Ann Thorac Surg 1998;65:1580-1587
© 1998 The Society of Thoracic Surgeons
Original articles: cardiovascular
Terminal Warm Blood Cardioplegia Improves Cardiac Function Through Microtubule Repolymerization
Hironori Tenpaku, MDa,
Koji Onoda, MDa,
Kyoko Imanaka-Yoshida, MDb,
Toshimichi Yoshida, MDb,
Takatsugu Shimono, MDa,
Hideto Shimpo, MDa,
Isao Yada, MDa
a Department of Thoracic and Cardiovascular Surgery, Mie University School of Medicine, Mie, Japan
b Department of Pathology, Mie University School of Medicine, Mie, Japan
Accepted for publication January 7, 1998.
Address reprint requests to Dr Onoda, Department of Thoracic and Cardiovascular Surgery, Mie University School of Medicine, 2-174 Edobashi, Tsu, Mie 514, Japan
e-mail: (k-onoda{at}clin.medic.mie-u.ac.jp)
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Abstract
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Background. To elucidate the mechanisms responsible for the beneficial effects of terminal warm blood cardioplegia, we studied dynamic change in microtubules induced by cold cardioplegia followed by rewarming. Further, we investigated the relationship between cardiac function and morphologic changes in microtubules caused by hyperkalemic, hypocalcemic warm cardioplegia during initial reperfusion.
Methods. In protocol 1 isolated rat hearts were perfused at 37°C with Krebs-Henseleit buffer (KHB). After 3 hours of hypothermic cardiac arrest at 10°C, hearts were reperfused at 37°C with one of two buffers: group C, 60-minute reperfusion with KHB (K+, 5.9 mmol/L; Ca2+, 2.5 mmol/L); and group TC, 10-minute initial reperfusion with modified KHB (K+, 15 mmol/L; Ca2+, 0.25 mmol/L), followed by 50 minutes of reperfusion with KHB. Cardiac function after reperfusion was determined as a percentage of the prearrest value. In protocol 2 hearts were perfused at 37°C with KHB containing colchicine (10-5 mol/L) for 60 minutes.
Results. There was spontaneous contractile recovery after 10 minutes of initial reperfusion in hearts from group TC as well as improved cardiac function after 15, 30, and 60 minutes of reperfusion compared with that in group C. Immunohistochemical staining and immunoblot analysis demonstrated microtubule depolymerization during hypothermic cardiac arrest and complete repolymerization after 10 minutes of reperfusion with warm buffers in both groups. Colchicine-induced microtubule depolymerization is associated with deterioration of cardiac function.
Conclusions. One mechanism responsible for improved cardiac function mediated by terminal warm blood cardioplegia is the restart of contraction after complete microtubule repolymerization.
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Introduction
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Open heart operations are currently performed using hypothermic cardiac arrest, and recent advances in myocardial protection during hypothermic arrest have made this procedure much safer. Among these advances, the introduction of terminal warm blood cardioplegia to cardiac operations has had a profound impact. Terminal warm blood cardioplegia has been used to prevent reperfusion injury and to improve the recovery of cardiac function after hypothermic arrest. The concept is based on the maintenance of cardiac arrest using hyperkalemic, hypocalcemic warm perfusate during the initial reperfusion period. Use of terminal warm blood cardioplegia favors repletion of high-energy phosphate stores and cellular repair over the generation of electromechanical work [1–3]. However, the mechanisms responsible for improved recovery of contractile function in the setting of terminal warm blood cardioplegia are still unclear.
In the current study, we focused on changes in microtubules, a cytoskeletal component, in the protective effect of terminal warm blood cardioplegia. Microtubules are conserved structures that are composed primarily of the protein tubulin, and a dynamic equilibrium exists between free tubulin and polymerized tubulin of microtubules [4, 5]. In cardiac myocytes, microtubules are involved in the intracellular arrangement of mitochondria, the sarcoplasmic reticulum, plasma membranes, and other organelles and the transport of substances between the organelles through the sarcoplasm [6–9].
Previous studies have shown that microtubules in various types of cells are disrupted by cold treatment and then repolymerized by rewarming [10–13]. Using isolated cardiomyocytes, the effects of cold treatment at 0°C followed by reversal on changes in microtubules and the relation to contractile function have been shown by Tsutsui and colleagues [14, 15]. However, changes in the structure of microtubules during hypothermic cardiac arrest and subsequent reperfusion, which is a conventional procedure employed in cardiac operations, have not been studied in isolated perfused hearts. In the current report, we characterized changes in microtubules and studied the relationship between cardiac function and changes in microtubule morphology caused by hyperkalemic, hypocalcemic warm perfusate during the initial reperfusion.
