ATS
HOME HELP FEEDBACK SUBSCRIPTIONS ARCHIVE SEARCH TABLE OF CONTENTS
QUICK SEARCH:   [advanced]


     

Ann Thorac Surg 2008;85:1367-1373. doi:10.1016/j.athoracsur.2007.12.053
© 2008 The Society of Thoracic Surgeons
This Article
Right arrow Abstract Freely available
Right arrow Full Text (PDF)
Right arrow Alert me when this article is cited
Right arrow Alert me if a correction is posted
Right arrow Citation Map
Services
Right arrow Email this article to a friend
Right arrow Similar articles in this journal
Right arrow Similar articles in PubMed
Right arrow Alert me to new issues of the journal
Right arrow Add to Personal Folders
Right arrow Download to citation manager
Right arrow Author home page(s):
Goro Matsumiya
Norihide Fukushima
Hajime Ichikawa
Yoshiki Sawa
Right arrow Permission Requests
Citing Articles
Right arrow Citing Articles via HighWire
Right arrow Citing Articles via Google Scholar
Google Scholar
Right arrow Articles by Higuchi, T.
Right arrow Articles by Sawa, Y.
Right arrow Search for Related Content
PubMed
Right arrow PubMed Citation
Right arrow Articles by Higuchi, T.
Right arrow Articles by Sawa, Y.
Related Collections
Right arrow Transplantation - heart

Original Articles: Adult Cardiac

Granulocyte Colony-Stimulating Factor Prevents Reperfusion Injury After Heart Preservation

Takuya Higuchi, MDa, Keiko Yamauchi-Takihara, MD, PhDb, Goro Matsumiya, MD, PhDa, Norihide Fukushima, MD, PhDa, Hajime Ichikawa, MD, PhDa, Toru Kuratani, MD, PhDa, Yoshito Maehata, MDa, Yoshiki Sawa, MD, PhDa,*

a Division of Cardiovascular Surgery, Department of Surgery, Osaka University Graduate School of Medicine, Osaka, Japan
b Division of Cardiovascular Surgery, Department of Cardiovascular Medicine, Osaka University Graduate School of Medicine, Osaka, Japan

Accepted for publication December 18, 2007.

* Address correspondence to Dr Sawa, Division of Cardiovascular Surgery, Department of Surgery, Osaka University Graduate School of Medicine, Suita Osaka 565-0871, Japan (Email: sawa{at}surg1.med.osaka-u.ac.jp).


   Abstract
Top
 Abstract
Introduction
 Material and Methods
 Results
 Comment
 References
 
Background: Heart transplantation is an accepted method of treatment for selected patients with end-stage heart disease. Making prolonged heart preservation safer will benefit patients awaiting heart transplantation. Granulocyte colony-stimulating factor (G-CSF) exhibited protective effects against myocardial ischemia–reperfusion injury mediated through the Janus kinase (Jak)/(signal transducer and activator of transcription (Stat) pathway. We examined whether pharmacologic preconditioning with G-CSF improves cardiac function after heart preservation.

Methods: Male rats were divided into four groups: group A, saline injection; group B, G-CSF, 10 µg/kg; group C, G-CSF, 100 µg/kg; and group D, G-CSF, 100 µg/kg plus AG490 (a selective Jak2 inhibitor), 1 mg/kg. The G-CSF and AG490 were given intravenously for 3 consecutive days. Four hours after the final treatment, isolated rat hearts underwent 12 hours of hypothermic (4°C) preservation, followed by 60 minutes of normothermic reperfusion.

Results: Stat3 phosphorylation was observed in the heart at 15 minutes after G-CSF treatment in group C, but this was attenuated by additional treatment with AG 490 in group D. Compared with group A, group C exhibited significant recovery of left ventricular pressure, maximum positive rate of left ventricular developed pressure (Max dP/dt), and coronary flow (p < 0.05, respectively), as well as lower creatine phosphokinase leakage during reperfusion (p < 0.05). Group B and group D did not show significant hemodynamic recovery during reperfusion. In group C, increased Bcl-xL and decreased Bax expressions as well as decreased terminal deoxynucleotide transferase-mediated deoxy uridine triphosphate nick-end labeling (TUNEL)-positive cardiomyocytes were observed after reperfusion. Immunohistochemical examination showed significantly increased capillary density before hypothermic preservation in group C, but not in other groups.

Conclusions: Pharmacologic preconditioning with G-CSF protected hearts from prolonged hypothermic ischemia–reperfusion injury.


