Differential regulation of macrophage arginine metabolism: a proposed role in wound healing.
Am J Physiol (UNITED STATES) Feb 1997, 272 (2 Pt 1) pE181-90
Nitric oxide (NO) and ornithine, products of NO synthase or arginase, respectively, have opposing biological activities. The effect of mediators of leukocyte activation and inhibition on arginine metabolism of resident mouse peritoneal exudate cells (MPEC) was determined. Factors that increased basal NO synthase activity, interferon (IFN)-gamma and lipopolysaccharide (LPS), decreased arginase activity in intact cells. Transforming growth factor (TGF)-beta1 decreased IFN-gamma-stimulated NO synthase activity and produced a reciprocal increase in urea and ornithine release. TGF-beta1 had no effect on the activity of these enzymes in LPS-stimulated MPEC. Corticosterone (Cort, 100 ng/ml) decreased the basal activity of both enzymes.However, Cort inhibited NO synthase activity and increased ornithine release in MPEC exposed to IFN-gamma or LPS. The difference between arginase activity in intact cells vs. that of cell lysates suggested intracellular inhibition of arginase activity. Products of NO synthase, NO and citrulline, were shown to inhibit MPEC arginase activity under maximal assay conditions. Intracellular pH was not altered by exposure of MPEC to LPS, IFN-gamma, TGF-beta, and Cort. This reciprocal change in arginine metabolism is proposed to be an important component of wound healing. Expression of NO synthase creates a cytotoxic environment that may be important to the early phase of wound healing. As wound healing progresses, increased arginase activity produces an environment favorable for fibroblast replication and collagen production.
Human dermal fibroblasts produce nitric oxide and express both constitutive and inducible nitric oxide synthase isoforms.
J Invest Dermatol (UNITED STATES) Mar 1996, 106 (3) p419-27
Nitric oxide (NO) is produced by a variety of human and animal cells and is involved in a broad rray of physiological and pathophysiological processes. It can cause vasodilation, serve as a neurotransmitter, and have anti-neoplastic, anti-microbial, and anti-proliferative effects. In this study, we have demonstrated that fibroblasts derived from human skin spontaneously produce NO and that this production can be enhanced by stimulating the cells with interferon-gamma and lipopolysaccharide. The production of NO by human dermal fibroblasts can be blocked by NG-monomethyl-L-arginine (L-NMMA). The inhibitory effect of L-NMMA on NO production was restored by addition of L-arginine but not D-arginine. By measuring the rate of conversion of [14C]L-arginine to [14C]L-citrulline, we show that unstimulated cells expressed only Ca2+-dependent NO synthase (NOS) activity (1.36 +/- 0.57 pmol/mg/min; n = 4) whereas stimulated cells expressed both Ca2+-dependent (2.60 +/- 0.54 pmol/mg/min; n = 4) and -independent (1.59 +/- 0.14 pmol/mg/min; n = 4) NOS activities. With reverse transcription polymerase chain reaction (RT-PCR), the 422-bp RT-PCR product for human endothelial constitutive NOS and the 462-bp RT-PCR product for human hepatocyte inducible NOS were detected in proportion to the amount of mRNA-related RT-cDNA added to the reaction mixture. Further evidence by immunocytochemistry demonstrated that human dermal fibroblasts express both constitutive and inducible NOS proteins. These data collectively suggest that in addition to macrophages and other inflammatory cells, nitric oxide production by dermal fibroblasts could be important during the inflammatory stages of wound healing and possibly also in the later stages of proliferation and tissue remodeling after skin injury in humans.
Role of lactose, arginine and lysine combination in fracture healing (an experimental study)
Ann Ital Chir (ITALY) Jan-Feb 1996, 67 (1) p77-82; discussion 82-3
L-arginine and L-lysine are essential amino acids which seem to possess some properties able to influence bone fractures healing. In fact, the increase of intestinal calcium adsorption but also in collagen synthesis, in insulin and growth hormone secretion and in osteoblastic activation. So, an experimental in vivo model was carried out by using 50 adult rabbits which, under general anaesthesia, were submitted to an osteotomy of the left fibula. Animals were divided into 5 groups and were daily treated with a mixture of lactose, L-arginine and L-lysine or with the only lactose (control group) at the same dosage as recommended for humans. They were sacrificed after 15, 30, 40, 50 and 60 days for radiological and histological studies. The results of the study showed that the pharmacological mixture containing L-arginine and L-lysine accelerates and ameliorates the healing processes and this positive effect was particularly evident from the 30th day after the osteotomy. We think that these results are linked not only to calcium metabolism but also to different biological properties which positively contribute to good healing of bone fractures.
Sepsis impairs anastomotic collagen gene expression and synthesis: A possible role for nitric oxide
Journal of Surgical Research (USA), 1997, 69/1 (81-86)
Although intra-abdominal sepsis is known to impair colon healing by inhibiting anastomotic collagen synthesis, the effect of systemic sepsis on this process is unknown. Endotoxins and cytokines associated with sepsis induce nitric oxide synthesis both systemically and locally within colonic tissue. We hypothesized that systemic sepsis impairs colonic healing and examined a possible correlation with nitric oxide expression. Male Sprague- Dawley rats received intraperitoneal injections of either saline (sham group) or Escherichia coli endotoxin (lipopolysaccharide 1 mg/100 g body weight) at Times - 24 and - 12 hr (LPS group). All animals underwent laparotomy and left colonic anastomosis at Time 0. At 24 and 96 hr postlaparotomy rats were sacrificed, the anastomoses excised, and (3H)-proline incorporation into protein measured as an index of total new protein synthesis (TNP). Digestion with purified collagenase yielded incorporation into the collagen fraction (CDP). Additional sham and LPS-treated rats were sacrificed at 24, 72, and 120 hr, the anastomoses excised, and nitric oxide synthase activity in the tissue measured by the conversion of (3H)-arginine to (3H)citrulline in an ex vivo culture system. Finally, sham and LPS rats were sacrificed at 120 hr for measurement of colon anastomotic bursting pressure. Systemic sepsis significantly impaired new collagen synthesis in anamotic tissue at 24 hr compared to control samples (P < 0.02). No difference was noted at 96 hr. TNP synthesis was similar in both groups at 24 or 96 hr. Northern blot analysis confirmed a significant decrease in Type I and Type III collagen mRNA expression at 24 hr in septic rats. Anastomotic bursting pressure was also decreased in the septic group (P < 0.003). Sepsis elevated nitric oxide synthase activity in anastomotic tissue 24 hr postanastomosis, when compared to sham tissue (P < 0.0001). These data suggest that systemic endotoxin induces nitric oxide synthesis at the anastomotic site. The simultaneous dysregulation of collagen gene expresion and synthesis with decreased anastomotic strength suggests a possible regulatory role for nitric oxide in gastrointestinal healing.
