Using platelet-rich plasma to treat jumper's knees: Exploring the effect of a second closely-timed infiltration.

OBJECTIVES: Some clinical series have evaluated the effect of platelet-rich plasma (PRP) in the treatment of proximal patellar tendinopathy. Although it is possible that a single infiltrative administration may prove to be an effective treatment for this indication, most of the existing studies evaluated the effects of two or three successive infiltrations. The aim of this study was to evaluate whether two infiltrations of PRP proves more effective than a single treatment. DESIGN: Prospective, randomized and comparative study of level 2. METHODS: Twenty patients suffering from chronic proximal patellar tendinopathy were enrolled into the study and split into two randomized groups (one or two infiltrations of PRP, respectively). The 3-month follow-up evaluation consisted of VAS, IKDC and VISA-P scores, along with algometer, isokinetic and ultrasounds evaluations. After 1 year, subjects were contacted to define their functional evolution. RESULTS: The concentration of the PRP used for each infiltration was similar in both groups, and contained no red or white cells. Results revealed no difference in treatment efficacy between the groups. CONCLUSIONS: The comparison between one or two infiltrations of PRP did not reveal any difference between the two groups at short to mid term. A second closely-timed infiltration of PRP to treat proximal patellar tendinopathies is not necessary to improve the efficacy of this treatment in the short term.

Ca2+-agonistic effect of a T-type Ca-channel blocker mibefradil (Ro 40-5967).

Here we report that a Ca2+ antagonist mibefradil (Ro 40-5967) which has been shown to be a selective inhibitor of T-type calcium channels increases free calcium concentration ([Ca2+]i) in the cytoplasm of cultured smooth muscle cells isolated from porcine coronary artery. Smooth muscle cells were loaded with Fura 2 and a videoimage system was used to follow the [Ca2+]i responses. It was shown that at a concentration of 1 nM mibefradil induced a transient [Ca2+]i elevation in individual cells and at a concentration of 100 nM this compound stimulated almost all the cells in monolayer. The [Ca2+]i response did not change with the further increase of the mibefradil concentration up to 10 microM. The half-maximal effect was observed at 10 nM. The increase in [Ca2+]i strongly depended on the presence of Ca in the extracellular medium. Calcium antagonists belonging to three different classes--verapamil (phenylalkylamines), diltiazem (benzothiazepines) and amlodipin (dihydropyridines) neither suppressed the mibefradil effect nor mimicked it. These data indicate that mibefradil increased [Ca2+]i acting via a distinct receptor site. We suggest that these receptors are coupled to calcium channels of plasma membrane.

Cardiovascular care with the new T-type calcium channel antagonist: possible role of attendant sympathetic nervous system inhibition.

DIFFERENCES AMONG TYPES OF CALCIUM ANTAGONISTS: Calcium antagonists lower blood pressure, relieve angina pectoris and improve chronic heart failure, primarily through peripheral and coronary vasodilation. The debate as to whether short-acting, long-lasting (L)-channel calcium influx antagonists of the 1,4-dihydropyridine type might be involved in excess cardiac mortality has raised new controversies with respect to the cardiac morbidity and mortality outcome for all calcium antagonists. Different pharmacodynamic effects (short-acting vasodilation inducing pulsatile sympathetic reflex stimulation) may explain differences in outcome with calcium antagonist therapies. Calcium antagonists also differ in their direct effects on the sympathetic nervous system, and on its long endocrine arm, the renin-angiotensin-aldosterone system. These differential effects relate to the cardiac conduction system and ventricular ectopic activity, to cardiac and vascular remodelling and hypertrophy, and perhaps also to the development of hypertension. L-CHANNEL CALCIUM ANTAGONISTS: Depending on their pharmacodynamic characteristics, L-channel calcium antagonists of the dihydropyridine, verapamil or diltiazem type reflexly activate the sympathetic nervous system and blunt beta-adrenoceptor-mediated calcium influx, thus eliciting negative inotropy and activation of the renin-angiotensin system. Both verapamil and diltiazem slow down pacemaker activity and atrioventricular conduction. MIBEFRADIL: The new-class T-channel blocker mibefradil exhibits vascular selectivity and induces peripheral and coronary vasodilation. There is no reflex sympathetic activation and no negative inotropic effect. It increases coronary blood flow without increasing oxygen consumption and causes a slight slowing of the heart rate, thereby inducing diastolic relaxation. The latter improves subendocardial and small artery perfusion. There is a sympatholytic effect, owing to T-channel expression in neurones, sinoatrial and atrioventricular nodes and Purkinje fibres. In experimental models, ventricular ectopic activity is reduced with mibefradil. The renin-angiotensin-aldosterone system and endothelin effects are blunted by T-channel inhibition. These and other factors reduce smooth muscle cell proliferation, hypertrophy and matrix deposition. T-type calcium channel inhibition, over and above its antihypertensive and anti-ischaemic effects, and afterload-reducing effects in chronic heart failure, offers the potential for a cardiovascular protective benefit, which may be critically related to interference with the sympathetic nervous system.

