Delayed apoptosis and modulation of phospholipase D activity by plasmid containing mammalian cDNA in human neutrophils.
Phospholipase D (PLD) has been reported to have an anti-apoptotic role in neutrophils. This study examined the effects of plasmids containing the cDNA of PLD on the apoptosis of neutrophils. The apoptotic rate of neutrophils treated with the pCDNA3.1 plasmid was similar to that of the untreated cells after 24 h culture. However, the addition of pCDNA3.1 containing the cDNA of either human PLD1 (pCDNA3.1-PLD1) or -PLD2 (pCDNA3.1-PLD2) to the culture media with or without transfection reagent significantly decreased the rate of spontaneous apoptosis but not Fas-stimulated apoptosis and the decreased apoptosis was blocked by 1-butanol. pCDNA3.1-PLD blocked the cleavage of procaspase-3 and -8. The phorbol myristate acetate stimulated the PLD activities of pCDNA3.1-PLD-treated neutrophils but did not stimulate the activities of untreated or pCDNA3.1-treated neutrophils. The level of the PLD1 protein was higher in the cultured neutrophils with pCDNA3.1-PLD than with the media or pCDNA3.1. The spontaneous apoptosis of neutrophils was inhibited and the PLD1 expression level was increased by the linearized or promoterless forms of pCDNA3.1-PLD1 and the plasmids containing the cDNA of the enhanced green fluorescent protein (pEGFP) and EGFP-PLD1. These results suggest that the plasmids containing mammalian cDNA inhibit the spontaneous apoptosis of neutrophils and modulate PLD.
Lee
Sun-Young
SY
Department of Biochemistry, College of Medicine and Medical Research Center for Cancer Molecular Therapy, Dong-A University, Busan 602-714, Republic of Korea.
Kim
Ja-Woong
JW
Jin
Jun-O
JO
Song
Min-Gyu
MG
Park
Joo-In
JI
Min
Do Sik
do S
Kwak
Jong-Young
JY
D017209:Apoptosis / Q000187:drug effects / Q000502:physiology*; D053148:Caspase 3; D020169:Caspases / Q000378:metabolism; D018076:DNA, Complementary; D004789:Enzyme Activation; D049452:Green Fluorescent Proteins / Q000235:genetics; D006801:Humans; D009504:Neutrophils / Q000166:cytology*; D010739:Phospholipase D / Q000235:genetics* / Q000378:metabolism*; D010957:Plasmids; D019014:fas Receptor / Q000502:physiology
D018076:DNA, Complementary; D019014:fas Receptor; D049452:Green Fluorescent Proteins; D010739:Phospholipase D; C505407:CASP3 protein, human; D053148:Caspase 3; D020169:Caspases