Why oral calcium supplements may reduce renal stone disease: report of a clinical pilot study.

AIMS: To investigate whether increasing the daily baseline of gut calcium can cause a gradual downregulation of the active intestinal transport of calcium via reduced parathyroid hormone (PTH) mediated activation of vitamin D, and to discuss why such a mechanism might prevent calcium oxalate rich stones. To demonstrate the importance of seasonal effects upon the evaluation of such data. METHODS: Within an intensive 24 hour urine collection regimen, daily calcium supplementation (500 mg) was given to five stone formers for a 10 week period during a six month crossover study. In a further population of patients on follow up for previous renal stone disease, observations were made on 1066 24 hour urine samples collected over five years in respect of seasonal effects relevant to the interpretation of the study. RESULTS: In the group of patients on calcium supplements the following results were found. During calcium supplementation, the proportion of urine calcium to oxalate was higher (increased calcium to oxalate molar ratio), the 24 hour urine product of calcium and oxalate did not rise, and urine oxalate was lower during the first six weeks of supplementation. Twenty four hour urine calcium was 10.2% higher than baseline in the final four weeks of the 10 weeks of supplementation. Twenty four hour urine phosphate was 11.4% lower during the first six weeks of supplementation, but then rose while the patients were still on supplementation; renal tubular reabsorption of phosphate (TmP/GFR) mirrored the urine phosphate changes inversely. PTH was higher after stopping supplementation, but 1,25-(OH)2-cholecalciferol changes were not detected. In the 1066 urine samples collected over five years the following results were found. Calcium and oxalate excretion correlated positively and not inversely. Urine calcium and phosphate excretion were 5.5% and 2.5% higher, respectively, in "light" months of the year compared with "dark" months. A post summer decline in both urine calcium and urine phosphate was relevant to the interpretation of the study. CONCLUSIONS: Regular calcium supplementation does not raise the product of calcium and oxalate in urine and the proportion of oxalate to calcium is reduced. The underlying mechanisms of the changes seen in phosphate, calcium, and PTH and the observations on 1,25-(OH)2-cholecalciferol are not clear. Observed changes in phosphate could possibly be part of a calcium regulating feedback loop operating over a period of weeks. In evaluating these mechanisms background seasonal effects are important. It is possible that "programming" of the gut mucosa in terms of calcium transport is a major determinant of the relation between calcium and oxalate concentrations in urine and their relative abundance. Increased oral calcium, in association with a reduction of the relative proportion absorbed, may be pertinent to the prevention of calcium oxalate rich stones.

Characterization of dopamine-mediated relaxation in experimental vein bypass grafts.

BACKGROUND: Dopamine is an endogenous inotropic agent commonly used during coronary artery surgery and in the medical therapy of a revascularized patient. In this study the responses of intimal hyperplastic vein grafts to dopamine are examined. METHODS: The in vitro isometric tension responses to dopamine of common carotid jugular vein bypass grafts in New Zealand White rabbits were determined. The responses were compared to those obtained in the jugular vein and in the common carotid artery. Both endothelialized and denuded vessels were precontracted with prostaglandin F(2alpha) and the responses to dopamine were assessed. The contributions of nitric oxide and prostanoids to the response were also determined. RESULTS: Each vessel showed a biphasic dose response to dopamine with relaxation at low concentrations followed by contraction at high concentrations. Dopamine relaxation in the jugular vein was endothelial independent while in the carotid artery it was endothelial dependent and decreased. The sensitivity of both vessels was significantly greater than the vein graft (6.62 +/- 0.12; P < 0. 05); however, after endothelial denudation, the sensitivity of dopamine-mediated relaxation of the vein graft (8.91 +/- 0.09) was significantly enhanced. Preincubation with L-NMMA (to block NO synthesis) inhibited vein graft relaxation to dopamine and preincubation with indomethacin (to block cyclooxygenase activity) inhibited carotid artery relaxation to dopamine. Addition of phenoxybenzamine, a broad alpha-adrenergic antagonist, enhanced dopamine relaxation in the jugular vein and depressed the relaxation in the carotid artery. There was no effect on the dopamine response in the vein graft. Jugular vein and carotid artery responded to dopamine with cholera toxin-sensitive (Galpha(s)) responses. In contrast, dopamine relaxation in the vein graft was enhanced by inhibition of Galpha(s). CONCLUSION: Dopamine relaxation in vein grafts is mediated in part by NO but not by either prostanoids or alpha-adrenergic receptor activation. It is diminished compared to native vessels due to an endothelium-dependent, Galpha(s)-mediated pathway.

