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J Thorac Cardiovasc Surg 2005;130:330-339
© 2005 The American Association for Thoracic Surgery

Cardiopulmonary Support and Physiology

Differential cardiac gene expression during cardiopulmonary bypass: Ischemia-independent upregulation of proinflammatory genes

Mihai V. Podgoreanu, MD a , * , Gregory A. Michelotti, PhD a , * , Yukie Sato, MD a , Michael P. Smith, MSc a , Simon Lin, MD d , Richard W. Morris, PhD a , Hilary P. Grocott, MD a , Joseph P. Mathew, MD a , Debra A. Schwinn, MD a , b , c

a Department of Anesthesiology, Duke University Medical Center, Durham, NC
b Department of Surgery, Duke University Medical Center, Durham, NC
c Department of Pharmacology/Cancer Biology, Duke University Medical Center, Durham, NC
d Duke Comprehensive Cancer Center (BioInformatics Shared Resource), Duke University Medical Center, Durham, NC

Read in part at Fourth Scientific Forum on Quality of Care and Outcomes Research in Cardiovascular Disease and Stroke, Washington, DC, October 12–14, 2002.

Received for publication June 11, 2004; revisions received November 2, 2004; accepted for publication November 5, 2004.

* Address for reprints: Mihai Podgoreanu, MD, Department of Anesthesiology, Duke University Medical Center, Genome Science Research Bldg 1, 595 LaSalle St, Ste 1027, Durham, NC 27710 (Email: podgo001{at}mc.duke.edu).


   Abstract
Top
 Abstract
Introduction
 Methods
 Results
 Discussion
 Conclusion
 References
 
OBJECTIVE: Cardiac surgery with cardiopulmonary bypass induces both systemic and local inflammatory responses implicated in the pathogenesis of myocardial dysfunction. Multifactorial perioperative sources of myocardial injury complicate understanding of the molecular mechanisms involved. By using microarray technology, this study examines myocardial gene expression responses to cardiopulmonary bypass in the absence of cardioplegic arrest and ischemia-reperfusion injury.

METHODS: We used a unique rat model of cardiopulmonary bypass in which sternotomy, direct operations on the heart, aortic crossclamping, and cardioplegic arrest were not performed. Hearts from 6 animals randomized to either 90 minutes of cardiopulmonary bypass or sham control animals were used to perform cDNA microarray analyses of 2343 genes. Real-time quantitative polymerase chain reaction was used to confirm the microarray results for a subset of genes.

RESULTS: Compared with sham-operated control animals, myocardium from animals undergoing cardiopulmonary bypass revealed 42 differentially expressed genes. Upregulated genes include the transcription activator nuclear factor {kappa}B, adhesion molecules (vascular cell adhesion molecule 1 and P-selectin), and interleukin 6 receptor subunits; downregulated genes include transforming growth factor ß receptor 2, tissue inhibitor of metalloproteinase 3, and mitogen-activated protein kinase 1. Distinct proinflammatory gene cascades were confirmed by means of category overrepresentation analysis.

CONCLUSIONS: This study represents an initial report on the use of microarray technology to elucidate cardiac transcriptional programs in response to cardiopulmonary bypass-specific injury in vivo. These preliminary findings, combined with future functional genomic studies superimposing ischemia and reperfusion and other inflammatory stimuli, should improve our understanding of the molecular regulatory networks involved in myocardial responses to injury and aid in the development of novel cardioprotective and perfusion strategies.


   Introduction
Top
 Abstract
 Introduction
Methods
 Results
 Discussion
 Conclusion
 References
 

Figure 1
Michelotti and Podgoreanu (left to right)


Almost 50 years after the first clinical application of cardiopulmonary bypass (CPB), the pathophysiology of injury-induced myocardial dysfunction after cardiac operations with CPB remains poorly characterized. Numerous experimental and clinical studies have demonstrated that CPB leads to a systemic inflammatory response triggered by bioincompatibility of blood-contacting surfaces, surgical trauma, ischemia and reperfusion, and endotoxemia. 1 Go It is increasingly recognized that this generalized inflammation is implicated in the pathogenesis of post-CPB cardiovascular dysfunction, including myocardial stunning, ischemia, and ß-adrenergic desensitization. 2,3 Go In addition to the well-described changes in circulating levels of proinflammatory mediators associated with CPB, it has been established that the heart itself participates in the host inflammatory response to cardiac surgery, 4 Go but underlying molecular regulatory mechanisms remain poorly characterized.

