Korean J Ophthalmol.  2004 Dec;18(2):132-140. 10.3341/kjo.2004.18.2.132.

The Effects of Glucose on the Expression of MMP and TIMP in Cultured Retinal Pigment Epithelial Cells

• Lee D
• Kim JS
• Cho HK
• Lee JH

• KMID: 754378
• DOI: http://doi.org/10.3341/kjo.2004.18.2.132

Abstract

This study evaluated the effects of glucose in human retinal pigment epithelial (RPE) cells to investigate the cause of diabetic retinal complications. Human RPE cells were cultured in media containing 5.5 mM, 11.0 mM, and 16.5 mM D-glucose. The present study performed proliferation and migration assays, and conducted western blotting for the protein expression, as well as RT-PCR for the mRNA expression, of MMP-2 and -9, and TIMP-1 and -2. The results of the western blotting analysis showed that increasing glucose concentration significantly increased the expression of MMP-2 and -9, but significantly decreased the expression of TIMP-1 and -2. Moreover, the RT-PCR results indicated significant increases in the mRNA expression of MMP-2 and -9, as well as of TIMP-1 and -2, by raising glucose concentration. This study provides fundamental data for future research on the mechanism of retinal complication in diabetic patients.

Keyword

glucose; human retinal pigment epithelial cell; MMP; TIMP

MeSH Terms

Blotting, Western
Cell Movement
Cell Proliferation
Cells, Cultured
Comparative Study
Dose-Response Relationship, Drug
Glucose/*pharmacology
Humans
In Vitro
Matrix Metalloproteinases/genetics/*metabolism
Pigment Epithelium of Eye/*drug effects/enzymology
RNA, Messenger/metabolism
Research Support, Non-U.S. Gov't
Reverse Transcriptase Polymerase Chain Reaction
Time Factors
Tissue Inhibitor of Metalloproteinases/genetics/*metabolism

Figure

• 

  Fig. 1 Effects of glucose concentration on the proliferation of human retinal pigment epithelial cells. Human retinal pigment cells were exposed to different concentrations of glucose for 2 hours. The cell proliferation was not affected by the glucose concentration. The mean of proliferation is represented by the height of each bar and the standard deviation is depicted on the graph by brackets at the top of each bar. Each bracket extends 1 standard deviation above and 1 standard deviation below the sample mean.

• 

  Fig. 2 Effects of glucose concentration on the proliferation of human retinal pigment epithelial cells. Human retinal pigment epithelial cells were exposed to different concentrations of glucose for 24 hours. The cell proliferation was not affected by the 5.0mM and 11.0mM concentrations of glucose. However, the cells cultured in the 16.5mM concentration of glucose showed significant decreases in the proliferation after 48 hours of incubation. The mean of proliferation is represented by the height of each bar and the standard deviation is depicted on the graph by brackets at the top of each bar. Each bracket extends 1 standard deviation above and 1 standard deviation below the sample mean (*: P <0.05).

• 

  Fig. 3 Effects of glucose concentration on the proliferation of human retinal pigment epithelial cells. Human retinal epithelial cells were exposed to different concentrations of glucose for 14 days. The cell proliferation was significantly affected by 11.0 mM and 16.5 mM concentrations of glucose. The mean of proliferation is represented by the height of each bar and the standard deviation is depicted on the graph by brackets at the top of each bar. Each bracket extends 1 standard deviation above and 1 standard deviation below the sample mean (*: P <0.05).

• 

  Fig. 4 Effects of increasing glucose concentration on the migration of human retinal pigment epithelial cells. The migration of human retinal pigment epithelial cells was not affected in the 2 hour-exposed group. It was significantly affected by the 11.0 mM and the 16.5 mM concentrations of glucose in the 24 hour- and 14 day-exposed groups. The mean of migration is represented by the height of each bar and the standard deviation is depicted on the graph by brackets at the top of each bar. Each bracket extends 1 standard deviation above and 1 standard deviation below the sample mean (*: P <0.05).

• 

  Fig. 5 Western blot analysis of MMP-2 and -9 secreted by retinal pigment epithelial cells. As the glucose concentration increased, the secretions of MMP-2 and -9 increased in the 24 hour-and 14 dayexposed groups.

• 

  Fig. 6 Western blot analysis of TIMP-1 and -2 secreted by retinal pigment epithelial cells. The expression of TIMP-1 and -2 was not affected in the 2 hour-exposed group. As the glucose concentration increased, the secretions of TIMP-1 and -2 decreased in the 24 hour-and 14 day-exposed groups.

• 

  Fig. 7 Analysis of MMP-2 and -9 genes out of cultured human retinal pigment epithelial cells that were produced using reverse transcription polymerase chain reaction. The gene expressions of MMP-2 were not increased, but of -9 were increased by raising glucose concentration, especially at higher concentration.

• 

  Fig. 8 Analysis of TIMP-1 and -2 genes out of cultured human retinal pigment epithelial cells that were produced using reverse transcription polymerase chain reaction. The gene expressions of TIMP-1 and -2 were increased by raising glucose concentration, especially at higher concentration.

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