Editorial I: The new age of medical genomics

D. A. Schwinn* and M. Podgoreanu

Departments of Anesthesiology, Pharmacology/Cancer Biology, and Surgery, Duke University Medical Center, Durham, NC 27710, USA

* E-mail: Schwi001{at}mc.duke.edu

"I would have everie man write what he knowes and no more."—Montaigne

Genomic tools have enhanced research across all of medicine, including anaesthesia. Our understanding of both acute responses of the heart (such as myocardial preconditioning, ischaemia–reperfusion, stunning and infarction), as well as the onset and development of chronic heart disease, is undergoing a paradigm shift as functional genomic and proteomic research is providing further insights into the complex dynamic regulation of myocardial gene interactions and pathways. In this issue of the British Journal of Anaesthesia, Lucchinetti and colleagues1 demonstrate the use of genome-based technologies to discover mechanisms underlying important clinical questions such as how best to protect the heart in settings of ischaemia. One protective strategy is ischaemic preconditioning, defined as a brief period of controlled ischaemia followed by reperfusion prior to a longer period of ischaemia; certain pharmacologic agents, such as {alpha}1-adrenergic receptor agonists, ATP-sensitive K+-channel activating agents and inhalational anaesthetics, have all been shown to be as effective as ischaemia in achieving this protective effect. However, except in highly controlled settings such as the coronary catheterization laboratory or the operating theatre, clinicians rarely have the opportunity to intervene before the onset of myocardial ischaemia. Usually, the best one can provide is post-ischaemic conditioning as reperfusion strategies are initiated. Suprisingly, several groups have demonstrated that myocardial infarct size is reduced significantly, and approximately equally, with both pre- and post-ischaemic intervention. In this study, Lucchinetti and colleagues demonstrate, using an isolated perfused rat heart model and isoflurane as the pharmacological conditioning agent, that gene programmes activated by pre-ischaemic conditioning differ from those activated by post-ischaemic conditioning. Thus the molecular signatures for pre-ischaemic compared with post-ischaemic conditioning differ. This finding suggests either differences in the mechanisms underlying these two protective strategies or intersection of a few key pathways leading to protection. The authors explore each possibility in their discussion. But before exploring these findings further, a brief review of genomic technologies, including their power and limitations, is presented below.

What is genomics? Genomics refers to the study of multiple genetic variants and/or molecular pathways simultaneously. As such, genomics often utilizes technologies designed to elucidate hundreds to thousands of DNA variants (e.g. single-nucleotide polymorphism [SNP] arrays/chips), alterations in RNA expression levels (e.g. microarrays), alterations in protein levels (e.g. proteomics) and even fundamental analysis of final metabolic products (e.g. metabolomics). Because thousands of gene products are analysed simultaneously, intense mathematical analysis is required to interpret the final results. Indeed, an entire field of mathematical genomics initially developed around microarray platforms and has now expanded to other genomic approaches. Some analysis methods have been standardized and ‘packaged’, and are available in genomic array cores at most academic institutions. Differential gene expression, which compares gene/protein/metabolite expression between condition A and condition B, is most commonly used to express the results of a genomics experiment. Various methods are used to determine whether resulting differences are significant enough to report. Such analysis approaches include volcano plots and Venn diagrams. It is also important to have several replications as well as independent confirmation of at least a sampling of mRNA, protein or metabolite changes to validate the findings. Finally, in order to suggest possible biological relevance, analysis often includes clustering genes into categories of biological processes (e.g. ion channels, inflammation, cell cycle, cell–cell interactions, etc.). While genomic approaches seem ‘glitzy’, one must remember that they are simply another technical approach to answering fundamental biomedical questions, just as molecular biology and transgenic/knockout mouse biology sped the last decades of research forward. Genomics cannot replace hypothesis-driven research. However, a clear advantage of genomics approaches is that they provide an ‘unbiased’ assessment of biological pathways. Previously, scientists could only investigate biological pathways that they thought might be relevant to a given disease. Genomics approaches have facilitated discovery of new biological/physiological pathways that were not previously intuitive. However, in the end, genomics experiments are descriptive and represent only a ‘snapshot’ in time unless investigators have been careful to provide a time course in experiments utilizing genomics techniques whenever possible. In summary, therefore, genomics tools are most helpful and powerful for hypothesis generation and/or as justification for why scientists have chosen to pursue a new pathway in a given disease.