Ligand selectivity of 105 kDa and 130 kDa lipoprotein-binding proteins in vascular-smooth-muscle-cell membranes is unique.

Using ligand blotting techniques, with low-density lipoprotein (LDL) as ligand, we have previously described the existence of atypical lipoprotein-binding proteins (105 kDa and 130 kDa) in membranes from human aortic medical tissue. The present study demonstrates that these proteins are also present in membranes from cultured human (aortic and mesenteric) and rat (aortic) vascular smooth-muscle cells (VSMCs). To assess the relationship of 105 and 130 kDa lipoprotein-binding proteins to known lipoprotein receptors, ligand binding specificity was studied. We tested effects of substances known to antagonize ligand binding to either the LDL [apolipoprotein B,E (apo B,E)] receptor (dextran sulphate, heparin, pentosan polysulphate, protamine, spermine, histone), the scavenger receptor (dextran sulphate, fucoidin), the very-low-density-lipoprotein (VLDL) receptor [receptor-associated protein (RAP)], or LDL receptor-related protein (RAP, alpha 2-macroglobulin, lipoprotein lipase, exotoxin-A). None of these substances, with the exception of dextran sulphate, influenced binding of LDL to either 105 or 130 kDa proteins. Sodium oleate or oleic acid, known stimuli for the lipoprotein binding activity of the lipolysis-stimulated receptor, were also without effect. LDL binding to 105 and 130 kDa proteins was inhibited by anti-LDL (apo B) antibodies. LDL and VLDL bound to 105 and 130 kDa proteins with similar affinities (approximately 50 micrograms/ml). The unique ligand selectivity of 105 and 130 kDa proteins supports the existence of a novel lipoprotein-binding protein that is distinct from all other currently identified LDL receptor family members. The similar ligand selectivity of 105 and 130 kDa proteins suggests that they may represent variant forms of an atypical lipoprotein-binding protein.

Prevention of neointima formation by mibefradil after vascular injury in rats: comparison with ACE inhibition.