Regionally different vascular response to vasoactive substances in the remodelled infarcted rat heart; Aberrant vasculature in the infarct scar
Journal of Molecular and Cellular Cardiology (United Kingdom), 1997, 29/5 (1487-1497)
Remodelling after myocardial infarction (MI) is associated with vascular adaption, increasing vascular capacity of non-infarcted myocardium, and angiogenesis in the infarcted part during wound healing and scarring. We investigated regional vascular reactivity in the infarcted rat heart. Transmural infarction of the left ventricular free wall was induced by coronary artery ligation. After 3 weeks, regional flow during maximal vasodilation (nitroprusside, NPR) and submaximal vasoconstriction (arginine-vasopressin, AVP) were studied in buffer-perfused hearts. The main findings were: (1) a reduced vasodilator response (NPR) in the viable part of the left ventricular free wall, where hypertrophy was most pronounced, resulting in reduced maximal tissue perfusion of the myocardium bordering the scar (19.7 plus or minus 0.6 v 25.7 plus or minus 1.2 ml/min.g), whereas peted regions was preserved. (2) A 54% lower vasodilator response (NPR) and a 25% stronger vasoconstriction (AVP) in scar tissue compared to viable parts of MI hearts. Microscopy showed thicker walls of resistance arteries in scar tissue than in viable parts of MI hearts or in sham hearts, morphometrically substantiated by two- to three-fold greater wall/lumen ratios. These data indicate a deviant response of scar vessels of MI hearts, and in the non-infarcted part, a reduced coronary reserve in the most hypertrophied region. Whereas the former may be caused by different vessel structure, the reduced vasodilator reserve of the spared part of the left ventricular free wall may indicate vasodilation at rest due to insufficient vascular growth. Thus, the most hypertrophied region would be at the highest risk of further ischemic damage.
Modulation of tendon healing by nitric oxide
Inflammation Research (Switzerland), 1997, 46/1 (19-27)
Nitric oxide (NO.) is a small, diffusible free radical that is generated from L-arginine by a family of enzymes, collectively termed the nitric oxide synthases. We investigated the role of NO. in tendon healing. NO. synthase activity and immunoreactivity was absent in un-injured rat Achilles tendon. After surgical division there was a five-fold increase in NO. synthase activity and immunoreactivity within the healing tendon at day 7, with a return to near baseline levels at day 14. Inhibition of NO. synthase activity with oral administration of Nomega-nitro-L-arginine methyl ester (L-NAME) resulted in a significant reduction in cross-sectional area (30% at day 7, p < 0.01, 50% at day 15, p < 0.001) and failure load (24% at day 7, p < 0.01) of the healing Achilles tendon constructs. Rats fed the same regimen of the enantiomer of L-NAME, (D-NAME) had normal tendon healing. These results indicate that nitric oxide synthase is induced during tendon healing and inhibition of nitric oxide synthase inhibits this tendon healing.
Acute protein-calorie malnutrition impairs wound healing: A possible role of decreased wound nitric oxide synthesis
Journal of the American College of Surgeons (USA), 1997, 184/1 (37-43)
BACKGROUND: Nitric oxide is synthesized in wounds. Systemic inhibition of wound nitric oxide synthesis decreases wound collagen accumulation and wound mechanical strength. The role of nitric oxide during impaired healing is not known. In a model of impaired wound healing induced by acute protein- calorie malnutrition, we correlated wound healing parameters with wound nitric oxide synthesis. STUDY DESIGN: One group of Sprague-Dawley rats was rendered acutely malnourished by restricting its food intake to 50 percent of the food intake of an ad libitum-fed control group. Wound collagen accumulation and types I and III collagen gene expression were measured 10 days postwounding in subcutaneously implanted polyvinyl alcohol sponges. Nitric oxide synthesis was determined in wound fluid and in supernatants of wound cell cultures. RESULTS: Animals with acute protein-calorie malnutrition lost 10.4plus or minus0.8 percent, while controls gained 17.5plus or minus1.2 percent of their original body weight. Protein-calorie malnutrition reduced sponge hydroxyproline contents (995plus or minus84 compare with 1,580plus or minus109 microg/100 mg sponge, p<.001), indicating diminished wound collagen accumulation. Gene expression of type III, but not type I, collagen was decreased in wounds of protein- calorie malnutrition animals. Nitrite/nitrate and citrulline concentrations in wound fluid (p<.01) and in wound cell supernatants (p<.001) were also lower in protein-calorie malnutrition animals, indicating a net decrease in nitric oxide production. CONCLUSIONS: Impaired wound collagen accumulation caused by protein-calorie malnutrition may be a reflection of reduced nitric oxide synthesis within the wound.
Dietary L-arginine in renal disease
Seminars in Nephrology (USA), 1996, 16/6 (567-575)
The amino acid L-arginine is a substrate for at least three products involved extensively in tissue injury and fibrosis. L-arginine is metabolized to L-proline, a major constituent of the collagen that makes up fibrotic extracellular matrix. L-arginine is a precursor for polyamines, which are required for proliferative responses characteristic of many renal diseases. L-arginine is also the sole substrate for generation of nitric oxide (NO) which, produced in large quantities by macrophages, has been implicated in tissue injury. On the other hand, NO produced in small quantities endothelium is a critical vasodilator. Given the importance of elevated intraglomerular pressure in renal injury, it is perhaps not surprising that dietary L-arginine supplementation increases NO generation and is beneficial in reducing intraglomerular pressure and subsequent disease. Other data, based on the therapeutic effects of low protein diets, have suggested that L- arginine restriction limits NO-mediated glomerular injury and greatly reduces matrix accumulation, consistent with the idea that limitation of substrate effectively diminishes injurious NO levels, polyamine synthesis, and collagen production.
