HOME HELP FEEDBACK SUBSCRIPTIONS ARCHIVE SEARCH TABLE OF CONTENTS		QUICK SEARCH:   [advanced] Author: Keyword(s): Year:  Vol:  Page:

This Article Abstract Alert me when this article is cited Alert me if a correction is posted Citation Map Services Email this article to a friend Similar articles in this journal Similar articles in PubMed Alert me to new issues of the journal Add to Personal Folders Download to citation manager Author home page(s): Mohammad Bashar Izzat Dheeraj Mehta Alan J. Bryan Gianni D. Angelini Permission Requests Citing Articles Citing Articles via HighWire Citing Articles via Google Scholar Google Scholar Articles by Jeremy, J. Y. Articles by Angelini, G. D. Search for Related Content PubMed PubMed Citation Articles by Jeremy, J. Y. Articles by Angelini, G. D.

Ann Thorac Surg 1997;63:470-476
© 1997 The Society of Thoracic Surgeons

---

Original Article: Cardiovascular

Nitric Oxide Synthase and Adenylyl and Guanylyl Cyclase Activity in Porcine Interposition Vein Grafts

Bristol Heart Institute, University of Bristol, Bristol, and Department of Physiology, Royal Free Hospital School of Medicine, London, United Kingdom

Accepted for publication September 9, 1996.

	Abstract

Top Footnotes Abstract Introduction Material and Methods Results Comment Acknowledgments References

 
Background. A high proportion of autologous saphenous vein grafts occlude as the result of intimal thickening. Blood vessels synthesize substances that may inhibit such intimal thickening. These include cyclic adenosine monophosphate (cAMP) and cyclic guanosine monophosphate (cGMP), which are stimulated by prostacyclin and nitric oxide, respectively. The prostacyclin–cAMP and nitric oxide–cGMP axes were therefore investigated in porcine vein grafts.

Methods. Saphenous vein–carotid artery interposition graft procedures were carried out in pigs. One month after the operation, ungrafted saphenous veins, vein grafts, and carotid arteries were excised, the formation of cAMP and cGMP was assessed by radioimmunoassay, and the nitric oxide synthase content was determined by autoradiography.

Results. The formation of cAMP and nitroprusside-stimulated cGMP was significantly diminished in vein grafts compared with ungrafted saphenous veins and carotid arteries. Calimycin-stimulated cGMP synthesis (nitric oxide release dependent) and the endothelial nitric oxide synthase content (autoradiography) were significantly elevated in vein grafts compared with ungrafted saphenous veins but were significantly less than those in carotid arteries.

Conclusions. Adenylyl and guanylyl cyclase activity are down-regulated in vein grafts, which may contribute to the development of intimal and medial thickening. Nitric oxide release and endothelial nitric oxide synthase content are up-regulated in vein grafts, which is indicative of an adaptation to the arterial conditions of shear stress and pulsatile pressure.

	Introduction

Top Footnotes Abstract Introduction Material and Methods Results Comment Acknowledgments References

 
Autologous saphenous vein continues to be the most commonly used conduit for coronary artery bypass grafting [1]. In a significant proportion of patients who have undergone coronary artery bypass grafting, however, vein grafts occlude as the result of accelerated intimal thickening and the subsequent development of superimposed atheroma [1]. Although it is known that intimal thickening involves the migration and proliferation of vascular smooth muscle cells (VSMCs), the mechanisms underlying these events remain to be fully elucidated [2–4]. On the basis of findings from experiments performed in animal models of arterial injury, it has been established, however, that both mitogens (eg, platelet-derived growth factor) and vasoconstrictors generated by platelets, leukocytes, and the vascular wall play key roles in initiating and maintaining intimal thickening [2–4]. In contrast, blood vessels possess the capacity to counter intimal hyperplasia. These "defense mechanisms" include prostacylin (PGI₂) and nitric oxide (NO) [4–6], the effects of which are mediated by adenylyl cyclase and guanylyl cyclase, respectively [4–9]. Thus, PGI₂ and stable analogues of cyclic adenosine monophosphate (cAMP) and cyclic guanosine monophosphate (cGMP) have been shown to influence numerous events associated with intimal thickening in vein grafts, including the inhibition of platelet and leukocyte activity, VSMC proliferation and migration, the modulation of tissue plasminogen activators, metalloproteinase expression, proteoglycan synthesis, and cholesterol metabolism [4–9].

