Effects of 5-azaC on methylation pattern of the perforin promoter of the perforin gene in normal human T cells / 中南大学学报(医学版)
Journal of Central South University(Medical Sciences)
;
(12): 843-847, 2006.
Artigo
em Chinês
| WPRIM
| ID: wpr-813589
ABSTRACT
OBJECTIVE@#To investigate the effects of DNA hypomethylation on mRNA and protein expression of perforin promotor in T cells.@*METHODS@#T cells were isolated from the peripheral venous blood of healthy donors by density gradient centrifugation. CD4(+) and CD8(+) subsets were isolated using Miltenyi beads and protocols provided by the manufacturer. Where indicated the T cells were stimulated with PHA for 24 h, then treated with 5-azaC for an additional 72 h. Genomic DNA, mRNA, and protein were isolated from untreated and 5-azaC-treated T cells. Purified DNA was treated with sodium bisulfite, the desired sequences were amplified in sequential fragments using nested PCR. The amplified fragments were cloned into bacteria DH5 alpha and 5 independent clones for each of the amplified fragments were sequenced. The expression of perforin was determined using real time RT-PCR and Western blot.@*RESULTS@#The perforin mRNA and protein in the CD4(+) and CD8(+) subsets treated with 5-azaC were significantly higher than those in the untreated subsets (P<0.05). The results of bisulfite genomic sequencing showed that the methylation of perforin promotor was significantly reduced in the treated cells compared with the untreated cells (P<0.05).@*CONCLUSION@#The mRNA and protein expression of perforin significantly increases in the CD4(+) and CD8(+) T cells treated with 5-azaC,which is associated with DNA hypomethylation of perforin promoter in T cells.
Texto completo:
DisponíveL
Índice:
WPRIM (Pacífico Ocidental)
Assunto principal:
Farmacologia
/
Azacitidina
/
Células Cultivadas
/
Subpopulações de Linfócitos T
/
Regiões Promotoras Genéticas
/
Metilação de DNA
/
Perforina
/
Genética
/
Metabolismo
Limite:
Adulto
/
Humanos
Idioma:
Chinês
Revista:
Journal of Central South University(Medical Sciences)
Ano de publicação:
2006
Tipo de documento:
Artigo
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