Transcriptional profiling has emerged as a powerful tool for delineating complex patterns of tissue-specific gene expression in response to severe systemic stimuli and injury, 5 Go and several recent studies have used microarray technology to examine the global myocardial stress response during cardiac surgery. 6,7 Go However, interpretation of such gene expression analyses is complicated by the activation of biologic cascades stemming from the combined insults of cardioplegic arrest, ischemia and reperfusion, and direct tissue injury in addition to the effects of CPB. Therefore, to begin dissecting the molecular mechanisms underlying myocardial responses to specific CPB-initiated injury, we used a unique model of rat CPB in which median sternotomy, direct operations on the heart, aortic crossclamping, and cardioplegic arrest were not performed. Specifically, we tested the hypothesis that nonpulsatile normothermic CPB alone (in the absence of cardioplegic arrest and ischemia-reperfusion) activates both systemic proinflammatory responses and myocardial inflammatory gene cascades. Such differential gene expression profiling in response to different myocardial stressors or insults might provide mechanistic insights that aid in the development and evaluation of novel cardioprotective and perfusion strategies.


   Methods
Top
 Abstract
 Introduction
 Methods
Results
 Discussion
 Conclusion
 References
 
Surgical Preparation and CPB
All animal experiments were performed in compliance with the "Guide for the Care and Use of Laboratory Animals." 8 Go Details of the closed-chest rat CPB model used in the current study (Figure 1) have been previously described. 9 Go Adult male Sprague-Dawley rats (325–375 g; Harlan, Indianapolis, Ind) were anesthetized with isoflurane, orotracheally intubated, and mechanically ventilated (40% O2/balance N2). Surgical preparation involved cannulation of the ventral tail artery (arterial inflow) by using a 20-gauge catheter and, after systemic heparinization (150 U/kg), insertion of a 4.5F multiorifice cannula through the external jugular vein into the right atrium (venous outflow). The anesthetic was then converted to fentanyl (150 µg/kg administered intravenously), diazepam (2 mg/kg administered intravenously), and pancuronium (0.2 mg/kg administered intravenously). Animals were randomized to 90 minutes of normothermic, nonpulsatile CPB (n = 3) or to serve as sham-operated control animals (n = 3) after undergoing identical surgical preparation and anesthetic regimen. The CPB circuit consisted of a venous reservoir, a peristaltic pump (Masterflex; Cole-Parmer Instrument Co, Vernon Hills, Ill), and a membrane oxygenator (Micro neonatal oxygenator; Cobe Cardiovascular, Inc, Arvada, Colo) connected through 1.6-mm internal diameter silicone tubing (Tygon, Cole-Parmer Instrument Co). The CPB circuit was primed with approximately 40 mL of whole blood obtained before the start of the experiment from 2 heparinized (100 U per rat, intravenous) donor rats that were exsanguinated during isoflurane-induced anesthesia; the activated clotting time was maintained at greater than 450 seconds for the duration of CPB. All sham-operated animals were given equivalent heparin doses. The targeted CPB flow was 160 to 180 mL · kg–1 · min–1, corresponding to normal cardiac output in the rat, 10 Go and the inflow temperature was maintained at 37.5°C by using a circulating water bath system. Whole hearts were then snap-frozen in liquid nitrogen and stored at –80°C.