To study the possible roles of the PGI₂–cAMP and NO–cGMP axes in the pathophysiology of vein graft failure, the generation of these cyclic nucleotides was assessed in porcine saphenous vein–carotid artery bypass grafts 4 weeks after implantation and compared with their generation in ungrafted saphenous veins and carotid arteries. The synthesis of cAMP was elicited with forskolin, which activates adenylyl cyclase directly, and prostaglandin E₁ (PGE₁; akin in its properties to those of PGI₂), which acts through surface receptors.

The synthesis of cGMP was elicited with sodium nitroprusside, which breaks down spontaneously to generate NO, thereby activating guanylyl cyclase directly [10], and the calcium ionophore A23187, which acts by means of nitric oxide synthase (NOS) activation and the generation of endothelial NO [5]. These biochemical measurements were corroborated by measurements of the NOS content and determinations of its distribution using autoradiography with [³H]L-N^G-nitroarginine (NO ARG) [11].

	Material and Methods

Top Footnotes Abstract Introduction Material and Methods Results Comment Acknowledgments References

 
Surgical Procedures
Studies were performed using white race pigs (weight, 25 to 30 kg) in accordance with British Home Office Animal Care regulations. Premedication, anesthesia, and autologous saphenous vein into carotid artery interposition grafts were performed as detailed previously [12]. Briefly, this involved first making a longitudinal incision on the outer aspect of the hind leg to expose approximately 10 cm of the long saphenous vein. The vein was then dissected free of surrounding tissue by means of a "no touch" technique [13], and all side branches were secured with a 6-0 Prolene ligature (Ethicon Inc, Somerville, NJ). After this the vein was removed from the animal and irrigated with heparinized isosmotic sodium chloride solution (0.9 g/L). Veins were grafted using an end-to-end anastomotic technique, as follows. A segment of one common carotid artery was exposed through a longitudinal neck incision (medial to the sternocleidomastoid muscle) and a 3- to 4-cm segment isolated between vascular clamps was excised; both cut ends were then beveled to approximately 45 degrees. A segment of saphenous vein was similarly beveled and then anastomosed to the carotid artery with continuous 7-0 Prolene suture. The neck and leg wounds were then closed in layers and the animals allowed to recover. After 4 weeks the pigs were reanesthetized, the neck wounds reopened, and the graft identified. The carotid artery was transected distal to the graft, and the absence of any blood flow was taken to indicate graft occlusion. Only patent grafts were used for further analysis. Intact carotid artery from the opposite ungrafted side was also excised, as was the saphenous vein from the contralateral hind leg. All vessels were gently irrigated with prewarmed (37°C) Dulbecco's minimum essential medium ([DMEM] Sigma Chemical Co, Poole, Dorset, UK) pregassed with 95 O₂ and 5% CO₂ and placed immediately in prewarmed DMEM for 20 to 30 minutes before the preparation of tissues for experimentation. Other segments of tissues were placed on aluminum foil on dry ice until frozen. These tissues were immediately transferred to a -80°C freezer before processing for autoradiography.

Preparation of Tissues and Assessment of Cyclic Nucleotide Synthesis
Vascular tissues were prepared for assessment of cyclic nucleotide synthesis according to a previously described method [14]. Adventitia was carefully removed from all vessels, which were opened with a longitudinal incision. Each vessel was then cut laterally into approximately 2-mm strips and further into approximately 2-mm squares. Tissues were then placed in DMEM, pregassed with 95% O₂ and 5% CO₂, and incubated at 37°C for 1 hour, with frequent changes of medium to allow for the equilibration of tissues to in vitro incubation conditions. After preincubation, tissues in duplicate were placed in polypropylene tubes containing DMEM, 250-µmol/L isobutylmethylxanthine (a phosphodiesterase inhibitor), and various concentrations of stimulators of cyclic nucleotide synthesis: PGE₁ and forskolin (cAMP synthesis) and sodium nitroprusside and A23187 (cGMP synthesis). Tubes were incubated for a further 20 minutes at 37°C. Reactions were stopped by the addition of 1 mol/L perchloric acid and the tissues sonicated (3 x 30 seconds; Soniprep, MSE). After centrifugation at 1,000 g for 15 minutes, supernatants were taken and neutralized with 1-mol/L potassium phosphate. Aliquots were then taken and acetylated with triethylamine–acetic anhydride ( vol/vol). Concentrations of cAMP and cGMP were measured using specific iodine 125–labeled radioimmunoassay kits (Amersham International, Aylesbury, Buckinghamshire, UK).