   Introduction
Top
 Abstract
 Introduction
Material and Methods
 Results
 Comment
 References
 
Heart transplantation is an accepted method of treatment for patients with end-stage heart disease [1]. Although the liver and kidney can undergo 12- to 24-hour preservation before transplantation, the preservation of the heart for transplantation is limited to 4 hours of cold ischemic storage [2]. Recent studies using a model of prolonged preservation revealed that apoptosis may play an important role in reperfusion injury after prolonged hypothermic ischemia [3]. Many investigators have improved the conditions for preservation in attempts to overcome this problem [4–6].

Granulocyte colony-stimulating factor (G-CSF), a hematopoietic cytokine, induces bone marrow stem cells into the peripheral circulation [7]. Recently, G-CSF has been reported to prevent left ventricular (LV) remodeling and dysfunction after acute myocardial infarction by activating the Janus kinase 2 (Jak2) /signal transducer and activator of transcription 3 (Stat3) pathway [8]. G-CSF receptor was found expressed on cardiomyocytes [8]. G-CSF significantly induced phosphorylation of Jak2 and Stat3 in a dose-dependent manner and protected cultured cardiomyocytes from apoptotic cell death through upregulation of Bcl-2 and Bcl-xL expression [8]. G-CSF also reduced apoptosis of endothelial cells and increased vascularization [8].

Numerous experimental studies have focused on the beneficial effects of G-CSF against myocardial ischemia–reperfusion injury [9–11]. This study examined whether pharmacologic preconditioning with high-dose G-CSF protected the heart from reperfusion injury through its antiapoptotic effect even after 12 hours of myocardial preservation.


   Material and Methods
Top
 Abstract
 Introduction
 Material and Methods
Results
 Comment
 References
 
This study was performed under the supervision of the Animal Research Committee in accordance with the Guidelines on Animal Experiments of Osaka University Medical School and the Japanese Government Animal Protection and Management Law.

Experimental Protocol
This experiment used 58 Sprague-Dawley male rats (weight, 250 g; Nippon Dobutsu, Osaka, Japan). In experiment 1, 40 rats were randomized into four groups of 10 rats each to examine physiologic data (n = 5 per group) and apoptosis (n = 5 per group) after 60 minutes of normothermic reperfusion, followed by 12 hours of hypothermic preservation. In group A, 1 mL of saline was given intravenously for 3 consecutive days. Group B received 10 µg/kg/day G-CSF (generous gift from Kyowa Hakko Kogyo Co, Tokyo, Japan), and group C received 100 µg/kg/day, both in the same manner as given in group A. Group D received 1 mg/kg AG490 (Wako Pure Chemical Industries, Osaka, Japan), a selective Jak2 inhibitor, in addition to 100 µg/kg G-CSF.

As shown in Figure 1, 4 hours after the third injection of G-CSF or saline, rat hearts were excised and perfused using a Langendorff perfusion system [4]. The rats were anesthetized with an intraperitoneal injection of sodium pentobarbital (50 mg/kg) and heparin (200 USP units). The hearts were perfused with modified Krebs-Henseleit buffer (120mM NaCl, 4.5mM KCl, 20.0mM NaHCO3, 1.2mM KH2PO4, 1.2mM MgCl2, 2.5mM CaCl2, and 10mM glucose, and aerated with 95% O2 and 5% CO2 to obtain pH 7.4 at 37°C) at 74 mm Hg for 20 minutes. The hearts were then arrested with 1 mL of St. Thomas solution, dipped into 20 mL of Euro-Collins solution, and preserved at 4°C for 12 hours. After hypothermic preservation, the hearts were again perfused with 37°C Krebs-Henseleit buffer for 60 minutes.


Figure 1
View larger version (13K):
[in this window]
[in a new window]

  		Fig 1. Experimental protocol. Drugs were given intravenously (iv) for 3 consecutive days. Isolated rat hearts were perfused with Langendorff perfusion system for 20 minutes. Hearts were dipped into Euro-Collins solution and preserved for 12 hours after cardioplegic arrest. Thereafter, the hearts were perfused again for 60 minutes. Hemodynamic measurements were done after 20 minutes of initial perfusion and 10, 30, and 50 minutes after 12 hours of hypothermic preservation. Expression of Bcl-xL and Bax, and terminal deoxynucleotide transferase-mediated deoxy uridine triphosphate nick-end labeling (TUNEL)–positive cells were examined 60 minutes after reperfusion. Capillary density was examined 20 minutes before initial perfusion.