Figure 1
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  		Figure 1. Schematic diagram of the surgical preparation and rat CPB apparatus. (From Mackensen GB, Sato Y, Nellgard B, Pineda J, Newman MF, Warner DS, Grocott HP. Cardiopulmonary bypass induces neurologic and neurocognitive dysfunction in the rat. Anesthesiology. 2001;95:1485–91. Reproduced with permission of Lippincott Williams & Wilkins.)

 
Quantification of Plasma Cytokine Levels
Characterization of the systemic inflammatory response in this closed-chest small-animal model of CPB was performed by quantifying plasma cytokine levels (interleukin [IL] 6 and IL-10) at the end of the experimental period (90 minutes). Evaluations were performed in duplicate by using commercially available enzyme-linked immunosorbent assay kits (Pierce Endogen, Inc, Rockford, Ill), according to the manufacturer’s protocol. Changes in IL-6 and IL-10 levels (plasma concentration after surgical intervention minus baseline plasma concentration) were compared between experimental groups by using the Wilcoxon rank sum test in SAS (version 8; SAS Institute, Cary, NC). Physiologic parameters were compared between groups by using repeated-measures analysis of variance, followed by Newman-Keuls tests for multiple comparisons as appropriate.

RNA Isolation, Labeling, and cDNA Array Hybridization
Total RNA was extracted from tissue samples (whole hearts) by using the Trizol method (Gibco BRL Life Technologies, Rockville, Md). Genomic DNA contamination was removed by means of digestion of RNA samples with DNase I (Ambion, Inc, Austin, Tex) and confirmed by means of polymerase chain reaction (PCR) using glyceraldehyde-phosphate dehydrogenase (GAPDH)–specific primers. Radiolabeled cDNA probes were synthesized by means of reverse transcription of total RNA (5 µg) in the presence of {alpha}-[32P]dCTP and hybridized to Atlas Rat cDNA expression nylon arrays (Rat 1.2 and 1.2II; Clontech, BD Biosciences, Palo Alto, Calif) for 12 hours at 68°C, according to the manufacturer’s protocol. The arrays were washed in 2x saline sodium citrate/1% sodium dodecylsulfate at 68°C for 2 hours and in 0.1x saline sodium citrate/0.5% sodium dodecylsulfate at 68°C for 30 minutes and exposed to low-energy storage phosphor screens (Molecular Dynamics, Sunnyvale, Calif) for 24 hours.

Microarray Data Analysis
Phosphor images were acquired with a PhosphorImager Storm system (Molecular Dynamics) at 100 µm resolution. Hybridized spots intensities on the microarray were quantified with ImageQuant (Molecular Dynamics) and AtlasImage version 2.02 (Clontech, BD Biosciences) software. Background intensities were subtracted from the hybridization signal, and genes with background-adjusted intensities of less than twice the background value were filtered out. The annotation of genes spotted on the Clontech Atlas Rat v1.2 (I and II) microarray was updated by querying the National Center for Biotechnology Information (NCBI) Unigene database and using the 2370 GenBank identifiers provided by the manufacturer. To remove systematic intensity differences among arrays caused by differences in film exposure, labeling efficiency, and hybridization-washing conditions, a variance stabilizing transformation and normalization (VSN) method 11 Go was applied to each hybridization image as follows:


Formula

where yki is the background-corrected intensity of gene k on array i, and ai and bi are affine parameters optimized by the VSN algorithm by using maximum-likelihood estimations for array i. All data analysis was in compliance with the Minimum Information About Microarray Experiments standards (http://www.mged.org/Workgroups/MIAME/miame.html), and the complete data sets were deposited in the NCBI Gene Expression Omnibus database (http://www.ncbi.nlm.nih.gov/geo/) under accession numbers GSM14486-14497 for the individual samples and GSE921, GSE922, and GSE924 for the experimental series. Detection of differentially expressed genes was performed by using the method developed by Lonnstedt and Speed 12 Go in the statistical package R (http://lib.stat.cmu.edu/R/CRAN). A Bayesian B-score defined as


Formula 1

(1)
was used in conjunction with the empirical M score (the average difference between the 2 experimental groups) to select the best candidate genes modulated by CPB. Gene-to-gene correlations were calculated by using the pairwise correlation function in the statistical package R without missing value estimation. Hierarchical clustering was used to group genes on the basis of similarity in expression across the samples by using the Cluster software package (http://rana.lbl.gov/EisenSoftware.htm).