Combination therapy of cholesterol reduction and L-arginine supplementation controls accelerated vein graft atheroma.

Hyperlipidemia contributes to the development of intimal hyperplasia and accelerated atheroma in vein bypass grafts. Dietary cholesterol reduction and oral supplementation with L-arginine have been shown to reduce accelerated atheroma in experimental vein grafts. This study extends these observations by examining the effect of the combination therapy of cholesterol reduction and L-arginine supplementation on the development of intimal hyperplasia in vein grafts in hypercholesterolemic animals. Thirty New Zealand White rabbits had a carotid vein bypass graft performed and were sacrificed at 28 days postoperatively either for morphology (light and electron microscopy) and videomorphometry, or for in vitro contractile studies. Twenty animals received a 1% cholesterol diet for 4 weeks prior to surgery. This diet was continued until harvest in ten animals. Ten cholesterol-fed animals received L-arginine supplementation (2 g/kg/day, p.o.) for 7 days preoperatively and thereafter until harvest and in addition were returned to a normal diet on the day of surgery. The last ten animals were controls (normal diet). Combined cholesterol reduction and L-arginine supplementation prevented accelerated atheroma in vein grafts, halted the change in enhanced smooth muscle cell contractility, and improved endothelial cell function. Early postoperative therapy targeting atheroma development in the high-risk patient could offer significant morphological and functional benefits.

Local effects of nitric oxide supplementation and suppression in the development of intimal hyperplasia in experimental vein grafts.

OBJECTIVES: The universal response of vein grafts after insertion into the arterial circulation is the development of intimal hyperplasia; smooth muscle cell proliferation and connective tissue deposition, which may be modulated in part by dysfunctional endothelial nitric oxide (NO) metabolism. This study examines the effects of single dose, local application by pluronic gel of a NO donor, S-nitroso-N-acetylpenicillamine (SNAP) and an NO synthase inhibitor nitro-L-arginine methyl ester (L-NAME) on the formation of intimal hyperplasia. MATERIALS: Forty New Zealand white rabbits underwent jugular vein interposition grafting of the common carotid artery. DESIGN: Ten animals were controls, 10 animals had the outer surface of the vein graft coated with 30% pluronic gel (2.5 ml), and 10 each were immersed for 15 min prior to insertion in Ringer lactate containing 10(-3) M of SNAP or L-NAME and then had their vein grafts coated with 2.5 ml of gel containing either SNAP (10(-3) M) or L-NAME (10(-3) M), which allows for sustained delivery for up to 6 h. On the 28th post operative day, the animals were sacrificed and vein grafts were harvested for morphology by electron microscopy (SEM and TEM) and dimensional analysis by videomorphometry. RESULTS: All vein grafts developed intimal hyperplasia. On SEM the vein grafts had a confluent layer of endothelial cells with multiple layers of smooth muscle cells representing intimal hyperplasia in TEM. There were no demonstrable morphological differences between the four groups. Local treatment with SNAP produced a significant 36% decrease in mean intimal thickness (72 +/- 4 microns vs. 45 +/- 4 microns; mean +/- S.E.M.; p < 0.01) without a change in medial thickness compared to gel-only treated groups (58 +/- 6 microns vs. 61 +/- 7 microns; p = ns). Inhibition of NO synthase by L-NAME had no effect on the development of intimal hyperplasia (72 +/- 4 microns vs. 79 +/- 10 microns; p = ns); medial thickness was also unchanged. CONCLUSION: These data confirm the protective effect of NO in vascular injury and suggest that NO synthase activity is either absent or reduced to such a level that further inhibition in this short time course is not relevant to the pathophysiology of vein graft intimal hyperplasia.

Oral monoterpene therapy (perillyl alcohol) reduces vein graft intimal hyperplasia.

