Parathyroid hormone-related protein and its receptors: nuclear functions and roles in the renal and cardiovascular systems, the placental trophoblasts and the pancreatic islets.

The cloning of the so-called 'parathyroid hormone-related protein' (PTHrP) in 1987 was the result of a long quest for the factor which, by mimicking the actions of PTH in bone and kidney, is responsible for the hypercalcemic paraneoplastic syndrome, humoral calcemia of malignancy. PTHrP is distinct from PTH in a number of ways. First, PTHrP is the product of a separate gene. Second, with the exception of a short N-terminal region, the structure of PTHrP is not closely related to that of PTH. Third, in contrast to PTH, PTHrP is a paracrine factor expressed throughout the body. Finally, most of the functions of PTHrP have nothing in common with those of PTH. PTHrP is a poly-hormone which comprises a family of distinct peptide hormones arising from post-translational endoproteolytic cleavage of the initial PTHrP translation products. Mature N-terminal, mid-region and C-terminal secretory forms of PTHrP are thus generated, each of them having their own physiologic functions and probably their own receptors. The type 1 PTHrP receptor, binding both PTH(1-34) and PTHrP(1-36), is the only cloned receptor so far. PTHrP is a PTH-like calciotropic hormone, a myorelaxant, a growth factor and a developmental regulatory molecule. The present review reports recent aspects of PTHrP pharmacology and physiology, including: (a) the identification of new peptides and receptors of the PTH/PTHrP system; (b) the recently discovered nuclear functions of PTHrP and the role of PTHrP as an intracrine regulator of cell growth and cell death; (c) the physiological and developmental actions of PTHrP in the cardiovascular and the renal glomerulo-vascular systems; (d) the role of PTHrP as a regulator of pancreatic beta cell growth and functions, and, (e) the interactions of PTHrP and calcium-sensing receptors for the control of the growth of placental trophoblasts. These new advances have contributed to a better understanding of the pathophysiological role of PTHrP, and will help to identify its therapeutic potential in a number of diseases.

The C-terminal region of PTHrP, in addition to the nuclear localization signal, is essential for the intracrine stimulation of proliferation in vascular smooth muscle cells.

PTHrP is secreted by most cell types. In addition to a paracrine/autocrine role, PTHrP has "intracrine" actions, entering the nuclear compartment under the direction of a classic bipartite nuclear localization signal. In vascular smooth muscle cells, nuclear entry stimulates mitogenesis. In the current study, we sought to more precisely define the regions of PTHrP required for the activation of mitogenesis in vascular smooth muscle cells. PTHrP deletion mutants missing large regions [i.e. the signal peptide, N terminus (1--36), mid region (38--86), nuclear localization signal, C terminus (108--139), or combinations of the above] were expressed in A-10 vascular smooth muscle cells. The consequences on nuclear localization and proliferation were examined. Deletion of the nuclear localization signal prevented nuclear entry and slowed proliferation. Deletion of the highly conserved N terminus or mid region had no impact on nuclear localization or on proliferation. Deletion of the C terminus had no deleterious effect on nuclear localization but dramatically reduced proliferation. Thus, the nuclear localization signal is both necessary and sufficient for nuclear localization of PTHrP. In contrast, activation of proliferation in vascular smooth muscle cells requires both an intact nuclear localization signal and an intact C terminus. Whereas the nuclear localization signal is required for nuclear entry, the C terminus may serve a trans-activating function to stimulate mitogenesis once inside the nucleus of vascular smooth muscle cells.

Paradoxical actions of exogenous and endogenous parathyroid hormone-related protein on renal vascular smooth muscle cell proliferation: reversion in the SHR model of genetic hypertension.

In previous studies, added parathyroid hormone-related protein (PTHrP) inhibits whereas transfected PTHrP stimulates the proliferation of A10 aortic smooth muscle cells by nuclear translocation of the peptide. In the present studies, we asked whether these paradoxical trophic actions of PTHrP occur in smooth muscle cells (SMC) cultured from small intrarenal arteries of, and whether they are altered in, 12-wk-old spontaneously hypertensive rats (SHR) as compared to normotensive Wistar-Kyoto (WKY) rats. SHR cells grew faster than WKY cells. PTHrP transcript was increased in SHR-derived cells whereas PTH1 receptor (PTH1R) transcripts were similar in both cell lines. In both strains of cells, stable transfection with human PTHrP(1-139) cDNA did not further induce proliferation, suggesting maximal effect of endogenous PTHrP in wild cells. In contrast, transfection with antisense hPTHrP(1-139) cDNA, which abolished PTHrP mRNA, decreased WKY but increased SHR cell proliferation. Added PTHrP(1-36) (1-100 pM) decreased WKY and increased SHR cell proliferation. Additional studies indicated that the preferential coupling of PTH1-R to G-protein Gi was responsible for the proliferative effect of exogenous PTHrP in SHR cells. Moreover, PTHrP was detected in the nucleolus of a fraction of WKY and SHR renal SMC, in vitro as well as in situ, suggesting that the nucleolar translocation of PTHrP might be involved in the proliferative effects of endogenous PTHrP. In renovascular SMC, added PTHrP is antimitogenic, whereas endogenously produced PTHrP is mitogenic. These paradoxical effects of PTHrP on renovascular SMC proliferation appear to be reversed in the SHR model of genetic hypertension. A new concept emerges from these results, according to which a single molecule may have opposite effects on VSMC proliferation under physiological and pathophysiological conditions.