Category overrepresentation analysis was performed with the Expression Analysis Systematic Explorer software, as previously described, to identify and prioritize biologic themes within the lists of differentially expressed genes. 13 Go By using the 3 systems of Gene Ontology (http://www.geneontology.org/) as categorization systems, the Expression Analysis Systematic Explorer software calculates the statistical measure of overrepresentation of differentially expressed genes with respect to the total number of genes assayed and annotated within each system. This is reported as the 1-tailed Fisher exact probability corrected for multiple comparisons by using a bootstrap function with 10,000 random trials. 13 Go

Real-Time Reverse Transcriptase-Polymerase Chain Reaction
Reverse transcriptase-polymerase chain reaction (RT-PCR) was used to validate microarray data for 7 genes selected on the basis of involvement in recognized proinflammatory and immunologic pathways: IL-6 signal transducer (gp130), mitogen-activated protein kinase 1 (MAPK1), nuclear factor {kappa}B p105 subunit (NFKB1), platelet selectin (SELP), tissue inhibitor of metalloproteinase 3 (TIMP3), transforming growth factor ß receptor 2 (TGFBR2), and vascular cell adhesion molecule 1 (VCAM1). Total heart RNA (3 µg) from each animal was reverse transcribed with commercially available reagents (Perkin Elmer Life Sciences, Boston, Mass). RT-PCR was performed in duplicate by using the LightCycler Instrument (Roche Diagnostics, Indianapolis, Ind) and SYBR Green I fluorescence for detection. Primer sequences for the validation genes were as follows: gp130 (5'-gtggcccagcatcaatgtgtcatcc; 3'-agaacttccgtactgatcctcgtgg), MAPK1 (5'-gggccgcgctacactaatctctc; 3'-ccggatgatgtcattgatgccgatg), NFKB1 (5'-tcttcgactacgcggttacgggag; 3'-gatcacggccaagtgcaaaggtgtc), SELP (5'-caataagactctcacggcggaggc; 3'-caggtgtagctcccaatggtctcg), TIMP3 (5'-gtacacagggctgtgcaactttgtg; 3'-cttctgccggatgcaggcgtagtg), TGFBR2 (5'-ggagtccttcaagcagacggatgtc; 3'-cagcactcggtcagtgtctcacac), VCAM1 (5'-ggctacatccacactgacgctgag; 3'-cccttcagtagttcaatctccagatgg), GAPDH (5'-gaccccttcattgacctcaac; 3'-cttctccatggtggtgaaga). PCR conditions were 0.2 U of Platinum Taq (Hot Start; Invitrogen Life Technologies, Carlsbad, Calif) with supplied buffer, 1 µL of SYBR Green I (Molecular Probes, Inc, Eugene, Ore) diluted 1:1500, 0.25 µmol/L of each PCR primer, 200 µmol/L of each deoxyribonucleoside triphosphate, and 120 ng of cDNA in a final volume of 20 µL. The amplification profile was as follows: initial denaturation at 95°C for 2 minutes and then 40 cycles of 95°C for 5 seconds, 58°C for 10 seconds, and 72°C for 5 seconds. Tests of optimal annealing conditions and melting-curve analysis were conducted for each set of gene-specific primers; typically, fluorescence was acquired on channel F1 at 85°C for 2 seconds. Fluorescence curves were analyzed by means of a second derivative method with the LightCycler Quantification Software v1.0 (Roche Diagnostics). Relative transcript abundance (normalized to GAPDH) was determined by means of comparison with 3 control samples serially diluted 10-fold. Amplicons were recovered after separation by means of electrophoresis on a 2% Tris-borate agarose gel, cloned into a plasmid, and sequenced to verify amplification of the correct PCR product.


