ventricular hypertrophy (ATP/cardioplegia/cardiopulmonary bypass/inosine)

Transcription

1 Proc. NatL Acad. Sci. USA Vol. 79, pp , January 1982 Medical Sciences Derangements in myocardial purine and pyrimidine nucleotide metabolism in patients with coronary artery disease and left ventricular hypertrophy (ATP/cardioplegia/cardiopulmonary bypass/inosine) JUDITH L. SWAIN, RICHARD L. SABINA, ROBERT B. PEYTON, ROBERT N. JONES, ANDREW S. WECHSLER, AND EDWARD W. HOLMES The Howard Hughes Medical Institute Laboratories, Departments of Medicine and Surgery, Duke University Medical Center, and The Veterans Administration Hospital, Durham, North Carolina 2771 Communicated by James B. Wyngaarden, October 5, 1981 ABSTRACT Studies in animal models of myocardial ischemia and left ventricular hypertrophy have demonstrated a number of derangements in purine and pyrimidine nucleotide content of myocardium that are postulated to play a role in the pathogenesis of muscle dysfunction in these disorders. The present study examined myocardium of patients with coronary artery disease, left ventricular hypertrophy, or neither of these two abnormalities, to determine whether derangements in purine and pyrimidine nucleotide metabolism occur in humans. In patients with coronary artery disease, endocardial content of ATP, GTP, UTP, CTP, and creatine phosphate was reduced and ranged between 6% and 86% of the amount found in the epicardium. In patients without coronary artery disease or ventricular hypertrophy, endocardial content of these nucleotides was equal to or greater than that of epicardium. Endocardial and epicardial content of inosine was increased in patients with coronary artery disease, and after vein bypass grafting inosine content fell to the levels observed in myocardium of patients with normal coronary arteries. In patients with left ventricular hypertrophy, endocardial content of ATP, GTP, UTP, CTP, and creatine phosphate was also reduced and ranged between 64% and 88% of the epicardial content. In contrast to results obtained in patients without left ventricular hypertrophy, epicardial content of GTP, UTP, and CTP was increased by 131%, 123%, and 132% in hypertrophied myocardium. Thus the changes noted in myocardial nucleotide content in patients are similar to those noted in animal models of occlusive coronary disease and ventricular hypertrophy. These results suggest that the pathophysiological abnormalities in nucleotide metabolism noted in animal models also occur in human myocardium. Studies with animal models of occlusive coronary disease and left ventricular hypertrophy have documented changes in the myocardial content and metabolism of purine and pyrimidine nucleotides (1-12). In animal models of ischemia it has been shown that coronary occlusion leads to an abrupt and prolonged depletion of purine, pyrimidine, and pyridine nucleotide content of the myocardium (1-1) and that the magnitude of these changes in nucleotide content is greater in the endocardium than in the epicardium (2, 4-6). In animal models ofventricular hypertrophy, myocardial content of adenine nucleotides has been shown to be decreased (11, 12), with the greatest depression occurring in the subendocardial region of the heart (12). In contrast, pyrimidine nucleotide content of hypertrophied myocardium is increased (11). The derangements in nucleotide metabolism observed in these animal models may provide insight into the pathophysiological events occurring in human myoeardium during ischemia and ventricular hypertrophy. The publication costs ofthis article were defrayed in part by page charge payment. This article must therefore be hereby marked "advertisement" in accordance with 18 U. S. C solely to indicate this fact. However, no data are available to document whether similar changes in purine and pyrimidine nucleotide content are found in the myocardium of patients with coronary artery disease or left ventricular hypertrophy. In the present study we have quantified the endocardial and epicardial content of purine, pyrimidine, and pyridine nucleotides and nucleosides, as well as creatine phosphate (creatine- P), in patients with coronary artery disease and left ventricular hypertrophy during open-heart surgery. Patients were divided into three groups for this study: group 1, those patients with an area of myocardium that was neither supplied by an