Cilazapril, an angiotensin-converting enzyme inhibitor, and mibefradil, a selective T-type voltage-operated calcium channel blocker, have been shown to prevent neointima formation after vascular injury. The goal of the present study was to evaluate the mechanism of action of both drugs. For this purpose, the influence of the renin angiotensin system on the effects of mibefradil (30 mg/kg po) and cilazapril (10 mg/kg po) on neointima formation after carotid injury were evaluated in normotensive rats (normal renin angiotensin system) and DOCA hypertensive rats (suppressed renin angiotensin system). In addition, in order to differentiate an effect on cell migration or cell proliferation, both drugs were given either before or after the smooth muscle migration phase. Finally, cilazapril and mibefradil were given in combination. In normotensive rats, mibefradil and cilazapril decreased neointima formation, resulting in neointima/media ratios of 38% (p < 0.05) and 53% (p < 0.01), respectively. However, in DOCA hypertensive rats, mibefradil was active, with a reduction of the neointima/media ratio by 63% (p < 0.001), whereas cilazapril reduced it only slightly (19%) and not significantly. In addition, cilazapril was active only when treatment started before the migration phase (63%, reduction in neointima/media ratio, p < 0.001) but not when started thereafter (13% reduction in neointima/media ratio, n.s.). In contrast, treatment with mibefradil was also active when started after the migration phase (51% reduction in neointima/ media ratio, p < 0.001 when treatment started 1 day before balloon injury and 41%, p < 0.01 when treatment started 5 days after balloon injury). The combination of both drugs was additive (67% reduction in neointima/media ratio, p < 0.001 vs. control). These experiments clearly show that mibefradil and cilazapril have a different mechanism of action after vascular injury. Mibefradil most likely prevents the proliferation of smooth muscle cells. In contrast, cilazapril most likely inhibits the migration of smooth muscle cells. These two different mechanisms of action explain why the effects of both drugs are additive.

Low- and high-density lipoproteins as mitogenic factors for vascular smooth muscle cells: individual, additive and synergistic effects.

The mitogenic activities of low (LDL)- and high (HDL)-density lipoproteins have been examined in cultures of human vascular smooth muscle cells (VSMC). LDL and HDL3 dose-dependently (EC50 values approximately 50 micrograms/ml) stimulated DNA and protein synthesis ([3H]-thymidine and [3H]-leucine incorporation, respectively) in the absence of exogenously added mitogens. The synthetic responses of VSMC to combinations of LDL and HDL3 were additive, indicating that each lipoprotein mediates discrete effects. LDL or HDL3 promoted VSMC proliferation under strict mitogen-free conditions, but this growth response was not sustained. VSMC exposed to combinations of lipoproteins (either LDL or HDL3) and growth factors (either PDGF-BB, EGF, bFGF or IGF) exhibited synergistic DNA synthesis responses. In the combined presence of PDGF-BB and either LDL or HDL3, VSMC proliferation was sustained. Anionized lipoprotein preparations (oxidized, acetylated, carbamylated or malonimylated) also stimulated DNA and protein synthesis. Since the antioxidant beta-hydroxylated toluene did not block the effect of native LDL on DNA synthesis, and fucoidin, a specific competitor for the 'scavenger' receptor, did not inhibit oxidized LDL-induced DNA synthesis, activation of mitogenic signals by lipoproteins does not depend on lipid peroxidation. Rather, the apparent intrinsic mitogenic potential of lipoproteins may depend upon their direct activation of replication-coupled signal transduction systems.

Mibefradil prevents neointima formation after vascular injury in rats. Possible role of the blockade of the T-type voltage-operated calcium channel.

Mibefradil is a novel calcium antagonist that is selective for the T-type voltage-operated calcium channel rather than the L type. Because T-type calcium channels are present on rapidly proliferating cells and mediate the increase of intracellular calcium induced by some growth factors, such as platelet-derived growth factor, we hypothesized that the blockade of T channels could prevent the excessive smooth muscle cell proliferation that occurs in conditions such as vascular injury. To test this hypothesis, we evaluated in rats the effects of mibefradil (which blocks both L- and T-type channels) on neointima formation after vascular injury, and we compared them with those of equihypotensive doses of amlodipine and verapamil (which block only L-type channels). Mibefradil (30 mg/kg) decreased the area of neointima formed 14 days after balloon injury by 54% (P < .001). In contrast, neither verapamil nor amlodipine had an effect despite a blood pressure reduction at least equal to that of mibefradil. The in vivo effect of mibefradil was indeed an inhibition of smooth muscle cell proliferation, as shown by thymidine incorporation experiments. This antiproliferative effect of mibefradil was also observed in vitro in smooth muscle cells stimulated by fetal calf serum. In this condition also, verapamil was ineffective. We conclude that in rats mibefradil has a potent antiproliferative effect on smooth muscle cells after vascular injury. This effect might be due to blockade of voltage-operated T channels.