The development of intimal hyperplasia is recognized as a major impediment to graft patency. D-Limonenes are monoterpenes with a recognized cytostatic effect on cell proliferation by inhibiting posttranslational isoprenylation of p21ras and other small G-proteins. This study examines the effect of perillyl alcohol, an oral hydroxylated D-limonene, on the development of intimal hyperplasia and its associated smooth muscle cell physiological responses in an experimental model of vein bypass grafting. Twenty New Zealand white rabbits had a right carotid interposition bypass graft using the ipsilateral external jugular vein. Ten animals received chronic oral therapy with a perillyl alcohol (200 mg/kg/day; begun 5 days before surgery and continued until harvest) and 10 control animals received vehicle only. All animals were sacrificed on the 28th postoperative day. Vein grafts were harvested either for morphology/videomorphometry (n = 6 per group) or for in vitro isometric tension studies (n = 4; four 5-mm rings per graft). The cell proliferation and incorporation of [3H]thymidine into the cellular DNA of serum-stimulated rabbit aortic smooth muscle cells was also assessed in the presence of increasing concentrations of perillyl alcohol (10(-9)-10(-4) M). Perillyl alcohol treated vein grafts showed a 22% reduction in overall mean intimal thickness (54 +/- 4 microns vs 69 +/- 3 microns; P = 0.006) but a 25% increase in overall mean medial thickness (86 +/- 4 microns vs 61 +/- 3 microns). The intimal ratio of the perillyl alcohol treated vein grafts decreased by 27% compared to controls. Perillyl alcohol induced norepinephrine and serotonin hypersensitivity in vein grafts compared to controls. The IC50 for perillyl alcohol was 176 nM with maximal inhibition at 5 microM. Incubation of smooth muscle cell cultures with increasing concentrations of perillyl alcohol showed a dose-dependent decrease in in vitro cellular proliferation, maximal at 1 microM. Therapy with perillyl alcohol alters the early development of intimal hyperplasia reducing the intimal response but increasing the medial response without significant changes in the physiological responses of the smooth muscle cells. Modulating G-proteins will affect the intimal hyperplastic response in vein grafts.

L-arginine supplementation improves venous endothelial cell but not smooth muscle cell dysfunction induced by prolonged diet-induced hypercholesterolemia.

Hypercholesterolemia induces venous vasomotor dysfunction. This study examines the endothelial and smooth muscle cell vasoreactivity of external jugular veins from rabbits fed either a normal or a 1% cholesterol diet for 8 weeks with and without L-arginine supplementation (2 g/kg day-1 orally for the last 5 weeks). Isometric tension studies were performed on harvested jugular veins. Concentrations of serum cholesterol were 20-fold higher than controls and serum L-arginine twofold higher than untreated animals. Hypercholesterolemia induced hypersensitivity to norepinephrine (p < .05), bradykinin (p < .05), and histamine (p < .05) with a contractile response to serotonin compared to controls. L-Arginine supplementation decreased bradykinin hypersensitivity but had no effect on the changes in norepinephrine serotonin and histamine responses compared to controls. Hypercholesterolemia interfered with relaxation induced by acetylcholine but with L-arginine, normal acetylcholine-induced, endothelium-dependent relaxation returned (54 +/- 10%, compared to 40 +/- 14% in control veins; p > .05). Non-endothelium-dependent relaxation to sodium nitroprusside of precontracted veins was unaffected by the presence of high cholesterol concentrations. This study suggests that L-arginine therapy may ameliorate hypercholesterolemia-induced functional abnormalities in endothelial cells.

Control of accelerated vein graft atheroma with the nitric oxide precursor: L-arginine.