Deoxyribonucleic acid assays were carried out using previously described and validated methods [15]. Vessel segments in which cyclic nucleotide synthesis had been assessed were weighed and frozen in liquid nitrogen. Tissues were crushed (in liquid nitrogen) in a pestle and mortar and lipids extracted with 3 x 500 µL absolute ethanol. Eighty microliters of diaminobenzoic acid was added to the residue and the tubes incubated at 60°C for 30 minutes. Perchloric acid (800 µL) was then added, vortexed, and centrifuged at 1000 g for 3 minutes. Aliquots of supernatant were taken and placed in borosilicate tubes for the measurement of transmission in a fluorometer. Concentrations of DNA were calculated from a concomitantly prepared standard curve.

Samples of ungrafted saphenous veins, carotid arteries, and vein grafts from each animal were placed in phosphate-buffered formalin for histologic appraisal, using a previously described method [12].

Autoradiography of Nitric Oxide Synthase
Transverse 10-µm sections of the carotid artery, ungrafted saphenous vein, and vein graft were cut in a cryostat at approximately -20°C and thaw-mounted onto gelatinized microscope slides, which were stored at -70°C in air-tight containers until use. Localization of NOS was carried out using the the technique described by Kidd and associates [11]. Slide-mounted tissue was allowed to equilibrate to room temperature for at least 30 minutes before incubations were performed. Consecutive sections were incubated for 60 minutes at 22°C in buffer containing 10-nmol/L [³H]L-NG-nitroarginine (³H-NOARG; specific activity, 55 Ci/mmol; Amersham), the degree of nonspecific binding being established by the incubation of alternate sections in the presence of 10-µmol/L unlabeled L-arginine. Slides were washed in buffer (four times in 2 minutes) and dried in a stream of cold air. Low-resolution autoradiography was carried out by exposing sections to ³H-Hyperfilm (Amersham) in x-ray cassettes for 3 months. Films were processed in accordance with the manufacturer's instructions and autoradiographs photographed when appropriate. The microscopic localization (high-resolution autoradiography) of binding was performed by postfixing tissue in paraformaldehyde vapor (2 hours at 80°C) and coating slides in nuclear emulsion (LM-L; Amersham). Slides were then stored in light-proof boxes for 12 weeks at 4°C, after which they were processed in D19 high-contrast developer (Kodak, Hemel Hempstead, UK) and fixed (Hypam, Ilford, UK). Underlying tissue was stained with hemoxylin-eosin, high-resolution autoradiographs were viewed on an Olympus Vanox microscope, and selected sections were photographed when appropriate. Selected sections were incubated in primary antibodies against PECAM and anti–smooth muscle actin (Daco, High Wycombe, Buckinghamshire, UK) to identify smooth muscle and endothelial cells using standard immunohistochemical techniques (avidin-biotin complex peroxidase method; Vector Labs, UK). Receptor binding was determined by counting autoradiographic grains under dark-field illumination (x400 on an Olympus Vanox microscope) overlying luminal endothelial cells and binding expressed in terms of grains per endothelial cell.

Specific ³H-NOARG binding was calculated by subtracting the grain counts of nonspecific binding (ie, sections incubated in 10-nmol/L ³H-NOARG in the presence of 10-µmol/L L-arginine) from the grain counts of total binding (ie, sections incubated in 10-nmol/L ³H-NOARG alone). Values were derived by counting grains from a minimum of 24 sections per vessel (n = 3 pigs).