obstructed coronary artery nor hypertrophied (non-cad-lvh); group 2, those with an area of myocardium supplied by a coronary artery with greater than 75% stenosis (CAD); and group 3, those with left ventricular hypertrophy secondary to aortic stenosis (LVH). Transmural tissue samples were taken at three times: within 5 min of initiation of cardiopulmonary bypass, at the end of cardioplegic arrest, and 3 min after institution of myocardial reperfusion. Each tissue sample was divided into epicardial and endocardial sections and analyzed for ATP, ADP, AMP, GTP, UTP, CTP, NAD, nucleosides, and creatine-p. 655 METHODS Patient Selection. The cardiac catheterization data of all patients to be included in the study were reviewed by a cardiologist and cardiovascular surgeon prior to operation. The patients were then assigned to one of the three groups mentioned above on the basis of the hemodynamic data and coronary arteriograms. A total of 33 patients undergoing cardiac surgery at Duke University Medical Center and the Durham Veterans Administration Hospital were included in the study. Five patients were included in the non-cad-lvh category (group 1); all 5 had at least one tissue sample taken during bypass prior to cardioplegia (this time will subsequently be referred to as the "'bypass" period). The CAD category (group 2) consisted of 18 patients; all ofthese patients had at least one tissue sample taken during cardiopulmonary bypass prior to cardioplegia (bypass), and 6 patients in this group had additional tissue samples taken at the end of cardioplegic arrest (this period is termed "cardioplegia") and after 3 min of reperfusion after cardioplegic arrest (the material taken in the latter biopsy is referred to as the "reperfusion" sample). There were 1 patients in the LVH category (group 3); 9 had at least one tissue sample taken during bypass prior to cardioplegia, and 5 had tissue samples taken Abbreviations: creatine-p, creatine phosphate; CAD, coronary artery disease; LVH, left ventricular hypertrophy; TAN, total adenine nucleotides.

2 656 Medical Sciences: Swain et al P during at least two of the three subsequent study intervals. In the non-cad-lvh group there were two patients with mitral regurgitation, one patient with aortic regurgitation, one with tricuspid stenosis, and one patient with. significant CAD but whose tissue samples were taken from an area perfused by a nonobstructed coronary artery. In the CAD group, there were 3 patients with significant involvement of one coronary artery (narrowing equal to or greater than 75% of the diameter), 5 patients with significant two-vessel involvement, and 1 patients with significant disease of the three major coronary arteries. The LVH group consisted of patients classified as having severe aortic stenosis with valve areas ranging between.4 and.8 cm2 and peak aortic gradients between 5 and 136 mm Hg ( kpa). All had LVH by electrocardiography, ventriculography, or both. Informed consent was obtained from each patient according to the guidelines approved by the Duke University and Durham Veterans Administration Human Experimentation Committees. Operative Procedure. At the time of operation the patients were placed on cardiopulmonary bypass and transmural left ventricular tissue samples were taken at specific stages of the operative procedure. Single or multiple samples were taken 5 min after the establishment of stable cardiopulmonary bypass and prior to cardioplegic arrest (bypass sample), during aortic cross-clamping and cardioplegic arrest (cardioplegia sample), and during the reperfusion period after release of aortic crossclamping but while the patient was still on cardiopulmonary bypass (reperfusion sample). Cardioplegic arrest was achieved by using cold (5C) Ringer's lactate solution containing 2 mm potassium. Eight hundred milliliters of this solution was injected into the aortic root via a 14-gauge needle to initiate arrest, and supplemental infusions were made every 2-3 min during aortic cross-clamping. The myocardial temperature fell from 33 ± 1C at the initiation of cardiopulmonary bypass to 13 ± 2C during cardioplegic arrest.. Hypothermia was augmented by the addition of a topical cold saline slush. Myocardial temperature increased to 37 ± 1PC during the reperfusion period. All tissue samples obtained during aortic cross-clamping and cardioplegia (cardioplegic