UofMN
`Bio-Medical Library
`
`3/5/97
`
`American Heart
`AssociationsM
`
`Fighting Heart Disease
`and Stroke
`
`Volume 95, Number 5 March 4, 1997
`
`Cardiovascular News.
`
`Awards and Named Lecturers at the 69th Scientific Sessions
`
`Editorials
`
`,8-Radiation to Reduce Stenosis • Stent Thrombosis • Mechanisms for Anti proliferative
`Effects of NO and cGMP • Downregulation of Ang II Receptor Type 1 in Heart Failure •
`Vasopressin Deficiency
`
`Brief Rapid Communications
`
`Nitric Oxide and Myocyte Contractility in Heart Failure • ACE Inhibition and Bradykinin in
`Humans • Hyperhomocyst(e)inemia and Endothelial Dysfunction • Vasopressin and
`Vasodilatory Shock
`
`Clinical Investigation and Reports
`
`Cholesterol Lowering and Vascular Function • Risks of Borderline Isolated Systolic
`Hypertension •
`lntracoronary ,B-Irradiation to Reduce Restenosis • Aspirin vs Ticlodipine(cid:173)
`Aspirin After Optimal Stenting • Assessment of CBF After Rotablator Atherectomy • Basic
`Fibroblast Growth Factor and Ischemia • Smooth Muscle Phenotype in Angina and Resteno~is
`• Exercise Echocardiography and PTCA • Pathophysiology of TMI in Acute Coronary
`Syndromes • Angiotensin II Receptors in Failing Human Heart • AT1- and AT2-Receptor Gene
`Expression in Heart Failure • LV-Infundibular Apical Ventricular Septal Defect • Vasodilatory
`Effects of Adrenomedullin in Humans • Mitral Flow Changes and Prognosis in Heart Failure •
`Configuration of AF Electrograms • Heparin-Associated Antibodies and Cardiopulmonary
`Bypass
`
`Basic Science Reports
`
`Tumor Necrosis Factor-a and Protein Synthesis • Myosin Gene Expression in Cardiac
`Increasing cGMP Inhibits EGF
`Hypertrophy • Norepinephrine-Induced Cardiac Hypertrophy •
`Proliferation • KATP Channels and Chronic Hypoxia • Preconditioning and Critical Coronary
`Stenoses • Collagen Metabolism and Restenosis • Estrogen Lacks Vasoprotection in Male
`Rats • Nitroglycerin and Platelet Interaction During Tolerance • Early vs Delayed ACE
`Inhibition in Heart Failure • Septum During LV Unloading • L-Arginine Analogues Reduce
`Myocardial 0 2 Consumption • Lifestick CPR
`Current Perspectives
`
`GISSI Trials and Postinfarction Prognosis
`
`Images in Cardiovascular Medicine
`
`Carcinoid Heart Disease
`
`73-3234(SP) .ISSN 0009-7322
`
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`

`
`Circulation volumees Numbers March4,1997
`
`Cardiovascular News
`
`Awards and Named Lecturers at the American Heart Association 69th Scientific Sessions
`the Editors of Circulation ............................................................................... 1091
`
`Editorials
`
`/3-Radiation to Reduce Restenosis: Too Little, Too Soon?
`Paul Teirstein, MD ..................................................................................... 1095
`Stent Thrombosis: Closing in on the Best Preventive Treatment
`Donald S. Bairn, MD; Joseph P. Carrozza, Jr, MD ......................................................... 1 098
`What Are the Molecular Mechanisms for the Antiproliferative Effects of Nitric Oxide and cGMP
`in Vascular Smooth Muscle?
`Ferid Murad, MD, PhD ................................................................................. 11 01
`
`Downregulation of Angiotensin II Receptor Type 1 in Heart Failure: A Process of Adaptation
`or Deterioration?
