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¹ Laboratory of Sports Physiology, Research and Education Center for Winter Sports, Hokkaido University of Education, Ainosato 5-3, Kita-ku, Sapporo, Hokkaido, 002-8502, Japan

	Abstract

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This study was designed to examine whether dietary L-arginine supplementation modulates exercise-induced angiogenesis and vascular endothelial growth factor (VEGF) expression in female Wistar rats. Exercise training (running) lasted for 6 weeks at 25 m min^–1 on a 20% gradient for 10–60 min day^–1. Rats in the L-arginine-treated groups drank water containing 4% L-arginine. Histochemical identification of capillary profiles showed that training with L-arginine significantly increased the capillary/fibre (C/F) ratio in the subendocardium of the left ventricle, whereas training alone did not. Because of a significantly higher fibre cross-sectional area, a concomitant, but not significant, decrease in capillary density was also observed. In the hindleg muscles, training with L-arginine significantly increased the C:F ratio, although the degree of change was the same as that observed after training alone. Western blot analysis showed that training with L-arginine significantly increased VEGF protein expression by 1.7-fold in the left ventricle, while the increase with training alone was insignificant. In the soleus muscle, although VEGF protein expression was elevated insignificantly after training (2.8-fold), training with L-arginine significantly increased the protein levels (3.8-fold). Tissue endothelial nitric oxide synthase protein levels did not changed after either training or L-arginine treatment. The present results suggest that L-arginine supplementation causes additional effects on exercise-induced angiogenesis in the rat heart by promoting VEGF expression.

(Received 31 May 2005; accepted after revision 6 July 2005; first published online 7 July 2005)
Corresponding author J. Suzuki: Research and Education Center for Winter Sports, Hokkaido University of Education, Ainosato 5-3, Kita-ku, Sapporo, Hokkaido 002-8502, Japan. Email: suzuki{at}sap.hokkyodai.ac.jp

	Introduction

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Endurance exercise training induces a marked increase in capillarity in skeletal (e.g. Hermansen & Wachtlova, 1971) and cardiac muscles (e.g. Tomanek, 1970). Reduced oxygen tension (Yang et al. 1991; Gustafsson et al. 1999) and/or mechanical factors, such as shear stress and tissue stretching (Ziada et al. 1984; Milkiewicz et al. 2001), may stimulate the expression of peptide angiogenic factors such as vascular endothelial growth factor (VEGF). VEGF is thought to be an important regulator of angiogenesis during exercise training (Hoffner et al. 2003; Jensen et al. 2004; Kraus et al. 2004; Wittwer et al. 2004), as well as during embryonic development (Jakeman et al. 1993), wound healing (Brown et al. 1992) and tumour growth (Celec et al. 2005).

VEGF gene expression is increased in response to hypoxia (Goldberg & Schneider, 1994). Recently, several other factors, including nitric oxide (NO), have been shown to regulate VEGF gene expression. Regulation of VEGF gene expression induced by NO has been demonstrated in numerous cell types, including vascular smooth muscle cells (Dulak et al. 2000). Inhibition of nitric oxide synthase (NOS) abolished exercise-induced capillary angiogenesis in skeletal muscle (Hudlicka et al. 2000). Neonatal endothelial NOS-deficient (eNOS^–/–) mice had significantly less myocardial capillary density and VEGF mRNA expression than neonatal wild-type mice (Zhao et al. 2002). Moreover, the mRNA level and protein expression of VEGF in ischaemic skeletal muscle showed no apparent difference between eNOS^–/– and control wild-type mice, whereas angiogenesis in response to ischaemia was severely inhibited in eNOS^–/– mice (Murohara et al. 1998). These findings suggest that endogenous NO plays a critical role in exercise-induced angiogenesis in skeletal muscles, as well as in the heart.

Chronic treatment with L-arginine, a substrate for NOS, enhanced exercise-induced endothelial NO synthesis and aerobic capacity (Maxwell et al. 2001). Moreover, L-arginine is known to stimulate growth hormone (Korbonits et al. 1996) and insulin release (Giugliano et al. 1997). As both growth hormone and insulin are known to stimulate VEGF expression (Miele et al. 2000; Kobayashi & Kamata, 2002), increases in the levels of these hormone may contribute to VEGF expression during exercise training. It therefore appears possible that L-arginine supplementation causes additional effects on exercise-induced angiogenesis.

