The effect of a single nucleotide polymorphism in the matrix metalloproteinase-1 (MMP-1) promoter on force-induced MMP-1 expression in human periodontal ligament cells.

Matrix metalloproteinase-1 (MMP-1) 1G/2G (-1,607) polymorphisms have been identified and shown to influence the transcription of the MMP-1 gene. In order to compare the expression of MMP-1 with different MMP-1 gene promoter alleles after force loading, human periodontal ligament (PDL) cells were cultured and genotyped into three alleles by polymerase chain reaction and restriction endonuclease cleavage. The three genotypes of PDL cells were centrifuged and the expression of MMP-1 mRNA and protein were determined by reverse transcription-polymerase chain reaction and enzyme-linked immunosorbent assay. The results showed that centrifugal force upregulated the expression of both MMP-1 mRNA and protein in all three genotypes of PDL cells. The induction of MMP-1 by force was significantly greater in cells with a 2G/2G genotype or a 1G/2G genotype than in cells homozygous for the 1G allele. The MMP-1 mRNA and protein levels were significantly higher for cells with the 2G allele than for cells with the 1G/2G allele or the 1G allele. These results suggest that a single nucleotide polymorphism in the -1,607 bp MMP-1 promoter region might be associated with the difference observed in the endogenous expression of MMP-1 in PDL cells under mechanical force.

Dentin matrix protein 1 and phosphate homeostasis are critical for postnatal pulp, dentin and enamel formation.

Deletion or mutation of dentin matrix protein 1 (DMP1) leads to hypophosphatemic rickets and defects within the dentin. However, it is largely unknown if this pathological change is a direct role of DMP1 or an indirect role of phosphate (Pi) or both. It has also been previously shown that Klotho-deficient mice, which displayed a high Pi level due to a failure of Pi excretion, causes mild defects in the dentinal structure. This study was to address the distinct roles of DMP1 and Pi homeostasis in cell differentiation, apoptosis and mineralization of dentin and enamel. Our working hypothesis was that a stable Pi homeostasis is critical for postnatal tooth formation, and that DMP1 has an antiapoptotic role in both amelogenesis and dentinogenesis. To test this hypothesis, Dmp1-null (Dmp1(-/-)), Klotho-deficient (kl/kl), Dmp1/Klotho-double-deficient (Dmp1(-/-)/kl/kl) and wild-type (WT) mice were killed at the age of 6 weeks. Combinations of X-ray, microcomputed tomography (µCT), scanning electron microscopy (SEM), histology, apoptosis and immunohistochemical methods were used for characterization of dentin, enamel and pulp structures in these mutant mice. Our results showed that Dmp1(-/-) (a low Pi level) or kl/kl (a high Pi level) mice displayed mild dentin defects such as thin dentin and a reduction of dentin tubules. Neither deficient mouse line exhibited any apparent changes in enamel or pulp structure. However, the double-deficient mice (a high Pi level) displayed severe defects in dentin and enamel structures, including loss of dentinal tubules and enamel prisms, as well as unexpected ectopic ossification within the pulp root canal. TUNEL assay showed a sharp increase in apoptotic cells in ameloblasts and odontoblasts. Based on the above findings, we conclude that DMP1 has a protective role for odontoblasts and ameloblasts in a pro-apoptotic environment (a high Pi level).

[Protective effect of baicalin on experimental periodontitis in rats and its possible mechanisms].

