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Methylglyoxal induces apoptosis mediated by reactive oxygen species in bovine retinal pericytes.

Kim J, Son JW, Lee JA, Oh YS, Shinn SH - J. Korean Med. Sci. (2004)

Bottom Line: NF-kappaB activation and increased caspase-3 activity were detected.Apoptosis was also inhibited by the caspase-3 inhibitor, Z-DEVD-fmk, or the NF-kappaB inhibitor, pyrrolidine dithiocarbamate.These data suggest that elevated MGO levels observed in diabetes may cause apoptosis in bovine retinal pericytes through an oxidative stress mechanism and suggests that the nuclear activation of NF-kappaB are involved in the apoptotic process.

View Article: PubMed Central - PubMed

Affiliation: Division of Endocrinology and Metabolism, Department of Internal Medicine, College of Medicine, Chung-Ang University, Seoul, Korea. jtkim@cau.ac.kr

ABSTRACT
One of the histopathologic hallmarks of early diabetic retinopathy is the loss of pericytes. Evidences suggest that the pericyte loss in vivo is mediated by apoptosis. However, the underlying cause of pericyte apoptosis is not fully understood. This study investigated the influence of methylglyoxal (MGO), a reactive alpha-dicarbonyl compound of glucose metabolism, on apoptotic cell death in bovine retinal pericytes. Analysis of internucleosomal DNA fragmentation by ELISA showed that MGO (200 to 800 microM) induced apoptosis in a concentration-dependent manner. Intracellular reactive oxygen species were generated earlier and the antioxidant, N-acetyl cysteine, inhibited the MGO-induced apoptosis. NF-kappaB activation and increased caspase-3 activity were detected. Apoptosis was also inhibited by the caspase-3 inhibitor, Z-DEVD-fmk, or the NF-kappaB inhibitor, pyrrolidine dithiocarbamate. These data suggest that elevated MGO levels observed in diabetes may cause apoptosis in bovine retinal pericytes through an oxidative stress mechanism and suggests that the nuclear activation of NF-kappaB are involved in the apoptotic process.

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Effects of MGO on NF-κB binding. Pericytes were treated with MGO (800 µM) for 6 hr. Nuclear extracts from the treated and untreated control cells were isolated and used in an EMSA with 32P-labeled NF-κB oligonucleotide as a probe. The arrow indicates the NF-κB binding complex. Competitor, 100-fold molar excess of unlabeled NF-κB probe. Data are representative results from three separate experiments.
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Figure 5: Effects of MGO on NF-κB binding. Pericytes were treated with MGO (800 µM) for 6 hr. Nuclear extracts from the treated and untreated control cells were isolated and used in an EMSA with 32P-labeled NF-κB oligonucleotide as a probe. The arrow indicates the NF-κB binding complex. Competitor, 100-fold molar excess of unlabeled NF-κB probe. Data are representative results from three separate experiments.

Mentions: Subcellular immunolocalization of the p65 subunit of NF-κB demonstrated a cytoplasmic distribution in controls (Fig. 4A, B). However, after incubation with MGO, nuclear translocation of NF-κB was evident (Fig. 4C, D). The EMSA demonstrated more NF-κB binding was evident after 6 hr of MGO treatment (Fig. 5).


Methylglyoxal induces apoptosis mediated by reactive oxygen species in bovine retinal pericytes.

Kim J, Son JW, Lee JA, Oh YS, Shinn SH - J. Korean Med. Sci. (2004)

Effects of MGO on NF-κB binding. Pericytes were treated with MGO (800 µM) for 6 hr. Nuclear extracts from the treated and untreated control cells were isolated and used in an EMSA with 32P-labeled NF-κB oligonucleotide as a probe. The arrow indicates the NF-κB binding complex. Competitor, 100-fold molar excess of unlabeled NF-κB probe. Data are representative results from three separate experiments.
© Copyright Policy - open-access
Related In: Results  -  Collection

License
Show All Figures
getmorefigures.php?uid=PMC2822272&req=5

Figure 5: Effects of MGO on NF-κB binding. Pericytes were treated with MGO (800 µM) for 6 hr. Nuclear extracts from the treated and untreated control cells were isolated and used in an EMSA with 32P-labeled NF-κB oligonucleotide as a probe. The arrow indicates the NF-κB binding complex. Competitor, 100-fold molar excess of unlabeled NF-κB probe. Data are representative results from three separate experiments.
Mentions: Subcellular immunolocalization of the p65 subunit of NF-κB demonstrated a cytoplasmic distribution in controls (Fig. 4A, B). However, after incubation with MGO, nuclear translocation of NF-κB was evident (Fig. 4C, D). The EMSA demonstrated more NF-κB binding was evident after 6 hr of MGO treatment (Fig. 5).

Bottom Line: NF-kappaB activation and increased caspase-3 activity were detected.Apoptosis was also inhibited by the caspase-3 inhibitor, Z-DEVD-fmk, or the NF-kappaB inhibitor, pyrrolidine dithiocarbamate.These data suggest that elevated MGO levels observed in diabetes may cause apoptosis in bovine retinal pericytes through an oxidative stress mechanism and suggests that the nuclear activation of NF-kappaB are involved in the apoptotic process.

View Article: PubMed Central - PubMed

Affiliation: Division of Endocrinology and Metabolism, Department of Internal Medicine, College of Medicine, Chung-Ang University, Seoul, Korea. jtkim@cau.ac.kr

ABSTRACT
One of the histopathologic hallmarks of early diabetic retinopathy is the loss of pericytes. Evidences suggest that the pericyte loss in vivo is mediated by apoptosis. However, the underlying cause of pericyte apoptosis is not fully understood. This study investigated the influence of methylglyoxal (MGO), a reactive alpha-dicarbonyl compound of glucose metabolism, on apoptotic cell death in bovine retinal pericytes. Analysis of internucleosomal DNA fragmentation by ELISA showed that MGO (200 to 800 microM) induced apoptosis in a concentration-dependent manner. Intracellular reactive oxygen species were generated earlier and the antioxidant, N-acetyl cysteine, inhibited the MGO-induced apoptosis. NF-kappaB activation and increased caspase-3 activity were detected. Apoptosis was also inhibited by the caspase-3 inhibitor, Z-DEVD-fmk, or the NF-kappaB inhibitor, pyrrolidine dithiocarbamate. These data suggest that elevated MGO levels observed in diabetes may cause apoptosis in bovine retinal pericytes through an oxidative stress mechanism and suggests that the nuclear activation of NF-kappaB are involved in the apoptotic process.

Show MeSH
Related in: MedlinePlus