In the present study, experiments were designed to examine the combined effects of exercise training and L-arginine supplementation on capillary geometry and VEGF protein expression in left ventricle and hindleg muscles. The present results demonstrate that L-arginine administration caused additional effects on exercise-induced angiogenesis by promoting VEGF expression in the left ventricle.

	Methods

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This study was approved by the Animal Care and Use Committee of Hokkaido University of Education and performed in accordance with the Guiding Principles for the Care and Use of Animals in the Field of Physiological Sciences of the Physiological Society of Japan.

Animals, experimental conditions and muscle samples

Twenty-eight, 10-week-old female Wistar rats were purchased from Clea Japan Inc. (Tokyo, Japan). After the rats were fed for 2 weeks to allow adaptation to the new environment, they were randomly divided into the following four groups according to treatment and exercise conditions: non-treated sedentary (Cnt, n = 7); non-treated training (TR, n = 8); L-arginine-treated sedentary (Arg, n = 7); and L-arginine-treated training (ArTR, n = 8). The L-arginine administration and exercise training were started at 12 weeks of age and lasted for 6 weeks. During the second week of the adaptation period, all rats were subjected to treadmill running for 2 min day^–1 for 3 days. All rats were housed under controlled temperature conditions (24 ± 1°C) and a relative humidity of approximately 50%. Lighting (07.00–19.00 h) was controlled automatically. All rats were given commercial laboratory chow (CE-2, Clea Japan Inc.) ad libitum. Rats in the non-treated groups drank tap water, whereas rats in the L-arginine-treated groups drank water containing 4% (w/v) L-arginine (A5006, Sigma-Aldrich, St Louis, MO, USA). The average L-arginine intake, estimated from water intake, was 4.57 g kg ^–1 day^–1. Because the rats were housed two to three per cage, the water intake per individual rat was estimated from the water intake per cage.

Rats in the training groups were subjected to treadmill running using a rodent treadmill (KN-73, Natsume Co. Tokyo, Japan). The rats ran 10 min day^–1 at 25 m min^–1 with a 20% gradient on the first day of training. Then, the duration was increased by 3 min day^–1. The speed and gradient were maintained throughout the remaining training period. At the end of the 4th week, the rats were running continuously for 1 h day^–1. This final duration was maintained throughout the remaining training period. The training session was carried out 5 days week^–1.

The animals were not exercised for at least 48 h prior to killing. Under light anaesthesia with ether, the rats were anaesthetized with pentobarbital sodium (50 mg kg^–1 I.P.). A toe-pinch response was used to validate adequate anaesthesia. Then, the left soleus (SOL) and plantaris (PL) muscles were excised and weighed. The muscles were fixed at the length measured when the knee joint was maximally extended, and the tibiotarsal joint was fixed at 90 deg. The tissues were placed in embedding medium, an optimal cutting temperature (O.C.T.) compound (Sakura Finetechnical Co., Ltd, Tokyo, Japan), and rapidly frozen in isopentane cooled to its melting point (–160°C) with liquid nitrogen. For biochemical analyses, the remaining muscles (i.e. the right side, and the apex half of the left ventricle (LV)) were excised and frozen in liquid nitrogen. The LV was excised and weighed. The apex half of the LV was used for biochemical analyses, while the other half was used for histochemical analyses. All LV samples were treated in a similar fashion. The tissue samples were stored at –80°C until analyses.

Histological analyses

Tissue cross-sections were obtained by using a cryotome (CM-1500; Leica Japan, Tokyo, Japan) at –20°C. To determine capillary profiles, the sections were double-stained for alkaline phosphatase (AP) and dipeptidyl peptidase IV (DPP IV) in the capillary endothelium. Although the original staining protocol was described by Lojda (1979), a slightly modified protocol was used, as described previously(Suzuki, 2004). The validity of this double-staining method for differentiation of arteriolar and venular capillary portions has been confirmed previously (Lojda, 1979; Koyama et al. 1998). The images of incubated sections were digitized using a digital microscope camera (PDMC le, Polaroid, Cambridge, MA, USA) attached to a light microscope (BX-50, Olympus, Tokyo, Japan) and were stored on computer disc. The capillary profiles were identified as either arteriolar (blue), venular (red) or intermediate (violet). Non-overlapping microscopic fields were selected at random from each muscle sample when the microscope was set to phase-contrast; that is, the observer did not observe the colour of capillaries. During the measurements, the observer was blind as to the source (groups) of each slide. Morphological analyses were obtained from the SOL, mixed fibre (PLm) and predominantly fast fibre (PLs) portions of the PL, and subendocardium and subepicardium of the LV. Total cross-sectional areas used for each morphological measurement were up to 0.69 mm² per tissue region.