`Masahiko Kurabayashi, MD, PhD; Yoshio Yazaki, MD, PhD .................................................. 1104
`
`Role of Vasopressin Deficiency in the Vasodilation of Septic Shock
`fan A. Reid, PhD ...................................................................................... 11 08
`
`Brief Rapid Communications
`
`Role of Myocyte Nitric Oxide in /3-Adrenergic Hyporesponsiveness in Heart Failure
`Shimako Yamamoto, MD; Hiroyuki Tsutsui, MD; Hirofumi Tagawa, MD; Keiko Saito, MD; Masaru Takahashi, MD;
`Hideo Tada, MD; Mitsutaka Yamamoto, MD; Makoto Katoh, BS; Kensuke Egashira, MD; Akira Takeshita, MD ..... 1111
`
`Role of Bradykinin in Mediating Vascular Effects of Angiotensin-Converting Enzyme
`Inhibitors in Humans
`Burkhard Hornig, MD; Christoph Kohler, BS; Helmut Drexler, MD ............................................ 1115
`Hyperhomocyst(e)inemia Is Associated With Impaired Endothelium-Dependent
`Vasodilation in Humans
`Ahmed Tawako/, MD; TorbjGJrn Om/and, MD, PhD; Marie Gerhard, MD; James T. Wu, PhD;
`Mark A. Creager, MD . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 1119
`Vasopressin Deficiency Contributes to the Vasodilation of Septic Shock
`Donald W. Landry, MD, PhD; Howard R. Levin, MD; Ellen M. Gallant, MD; Robert C. Ashton, Jr, MD;
`Susan Sea, BA; David D'Aiessandro, BA; Mehmet C. Oz, MD; Juan A. Oliver, MD ............................. 1122
`
`Clinical Investigation and Reports
`
`Prevention of Cardiovascular Disease*
`Simvastatin, an HMG-Coenzyme A Reductase Inhibitor, Improves Endothelial Function
`Within 1 Month
`Gerard O'Driscoll, MB, BCh, BAO; Danny Green, PhD; Roger R. Taylor, MBBS, FRACP ........................ 1126
`
`*Merck & Co provide unrestricted funds to Circulation in support of the prevention of cardiovascular disease, and Eli Lilly &
`Co provide subscriptions to Circulation to Cardiology Fellows in training.
`
`CIRCULATION (ISSN 0009-7322) is published twice monthly by the American Heart Association, 7272 Greenville Avenue, Dallas, TX 75231-4596. Individuals
`may subscribe for their personal use at the following rates: $132 for members of an American Heart Association scientific council and $176 for nonmembers.
`Outside the United States, add $94 for postage. Contact AHA for single copy rates and subscription rates for medical professionals in training and for libraries,
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`
`A3
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`

`
`1126
`
`Clinical Investigation and Reports
`
`Simvastatin, an HMG-Coenzyme A Reductase
`Inhibitor, Improves Endothelial Function
`Within 1 Month
`
`Gerard O'Driscoll, MB, BCh, BAO; Danny Green, PhD; Roger R. Taylor, MBBS, FRACP
`
`Background Cholesterol-lowering therapy can improve car(cid:173)
`diovascular morbidity and mortality in patients with atheroscle(cid:173)
`rosis. Although the mechanisms responsible are unclear, these
`benefits precede macroscopic changes in the vasculature. Emerg(cid:173)
`ing evidence that improvement in endothelial function may occur
`requires substantiation; in particular, it is unclear how early any
`such improvement would be detectable after initiation of therapy.
`Methods and Results This randomized, double-blind, pla(cid:173)
`cebo-controlled crossover study evaluated the effect of simvas(cid:173)
`tatin (20 mg daily for 4 weeks) on endothelium-dependent and
`endothelium-independent vasodilation and on the response to the
`inhibitor of nitric oxide synthesis, N°-monomethyl-L-arginine
`(L-NMMA), in the forearm vasculature of subjects with moderate
`elevation of total serum cholesterol (6.0 to 10.0 mmol/L) by use
`of strain-gauge plethysmography. Studies were repeated after 3
`more months of open therapy. When the results are expressed as
`
`Hypercholesterolemia is a risk factor for cardio(cid:173)
`
`vascular disease, particularly ischemic heart
`disease, and several recent studies have indi(cid:173)
`cated that cholesterol reduction is associated with de(cid:173)
`creased cardiovascular morbidity and mortality in pa(cid:173)
`tients with established disease 1·2 and in subjects with mild
`hypercholesterolemia. 3 In other studies, clinical improve(cid:173)
`ment has been documented in response to lipid-lowering
`therapy despite relatively minor angiographic changes,
`suggesting that benefits are not causally related only to
`regression of obstructive disease. 4·5 It has been proposed
`that enhanced endothelial function might contribute to the
`improvement in clinical status. 6-8
`Hypercholesterolemia is associated with impaired en(cid:173)
`dothelial function, 9-1 2 and animal13,14 and human 7,15.16
`studies indicate that endothelial function improves with
`lowering of serum cholesterol. In human large coronary
`arteries, NO-dependent, ACh-stimulated vasoconstriction
`was attenuated after 6 months. 6,15 Although a recent re(cid:173)
`port indicated that lipid lowering increased forearm va(cid:173)
`sodilation in response to serotonin after 3 months of treat(cid:173)
`ment, 16 a previous study found that despite significant
`reduction of serum cholesterol after 12 days of therapy,
`no difference in coronary vascular function was evident
`
`Received July 15, 1996; revision received October 2, 1996; ac(cid:173)
`cepted October 13, 1996.