Endothelium-modulated proliferation of medial smooth muscle cells: influence of angiotensin II and converting enzyme inhibition.

This study investigated the role of the endothelium and angiotensin II (Ang II) in regulating medial smooth muscle cell (SMC) proliferation. [3H]-thymidine incorporation into medial SMC of rat arteries was examined in vivo, using ballooned rat carotid arteries, as well as in vitro, using cultures of aortic tissue rings (organoids). In vivo, maximal medial [3H]-thymidine incorporation occurred within 3 days post-ballooning. In endothelium-denuded organoids, maximum medial DNA synthesis was achieved after 7 days of culture. [3H]-thymidine-labelling of SMC in intact organoids (with endothelium) increased minimally during culture, indicating that the endothelium provided protection with respect to medial proliferation under basal conditions (culture in the presence of 1% plasma-derived serum). Inclusion of 10(-7) M Ang II significantly elevated medial [3H]-thymidine incorporation above that in control cultures. The stimulatory effect of Ang II was much more pronounced in intact organoids that in endothelium-denuded organoids, indicating synergistic growth regulation by Ang II and endothelium-derived factors. When organoids were cultured in the combined presence of Ang II and the ACE inhibitor cilazaprilat, labelling indices of intact organoids were also significantly increased above control, but to a lower level than those obtained in the presence of Ang II alone. However, for endothelium-denuded organoids, medial [3H]-thymidine incorporation in the combined presence of Ang II and cilazaprilat was not significantly different from that in untreated controls. Thus, cilazaprilat exerts both endothelium-dependent and endothelium-independent negative regulatory effects on medial SMC proliferation.

Functional aspects of vascular tenascin-C expression.

The arterial tenascin C expression in vivo and in vitro has been studied using immunohistochemistry. The functional relevance of localized tenascin C expression was assessed in vitro using various human cell types involved in the progression of vascular disease. Normotensive and hypertensive rats exhibited age-dependent patterns of vascular (aorta) tenascin expression, but the lumen-to-media-directed progression of tenascin induction was accelerated in hypertensive rats. Tenascin-rich neointimal lesions (spontaneous) were observed at branching sites of aorta from aged (80 weeks) hypertensive rats. Subendothelial tenascin foci contained lipid-laden smooth muscle cells and monocytes/macrophages. Medial tenascin foci encaged smooth muscle cells which synthesized DNA. Tenascin was expressed both in vivo and in vitro by endothelial and smooth muscle cells but not by monocytes/macrophages; angiotensin II, oxidized-low density lipoprotein and transforming growth factor beta 1 induced expression of tenascin transcripts and glycoprotein in vitro. Endothelial and smooth muscle cells, but not monocytes, adhered to tenascin substrata. Tenascin reduced focal adhesion integrity in confluent endothelial and smooth muscle cell cultures. Angiotensin II-induced migration of endothelial and smooth muscle cells was accompanied by tenascin deposition within extracellular matrix migration trails. Tenascin may function both as a defense against monocyte invasion and medial smooth muscle replication, as well as a substratum for directed endothelial and smooth muscle cell migration.

Diminished vascular response to inhibition of endothelium-derived nitric oxide and enhanced vasoconstriction to exogenously administered endothelin-1 in clinically healthy smokers.