Western blot analysis

Tissue homogenates were obtained from approximately 50 mg frozen tissue homogenized for three interrupted 15-s bursts with Polytron homogenizer (set at 15 000 r.p.m.) in ice-cold medium (100 mM potassium phosphate buffer (pH 7.2), 0.1% Triton X-100, 1 mM dithiothreitol and 5% (v/v) protease inhibitor cocktail (Sigma-Aldrich)). After centrifugation at 1500 g for 15 min at 0°C, the supernatant was used for protein analysis. A portion (20 µg) of protein was fractionated by SDS-PAGE on 7.5 or 12% (w/v) polyacrylamide gels, electrically transferred to a polyvinylidene fluoride (PVDF) membrane and blocked with 1% (w/v) bovine serum albumin for 1 h. The blots were exposed to the specific primary antibodies (Santa Cruz Biotech, Santa Cruz, CA, USA) against VEGF (1: 1000) or eNOS (1: 1000) diluted in blocking solution. After the blots were incubated with an AP-conjugated secondary antibody (Santa Cruz, 1: 5000) diluted in blocking solution, the required proteins were detected with a 5-bromo-4-chloro-3-indolyl phosphate/Nitro blue tetrazolium (BCIP/NBT) reaction. The membranes were scanned (GT-8200UF, Seiko Epson, Tokyo, Japan) and densities of bands were quantified using Image J public domain software (written by W. Rasband, National Institutes of Health, USA). Negative controls without primary or secondary antibodies were run to validate the results.

Biochemical analyses

Using the muscle homogenate aliquots, metabolic enzyme activities were measured. The activity of ß-hydroxyacyl-CoA-dehydrogenase (HAD) was assayed according to the method of Bass et al. (1969). The activity of citrate synthase (CS) was determined according to the methods of Srere (1969). All measurements were carried out at 25°C with a spectrophotometer (U-2001, Hitachi, Tokyo, Japan).

Statistical analyses

All values are expressed as means ± S.E.M. Using the Kolmogrov-Smirnoff test, the distribution of all parameters was tested to determine whether it was compatible with a normal distribution. Two-way analysis of variance (ANOVA) was used to test the effects of training, drugs and their interactions. If the two-way ANOVA was significant, differences among the four groups were analysed using one-way ANOVA and Fisher's PLSD post hoc test. Differences were considered to be statistically significant at P < 0.05.

	Results

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Mean body mass at the end of treatment was not significantly different among the four groups (Table 1). Training with or without L-arginine significantly increased the tissue mass and the tissue mass/body mass ratio in all organs examined in the present study (P < 0.05). After L-arginine treatment alone, the mass of the hindleg muscles was enhanced, whereas the mass/body weight ratio did not change. Dietary L-arginine supplementation did not affect exercise-induced hypertrophy, whereas L-arginine caused modest hypertrophy in hindleg muscles.

View larger version (161K): [in this window] [in a new window]   Figure 1.  Micrographic images of the subendocardium stained with alkaline phosphatase and dipeptidylpeptidase IV A, non-treated sedentary group. B, trained group. C, L-arginine treated group. D, L-arginine-treated trained group. Types of capillaries cannot be distinguished because they are grey scale images. All images are at the same magnification.

View larger version (32K): [in this window] [in a new window]   Figure 2.  The capillary/fibre ratio of arteriolar, intermediate and venular capillaries All values are represented as means. *, # and significantly different from the control (Cnt), training (TR) and L-arginine-treated (Arg) groups, respectively, at P < 0.05. ArTR, training and L-arginine-treated.

View larger version (18K): [in this window] [in a new window]   Figure 3.  Western blot analysis demonstrating effects of exercise training or L-arginine supplementation on VEGF and eNOS protein expressions All values are represented as means ± S.E.M. *Significantly different from the control group at P < 0.05. Cnt, control; TR, training; Arg, L-arginine-treated; ArTR, training and L-arginine-treated.