 
Experiment 2 (n = 18) was performed to examine Stat3 phosphorylation before preservation at three different time points in groups C and D (n = 3 per time point, per group). Stat3 phosphorylation after hemodynamic estimation at reperfusion was also evaluated in each group. In experiment 3 (n = 20), capillary density was examined in rat hearts after three consecutive G-CSF treatments (n = 5 per group).

Hemodynamic Measurements
A thin-walled latex balloon was inserted into the LV through the left atrium to monitor the LV pressure and control the LV volume. The balloon pressure was then adjusted to 10 mm Hg of the LV end-diastolic pressure. After 20 minutes of stabilization with the Langendorff apparatus, the heart rate, LV developed pressure (LVDP), maximum positive rate of LVDP (Max dP/dt), maximum negative rate of LVDP (Min dP/dt), and coronary flow were all measured with the LV diastolic pressure stabilized at 10 mm Hg. The indices of the cardiac function were continuously measured and analyzed after reperfusion (Polygraph System, Nihon Koden, Japan).

Laboratory Measurements
Four hours after the third injection of G-CSF or saline, 1 mL of peripheral blood was drawn from each rat by aspiration from the inferior vena cava, and the white blood cells (WBC) and platelets were counted. The coronary effluent during reperfusion was collected in chilled vials, and creatine phosphokinase (CPK) leakage was determined.

Western Blot Analysis
The samples were prepared by tissue homogenization using Cell Lysis Buffer (Cell Signaling Technology, Boston, MA). The protein concentration was measured using the DC Protein Assay (Bio-Rad Laboratories, Hercules, CA), and protein extracts were separated by sodium dodecylsulfate polyacrylamide gel electrophoresis (4% to 20% gradient gels) and then transferred to the polyvinylidene fluoride membrane (Invitrogen Corp., Carlsbad, CA), followed by staining with primary antibodies. Affinity purified phospho-Stat3 (p-Stat3), Stat3, Bax, or Bcl-xL (Santa Cruz Biotechnology, Santa Cruz, CA) antibody (1:1000) was used along with an antimouse secondary antibody conjugated to horseradish peroxidase (1:2000) and the Phototope-HRP Western detection Kit (Cell Signaling). A biotinylated protein marker detection pack (Cell Signaling) was used to detect the molecular weight markers on Western blots, which were visualized with the Phototop-HRP Western Blot Detection System. The images of Western blot studies were quantified by plotting a two-dimensional densitogram using the NIH image 1.63 (Research Services Branch, NIMH; National Institute of Health) image analysis program.

Staining
The samples were cut into thin, 5-mm sections and immediately frozen in embedded medium. Hematoxylin and eosin, and terminal deoxynucleotide transferase-mediated deoxy uridine triphosphate nick-end labeling (TUNEL Intergen Kit, Intergen, Purchase, NY) staining, according to the manufacturer’s instruction, were performed to evaluate apoptotic cells.

Immunohistochemical Analysis
The primary mouse antibody (1:50) was used for the immunohistochemical analysis of myocardial tissue. Tissue slides with paraformaldehyde-fixed and paraffin-embedded myocardium were washed several times with cold phosphate-buffered saline (PBS) solution, followed by blocking solution (PBS containing 5% albumin [Sigma, St Louis, MO], 0.1% Triton X-100, and 0.01% sodium acid). The sections were incubated with anti-CD34 antibody (Santa Cruz Biotechnology) as the primary antibody and diluted goat antimouse immunoglobulin G (Vector Laboratories Inc, Burlingame, CA) as the secondary antibody. During the immunohistochemical analysis, 3-3,-diaminobenzidine (DAB) was used for cell staining, with a laser scanning confocal microscope (LSM-510, Carl Zeiss, Oberkochen, Germany). The capillaries were detected by the ratio of CD34-positive area. Twenty different fields at x100 original magnification were randomly selected, and the capillary densities in each field were counted and results expressed as the number of capillaries/mm2.

Statistical Analysis
All values are the mean ± standard deviation. The significance of differences in the functional recovery was determined with repeated-measures analysis of variance. The differences in CPK, the expression of BcL-xL and Bax, and capillary density were determined by means of the one factor analysis of variance. A value of p < 0.05 was considered to be statistically significant.