sample) were taken just prior to release of the cross-clamp. The length of cardioplegia varied between 1 and 94 min, and there was no detectable relationship between the duration of cardioplegia and changes in nucleotide or creatine- P content of the samples obtained during this portion of the operative procedure. All tissue samples taken during reperfusion (reperfusion sample) were obtained 3 min after release of aortic cross-clamping. In one patient, postoperative bleeding occurred from the biopsy site, but this was accompanied by a diffuse bleeding disorder. No other complications occurred. Tissue samples weighing 5-2 mg were taken with a disposable Travenol Tru-Cut biopsy needle. In the CAD patients, the tissue samples were taken from an area of the left ventricle that appeared to be perfused by a significantly narrowed coronary artery identified at the time of coronary arteriography. Samples were not taken from areas where fibrosis was apparent. In the other groups, tissue samples were routinely taken from the anterolateral region of the left ventricle. Each successive sample was taken from an area proximal with, respect to blood supply to the previous samples. This was done to ensure that perfusion to an area for subsequent sampling had not been altered by previous tissue sampling. After removal of the sample from the heart, the needle was blotted on filter paper and plunged into liquid nitrogen within 2 sec. The samples were stored in liquid nitrogen for future analysis. Any sample too small to differentiate epicardium from endocardium was discarded. Analysis of Tissue Samples. Each frozen sample was divided into endocardial and epicardial sections under liquid nitrogen. Proc. Natl. Acad. Sci. USA 79 (1982) Purines, pyrimidines, and creatine-p were isolated by homogenizing the tissue in cold trichloroacetic acid as described (13). The protein pellet that was left after the acid extraction was resuspended in 2 ml of.5 M NaOH and left standing at 4C overnight. After thorough mixing, the protein content of the solubilized pellets was determined by the method of Lowry; et al. (14). Results are expressed in jumol or nmol/g of protein. Nucleotide analyses were carried out by using a Partisil-1 SAX anion-exchange column (Whatman) and a Waters highpressure liquid chromatograph (13). The following nucleotides were routinely quantified: ATP, ADP, AMP, NAD, GTP, UTP, and CTP. The nucleosides adenosine and inosine were quantitated by using a Waters C18 reverse-phase column (13), and the results are expressed in nmol/g of protein. A small change in hypoxanthine content of the samples would have been difficult to detect because of coelution of another compound that also absorbed at 254 nm; however, the absence from all biopsy samples of absorbance at 28 nm in the region of the chromatogram where hypoxanthine eluted excludes a large accumulation of this base. The same extract used for nucleotide and nucleoside analyses was also used for creatine-p determination. Twenty-five microliters of extract was mixed with 25 /,l of 1mM Tris HCl buffer (ph 7.4) that contained 4. mm [14C]ADP (2.5,Ci/,umol; 1 Ci = 3.7 X 11 becquerels), 1 mm MgCl2, and 1.6 units of creatine kinase (Sigma). After incubation at 37 C for 3 min, a 5-,A aliquot ofthe reaction mixture was applied to polyethyleneimine-cellulose F TLC plates (E. Merck, Darmstadt, Federal Republic of Germany) and developed in.8 M LiCl for 8 min. The ATP spot was located by UV light and cut out, and its radioactivity was counted at 79% efficiency in Triton scintillation fluid, using a Packard Tri-Carb liquid scintillation spectrometer. Creatine-P content was calculated from the amount of ATP produced in this reaction. The results are expressed in,tmol/ g of protein. All values are given as the mean ± SEM. Comparisons between individual samples in the same patients were performed by using paired, t analysis. Comparisons between different groups of patients were performed by using unpaired analyses. RESULTS An obvious limitation in a human study of this type is the lack of availability of a true control, either a tissue sample taken under normal conditions (i.e., from a patient not undergoing cardiac surgery) or a sample taken from a completely normal heart. One might hope to circumvent the first problem by percutaneous biopsy through a catheter. However, this has been unsuccessful because