View larger version (18K): [in this window] [in a new window]   Figure 4.  Enzyme activities in the soleus and plantaris muscles All values are represented as means ± S.E.M. *, # and significantly different from the control (Cnt), training (TR) and L-arginine-treated (Arg) groups, respectively, at P < 0.05. ArTR, training and L-arginine-treated, CS, citrate synthase; HAD, ß-hydroxyacyl-CoA-dehydrogenase.

	Discussion

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In the present study, exercise training with and without L-arginine administration caused marked angiogenesis in hindleg muscles. Although exercise-induced angiogenesis can be induced by several growth factors, hormones and cytokines (Prior et al. 2004; Lloyd et al. 2005), VEGF is thought to be an important regulator (Hoffner et al. 2003; Celec & Yonemitsu, 2004; Jensen et al. 2004; Kraus et al. 2004; Wittwer et al. 2004; Lloyd et al. 2005). VEGF mRNA is induced by hypoxia in cultured cells (Shweiki et al. 1992) and in skeletal muscles in vivo (Breen et al. 1996). Reduced oxygen tension in active muscles may stimulate the cellular oxygen-sensing mechanisms (i.e. sensing low oxygen), stabilizing hypoxia inducible factor (HIF)-1 and allowing cells to adapt to hypoxia (Goldberg et al. 1987). HIF-1 is necessary for the hypoxia-induced VEGF expression (Forsythe et al. 1996). Exercise-induced increase in VEGF mRNA was correlated to the exercise-induced changes in HIF-1 and -1ß mRNA (Gustafsson et al. 1999). Moreover, mechanical factors during exercise, such as shear stress and tissue stretching (Ziada et al. 1984; Milkiewicz et al. 2001), may stimulate the expression of VEGF. Chronic prazosin treatment caused 4-fold increase in shear stress and increased VEGF protein expression in rat skeletal muscle (Milkiewicz et al. 2001). A recent study showed that shear stress activated the VEGFR-2 pathway, independent of VEGF (Wang et al. 2004). Recent studies have shown that NO is both an upstream and downstream mediator of VEGF-dependent angiogenesis (Dulak & Jozkowicz, 2003; Kimura & Esumi, 2003). Zhao et al. (2002) showed that myocardial capillary density and VEGF mRNA expression were markedly lower in eNOS^–/– than in wild-type mice. Inhibition of NOS attenuated the exercise-induced increase in VEGF mRNA (Gavin et al. 2000), VEGFR-1 mRNA (Gavin & Wagner, 2002) and in protein levels of VEGF and VEGFR-2 (Milkiewicz et al. 2005) in rat skeletal muscles. Moreover, NOS inhibition abolished capillary angiogenesis induced by chronic electrical stimulation in rat skeletal muscle (Hudlicka et al. 2000; Milkiewicz et al. 2005). Thus, endogenous NO produces capillary angiogenesis by promoting the VEGF system in both skeletal and cardiac muscles.

In the present study, however, expression of eNOS protein did not change after either training or L-arginine administration (Fig. 3). Chronic exercise training, with a greater intensity than the present study, increased the expression of eNOS protein (Balon & Nadler, 1997; Vassilakopoulos et al. 2003). In contrast, training with an intensity lower than in the present study, decreased eNOS protein expression (Iemitsu et al. 2000). Thus, eNOS protein expression after exercise training may depend on the intensity of daily exercise.

Effects of L-arginine supplementation

The present study showed that training itself did not cause significant cardiac angiogenesis, whereas training with L-arginine supplementation caused exercise-induced cardiac angiogenesis. Although the effects of exercise training on cardiac capillaries have been intensively studied, controversial results have been reported (Tomanek, 1970; Tharp & Wagner, 1982; Jacobs et al. 1984; Hudlicka et al. 1992; Koyama et al. 1998). Most studies that have shown cardiac capillary growth induced by exercise training have been based on relatively young animals. In rats, exercise training beginning at younger than 5 weeks of age caused significant capillary angiogenesis, whereas that beginning at older than 14 weeks of age failed to cause capillary growth (Tomanek, 1970; Jacobs et al. 1984; Koyama et al. 1998). Thus, in the present study, the training started at 12 weeks of age did not induce significant capillary angiogenesis in cardiac muscle. However, L-arginine administration caused additional effects on exercise-induced capillary angiogenesis, possibly by promoting VEGF expression.