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Material and methods
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Preparation of isolated rat hearts
All experiments were performed in accordance with the guidelines of the committee on animals of Mie University School of Medicine and with the "Guide for the Care and Use of Laboratory Animals" published by the National Institutes of Health (NIH publication 85-23, revised 1985). Wistar male rats (350 to 400 g) were anesthetized by intraperitoneal administration of sodium pentobarbital (50 mg/kg) and given heparin (200 IU). Subsequently, hearts were removed, and the ascending aorta was cannulated [16]. Hearts were then perfused under constant pressure (90 cm H2O) with Krebs-Henseleit buffer (KHB) with the following composition (mmol/L): NaCl, 115; KCl, 4.7; CaCl2 · 2 H2O, 2.5; MgCl2 · 6 H2O, 1.2; NaHCO3, 25; KH2PO4, 1.2; glucose, 10; pH 7.4, [17]. The perfusate was equilibrated with a gas mixture containing 95% O2 and 5% CO2.
To assess contractile function in the perfused rat heart, a fluid-filled latex balloon was inserted into the left ventricular cavity through the mitral valve. The balloon was connected to a transducer (RPM-6004M, Nihon Koden, Japan), and left ventricular pressure was recorded using a four-channel polygraph (RTA-1100, Nihon Koden). The balloon was inflated with a volume of saline sufficient to produce an end-diastolic pressure of 10 mm Hg. Left ventricular developed pressure (LVDP) was calculated as the difference between peak systolic and end-diastolic pressures. Coronary flow (CF) measurements were based on the volume of perfusate released from the pulmonary artery. After a 15-minute equilibration period, heart rate, LVDP, and CF were recorded as control measurements.
Experimental protocols
Protocol 1. effects of terminal warm cardioplegia on cardiac function
Hearts were perfused retrograde with KHB at 4°C to induce cardiac arrest. Total interruption of retrograde aortic perfusion was then accomplished by cross-clamping the perfusion line. During the 3 hours of cardiac arrest, hearts were maintained at 10°C in cold KHB. The intraventricular balloon was deflated during this period.
Hearts were then divided into two experimental groups. In group C, six hearts were subjected to 60 minutes of reperfusion at 37°C with KHB (K+, 5.9 mmol/L; CA2+, 2.5 mmol/L). In group TC, six hearts were subjected to 10 minutes of initial reperfusion at 37°C with modified KHB (K+, 15 mmol/L; Ca2+, 0.25 mmol/L) followed by 50 minutes of reperfusion at 37°C with KHB.
Protocol 2. association between cardiac function and microtubular alteration
To elucidate the relationship between cardiac function and microtubular alterations, a microtubule depolymerizing agent, colchicine (Sigma Chemical Co), was added to the isolated perfused rat hearts [14, 15, 18–20]. After 15 minutes of perfusion at 37°C with KHB, hearts were divided into two experimental groups. In group N, four hearts were subjected to 60 minutes of perfusion at 37°C with KHB. In group CO, four hearts were subjected to 60 minutes of perfusion at 37°C with KHB containing colchicine (10-5 mol/L).
Immunohistochemical staining of tissue sections from rat hearts
Immunohistochemical staining was performed as previously described [9]. Briefly, hearts were fixed by retrograde perfusion with 4% paraformaldehyde in PEM buffer (0.1 mol/L PIPES-NaOH, pH 6.9, 1 mmol/L EGTA, 2 mmol/L MgCl2). The hearts were cut into small pieces and incubated with 4% paraformaldehyde in PEM buffer at 4°C for 12 hours. Three-micrometer thin sections were placed on glass slides, and incubated for 12 hours at 4°C with a monoclonal antibody directed against 
-tubulin (1:100 dilution; Cederlane Laboratories, Hornby, Ontario, Canada). After several washes with phosphate-buffered saline, the specimens were incubated with fluorescein isothiocyanate-conjugated goat anti–mouse immunoglobulin G (1:100 dilution; MBL, Nagoya, Japan) for 1 hour at room temperature. To stain for both -tubulin and F-actin, selected sections were also incubated with rhodamine-phalloidin (Molecular Probes, Eugene, OR) for 1 hour. The specimens were mounted and viewed using a Zeiss microscope equipped with an epifluorescence system. Photomicrographs were made with Kodak Tri-X film using 40x or 100x oil immersion objective lens.