   Results
Top
 Abstract
 Introduction
 Methods
 Results
Discussion
 Conclusion
 References
 
Physiologic Characterization of the Rat CPB Model
There were no significant differences in hemodynamic, gas exchange, and acid-base parameters between CPB animals and sham control animals throughout the experimental protocol. After an initial decrease in the CPB group, differences in hemoglobin levels between experimental groups remained not significant (Table 1). Plasma cytokine levels increased to greater than baseline values significantly more in the CPB group than in the sham-operated animals for both IL-6 (median [interquartile range]: 14,307 [10,006–23,620] vs 1753 [0-2160] pg · mL–1, P < .01) and IL-10 (median [interquartile range]: 770 [638–952] vs 111 [69–182] pg · mL–1, P < .01). However, the average relative increase from baseline concentrations was larger for IL-6 (18.9-fold) than for IL-10 (5.2-fold) in CPB animals compared with that seen in sham control animals, which is consistent with a shift in circulating cytokine balance toward a proinflammatory response.


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  		TABLE 1. Physiologic parameters in the experimental model of rat CPB-treated animals and sham-operated control animals
 
Identification of Differentially Expressed Genes
Of the 2370 genes spotted on the Atlas Rat microarray, 2343 unique genes were identified after querying the NCBI Unigene database. Of these, 618 (approximately 26%) had detectable hybridization levels (ie, threshold intensity at least twice the background value) under at least one of the 2 experimental conditions and were therefore scored as a genuine signal. After variance stabilizing transformation and normalization, the Pearson correlation coefficients for the pairs of replicates ranged from 0.90 to 0.94 (P < .001), indicating a high level of reproducibility. To identify genes differentially expressed in the heart in response to CPB, we compared triplicate microarray hybridizations from the 2 experimental groups. A volcano plot 14 Go of average difference (M score) under the 2 conditions and its statistical confidence (B score) for each gene is shown in Figure 2. The B score provides a stringent threshold to prevent false-positive identifications by taking the measurement variations into consideration and discounting transcripts with large average differences but also large variations between animals within experimental groups. 12 Go With an empiric rule of M greater than 0.5 and B greater than 0, we identified 17 upregulated and 25 downregulated genes, which are listed alphabetically in Table 2. The hierarchical clustering shows that the expression profiles of the 3 bypass-treated animals belong to 2 distinct clusters (Figure 3).


Figure 2
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  		Figure 2. Volcano plot of significance against effect. Bayesian statistical approaches were used to assess the statistical significance of microarray data, as described in the "Methods" section. For each gene, a Bayesian B score is calculated and plotted versus the average difference between the 2 experimental groups (M score). By using stringent statistical cutoff values, 25 downregulated (green) and 17 upregulated genes (red) were identified in response to CPB.

 

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  		TABLE 2. Summary of genes differentially expressed with CPB
 

Figure 3
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  		Figure 3. Unsupervised hierarchical clustering of 42 myocardial genes differentially expressed with CPB. Data from animals undergoing CPB are presented in the left 3 columns, and data on sham control animals are presented in the right 3 columns. The upper panel demonstrates genes downregulated (green), whereas the lower panel shows genes upregulated (red) compared with sham control animals. For each gene, the mean normalized hybridization intensity is calculated within each experimental group, and relative expression of each animal is presented relative to the calculated mean. A list of the exact genes shown is further detailed in Table 2.

 
Validation of mRNA Changes
We confirmed the technical quality of the microarray data in 7 selected transcripts, with fold changes in expression level by means of real-time quantitative RT-PCR correlating highly with fold changes in background-adjusted intensities on the microarray (Pearson correlation coefficient, 0.946; Figure 4). Confirmed genes correspond to the 2 clusters identified by means of hierarchical analysis. The upregulated genes include VCAM1, SELP, NF-{kappa}B (NFKB1), and IL-6 signal transducer (gp130). The second cluster corresponds to genes identified as downregulated in the CPB group and includes MAPK1, TGFBR2, and TIMP3. Although most of these transcripts are known to be involved in immune and inflammatory responses, they have not previously been reported as being differentially expressed at the level of the myocardium in response to CPB.