Arginine-enriched diets: Rationale for use and experimental data
Nutrition Clinique et Metabolisme (France), 1996, 10/2 (89-95)
Since the pioneering work of Rose who classified arginine as a non- essential amino acid, subsequent works have revealed that arginine can become an essential amino acid in stress situations. In septic rats, arginine- enriched nutrition (either enteral or parenteral) improves nitrogen balance and total body and liver protein synthesis. In addition, arginine stimulates growth hormone and insulin secretion. The most remarkable action of arginine is certainly that exerted on cellular immunity. This action concerns thymus and extra-thymus areas. Finally, arginine favours wound healing improves host defenses in cancer and slows tumour growth. The pharmacological action of arginine probably depends upon various mechanisms: its action on immunity may be mediated by the synthesis of nitric oxide and polyamines (via ornithine synthesis). The effect on wound healing may be related to proline synthesis. The effects on nitrogen metabolism may be linked to growth hormone secretion. These observations form the rationale for the administration of arginine- enriched diets to injured patients.
Effects of an arginine-glycine-aspartic acid peptide-containing artificial matrix on epithelial migration in vitro and experimental second-degree burn wound healing in vivo
Journal of Burn Care and Rehabilitation (USA), 1996, 17/3 (199-206)
Cells central to dermal tissue repair such as dermal fibroblasts and keratinocytes interact with arginine-glycine-aspartic acid (RGD)-containing proteins of the extracellular matrix such as fibronectin. It has been shown that synthetic peptides containing this RGD sequence can also support cell attachment and migration in vitro. We therefore set out to test whether the use of these peptides, when formulated as a synthetic RGD-peptide matrix consisting of peptide complexed with hyaluronic acid, would have an effect on the rate of epithelial migration and hounds. Evaluation consisted of measuring the extent of epithelial outgrowth from human dermal explants and the epithelization of experimental second-degree burn wounds in pigs. We show here that the RGD-peptide matrix supports epithelial sheet migration from explants in a dose-dependent manner. In second-degree burn wounds in pigs, wounds treated with daily applications of the RGD peptide matrix under occlusion resurfaced at a significantly faster rate (day 7 = 57% completely epithelized) than wounds treated with hyaluronic acid under occlusion (day 7 = 13% completely epithelized, p < 0.01), occlusion alone (day 7 = 13% completely epithelized, p < 0.01), or air exposed (day 7 = 0% completely epithelized, p < 0.001). Histologic examination showed that wounds treated with the RGD-peptide matrix also had thicker epithelial covering and greater granulation tissue deposition than occluded, air-exposed, and hyaluronate-treated wounds. These data therefore show that the use of RGD-peptide matrix induces faster explant epithelial migration and results in faster healing of experimental second-degree burns.
Spontaneously increased production of nitric oxide and aberrant expression of the inducible nitric oxide synthase in vivo in the transforming growth factor beta1 null mouse
Journal of Experimental Medicine (USA), 1996, 183/5 (2337-2342)
Transforming growth factor beta1 null mice (TGF-beta1(-/-)) suffer from multifocal inflammation and die by 3-4 wk of age. In these mice, levels of nitric oxide (NO) reaction products in serum are elevated approximately fourfold over levels m controls, peaking at 15-17 d of life. Short-term treatment of TGF-beta1(-/-) mice with N(G)-monomethyl-L-arginine suppressed this elevated production of NO. Expression of inducible NO synthase (iNOS) mRNA and protein is increased in the kidney and heart of TGF-beta1(-/-) mice. These findings demonstrate that TGF-betaerred from mechanistic studies o n the control of iNOS expression by TGF-beta1 in vitro.
Nitric oxide is necessary for a switch from stationary to locomoting phenotype in epithelial cells
American Journal of Physiology - Cell Physiology (USA), 1996, 270/3 39-3
The restitution of epithelial integrity is accomplished in part by cell migration. Studying this process, we have found that nitric oxide (NO) release from migrating epithelial BSC-1 cells displayed a biphasic response to the inflicted wounds; an initial transient release of NO is followed by a delayed sustained elevation. Whereas the constitutive endothelial NO synthase (NOS) did not show any spatial or temporal changes associated with wounding, the inducible NOS became expressed 3 h after wounding and showed higher abundance at the edges of epithelial wounds. L-Arginine (L-Arg) or NO-donor, S-nitroso-N-acetyl-DL-penicillamine, exerted motogenic effect in epithelial and endothelial cells. Inhibition of NOS with N(G)-nitro-L-arginine methyl ester (L-NAME) or a selective knockout of inducible NOS with antisense oligodeoxynucleotides reduced the rate of spontaneous or epidermal growth factor (EGF)-induced BSC-1 cell migration. Migrating cells showed the polarized expression of NOS, suggesting a head-to-rear NO gradient. Several growth factors (EGF, insulin-like growth factor I, hepatocyte growth factor, and fibroblast growth factor) were motogenic for BSC-1 cells, but this effect was abrogated by pretreatment with L-NAME. We conclude that endogenous NO production is a prerequisite for BSC-1 cell migration. A vectorial NO release may be essential for the spatially and temporally coordinated reciprocal phenomena that occur at the leading and trailing edge of locomoting epithelial cells. Although the exact mode of NO action remains uncertain, it is conceivable that the production of NO serves as a cellular switch from the stationary to the locomoting epithelial phenotype.