Quantitative western blot analysis of tubulin
Free and polymerized tubulin fractions were obtained from left ventricular free wall samples using the procedure described by Tsutsui and coworkers [14, 15]. Briefly, a sample of left ventricular tissue (250 mg) was homogenized in 10 mL of microtubule stabilizing buffer, and centrifuged at 100,000 g at 25°C for 15 minutes. The supernatant was saved as the free tubulin fraction. The pellet was resuspended at 0°C in 8 mL of microtubule depolymerizing buffer. After a 1-hour incubation on ice, the resuspended pellet was centrifuged at 100,000 g at 4°C for 15 minutes and the supernatant saved as the polymerized tubulin fraction. Equal amounts of protein from the free and polymerized samples were subjected to electrophoresis on 12.5% polyacrylamide gels. The samples were transferred to nitrocellulose membranes (Schleicher & Schuell, Dassel, Germany), and probed with a monoclonal -tubulin antibody. The bound antibody was labeled using a horseradish peroxidase-conjugated horse anti–mouse IgG (Vector Laboratories, Inc, Burlingame, CA) and visualized with a chemiluminescence system (Amersham International Plc, Buckinghamshire, UK). Purified bovine brain tubulin (Molecular Probes, Inc) was run in a separate lane as a standard. The densities of the bands corresponding to -tubulin were quantified using a densitometer (AE-6900, Atto Corporation, Japan). Values were normalized as a percentage of the control value.
Data analysis
Data are expressed as the mean ± standard error of the mean. All statistical comparisons were performed using a commercially available statistical package for the Macintosh personal computer (StatView, version 4.0, Abacus Concepts). Data were analyzed by one- or two-way analysis of variance. If a significant difference was found, then experimental groups were compared using the appropriate post hoc test. Differences were considered significant at a level of p less than 0.05.
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Results
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Changes in cardiac function and microtubule formation mediated by terminal warm cardioplegia in isolated rat hearts
Changes in cardiac function
In hearts from group TC, rhythmic contractions were observed soon after replacing the modified KHB with KHB. In this group, heart rate recovered to 89.6% ± 2.3% of the control value 15 minutes after reperfusion and returned to control values 60 minutes after reperfusion (Fig 1a). In contrast, in group C, three hearts recovered spontaneous rhythmic contractions by 10 minutes of reperfusion, whereas ventricular fibrillation or asystole continued in the three remaining hearts even after 10 minutes of reperfusion; these hearts recovered rhythmic contractions by 30 minutes of reperfusion. Heart rate recovered to only 38.0% ± 17.3% of the control value after 15 minutes of reperfusion and to only 81.0% ± 2.8% of the control value after 60 minutes of reperfusion (Fig 1a).
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Fig 1. Changes in cardiac function in isolated perfused rat hearts. (a) Changes in heart rate; (b) Changes in coronary flow; (c) Changes in left ventricular developed pressure. Values are expressed as the percent recovery compared with control values and represent the mean ± standard error of the mean for six hearts. (*p < 0.01 compared with group C at the same time point.)
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Coronary flow recovered to 67.5% ± 4.3% and 45.6% ± 2.4% of the control values 15 minutes after reperfusion in group TC and group C, respectively. After 60 minutes, CF had returned to 82.2% ± 4.5% of the control value in group TC. However, CF did not increase after 60 minutes of reperfusion in hearts from group C, and there was a significant difference in the recovery of CF between the two groups during reperfusion (Fig 1b).
Furthermore, in group TC, LVDP recovered to 36.2% ± 6.1% of the control value 15 minutes after reperfusion, and after 60 minutes of reperfusion, the value increased to 66.4% ± 5.5% of the control value. In group C, however, LVDP was less than 10% of the control value after 15 minutes of reperfusion. The LVDP recovered to only 35.1% ± 4.6% of the control value after 60 minutes of reperfusion. These differences in the LVDP between the two groups were significant after 15, 30, and 60 minutes of reperfusion (Fig 1c). These results indicate that initial reperfusion with hyperkalemic, hypocalcemic cardioplegia improves the recovery of cardiac function.
Morphologic changes in microtubules
Immunohistochemical study of isolated rat hearts subjected to 15 minutes of perfusion with KHB at 37°C (control) demonstrated a microtubular network throughout the cytoplasm with a circular formation surrounding the nuclei (Fig 2a). After 3 hours of hypothermic cardiac arrest, disappearance of the cytoplasmic microtubule array was noted in about half of the areas analyzed in tissue sections (Figs 2b and 3a, low magnification).