Data Analysis and Statistics
Data are related to micrograms of deoxyribonucleic acid (rather than protein or wet weight), because this varies for the different vascular tissues. Each data point is expressed as the mean ± standard error of the mean, with numbers of observations in parentheses. Data were analyzed using analysis of variance for multiple comparisons. Paired comparisons between two groups were performed using the paired Student's t test when analysis of variance indicated significance for the multiple comparison. Statistical significance was accepted when p was less than 0.05.

	Results

Top Footnotes Abstract Introduction Material and Methods Results Comment Acknowledgments References

 
The time course of medial and intimal thickening in this pig venous arterial graft model has been reported previously [13]. Consistent with the previous findings, 4 weeks after implantation of grafts, there was an increase in the medial thickness as compared with that in ungrafted veins and the formation of a detectable neointima composed of large amounts of extracellular matrix and smooth muscle cells (n = 8). Investigations using specific immunocytochemical markers and conducted in 4 animals revealed the presence of leukocytes in the media of vein grafts but not in ungrafted saphenous veins or carotid arteries (Southgate and colleagues, unpublished observations). Histologic studies also demonstrated that the endothelium remained intact in all tissues and therefore that harvesting and flushing procedures had no deleterious effect on these cells.

The synthesis of cAMP in response to both forskolin and PGE₁ was significantly lower in vein grafts than that in either ungrafted saphenous veins or carotid arteries (Figs 1, 2). The similarity of the patterns of response seen with both forskolin and PGE₁ indicates that the alterations occur at the level of adenylyl cyclase rather than at the level of prostaglandin receptors and linked systems (eg, G proteins). Similarly, cGMP synthesis in response to sodium nitroprusside was significantly lower in vein grafts than that in carotid arteries but not in ungrafted saphenous veins (Fig 3). In contrast, despite a reduction in guanylyl cyclase activity, A23187-stimulated cGMP synthesis was significantly enhanced in vein grafts compared with that in ungrafted saphenous veins (Fig 4).

View larger version (19K): [in this window] [in a new window]   Fig 1. . Prostaglandin E1 [PGE1]–stimulated cyclic adenosine monophosphate (cAMP) synthesis by porcine ungrafted saphenous vein (), carotid artery (circles), and saphenous vein graft (diamonds). Each point = mean ± standard error of the mean (n = 8). (#p < 0.05, saphenous vein graft versus ungrafted saphenous vein and carotid artery.)

View larger version (17K): [in this window] [in a new window]   Fig 2. . Forskolin-stimulated cyclic adenosine monophosphate (cAMP) synthesis by porcine ungrafted saphenous vein (triangles), carotid artery (circles), and saphenous vein graft (diamonds). Each point = mean ± standard error of the mean (n = 8). (#p < 0.05, saphenous vein graft versus ungrafted saphenous vein and carotid artery.)

View larger version (20K): [in this window] [in a new window]   Fig 3. . Nitroprusside-stimulated cyclic guanosine monophosphate (cGMP) synthesis by porcine ungrafted saphenous vein (triangles), carotid artery (circles), and saphenous vein graft (diamonds). Each point = mean ± standard error of the mean (n = 8). (black star = p < 0.05 [saphenous vein graft versus ungrafted saphenous vein]; #p < 0.05 [ungrafted saphenous vein versus carotid artery]; white star = p < 0.05 [saphenous vein graft versus carotid artery].)

View larger version (21K): [in this window] [in a new window]   Fig 4. . A23187-stimulated cyclic guanosine monophosphate (cGMP) synthesis by porcine ungrafted saphenous vein (triangles), carotid artery (circles), and saphenous vein graft (diamonds). Each point = mean ± standard error of the mean (n = 8). (black star = p < 0.05 [saphenous vein graft versus ungrafted saphenous vein]; # = p < 0.05 [ungrafted saphenous vein versus carotid artery]; white star = p < 0.05 [saphenous vein graft versus carotid artery].)

View larger version (169K): [in this window] [in a new window]   Fig 5. . [3H]L-NG-nitroarginine binding (low resolution) to porcine ungrafted saphenous vein (SV), carotid artery (CC), and vein graft (VG). Autoradiographs, generated on film, from sections incubated in 10-nmol/L 3HL-NG-nitroarginine alone (total binding [TOT]). Nonspecific binding (NSB) was established in the presence of 10 µmol/L L-arginine. Middle panel shows hematoxylin and eosin–stained sections underlying the autoradiographs. (Scale bar = 2.5 mm.)

