MGST1
Microsomal glutathione S-transferase 1 (EC 2.5.1.18) (Microsomal GST-1) (Microsomal GST-I) [GST12] [MGST]
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Despite the known longevity of human hematopoietic stem and progenitor cells (HSC), numerous functional impairments of these cells can be observed in an age-dependent manner. However, the molecular alterations associated with aging of HSC are largely unknown. Therefore, we scrutinized gene expression patterns of HSC from newborn, young and old healthy donors. CD34 HSC were isolated via immuno-magnetic separation and evaluated by FACS analysis. We performed cDNA macroarray analyses on a first set of CD34 samples (n=13). We found the genes encoding KU-antigen 70 kD (KU70), microsomal glutathione S-transferase 1 (MGST1) and BCL2-interacting killer (BIK) to possess age-related mRNA expression levels. KU70 is a DNA repair gene and part of the DNA-dependent protein kinase (DNA-PK) complex. Its expression was negatively correlated with donor age showing highest expression levels in newborn, 2.6-fold lower levels in young and 6.3-fold lower levels in old donors. The transcription levels of MGST1, a gene protecting against oxidative stress, progressively increased with age. Expression was lowest in newborn, 2.6-fold higher in young and 4.3-fold higher in old donors. BIK is a proapoptotic gene and its expression was positively correlated with donor age: lowest in newborn, 1.8-fold higher in young and 4.1-fold higher in old donors. These findings were confirmed with an independent, second set of CD34 samples (n=16) by means of quantitative real-time RT-PCR. Elucidation of age-dependent molecular alterations in healthy HSC facilitate a better understanding of functional impairments in hematopoiesis and may become valuable for anti-aging drug development and the emerging field of regenerative medicine.
MeSH Terms
- Adult
- Adult Stem Cells
- Age Factors
- Aged
- Aged, 80 and over
- Aging
- Antigens, CD34
- Antigens, Nuclear
- Apoptosis Regulatory Proteins
- Cellular Senescence
- Cluster Analysis
- DNA-Binding Proteins
- Fetal Blood
- Gene Expression Profiling
- Glutathione Transferase
- Hematopoietic Stem Cells
- Humans
- Infant, Newborn
- Ku Autoantigen
- Membrane Proteins
- Middle Aged
- Mitochondrial Proteins
- Oligonucleotide Array Sequence Analysis
- Polymerase Chain Reaction
- RNA, Messenger
- Reproducibility of Results
- Transcription, Genetic
High oxygen tension, exposure to light, and the biochemical events of vision generate significant oxidative stress in the retina and the retinal pigment epithelium (RPE). Understanding the mechanisms and basis of susceptibility to progressive retinal diseases involving oxidative damage such as age-related macular degeneration (AMD) remains a major challenge. Here microsomal glutathione S-transferase (MGST1) is shown to be a dominant, highly expressed enzyme in bovine and mouse RPE microsomes that displays significant reduction activity toward synthetic peroxides, oxidized RPE lipids, and oxidized retinoids. This enzymatic reduction activity (GPx) can be partially neutralized with a monoclonal anti-MGST1 antibody developed in this study. MGST1-transfected HEK293 cells exhibited greater viability (70 /- 4% survival) compared with untransfected control cells (46 /- 4% survival) when challenged with 20 microM H(2)O(2), and greater viability of MGST1-transfected cells following challenge with oxidized docosahexaenoic acid was also observed. Cultured ARPE19 cells transfected with silencing MGST1 siRNAs exhibited lower expression of MGST1 (12% and 26% of the controls) and significantly lower GPx activity (44 /- 13%) and, thus, were more susceptible to oxidative damage. Immunoblotting revealed that the in vivo expression of MGST1 in mouse RPE decreases 3-4-fold with age, to trace levels in 18-month-old mice. GPx activity in the RPE was also found to be reduced in 12-month-old mice to approximately 67%. These results support an important protective function for MGST1 against oxidative insult in the RPE that decreases with age and suggest that this enzyme may play a role in the development of age-related diseases such as AMD.
MeSH Terms
- Aging
- Amino Acid Sequence
- Animals
- Antibodies, Monoclonal
- Cattle
- Cell Line
- Cell Survival
- Cross Reactions
- Enzyme Activation
- Gene Expression Regulation
- Glutathione Transferase
- Mice
- Microsomes
- Molecular Sequence Data
- Molecular Weight
- Oxidative Stress
- Pigment Epithelium of Eye
- Transfection