Activation and up-regulation of phospholipase D expression by lipopolysaccharide in human peripheral T cells.
In a previous study, we showed that bacterial LPS activates protein kinase C (PKC) and causes an intracellular pH (pHi) increase, but does not elevate intracellular calcium ([Ca2+]i) in human peripherial T cells. Hence this study aimed to investigate whether the activation of PKC was resulted from phospholipase D (PLD) catalysis by LPS. The activity of PLD was measured by the production of 3H-phosphatidylethanol from phosphatidic acid (PA), and the expression of PLD or IL-2 Ralpha was determined by reverse transcription-polymerase chain reaction (RT-PCR) analysis. Enzyme-linked immunosorbent assay (ELISA) was used to analyze IL-2 and IL-4. Phytohemagglutinin (PHA), and phorbol 12-myristate 13-acetate (PMA) were used as controls. Our results indicated that (1) LPS-stimulated pHi elevation was PKC dependent; (2) After 30 min stimulation, LPS increased PLD activity via a measured production of 3H-phosphatidylethanol from phosphatidic acid and the initiation of PLD1a mRNA expression started; (2) LPS stimulated IL-2 R expression but not IL-2 and IL-4 secretion. Our findings suggested that the stimulation of PLD activity and its mRNA expression by LPS might be required for IL-2 R expression and a sustained PKC dependent pHi elevation but not for the secretion of IL-2 or IL-4 in human T cells. This indicated that LPS might enhance T cell adaptive immunity to resist Gram-negative bacterial infection.
Chien
Eileen Jea
EJ
Department of Physiology, School of Medicine, National Yang-Ming University, Taipei 11221, Taiwan, Republic of China. eileen@ym.edu.tw
Chen
Chien-Ching
CC
Chien
Chau-Heng
CH
Yeh
Tzu-Pei
TP
Lu
Li-Ming
LM
D000328:Adult; D015536:Down-Regulation / Q000187:drug effects; D004797:Enzyme-Linked Immunosorbent Assay; D015971:Gene Expression Regulation, Enzymologic / Q000187:drug effects; D020404:Glycerophospholipids / Q000378:metabolism; D006801:Humans; D006863:Hydrogen-Ion Concentration; D007376:Interleukin-2 / Q000378:metabolism; D015847:Interleukin-4 / Q000378:metabolism; D008070:Lipopolysaccharides / Q000494:pharmacology*; D008297:Male; D010712:Phosphatidic Acids / Q000378:metabolism; D010739:Phospholipase D / Q000096:biosynthesis*; D010835:Phytohemagglutinins / Q000494:pharmacology; D015375:Receptors, Interleukin-2 / Q000096:biosynthesis; D020133:Reverse Transcriptase Polymerase Chain Reaction; D013601:T-Lymphocytes / Q000187:drug effects / Q000201:enzymology*; D013755:Tetradecanoylphorbol Acetate / Q000494:pharmacology; D013936:Thymidine / Q000378:metabolism; D015854:Up-Regulation / Q000187:drug effects
D020404:Glycerophospholipids; D007376:Interleukin-2; D008070:Lipopolysaccharides; D010712:Phosphatidic Acids; D010835:Phytohemagglutinins; D015375:Receptors, Interleukin-2; C051521:phosphatidylethanol; D015847:Interleukin-4; D010739:Phospholipase D; C106801:phospholipase D1; D013755:Tetradecanoylphorbol Acetate; D013936:Thymidine