Expression of VEGF mRNA was increased after a single acute bout of exercise (Breen et al. 1996; Richardson et al. 1999), whereas chronic exercise training attenuated its expression in response to acute exercise in human skeletal muscle (Richardson et al. 2000). In rat skeletal muscle, expression of VEGF protein at capillary sites was significantly increased on the 6th and 10th days of the training (Suzuki, 2004), whereas after 5 weeks of training it still showed a high value, but was not significantly different from a sedentary control (Suzuki, 2002). In the present study, the VEGF level was still markedly higher after 6 weeks of training with L-arginine (Fig. 3). Therefore, combined effects of exercise and L-arginine supplementation can sustain VEGF expression for longer periods.

Although in the present study, vessel diameter and blood flow were not measured, oral L-arginine supplementation at the same amount (gram per body mass) as used in the present study caused arteriolar enlargement and increased blood flow in the cerebrum of stroke-prone spontaneously hypertensive rats (Noguchi et al. 1999). Daily handgrip training and L-arginine administration caused an additional effect on acetylcholine (Ach)-induced hyperemia (Hambrecht et al. 2000). However, infusion of L-arginine increased Ach-induced vasodilatation, although it did not affect exercise-induced vasodilatation (Kubota et al. 1997). Thus, it is unlikely that L-arginine supplementation facilitates flow-induced VEGF expression during exercise in the present study.

L-arginine is known to stimulate growth hormone (Korbonits et al. 1996), insulin (Giugliano et al. 1997) and insulin-like growth factor (IGF)-1 release (Visser & Hoekman, 1994). Blood hormone levels were not measured in the present study. The significant increase in muscle mass after L-arginine supplementation (Table 1) raises the possibility that L-arginine promotes protein synthesis in hindleg muscles. As both growth hormone and insulin are known to stimulate VEGF expression (Miele et al. 2000; Kobayashi & Kamata, 2002), increases in these hormone levels may contribute to enhanced VEGF expression observed after training with L-arginine administration.

Because supplementation of L-arginine attenuated hypoxia/reoxygenation-induced injury, L-arginine is thought to be an antioxidant (Akisu et al. 2002). However, a recent study showed that L-arginine did not have an antioxidant effect, even though it improved exercise capacity in patients with congestive heart failure (Bednarz et al. 2004). Thus, L-arginine supplementation may not attenuate exercise-induced production of reactive oxygen species, which have been reported to stimulate VEGF production through the phosphatidylinositol 3-kinase/Akt pathway (Kosmidou et al. 2001).

Infusion of L-arginine increased plasma growth hormone (Korbonits et al. 1996) and glucagon levels (MacAllister et al. 1995). Because these hormones stimulate lipolysis, L-arginine supplementation may possibly affect fat metabolism in skeletal muscle. However, in the present study, L-arginine did not enhance enzyme activity of ß-oxidation (Fig. 4). Thus, L-arginine administration may not facilitate fatty acid utilization in hindleg muscles during exercise.

In summary, the present study demonstrated that L-arginine supplementation caused additional effects on exercise-induced capillary angiogenesis in the rat left ventricle. The combined effect of exercise and L-arginine induced marked VEGF expression in both heart and skeletal muscle. The present results suggest that exercise training with L-arginine administration causes capillary angiogenesis in the rat heart by promoting VEGF expression.

	References

Top Abstract Introduction Methods Results Discussion References

 
Akisu M, Ozmen D, Baka M, Habif S, Yalaz M, Arslanoglu S, Kultursay N & Bayindir O (2002). Protective effect of dietary supplementation with L-arginine and L-carnitine on hypoxia/reoxygenation-induced necrotizing enterocolitis in young mice. Biol Neonate 81, 260–265.[Medline]

Balon TW & Nadler JL (1997). Evidence that nitric oxide increases glucose transport in skeletal muscle. J Appl Physiol 82, 359–363.[Abstract/Free Full Text]

Bass A, Brdiczka D, Eyer P, Hofer S & Pette D (1969). Metabolic differentiation of distinct muscle types at the level of enzymatic organization. Eur J Biochem 10, 198–206.[Medline]

Bednarz B, Jaxa-Chamiec T, Gebalska J, Herbaczynska-Cedro K & Ceremuzynski L (2004). L-arginine supplementation prolongs exercise capacity in congestive heart failure. Kardiol Pol 60, 348–353.[Medline]

Breen EC, Johnson EC, Wagner H, Tseng HM, Sung LA & Wagner PD (1996). Angiogenic growth factor mRNA responses in muscle to a single bout of exercise. J Appl Physiol 81, 355–361.[Abstract/Free Full Text]