Hyperlipidemia contributes to the development of intimal hyperplasia and subsequent accelerated atherosclerosis in vein bypass grafts. This study examines the effect of dietary supplementation with L-arginine on the development of intimal hyperplasia and the vasomotor function of vein grafts in hypercholesterolemic animals. Thirty male New Zealand White rabbits had a right carotid vein bypass graft and were sacrificed at 28 days postoperatively. Twenty animals received a 1% cholesterol diet for 4 weeks prior to surgery and this diet was continued until harvest. Of these, 10 also received L-arginine (2.25%, 2 g/kg, p.o.) 7 days preoperatively and thereafter until harvest. The last 10 animals were controls. Vein grafts were harvested either for morphology or for in vitro isometric tension studies. Cumulative dose-response curves to norepinephrine, serotonin, and bradykinin were recorded, and following norepinephrine precontraction, relaxation to acetylcholine and sodium nitroprusside were determined. After in situ pressure fixation, intimal thicknesses of the vein grafts were measured by videomorphometry. The addition of L-arginine doubled the serum arginine concentrations. Intimal hyperplasia of both groups of hypercholesterolemic vein grafts contained foam cells and lipid-laden endothelial and smooth muscle cells. There was a 24% reduction in the intimal thickness of vein graft intimal hyperplasia in the L-arginine group compared to that in the hypercholesterolemia group (P < 0.05). All hypercholesterolemic vein grafts were two-fold thicker than in the control group. L-arginine supplementation resulted in the preservation of acetylcholine-mediated relaxation but did not change hypercholesterolemia-induced contractile agonist supersensitivity.(ABSTRACT TRUNCATED AT 250 WORDS)

Ramipril and experimental vein graft intimal hyperplasia.

Angiotensin-converting enzyme (ACE) inhibitors have been shown to reduce the intimal proliferation in animal models of arterial angioplasty and vein bypass grafting. This study examines the effect of high-dose ramipril, an ACE inhibitor that does not contain a sulfhydryl group, on the development of intimal hyperplasia in experimental vein bypass grafts. Twenty New Zealand White rabbits underwent common carotid interposition bypass grafting. Twelve were treated with ramipril (2 mg/kg/day; po) five days prior to surgery and thereafter until harvest. The remaining 8 animals were used as controls. Vein grafts were harvested at twenty-eight days by pressure fixation (80 mmHg). The grafts were sectioned into proximal, middle, and distal thirds, and the thickness of the intima and the media and the area of the lumen from each segment were determined by videomorphometry. The effect of ramipril on the [H3]thymidine incorporation into DNA of serum-stimulated smooth muscle cells (culture passage 6 to 12) was also assessed. There was a 50% mortality rate in the rabbits that received ramipril, and this was assumed to be related to the high dose of the drug. Ramipril treatment reduced mean vein graft intimal area by 34% (P > 0.05), but this was accompanied by an increase of 73% in the mean medial area of the vein grafts as compared with controls. These changes resulted in a decrease in the mean intimal ratio (intima/[intima + media]) by 39% in the ramipril group as compared with controls. Ramipril did not inhibit [H3]thymidine incorporation into DNA of serum-stimulated smooth muscle cells.(ABSTRACT TRUNCATED AT 250 WORDS)

Reduction of experimental vein graft intimal hyperplasia and preservation of nitric oxide-mediated relaxation by the nitric oxide precursor L-arginine.

BACKGROUND: Previous studies in animals and human beings have shown that vein bypass grafts exhibit diminished endothelium-dependent relaxation and the development of intimal hyperplasia. This study examines the effect of L-arginine on experimental vein graft endothelial cell function and the development of intimal hyperplasia. METHODS: Common carotid vein bypass grafts were performed in 24 New Zealand White rabbits: 12 were controls and 12 received L-arginine (2.25%) orally 7 days before operation and thereafter until harvest 28 days after operation. Intimal and medial dimensions were determined by planimetry on pressure-fixed vessels. Relaxation to acetylcholine, serotonin, calcium ionophore (A23187), and sodium nitroprusside was performed on precontracted vessel rings. RESULTS: Arginine-treated vein grafts showed a 47% reduction in mean intimal thickness (p < 0.001) compared with controls. By scanning and transmission electron microscopy, all vein grafts showed a confluent endothelium. In contrast to control grafts, which do not relax to acetylcholine and serotonin, arginine-treated vein grafts relaxed in response to both agonists. There was a significant increase (p < 0.05) in the maximal relaxation to calcium ionophore (A23187) in arginine-treated vein grafts compared with control grafts. Non-endothelium-dependent responses to sodium nitroprusside were equivalent in all vein grafts. CONCLUSIONS: This study shows that oral L-arginine supplementation significantly reduces intimal hyperplasia and preserves nitric oxide-mediated relaxation in experimental vein grafts, suggesting a role for nitric oxide in the regulation of the cellular events that lead to intimal hyperplasia.