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Fig 2. Immunohistochemical staining of microtubules from isolated perfused rat hearts from group C. (a) Longitudinal section from a control rat heart. Microtubules are seen as a filamentous network in the cytoplasm and a circular formation around the nuclei. (b) A section from a heart during hypothermic arrest. Microtubules are not seen in this section. (c) A section from a heart 5 minutes after reperfusion. Microtubules have repolymerized around the nuclei. (d) A section from a heart 10 minutes after reperfusion. The cytoplasmic microtubules forming the reticular filaments has been reconstructed. (e) A section from a heart 60 minutes after reperfusion. The microtubules are similar to those seen in control hearts. Scale bar represents 10 µm.
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Fig 3. Double staining for microtubules and actin filaments in isolated perfused rat hearts. (a and b) Longitudinal sections from hearts during hypothermic cardiac arrest. (a) Staining for microtubules. Microtubules are not seen in about half of the area of the tissue sections analyzed (asterisk). (b) Staining for actin filaments by rhodamine-phalloidin localizes to the cross striations of the sarcomeres. No morphologic changes in the actin filaments are seen during hypothermic cardiac arrest. Scale bar represents 10 µm.
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In hearts from group C, repolymerization of the microtubules was observed after 5 minutes of reperfusion with KHB at 37°C. However, despite circular formations around the nuclei, the cytoplasmic network was not completely repolymerized (Fig 2c). After 10 minutes of reperfusion, the cytoplasmic network, which was mainly composed of longitudinal and transverse filaments, had repolymerized completely (Fig 2d). The morphology of the microtubules after 60 minutes of reperfusion was similar to that seen in control sections (Fig 2e). In hearts from group TC, the morphology of microtubules after 5, 10, and 60 minutes of reperfusion was similar to that in hearts from group C (Fig 4). These results suggested that morphologic changes in the microtubule structure were apparently temperature-dependent, and that 10 minutes of reperfusion with modified KHB was enough for the reorganization of cytoplasmic microtubular networks. However, the findings also indicated that these changes were not dependent on the concentration of either potassium or calcium in the cardioplegic solutions commonly used in cardiac operations.
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Fig 4. Immunohistochemical staining of microtubules from isolated perfused rat hearts from group TC. (a) Longitudinal section from a heart 5 minutes after reperfusion. (b) A section from a heart 10 minutes after reperfusion. (c) A section from a heart 60 minutes after reperfusion. The morphology of the microtubules is similar to that in hearts from group C at the same time point. Scale bar represents 10 µm.
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To evaluate the morphology of other cytoskeletal components, double staining for actin filaments and microtubules was performed. In sections from control hearts, actin filaments were stained in a pattern along the cross striations of sarcomeres. This pattern did not change during 3 hours of hypothermic cardiac arrest (Fig 3b), in contrast to the changes in microtubule morphology described previously (Fig 3a).
Quantitation of tubulin
Quantitative densitometric analysis for immunoblots was performed to estimate the amounts of polymerized and depolymerized tubulin more accurately (Fig 5). The amount of free tubulin increased to 162.1% ± 9.4% of the control value after hypothermic cardiac arrest. The amounts of free tubulin after 2, 5, 8, 10, and 60 minutes of reperfusion with KHB (group C) were 136.6% ± 9.4%, 131.9% ± 9.1%, 104.4% ± 2.1%, 103.3% ± 2.5%, and 102.0% ± 2.1% of the control value, respectively (Fig 5a). The amount of free tubulin after 2, 5, 8, and 10 minutes of reperfusion with modified KHB and the following 50 minutes with KHB (group TC) were 142.6% ± 8.9%, 134.1% ± 9.0%, 105.1% ± 2.2%, 103.8% ± 2.3%, and 102.2% ± 2.0% of the control value, respectively (Fig 5a). Conversely, the amount of polymerized tubulin decreased to 54.9% ± 2.6% after hypothermic cardiac arrest. The amounts of polymerized tubulin after 2, 5, 8, 10, and 60 minutes of reperfusion with KHB (group C) were 81.5% ± 1.8%, 81.8% ± 1.9%, 83.4% ± 1.4%, 105.3% ± 3.7%, and 111.3% ± 4.0%, respectively (Fig 5b). The amounts of polymerized tubulin after 2, 5, 8, and 10 minutes of reperfusion with modified KHB and the following 50 minutes with KHB (group TC) were 75.1% ± 2.0%, 80.1% ± 1.9%, 82.9% ± 2.2%, 103.1% ± 4.0%, and 110.1% ± 3.9%, respectively (Fig 5b). There was no significant difference between group C and group TC at their corresponding time points. These results indicated that polymerized tubulin molecules were decreased during hypothermic cardiac arrest but were restored to a level similar to that in the control hearts after 10 minutes of reperfusion.
