S. Joseph Leibovich, Ph.D.
Professor

Office: MSB G677
Lab: MSB G606 / G607
Tel: (973) 972-5404
Fax: (973) 972-7489
Email: leibovic@umdnj.edu

CELL & MOLECULAR BIOLOGY OF WOUND REPAIR AND ANGIOGENESIS

The research interests of my laboratory are concerned with the role of macrophages and macrophage-derived factors in controlling wound healing and angiogenesis. Macrophages are recruited to sites of injury and inflammation, where they are activated in response to micro-environmental conditions to express either inflammatory cytokines such as TNF?, IL-1 and IL-12, or to express angiogenic growth factors such as VEGF. We have recently discovered a novel pathway that regulates the phenotype of macrophages. This pathway involves a synergistic interaction between Toll-like receptor (TLR) agonists (such as endotoxins and non-methylated CpG DNA) and adenosine A2A receptor (A2AR) agonists, and results in the strong up-regulation of VEGF expression and the strong down-regulation TNF? and Il-12 expression, thus acting as an angiogenic “switch”. This “switch” changes macrophages from an inflammatory (M1) to an anti-inflammatory, angiogenic (M2-like) phenotype. This pathway is independent of hypoxia and nitric oxide, and involves signaling through both G-protein coupled receptors and TLRs via a MyD88-dependent pathway. We are currently studying how these pathways interact to mediate the angiogenic “switch”.

Analyses using VEGF promoter constructs indicate an important role for the HIF1 transcription factor in this switch, and we are studying this requirement in detail. Also, up-regulation of A2AR expression by TLRs plays an important role in this pathway. Resting macrophages express very low levels of A2ARs, and TLR agonists strongly up-regulate A2AR expression.

We have found using global gene expression analyses that, in addition to VEGF, a small group of genes is regulated by the TLR/A2AR-mediated angiogenic switch. Most notably, sphingosine kinase-1 (SK-1), an enzyme that generates the angiogenic lipid sphingosine-1-phosphate from sphingosine, is strongly upregulated in parallel with VEGF. We are currently studying the role of SK-1 in macrophage-dependent angiogenesis. We are also studying a nitric oxide (NO)-dependent pathway in macrophages that regulates the expression of VEGF. Analyses using VEGF promoter constructs indicate that NO regulates VEGF gene transcription, in a HIF1-dependent manner. However, major regulation of VEGF expression also involves post-transcriptional regulation. The mechanisms involved in these transcriptional and post-transcriptional regulatory pathways are currently under investigation.

The importance of this TLR/A2AR-mediated angiogenic switch in the regulation of wound healing is currently under investigation, using various specific gene knockout mouse models. We are also initiating studies using mouse models to determine the role of this angiogenic switch in macrophages in the development of atherosclerosis, cardiac fibrosis and cancer.

Representative Publications:

Leibovich, S.J. The Role of Cytokines and Growth Factors in Tumor Angiogenesis. In: Human Cytokines: Their Role in Disease and Therapy:. Eds. Aggarwal, B. and Puri, R. Blackwell Scientific, pp.539-564, 1996.

Xiong, M., Elson, E. Legarda, D. and Leibovich, S.J. Production of Vascular Endothelial Growth Factor (VEGF) by Murine Macrophages: Regulation by Hypoxia, Lactate and the Inducible Nitric Oxide Synthase Pathway. Am. J. Pathol. 153, 587-598, 1998.

Leibovich, S.J., Chen, J.F., Pinhal-Enfield, G., Belem, P.C., Elson, G., Rosania, A., Ramanathan, M., Montesinos, C., Jacobson, M., Schwarzschild, M.A., Fink, J.S., Cronstein, B. Synergistic up-regulation of vascular endothelial growth factor (VEGF) expression in murine macrophages by adenosine A2A receptor agonists and endotoxin. Am. J. Pathol. 160, 2231, 2002.

Bohl-Masters, K.S., Leibovich, S.J., Belem, P., West, J.L., Poole-Warren, L.A. The effects of nitric oxide releasing hydrogel dressings on dermal wound healing in diabetic mice. Wound Rep. Regen. 10(5), 286-294, 2002.

Ramanathan, M., Giladi, A., Leibovich, S.J. Regulation of VEGF gene expression in murine macrophages by nitric oxide and hypoxia. Exp. Biol. Med. 228, 697, 2003.

Pinhal-Enfield, G., Ramanathan, Hasko, G., Vogel, S.N., Salzman, A.L., Boons, G.J., Leibovich, S.J. An angiogenic switch in macrophages involving synergy between Toll-like receptors 2, 4 and 9 and adenosine A2A receptors. Am. J. Pathol. 163, 711-721, 2003.

Nemeth, Z.H., Leibovich, S.J., Deitch, E.A., Szabo, C., Hasko, G. Microarray analysis reveals a NF-kB-independent regulation of macrophage function by adenosine. J. Pharmacol. Exp. Therapeut. 306, 1042-1049, 2003.

Nemeth, Z.H., Leibovich, S.J., Deitch, E.A., Sperlagh, B., Virag, L., Vizi, E.S., Szabo, C., Hasko, G. Adenosine stimulates CREB activation in macrophages via a p38-MAPK-mediated mechanism. Biochem. Biophys. Res. Comms. 312, 883-888, 2003.

Martin, P.A. and Leibovich, S.J. Inflammatory Cells During Wound Repair: The Good, The Bad and The Ugly. Trends in Cell Biol. 15, 599-607, 2005.

Leibovich, S.J. Vasculopathy and renal injury in lupus erythematosus: Does shedding of the endothelial protein C receptor play a role? Kidney Int. 68, 1-2, 2005.

Leibovich, S.J. Macrophages, Adenosine and Toll-Like Receptors: Their Role in the Regulation of Atherosclerotic Lesion Development. In: “Proteins Involved in the Pathogenesis of Atherosclerosis” Eds. Reiss, A.B. & Carsons, S., Pub. Research Signpost, pp. 35-62, 2006.

Ramanathan, M., Hasko, G., Leibovich, S.J. Analysis of signal transduction pathways in macrophages using expression vectors with CMV promoters: A cautionary tale. Inflammation. 29, 94-102, 2006.

Ramanathan, M., Pinhal-Enfield, G., Hao, I., Leibovich, S.J. Synergistic up-regulation of Vascular Endothelial Growth Factor (VEGF) expression in macrophages by adenosine A2A receptor agonists and endotoxin involves transcriptional regulation via the hypoxia response element (HRE) in the VEGF promoter. Mol. Biol. Cell. 18, 14-23, 2007.

Csoka, B., Nemeth, Z.H., Virag, L., Gergeley, P., Leibovich, S.J., Pacher, P., Sun, C.X., Blackburn, M.R., Vizi, E.S., Deitch, E.A., Hasko, G. A2A adenosine receptors and C/EBP???are crucially required for IL-10 production by macrophages exposed to E. coli. Blood 110, 2685-2695, 2007.

Macedo, L., Pinhal-Enfield, G., Alshits, V., Elson, G., Cronstein, B.N., Leibovich, S.J. Wound healing is impaired in MyD88-deficient mice: A role for MyD88 in the regulation of wound healing by adenosine A2A receptors. Am. J. Pathol. 171, 1774-1788, 2007.