the catabolism that results from failure to freeze the sample immediately in liquid nitrogen leads to significantly lower estimates of myocardial nucleoside triphosphate content (15). Performing biopsies at surgery obviates the problem of in vitro catabolism. The second problem cannot be completely overcome, however, because patients with normal hearts do not undergo open-heart surgery. Having to accept this limitation, yet desiring to have analyses from human myocardium for comparison with the CAD and LVH patients, we studied a group of subjects who had an area ofmyocardium that was neither hypertrophied nor supplied by an obstructed coronary artery. Results obtained from the non-cad-lvh. group (Table 1) are similar with respect to absolute nucleotide content, as well as relative endocardial to epicardial content, to the published results obtained for normal myocardium in the intact open-chest dog (2, 4-6). Studies in Patients with CAD. There are additional considerations in studying the group of patients with CAD, because

3 Medical Sciences: Swain et al. Table 1. Myocardial nucleotide and creatine-p contents (,Qmol/g of protein) in the three patient groups during bypass prior to cardioplegia Non- CAD-LVH CAD LVH (n= 5) (n= 18) (n = 9) ATP epi endo endo/epi P TAN epi 39 ± 5 41 ± 2 39 ± 2 endo 41 ± 5 34 ± ± endo/epi 1.3 ±.3.81 ±.4.84 ±.4 P GTP epi 1.3 ± ± ±.1 endo 1.6 ± ± ±.1 endo/epi 1.13 ±.8.9 ±.5.87 ±.5 P UTP epi 1.3 ± ± ±.1 endo 1.5 ± ± ±.1 endo/epi 1.7 ±.9.89 ±.4.85 ±.9 P CTP epi.44 ±.5.48 ±.4.58 ±.4 endo.45 ±.8.4 ±.2.41 ±.4 endo/epi 1.1 ±.9.88 ±.4.74 ±.7 P NAD epi 3.7 ± ± ±.2 endo 3.6 ±.3 3. ± ±.2 endo/epi.99 ±.3.83 ±.4.84 ±.5 P Cre- epi 32 ± 5 25 ± 4 25 ± 3 atine-p endo 35 ± 7 15 ± 3 16 ± 3 endo/epi 1.9 ±.5.71 ±.14.7 ±.12 P P values are the results of the paired ttest when endocardium (endo) was compared to epicardium (epi) in the same patient. TAN, total adenine nucleotides. the left ventricle is not homogeneously affected in this disorder. We have attempted to avoid sampling errors in this group by only reporting data for samples taken from areas that were thought to be supplied by a diseased coronary artery and that did not appear to be fibrotic. The areas to be biopsied were chosen by matching the coronary arteriograms with the anatomy observed at the time of surgery. Preliminary studies documented that we could obtain reproducible results when multiple samples were taken from a suspected ischemic area in the same patient. The mean variation in ATP content for each set Proc. Natl. Acad. Sci. USA 79 (1982) 657 of three samples taken from eight patients was 11%. Table 1 gives the mean ± SEM for epicardial and endocardial content of all classes of nucleotides plus creatine-p in patients with CAD. The data in Table 1 were obtained from tissue samples taken at the initiation of cardiopulmonary bypass. In patients with CAD the endocardial content of ATP, TAN, GTP, UTP, CTP, NAD, and creatine-p is significantly lower than the epicardial content of these nucleotides (all P <.5), leading to endocardial/epicardial ratios of less than 1 for each of these compounds. The magnitude of the reductions in endocardial nucleotide content compared to epicardial content range from 14% for GTP and UTP to 4% for creatine-p. In contrast, in the non-cad-lvh group of patients the endocardial content of these nucleotides is not significantly different from the epicardial content (all P >.1), with endocardial/epicardial ratios approximately equal to 1 for each of these compounds. The lower endocardial content relative to epicardial content of nucleotides and creatine-p in the CAD patients could be the result of either a decrease in the endocardial content or an increase in the epicardial content of the tissue samples. A comparison of the nucleotide and creatine-p values obtained for the epicardium and endocardium in the CAD group to the results obtained for the same myocardial regions in the non-cad-lvh group indicates that the lower ratios obtained in the CAD patients are the result of a decrease in nucleotide and creatine-p content of the endocardial region in these patients. Six patients in the CAD group were biopsied during the periods of cardioplegic arrest and reperfusion, as well as at the initiation of cardiopulmonary bypass. The mean ATP and creatine-p contents obtained for the bypass samples in this subgroup (Table 2) are