Diane S. Keeney, Ph.D.
Assistant Professor of Medicine (Dermatology) and Biochemistry
Vanderbilt University Medical Center
607 Light Hall
Nashville, TN 37232-0146
We study roles for cytochrome P450 enzymes in the regulation of epithelial differentiation.
Stratified squamous epithelia are self-renewing and directly interface the environment. In mammals, they comprise competent barriers that provide protection against hostile environmental elements; consequently, they are primary sites of disease, including cancer. Proliferating keratinocytes from different stratified squamous epithelia have many similarities, but their terminal differentiation programs are quite distinct. For example, the outer cell layer of the epidermis is anucleated, dry, and relatively impermeable; that in oral epithelium is nucleated, wet, and relatively permeable. Phenotype reversal at these sites would be fatal. We are studying mechanisms that regulate these disparate phenotypes during terminal differentiation and role for cytochrome P450-derived metabolites in these processes.
Skin expresses many cytochromes P450 (CYP genes) having critical roles in endogenous and exogenous substrate metabolism (e.g., therapeutic drugs, xenobiotics, fatty acids, eicosanoids, sterols, steroids, vitamins A & D). We characterized three homologous P450s in rodent skin (CYP2B19, 2B15, 2B12). Mouse CYP2B19 and rat CYP2B12 were found to be arachidonic acid epoxygenases, which generate epoxyeicosatrienoic (EET) acids. These P450 enzymes were detected only in differentiated keratinocytes comprising the outer epithelial cell layers, and this cell-specific transcriptional regulation suggested important roles for these enzymes in the production and/or disposition of molecules affecting keratinocyte differentiation and epidermal functions.
EETs are natural epoxy fatty acids formed in cutaneous tissues and in cultured keratinocytes. In keratinocyte cultures, the CYP2B19 metabolite 14,15-EET activated transglutaminase enzyme activities and promoted cell cornification. In present studies, we aim to understand the mechanisms how P450-derived epoxy fatty acids regulate these cellular processes, which are critical to establishing a competent epidermal barrier. In other ongoing projects, we aim to identify functional homologues in human epidermis responsible for generating epoxy fatty acids and to develop a Cyp2b19 null mouse. These in vitro and in vivo studies will generate new insight into how endogenously formed lipid mediators (EETs) regulate functions of epidermal keratinocytes that are responsible for forming the stratum corneum and the water permeability barrier in mammals.
- Du, Liping, Hoffman, Susan M G, Keeney, Diane S Epidermal CYP2 family cytochromes P450. Toxicol Appl Pharmacol, 195(3), 278-87, 2004.
- Du, L., Hoffman, S.M.G., and Keeney, D.S. "Epidermal CYP2 family cytochromes P450.." Toxicol. Appl. Pharmacol. , in press2003.
- Ladd, Patricia A., Du, Liping, Capdevila, Jorge H., Mernaugh, Raymond, Keeney, Diane S. Epoxyeicosatrienoic acids activate transglutaminases in situ and induce cornification of epidermal keratinocytes.. J Biol Chem, 2003.
- Wang, H., Donley, K.M., Keeney, D.S., and Hoffman, S.M.G. "Organizaton and evolution of the CYP2 gene cluster on mouse chromosome 7, and a comparison to the syntenic human cluster.." Environ. Health Perspec. Toxicogenomics , in press2003.
- Hoffman, Susan M G, Keeney, Diane S Fine-scale mapping of CYP gene clusters: an example from human CYP4 family.. Methods Enzymol, 35736-44, 2002.
- Shappell, S B, Keeney, D S, Zhang, J, Page, R, Olson, S J, Brash, A R 15-Lipoxygenase-2 expression in benign and neoplastic sebaceous glands and other cutaneous adnexa.. J Invest Dermatol, 117(1), 36-43, 2001.
