Table 1.
List of primer couples for PLC isoforms.
Fig 1.
Characterization of phospholipase C β isoforms in native arteries.
A. The presence of mRNA for phospholipase C (PLC) β isoforms was determined in mesenteric arteries (MA), pulmonary arteries (PA) and middle cerebral arteries (MCA) by PCR. Typical agarose gel electrophoresis of the PCR products showed the expression profile in different vascular beds. Brain and blood were used as positive control tissues. n = 3. B. Quantitative real time PCR analysis of mRNA expression levels of PLCβ isoforms in MA and freshly isolated endothelial cells (ECs) from MA, PA and MCA. Bar graphs show the expression profile of PLCβ1 (a), β2 (b), β3 (c) and β4 (d) isoforms in MAECs, PAECs, MCAECs, MA and blood as control for β2. n = 3. * P<0.05 between MAECs and MCAECs; # P<0.05 between PAECs and MCAECs; † P<0.05 between MCAECs and MAs; ‡ P<0.05 between control tissue and MA. C. (a) Representative immunoblots of murine MA and brain that were obtained using the primary antibody anti-PLC β3 (Abcam #ab52199). GAPDH was used as reference protein. Relevant molecular weight markers are indicated on the left. n = 3. (b) Intracellular distribution of PLCβ3 immunoreactivity in ECs. (Left) Typical image showing labelling of PLC β3 in red and nuclei in blue; scale = 10 μm. (Right) Labelling of PLC β3 (red) overlay with internal elastic lamina (IEL; green) where voids correspond to potential myoendothelial projections; nucleus in blue; scale = 10 μm; n = 4.
Fig 2.
Characterization of phospholipase C γ isoforms in native arteries.
A. The presence or mRNA for phospholipase C (PLC) γ isoforms was determined in mesenteric arteries (MA), pulmonary arteries (PA) and middle cerebral arteries (MCA) by PCR. Typical agarose gel electrophoresis of the PCR products showed the expression profile in the different vascular beds and brain was used as control tissue. n = 3. B. Quantitative real time PCR analysis of mRNA expression levels of PLCγ isoforms in MA and freshly isolated endothelial cells (ECs) from MA, PA and MCA. Bar graphs show the expression profile of PLCγ1 (a) and γ2 (b) isoforms in MAECs, PAECs, MCAECs and MA. n = 3. C. (a) Representative immunoblots of murine MA and brain that were analyzed using the primary antibody anti-PLC γ2 (Abcam #ab18983). GAPDH was used as reference protein. Relevant molecular weight markers are indicated on the left. n = 3. (b) Intracellular distribution of PLCγ2 immunoreactivity in ECs. (Left) Typical image showing labelling of PLCγ2 in red and nuclei in blue; scale = 10 μm. (Right) Labelling of PLCγ2 (red) overlay with internal elastic lamina (IEL; green) where voids correspond to potential myoendothelial projections; nucleus in blue; scale = 10 μm; n = 4.
Fig 3.
Characterization of phospholipase C δ isoforms in native arteries.
A. The presence of mRNA for phospholipase C (PLC) δ isoforms was determined in mesenteric arteries (MA), pulmonary arteries (PA) and middle cerebral arteries (MCA) by PCR. Typical agarose gel electrophoresis of the PCR products showed the expression profile of PLCδ isoforms in the different vascular beds and brain was used as control tissue. n = 3. B. Quantitative real time PCR analysis of mRNA expression levels of PLCδ isoforms in MA and freshly isolated endothelial cells (ECs) from MA, PA and MCA. Bar graphs showed the expression profile of PLCδ1 (a), δ3 (b) and δ4 (c) isoforms in MAECs, PAECs, MCAECs and MA. n = 3. * P<0.05 between MAECs and MCAECs; † P<0.05 between MCAECs and MA. C. (a) Representative immunoblots of murine MA and brain that were analyzed using the primary antibody anti-PLCδ1 (Abcam #ab154610). GAPDH was used as reference protein. Relevant molecular weight markers are indicated on the left. n = 3. (b) Intracellular distribution of PLCδ1 immunoreactivity in ECs. (Left) Typical image showing labelling of PLCδ1 in red and nuclei in blue; scale = 10 μm. (Right) Labelling of PLCδ1 (red) overlay with internal elastic lamina (IEL; green) where voids correspond to potential myoendothelial projections; nucleus in blue; scale = 10 μm; n = 4.
Fig 4.
Characterization of phospholipase C ε, ζ and η isoforms in native arteries.
A. The presence of mRNA for phospholipase C (PLC) ε, ζ, η1 and η2 isoforms was determined in mesenteric arteries (MA), pulmonary arteries (PA) and middle cerebral arteries (MCA) by PCR. Typical agarose gel electrophoresis of the PCR products showed the expression profile of PLCε, ζ, η1 and η2 isoforms in the different vascular beds and brain or testis were used as control tissue. n = 3. B. Quantitative real time PCR analysis of mRNA expression levels of PLCε, ζ, η1 and η2 isoforms in MA and freshly isolated endothelial cells (ECs) from MA, PA and MCA. Bar graphs show the expression of PLCε (a), ζ (b), η1 (c) and η2 (d) isoforms in MAECs, PAECs, MCAECs, MA and testis as positive control for ζ. n = 3. * P<0.05 between MAECs and MCAECs; # P<0.05 between PAECs and MCAECs; † P<0.05 between MCAECs and MA.
Table 2.
Summary of mRNA expression for phospholipase C isoforms.