lower than those obtained for the comparable samples of the entire group of CAD patients (Table 1). The mean values for the subgroup of CAD patients undergoing repetitive biopsies was brought down because two of the six patients in this subgroup had the lowest ATP and creatine-p values of all CAD patients. Clinically, this subgroup was indistinguishable from the remainder of the CAD patients, and the operative course was no different. Thus, we feel the results obtained in this subgroup are indicative of the changes in nucleotide, nucleoside, and creatine-p content that occur during cardioplegia and reperfusion. We expected nucleotide and creatine-p content of epicardium and to a greater extent endocardium to fall during cardioplegic arrest, but the results presented in Table 2 indicate that nucleotide and creatine-p content did not change significantly in either the endocardium or epicardium with cardioplegic arrest or reperfusion. There was a tendency for nucleo- Table 2. Nucleotide and creatine-p content (plmol/g of protein) in CAD and LVH patients during bypass, cardioplegia, and reperfusion CAD (n = 6) LVH (n = 5) Bypass Cardioplegia Reperfusion Bypass Cardioplegia Reperfusion ATP epi 26 ± 4 35 ± 1 28 ± 3 3 ± 3 33 ± 5 26 ± 4 endo 24 ± 4 31 ± 4 28 ± 3 26 ± 2 31 ± 7 22 ± 2 TAN epi 33 ± 4 42 ± 2 33 ± 2 4 ± 4 39 ± 8 32 ± 4 endo 3 ±5 37 ± 5 34 ± 3 34 ± 3 39 ± ± 2 GTP epi 1.2 ± ± ± ± ± ±.3 endo 1.2 ± ± ± ± ± ±.2 UTP epi 1. ± ±.1.9 ± ± ± ±.3 endo 1. ± ±.2.9 ± ±.1 2. ±.5.9 ±.1 CTP epi.31 ±.4.39 ±.4.31 ±.6.68 ±.6.58 ± ±.11 endo.29 ±.4.37 ±.5.32 ±.6.43 ±.3.51 ±.8.31 ±.4 NAD epi 3.1 ± ± ± ± ±.5 3. ±.5 endo 2.8 ± ± ± ± ± ±.2 Cre- epi 12 ± 3 15 ± 4 22 ± 4 16 ± 3 15 ± 4 24 ±9 atine-p endo 8 ± 3 14 ± 5 11 ± 3 12 ± ±3 epi, Epicardium; endo, endocardium.

4 658 Medical Sciences: Swain et al. tide and creatine-p content to increase in both endocardium and epicardium during cardioplegic arrest, but the changes were not statistically significant. During reperfusion mean nucleotide and creatine-p content of both layers of myocardium appeared to decrease toward the levels found at the initiation of cardiopulmonary bypass, but again the changes were not significant. Inosine content ofthe myocardium was quantified in the first sample obtained after initiation of bypass and in the sample obtained during reperfusion in patients from all three groups. In the interval between the bypass and reperfusion biopsies, vein grafting was performed in the CAD group. None of the patients in the other two groups who had inosine determinations underwent vein bypass grafting. The myocardial inosine content in the initial bypass samples for all three groups ofpatients is shown in Fig. 1. The mean inosine content was 5-fold higher in the epicardium (392 ± 137 vs. 8 ± 18 nmovg) and 7-fold higher in the endocardium (464 ± 158 vs. 66 ± 11 nmovg) in the CAD versus the non-cad-lvh group. Fig. 2 illustrates the change in myocardial inosine content between the time when the bypass and reperfusion samples were obtained in the CAD and non-cad-lvh patients. In the four CAD patients mean epicardial inosine content fell by 441 nmovg and endocardial content fell by 71 nmol/g. In contrast, there were only small changes in the epicardial (+24 nmovg) and endocardial (-1 nmovg) inosine contents in the three non-cad-lvh patients. The myocardial adenosine content was less than 18 nmovg in samples obtained at all time points in each of the three groups of patients. Adenosine content did not change in any detectable pattern during cardioplegic arrest or reperfusion. Studies in Patients with LVH. Unlike the regional changes seen in CAD, hypertrophy secondary to pressure overload should affect the left ventricle uniformly. Because tissue sampling in this group of patients does not present the potential sampling errors due to inhomogeneity, all samples were obtained from the anterior lateral region of the left ventricle. In the initial biopsies taken during cardiopulmonary bypass from the patients with LVH, endocardial content ofatp, TAN, GTP, UTP, CTP, NAD, and creatine-p is significantly lower bfj F 6 F 4 2,1 I- 8 I 1+ Non-CAD-LVH (n = 5) CAD (n = 12) LVH (n = 6) FIG. 1. Inosine content during bypass in the epicardium (o) and endocardium (-) of the non-cad-lvh, CAD, and LVH patients. Mean content for each group is given +SEM. I a Proc. Natl. Acad. Sci. USA 79 (1982) B R B R