Brown LF, Yeo KT, Berse B, Yeo TK, Senger DR, Dvorak HF & van de Water L (1992). Expression of vascular permeability factor (vascular endothelial growth factor) by epidermal keratinocytes during wound healing. J Exp Med 176, 1375–1379.[Abstract/Free Full Text]

Celec P, Gardlik R, Palffy R, Hodosy J, Stuchlik S, Drahovska H et al. (2005). The use of transformed Escherichia coli for experimental angiogenesis induced by regulated in situ production of vascular endothelial growth factor – an alternative gene therapy. Med Hypotheses 64, 505–511.[Medline]

Celec P & Yonemitsu Y (2004). Vascular endothelial growth factor – basic science and its clinical implications. Pathophysiology 11, 69–75.[Medline]

Dulak J & Jozkowicz A (2003). Regulation of vascular endothelial growth factor synthesis by nitric oxide: facts and controversies. Antioxid Redox Signal 5, 123–132.[CrossRef][Medline]

Dulak J, Jozkowicz A, Dembinska-Kiec A, Guevara I, Zdzienicka A, Zmudzinska-Grochot D, Florek I, Wojtowicz A, Szuba A & Cooke JP (2000). Nitric oxide induces the synthesis of vascular endothelial growth factor by rat vascular smooth muscle cells. Arterioscler Thromb Vasc Biol 20, 659–666.[Abstract/Free Full Text]

Forsythe JA, Jiang BH, Iyer NV, Agani F, Leung SW, Koos RD & Semenza GL (1996). Activation of vascular endothelial growth factor gene transcription by hypoxia-inducible factor 1. Mol Cell Biol 16, 4604–4613.[Abstract]

Gavin TP, Spector DA, Wagner H, Breen EC & Wagner PD (2000). Nitric oxide synthase inhibition attenuates the skeletal muscle VEGF mRNA response to exercise. J Appl Physiol 88, 1192–1198.[Abstract/Free Full Text]

Gavin TP & Wagner PD (2002). Attenuation of the exercise-induced increase in skeletal muscle Flt-1 mRNA by nitric oxide synthase inhibition. Acta Physiol Scand 175, 201–209.[CrossRef][Medline]

Giugliano D, Marfella R, Verrazzo G, Acampora R, Coppola L, Cozzolino D & D'Onofrio F (1997). The vascular effects of L-arginine in humans. J Clin Invest 99, 433–438.[Medline]

Goldberg MA, Glass GA, Cunningham JM & Bunn HF (1987). The regulated expression of erythropoietin by two human hepatoma cell lines. Proc Natl Acad Sci U S A 84, 7972–7976.[Abstract/Free Full Text]

Goldberg MA & Schneider TJ (1994). Similarities between the oxygen-sensing mechanisms regulating the expression of vascular endothelial growth factor and erythropoietin. J Biol Chem 269, 4355–4359.[Abstract/Free Full Text]

Gustafsson T, Puntschart A, Kaijser L, Jansson E & Sundberg J (1999). Exercise-induced expression of angiogenesis-related transcription and growth factors in human skeletal muscle. Am J Physiol 276, H679–H685.

Hambrecht R, Hilbrich L, Erbs S, Gielen S, Fiehn E, Schoene N & Schuler G (2000). Correction of endothelial dysfunction in chronic heart failure: Additional effects of exercise training and oral L-arginine supplementation. J Am Coll Cardiol 35, 706–713.[Abstract/Free Full Text]

Hermansen L & Wachtlova M (1971). Capillary density of skeletal muscle in well-trained and untrained men. J Appl Physiol 30, 860–863.[Free Full Text]

Hoffner L, Nielsen JJ, Langberg H & Hellsten Y (2003). Exercise but not prostanoids enhance levels of vascular endothelial growth factor and other proliferative agents in human skeletal muscle interstitium. J Physiol 550, 217–225.[Abstract/Free Full Text]

Hudlicka O, Brown M & Egginton S (1992). Angiogenesis in skeletal and cardiac muscle. Physiol Rev 72, 369–417.[Free Full Text]

Hudlicka O, Brown MD & Silgram H (2000). Inhibition of capillary growth in chronically stimulated rat muscles by N(G)-nitro-L-arginine, nitric oxide synthase inhibitor. Microvasc Res 59, 45–51.[CrossRef][Medline]