The effect of an arginine-glycine-aspartic acid peptide and hyaluronate synthetic matrix on epithelialization of meshed skin graft interstices
Journal of Burn Care and Rehabilitation (USA)
Keratinocytes and fibroblasts interact with proteins of the extracellular matrix such as fibronectin and vitronectin through RGD (arginine-glycine- aspartic acid) cell-attachment sequences. This study evaluated the ability of a provisional synthetic matrix composed of an RGD peptide and hyaluronic acid to accelerate the epithelialization of the interstices of meshed, human, split-thickness skin when placed on full-thickness wounds of athymic mice. Full-thickness skin defects, sparing the panniculus carnosus, were created on athymic mice and 3:1 meshed, human skin was placed on them. The grafts had four central, isolated interstices, which epithelialized by migration of human keratinocytes. Conditions were either the addition to the wound of the synthetic matrix or a matrix of hyaluronic acid alone. The time to closure of the graft interstices was decreased (p < 0.02) in the wounds treated with the RGD peptide-hyaluronic acid provisional matrix. The resultant epithelium of the closed interstices was significantly thicker 8 days after surgery for the RGD-treated wounds. Basement membrane proteins (laminin and type IV collagen) were also found to be present at the dermoepidermal junction earlier in the RGD-treated wounds. These results imply that use of the RGD peptide conjugate to effect-cell-matrix interactions may have clinical significance in the field of wound healing.
Clinical application of basic arginine amidase in human male urine.
Biol Pharm Bull (JAPAN) Aug 1996, 19 (8) p1083-5
Basic human urinary arginine amidase (or esterase, called BHUAE) which is only found in male urine, was measured from normal volunteers between the age of 4 and 70 years using D-valyl-L-leucyl-L-arginine-p-nitroanilide as a substrate. BHUAE increases during early adolescence, between 8 to 17 years of age. Then, BHUAE decreases in the twenties and takes a certain range of value in the mature age group, between the late thirties and fifties. In patients with prostate cancer, a significant increase in BHUAE was demonstrated in comparison with the healthy male group (control) over 55 years old. On the other hand, patients with benign prostatic hypertrophy showed no significant elevation of this enzyme activity. It would appear that the measurement of BHUAE in urine can be used as a marker of prostate cancer in an advanced age group.
Arginine needs, physiological state and usual diets. A reevaluation.
J Nutr (UNITED STATES) Jan 1986, 116 (1) p36-46
Evidence is discussed that puts in question the widely held belief that adult mammals, including human beings, can meet all of their arginine needs by endogenous synthesis. Arginine, used in synthesis of body proteins, is essential for ammonia detoxification via urea synthesis, which prevents metabolic derangements caused by elevations in tissue ammonia. It is a precursor for polyamine synthesis and is the only source of amidino groups for the formation of creatine, a major source of high energy phosphate for regeneration of ATP in muscle. Arginine at supraphysiologic doses is thymotropic and a secretagogue for hormones that control growth and metabolism. Studies in mature rats show that glucose tolerance, the rate of repletion from severe protein undernutrition and recovery from trauma are significantly accelerated by dietary arginine. Oral or intravenous administration of excessive arginine reverses nitrogen loss and immune suppression after trauma in rats, and healthy human volunteers consuming 30 g of oral supplements or arginine have shown significantly enhanced immunoreactivity of the lymphocytes of their peripheral blood. Calculations based on creatinine excretion show that 0.8 g of protein/kg body weight of the quality supplied by the usual American diet barely provides sufficient arginine for synthesizing the quantity of creatinine excreted daily in the urine of 70-kg adults. Human patients who often consume less than this amount of protein show a decline in creatinine excretion during illness; the decrease suggests that their intake of arginine is less than optimal. Recent studies of intraspecies and interspecies differences in responses to arginine reemphasize that dispensability or indispensability of arginine is a matter of definition and that growth and nitrogen balance data impose significant limitations on the drawing of far-reaching conclusions about the needs for arginine by mammalian adults including humans. Orotic acid excretion, immune responsiveness and circulating hormone levels are measures that should be evaluated for identifying when enhancement of arginine intakes might prove beneficial.
Modulation of immune function and weight loss by L-arginine in obstructive jaundice in the rat.
Br J Surg (ENGLAND) Aug 1994, 81 (8) p1199-201
Jaundiced surgical patients have a high incidence of postoperative complications. Many causative factors have been identified including cachexia and immune suppression. The amino acid L-arginine has anabolic and immunostimulatory properties. It was hypothesized that dietary supplementation with L-arginine would diminish the weight loss and immune suppression of obstructive jaundice. Sixteen male Wistar rats rendered jaundiced by bile duct ligation were allocated to two groups. The test group (n = 8) received drinking water supplemented with 1.8 percent L-arginine ad libitum and the control group (n = 8) received a solution of isonitrogenous glycine. Both groups had free access to standard chow. Body-weight, and fluid and food intake were recorded. After 21 days, delayed-type hypersensitivity to 2,4-dinitrofluorobenzene was assessed. Animals receiving L-arginine consumed more food than controls (mean(s.e.m.) 414(16) versus 360(13) g, P < 0.05) and lost less weight (mean(s.e.m.) proportion of initial body-weight lost 7.8(1.2) versus 14.8(1.4) percent, P < 0.05). The delayed-type hypersensitivity response was significantly greater in rats receiving L-arginine (mean(s.e.m.) increase in ear thickness 23.9(2.7) versus 9.4(2.1) percent, P < 0.05). In this animal model of obstructive jaundice dietary supplementation with L-arginine diminished both weight loss and immune suppression.
Can the length of hospital stay be influenced by enteral immunonutrition?
Anasthesiologie und Intensivmedizin (Germany), 1997, 38/3 (137-147)
The balance of current clinical data suggests that early enteral nutrition may influence infectious complications in the critically ill patients. Certain nutrients may affect organ function, independent of their general nutritional effects. Four of these nutrients are arginine, nucleotides, omega-3-fatty acids and glutamine. The target cells for the action of these nutrients appear to be T-lymphocytes and macrophages. An enteral nutrition enriched with such nutrients is called 'immunonutrition'. Recent evidence has suggested that an immunonutrition can have a beneficial effect on the prevention of infectious complications and SIRS, reduction of ventilator days, ICU- and hospital stay. This seems to be translated into a reduction in hospital charges. Beside a therapeutic approach with specific inhibitors and receptor antagonists the so called 'immunonutrition' seems to have a place in the therapy of the critically ill patient.