OBJECTIVE: To investigate the effect of baicalin on the experimental periodontitis in rats, as well as the expression of MMP-1, MMP-2, MMP-9. METHODS: Twenty-seven adult male Sprague-Dawley rats were divided into three groups, with 9 rats in each group. A nylon thread was placed around the lower first molars of rats, which were sacrificed after 7 days. Baicalin (200 mg/kg) was administered to the experimental group by oral gavage, starting one day before the induction of periodontitis. The negative control group received vehicle (0.5% carboxymethylcellulose) alone. The blank control group did not get induction of periodontitis. The alveolar bone loss (ABL) and the area fraction (AA% ) occupied by collagen fibers were assessed. MMP-1, MMP-2 and MMP-9 protein expressions in the gingiva were detected by immunohistochemistry. RESULTS: Baicalin treatment significantly decreased ABL compared with the negative control group (P = 0.009). AA% of collagen fibers was significantly higher in baicalin-treated group than in the negative control group (P = 0.047). Baicalin treatment significantly down-regulated the protein expression for MMP-1 (P = 0.023) and MMP-9 (P = 0.042) and decreased the expression for MMP-2 (P = 0.099) compared with the negative control group. CONCLUSIONS: Baicalin protects against tissue damage in ligature-induced periodontitis in rats, which might be mediated in part by its inhibitory effect on the expression of MMP-1, MMP-2 and MMP-9.

[Preparation and feasibility of superparamagnetic dextran iron oxide nanoparticles as gene carrier].

BACKGROUND & OBJECTIVE: Application of magnetic nano- particles as gene carrier in gene therapy of tumor has developed quickly. To obtain a new type non-viral gene introduction and therapy system,which is convenient,and can drive target gene to express highly and stably,this study was designed to explore the preparation of superparamagnetic dextran iron oxide nanoparticles(SDION),and the feasibility of SDION used as gene carrier in vitro. METHODS: SDION were prepared by chemical co-precipitation,separated by gel filtration chromatography on Sephacryl S-300HR,and centrifugation techniques,characterized by transmission electron microscopy,laser scattering system,and vibrating sample magnetometer signal processor. The green fluorescent protein-C2 (GFP-C2) plasmid was used as target gene. SDION-GFP- C2 compounds were synthesized by oxidation-reduction reaction. The connection rate of SDION and GFP-C2 was analyzed by agarose electrophoresis,and evaluated by measuring concentration of GFP in the supernatant after centrifugation. Liposome transfection was used as control,the efficiencies of SDION and liposome in transferring GFP gene into human bladder cancer BIU-87 cells were evaluated under fluorescence microscope in vitro. RESULTS: The diameter of SDION ranged from 3 nm to 8 nm, the effective diameter was 59.2 nm, and the saturation magnetization was 0.23 emu/g. After oxidized by sodium periodate of 10 mmol/L,and deoxidized by sodium hydride boron of 0.5 mol/L, SDION could connect with GFP in maximum degree,the transfection efficiency of SDION as gene carrier was about 45%, even higher than that of liposome (about 30%). CONCLUSION: SDION could connect with GFP plasmid by oxidation- reduction reaction,and success to transfer GFP gene into human bladder cancer BIU-87 cells in vitro.

[Effects of baicalin on the expression of pro-MMP-1 and MMP-3 in human gingival fibroblasts and periodontal ligament cells].

OBJECTIVE: To investigate the influence of baicalin on the IL-1beta induced pro-MMP-1 in HGF and the effects of baicalin on MMP-3 expression in periodontal ligament cells (PDLCs). METHODS: The amount of secreted pro-MMP-1 and MMP-3 expression was detected by ELISA and cell immunochemistry. RESULTS: (1) The amount of secreted pro-MMP-1 (3.333 +/- 0.123) microg/L increased significantly following 1 microg/L of IL-1beta, compared with control group (1.960 +/- 0.180) microg/L. Addition of baicalin to cell culture medium for 1 hour following IL-1beta decreased pro-MMP-1 secretion in a dose-dependent manner in the range of 10 approximately 1,000 microg/L. (2) 1 microg/L IL-1beta could significantly stimulate the synthesis and secretion of MMP-3 in PDLCs. (3) The baicalin could not interfere the synthesis of MMP-3, but could inhibit the release of MMP-3 from PDLCs. CONCLUSIONS: Baicalin could inhibit the secretion of pro-MMP-1 and MMP-3 expression in IL-1beta induced HGF and PDLCs, which suggests that baicalin may play an important role in preventing and treating periodontal disease.