View larger version (90K): [in this window] [in a new window]   Fig 6. . Representative autoradiographs generated on nuclear emulsion (dark-field illumination, where white grains represent binding sites [left panels]]) of binding to sections of porcine ungrafted saphenous vein (SV), carotid artery (CC), and vein graft (VG) incubated in 10-nmol/L [3H]L-NG-nitroarginine alone. Photomicrographs in righthand panels are of hematoxylin and eosin–stained sections underlying total autoradiographs. The relative density of [3H]L]-NG-nitroarginine binding determined by grain counting (carotid artery > vein graft > ungrafted saphenous vein) corresponds to A23187-stimulated cyclic guanosine monophosphate synthesis (see Fig 4). (Scale bar = 2.5 µm; TM = tunica media; L = lumen.)

	Comment

Top Footnotes Abstract Introduction Material and Methods Results Comment Acknowledgments References

 
This study demonstrated that, in porcine vein grafts, there is a significant reduction in the synthesis of cAMP in response to forskolin, which directly activates adenylyl cyclase activity, and PGE₁, which is mediated by surface receptor activation as compared with that occurring in ungrafted saphenous veins and carotid arteries. These data imply that there is a reduction in adenylyl cyclase activity rather than a change in PGE₁ receptor integrity and linked signal transduction mechanisms. We have also recently reported on our findings in the same model, in which we found that there is a marked reduction of PGI₂ synthesis, which appears to be due to a reduction in the activity of cyclooxygenase synthase [16]. The down-regulation of the cAMP and PGI₂ synthesizing capacity may promote intimal hyperplasia, because the PGI₂-cAMP axis inhibits several key components, including VSMC proliferation, cholesterol accumulation, proteoglycan synthesis, and metalloproteinase activity [4]. The determination of the mechanisms responsible for the reduction in adenylyl cyclase activity was beyond the scope of this study. However, it has been demonstrated that subjecting porcine coronary smooth muscle cells to strain results in a decrease in adenylyl cyclase activity [17], which is consistent with the present findings. Growth factors and the phenotypic transformation of VSMCs from a contractile to a proliferative-secretory phenotype are also possible determinants, but the impact of these remains to be investigated.

The present study also demonstrated that cGMP synthesis is differentially altered in porcine vein grafts, ungrafted saphenous veins, and carotid arteries. Cyclic GMP synthesis by vein grafts was significantly diminished in response to nitroprusside, in comparison with its synthesis by ungrafted saphenous veins and carotid arteries (carotid artery = saphenous vein > vein graft). Because nitroprusside breaks down spontaneously in aqueous solution to liberate NO [10], responses to nitroprusside thus reflect the NO–guanylyl cylase axis in the VSMC component of the vessel. We conclude, therefore, that there is a decrease in guanylyl cyclase activity in the VSMC of the medial-neointimal region of the vein graft. In contrast, there was a marked disparity in the synthesis of cGMP in response to the calcium ionophore A23187 (carotid artery > vein graft > saphenous vein). Because A23187 elicits cGMP synthesis through stimulation of NO release (via NOS) from the endothelium [5] we conclude that NOS activity is induced in the endothelium of vein graft, despite there being a concomitant reduction in guanylyl cyclase activity in the smooth muscle cell component of the graft. Furthermore, the NOS present in these tissues is undoubtedly constitutive NOS, because the activity of this isoform is calcium dependent and occurs exclusively in the endothelium [5]. This conclusion is confirmed by the autoradiographic data, which show that there is an increase in the NOS content in the endothelium of vein grafts compared with its content in ungrafted saphenous veins.