FIG. 2. Inosine content during bypass (B) and reperfusion (R) in the epicardium (o) and endocardium (e) of the non-cad-lvh and CAD patients. Mean content for each group is given ±SEM. than the concentration of these compounds in the epicardium (all P <.5) (Table 1). The magnitude of the reduction in endocardial content compared to epicardial content ranged between 12% for UTP and 36% for creatine-p. When endocardial content of nucleotides and creatine-p in the LVH group is compared to the concentration of these compounds in the endocardium of the non-cad-lvh group, it is apparent that the mean value for each compound is lower in the endocardium of the LVH patients. Thus, the patients with LVH are similar to those with CAD in exhibiting a reduction in endocardial content of all classes of nucleoside triphosphates, NAD, and creatine- P. However, the LVH patients differ from the CAD patients in that epicardial content of the purine nucleotide GTP and pyrimidine nucleotides UTP and CTP is greater in the LVH patients than in the non-cad-lvh patients (Table 1). Mean GTP, UTP, and CTP concentrations were 31%, 23%, and 32% greater in the epicardium of the LVH group compared to the non-cad-lvh group (P <.5). During cardioplegic arrest and the subsequent reperfusion period, myocardial content ofnucleotides and creatine-p did not change significantly in the patients with LVH (Table 2). As in the CAD group, there was a tendency for nucleotide and creatine-p content to increase in the cardioplegia samples and fall in the reperfusion samples in the LVH group, but these changes were not statistically significant. DISCUSSION This study demonstrates that the myocardium of patients with CAD and LVH exhibits many ofthe derangements in nucleotide metabolism that have been observed previously in animal models of these diseases. Comparison of the present study to experiments performed with animal models provides insight into the basis for the changes in myocardial coitent of these nucleotides in patients with CAD and LVH. In animal models it has been demonstrated that the myocardial content of ATP, GTP, UTP, CTP, NAD, and creatine- P (1-1) is reduced after brief coronary occlusion. The magnitude of the decrease in these compounds is greater in the endocardium than in the epicardium (2, 4-6). The results of the present study provide direct evidence for a reduction in endocardial content of these compounds in the region of human

5 Medical Sciences: Swain et al. myocardium served by a diseased coronary artery. Reduction in nucleotide content of the endocardium could be the result of ischemia or replacement of myocardium with fibrotic tissue. If the latter explanation were correct we would not expect vein bypass grafting to increase nucleotide or decrease nucleoside content of the endocardium. Given the brief period of time available for study after vein bypass grafting-i.e., 3 min of myocardial reperfusion-it is unrealistic to expect nucleotide content of the endocardium to have increased, because studies with animal models have shown that nucleotide repletion of noninfarcted myocardium is not complete for many hours after return of normal blood flow (1, 2). On the other hand, nucleoside content, and inosine content in particular, drops very rapidly after restoration of normal blood flow in experimental models of ischemia (1, 2). In patients inosine concentration of coronary sinus blood increases during an attack of angina and returns to normal rapidly after the pain subsides (16). In the present study inosine content of both endocardium and epicardium was increased in the samples obtained from the CAD patients prior to vein bypass grafting. This is the result we would expect if these myocardial regions were ischemic. In the samples obtained after vein bypass grafting the inosine content of both epicardium and endocardium fell in all CAD patients and the concentration of this nucleoside was in the same range observed in the samples from the non-cad-lvh patients. This decrease in inosine content could have resulted from a washout of the nucleoside with improved coronary flow, enhanced reutilization of this nucleotide precursor through the salvage pathway, or a combination of these two processes. The observation that myocardial inosine content decreases with vein bypass grafting suggests that the decrease in endocardial content of nucleotides