J Korean Acad Conserv Dent.  2004 Sep;29(5):470-478. 10.5395/JKACD.2004.29.5.470.

MMP-1 and TIMP-1 production in MG-63 cells stimulated with Prevotella nigrescens lipopolysaccharide

• Yang WK
• Kim MR
• Shon WJ
• Lee IB
• Cho BH
• Um CM
• Son HH

• KMID: 1987127
• DOI: http://doi.org/10.5395/JKACD.2004.29.5.470

Abstract

The purpose of this study is to monitor the secretion of matrix metalloproteinase-1 (MMP-1) and tissue inhibitor of metalloproteinase-1 (TIMP-1) produced by human osteosarcoma cell line (MG63) stimulated with Prevotella nigrescens lipopolysaccharides (LPS), and to compare the level of secretion before and after the treatment of calcium hydroxide on P. nigrescens LPS. LPS was extracted and purified from anaerobically cultured P. nigrescens. MG63 cells were stimulated by the LPS (0, 1, 10 microg/ml) or LPS (10 microg/ml) pretreated with 12.5 mg/ml of Ca(OH)2 for 3 days. Total RNA was isolated from the cell, and real-time quantitative polymerase chain reaction (PCR) was performed for quantification of MMP-1 and TIMP-1. The results were as follows. 1. MMP-1 mRNA expression at 48 hr was highly increased by stimulation with P. nigrescens LPS. The increase was dose-dependent. 2. When stimulated with 1 microg/ml of LPS, TIMP-1 mRNA expression was highly increased at 24 hr and 48 hr. However, TIMP-1 expression was suppressed at higher concentration (10 microg/ml). 3. When P. nigrescens LPS was pretreated with Ca(OH)2, MMP-1 and TIMP-1 gene expression was downregulated. The results of this study suggest that transcriptional regulation of MMP-1 and TIMP-1 by P. nigrescens LPS could be one of the important mechanisms in bone resorption of periapical inflammation. The result of calcium hydroxide on MMP-1 and TIMP-1 gene expression suppression shows that calcium hydroxide detoxified bacterial LPS and thus should be used the medication of choice for intracanal dressings in root canal infected with black-pigmented bacteria.

Keyword

MMP-1; TIMP-1; P. nigrescens; LPS; Ca(OH)2; MG63

MeSH Terms

Bacteria
Bandages
Bone Resorption
Calcium Hydroxide
Cell Line
Dental Pulp Cavity
Gene Expression
Humans
Inflammation
Lipopolysaccharides
Matrix Metalloproteinase 1
Osteosarcoma
Polymerase Chain Reaction
Prevotella nigrescens*
Prevotella*
RNA
RNA, Messenger
Tissue Inhibitor of Metalloproteinase-1*
Calcium Hydroxide
Lipopolysaccharides
Matrix Metalloproteinase 1
RNA
RNA, Messenger
Tissue Inhibitor of Metalloproteinase-1

Figure

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  Figure 1 Amplification plots of real time PCR for MMP-1. Dashed line (--x--) indicates the threshold cycle.

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  Figure 2 Amplification plots of real time PCR for TIMP-1. Dashed line (--x--) indicates the threshold cycle.

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  Figure 3 MMP-1 gene expression relative to GAPDH at different LPS concentrations (µg/ml) and different periods of time.

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  Figure 4 TIMP-1 gene expression relative to GAPDH at different LPS concentrations (µg/ml) and different periods of time.

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  Figure 5 MMP-1 gene expression relative to GAPDH in osteoblasts stimulated with 10 µg/ml of P. nigrescens LPS pretreated with Ca(OH)2 at different periods of time.

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  Figure 6 TIMP-1 gene expression relative to GAPDH in osteoblasts stimulated with 10 µg/ml of P. nigrescens LPS pretreated with Ca(OH)2 at different periods of time.

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