Differential effects of somatostatin and angiopeptin on cell proliferation

Abstract

Somatostatin (SRIF) exerts antiproliferative effects, and angiopeptin (an sst2/sst5 receptor‐selective analogue) has recently been evaluated in clinical trials for the prophylaxis of restenosis following coronary angioplasty. Using an in vitro model of cell growth we have examined the effects of SRIF and angiopeptin on cell proliferation in CHO‐K1 cells stably transfected with the human or rat recombinant sst2 or sst5 receptor and compared these with their effects on rat aortic vascular smooth muscle cells (VSMC) expressing endogenous somatostatin receptors. In CHO‐K1 cells, expressing either human or rat recombinant sst2 or sst5 receptors, or in rat aortic VSMC, SRIF and angiopeptin (0.1–1000 nM) had no effect on basal re‐growth of cells into a denuded area of a previously confluent monolayer. In contrast, basic fibroblast growth factor (bFGF, 10 ng ml−1) stimulated re‐growth of these cells. SRIF (0.1–1000 nM) caused a concentration‐dependent inhibition of the bFGF‐stimulated re‐growth in CHO‐K1 cells expressing human sst2 (h sst2) or sst5 (h sst5) receptors (pIC50=8.05±0.03 and 8.56±0.12, respectively). In contrast, angiopeptin (0.1–1000 nM) acted as a partial agonist at the h sst2 receptor (44.6±2.7% inhibition of the bFGF‐stimulated re‐growth at 100 nM; pIC50=8.69±0.25) but was devoid of any agonist activity at the h sst5 receptor. In CHO‐K1 cells stably expressing rat recombinant sst2 (r sst2) or sst5 (r sst5) receptors, SRIF (0.1–1000 nM) was able to inhibit the bFGF‐stimulated re‐growth (pIC50=7.98±24 and 8.50±0.12, respectively). Angiopeptin (0.1–1000 nM) caused a concentration‐dependent inhibition of bFGF‐stimulated re‐growth at the r sst2 receptor (pIC50=8.08±0.24) but acted as a partial agonist at the r sst5 receptor (maximum response=57.7±3.6% inhibition of bFGF‐stimulated re‐growth at 100 nM; pIC50=8.60±0.16). Although angiopeptin was inactive as an agonist at the h sst5 receptor, 100 nM angiopeptin potently antagonized the SRIF‐induced inhibition of proliferation in CHO h sst5 (estimated pKB=10.4±0.3). 5‐Hydroxytryptamine (0.1 nM–10 μM) also inhibited bFGF‐stimulated re‐growth (pIC50=8.36±0.11) and angiopeptin had no effect on this response (pKB<7). SRIF (0.1–1000 nM) caused a concentration‐dependent (pIC50=8.04±0.08) inhibition of bFGF‐stimulated re‐growth in VSMC, whereas angiopeptin displayed weak agonist activity, only inhibiting bFGF‐stimulated re‐growth at concentrations greater than 100 nM. Angiopeptin (100 nM) caused a rightward displacement of the concentration‐effect curve to SRIF with an estimated pKB value of 7.70±0.12. These findings suggest that the low intrinsic activity of angiopeptin at the h sst2 receptor, combined with its lack of agonist activity at the h sst5 receptor, may explain the poor clinical efficacy of angiopeptin in trials for coronary artery restenosis, which contrasts with encouraging data found in equivalent in vivo animal studies.

Rat somatostatin sst2(a) and sst2(b) receptor isoforms mediate opposite effects on cell proliferation

“…However, studies performed in our laboratory have shown that the two isoforms display similar ligand binding pro®les and transduction characteristics, functionally coupling to the inhibition of adenylate cyclase and activation of increased extracellular acidi®cation (Schindler et al, 1998c). Although it is well known that SRIF can inhibit cell proliferation through the rat recombinant sst 2(a) receptor (Alderton et al, 1998), the eect upon cell growth and proliferation of the rat sst 2(b) splice variant has not yet been examined. In the present study we describe the unexpected eects of SRIF, as well as the synthetic peptide angiopeptin (BIM-23014; under clinical investigation for the treatment of acromegaly and inhibition of tumour growth; see Gillespie et al, 1998), on cell proliferation mediated through the rat recombinant sst 2(b) receptor expressed in CHO-K1 cells.…”

“…Results We have previously shown in CHOsst 2(a) cells that SRIF has no eect on basal re-growth but can inhibit bFGFstimulated re-growth after partial denudation of a con¯uent cell monolayer (see Figure 1a,b; Alderton et al, 1998). In marked contrast to these eects, basal re-growth in CHOsst 2(b) cells (10.6+0.4%) was stimulated by SRIF (0.1 ± 1000 nM; Figure 1a) and angiopeptin (0.1 ± 1000 nM) in a concentration-dependent manner with the percentage regrowth at 1 mM not reaching a fully de®ned maximum (pEC 50 values of 7.79+0.20 and 7.94+0.29 estimated by curve ®tting; maxima of 20.2+0.7% and 18.8+0.8%, respectively).…”

“…In addition, SRIF (0.1 ± 1000 nM) was unable to aect either )-stimulated re-growth at 24 h, respectively. Data for the eects of SRIF on basal and bFGF-stimulated re-growth in CHOsst 2(a) cells are from Alderton et al (1998) and are provided for comparison with equivalent data in CHOsst 2(b) cells. In addition, the eects of pertussis-toxin pretreatment (PTx; 100 ng ml 71 ; 18 h) were determined on basal, bFGF-and SRIF-mediated inhibition or stimulation of re-growth in (c) CHOsst 2(b) cells and (d) CHOsst 2(a) cells.…”

“…Image analysis was carried out with a Seescan TM semi-automated, image analysis machine (Seescan, Cambridge, U.K.). For each coverslip ®ve ®elds of view selected at random were analysed and data expressed as the mean percentage of the area recovered following the period of re-growth as previously described (Alderton et al, 1998). All values are means+s.e.mean from a minimum of three experiments with four replicates per test group.…”

Cortistatin Inhibits Migration and Proliferation of Human Vascular Smooth Muscle Cells and Decreases Neointimal Formation on Carotid Artery Ligation

“…9 Ssts are widespread distributed in many cell types, including SMCs, and regulate multiple signal transduction pathways that lead to inhibition of cell proliferation. [9][10][11] Although various evidences indicate that somatostatin inhibits proliferation of SMCs in vitro, 7,11 there is still controversy regarding the failure of somatostatin or its synthetic analogues to regulate SMC proliferation in vivo. [12][13][14] Moreover, there is no actual evidence for the ability of somatostatin to regulate SMC migration.…”