Figure 4
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  		Figure 4. Confirmation of microarray data by means of real-time quantitative RT-PCR in a subset of 7 genes differentially expressed with CPB. Primers for each gene are listed in the "Methods" section. GAPDH was used to normalize mRNA levels between samples, and values for each gene are shown as fold change in CPB-treated animals versus those seen in sham-operated control animals (mean ± SEM; results represent n = 3 for 3 independent experimental groups).

 
Exploring Biologic Themes in Differentially Expressed Myocardial Genes
Themes of genes upregulated with CPB, as determined by means of overrepresentation analysis, included the following: inflammatory response (P = .015), response to external stimulus (P = .016), response to abiotic stimulus (P = .029), response to biotic stimulus (P = .039), and chemokine activity (P = .042). Significant themes among the gene categories downregulated with CPB include calcium ion binding (P = .004) and nucleocytoplasmic transport (P = .01).


   Discussion
Top
 Abstract
 Introduction
 Methods
 Results
 Discussion
Conclusion
 References
 
CPB causes a systemic inflammatory response, which can lead to organ failure and increased postoperative morbidity. 1 Go However, direct effects of CPB on the myocardium, particularly in the absence of ischemia and reperfusion and cardioplegic arrest, remain poorly understood. We used a previously described rat model of CPB and microarray technology to investigate myocardial gene expression changes in response to the stimulus of CPB. First, validity of this unique closed-chest small-animal model of CPB in eliciting a systemic inflammatory response was determined; we show that rats exposed to CPB display a robust increase in circulating levels of IL-6 and IL-10 and a shift toward a proinflammatory response consistent with previous findings in patients and larger animal models. Furthermore, microarray-based gene expression profiling suggests that CPB induces ischemia-independent transcriptional activation of myocardial proinflammatory gene programs in the rat.

Category overrepresentation analysis identified 4 biologic themes among the upregulated transcripts annotated to main Gene Ontology biologic processes, such as inflammatory response, response to external stimulus, response to abiotic stimulus, and response to biotic stimulus. Such upregulated genes include neutrophil and endothelial adhesion molecules (VCAM1 and SELP), cytokine receptors (IL6RA and gp130), proinflammatory chemokines (IP10), and inflammatory enzymes (phospholipase A2). Coregulation of these genes conceivably could be explained by activation of the common transcriptional inducer NF-{kappa}B because they all share NF-{kappa}B-binding motifs in their promoter regions. 15–19 Go NF-{kappa}B is a transcriptional regulator consisting of homodimers and heterodimers of 5 proteins of the Rel family, including p65/RelA, p105/p50/NFKB1, p100/p52/NFKB2, c-Rel, and RelB. Regulation of NF-{kappa}B activation is exceedingly complex, involving integration of transcriptional, posttranscriptional, and cytoplasmic events, culminating in nuclear translocation of NF-{kappa}B dimers and binding to specific promoter elements of target genes. 20 Go Although the processing mechanisms that yield active p50 from the precursor p105 molecule remain to be fully elucidated, previous studies indicate that p105 gene expression directly correlates with NF-{kappa}B transcriptional activity 18,21 Go and is upregulated in response to different inflammatory stimuli. 22 Go We find a strong (8.2-fold) induction of p105 gene expression in response to CPB, which is highly consistent with other proinflammatory and immunomodulatory genes identified in our model system. Although previous studies have demonstrated NF-{kappa}B involvement in the pathophysiology of myocardial injury associated with cardiac surgery, 23 Go they lacked the ability to differentiate between the roles of cardioplegic arrest and ischemia and reperfusion versus CPB exposure. Our findings suggest that myocardial NF-{kappa}B transcriptional activity might also be influenced by ischemia-independent mechanisms, leading to local activation of proinflammatory cascades. However, these data do not allow differentiation between the roles of nonpulsatile flow and exposure to CPB itself in explaining the observed changes in myocardial gene expression, and further studies comparing pulsatile and nonpulsatile CPB are required.

