ADP-Ribosyl Cyclase in Rat Vascular Smooth Muscle Cells

Abstract

Abstract-We investigated whether ADP-ribosyl cyclase (ADPR-cyclase) in rat vascular smooth muscle cells (VSMCs) has enzymatic properties that differ from the well-characterized CD38-antigen ADPR-cyclase, expressed in HL-60 cells. ADPR-cyclase from VSMCs, but not CD38 ADPR-cyclase from HL-60 cells, was inhibited by gangliosides (10 mol/L) GT 1B , GD 1 , and GM 3 . Preincubation of membranes from CD38 HL-60 cells, but not from VSMCs, with anti-CD38 antibodies increased ADPR-cyclase activity; CD38 antigen was detected both in VSMCs and in HL-60 cells. ADPR-cyclase in VSMC membranes was more sensitive than CD38 HL-60 ADPR-cyclase to inactivation by N-endoglycosidase F and to thermal inactivation at 45°C. The specific activity of ADPR-cyclase in membranes from VSMCs was Ͼ20-fold higher than in membranes from CD38 HL-60 cells. Most importantly, VSMC ADPR-cyclase was inhibited by Zn 2ϩ and Cu 2ϩ ions; the inhibition by Zn 2ϩ was dose dependent, noncompetitive, and reversible by EDTA. In contrast, Zn 2ϩ stimulated the activity of CD38 HL-60 ADPR-cyclase and other known types of ADPR-cyclases. Retinoids act either via the nuclear receptor retinoic acid receptor or retinoid X receptor, including all-trans retinoic acid (atRA), and panagonist 9-cis-retinoic acid-upregulated VSMC ADPR-cyclase; the stimulatory effect of atRA was blocked by actinomycin D and cycloheximide. 1,25(OH) 2 -Vitamin D 3 (calciferol) stimulated VSMC ADPR-cyclase dose dependently at subnanomolar concentrations (ED 50 Х56 pmol/L). Oral administration of atRA to rats resulted in an increase of ADPR-cyclase activity in aorta (Хϩ60%) and, to a lesser degree, in myocardium of left ventricle (ϩ18%), but atRA had no effect on ADPR-cyclases in lungs, spleen, intestinal smooth muscle, skeletal muscle, liver, or testis. Administration of 3,5,3Ј-triiodothyronine (T 3 ) to rats resulted in an increase of ADPR-cyclase activity in aorta (Хϩ89%), but not in liver or brain. We conclude the following: (1) ADPR-cyclase in VSMCs has enzymatic properties distinct from "classic" CD38 ADPR-cyclase, especially sensitivity to inhibition by Zn 2ϩ and Cu 2ϩ ; (2) ADPR-cyclase in VSMCs is upregulated by various retinoids, calcitriol, and T 3 in vitro; and (3) administration of atRA and T 3 increases ADPR-cyclase in aorta in vivo. We suggest that the cADPR signaling system plays an important role in the regulation of VSMC functions in response to steroid superfamily hormones.

Ryanodine receptor subtype 2 encodes Ca2+ oscillations activated by acetylcholine via the M2 muscarinic receptor/cADP-ribose signalling pathway in duodenum myocytes

“…Activation of the cADPR pathway by ACh in duodenum myocytes is shown by: (1) inhibition of Ca 2+ oscillations by application of the cADPR competitive antagonist (8Br-cADPR), (2) inhibition of ACh-induced Ca 2+ oscillations by inhibitors of ADP-ribosyl cyclase (ZnCl 2 , anti-CD38 antibody) and (3) detection of ADP-ribosyl cyclase activity by fluorescence experiments as the enzyme cyclizes NGD+ (non fluorescent) to produce cGDPR, a fluorescent compound (Graeff et al, 1994). This method has been used successfully in microsomes and cellular homogenates from bovine chromaffin cells (Morita et al, 1997), rat vascular myocytes (de Toledo et al, 2000) and human myometrium (Chini et al, 2002). We were able to detect ADP-ribosyl cyclase activity in single permeabilized cells under the same conditions that we used for Ca 2+ measurements.…”

“…To confirm the involvement of cADPR, ADP-ribosyl cyclase activity was inhibited in permeabilized cells using ZnCl 2 (de Toledo et al, 2000), high concentrations of NGD+ and an anti-CD38 antibody (Sternfeld et al, 2003). Application of 2 mM ZnCl 2 decreased significantly the amplitude of ACh-induced Ca 2+ responses and suppressed Ca 2+ oscillations (Fig.…”

Modulation of store-operated Ca2+ entry by cyclic-ADP-ribose

“…The components of this pathway are present and functional in myometrial, vascular and airway smooth muscle cells (18)(19)(20)(21)(22)(23)(24). One important observation is the fact that the intracellular Ca 2+ transients induced by oxytocin are dependent on both the CD38/cADPR signaling pathway and extracellular Ca 2+ in human myometrial cells.…”

“…We have shown that cADPR is important for agonist-stimulated Ca 2+ transients in smooth muscle cells (18)(19)(20)(21). Since RyR activity appears to be important for the gating of the store-operated Ca 2+ entry channels and A, Cell extracts were prepared in RIPA buffer, proteins adjusted to 200 µg/vial and 2 µg rabbit polyclonal anti-TRPC antibodies added overnight.…”

Kidney ADP-Ribosyl Cyclase Inhibitors as a Therapeutic Tool for Diabetic Nephropathy

“…Mounting evidence has indicated that ADPR-cyclase(s) other than CD38 may exist in the kidney, brain, and the heart (40,45), including various cells (30,(45)(46)(47). The first clues to the existence of novel ADPR-cyclase(s) emerged from experiments of the comparison of tissue cADPR levels in CD38 wild type and knockout mice (40).…”