`From the Department of Cardiology and Medicine, Royal Perth ·
`(Australia) Hospital and the Department of Human Movement, Uni(cid:173)
`versity of Western Australia, Perth (D.G.).
`Correspondence to Gerard O'Driscoll, Department of Cardiology,
`Royal Perth Hospital, Wellington St, Perth, Western Australia 6000.
`© 1997 American Heart Association, Inc.
`
`percentage changes in flow in the infused arm relative to the
`noninfused arm, the vasodilator response to acetylcholine was
`significantly increased after 4 weeks of treatment with simvas(cid:173)
`tatin (P<.0005), and this improvement was further enhanced af(cid:173)
`ter 3 months (P<.005). Concurrently, simvastatin augmented the
`vasoconstrictor response to L-NMMA, an effect that was main(cid:173)
`tained at 3 months (P<.0005). The response to the endothelium(cid:173)
`independent vasodilator sodium nitroprusside was unaltered.
`Conclusions These observations indicate that within 1 month
`of treatment with simvastatin, both the stimulated and basal nitric
`oxide dilator functions of the endothelium are augmented, and
`the benefits of this HMG-coenzyme A reductase inhibitor persist
`with continued therapy. (Circulation. 1997;95:1126-1131.)
`
`Key Words • blood flow • cholesterol • endothelium •
`hypercholesterolemia
`
`between patients treated with lovastatin and those given
`placebo. 7 Thus, it remains unclear how early improve(cid:173)
`ment in endothelial function might occur in response to
`lipid-lowering therapy.
`In this study, we documented an effect on both basal
`NO-mediated dilatation and ACh-stimulated dilatation,
`largely NO dependent, in the forearm vasculature after 4
`weeks of therapy with the HMG-CoA reductase inhibitor
`simvastatin.
`
`Methods
`
`Subjects
`Subjects were eligible for entry if they had a total serum cho(cid:173)
`lesterol of 6.0 to 10.0 mmol/L and were not receiving cholesterol(cid:173)
`lowering therapy. All undertook a screening program consisting
`of a medical history and examination and full hematologic and
`biochemical profiles, including measurement of serum electro(cid:173)
`lytes, liver function, uric acid, creatine phosphokinase, and serum
`lipids. The following were excluded: cigarette smokers; premen(cid:173)
`opausal women; those with renal or hepatic impairment, hyper(cid:173)
`tension, or diabetes mellitus; and patients· taking vitamin supple(cid:173)
`ments or vasoactive drugs such as nitrates, calcium channel
`antagonists, ,8-blockers, or ACE inhibitors. The study protocol
`was approved by Royal Perth Hospital Ethics Committee, and
`subjects gave written informed consent.
`Ten subjects (8 men, 2 women; age, 50±2 years [mean±SE])
`with total serum cholesterol concentrations ranging from 6.2 to
`7.5 mmol/L were enrolled. Medical histories and examinations
`were unremarkable except for 2 men, 1 of whom had undergone
`coronary artery bypass surgery and another who had surgically
`treated peripheral vascular disease several years earlier. Both
`were asymptomatic. Apart from 1 patient who was on aspirin 150
`mg daily, no subject was taking medication before or for the
`duration of the experimental period.