BACKGROUND: Smoking is a major risk factor for the development of atherosclerosis. Because endothelial dysfunction may be a marker for future atherosclerosis, we investigated the effects of smoking on endothelium-dependent control of vascular tone. METHODS AND RESULTS: The effects of brachial arterial infusions of NG-monomethyl-L-arginine (L-NMMA), a nitric oxide synthesis inhibitor; sodium nitroprusside; endothelin-1; and norepinephrine on forearm blood flow (strain-gauge plethysmography) were compared in 29 long-term smokers and 16 nonsmokers. The acute effects of smoking on systemic hemodynamics, plasma catecholamines, and forearm vascular responses to these compounds were investigated in smokers only. Smokers did not differ from nonsmokers (n = 16) regarding the vascular effects of sodium nitroprusside (n = 13) or vasoconstriction due to norepinephrine and endothelin-1 (n = 16). Low-dose endothelin-1-induced vasodilation, believed to reflect endothelial prostacyclin or nitric oxide release, was absent in smokers (n = 16), and their increase of forearm vascular resistance (FVR) after L-NMMA (n = 13) was impaired (35.6 +/- 27.9% versus 118.8 +/- 43.2%, P < .001). Short-term smoking (n = 11) increased blood pressure, heart rate, and plasma epinephrine concentrations (P < .05 or less); enhanced endothelin-1-induced vasoconstriction (delta FVR, 457 +/- 192% versus 254 +/- 143%, P < .01); and decreased norepinephrine-induced vasoconstriction (P < .05), but had no effect on the other interventions. CONCLUSIONS: Long-term smoking is associated with a diminished nitric oxide-dependent component of basal vascular tone and an impaired endothelium-dependent vasodilator response to low-dose endothelin-1 and short-term smoking enhances endothelin-1-induced vasoconstriction. Impaired endothelial control of vascular tone might reflect impairment of normal antiatherosclerotic endothelial functions in smokers, but the relevance of smoking-induced enhancement of endothelin-1 vasoconstriction remains to be determined.

Activation of human peripheral monocytes by angiotensin II.

This study has investigated the ability of the vasoconstrictor peptide angiotensin II to activate human peripheral blood monocytes. Activation was monitored by measuring both the release of tumor necrosis factor alpha from monocytes and their adhesion to monolayers of human endothelial cells. Angiotensin II-elicited activation of monocytes was dose-dependent (half-maximally effective concentration approximately 0.2 nM), saturable (maximally effective concentration approximately 5 nM), and sensitive to inhibition by the angiotensin type 1 receptor antagonist ZD 7155. Such direct actions imply that angiotensin II is an important candidate stimulus for the subendothelial infiltration of monocytes observed in atherogenesis and hypertension.

A repressive coping style protecting from emotional distress in low-renin essential hypertensives.

OBJECTIVE: To investigate the relationship between the behavioural characteristics and specified subgroups of patients with essential hypertension. DESIGN AND METHODS: Fifty-four patients were classified into groups with a high (n = 9), normal (n = 35) or low plasma renin activity (n = 10), and were compared with 20 normotensive subjects by psychological tests. Standardized tests were used to measure anger expression, defensiveness and the subjects' psychological status (e.g. anxiety, depression). RESULTS: A repressive coping style, defined by a high defensiveness and low anxiety levels, was found significantly more often in patients with low than in patients with high plasma renin activity and normotensive subjects. The patients with high plasma renin activity scored significantly higher on suppressed anger, anxiety and interpersonal sensitivity than did those with low plasma renin activity. The scores of the normal plasma renin activity group were similar to those of the normotensive group. CONCLUSIONS: The results underline that there is not one hypertensive 'personality'. Whereas the patients with a high plasma renin activity appear to be more susceptible to emotional conflicts, the patients with low plasma renin activity report low emotional distress and maintain an apparently well-adjusted facade.

Angiotensin-II-induced expression of laminin complex and laminin A-chain-related transcripts in vascular smooth muscle cells.