- Holla, V R, Adas, F, Imig, J D, Zhao, X, Price, E, Olsen, N, Kovacs, W J, Magnuson, M A, Keeney, D S, Breyer, M D, Falck, J R, Waterman, M R, Capdevila, J H Alterations in the regulation of androgen-sensitive Cyp 4a monooxygenases cause hypertension.. Proc Natl Acad Sci U S A, 98(9), 5211-6, 2001.
- Schneider, C, Keeney, D S, Boeglin, W E, Brash, A R Detection and cellular localization of 12R-lipoxygenase in human tonsils.. Arch Biochem Biophys, 386(2), 268-74, 2001.
- Hoffman, S M, Nelson, D R, Keeney, D S Organization, structure and evolution of the CYP2 gene cluster on human chromosome 19.. Pharmacogenetics, 11(8), 687-98, 2001.
- Keeney, D.S "Radiolabeled cRNA and In Situ Hybridization." Systematic Approach (Sundberg, J.P. and Bogess, D., eds.) , 145-167 , 2000.
- Brash, A R, Jisaka, M, Boeglin, W E, Chang, M S, Keeney, D S, Nanney, L B, Kasper, S, Matusik, R J, Olson, S J, Shappell, S B Investigation of a second 15S-lipoxygenase in humans and its expression in epithelial tissues.. Adv Exp Med Biol, 46983-9, 1999.
- Keeney, D.S. and Waterman, M.R "Two novel sites of expression of NADPH cytochrome P450 reductase during murine embryogenesis: limb mesenchyme and developing olfactory neuroepithelia." Develop. Dynamics , 216511-517, 1999.
- Keeney, D S, Waterman, M R Two novel sites of expression of NADPH cytochrome P450 reductase during murine embryogenesis: limb mesenchyme and developing olfactory neuroepithelia.. Dev Dyn, 216(4-5), 511-7, 1999.
- Keeney, D.S., Skinner, C., Wei, S., Friedberg, T., and Waterman, M.R "A keratinocyte-specific epoxygenase, CYP2B12, metabolizes arachidonic acid with unusual selectivity, producing a single major epoxyeicosatrienoic acid." J. Biol. Chem., 2739279-9284, 1998.
- Keeney, D S, Skinner, C, Wei, S, Friedberg, T, Waterman, M R A keratinocyte-specific epoxygenase, CYP2B12, metabolizes arachidonic acid with unusual selectivity, producing a single major epoxyeicosatrienoic acid.. J Biol Chem, 273(15), 9279-84, 1998.
- Keeney, D.S., Skinner, C., Travers, J.B., Capdevila, J.H., Nanney, L.B., King Jr., L.E., and Waterman, M.R "Differentiating keratinocytes express a novel cytochrome P450 enzyme, CYP2B19, having arachidonate oxygenase activity." J. Biol. Chem., 27332071-32079, 1998.
- Keeney, D S, Skinner, C, Travers, J B, Capdevila, J H, Nanney, L B, King, L E, Waterman, M R Differentiating keratinocytes express a novel cytochrome P450 enzyme, CYP2B19, having arachidonate monooxygenase activity.. J Biol Chem, 273(48), 32071-9, 1998.
- Keeney, D.S "The novel skin-specific cytochrome P450, Cyp2b19, maps to proximal Chromosome 7 in the mouse, near a cluster of Cyp2 family genes." Genomics, 53417-419, 1998.
- Keeney, D S The novel skin-specific cytochrome P450 Cyp2b19 maps to proximal chromosome 7 in the mouse, near a cluster of Cyp2 family genes.. Genomics, 53(3), 417-9, 1998.
- Mason, J I, Keeney, D S, Bird, I M, Rainey, W E, Morohashi, K, Leers-Sucheta, S, Melner, M H The regulation of 3 beta-hydroxysteroid dehydrogenase expression.. Steroids, 62(1), 164-8, 1997.
- Funk, C D, Keeney, D S, Oliw, E H, Boeglin, W E, Brash, A R Functional expression and cellular localization of a mouse epidermal lipoxygenase.. J Biol Chem, 271(38), 23338-44, 1996.