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`

`
`O'Driscoll et al Cholesterol Lowering and Vascular Function
`
`1127
`
`Selected Abbreviations and Acronyms
`ACh = acetylcholine
`CoA = coenzyme A
`forearm blood flow
`FBF
`L-NMMA = N°-monomethyl-L-arginine
`NO = nitric oxide
`SNP
`sodium nitroprusside
`
`Study Design
`The effect of 4 weeks of simvastatin therapy was studied by
`use of a randomized, double-blind, placebo-controlled crossover
`protocol that was followed by 3 months of open therapy. Subjects
`were randomized in equal numbers to receive simvastatin 20 mg
`daily (Zocor, Merck Sharp & Dohme) or a similarly packaged
`placebo. After 4 weeks, each subject·retumed for a study of fore(cid:173)
`arm vascular function. Then, after crossover, they were restudied
`4 weeks later. Of the 10 subjects, 2 withdrew at this time, and
`the remainder continued on simvastatin 20 mg daily for 3 more
`months and returned for a final study. Subjects refrained from
`drinking alcohol or caffeine-containing beverages 12 hours be(cid:173)
`fore the procedure. At each visit, the biochemical and hemato(cid:173)
`logic parameters were repeated. There were no adverse side
`effects.
`
`Vascular Function Protocol
`Investigations were conducted in a quiet, climate-controlled
`laboratory (24°C; relative humidity, 55%) with subjects lying
`supine and both forearms resting above heart level. A 20-gauge
`arterial cannula (Arrow) was introduced into the brachial artery
`of the nondominant arm under local anesthesia with <2 mL of
`1% lidocaine (Astra Pharmaceuticals) to transduce pressure and
`infuse drugs or physiological saline. FBF (mL/100 mL forearm
`tissue per minute) was measured simultaneously in both arms
`by galliuin/indium strain-gauge (SG24, Medasonics) plethys(cid:173)
`mography. Each wrist was connected to a flow-regulated source
`of compressed air, and the arm cuffs were connected to a rapid(cid:173)
`inflation device (E20, D.E. Hokanson). Output from the strain
`gauges passed through an amplifier (SPG 16, Medasonics) and
`was sampled by an on-line microcomputer at 75Hz before be(cid:173)
`ing displayed on a monitor in real time. A software program
`coordinated the acquisition, storage, and display of data, as well
`as inflation and deflation of the arm cuffs, ensuring that blood
`flow measures were synchronized with cuff inflation during re(cid:173)
`cording periods. Intra-arterial pressure was measured continu(cid:173)
`ous! y (Transpac, Abbot Laboratories) throughout the study.
`Drug infusions were administered with a constant-rate infusion
`pump (IVAC 770, IVAC Corp).
`Baseline measurements started at least 20 minutes after can(cid:173)
`nulation of the brachial artery, when FBF had stabilized. Blood
`flow measurements were taken by inflating the wrist cuffs to 220
`mm Hg to exclude the hands from the circulation and rapidly
`inflating the upper arm cuffs to 45 mm Hg for 10 of every 15
`seconds throughout the baseline and drug infusion periods. Out(cid:173)
`put from the strain gauges was stored, and the average of the last
`five flow measurements from each period was used for analysis.
`Between infusions, the cuffs were deflated, allowing at least 15
`minutes for FBF to recover before further baseline measures.
`All solutions were prepared aseptically from sterile stock so(cid:173)
`lutions or ampoules immediately before infusion into the brachial
`artery. ACh (Miochol, Johnson & Johnson) was infused at 10,
`20, and 40 p,g/min, each for 3 minutes, to produce a cumulative
`dose-response curve. This was followed by SNP (David Bull
`Laboratories) infusion at 2, 4, and 8 p,g/min, each for 3 minutes,
`and then L-NMMA (Clinalfa) at 2, 4, and 8 p,mol/min, each for
`5 minutes.
`
`Statistical Analysis
`Absolute measurements of FBF are subject to error due to the
`lack of precise standards for calibration purposes, so it is appro-
`
`priate to describe induced changes relative to baseline measure(cid:173)
`ments. Furthermore, although the low doses of drugs infused in
`the study produced negligible systemic effects and showed no
`effect on blood pressure or heart rate, it was necessary to exclude
`an alteration in overall hemodynamics as a cause of the flow
`changes seen in the infused forearm. Thus, FBF was measured
`simultaneously in both arms (although only one arm was in(cid:173)
`fused), and the noninfused arm served as a control. As in pre(cid:173)
`vious articles, 17-zo FBF in the infused arm is described as a
`ratio to that in the noninfused arm. Changes in these ratios
`during ACh, SNP, and L-NMMA infusions are expressed as
`percentage changes from the baseline immediately preceding
`each drug administration.