   Results
Top
 Abstract
 Introduction
 Material and Methods
 Results
Comment
 References
 
Effects of G-CSF Before Hypothermic Preservation
The phosphorylation of Stat3 after G-CSF treatment was first examined in the rat hearts. As shown in Figure 2, Stat3 was phosphorylated to a significant extent 15 minutes after G-CSF 100-µg/kg treatment in group C, although the extent of this phosphorylation decreased at 30 minutes. Stat3 phosphorylation exhibited by G-CSF treatment was significantly inhibited by additional treatment with AG490 (group D). The expression level of Stat3 was identical in both groups. Stat3 phosphorylation was not detected with the low-dose G-CSF treatment (10 µg/kg) in group B (data not shown).


Figure 2
View larger version (41K):
[in this window]
[in a new window]

  		Fig 2. Phosphorylation of signal transducer and activator of transcription 3 (Stat3) in rat hearts. (A) Tyrosin phosphorylation of Stat3 was examined by Western blot analysis as described in "Material and Methods." The hearts were excised 0, 15, and 30 minutes after intravenous administration of 100-µg/kg G-CSF (group C) or 100-µg/kg G-CSF plus 1-mg/kg AG490 (group D). A representative blot is shown. (G-CSF = granulocyte colony-stimulating factor.) (B) Quantitative analysis showed significant phosphorylation of Stat3 at 15 minutes in group C, but this was abolished by additional treatment of AG490 in group D (n = 3, *p < 0.05). All values are mean ± standard deviation.

 
As summarized in Table 1, the WBC count was higher in group C than in the other groups, although not to a significant extent (p = 0.07). Similarly, there was no significant difference in the number of platelets among the four groups (p = 0.66). Cardiac function was also examined before the hypothermic preservation. The LVDP, Max dP/dt, coronary flow, and heart rate were comparable in each group. AG490 treatment did not induce any declining effects on these hemodynamic variables.


View this table:
[in this window]
[in a new window]

  		Table 1 Results of Hemodynamic Indicators and Laboratory Examinations Before Hypothermic Preservation in the Study Groups a
 
Cardiac Function After Hypothermic Preservation
Cardiac function at reperfusion was evaluated as the percentage change of the values obtained before hypothermic preservation. As shown in Figure 3A, LVDP in group A decreased to 44.4% ± 10.0% at 10 minutes and was unchanged during reperfusion. On the other hand, the LVDP did not decrease in group C during reperfusion and was thereafter significantly sustained at 30 and 50 minutes after reperfusion (101.3% ± 17.2%). Treatment with 10 µg/kg G-CSF (group B) did not exhibit any beneficial effects on the recovery of LVDP during reperfusion as 100 µg/kg did (group C). Moreover, additional treatment with AG490 completely inhibited the beneficial effects of G-CSF on LVDP recovery (group D).


Figure 3
View larger version (20K):
[in this window]
[in a new window]

  		Fig 3. Cardiac function during reperfusion (all values are mean ± standard deviation; n = 5 in each group; *p < 0.05). (A) Left ventricular developed pressure (LVDP) was expressed as percentage change of values before hypothermic preservation. Percentage recovery of LVDP was about 50% in group A (vertical fill), whereas in group C (horizontal fill), LVDP recovered at an early stage of reperfusion and reached the levels comparable with those before preservation. AG490 abolished the effect of granulocyte colony-stimulating factor (G-CSF; group D, solid fill). A low dose of G-CSF (group B, clear bars) did not present beneficial effects on LVDP. (B) Coronary flow during reperfusion was significantly higher in group C than that observed in groups A, B, and D. (C) The heart rate during reperfusion was similar among the four groups.

 
The coronary flow during reperfusion was compared among the four groups (Fig 3B). The coronary flow in group A was 2.0 ± 0.7 mL/min at 30 minutes after reperfusion, which was less than 30% of that observed before hypothermic preservation. The coronary flow in group C at 30 minutes after reperfusion was more than 60% of that observed before hypothermic preservation and was maintained throughout reperfusion. The coronary flow in groups B and D was almost same as that observed in group A. There was no significant difference in the heart rate among four groups during reperfusion (Fig 3C).

In addition, for the distinctive estimation of cardiac function, Max dP/dt and Min dP/dt was estimated during reperfusion. Group C had a significantly higher Max dP/dt than group A (p < 0.05), which showed 96.1% recovery of Max dP/dt during reperfusion, whereas groups A, B, and D only demonstrated approximately 50% recovery (Fig 4A). The results of Min dP/dt were similar to those of Max dP/dt (Fig 4B). The CPK concentration in the coronary flow during the 60-minute reperfusion period was compared among four groups (Fig 4C). The CPK concentration in group C was significantly lower than that in group A (2.7 ± 0.8 vs 21.8 ± 7.6 IU, p < 0.05), whereas the CPK concentrations in groups B and D were similar to those in group A.