in the CAD patients is likely to be the result of ischemia rather than replacement of myocardium with fibrotic tissue. It also provides biochemical evidence that vein bypass grafting improves flow and substrate delivery to the previously ischemic segment of myocardium. Animal models of ventricular hypertrophy have also demonstrated changes in myocardial nucleotide content. Studies utilizing different models of hypertrophy have shown that the adenine nucleotide content of the myocardium is decreased in the hypertrophied heart (11, 12). Where examined, the decrease in adenine nucleotide content is greater in the endocardium than in the epicardium (12). In the present study of patients with LVH secondary to pressure overload, endocardial content of adenine nucleotides was found to be decreased. The decrease in nucleotide content was not restricted to the adenine nucleotide pool, however, as is evident from the decrease in endocardial content of GTP, UTP, CTP, and NAD. It is not known whether the decrease in these other nucleotide pools is unique to the type ofhypertrophy seen in this group ofpatients, because studies in animal models either have not included quantitation of these nucleotides or have not compared endocardial to epicardial changes. In hypertrophy studies in which pyrimidine nucleotides have been quantitated, endocardium and epicardium were not analyzed separately and data are reported only for the entire heart (11). In this particular model UTP content of hypertrophied myocardium was increased by 5%. In the present study epicardial content of CTP and UTP was increased by 32% and 23%, respectively, in the LVH patients compared to the non-cad-lvh patients. This finding was unique to the LVH group; epicardial content of CTP was not different and UTP was only slightly increased in the CAD group compared to the non-cad-lvh group. The increase in epicardial content of pyrimidine nucleotides in the LVH group is in animal models is associated with an increase in the rate of consistent with the observation that myocardial hypertrophy pyrimidine nucleotide synthesis (11). An increase in epicardial Proc. Natl. Acad. Sci. USA 79 (1982) 659 GTP was also seen in the hypertrophy patients. There are no reports in the literature of GTP content in animal models of ventricular hypertrophy. When this study was initiated we expected to see some drop in nucleotide and creatine-p content with cardioplegic arrest. However, the results demonstrate that high-energy phosphate stores are well preserved by the cardioplegia procedure. Preservation of these energy stores may be explained by the reduction in temperature and cessation of contractility during cardioplegia such that the decrease in energy utilization is equal to or greater than the decrease in energy production. Because the cardioplegia solution used for these studies was asanguineous, the results indicate that it is not necessary to perfuse the heart with blood during cardioplegia to preserve myocardial content of nucleotides and creatine-p, as suggested by previous investigators (17). The results described in this report demonstrate that there are a number of similarities in the changes in nucleotide metabolism observed in animal models of ischemia and hypertrophy and in patients with either occlusive coronary disease or ventricular hypertrophy. The establishment ofthis link between derangements in nucleotide metabolism in patients and the animal counterparts of these disorders is a necessary prerequisite if future studies with animal models are to provide relevant information regarding the pathogenesis of myocardial dysfunction in patients with occlusive coronary disease and ventricular hypertrophy. The authors thank Mrs. Jean C. Meade and Margaret C. Evans for technical assistance and Mrs. Carolyn S. Mills for secretarial assistance. The work was supported by the Howard Hughes Medical Institute, and by National Institutes of Health Grants ROlAM (to E. W. H.) and R23HL26831 (to J.L. S.). 1. Swain, J. L., Sabina, R. L., McHale, P. A., Greenfield, J. C., Jr., & Holmes, E. W. (1982) Am. J. Physiol., in press. 2. DeBoer, L. M. V., Ingwall, J. S., Kloner, R. A. & Braunwald, E. (198) Proc. Natl. Acad. Sci. USA 77, Reibel, D. K. & Rovetto, M. J. (1979) Am. J. Physiol. 237, H247-H Griggs, D. M., Tchokoev, V. V. & Chen, C. C. 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