`Results are expressed as mean±SE. Two-way ANOV A with
`repeated measures was used to compare simvastatin and placebo
`treatments at the three doses. When a significant difference was
`revealed by this analysis, comparisons at each drug infusion level
`were made by use of two-tailed t tests. A value of P< .05 was
`considered significant.
`
`Results
`Table 1 presents the serum lipid levels, heart rate, and
`blood pressure data at the time of the placebo study and
`after 4 weeks and 3 additional months of simvastatin ther(cid:173)
`apy. After 4 weeks of therapy, total serum cholesterol was
`19% lower than after placebo (P< .02), LDL cholesterol
`was 28% lower (P<.05), triglycerides were 21% lower
`(P=NS), and HDL cholesterol was not significantly dif(cid:173)
`ferent. The 3-month levels were similarly depressed rela(cid:173)
`tive to the placebo measures. Other biochemical and he(cid:173)
`matologic variables were unaltered.
`The baseline FBF values preceding infusion of ACh,
`SNP, and L-NMMA were not different in either arm, in(cid:173)
`dicating an adequate washout period between drug infu(cid:173)
`sions. The responses to intra-arterial drug infusions in
`terms of measured absolute FBF (Table 2) were not dif(cid:173)
`ferent between simvastatin and placebo treatment periods.
`However, such direct comparisons are not the most appro(cid:173)
`priate. The percentage changes in flow, from the preceding
`baseline and relative to the noninfused arm, induced by
`the drug infusions are presented in Table 3 and graphically
`shown in Fig 1. The vasodilation induced by ACh in(cid:173)
`creased significantly between placebo and 4 weeks
`(P<.0005, ANOVA) and between placebo and 3 months
`of simvastatin therapy (P<.0001). The ACh response was
`also significantly enhanced at 3 months relative to 4 weeks
`(P<.01). The decrease in FBF induced by L-NMMA was
`significantly greater after 4 weeks (P<.01) and 3 months
`(P<.0005) of therapy relative to placebo. The L-NMMA
`response at 3 months was slightly but not significantly
`greater than at 4 weeks. The results of paired t tests be-
`
`TABLE 1.
`
`lipid and Hemodynamic Values at Studies
`
`Simvastatin
`
`Placebo
`
`4 wk (n=10)
`
`3 mo (n=8)
`
`Cholesterol, mmoi/L
`Total
`LDL
`HDL
`Triglycerides, mmoi/L
`Blood pressure
`Systolic
`Diastolic
`Mean
`Heart rate, bpm
`
`6.6±0.3
`4.5±0.4
`1.2±0.1
`2.4±0.4
`
`131±1
`76±1
`99±1
`67±2
`
`Plus-minus values are mean±SE.
`
`5.4±0.3
`3.2±0.3
`1.3±0.1
`1.9±0.2
`
`131 ±1
`78±1
`100±1
`64±1
`
`5.3±0.2
`3.4±0.3
`1.2±0.1
`1.8±0.3
`
`128±1
`74±1
`96±1
`64±1
`
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`
`1128
`
`Circulation Vol 95, No 5 March 4, 1997
`
`TABLE 2. FBFs in Infused and Non infused Arms During Simvastatin
`and Placebo Administration
`
`Infused Arm, ml · min-1 ·1 00 ml - 1
`Forearm Tissue
`
`Noninfused Arm, ml·min-1 ·100 ml-1
`Forearm Tissue
`
`Simvastatin
`
`Simvastatin
`
`Placebo
`2.57±0.22
`
`4wk
`1.76±0.15
`
`3mo
`2.22±0.27
`
`Placebo
`3.00±0.51
`
`4wk
`2.51 ±0.35
`
`3mo
`2.49±0.46
`
`3.52±0.39
`5.24±0.3.9
`6.95±0.66
`2.55±0.27
`
`5.90±0.85
`9.20±1.64
`11.29±2.02
`2.57±0.38
`
`1.81±0.33
`1.43±0.27
`1.20±0.26
`
`2.83±0.30
`3.87±0.42
`5.01±0.40
`1.85±0.13
`
`4.65±0.48
`6.32±0.82
`7.93±1.01
`2.51±0.26
`
`1.62±0.39
`1.38±0.35
`1.19±0.29
`
`3.75±0.44
`5.36±0.65
`7.26±1.03
`2.14±0.30
`
`4.90±0.77
`6.52±0.99
`7.59±1.20
`2.66±0.35
`
`1.31±0.16
`1.10±0.12
`1.02±0.13
`
`2.79±0.44
`2.72±0.43
`2.74±0.42
`2.66±0.44
`
`2.72±0.46
`2.80±0.53
`2.74±0.54
`2.51 ±0.44
`
`2.47±0.44
`2.48±0.43
`2.49±0.45
`
`2.31±0.34
`2.17±0.31
`2.08±0.28
`2.20±0.29
`
`2.15±0.30
`2.11 ±0.27
`2.18±0.30
`2.76±0.86
`
`2.81±0.59
`2.83±0.48
`3.18±0.56
`
`2.17±0.42
`2.15±0.37
`2.07±0.42
`2.03±0.34
`
`2.05±0.38
`2.06±0.38
`1.88±0.34
`1.69±0.23
`
`1.78±0.20
`1.84±0.22
`2.12±0.16
`
`Baseline
`ACh, J.LQ/min
`10
`20
`40
`Baseline
`SNP, J.LQ/min
`2
`4
`8
`Baseline
`L-NMMA, J.Lmol/min
`2
`4
`8
`
`Values are mean±SE.