L-arginine restores dilator responses of the basilar artery to acetylcholine during chronic hypertension.
Hypertension (UNITED STATES) Apr 1996, 27 (4) p893-6
The objective of this study was to test the hypothesis that administration of L-arginine, a substrate for nitric oxide synthase, restores acetylcholine-induced dilatation of the basilar artery in chronically hypertensive rats. Basilar artery diameter was measured through a cranial window in anesthesized stroke-prone spontaneously hypertensive rats (SHRSP) and normotensive Wistar-Kyoto rats (WKY) aged 6 to 7 months (adult) and 12 months (older adult). Under control conditions, baseline basilar artery diameter was smaller in SHRSP (adult, 239 +/- 30 micron; older adult, 198 +/- 13 micron) (mean +/- SE) than in WKY (adult, 261 +/- 10 micron; older adult, 259 +/- 7 micron) (P <.05 versus SHRSP). Topical application of acetylcholine (10(-5) mol/L) produced dilatation of the basilar artery in WKY, which was impaired in both adult and older SHRSP (P <.05). Topical L-arginine (10(-3) mol/L for 30 minutes) did not affect responses to acetylcholine in adult SHRSP but enhanced vasodilatation in response to acetylcholine (10(-5) mol/L) in older SHRSP without affecting responses to sodium nitroprusside. In contrast, D-arginine did not affect acetylcholine-induced vasodilatation in older SHRSP. These results suggest that impaired dilatation of the basilar artery in response to acetylcholine in older SHRSP is restored toward normal by L-arginine, a substrate for nitric oxide synthase.
Dietary L-arginine attenuates blood pressure in mineralocorticoid-salt hypertensive rats.
Clin Exp Hypertens (UNITED STATES) Oct 1995, 17 (7) p1009-24
The present study was designed to investigate the influence of dietary L-arginine supplementation on blood pressure and on ex vivo vascular reactivity in mineralocorticoid-salt (DOCA-salt) hypertensive rats. Systolic blood pressure and heart rate were determined throughout the experimental period in unanaesthetized rats. Plasma and urine electrolyte levels were measured. Vasoconstrictor response to noradrenaline and vasodilator responses to acetylcholine and sodium nitroprusside were evaluated in the isolated perfused mesenteric vascular bed. DOCA-salt hypertensive rats were divided into 2 groups: a control group and a treated group receiving 0.8% L-arginine supplementation in drinking water. Dietary L-arginine supplementation attenuated systolic blood pressure in conscious DOCA-salt hypertensive rats, but did not modify heart rate. Plasma calcium and sodium concentrations and urinary magnesium excretion were decreased by L-arginine supplementation. Noradrenaline-induced vasoconstriction decreased and acetylcholine-induced vasodilatation increased, whereas sodium nitroprusside-induced vasodilatation was not modified, in the L-arginine-supplemented rats. It is concluded that dietary L-arginine supplementation in the diet lowers systolic blood pressure in DOCA-salt hypertensive rats, probably through vascular action.
Role of exogenous L-arginine in hepatic ischemia-reperfusion injury
Journal of Surgical Research (USA), 1997, 69/2 (429-434)
Plasma L-arginine is usually deficient immediately after hepatic reperfusion in orthotopic liver transplantation, which may also contribute to the occurrence of either hepatic ischemia-reperfusion injury or pulmonary hypertension. In this study, exogenous L-arginine was thus experimentally used to reverse the deficient status of the L-arginine/NO pathway. An in vivo model of 1 hr hepatic ischemia and reperfusion was thus tested in both rats (Experiment A) and pigs (Experiment B). In Experiment A, 10 mg/kg of L- arginine (group 1, n = 7), D-arginine (group 2, n = 7), or saline (group 3, n = 7) was administered through the portal vein. The hepatic tissue blood flow, at 20 min after reperfusion, improved in group 1 (70.7 plus or minus 7.0% of the preclamp levels) compared to groups 2 and 3. The serum glutamate oxaloacetate transaminase levels at 24 hr after reperfusion were also lower in group 1 (320 plus or minus 22.2 IU/L) than in either group 2 or group 3. The intrahepatic NO levels showed a temporal burst (>15,000 pA current) after reperfusion only in group 1. In Experiment B, 10 mg/kg of L-arginine (group 4, n 5), D-arginine (group 5, n= 5), or 10 ml of saline (group 6, n= 5) was administered through the portal vein. In group 4, the MPAP (mean pulmonary arterial pressure)/MAP (mean arterial pressure) was lower than that observed in groups 5 and 6. In conclusion, exogenous L-arginine administered from the portal vein was thus found to be effective in mitigating both portal hypertension and reperfusion injury by producing an increased amount of NO immediately after reperfusion.
Can the kidney prevent cardiovascular diseases?
Clin Exp Hypertens (UNITED STATES) Apr-May 1996, 18 (3-4) p501-11
The kidneys play an important role in the development of cardiovascular risk factors. It is well known that heavy proteinuria can induce hyperlipidemia, the uric acid is elevated in some renal deficiencies and that hypertension develops in most end stage renal diseases. In prehypertensive states, specially in subjects with a family history of hypertension, some hemodynamic changes take place, characterized by an increase in renal vasoconstriction with a reduction in renal plasma flow and an elevation of sodium reabsorption. The mechanisms for these alterations are not well understood, but an increase in intracytosolic calcium in vascular smooth muscle cells, a reduction in vasodilatory substances such as nitric oxide and an increased sympathetic nervous activity have been proposed. In normotensive subjects with two hypertensive parents a reduction in sodium diet, an increase in protein intake or in arginine diet could prevent established essential hypertension from developing. In borderline hypertension an early therapy with low doses of calcium antagonists, ACE inhibition or diuretics could be indicated. (31 Refs.)