Figure 4
View larger version (20K):
[in this window]
[in a new window]

  		Fig 4. (A, B) Cardiac function as measured by maximum positive and negative dP/dt (Max dP/dt and Min dP/dt) during reperfusion. Significantly higher Max dP/dt was observed during reperfusion in group C (horizontal fill) than those in groups A (vertical fill), B (clear bars) and D (solid fill). (C) Creatine phosphokinase (CPK) concentration in the coronary flow. Release of CPK during 60 minutes reperfusion was significantly suppressed in group C compared with those obtained in Groups A, B, and D (n = 5 in each group; *p < 0.05).

 
Apoptosis After Reperfusion
The expression levels of Bcl-xL and Bax were examined in rat hearts by Western blot analysis after 60 minutes of reperfusion followed by 12 hours of hypothermic preservation (Fig 5A). Compared with group A, Bcl-xL expression was significantly increased and Bax expression was significantly decreased in group C. These effects were not observed in group B. Additional treatment with AG490 inhibited the effect of G-CSF as shown in group D.


Figure 5
View larger version (52K):
[in this window]
[in a new window]

  		Fig 5. Apoptosis in the hearts after reperfusion. (A) Bcl-xL and Bax expression was examined by Western blot analysis. Relative intensity showed the ratio of that expression compared with that of β-actin. (B) Terminal deoxynucleotide transferase-mediated deoxy uridine triphosphate nick-end labeling (TUNEL) assay. The number of TUNEL positive cells was expressed as percentages compared with all nuclei. Percentage of TUNEL-positive cells from four groups was compared. TUNEL-positive cells were significantly lower in group C than other groups. (Bar = 100 µm; n = 5 in each group; *p < 0.05.)

 
The number of apoptotic cardiomyocytes was examined in rat hearts after 60 minutes of reperfusion (Fig 5B). Group A had 13.4% ± 3.8% TUNEL-positive cells, group C had 3.8% ±1.6% (p < 0.05), group B had 13.0% ± 3.2%, and group D had 15.2% ± 2.3% (Fig 5C).

Immunohistochemical Examination
The hearts were isolated before the initial perfusion, and samples were fixed and immunostained with anti-CD34 antibody. Capillaries are stained as dark brown. As shown in Figure 6, group C showed a significantly higher capillary density than did the other groups (p < 0.05). In addition, the effect of G-CSF was abolished by additional treatment with AG490.


Figure 6
View larger version (75K):
[in this window]
[in a new window]

  		Fig 6. Capillary density in the hearts after granulocyte colony-stimulating factor (G-CSF) treatment. (A) Capillary density was examined in rat hearts after three consecutive G-CSF treatments 20 minutes before initial perfusion. Immunohistochemical staining was proceeded as described in "Material and Methods." Capillaries are stained dark brown by 3-3'-diaminobenzidine and H2O2. The density of CD34-positive capillaries were markedly increased in group C (bar = 100 µm). (B) Data are expressed as the number of CD34 positive cells/mm2 in the 20 random fields from the each heart. Capillary density was a significantly higher in Group C than any other groups. (All values are mean ± standard deviation; n = 5 in each group; *p < 0.05.)

 

   Comment
Top
 Abstract
 Introduction
 Material and Methods
 Results
 Comment
References
 
One of the most important issues in heart transplantation is ensuring that donor grafts do not exhibit a loss of physiologic activity [12]. A longer cold ischemic preservation time is a significant predictor death at 1 year in patients undergoing heart transplantation [13]. Although many investigators have attempted to extend this time [4–6, 14], the precise mechanism of reperfusion injury after prolonged hypothermic preservation is still not clearly known.

Recently, G-CSF has been shown to act directly on cardiomyocytes and to activate the Jak/Stat pathway, which plays a critical role in up-regulating the expression of antiapoptotic proteins and angiogenetic factors in hearts after myocardial infarction [8]. Protection of the heart from normothermic ischemia–reperfusion injury by G-CSF has been demonstrated in short-term preservation [8]. In addition, Stat3 protects the heart against pathophysiologic stress, such as ischemia, mechanical stress, and cytotoxic agents [15–18]. The phosphorylation level of Stat3 was reported to be severely depressed in the myocardium obtained from patients with dilated cardiomyopathy, thus suggesting the possibility that a decrease in Stat3 activation may contribute to the development of cardiac failure [19].