`
`tween placebo and treatment groups at each drug dose are
`listed in Table 3. Forearm vascular responses in each in(cid:173)
`dividual subject improved in individuals while on treat(cid:173)
`ment relative to responses obtained from subjects on pla(cid:173)
`cebo (Fig 2). Fig 2 also shows that there was no relation
`between the percentage increase in blood flow ratio in re(cid:173)
`sponse to ACh and the decrease in serum cholesterol in
`individuals at either 4 weeks or 3 months. Similarly, the
`change in response to L-NMMA was not related to the
`decrease in cholesterol. Responses to the endothelium-in(cid:173)
`dependent vasodilator SNP were not significantly altered
`by simvastatin therapy.
`
`Discussion
`This study examined the effects of cholesterol-lowering
`therapy with the HMG-CoA reductase inhibitor simvas(cid:173)
`tatin on the endothelial function of patients with moder(cid:173)
`ately elevated serum cholesterol. After 4 weeks of
`treatment with simvastatin (20 mg daily), the vasodilator
`response to ACh was significantly increased, as was
`L-NMMA-induced vasoconstriction, indicating that both
`stimulated and basal vasodilator function of the endothe-
`
`lium were enhanced. By contrast, the response to the en(cid:173)
`dothelium-independent vasodilator SNP was unchanged.
`Improvement in endothelial function increased with con(cid:173)
`tinued administration of simvastatin despite the absence of
`further reduction in serum cholesterol, and there was no
`relation between the decrease in cholesterol and the im(cid:173)
`provement in endothelial function. FBF responses to in(cid:173)
`fused agents were analyzed by reference to the basal flow
`in the infused and noninfused arms, as in previous stud(cid:173)
`ies.17-2o This corrects first for inaccuracy in strain-gauge
`calibration in the absence of independent absolute flow
`measurements and second for any other changes occurring
`in hemodynamics. Examination of measured values of ab(cid:173)
`solute FBF did not reveal the differences between simvas(cid:173)
`tatin and placebo periods that did exist.
`It is now known that abnormal function of the endothe(cid:173)
`lium is detectable before the establishment of obvious in(cid:173)
`timal lesions in patients with risk factors for atheroscle(cid:173)
`rosis. 21-23 A number of studies performed in animals 12·24
`and in the coronary9,10,2s and peripheral 11·26 vascular beds
`of humans indicate that hypercholesterolemia impairs en(cid:173)
`dothelial fu:p.ction and that the degree of impairment is
`
`TABLE 3. Percentage Changes From Baseline in the Flow Ratio Induced by Drug
`Infusions During Simvastatin and Placebo Administration
`
`Placebo
`
`4wk
`
`P"
`
`Placebo
`
`3mo
`
`p
`
`ACh, J.LQ/min
`10
`20
`40
`SNP, J.LQ/min
`2
`4
`8
`L-NMMA, J.Lmol/min
`2
`4
`8
`
`43±10
`121 ±9
`187±14
`
`150±17
`241 ±30
`333±36
`
`-30±3
`-46±3
`-55±3
`
`79±15
`152±1 1
`249±23
`
`156±26
`248±38
`324±37
`
`-46±5
`-57±3
`-66±3
`
`<.005
`<.0001
`<.001
`
`NS
`NS
`NS
`
`<.005
`<.05
`<.01
`
`36±10
`114±7
`184±16
`
`144±16
`225±28
`319±39
`
`-31±3
`-46±3
`-54±3
`
`108±20
`188±20
`297±28
`
`134±16
`222±26
`315±43
`
`-53±3
`-62±2
`-70±2
`
`<.005
`<.005
`<.0005
`
`NS
`NS
`NS
`
`<.0005
`<.005
`<.005
`
`Values are mean±SE.