L-arginine prevents corticotropin-induced increases in blood pressure in the rat.
Hypertension (UNITED STATES) Feb 1996, 27 (2) p184-9
In this study we examined whether L-arginine treatment could prevent corticotropin (ACTH)-induced increases in blood pressure in the Sprague-Dawley rat. Sixty rats were randomly divided into six groups (n = 10): sham injection, ACTH injection (0.5 mg/kg per day in divided doses), L-arginine (0.6%) in food plus sham injection, L-arginine plus ACTH treatment, D-arginine (0.6%) in food plus sham injection, and D-arginine plus ACTH. Systolic pressure, water intake, urine volume, body weight, plasma and urinary electrolytes, and serum corticosterone concentrations were measured. ACTH increased systolic pressure (from 127 +/- 2 to 165 +/- 6 mm Hg, P < .001), water intake, and urine volume and decreased body weight body weight. L-Arginine reduced ACTH-induced blood pressure rises (130 +/- 3 mm Hg, P < .001) but had no effect on blood pressure in sham-treated rats. D-Arginine did not affect blood pressure in sham-treated rats, and systolic pressure in D-arginine+ACTH-treated rats was similar to that of ACTH-treated rats. L-Arginine decreased serum corticosterone concentrations in sham-treated rats (424 +/- 42 versus 238 +/- 25 ng/mL, P < .01), but D-arginine had no effect. However, both drugs decreased serum corticosterone concentrations in ACTH-treated rats (1071 +/- 117 versus 739 +/- 95 and 695 +/- 72 ng/mL for L- and D-arginine, respectively; both P < .05). As L-arginine but not D-arginine prevented ACTH-induced increases in blood pressure in Sprague-Dawley rats and both L- and D-arginine reduced serum corticosterone concentrations in ACTH-treated rats, the effects of L-arginine in preventing ACTH-induced hypertension were not simply a consequence of decreased corticosterone secretion.
Improvement of cardiac output and liver blood flow and reduction of pulmonary vascular resistance by intravenous infusion of L-arginine during the early reperfusion period in pig liver transplantation.
Transplantation (UNITED STATES) May 15 1997, 63 (9) p1225-33
BACKGROUND: The release of liver arginase after orthotopic liver transplantation (OLT) causes a deficiency of L-arginine and nitrite in the plasma. This deficiency is possibly related to pulmonary hypertension and reduced hepatic blood flow, which are commonly observed in the immediate reperfusion period. The aim of this study was to evaluate the impact of L-arginine supplementation on metabolic and hemodynamic parameters during liver reperfusion after OLT in pigs. METHODS: Thirteen pig OLTs (control group, n=6; arginine group, n=7) were performed by a standard technique. Cold ischemic time was 20 hr. L-Arginine was infused at a dosage of 500 mg/kg body weight into the donor pigs (30 min before liver explantation) and also into the recipients (over a period of 3 hr from the beginning of the reperfusion period). At the end of the experimental study, the pigs were killed with an overdose of potassium. RESULTS: In the control group, liver revascularization increased plasma arginase concentrations (+615%) and reduced plasma levels of L-arginine (-87%), nitrite (-82%), and nitrate (-53%). Infusion of L-arginine increased plasma levels of L-arginine from 94+/-21 micromol/L to 1674+/-252 micromol/L (P<0.001), L-ornithine from 46+/-8 micromol/L to 2215+/-465 micromol/L (P<0.001), and L-citrulline from 58+/-8 micromol/L to 116+/-34 micromol/L (P<0.001), but had no effect on plasma levels of nitrite and nitrate. Administration of L-arginine in the donor pigs did not produce any systemic or organ-specific hemodynamic alterations. Infusion of L-arginine into the recipient pigs improved cardiac performance (increase in heart rate [+61%, P=0.017] and cardiac index [+53%, P=0.005], reduction in pulmonary capillary wedge pressure [-54%, P=0.014]). Moreover L-arginine infusion increased oxygen consumption (+65%, P=0.003), reduced pulmonary vascular resistance index (P=0.001), stimulated portal venous blood flow (P=0.014), and elevated body temperature during the reperfusion period (P=0.007). CONCLUSIONS: From these data, we conclde that the infusion of L-arginine during OLT improves the hemodynamic performance of the heart, lung, and liver.
Renal response to L-arginine in salt-sensitive patients with essential hypertension.
Hypertension (UNITED STATES) Mar 1996
This study examined whether disturbances in nitric oxide formation contribute to renal dysfunction in salt-sensitive essential hypertensive patients. We evaluated the effects of intravenous administration of L-arginine (500 mg/kg given over 30 minutes) on systemic and renal hemodynamics in 23 patients with mild essential hypertension during 1 week of a low NaCl diet (50 mmol/d) followed by 1 week of a high NaCl diet (340 mmol/d). Patients were classified as salt sensitive (n=10) or salt resistant (n=13) based on salt-induced changes in their blood pressures. Salt loading increased renal vascular resistance but not renal plasma flow in salt-sensitive patients. The L-arginine-induced renovascular relaxation was significantly reduced by a high NaCl diet (renal vascular resistance: low NaCl -12.4 +/- 2.3% versus high NaCl -7.1 +/- 1.8%, P < .001) in salt-sensitive patients, whereas it was unchanged in salt-resistant patients. The increase in plasma cGMP in response to L-arginine was also reduced by a high NaCl diet in the salt-sensitive patients (low NaCl 49 +/- 7% versus high NaCl 36 +/- 8%, P < .05) but not in the salt-resistant patients (low NaCl 51 +/- 6% versus high NaCl 58 +/- 6%). These findings suggest that NaCl loading in salt-sensitive patients with mild essential hypertension reduces the ability of L-arginine to produce nitric oxide in the endothelium of the renal vasculature.