Culture of vascular smooth muscle cells from small arteries of the rat kidney.

BACKGROUND: In contrast to arterioles, small arteries appear to be the preferential site of renal vascular smooth muscle cell (VSMC) proliferation under pathophysiological conditions. To date, techniques have been described to isolate renal arterioles and to culture VSMCs. The aim of the present study was to develop a method of culturing VSMCs from isolated small arteries of the rat kidney and to characterize their growth as compared with that of aortic VSMCs. METHODS: Renal vascular trees were isolated from kidneys of male Wistar rats by a sieving technique. VSMCs were grown from explants of collagenase-treated renal vascular trees and thoracic aorta. Growth curves and proliferation of renal and aortic VSMCs in response to fetal bovine serum (FBS) were compared by determination of cell number and DNA synthesis, measured as incorporation of 5-bromo-2'-deoxyuridine. RESULTS: Renal vascular trees consisted mainly of small arteries with a diameter of 80 to 400 microm (interlobar and arcuate arteries). As compared with total kidney or renal cortex, alkaline phosphatase activity was decreased by 81%, and vasopressin (10 micromol/L) was unable to stimulate adenylyl cyclase in renal vascular trees, indicating little tubular contamination. A homogenous population of spindle-shaped cells was cultured from renal vascular trees, which grew in a hill-and-valley pattern and stained positively for smooth muscle alpha-actin, according to the characteristics of VSMC phenotype. Renal VSMCs proliferated more slowly than aortic VSMCs and reached the plateau of growth at about half of the cell density of aortic VSMCs. Furthermore, proliferation of renal VSMCs depended more heavily on FBS concentration, since about threefold higher concentrations of FBS were needed for renal VSMCs to multiply at the same rate and to similarly stimulate DNA synthesis as compared with aortic VSMCs. CONCLUSIONS: We present a method to culture renal VSMCs from small arteries of the rat kidney, which possess distinct growth characteristics as compared with aortic VSMCs.

Renovascular parathyroid hormone-related protein in spontaneously hypertensive rats: dilator or trophic factor?

Parathyroid hormone-related protein (PTHrP) is expressed throughout the renovascular system, and it dilates renal vessels, increases renal blood flow and glomerular filtration rate, and stimulates renin release. Mechanical forces and experimental hypertension have been shown to stimulate PTHrP expression in smooth muscles, suggesting a negative feedback control of vascular tone by PTHrP in hypertension. In this study, we compared the impact of a PTHrP receptor antagonist, PTHrP (7-34), and a PTHrP receptor agonist, PTHrP (1-36), on the vascular resistance of perfused kidneys isolated from spontaneously hypertensive rats (SHR) and Wistar Kyoto rats (WKY). Endogenous PTHrP appears not to act as a renal vasodilator in either WKY or SHR. However, the vasodilation following infused PTHrP (1-36) is blunted markedly in SHR, possibly due to desensitization or down-regulation of PTH/PTHrP receptors. Negative feedback control of vascular tone by PTHrP in SHR thus appears unlikely. The results raise the question of whether endogenous renovascular PTHrP behaves rather as a growth factor than as a vasodilator.

Parathyroid hormone-related peptide--a smooth muscle tone and proliferation regulatory protein.

Parathyroid hormone-related protein (PTHrP) appears to play crucial roles in the cardiovascular system. Over the past few years it has become apparent that there is more than one receptor recognizing parathyroid hormone or PTHrP, or both, and that PTHrP is not only a potent vasodilator of vascular smooth muscle cell tone, but is also a regulator of vascular smooth muscle cell proliferation and a secretagogue of renin and vasopressin. Investigators in several laboratories have started to query whether PTHrP intervenes in vascular diseases such as hypertension, (re)stenosis-atherosclerosis and endotoxaemia.