`*Values are for the comparisons between placebo and active treatment periods at each drug infusion level with
`two-tailed t tests.
`
`CFAD v. Anacor, IPR2015-01776 ANACOR EX. 2123 - 5/8
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`

`
`0 'Driscoll et al Cholesterol Lowering and Vascular Function
`
`1129
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`
`300
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`200
`
`150
`
`100
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`50
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`
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`
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`400
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`350
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`200
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`0
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`in
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`c
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`.c
`0
`
`40
`20
`10
`Acetylcholine Dose (llg/mln)
`
`0
`
`8
`4
`2
`Sodium Nitroprusside Dose (llg/min)
`
`c
`
`0
`
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`
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`
`-40
`
`-30 ~;t
`
`-50
`
`-60
`
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`
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`
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`
`4
`L-NMMA Dose (llmol/min)
`
`FIG 1. Percentage changes in blood flow ratio (infused/non infused arm) from the baseline preceding each drug infusion for three dose
`levels of ACh (A), SNP (B), and L-NMMA (C). Results are expressed as mean±SE. The response to ACh was significantly greater after
`4 weeks of therapy (*P<.0005), and there was a significant increase with an additional3 months of treatment (tP<.005). The response
`to L-NMMA was significantly different after 4 weeks (tP<.01), and the subsequent increase was not significant. There was no change
`in the response to SNP. • indicates placebo; II, 1 month of simvastatin; and D, 3 months of simvastatin.
`
`correlated with serum cholesterol concentration. 15 In ad(cid:173)
`dition, cholesterol-lowering interventions can reverse en(cid:173)
`dothelial dysfunction in both the coronary and peripheral
`vasculature. 6-8,15.16·27 However, these studies were not ran(cid:173)
`domized, 6,15,27 were performed in patients with established
`atherosclerosis, 7·8 or documented
`improvement after
`longer periods of treatment (6 months in the coronary cir(cid:173)
`culation6·7·15 and 3 months16 or at least 8 weeks27 in the
`peripheral circulation). We have demonstrated, after 4
`weeks of therapy, improvement in endothelial function in
`the forearm vascular bed, one seldom prone to atheroscle(cid:173)
`rosis, in previously untreated subjects with hypercholes(cid:173)
`terolemia using a randomized, double-blind, placebo-con(cid:173)
`trolled crossover design.
`The present study is the first to report that vascular
`changes occur rapidly and to indicate that L-NMMA-me(cid:173)
`diated vasoconstriction is enhanced after cholesterol-low(cid:173)
`ering therapy, indicative of an augmented basal effect of
`NO. The disparity between this finding and that of Stroes
`et al, 16 who reported unchanged L-NMMA responses after
`3 months of combined simvastatin and cholestyramine ad(cid:173)
`ministration, may be attributable to differences in patient
`characteristics, study design, or analysis. Stroes et al 16 re(cid:173)
`cruited predominantly premenopausal women suffering
`from familial hypercholesterolemia who had higher cho(cid:173)
`lesterol levels and had been treated previously with lipid(cid:173)
`lowering therapy. The study was not placebo controlled
`and did not construct a dose-response curve to L-NMMA,
`although the single dose used was the same as our highest
`infusion level. That study, however, found a convincing
`difference in dose-response curves to the endothelium-de(cid:173)
`pendent serotonin according to treatment status, the dose
`being the calculated plasma concentration of serotonin in
`the FBF and the response being expressed as the percent(cid:173)
`age decrease in vascular resistance induced.