In this context, where do the findings of Lucchinetti and colleagues1 fit? Several studies have profiled myocardial gene expression in the ischaemic heart, demonstrating alterations in the expression of immediate–early genes and genes coding for calcium-handling, extracellular matrix and cytoskeletal proteins.2 3 Studies in stunned myocardium have identified upregulation of transcripts associated mechanistically in cytoprotection (heat shock proteins), resistance to apoptosis and cell growth, as well as previously uncharacterized genes.4 Microarray technology has also been utilized in the quest for novel cardioprotective genes,5 with the ultimate goal of devising strategies to activate these genes and prevent myocardial injury. Preconditioning is one of the well-studied models of cardioprotection, which can be induced (as described above) by diverse triggers including intermittent ischaemia, osmotic or redox stress, heat shock or toxins. Although acute preconditioning seems to be mediated by post-translational modifications and does not require new gene synthesis, delayed preconditioning depends on enhanced transcription of genes that lead to cardioprotection. Putative functional categories of genes involved in cardioprotective pathways include a host of transcription factors, heat shock proteins, antioxidant genes and growth factors. Results from transgenic studies have also suggested that, depending upon the level and duration of expression, certain genes (e.g. protein kinase C{varepsilon}) can have either myocardial protective or hypertrophic effects, suggesting an intersection of the molecular pathways involved in the two phenotypes.6 A preliminary step in identifying candidate genes potentially involved in cardioprotection would be to determine patterns of genes expressed in a coordinated manner that are shared across different models of cardioprotection, like preconditioning (ischaemic vs pharmacological), endogenous protection (male–female differences) or transgenic animals.

The report by Lucchinetti and colleagues1 adds to this literature by providing data on isoflurane-mediated pre- and post-ischaemic myocardial preconditioning. Their study is generally strong, utilizes cutting-edge analysis approaches and has incorporated independent validation of several mRNA changes in expression between conditions. As such, it moves the field forward. However, as the authors themselves caution, changes in mRNA expression do not always correlate with protein expression; indeed, there are some notable exceptions. Hence, some researchers now perform such experiments using proteomic approaches first, to see alterations in final proteins, and then add microarray experiments in order to determine the mechanism by which such protein changes occur. Other investigators use integrated genomic and proteomic analyses and systems biology approaches to infer genome-wide molecular networks that open a new window in the study of evolution and gene function in complex organisms. Clearly, the findings presented document that the transcriptional responses of isoflurane-mediated pre- and post-ischaemic conditioning differ. Perhaps the most exciting result is that, despite these differences, a small subset of gene alterations (22 out of 8000 genes analysed) are identified in both settings. Since approximately a quarter of the rat genome has been analysed in this study, this suggests that, in the end, perhaps as few as 100 genes might be involved in myocardial protection in both settings, despite the vast differences seen in gene programmes. Are these the key genes (or gene products at a protein level) that might suggest novel targets for intervention in the future? After all, both methods decrease myocardial injury to the same extent. Having said this, it is very interesting to note that clearly different overall mechanisms are in place for each method of anaesthetic preconditioning. Whether pre-ischaemic conditioning is better because it mimics the ‘natural’ pattern of gene expression in non-ischaemic myocardium (as the authors suggest) remains to be determined. Indeed, cardiac remodelling is not all bad: both compensatory (‘good’) and non-compensatory (‘bad’) myocardial remodelling are known to exist. With this in mind, the post-ischaemic conditioning data presented by Lucchinetti and colleagues1 may be helpful in teasing out mechanisms underlying the branch point between these two myocardial remodelling pathways. Furthermore, the development of molecular signatures that define the magnitude of myocardial injury and the response to various cardioprotective strategies may ultimately contribute to improved outcome prediction in patients undergoing coronary revascularization procedures.

In conclusion, genomic science is here to stay. While not a means unto themselves, genomic approaches effectively complement classical studies designed to examine hypothesis-driven questions, or even help to generate new hypotheses for the future.

References

1 Lucchinetti E, da Silva R, Pasch T, Schaub MC, Zaugg M. Anaesthetic preconditioning but not postconditioning prevents early activation of the deleterious cardiac remodelling programme evidence of opposing genomic responses in cardioprotection by pre- and postconditioning. Br J Anaesth 2005; 95: 140–52[Abstract/Free Full Text]

2 Deindl E, Schaper W. Gene expression after short periods of coronary occlusion. Mol Cell Biochem 1998; 186: 43–51[CrossRef][ISI][Medline]

3 Stanton LW, Garrard LJ, Damm, D, et al. Altered patterns of gene expression in response to myocardial infarction. Circ Res 2000; 86: 939–45[Abstract/Free Full Text]

4 Depre C, Tomlinson JE, Kudej RK, et al. Gene program for cardiac cell survival induced by transient ischemia in conscious pigs. Proc Natl Acad Sci USA 2001; 98: 9336–41[Abstract/Free Full Text]

5 Steenbergen C, Afshari CA, Petranka JG, et al. Alterations in apoptotic signaling in human idiopathic cardiomyopathic hearts in failure. Am J Physiol Heart Circ Physiol 2003; 284: H268–76[Abstract/Free Full Text]

6 Pass JM, Zheng Y, Wead WB, et al. PKCepsilon activation induces dichotomous cardiac phenotypes and modulates PKCepsilon–RACK interactions and RACK expression. Am J Physiol Heart Circ Physiol 2001; 280: H946–55[Abstract/Free Full Text]





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