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Fig 5. Densitometric quantification of immunoblots for free and polymerized tubulin fractions from isolated rat left ventricles before and after hypothermic arrest and after reperfusion. (a) Changes in free tubulin. (b) Changes in polymerized tubulin. Values are reported as the mean ± standard error of the mean for four experiments. (*p < 0.05 versus the respective control value;  p < 0.05 versus the respective control value.)
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Relationship between cardiac function and alterations in microtubule morphology in isolated rat hearts
Figure 6 illustrates changes in cardiac function for isolated rat hearts perfused with KHB or with KHB containing colchicine. In group N, there was no change in heart rate, CF, or LVDP during 60 minutes of perfusion (Fig 6). In group CO, heart rate did not change during 60 minutes of perfusion (Fig 6a); however, at 60 minutes of perfusion, CF and LVDP decreased to 61.1% ± 4.6% and 72.4% ± 4.8% of the control values, respectively (Figs 6b, 6c).
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Fig 6. Effect of colchicine on cardiac function in isolated perfused rat hearts. (a) Changes in heart rate; (b) Changes in coronary flow; (c) Changes in left ventricular developed pressure. Values are expressed as the percentage compared with control values and represent the mean ± standard error of the mean for four hearts. (*p < 0.01 compared with group N at the same time point.)
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Figure 7 showed representative immunoblots for free (Fig 7a) and polymerized (Fig 7b) tubulin, as well as immunohistochemical studies (Figs 7c, 7d) of isolated rat hearts after 60 minutes of perfusion with KHB or with KHB containing colchicine. The amounts of free tubulin in group N and group CO were 100% ± 1.7% and 131% ± 4.0% of the amount before perfusion, respectively (p < 0.05). Conversely, the amounts of polymerized tubulin in group N and group CO were 101% ± 1.6% and 70.2% ± 3.5% of the amount before perfusion, respectively (p < 0.05).
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Fig 7. Immunoblot analysis of free and polymerized tubulin fractions and immunohistochemical staining of microtubules from isolated perfused rat hearts 60 minutes after perfusion with Krebs-Henseleit buffer or with Krebs-Henseleit buffer containing colchicine. (a) Immunoblots of free tubulin fraction. (b) Immunoblots of polymerized tubulin fraction. Lanes 1 and 6, purified bovine brain tubulin; lanes 2 and 4, tubulin from group N; lanes 3 and 5, tubulin from group CO. (c) Longitudinal section from a heart in group N. The morphology of the microtubules is similar to that in control hearts (see Fig 2a). (d) Longitudinal section from a heart in group CO. Cytoplasmic microtubules have decreased; however, microtubules around the nuclei have not disappeared. Arrows indicate nuclei. Scale bar represents 10 µm.
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In hearts from group N, the morphology of the microtubules was similar to that seen in the control hearts (Fig 7c). However, reticular formation of microtubules disappeared from the cytoplasm in hearts from group CO, although perinuclear microtubules did not disappear completely (Fig 7d). These results suggest that the cytoplasmic microtubule network is important for contractile function.
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Comment
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The effects of terminal warm blood cardioplegia have been studied both experimentally and clinically since Lazar and coworkers [21, 22] and Follette and associates [1, 2, 23] established the basic concept of maintaining cardiac arrest with hyperkalemic, hypocalcemic warm blood perfusate during initial reperfusion. The mechanisms responsible for the beneficial effects of this technique are believed to include restoration of high-energy phosphate stores and enhancement of cellular repair [1, 2]. However, there have been no studies concerning the effects of warm blood cardioplegia on the cytoskeletal components. The current study was therefore performed to characterize the dynamic changes in microtubule polymerization and depolymerization in the setting of cardioplegic arrest.
Immunostaining of control hearts demonstrated that microtubules in the perinuclear space form a basketlike structure around the nucleus. Cytoplasmic microtubules, in contrast, form tortuous filaments organized throughout the cytoplasm. These cytoplasmic structures are composed mainly of longitudinal and transverse filaments arranged in reticular networks [24, 25].