We demonstrated a significant increase in plasma IL-6 levels after CPB previously associated with circulatory dysregulation and myocardial dysfunction. 24 Go IL-6 is a pleiotropic cytokine that exerts its many actions through a heterodimeric receptor consisting of 2 membrane-bound glycoproteins (IL-6 binding subunit [IL6RA] and IL-6 signal transducer [gp130]), which are responsible for signal transduction and orchestration through the JAK/STAT pathway, and shared by other cytokines of the IL-6 family. 25 Go Myocardial mRNA expression of both IL-6 receptor subunits increased in the CPB group, making them attractive targets for attenuation of IL-6–mediated injury during cardiac surgery.

In addition to the observed activation of proinflammatory genes, suppression of local anti-inflammatory activity is suggested by the significant downregulation of myocardial transforming growth factor receptor 2 (TGFBR2) mRNA, a key molecule modulating various TGF-ß–mediated immunosuppressive and immunomodulatory effects. 26 Go Interestingly, 2 novel biologic themes, namely calcium ion binding and nucleocytoplasmic transport, have been identified among the genes downregulated in response to CPB and prioritized for further analysis and functional characterization.

This work has several limitations apart from those discussed above. First, our findings provide only preliminary insight into potential pathways that are activated in a defined temporal window and suggest orchestration of a programmed response to upregulate proinflammatory genes, likely in an NF-{kappa}B-dependent mechanism. Confirming the physiologic significance of the observed transcriptional changes requires the analysis of temporal gene expression profiles and the association with changes in protein expression. Second, the presence of heterogeneous myocardial cell populations in a whole-heart preparation is partially counterbalanced by the fact that the biologic relevance of our findings (the proposed physiome) might be lost in the absence of cell-cell interactions. A third limitation of this study involves the relevance of the small-animal model to the human clinical condition. Although hemodynamic, gas exchange, and acid-base parameters are similar to clinical CPB in human subjects, the higher proportion of blood-foreign surface interaction relative to body weight in the rat model and the use of donor rat blood to prime the CPB circuit are current technologic limitations of rat CPB, which could conceivably alter patterns of gene expression. Despite this, a number of factors suspected to induce myocardial injury during cardiac surgery, including the inflammatory response, are replicated by this model.


   Conclusion
Top
 Abstract
 Introduction
 Methods
 Results
 Discussion
 Conclusion
References
 
This study represents an initial report on the use of broad-scale genomic technology to elucidate ischemia-independent myocardial transcriptional programs in response to CPB-induced injury in vivo. Consistent with recent data showing a robust systemic inflammatory response during CPB in both human and animal models, we have shown significant increases in circulating IL-6 and IL-10 levels balanced toward inflammation. Furthermore, microarray-based gene expression analysis identified a number of proinflammatory genes upregulated in response to CPB. The transcription factor NF-{kappa}B might be a nexus of control in the myocardial inflammatory activation because most genes identified appear to be regulated in an NF-{kappa}B-dependent manner. These preliminary findings, combined with future studies superimposing ischemia-reperfusion injury with or without CPB, should help elucidate the role played by (dys)regulation of inflammatory pathways in the response to myocardial injury. Studies such as these should ultimately aid in the development of novel pharmacologic and genetic cardioprotective strategies for cardiac surgical patients.


   Footnotes
 
Funded by National Institutes of Health grant HL57447 (D.A.S.) and American Heart Association grant 0120492U (M.V.P.). Dr Schwinn is a senior fellow of the Duke University Center for the Study of Aging and Human Development.

* M.V.P. and G.A.M. contributed equally to this article. Back


   References
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 Abstract
 Introduction
 Methods
 Results
 Discussion
 Conclusion
 References
 

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