The findings from the present study appear to conflict with those from studies using rabbit vein grafts. Specifically, Cross and associates [18] found that vein graft rings precontracted with norepinephrine failed to relax in response to both acetylcholine and histamine (NO release dependent) but relaxed in response to PGI₂, sodium nitroprusside, and atrial natriuretic peptide (NO release independent). Although Cross and associates suggested that reduced endothelial NO production might be responsible, no direct analyses (eg, autoradiography, cGMP synthesis) were conducted. In a more recent study performed by the same group [19], endothelium-dependent relaxation in explanted human coronary vein grafts was also shown to be impaired, with no response to acetylcholine but a significant relaxation in response to A23187. Because acetylcholine operates by means of receptor-mediated NO release and A23187 operates by means of receptor-independent mechanisms, these data indicate that endothelial NO synthesis may be preserved in human vein grafts, as it is in porcine grafts, but disruption may occur at some point upstream in the signaling pathway linked to the acetylcholine receptor. Indeed, in a recent study on human aortocoronary vein grafts, it was demonstrated that the acetylcholine-stimulated relaxation of vein grafts was impaired, even though constitutive NOS activity was preserved [20].

It is well established that increased shear stress induces endothelial NOS activity in animal models and in vitro systems [21]. It is reasonable to suggest therefore that the increased NOS content and activity in vein grafts compared with ungrafted saphenous veins represents an adaptation to shear stress and other hemodynamic forces. It has also been demonstrated that proliferating endothelial cells show a marked increase in the expression of NOS [22]. Taken together, these observations seem to militate against alterations in NOS expression as playing an etiologic role in the intimal thickening that occurs in vein grafts. However, it has also become increasingly apparent that NO is a double-edged sword, in that an increase in its synthesis may, under certain circumstances, be deleterious to the graft. For example, it has been established that NO reacts with other free radical species to generate the cytotoxic free radical peroxynitrite [23]. Because there is evidence that there is an increased accumulation and infiltration of leukocytes (monocytes, neutrophils) in vein grafts [24], which in turn generate large amounts of superoxide [23], it is reasonable to speculate that the normally protective NO is transformed into peroxynitrite in this "microenvironment." A recent study has also demonstrated that NO interacts with basic fibroblast growth factor to augment the proliferation of VSMCs in culture [25]. Because the expression of growth factors occurs rapidly in recently implanted vein grafts [3], it is possible that the NO interacts with these growth factors to actually enhance VSMC proliferation.

In conclusion, the present study demonstrates that there is a significant down-regulation of adenylyl and guanylyl cyclase activity in porcine saphenous vein–carotid artery interposition grafts at 1 month after implantation. These observations shed further light on the basic mechanisms underlying vein graft failure and may lend support to the use of therapeutic intervention (eg, the administration of PGI₂ analogues, enhancers of cyclic nucleotide synthesis, and NO donors). The present study also demonstrates that there is an up-regulation of NOS activity in the endothelium of the porcine vein graft, which is probably induced by mechanical forces, including shear stress and pulsatile flow. This finding seems to militate against NO playing a significant role in the etiology of intimal thickening; a protective role is more likely. However, given the unique local environment of vein grafts (eg, alterations in shear stress and pulsatile flow, increased leukocyte and platelet adhesion, increased growth factor expression), enhanced NO release, occurring as the result of its interactions with free radical reactions and growth factors, may actually be deleterious. If this is the case, then the administration of NO donors and enhancers of cGMP synthesis (eg, specific phosphodiesterase inhibitors) to ameliorate intimal thickening may actually be contraindicated.

	Acknowledgments

Top Footnotes Abstract Introduction Material and Methods Results Comment Acknowledgments References

 
This research was supported by the British Heart Foundation and the Wellcome Trust.

	Footnotes

Top Footnotes Abstract Introduction Material and Methods Results Comment Acknowledgments References

 
Address reprint requests to Dr Jeremy, Bristol Heart Institute, Bristol Royal Infirmary, Bristol BS2 8HW, United Kingdom.

