Research interests in my laboratory center around two general areas. The first deals with the molecular and functional characterization of receptors for the endogenous nucleoside adenosine. The second addresses the potential for cardiac regeneration following myocardial infarction using embryonic stem cells.
Techniques in use:
1. Molecular biology - receptor cloning and expression, radioligand binding, siRNA, real-time PCR, western immunoblotting, ELISA, electromobility shift assay
2. Cell biology – isolation and functional analysis of mouse macrophages, neutrophils, endothelial cells, and lymphocytes
3. Mouse genetics - generation of global and tissue-specific gene 'knock-out' mouse lines, bone marrow transplantation
4. Mouse disease models - In vivo mouse infarction model, aortic banding model of cardiac hypertrophy, Langendorff-perfused isolated mouse heart model of ischemia/reperfusion injury
5. Mouse physiology – systemic blood pressure and left ventricular pressure measurements, echocardiography
Molecular and Functional Characterization of Adenosine Receptors. Adenosine is an important signaling molecule that exerts effects in essentially every organ system by interacting with four G protein-coupled receptors designated A1, A2A, A2B, and A3. Our laboratory employs modern techniques of molecular pharmacology to understand adenosine receptor function at both the molecular and physiological level. Current emphasis is focused on the two most recently identified receptors for adenosine, the A2B and A3 receptors. Studies underway in the laboratory include cloning and pharmacological characterization of adenosine receptors, analysis of adenosine receptor signaling in cells, and functional characterization of adenosine receptors in mouse models of pathogenesis using genetically modified mice.
We are currently assessing the importance of the newly discovered A3 adenosine receptor during myocardial infarction. Our laboratory has previously shown that administering agonists with high affinity for the A3 adenosine receptor reduces injury from myocardial ischemia and reperfusion in dogs, rabbits, and mice if given prior to the ischemic event or if given only during reperfusion. Using global and cardiac-specific A3 adenosine receptor gene 'knock-out' mice (Cre recombinase-loxP strategy) and bone marrow chimeric mice lacking the expression of the A3 adenosine receptor in bone marrow-derived cells, we are testing the hypothesis (see Figure 1) that activating the A3 adenosine receptor in cardiomyocytes protects against ischemic injury by improving mitochondrial function and reducing apoptosis, whereas activating the A3 adenosine receptor in immune cells during reperfusion is protective by suppressing inflammatory responses. Correlative molecular/cellular studies are underway to examine the function of the A3 adenosine receptor in specific populations of inflammatory cells isolated from the mouse including neutrophils, monocytes, macrophages, and endothelial cells.
Cardiac Regeneration Using Embryonic Stem Cells. Embryonic stem cells have the potential to differentiate into any cell type in the body and are therefore attractive for application to tissue regeneration. We and others have shown that injecting small numbers of pluripotent mouse embryonic stem cells results in cell engraftment and functional improvement of post-infarcted mouse myocardium (Figure 2),
although the mechanism for improvement is uncertain and the potential for tumorogenesis remains a significant concern. In collaboration with the Lough lab (Cell Biology, Neurobiology and Anatomy), Misra lab (Biochemistry), and others, we are addressing the hypothesis that transplantation of mouse embryonic stem cells pre-differentiated in cell culture to specific cardiac lineages (i.e., cardiomyocytes, vascular precursor cells) will reduce the potential for tumor formation and improve functional recovery due to the incorporation of functional, electrically coupled myocytes into the myocardium.
van der Hoeven D, Gizewski E, Hoshino M, Auchampach JA (2010). Activation of the A3 adenosine receptor inhibits fMLP-induced Rac activation in mouse bone marrow neutrophils. Biochemical Pharmacology 79:1667-1673.
Auchampach JA, Gizewski ET, Wan TC, de Castro S, Brown GG, Jacobson KA (2010). Synthesis and pharmacological characterization of [125I]MRS5127, a high affinity selective agonist radioligand for the A3 adenosine receptor. Biochemical Pharmacology 79:967-973.
Ge ZD, van der Hoeven D, Maas JE, Wan TC, Auchampach JA (2010). A3 adenosine receptor activation during reperfusion reduces infarct size through actions on bone marrow-derived cells. Journal of Molecular and Cellular Cardiology 49:280-286.
Kreckler LM, Wan TC, Gizewski E, Auchampach JA (2009). Adenosine suppresses LPS-induced TNF-α production from murine macrophages by inhibiting gene transcription through a PKA- and EPAC-independent signaling pathway. Journal of Pharmacology and Experimental Therapeutics 331:1051-1061.
Auchampach JA, Kreckler LM, Wan TC, Maas JE, van der Hoeven D, Gizewski E, Narayanan J, Maas GE. (2009). Pharmacological characterization of A2B adenosine receptors from mouse, rabbit, and dog. Journal of Pharmacology and Experimental Therapeutics 329:2-13.
van der Hoeven D, Wan TC, and Auchampach JA (2008). Activation of A3 adenosine receptors in mouse bone marrow neutrophils inhibits superoxide production and chemotaxis. Molecular Pharmacology 74:685-696.
van der Hoeven D, Wan TC, and Auchampach JA (2008). Activation of A3 adenosine receptors in mouse bone marrow neutrophils inhibits superoxide generation and chemotaxis. Molecular Pharmacology 74:685-696.