Laminin, a major structural glycoprotein complex of basement membranes has been found to be modulated by angiotensin II in vitro and in vivo. In cultures of aortic organoids and vascular smooth muscle cells, expression of laminin was stimulated by exogenous vasoconstrictor peptide. Stimulation of laminin protein and mRNA expression was observed for both laminin B1/B2-chains and an unknown laminin heavy chain. Compared with PYS-2 cells, a mouse teratocarcinoma cell line which constitutively expresses a 10-kb mRNA transcript for 'classical' laminin A-chain, cultured vascular smooth muscle cells (VSMC) did not express a corresponding mRNA. However, cultured VSMC were found to express laminin A-chain-related mRNAs of approximately 1.8 kb and approximately 3.8 kb, respectively. The 1.8-kb species of transcript was expressed in a constitutive manner, whereas the 3.8-kb mRNA was found to be regulated by angiotensin II. Laminin complexes secreted by cultured cells contained a approximately 300 kD heavy chain which did not immunoreact with immunoreagents raised against either the classical laminin complex secreted by EHS tumor cells or the merosin heavy chain. The putative A-chain analogue possibly represents a new form of a tissue-specific laminin heavy chain, distinct from the A- and M-chains thus far described. Translation products encoded by the A-chain-related transcripts of cultured smooth muscle cells could not be specified using currently available antibodies. The putative protein(s) is speculated to contain the biological features of the N-terminus of the laminin A-chain, namely self-assembly and association with collagen type IV.

Characteristics of low and high density lipoprotein binding and lipoprotein-induced signaling in quiescent human vascular smooth muscle cells.

Low density lipoprotein (LDL) and high density lipoprotein (HDL) have been shown to stimulate signal transduction events in a number of cell types, including cultured vascular smooth muscle cells (VSMC), but it is not known whether these events are mediated through distinct lipoprotein receptors for transmembrane signaling. This study has used confluent quiescent cultures of human microarteriolar VSMC to investigate the relationship between the characteristics of 125I-LDL and 125I-HDL3 binding and those of LDL- and HDL3-stimulated cell signaling. Two distinct binding sites for LDL (Kd1 approximately 2 micrograms/ml and Kd2 approximately 40 micrograms/ml) and a single class of sites for HDL3 (Kd approximately 30 micrograms/ml) were identified. The Kd1 for high affinity 125I-LDL binding in quiescent VSMC was comparable to the value for heparin-sensitive binding of 125I-LDL to apolipoprotein B/E receptors in fibroblasts (Kd approximately 1 microgram/ml). Concentrations of lipoproteins required for half-maximal stimulation (EC50) of phosphoinositide catabolism and intracellular calcium mobilization in VSMC were approximately 35 micrograms/ml for HDL3 and approximately 40 micrograms/ml for LDL. Both LDL- and HDL3-stimulated signaling responses in VSMC, as well as 125I-HDL3 binding and low affinity 125I-LDL binding to VSMC, were insensitive to heparin. Competition binding studies (with unlabeled lipoproteins at 2.5-200 micrograms/ml) showed partial displacement of 125I-LDL by HDL3 and of 125I-HDL3 by LDL, whereas complete displacement of 125I-LDL or 125I-HDL3 by their homologous lipoproteins was achieved. Thus, the binding sites for HDL3 are distinct from those for LDL. Because the response of VSMC to combinations of LDL and HDL3 was additive, LDL and HDL3 also exert their signaling effects through distinct sites. Further investigation is required to unequivocally demonstrate that the heparin-insensitive HDL3 and low affinity LDL binding sites in VSMC are those through which LDL and HDL3 stimulate transmembrane signaling.

Concerted effects of lipoproteins and angiotensin II on signal transduction processes in vascular smooth muscle cells.