	References

Top Footnotes Abstract Introduction Material and Methods Results Comment Acknowledgments References

 

1. Bryan AJ, Angelini GD. The biology of saphenous vein graft occlusion: etiology and strategies for prevention. Curr Opin Cardiol 1994;9:641–9.[Medline]
2. Davies MG, Hagen P-O. Pathobiology of intimal hyperplasia. Br J Surg 1994;81:1254–69.[Medline]
3. Newby AC, George SJ. Proposed roles for growth factors in mediating smooth muscle cell proliferation in vascular pathologies. Cardiovasc Res 1993;27:1173–83.[Free Full Text]
4. Jeremy JY, Jackson CL, Bryan AJ, Angelini GD. Eicosanoids, fatty acids and restenosis following coronary artery bypass surgery and balloon angioplasty. Prostaglandins Leukotr Essent Fatty Acids 1996;54:385–402.
5. Bredt DS, Snyder SH. Nitric oxide: a physiologic messenger molecule. Annu Rev Biochem 1994;63:175–95.
6. Jackson CL, Schwartz SM. Pharmacology of smooth muscle cell replication. Hypertension 1992;20:193–205.
7. Garg LC, Hassid S. Nitric oxide generating vasodilators and 8-bromo-cyclic guanosine monophosphate inhibit mitogenesis and proliferation of cultured rat vascular smooth muscle cells. J Clin Invest 1989;893:1774–7.
8. Assender JW, Southgate KM, Hallett MB, Newby AC. Inhibition of proliferation but not calcium mobilization by cyclic AMP and cyclic GMP in rabbit aortic smooth muscle cells. Biochem J 1992;288:527–32.
9. Bennett MR, Evan GI, Newby AC. Deregulated expression of the c-myc oncogene abolishes inhibition of proliferation of rat vascular smooth muscle cells by serum reduction, interferon-, heparin and cyclic nucleotide analogues and induces apoptosis. Circ Res 1994;74:525–36.[Abstract/Free Full Text]
10. Freelisch M, Noack E. Correlation between nitric acid formation during degradation of organic nitrates and activation of guanylate cyclase. Eur J Pharmacol 1987;139:19–30.[Medline]
11. Kidd EJ, Michel AD, Humphrey PPA. Autoradiographic distribution of [³H]L-N^G-nitro-arginine binding in rat brain. Neuropharmacology 1995;34:63–73.[Medline]
12. Angelini GD, Bryan AJ, Williams HMJ, et al. Time course of medial and intimal thickening in pig venous arterial grafts: relationship to endothelial injury and cholesterol accumulation. J Thorac Cardiovasc Surg 1992;103:1093–1103.[Abstract]
13. Gottlob R. The preservation of venous endothelium by dissection without touching and by an atraumatic technique of vascular anastomosis. Minerva Chir 1977;32:693–700.[Medline]
14. Miller MAW, Morgan RJ, Thompson CS, Mikhailidis DP, Jeremy JY. Adenylate and guanylate cyclase activity in the penis and aorta of the diabetic rat: an in vitro study. Br J Urol 1994;74:106–11.[Medline]
15. Kissane JM, Robins E. The fluorimetric measurement of deoxyribonucleic acid in animal tissues with special reference to the central nervous system. J Biol Chem 1979;233:184–8.
16. Jeremy JY, Izzat MB, Birkett SD, Knight DM, Bryan AJ, Angelini GD. Reduced prostacyclin and increased leukotriene B₄ synthesis in porcine venous-arterial grafts. Ann Thorac Surg 1996;61:143–8.[Abstract/Free Full Text]
17. Mills I, Letsou G, Rabban J, Sumpio B, Gerwitz J. Mechanosensitive adenylate cyclase activity in coronary artery vascular smooth muscle. Biochem Biophys Res Comm 1990;171:143–7.[Medline]
18. Cross KS, El-Sanadiki MN, Murray JJ, Mikat EM, McCann RL, Hagen P-O. Endothelium-derived relaxing factor production is absent in vein grafts. Surg Forum 1987;38:319–21.
19. Cross KS, Davies MG, El-Sanadiki MN, Murray JJ, Mikat EM, Hagen P-O. Long term human vein graft contractility and morphology: a functional and histopathological study of retrieved coronary artery vein grafts. Br J Surg 1994;81:699–705.[Medline]
20. Davies MG, Berkowitz DE, Hagen P-O. Constitutive nitric oxide synthase is expressed and nitric oxide–mediated relaxation is preserved in retrieved human aortocoronary vein grafts. J Surg Res 1995;58:732–8.[Medline]
21. Rubanyi GM, Romero JC, Vanhoutte P. Flow-induced release of endothelium-derived relaxing factor. Am J Physiol 1986;250:H1145–9.[Abstract/Free Full Text]
22. Arnal J-F, Yamin J, Dockery S, Harrison DG. Regulation of endothelial nitric oxide synthase mRNA, protein and activity during cell growth. Am J Physiol 1994;267:C1381–8.[Abstract/Free Full Text]
23. Darley-Usmar V, Radomski M. Free radicals in the vasculature: the good, the bad and the ugly. Biochemist 1994;Oct/Nov:15–18.
24. Angelini GD, Bryan AJ, Williams HMJ, et al. Distention promotes platelet and leukocyte adhesion and reduces short-term patency in pig arteriovenous bypass grafts. J Thorac Cardiovasc Surg 1990;99:433–9.[Abstract]
25. Hassid A, Arabshahi H, Bourcier T, et al. Nitric oxide selectively amplifies FGF-2 induced mitogenesis in primary rat aortic smooth muscle cells. Am J Physiol 1994;267:H1040–8.[Abstract/Free Full Text]

