively. In the hypoxic group, cells were incubated in a humidified atmosphere of 1% O2, 5% CO2, 94% N2 for 24 hours and 48 hours, respectively. The expression of HIF-1alpha and VEGF in hPDLCs in vitro was measured using reverse transcription-polymerase chain reaction. SP immunohistochemistry method was performed to localize the distribution of HIF-1alpha and VEGF in hPDLCs. The data was analyzed by Student's t test, one-way ANOVA and LSD test with SPSS 13.0 software package., Results: There was no significant difference between the hypoxic group and the normoxic control group in the expression of HIF-1alpha mRNA in hPDLCs. The expression of VEGF mRNA in the hypoxic group of 24 hours and 48 hours was both statistically higher than in the control group (P<0.05). The expression of HIF-1alpha protein was significantly increased with hypoxia time while the staining of HIF-1alpha was negative under normoxic condition. The expression of VEGF protein in hypoxic groups was significantly higher than in normoxic control groups. The staining of VEGF gradually enhanced in a time-dependent manner and was weakly positive after 48 hours under normoxic condition., Conclusion: Hypoxia could change the metabolic pathway of human periodontal ligament cells by upregulating the expression of HIF-1alpha and other relevant growth factors.

Purpose: To study the effects of epidermal growth factor (EGF) on the secretion of monocyte chemotactic protein-one (MCP-1) in rat dental follicle cells., Methods: Dental follicle was harvested from 4-6 days postnatal Wistar rats and cultured in alpha-MEM with 15% FBS. Cells in the third passage were incubated with EGF at different concentrations to choose the best dosage. MTT assays were used to evaluate cell proliferation. Using the suitable concentration, 10 ng/ml EGF cells were cultured for 0, 0.5, 1, 3 and 6 hours respectively, and then collected for RT-PCR analysis. The results were analyzed with SPSS13.0 software package., Results: EGF(ng/ml) at 5-10 ng/ml increased the cell proliferation (P<0.05), which reached the peak at 10 ng/ml according to MTT assay (P<0.01). 10 ng/ml and 3 hours EGF exposure up-regulated the cellular MCP-1 expression dramatically (P<0.01)., Conclusion: Proper concentration of EGF may promote dental follicle cell proliferation, EGF may up-regulate MCP-1 transcription in the rat dental follicle cells.

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.