Low-density (LDL) and high-density (HDL3) lipoproteins dose-dependently activate phosphoinositide turnover and elevate cytosolic free Ca2+ concentrations ([Ca2+]i) in cultured vascular smooth muscle cells (VSMCs) from either human (microarterioles and aorta) or rat (aorta) sources. High-performance liquid chromatography analysis of cell extracts revealed comparable spectra of inositol phosphate isomers generated in response to either LDL, HDL3, or angiotensin II (Ang II). Thus, lipoproteins and Ang II may use similar, if not identical, signal transduction pathways for the generation and metabolism of inositol phosphates and intracellular Ca2+ mobilization in VSMCs. When Ang II was added in combination with either LDL or HDL3, the phosphoinositide and [Ca2+]i responses of VSMCs were either equal to or even greater than the sum of the effects elicited by the agonists individually. This additivity/synergy between Ang II and the lipoproteins was not accompanied by alteration in the half-maximally effective dose requirements of VSMCs for either Ang II (approximately 2 nmol/L, with or without lipoproteins) or lipoproteins (approximately 50 micrograms/mL for LDL and HDL3, with or without Ang II). Neither short-term (up to 10 minutes) nor long-term (48 hours) exposure of VSMCs to lipoproteins caused desensitization of phospholipase C and intracellular Ca2+ mobilization responses to either Ang II or lipoproteins. Since constant exposure of VSMCs to lipoproteins is a physiological circumstance, and because elevation of [Ca2+]i and activation of phosphoinositide turnover are pivotal events for VSMC contraction and growth, we suggest that the low concentrations of lipoproteins in the vessel intima may play an important role in regulating the response of the vasculature to Ang II.

Atrial natriuretic peptide release and volume regulation following kidney transplantation.

The major stimulus for atrial natriuretic peptide (ANP) release is atrial stretch and increased values are observed in volume overload states such as chronic renal failure. Since successful kidney transplantation restores volume homeostasis, we compared the effects of human cadaveric kidney transplantation on time course and changes of plasma ANP in the early postoperative period in 4 patients with successful and 4 patients with failed transplantation. ANP concentrations were elevated before transplantation in both groups (91 +/- 16 and 70 +/- 32 pmol/l) and decreased after successful (50 +/- 27 pmol/l, day 16) but increased after failed transplantation (146 +/- 45 pmol/l, day 16). Moreover, there was a close correlation between changes of body weight and ANP concentrations. Plasma renin activity decreased and plasma noradrenaline increased non-significantly in both groups, the latter more so after failed transplantation (116 +/- 42 to 194 +/- 156 vs 156 +/- 157 to 425 +/- 287 ng/l). No correlation was found between changes of renin activity or plasma catecholamines and ANP concentrations. The results indicate that the mechanisms governing release of atrial natriuretic peptide are operative in patients with chronic end-stage renal failure and after successful kidney transplantation with a return of atrial natriuretic peptide concentrations towards normal in the latter.

Morphogenic effects of endothelin-1 on vascular smooth muscle cells.

Endothelin-1 (ET-1), a vasoconstrictor peptide produced by endothelial and vascular smooth muscle cells (VSMC) might play a role in vascular remodelling. To investigate the proposed 'mitogenic' potential of ET-1, we examined the effects of chronic exposure of VSMC to ET-1 on cell cycle, growth/proliferation and differentiation under essentially mitogen-free culture conditions. Bulk cultures of thoracic aortic VSMC of spontaneously hypertensive (SHR) and normotensive Wistar Kyoto (WKY) rats, although exhibiting genetically determined differences in growth/proliferation (due to shortened G1 and G2 phases in SHR VSMC), respond in a similar manner to ET-1 exposure: long-term exposure (12-15 days) of VSMC from both sources to ET-1 in nonmitogenic medium did not promote cycling of cells. On the contrary, ET-1 attenuated the cycling of VSMC which had already cycled beyond the S phase. For cells which had not cycled beyond the S phase, ET-1 interrupted progression through the cell cycle at the late G1/early S phase. The specific ability of SHR VSMC to grow in mitogen-free medium was abolished by ET-1, most likely via down-regulation of platelet-derived growth factor (PDGF)-alpha receptors. Subsequent to ET-1 exposure, VSMC expressed increased levels of mRNA and protein for smooth-muscle-specific alpha-actin. However, expression of smooth muscle alpha-actin did not predominate over beta-actin as observed for adult contractile VSMC in vivo. The ET-1-induced expression of smooth-muscle-specific alpha-actin mRNA was dose dependent (EC50 approx. 2 x 10(-9) M), and alpha-actin protein expressed was associated with organized actin fibers.