Ultrastructural localization of tyrosine hydroxylase in human peripheral blood mononuclear cells: effect of stimulation with phytohaemagglutinin.
Using immunocytochemistry coupled to fluorescence and electron microscopy, we investigated the expression and ultrastructural localization of tyrosine hydroxylase (TH, EC 1.14.16.2), the rate-limiting enzyme in the biosynthesis of catecholamines, in human peripheral blood mononuclear cells (PBMCs), with PC12 cells as positive controls. In unstimulated PBMCs, TH-specific immunoreactivity was localized to the plasma membrane. However, after stimulation with the polyclonal mitogen phytohaemagglutinin (PHA), TH immunoreactivity was almost completely localized to electron-dense cytoplasmic granules, which resembled those found in PC12. TH-positive granules, however, were larger (300-500 nm) than in PC12 cells (100-200 nm). Flow cytometry analysis of TH expression showed about 46-50% positive cells in unstimulated PBMCs and in PHA-stimulated PBMCs in the G0/G1 phase of the cell cycle, but more than 80% positive cells in PHA-stimulated PBMCs in the S+G2/M phase. In agreement with previous observations, PHA stimulation also induced de novo expression of TH mRNA as well as increased intracellular catecholamine content, suggesting the occurrence of TH upregulation at the level of both gene expression and enzyme activity. The ultrastructural localization of TH in human PBMCs seems therefore regulated by cell stimulation and related to the functional activity of the enzyme.
Reguzzoni
Marcella
M
Department of Clinical and Biological Sciences, Laboratory of Human Morphology Luigi Cattaneo, University of Insubria, Varese, Italy.
Cosentino
Marco
M
Rasini
Emanuela
E
Marino
Franca
F
Ferrari
Marco
M
Bombelli
Raffaella
R
Congiu
Terenzio
T
Protasoni
Marina
M
Quacci
Daniela
D
Lecchini
Sergio
S
Raspanti
Mario
M
Frigo
Gianmario
G
D006801:Humans; D007150:Immunohistochemistry; D007700:Kinetics; D007963:Leukocytes, Mononuclear / Q000187:drug effects / Q000201:enzymology* / Q000648:ultrastructure; D016253:Microscopy, Immunoelectron; D010835:Phytohemagglutinins / Q000494:pharmacology*; D012333:RNA, Messenger / Q000235:genetics; D020133:Reverse Transcriptase Polymerase Chain Reaction; D014158:Transcription, Genetic; D014446:Tyrosine 3-Monooxygenase / Q000097:blood* / Q000235:genetics / Q000648:ultrastructure
D010835:Phytohemagglutinins; D012333:RNA, Messenger; D014446:Tyrosine 3-Monooxygenase