`Decreased availability of the NO substrate L-arginine or
`impaired activity of the NO synthase enzyme is not pri(cid:173)
`marily responsible for the impaired endothelial function in
`hypercholesterolemia, 28 and although the effect of NO is
`impaired, its production is increased. 29 Excess endothelial(cid:173)
`derived superoxide generation occurs in hypercholester(cid:173)
`olemia, and these radicals can inactivate N0, 30 augment
`oxidation of LDL,31 and lead to increased endothelial cell
`
`membrane damage through generation of peroxynitrite and
`hydroxyl radicals. Oxidized LDL inhibits the production
`of NO from L-arginine, 32 a finding that may explain the
`observation in both humans 33 and animals 34 that impaired
`endothelium-dependent relaxation in hypercholesterol(cid:173)
`emia can be reversed by the provision of excess substrate
`(L-arginine ). 33,35,36 This benefit of L-arginine is lost after
`lipid-lowering therapy. 16 Further evidence that the oxida(cid:173)
`tion state is important is provided by the observation that
`despite a similar reduction in serum cholesterol with lipid(cid:173)
`lowering therapy alone or in combination with an anti ox(cid:173)
`idant, endothelium-dependent coronary vasomotion was
`improved significantly more with the latter regimen. 8 Sub(cid:173)
`sequently, the coronary vascular response to ACh was
`found to vary inversely with the in vitro oxidizability of
`LDL. 37 However, Garcia et al38 have shown that super(cid:173)
`oxide dismutase coinfusion had no effect on ACh re(cid:173)
`sponses in hypercholesterolemic subjects, failing to sup(cid:173)
`port the theory of extracellular destruction of NO by
`superoxide anions.
`The relative potency of the HMG-CoA reductase inhib(cid:173)
`itors cannot be assessed from the literature concerning en(cid:173)
`dothelial function, but simvastatin is more potent, perhaps
`as much as twice as potent, than pravastatin, fluvastatin,
`and lovastatin in lowering cholesterol. It was interesting
`that the improvement in endothelial function with simvas(cid:173)
`tatin therapy did not correlate with the decrease in serum
`cholesterol. This may be due to the relatively small num(cid:173)
`bers, the relative consistency of the decrease in cholesterol,
`and the imprecision in measurements. In addition to these
`factors, a biological lag phase might explain why the im(cid:173)
`provement between 4 weeks and 3 months was not asso(cid:173)
`ciated with a further decrease in serum cholesterol. How(cid:173)
`ever, there is the possibility that the improvement in
`endothelial function might not be due solely to a reduction
`in circulating cholesterol concentration. Administration of
`fish oils improved endothelial function without altering se(cid:173)
`rum cholesterol. 27 Perhaps the effects of HMG-CoA re(cid:173)
`ductase inhibitors, other than lipid lowering per se, such
`as inhibition of cholesterol synthesis or lowering oxidized
`LDL could improve endothelial function, whereas other
`effects such as reduced monocyte chemotaxis or smooth
`muscle cell migration and proliferation 39.4° and reduced
`
`CFAD v. Anacor, IPR2015-01776 ANACOR EX. 2123 - 6/8
`
`

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`1130
`
`Circulation Vol 95, No 5 March 4, 1997
`
`60 -~ c
`
`"()'50
`~
`CG
`0::
`3: 40
`0 u::
`-g 30
`0
`iii
`. 5 20
`Ul ns
`
`4weeks
`
`II
`
`II
`
`•
`•
`
`II
`
`II
`
`3. Shepherd J, Cobbe S, Ford I, Isles CG, Lorimar AR, MacFarlane P,
`McKillop JH, Packard CJ. Prevention of coronary heart disease with
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`5. Brown G, Albers JJ, Fisher L, Schaefer SM, Lin JT, Kaplan C, Zhao
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`artery disease as a result of intensive lipid lowering therapy in men
`with high levels of apolipoprotein B. N Eng! J Med. 1990;323:
`1289-1298.
`6. Egashira K, Hirooka Y, Kai H, Sugimachi M, Suzuki S, Inou T,
`Takeshita A. Reduction in serum cholesterol with pravastatin im(cid:173)
`proves endothelium-dependent coronary vasomotion in patients with
`hypercholesterolemia. Circulation. 1994;89:2519-2524.
`7. Treasure CB, Klein JL, Weintraub WS, Talley JD, Stillabower ME,
`Kosinski AS, Zhang J, Boccuzzi SJ, Cedarholm JC, Alexander RW.
`Beneficial effects of cholesterol-lowering therapy on the coronary
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`8. Anderson TJ, Meredith IT, Yeung AC, Frei B, Selwyn AP, Ganz P.
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`9. Vita JA, Treasure CB, Yeung AC, Vekshtein VI, Fantasia GM, Fish
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`endothelium-dependent vasodilation of forearm resistanc