During cardiac arrest a 10°C, immunohistochemical staining of rat hearts showed microtubule depolymerization in approximately half of the areas observed on tissue sections. Furthermore, immunoblot analysis confirmed the results of our immunohistochemical studies. It was previously reported that cold treatment at 0°C brings about a complete disappearance of the microtubule network in cultured fibroblasts [10–13] and isolated cardiomyocytes [14, 15, 24]. We maintained the rat hearts at 10°C during cardiac arrest because the myocardial temperature during cardiac arrest in cardiac operations is approximately 10°C. Differences in the degree of the microtubule depolymerization between our finding and previous reports may be explained by the difference in incubation conditions at 0°C or 10°C. The minimal temperature achieved in the current study is not that at which full microtubule depolymerization occurs in most types of cells, including cardiomyocytes. In addition, it is unlikely that the differences between our study and previous reports could be explained by inhomogeneity in microtubule staining in hypothermic hearts, because the staining of actin was homogeneous (Fig 3). However, by 10 minutes after reperfusion with warm buffers, the cytoplasmic microtubule network was reorganized. This phenomenon of microtubular repolymerization was also confirmed by immunoblot analysis at various time points. These findings indicate that microtubules in cardiomyocytes undergo depolarization and then repolymerization during the usual course in cardiac operations. In addition, terminal warm cardioplegia could be effective for microtubule reorganization.
With respect to the recovery of cardiac function, the contractile performance of hearts from group TC with terminal warm cardioplegia was much better than that of hearts from group C. Furthermore, spontaneous rhythmic contractions were observed in all hearts from group TC after initial reperfusion, whereas in group C, three hearts recovered spontaneous rhythmic contractions by 10 minutes of reperfusion, and ventricular fibrillation or asystole continued in the three remaining hearts even after 10 minutes of reperfusion. Cytoplasmic microtubules of the cardiomyocytes in three hearts of group C were still depolymerized at the restart of the beating, whereas in group TC, microtubules were fully reorganized. Therefore, the better contractile function of the hearts in group TC may be related to better reorganization of the microtubule networks.
The question arises as to whether depolymerization of microtubules is deleterious to cardiac function. Lampidis and associates [18] reported that application of low doses of colchicine, a microtubule depolymerizing agent, causes an early reversible increase in the beating rate of neonatal rat cardiac myocytes with a concomitant decrease in the amplitude of contraction. Further, Mery and coinvestigators [26] demonstrated that the intraperitoneal administration of colchicine markedly impaired the intrinsic contractility of left ventricular papillary muscles in adult rats. In contrast, Tsutsui and colleagues [14, 15] found that colchicine- or hypothermia-induced microtubule depolymerization did not significantly affect the extent or velocity of sarcomere shortening in isolated normal cat cardiomyocytes. Therefore, we evaluated cardiac function in the perfused rat hearts after microtubule depolymerization with colchicine. Colchicine (10-5 mol/L) treatment increased free tubulin to more than the control levels, and decreased both CF and LVDP in a time-dependent manner. Our findings showed that the contractile function of hearts with microtubule depolymerization is depressed and that the restart of cardiac beating in hearts, in which microtubules are not repolymerized completely during reperfusion after hypothermic arrest, may result in reduced cardiac function. Microtubules are well-known as tracts of intracellular transport and as a structure for moving and anchoring intracellular organelles [6–9]. Loss of cytoplasmic microtubules during hypothermic arrest could inactivate the transport of various substances and disorganize cytoplasmic organelles. Terminal warm cardioplegia facilitates the reorganization of microtubules, possibly followed by restoration of the active transport and rearrangement of the organelles before the restart of beating. Therefore, we conclude that one mechanism responsible for improved cardiac function mediated by terminal warm blood cardioplegia is the restart of contraction after complete microtubule repolymerization.
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Acknowledgments
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We thank Dr Hiroyuki Tsutsui for technical advice in the biochemical analysis of cardiac tubulin, and Dr Hiroyuki Okada and Dr Masakatsu Shimizu for technical advice in the immunohistochemical staining of cardiac tubulin. This study was supported by a Grant-in-Aid for Scientific Research from the Japanese Minister of Education, by a grant for Cardiac Hypertrophy from Mie University to Koji Onoda, MD, and by a Japan Heart Foundation and IBM Japan Research Grant for 1994 to Kyoko Imanaka-Yoshida, MD.
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Abstract
Introduction