This article has been cited by other articles:

				A. W.Y. Chung, P. Rauniyar, H. Luo, Y. N. Hsiang, C. van Breemen, and E. B. Okon Pharmacologic relaxation of vein grafts is beneficial compared with pressure distention caused by upregulation of endothelial nitric oxide synthase and nitric oxide production J. Thorac. Cardiovasc. Surg., October 1, 2006; 132(4): 925 - 932. [Abstract] [Full Text] [PDF]

				T. Schachner, G. Laufer, and J. Bonatti In vivo (animal) models of vein graft disease. Eur. J. Cardiothorac. Surg., September 1, 2006; 30(3): 451 - 463. [Abstract] [Full Text] [PDF]

				Y.-J. Wu, M. Bond, G. B. Sala-Newby, and A. C. Newby Altered S-Phase Kinase-Associated Protein-2 Levels Are a Major Mediator of Cyclic Nucleotide-Induced Inhibition of Vascular Smooth Muscle Cell Proliferation Circ. Res., May 12, 2006; 98(9): 1141 - 1150. [Abstract] [Full Text] [PDF]

				T. Schachner Pharmacologic inhibition of vein graft neointimal hyperplasia J. Thorac. Cardiovasc. Surg., May 1, 2006; 131(5): 1065 - 1072. [Abstract] [Full Text] [PDF]

				T. Sakaguchi, T. Asai, D. Belov, M. Okada, D. J. Pinsky, A. M. Schmidt, and Y. Naka Influence of ischemic injury on vein graft remodeling: Role of cyclic adenosine monophosphate second messenger pathway in enhanced vein graft preservation J. Thorac. Cardiovasc. Surg., January 1, 2005; 129(1): 129 - 137. [Abstract] [Full Text] [PDF]

				J. Y. Jeremy, D. Rowe, A. M. Emsley, and A. C. Newby Nitric oxide and the proliferation of vascular smooth muscle cells Cardiovasc Res, August 15, 1999; 43(3): 580 - 594. [Full Text] [PDF]

				N. Ohno, H. Itoh, T. Ikeda, K. Ueyama, K. Yamahara, K. Doi, J. Yamashita, M. Inoue, K. Masatsugu, N. Sawada, et al. Accelerated Reendothelialization With Suppressed Thrombogenic Property and Neointimal Hyperplasia of Rabbit Jugular Vein Grafts by Adenovirus-Mediated Gene Transfer of C-Type Natriuretic Peptide Circulation, April 9, 2002; 105(14): 1623 - 1626. [Abstract] [Full Text] [PDF]

This Article Abstract Alert me when this article is cited Alert me if a correction is posted Citation Map Services Email this article to a friend Similar articles in this journal Similar articles in PubMed Alert me to new issues of the journal Add to Personal Folders Download to citation manager Author home page(s): Mohammad Bashar Izzat Dheeraj Mehta Alan J. Bryan Gianni D. Angelini Permission Requests Citing Articles Citing Articles via HighWire Citing Articles via Google Scholar Google Scholar Articles by Jeremy, J. Y. Articles by Angelini, G. D. Search for Related Content PubMed PubMed Citation Articles by Jeremy, J. Y. Articles by Angelini, G. D.