In the present study, there was no remarkable difference in Stat3 activation between the control and G-CSF groups during the reperfusion period (data not shown), although the cardiac function in the G-CSF-treated group exhibited a significant extent of recovery at reperfusion. Stat3 was markedly phosphorylated at 15 minutes after treatment with 100-µg/kg G-CSF before hypothermic preservation and dephosphorylated thereafter, but this phosphorylation was abolished by the additional treatment of AG490, a selective Jak2 inhibitor. These results showed that the phosphorylation of Stat3 by G-CSF was partially mediated by Jak2 activation.

The administration of 100-µg/kg G-CSF before hypothermic preservation significantly improved cardiac functional recovery (LVDP, Max dP/dt, Min dP/dt, and coronary flow) and reduced CPK leakage during reperfusion. However, the recovery of the LVDP was completely inhibited in the presence of AG490. Although AG490 might have a detrimental effect on the cardiac function, no significant difference was found in the cardiac function among the four groups before hypothermic preservation. The expression level of Bcl-xL significantly increased, whereas that of Bax and TUNEL-positive cells significantly decreased in rats treated with 100-µg/kg G-CSF. These findings suggest that the protective effect of G-CSF in prolonged hypothermic preservation might be partly mediated by an antiapoptotic effect. These protective effects of G-CSF were completely inhibited by the additional treatment with AG490.

G-CSF has been found to mobilize bone marrow–derived stem cells and induce revascularizations [20]. Capillaries supply oxygen and metabolites locally and improve blood perfusion [21], thus probably playing a pivotal role in the cardioprotection observed in this study. These findings suggest that G-CSF increased capillary formation before hypothermic preservation and therefore might exhibit cardioprotective effects against reperfusion injury after prolonged hypothermic preservation.

Pharmacologic preconditioning with G-CSF attenuated reperfusion injury after prolonged hypothermic preservation. This effect may be related to late ischemic preconditioning, which has been demonstrated to be associated with Jak/Stat signaling [17, 18]. Bolli and colleagues [18] reported that the phosphorylation of Jak2 and Stat3 rapidly occurred after ischemia–reperfusion when mice hearts were subjected to a stimulus that elicits late preconditioning and that AG490 completely abolished the enhanced phosphorylation of Jak2 and Stat3. In addition, mice with ischemic preconditioning exhibited a reduced infarct size, and this protective effect was abolished by pretreatment with AG490 [18]. In the present study, pharmacologic preconditioning with G-CSF induced the phosphorylation of Stat3 and angiogenesis before hypothermic preservation, and this might also attenuate reperfusion injury after prolonged hypothermic preservation.

The effective dose of G-CSF administered in this study is nearly 10 times higher than that used in patients with bone marrow cell transplantation. The administration of G-CSF, which induces an increase in the WBC counts and platelet aggregation, has numerous possible complications [22]. In the present study, the WBC counts after 100-µg/kg G-CSF treatment were maintained at a level of approximately 2 x 104/µL. No significant difference was seen in the number of platelets among groups.

The life-threatening complications of G-CSF injection, including myocardial and cerebral infarctions and spontaneous rupture of the spleen, have been reported in peripheral blood stem cell transplantation in healthy humans [23]. However, no such complications were observed in this study.

The present study has some limitations. First, G-CSF may activate other intracellular signaling cascades such as that of Ras-Raf-mitogen-activated protein kinase (MAPK) kinase, as well as the Src family kinase pathways [13]. The reduction of the infarcted area obtained with G-CSF administration is abolished by the Akt inhibitor, LY294002, or the Jak2 inhibitor, AG490, but not by PD98059, a MAPK/mitogen-activated protein extracellular signal-regulated kinase (MEK) inhibitor [24]. The role of Akt and MAPK in the cardioprotective effect of G-CSF against reperfusion injury was not examined in the present study.

Second, Stat3 signaling is tightly controlled in normal cycling cells to maintain standard cellular responses [8]. Constitutively activated Stats have been detected in a wide variety of human cancer cell lines and primary tumors [18]. It was not possible to determine the tumorigenetic effects of G-CSF in this study, although such effects must be determined before the clinical application of G-CSF.

This experimental model is not clinically feasible because pretreatment with G-CSF for 2 days is required before heart transplantation. It is therefore necessary to improve and shorten the duration of G-CSF administration and to conduct more experiments before clinical application. Donor brain death is currently evaluated twice at a 6-hour interval before listing for transplantation. If less G-CSF could be administered than was used in this study, we could clinically apply this experimental model to heart transplantation. In addition, the preservation solution used in the experiment differs from clinical prevalence; however, this would not influence the results.

