Enhanced proliferation and altered calcium handling in RGS2-deficient vascular smooth muscle cells.
BACKGROUND
Regulator of G-protein signaling-2 (RGS2) inhibits Gq-mediated regulation of Ca(2+) signalling in vascular smooth muscle cells (VSMC).


OBJECTIVE
RGS2 knockout (RGS2KO) mice are hypertensive and show arteriolar remodeling. VSMC proliferation modulates intracellular Ca(2+) concentration [Ca(2+)]i. RGS2 involvement in VSMC proliferation had not been examined.


METHODS
Thymidine incorporation and 3-(4,5-dimethylthiazol-2-yl)-2,5-diphenyltetrazolium bromide (MTT) conversion assays measured cell proliferation. Fura-2 ratiometric imaging quantified [Ca(2+)]i before and after UTP and thapsigargin. [(3)H]-labeled inositol was used for phosphoinositide hydrolysis. Quantitative RT-PCR and confocal immunofluorescence of select Ca(2+) transporters was performed in primary aortic VSMC.


CONCLUSIONS
Platelet-derived growth factor (PDGF) increased S-phase entry and proliferation in VSMC from RGS2KO mice to a greater extent than in VSMC from wild-type (WT) controls. Consistent with differential PDGF-induced changes in Ca(2+) homeostasis, RGS2KO VSMC showed lower resting [Ca(2+)]i but higher thapsigargin-induced [Ca(2+)]i as compared with WT. RGS2KO VSMC expressed lower mRNA levels of plasma membrane Ca(2+) ATPase-4 (PMCA4) and Na(+) Ca(2+) Exchanger (NCX), but higher levels of sarco-endoplasmic reticulum Ca(2+) ATPase-2 (SERCA2). Western blot and immunofluorescence revealed similar differences in PMCA4 and SERCA2 protein, while levels of NCX protein were not reduced in RGS2KO VSMC. Consistent with decreased Ca(2+) efflux activity, (45)Ca-extrusion rates were lower in RGS2KO VSMC. These differences were reversed by the PMCA inhibitor La(3+), but not by replacing extracellular Na(+) with choline, implicating differences in the activity of PMCA and not NCX.


CONCLUSIONS
RGS2-deficient VSMC exhibit higher rates of proliferation and coordinate plasticity of Ca(2+)-handling mechanisms in response to PDGF stimulation.
Division of Experimental Therapeutics, Toronto General Research Institute , University Health Network, Toronto, Ontario , Canada and.
Abdul
A
Momen
Talat
T
Afroze
Al-Muktafi
AM
Sadi
Amir
A
Khoshbin
Hangjun
H
Zhang
Jaehyun
J
Choi
Steven
S
Gu
Syed H
SH
Zaidi
Scott P
SP
Heximer
Mansoor
M
Husain
D000818:Animals; D002118:Calcium / Q000378:metabolism*; D020013:Calcium Signaling / Q000187:drug effects / Q000502:physiology*; D049109:Cell Proliferation / Q000187:drug effects / Q000502:physiology; D002478:Cells, Cultured; D051379:Mice; D018345:Mice, Knockout; D009131:Muscle, Smooth, Vascular / Q000166:cytology* / Q000187:drug effects / Q000502:physiology*; D032389:Myocytes, Smooth Muscle / Q000166:cytology* / Q000187:drug effects / Q000502:physiology*; D010982:Platelet-Derived Growth Factor / Q000494:pharmacology; D020710:RGS Proteins / Q000235:genetics / Q000378:metabolism*
D010982:Platelet-Derived Growth Factor; D020710:RGS Proteins; C105345:Rgs2 protein, mouse; D002118:Calcium
Ca2+; PMCA4; RGS2; proliferation; vascular smooth muscle cell