Iemitsu M, Miyauchi T, Maeda S, Yuki K, Kobayashi T, Kumagai Y, Shimojo N, Yamaguchi I & Matsuda M (2000). Intense exercise causes decrease in expression of both endothelial NO synthase and tissue NOx level in hearts. Am J Physiol Regul Integr Comp Physiol 279, R951–R959.[Abstract/Free Full Text]

Jacobs TB, Bell RD & McClements JD (1984). Exercise, age and the development of the myocardial vasculature. Growth 48, 148–157.[Medline]

Jakeman LB, Armanini M, Phillips HS & Ferrara N (1993). Developmental expression of binding sites and messenger ribonucleic acid for vascular endothelial growth factor suggests a role for this protein in vasculogenesis and angiogenesis. Endocrinology 133, 848–859.[Abstract]

Jensen L, Bangsbo J & Hellsten Y (2004). Effect of high intensity training on capillarization and presence of angiogenic factors in human skeletal muscle. J Physiol 557, 571–582.[Abstract/Free Full Text]

Kimura H & Esumi H (2003). Reciprocal regulation between nitric oxide and vascular endothelial growth factor in angiogenesis. Acta Biochim Pol 50, 49–59.[Medline]

Kobayashi T & Kamata K (2002). Short-term insulin treatment and aortic expressions of IGF-1 receptor and VEGF mRNA in diabetic rats. Am J Physiol Heart Circ Physiol 283, H1761–H1768.[Abstract/Free Full Text]

Korbonits M, Trainer PJ, Fanciulli G, Oliva O, Pala A, Dettori A, Besser M, Delitala G & Grossman AB (1996). L-arginine is unlikely to exert neuroendocrine effects in human via the generation of nitric oxide. Eur J Endocrinol 135, 543–547.[Abstract]

Kosmidou I, Xagorari A, Roussos C & Papapetropoulos A (2001). Reactive oxygen species stimulate VEGF production from C₂C₁₂ skeletal myotubes through a PI3K/Akt pathway. Am J Physiol Lung Cell Mol Physiol 280, L585–L592.[Abstract/Free Full Text]

Koyama T, Xie Z, Gao M, Suzuki J & Batra S (1998). Adaptive changes in the capillary network in the left ventricle of rat heart. Jpn J Physiol 48, 229–241.[CrossRef][Medline]

Kraus RM, Stallings HW III, Yeager RC & Gavin TP (2004). Circulating plasma VEGF response to exercise in sedentary and endurance-trained men. J Appl Physiol 96, 1445–1450.[Abstract/Free Full Text]

Kubota T, Imaizumi T, Oyama J, Ando S & Takeshita A (1997). L-arginine increases exercise-induced vasodilation of the forearm in patients with heart failure. Jpn Circ J 61, 471–480.[CrossRef][Medline]

Lloyd PG, Prior BM, Li H, Yang HT & Terjung RL (2005). VEGF receptor antagonism blocks arteriogenesis, but only partially inhibits angiogenesis, in skeletal muscle of exercise-trained rats. Am J Physiol Heart Circ Physiol 288, H759–H768.[Abstract/Free Full Text]

Lojda Z (1979). Studies on dipeptidyl (amino) peptidase IV (glycyl proline norphthylamidase). II. Blood vessels. Histochemistry 59, 153–166.[CrossRef][Medline]

MacAllister RJ, Calver AL, Collier J, Edwards CM, Herreros B, Nussey SS & Vallance P (1995). Vascular and hormonal responses to arginine: provision of substrate for nitric oxide or non-specific effect? Clin Sci (Lond) 89, 183–190.[Medline]

Maxwell AJ, Ho HV, Le CQ, Lin PS, Bernstein D & Cooke JP (2001). L-arginine enhances aerobic exercise capacity in association with augmented nitric oxide production. J Appl Physiol 90, 933–938.[Abstract/Free Full Text]

Miele C, Rochfold JJ, Filippa N, Giorgetti-Peraldi S & Van Obberghen E (2000). Insulin and insulin-like growth factor-I induce vascular endothelial growth factor mRNA expression via different signaling pathways. J Biol Chem 275, 21695–21702.[Abstract/Free Full Text]

Milkiewicz M, Brown MD, Egginton S & Hudlicka O (2001). Association between shear stress, angiogenesis, and VEGF in skeletal muscles in vivo. Microcirculation 8, 229–241.[CrossRef][Medline]