In summary, this study demonstrated that pharmacologic preconditioning with G-CSF attenuated reperfusion injury after prolonged hypothermic preservation. The effects of G-CSF showed an enormous increase in the capillaries before hypothermic preservation and also exhibited antiapoptotic effects at reperfusion mainly mediated by the activation of Stat3. The present study provides evidence demonstrating the cardioprotective role of G-CSF in prolonged hypothermic preservation.


   References
Top
 Abstract
 Introduction
 Material and Methods
 Results
 Comment
 References
 

  1. Kim YI, Herijgers P, Laycock SK, Lommel AV, Verbeken E, Flameng WJ, et al. Na+/H+ exchange inhibition improves long-term myocardial preservation Ann Thorac Surg 1998;66:436-442.[Abstract/Free Full Text]
  2. Jahania MS, Sanchez JA, Narayan P, Lasley RD, Mentzer RM. Heart preservation for transplantation: principles and strategies Ann Thorac Surg 1999;68:1983-1987.[Abstract/Free Full Text]
  3. Masters TN, Fokin AA, Schaper J, Lorenz-Meyer S, Pool L, Gong G, et al. Extending myocardial viability during heart preservation with cyclosporine A J Card Surg 2000;15:392-402.[Medline]
  4. Ryugo M, Sawa Y, Ono M, Fukushima N, Aleshin AN, Mizuno S, et al. Myocaedial protective effect of human recombinant hepatocyte growth factor for prolonged heart graft preservation in rats Transplantation 2004;78:1153-1158.[Medline]
  5. Oriyanhan W, Yamazaki K, Miwa S, Takaba K, Ikeda T, Komeda Ml. Taurine prevents myocardial ischemia/reperfusion–induced oxidative stress and apoptosis in prolonged hypothermic rat heart preservation Heart Vessels 2005;20:278-285.[Medline]
  6. Fukushima N, Shirakura R, Nakata S, Kaneko M, Miyagawa S, Naka Y, et al. Study of efficacies of leukocyte-depleted terminal blood cardioplegia in 24-hour preserved hearts Ann Thorac Surg 1994;58:1651-1656.[Abstract/Free Full Text]
  7. Berliner N, Hsing A, Graubert T, Sigurdsson F, Zain M, Buruno E, et al. Granulocyte colony-stimulating factor induction of normal human bone marrow progenitors results in neutrophil-specific gene expression Blood 1995;85:799-803.[Abstract/Free Full Text]
  8. Harada M, Qin Y, Takano H, Minamino T, Zou Y, Toko H, et al. G-CSF prevents cardiac remodeling after myocardial infarction by activating the Jak-Stat pathway in cardiomyocytes Nat Med 2005;11:305-311.[Medline]
  9. Minatoguchi S, Takemura G, Chen XH, Wang N, Uno Y, koda M, et al. Acceleration of the healing process and myocardial regeneration may be important as a mechanism of improvement of cardiac function and remodeling by postinfarction granulocyte colony-stimulating factor treatment Circulation 2004;109:2572-2580.[Abstract/Free Full Text]
  10. Iwanaga K, Takano H, Ohtsuka M, Hasegawa H, Zou Y, Qin Y, et al. Effects of G-CSF on cardiac remodeling after acute myocardial infarction in swine Biochem Biophys Res Commun 2004;325:1353-1359.[Medline]
  11. Hilfiler-Kleiner D, Hilfiker A, Drexler H. Many good reasons to have STAT3 in the heart Pharmacol Ther 2005;107:131-137.[Medline]
  12. Ibrahim M, Masters RG, Hendry PJ, Davies RA, Smith S, Struthers S, et al. Determinants of hospital survival after cardiac transplantation Ann Thorac Surg 1995;59:604-608.[Abstract/Free Full Text]
  13. Hosenpud JD, Bennett LE, Keck B, Fiol B, Novick RJ. The registry of the International Society for Heart and Lung transplantation: fourteenth official report-1997 J Heart Lung Transplant 1997;16:691-712.[Medline]
  14. Belzer FO, Southard JH. Principles of solid organ preservation by cold overview Transplantation 1988;45:673-676.[Medline]
  15. Kunisada K, Negoro S, Tone E, Funamoto M, Osugi T, Yamada S, et al. Signal transducer and activator of transcription 3 in the heart transduces not only a hypertrophic signal but a protective signal against doxorubicin-induced cardiomyopathy Proc Natl Acad Sci USA 1999;97:315-319.
  16. Smith RM, Suleman N, Lacerda L, Opie LH, Akira S, Chien KR, et al. Genetic depletion of cardiac myocyte STAT-3 abolishes classical preconditioning Cardiovasc Res 2004;63:611-616.[Abstract/Free Full Text]
  17. Bolli R, Dawn B, Xuan YT. Role of the JAK-STAT pathway in protection against myocardial ischemia/reperfusion injury Trends Cardiovasc Med 2003;13:72-79.[Medline]
  18. Hilfiker-Kleiner D, Hilfiker A, Fuchs M, Schaefer A, Schieffer B, hillmer A, et al. Signal transducer and activator of transcription 3 is required for myocardial capillary growth, heart protection from ischemic injury Cir Res 2004;95:187-195.[Abstract/Free Full Text]
  19. Podewski EK, Hilfiker-Kleiner D, Hilfiker A, Morawietz H, Lichtenberg A, Wollert KC, et al. Alterations in Janus Kinase (JAK)-signal transducers and activators of transcription (STAT) signaling in patients with end-stage dilated cardiomyopathy Circulation 2003;107:798-802.[Abstract/Free Full Text]
  20. Kocher AA, Schuster, MD, Szabolcs MJ, Takuma S, Burkhoff D, Wang J, et al. Neovascularization of ischemic myocardium by human bone-marrow-derived angioblasts prevents cardiomyocyte apoptosis, reduces remodeling and improves cardiac function Nat Med 2001;7:430-436.[Medline]
  21. Deindle E, Zaruba MM, Brunner S, Huber B, Mehl U, Assmann G, et al. G-CSF administration after myocardial infarction in mice attenuates late ischemic cardiomyopathy by enhanced arteriogenesis FASEB J 2006;20:E27-E36.
  22. Falzetti F, Aversa F, Minelli O, Tabilio A. Spontaneous rupture of spleen during peripheral blood stem-cell mobilization in a healthy donor Lancet 1999;353:5.
  23. Becker PS, Wagle M, Matous S, Swanson RS, Pihan G, lowry PA, et al. Spontaneous splenic repture following administration of granulocyte colony-stimulating factor (G-CSF): Occurrence in an allogeneic donor of peripheral blood stem cells Biol Blood Marrow Transplantation 1997;3:45-49.[Medline]
  24. Logue SE, Gustafasson AB, Samali A, Gottlie b RA. Ischemia/reperfusion injury at the interaction with cell death J Mol Cell Cardiol 2005;38:21-33.[Medline]