Contrasting effect of antihypertensive treatment on the renal response to L-arginine
Hypertension (UNITED STATES) Dec 1995
We assessed the renal hemodynamic response to L-arginine infusion (30 g within 60 minutes) in normotensive subjects, patients with never-treated essential hypertension, and hypertensive patients controlled by long-term (more than 2 years) treatment with or without an angiotensin-converting enzyme inhibitor. The renal vasodilator response to L-arginine observed in normotensive subjects (15 +/- 4% increase in effective renal plasma flow) was abolished in untreated hypertensive patients and restored only in the group treated by angiotensin-converting enzyme inhibition. In the whole population a positive correlation between the change in effective renal plasma flow and the change in urinary cGMP was obtained. It is suggested that abnormalities of the renal nitric oxide pathway not corrected by increased availability of L-arginine and reversible only on long-term treatment by angiotensin-converting enzyme inhibition may underlie the abnormal renal resistance observed in essential hypertension.
Endothelial dysfunction: Clinical implications.
Drexler H. Germany Progress in Cardiovascular Diseases (USA), 1997, 39/4 (287-324)
The endothelium is involved in the control of vascular tone and homeostasis. Risk factors for arteriosclerosis, as well as other conditions have been shown to be associated with a dysfunctional endothelium. Clinically, endothelial function and dysfunction have been mostly evaluated by the assessment of endothelial dependent relaxation, for example in response to acetylcholine or increase inflow. The functional implications of endothelial dysfunction in cardiovascular disease are not well defined, but recent clinical trials have suggested that endothelial dysfunction may affect vascular tone and organ perfusion particularly during stress situations such as exercise. Moreover, endothelial dysfunction may represent an early event in the development of arteriosclerosis. Therefore, recent clinical studies have been performed to restore normal endothelial function in patients, using interventions such as L-arginine, lipid lowering drugs, vitamin C, other antioxidants, or exercise.
L-Arginine reduces lipid peroxidation in patients with diabetes mellitus.
Free Radic Biol Med (UNITED STATES) 1997, 22 (1-2) p355-7
A current concept for the development of diabetic long-term complications is the involvement of oxidative stress, as, e.g., lipid peroxidation, in the diabetic state. Data published recently show also oxidative damage to DNA, which might be one factor for accelerated aging and diabetic microangiopathy. In our study we tested the hypothesis that L-arginine can reduce lipid peroxidation in patients with diabetes. We performed a blind placebo controlled study with crossing over two treatment periods for 3 months. Thirty patients with diabetes mellitus were randomly assigned to treatment group A (first treatment then placebo) and B (first placebo then treatment). Treatment consisted of two daily dosages of 1 g L-arginine free base. Lipid peroxidation as reflected by malondialdehyde was evaluated in urine using a standard HPLC assay. After 3 months of treatment there was a significant reduction in malondialdehyde levels in group A (p < .0032), whereas there was no difference compared to the baseline values after three months of placebo treatment in group B (p < .97). After crossing over, there was a significant reduction in malondialdehyde levels in group B (p < .0002). Group A showed a significant increase in malondialdehyde levels (p < .0063) returning to baseline values. L-Arginine treatment was able to reduce the lipid peroxidation product malondialdehyde. This provides evidence that treatment with L-arginine may counteract lipid peroxidation and thus reduce microangiopathic long-term complications in diabetes mellitus.
Short-term oral administration of L-arginine reverses defective endothelium-dependent relaxation and cGMP generation in diabetes.
Eur J Pharmacol (NETHERLANDS) Dec 19 1996, 317 (2-3) p317-20
In the present study, we evaluated whether acute dietary supplementation with L-arginine in vivo could reverse the defective endothelium-dependent relaxation in diabetic blood vessels assessed ex vivo. At 8 weeks of diabetes, streptozotocin-induced diabetic rats were given 1.25% L-arginine in drinking water 3 days prior to isolation of aortic rings for evaluation ex vivo. Plasma arginine concentration was reduced by diabetes but restored to normal in diabetic rats receiving dietary L-arginine. In norepinephrine-contracted rings, relaxation to acetylcholine but not to nitroglycerin was reduced by diabetes. Dietary treatment with L-arginine restored relaxation to acetylcholine without altering relaxation to nitroglycerin and restored the defect in acetylcholine-stimulated cGMP generation. These data suggest that the substrate for nitric oxide synthesis by the endothelium is likely to be limited in diabetes but can be overcome by dietary supplementation with L-arginine.
Metformin improves hemodynamic and rheological responses to L-arginine in NIDDM patients
Diabetes Care (UNITED STATES) Sep 1996, 19 (9) p934-9
OBJECTIVE: The endothelium plays a pivotal role in the regulation of vascular tone by releasing nitric oxide (NO). Increased availability of L-arginine, the natural precursor of NO, induces vasodilatation and inhibits platelet activity. We studied the effect of metformin on hemodynamic and rheological responses to L-arginine in patients with NIDDM. RESEARCH DESIGN AND METHODS: Ten newly diagnosed NIDDM patients with mild fasting hyperglycemia (7.5 +/- 0.3 mmol/l) and without evidence of both micro- and macrovascular complications were investigated. They received an intravenous infusion of L-arginine (1 g/min for 30 min) with evaluation of plasma glucose and insulin, systolic (sBP) and diastolic (dBP) blood pressure, heart rate and plasma catecholamines, platelet aggregation, and blood viscosity and filterability. The L-arginine test was repeated after an 8-week treatment with metformin (850 mg b.i.d.). RESULTS: Metformin treatment significantly reduced basal fasting plasma glucose, HbA1c, and platelet aggregation to ADP (P < 0.05); the other parameters did not change. During pretreatment test, L-arginine infusion decreased sBP (from 137 +/- 4.1 to 129 +/- 4.5 mmHg, P < 0.01) and dBP (from 79 +/- 1.9 to 75 +/- 1.2 mmHg, P < 0.01) without affecting heart rate or plasma catecholamines. Both platelet aggregation and blood viscosity showed significant decrements after L-arginine, while blood filterability did not change. After metformin treatment, the decrease in blood pressure after L-arginine infusion was significantly enhanced, with a maximal decrease of sBP of 12 +/- 3.4 mmHg (8 +/- 2.5 mmHg pretreatment, P < 0.05) and dBP of 9.5 +/- 2.4 mmHg (4.5 +/- 1.9 mmHg pretreatment, P < 0.01). Heart rate, plasma norepinephrine levels, and blood filterability also rose significantly (P < 0.05-0.01). The decrease in both platelet aggregation and blood viscosity after L-arginine was significantly amplified after metformin. CONCLUSIONS: We conclude that L-arginine infusion in newly diagnosed NIDDM patients without vascular comlications produces relevant hemodynamic and rheological changes, which are amplified by an 8-week treatment with metformin. Whether these vascular effects of metformin will improve the poor cardiovascular outlook of the diabetic patient is still unknown.