- StrÃ¶mstedt, M, Keeney, D S, Waterman, M R, Paria, B C, Conley, A J, Dey, S K Preimplantation mouse blastocysts fail to express CYP genes required for estrogen biosynthesis. Mol Reprod Dev, 43(4), 428-36, 1996.
- Waterman, M R, Keeney, D S Signal transduction pathways combining peptide hormones and steroidogenesis.. Vitam Horm, 52129-48, 1996.
- Keeney, D S, Ikeda, Y, Waterman, M R, Parker, K L Cholesterol side-chain cleavage cytochrome P450 gene expression in the primitive gut of the mouse embryo does not require steroidogenic factor 1.. Mol Endocrinol, 9(8), 1091-8, 1995.
- Keeney, D S, Jenkins, C M, Waterman, M R Developmentally regulated expression of adrenal 17 alpha-hydroxylase cytochrome P450 in the mouse embryo.. Endocrinology, 136(11), 4872-9, 1995.
- Keeney, D S Perspectives in steroid hydroxylase gene expression: novel sites of expression during embryonic development. Endocr Res, 21(1-2), 103-7, 1995.
- Jarow, J P, Keeney, D S, Robaire, B, Ewing, L L, Zirkin, B R In vivo model for chronic direct intratesticular drug administration.. J Androl, 15(5), 456-61, 1994.
- Keeney, D S, Murry, B A, Bartke, A, Wagner, T E, Mason, J I Growth hormone transgenes regulate the expression of sex-specific isoforms of 3 beta-hydroxysteroid dehydrogenase/delta 5-->4-isomerase in mouse liver and gonads.. Endocrinology, 133(3), 1131-8, 1993.
- Keeney, D S, Naville, D, Milewich, L, Bartke, A, Mason, J I Multiple isoforms of 3 beta-hydroxysteroid dehydrogenase/delta 5-->4-isomerase in mouse tissues: male-specific isoforms are expressed in the gonads and liver.. Endocrinology, 133(1), 39-45, 1993.
- Keeney, D S, Waterman, M R Regulation of steroid hydroxylase gene expression: importance to physiology and disease.. Pharmacol Ther, 58(3), 301-17, 1993.
- Keeney, D S, Mason, J I Expression of testicular 3 beta-hydroxysteroid dehydrogenase/delta 5----4-isomerase: regulation by luteinizing hormone and forskolin in Leydig cells of adult rats.. Endocrinology, 130(4), 2007-15, 1992.
- Waterman, M R, Keeney, D S Genes involved in androgen biosynthesis and the male phenotype.. Horm Res, 38(5-6), 217-21, 1992.
- Naville, D, Keeney, D S, Jenkin, G, Murry, B A, Head, J R, Mason, J I Regulation of expression of male-specific rat liver microsomal 3 beta-hydroxysteroid dehydrogenase.. Mol Endocrinol, 5(8), 1090-100, 1991.
- Keeney, D S, Ewing, L L Effects of hypophysectomy and alterations in spermatogenic function on Leydig cell volume, number, and proliferation in adult rats.. J Androl, 11(4), 367-78, 1990.
- Keeney, D S, Sprando, R L, Robaire, B, Zirkin, B R, Ewing, L L Reversal of long-term LH deprivation on testosterone secretion and Leydig cell volume, number and proliferation in adult rats.. J Endocrinol, 127(1), 47-58, 1990.
- Mendis-Handagama, S M, Keeney, D S, Hardy, M P, Ewing, L L Application of the disector method to enumerate cells in the testis.. Ann N Y Acad Sci, 56486-98, 1989.
- Juniewicz, P E, Keeney, D S, Ewing, L L Effect of adrenocorticotropin and other proopiomelanocortin-derived peptides on testosterone secretion by the in vitro perfused testis.. Endocrinology, 122(3), 891-8, 1988.