Milkiewicz M, Hudlicka O, Brown MD & Silgram H (2005). Nitric oxide, VEGF and VEGFR–2: interactions in activity-induced angiogenesis in rat skeletal muscle. Am J Physiol Heart Circ Physiol 289, H336–H343.[Abstract/Free Full Text]

Murohara T, Asahara T, Silver M, Bauters C, Masuda H, Kalka C, Kearney M, Chen D, Symes JF, Fishman MC, Huang PL & Isner JM (1998). Nitric oxide synthase modulates angiogenesis in response to tissue ischemia. J Clin Invest 101, 2567–2578.[Medline]

Noguchi T, Sasaki Y, Seki J, Giddings JC & Yamamoto J (1999). Effects of voluntary exercise and L-arginine on thrombogenesis and microcirculation in stroke-prone spontaneously hypertensive rats. Clin Exp Pharmacol Physiol 26, 330–335.[CrossRef][Medline]

Prior BM, Yang HT & Terjung RL (2004). What makes vessels grow with exercise training? J Appl Physiol 97, 1119–1128.[Abstract/Free Full Text]

Richardson RS, Wagner H, Mudaliar SRD, Henry R, Noyszewski EA & Wagner PD (1999). Human VEGF gene expression in skeletal muscle: Effect of acute normoxic and hypoxic exercise. Am J Physiol 277, H2247–H2252.

Richardson RS, Wagner H, Mudaliar SRD, Saucedo E, Henry R & Wagner PD (2000). Exercise adaptation attenuates VEGF gene expression in human skeletal muscle. Am J Physiol Heart Circ Physiol 279, H772–H778.[Abstract/Free Full Text]

Shweiki D, Itin A, Soffer D & Keshet E (1992). Vascular endothelial growth factor induced by hypoxia may mediate hypoxia-initiated angiogenesis. Nature 359, 843–845.[CrossRef][Medline]

Srere PA (1969). Citrate synthase. Methods Enzymol 13, 3–5.

Suzuki J (2002). Microvascular remodelling after endurance training with Co²⁺ treatment in the rat diaphragm and hindleg muscles. Jpn J Physiol 52, 409–419.[Medline]

Suzuki J (2004). Time-course changes in VEGF expression and capillarity in the early stage of exercise training with Co²⁺ treatment in rat skeletal muscle. Acta Physiol Scand 181, 225–232.[CrossRef][Medline]

Tharp GD & Wagner CT (1982). Chronic exercise and cardiac vascularization. Eur J Appl Physiol Occup Physiol 48, 97–104.[CrossRef][Medline]

Tomanek RJ (1970). Effects of age and exercise on the extent of the myocardial capillary bed. Anat Rec 167, 55–62.[CrossRef][Medline]

Vassilakopoulos T, Deckman G, Kebbewar M, Rallis G, Harfouche R & Hussain SN (2003). Regulation of nitric oxide production in limb and ventilatiory muscles during chronic exercise training. Am J Physiol Lung Cell Mol Physiol 284, L452–L457.[Abstract/Free Full Text]

Visser JJ & Hoekman K (1994). Arginine supplementation in the prevention and treatment of osteoporosis. Med Hypotheses 43, 339–442.[CrossRef][Medline]

Wang Y, Chang J, Li YC, Li YS, Shyy JY & Chien S (2004). Shear stress and VEGF activate IKK via the Flk-1/Cbl/Akt signaling pathway. Am J Physiol Heart Circ Physiol 286, H685–H692.[Abstract/Free Full Text]

Wittwer M, Billeter R, Hoppeler H & Fluck M (2004). Regulatory gene expression in skeletal muscle of highly endurance-trained humans. Acta Physiol Scand 180, 217–227.[CrossRef][Medline]

Yang HT, Ogilve RW & Terjung RL (1991). Low-intensity training produces muscle adaptation in rats with femoral artery stenosis. J Appl Physiol 71, 1822–1829.[Abstract/Free Full Text]

Zhao X, Lu Z & Feng Q (2002). Deficiency in endothelial nitric oxide synthase impairs myocardial angiognesis. Am J Physiol Heart Circ Physiol 283, H2371–H2378.[Abstract/Free Full Text]

Ziada AM, Huclicka O, Tyler KR & Wright AJ (1984). The effect of long-term vasodilation on capillary growth and performance in rabbit heart and skeletal muscle. Cardiovasc Res 18, 724–732.[Medline]

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