Impairment of coronary blood flow regulation by endothelium-derived nitric oxide in dogs with alloxan-induced diabetes.
J Cardiovasc Pharmacol (UNITED STATES) Jul 1996, 28 (1) p60-7
Diabetes mellitus is a major cause of ischemic coronary artery disease. Endothelial dysfunction is implicated in the pathogenesis of diabetic vascular disease. To examine coronary blood flow (CBF) regulation with endothelium-derived nitric oxide (EDNO) in the diabetic state, we compared the effects of both acetylcholine (ACh) and adenosine (Ado) on left circumflex coronary artery (LCX) blood flow in 12 vehicle-treated and 21 dogs made diabetic with alloxan anesthetized with pentobarbital. All dogs were pretreated with aspirin to inhibit endogenous prostaglandins. None of the hemodynamic parameters were significantly different in the two groups. The percent change in coronary vascular resistance (CVR) after ACh (100 ng/kg) infusion was significantly attenuated in diabetic dogs (-56.5 +/- 1.4%) as compared with vehicle-treated dogs (-64.5 +/- 1.2%) (p < 0.01), whereas the effect of Ado (1 microgram/kg) was not different between the two groups (-71.1 +/- 1.5% in vehicle, -67.0 +/- 1.3% in diabetes). After infusion of incremental doses of NG-nitro-L-arginine methyl ester (L-NAME) 10(-5)-10(-3)M, the effect of ACh was progressively inhibited in both groups and was different no longer between the two groups after the maximal dose. L-Arginine (L-ARG), but not D-ARG, significantly restored the effect of ACh in diabetic dogs but did not affect vehicle-treated dogs. The effect of Ado did not change after L- and D-ARG administration. Cu, Zn-superoxide dismutase (Cu, Zn-SOD) had no effect on any of the effects of ACh and Ado in diabetic dogs. Regulation of CBF with EDNO is impaired in dogs with alloxan-induced diabetes, and this impairment is partially restored by L-ARG.
Involvement of the L-arginine-nitric oxide pathway in hyperglycaemia-induced coronary artery dysfunction of isolated guinea pig hearts.
Eur J Clin Invest (ENGLAND) Aug 1996, 26 (8) p707-12
The effects of hyperglycaemia and L-arginine on flow-induced reduction of coronary artery resistance were investigated in isolated guinea pig hearts. In the presence of indomethacin, hyperglycaemia caused an increase in flow-induced vasodilatation (P < 0.05). Hyperosmotic controls failed to mimic this effect. Addition of L-arginine strongly enhanced this effect. Addition of D-arginine failed to mimic the effects of L-arginine. The effect of L-arginine was abolished by co-administration of NG-nitro-L-arginine. In the absence of indomethacin and L-arginine, the effect of hyperglycaemia was blunted, suggesting the formation of vasoconstrictive prostanoids. Addition of L-arginine again resulted in a significant increase in flow-induced vasodilatation. In conclusion our results suggest that increased flow-induced vasodilatation under hyperglycaemic conditions depends on an adequate supply of L-arginine to maintain sufficient formation of nitric oxide.
Deficient nitric oxide responsible for reduced nerve blood flow in diabetic rats: effects of L-NAME, L-arginine, sodium nitroprusside and evening primrose oil.
Br J Pharmacol (ENGLAND) May 1996, 118 (1) p186-90
1. This study examined the potential role of impaired nitric oxide production and response in the development of endoneurial ischaemia in experimental diabetes. Rats were anaesthetized (Na pentobarbitone 45 mg kg-1, diazepam 2 mg kg-1) for measurement of sciatic nerve laser Doppler flux and systemic arterial pressure. Drugs were administered into the sciatic endoneurium via a microinjector attached to a glass micropipette.
2. In two separate studies comparing diabetic rats (streptozotocin-induced; 8-10 wk duration) with controls, nerve Doppler flux in diabetic rats (Study 1, 116.6 +/- 40.4 and Study 2, 90.1 +/- 34.7 (s.d.) in arbitrary units) was about half that measured in controls (219.6 +/- 52.4 and 212.8 +/- 95.5 respectively; P < 0.005 for both). There were no significant differences between the two in systemic arterial pressure.
3. Inhibition of nitric oxide production by microinjection of 1 nmol L-NAME into the endoneurium halved flux in controls (to 126.3 +/- 41.3 in Study 1 and 102.1 +/- 38.9 in Study 2; both P < 0.001), with no significant effect in diabetic rats, indicating markedly diminished tonic nitric oxide production in the latter. D-NAME was without effect on nerve Doppler flux.
4. L-Arginine (100 nmol), injected after L-NAME, markedly increased flux in controls (by 65.8% (P < 0.03) and 97.8% (P < 0.01) in the two studies) and by proportionally similar amounts in diabetic rats [75.8% (P < 0.001) and 60.2% (P < 0.02)]. The nitro-donor, sodium nitroprusside (SNP; 10 nmol) had similar effects to L-arginine in both groups (increases of 66.0% in controls and 77.5% in diabetics; both P < 0.002).
5. A second diabetic group, treated with evening primrose oil performed exactly like control rats in respect of responses to L-NAME, L-arginine and SNP.
6. These findings implicate deficient nitric oxide in nerve ischaemia of diabetes and suggest correction thereof as a mechanism of action of evening primrose oil.