This article has been cited by other articles:


Home page
 Eur J Cardiothorac SurgHome page
N. Ishida, H. Iwata, K. Shimabukuro, E. Murakami, S. Matsumoto, H. Manabe, and H. Takemura
Effects of omentopexy combined with granulocyte colony-stimulating factor in a rabbit heart model
Eur J Cardiothorac Surg, March 1, 2011; 39(3): 375 - 380.
[Abstract] [Full Text] [PDF]

Home page
 Am. J. Physiol. Heart Circ. Physiol.Home page
M. Kurdi and G. W. Booz
JAK redux: a second look at the regulation and role of JAKs in the heart
Am J Physiol Heart Circ Physiol, November 1, 2009; 297(5): H1545 - H1556.
[Abstract] [Full Text] [PDF]

This Article
Right arrow Abstract Freely available
Right arrow Full Text (PDF)
Right arrow Alert me when this article is cited
Right arrow Alert me if a correction is posted
Right arrow Citation Map
Services
Right arrow Email this article to a friend
Right arrow Similar articles in this journal
Right arrow Similar articles in PubMed
Right arrow Alert me to new issues of the journal
Right arrow Add to Personal Folders
Right arrow Download to citation manager
Right arrow Author home page(s):
Goro Matsumiya
Norihide Fukushima
Hajime Ichikawa
Yoshiki Sawa
Right arrow Permission Requests
Citing Articles
Right arrow Citing Articles via HighWire
Right arrow Citing Articles via Google Scholar
Google Scholar
Right arrow Articles by Higuchi, T.
Right arrow Articles by Sawa, Y.
Right arrow Search for Related Content
PubMed
Right arrow PubMed Citation
Right arrow Articles by Higuchi, T.
Right arrow Articles by Sawa, Y.
Related Collections
Right arrow Transplantation - heart

HOME HELP FEEDBACK SUBSCRIPTIONS ARCHIVE SEARCH TABLE OF CONTENTS
ANN THORAC SURG ASIAN CARDIOVASC THORAC ANN EUR J CARDIOTHORAC SURG
J THORAC CARDIOVASC SURG ICVTS ALL CTSNet JOURNALS