Abstract
Aging is understood as the result of a complex interaction of biological processes that are caused by both environmental processes (extrinsic aging) and genetic processes (intrinsic aging). Research into the biology of aging has provided detailed insight into the molecular mechanisms of age-related changes in organs, tissues, and cells. Most information relating to intrinsic aging processes comes from tissues other than the skin. This is in part due to the fact that clinically manifest diseases such as type 2 diabetes or neurodegenerative disease are often correlated with aging of cells. In part it is also due to the fact that substantial amounts of primary cells and organelles for biochemical analyses can be more easily isolated from other organs such as the muscle, brain, or liver, as compared with skin. Nevertheless, intrinsic aging is based on general biological processes that apply more or less to all proliferating cells and terminally differentiated cells as well. Therefore, general intrinsic aging processes seen in a liver cell, muscle cell, or neuron can be expected also to apply more or less to skin cells. In fact, most of the aging processes identified and studied with other cells could also be confirmed with keratinocytes or dermal fibroblasts, even though some downstream details may be different.
References
Birch-Machin MA. The role of mitochondria in ageing and carcinogenesis. Clin Exp Dermatol. 2006;31:548–52.
Jendrach M, Pohl S, Voth M, et al. Morpho-dynamic changes of mitochondria during ageing of human endothelial cells. Mech Ageing Dev. 2005;126:813–21.
Hansford RG. Bioenergetics in aging. Biochim Biophys Acta. 1983;726:41–80.
Goldstein S, Moerman EJ, Porter K. High-voltage electron microscopy of human diploid fibroblasts during ageing in vitro. Morphometric analysis of mitochondria. Exp Cell Res. 1984;154:101–11.
Brantova O, Tesarova M, Hansikova H, et al. Ultrastructural changes of mitochondria in the cultivated skin fibroblasts of patients with point mutations in mitochondrial DNA. Ultrastruct Pathol. 2006;30:239–45.
Feldman D, Bryce GF, Shapiro SS. Mitochondrial inclusions in keratinocytes of hairless mouse skin exposed to UVB radiation. J Cutan Pathol. 1990;17:96–100.
Guillery O, Malka F, Frachon P, et al. Modulation of mitochondrial morphology by bioenergetics defects in primary human fibroblasts. Neuromuscul Disord. 2008;18:319–30.
Lazarou M, McKenzie M, Ohtake A, Thorburn DR, Ryan MT. Analysis of the assembly profiles for mitochondrial- and nuclear-DNA-encoded subunits into complex I. Mol Cell Biol. 2007;27:4228–37.
Papa S. Mitochondrial oxidative phosphorylation changes in the life span. Molecular aspects and physiopathological implications. Biochim Biophys Acta. 1996;1276:87–105.
Zwerschke W, Mazurek S, Stockl P, et al. Metabolic analysis of senescent human fibroblasts reveals a role for AMP in cellular senescence. Biochem J. 2003;376:403–11.
Sugrue MM, Tatton WG. Mitochondrial membrane potential in aging cells. Biol Signals Recept. 2001;10:176–88.
Rottenberg H, Wu S. Mitochondrial dysfunction in lymphocytes from old mice: enhanced activation of the permeability transition. Biochem Biophys Res Commun. 1997;240:68–74.
Yen TC, Chen YS, King KL, Yeh SH, Wei YH. Liver mitochondrial respiratory functions decline with age. Biochem Biophys Res Commun. 1989;165:944–1003.
Trounce I, Byrne E, Marzuki S. Decline in skeletal muscle mitochondrial respiratory chain function: possible factor in ageing. Lancet. 1989;1:637–9.
Mammone T, Gan D, Foyouzi-Youssefi R. Apoptotic cell death increases with senescence in normal human dermal fibroblast cultures. Cell Biol Int. 2006;30:903–9.
Greco M, Villani G, Mazzucchelli F, et al. Marked aging-related decline in efficiency of oxidative phosphorylation in human skin fibroblasts. FASEB J. 2003;17:1706–8.
Lenz H, Schmidt M, Welge V, et al. The creatine kinase system in human skin: protective effects of creatine against oxidative and UV damage in vitro and in vivo. J Invest Dermatol. 2005;124:443–52.
Paz ML, Gonzalez Maglio DH, Weill FS, Bustamante J, Leoni J. Mitochondrial dysfunction and cellular stress progression after ultraviolet B irradiation in human keratinocytes. Photodermatol Photoimmunol Photomed. 2008;24:115–22.
Jongkind JF, Verkerk A, Poot M. Glucose flux through the hexose monophosphate shunt and NADP(H) levels during in vitro ageing of human skin fibroblasts. Gerontology. 1987;33:281–6.
Stucker M, Struk A, Altmeyer P, et al. The cutaneous uptake of atmospheric oxygen contributes significantly to the oxygen supply of human dermis and epidermis. J Physiol. 2002;538:985–94.
Ronquist G, Andersson A, Bendsoe N, Falck B. Human epidermal energy metabolism is functionally anaerobic. Exp Dermatol. 2003;12:572–9.
Hornig-Do HT, von Kleist-Retzow JC, Lanz K, et al. Human epidermal keratinocytes accumulate superoxide due to low activity of Mn-SOD, leading to mitochondrial functional impairment. J Invest Dermatol. 2007;127:1084–93.
Dzeja PP, Terzic A. Phosphotransfer networks and cellular energetics. J Exp Biol. 2003;206:2039–47.
Bessman SP, Carpenter CL. The creatine-creatine phosphate energy shuttle. Annu Rev Biochem. 1985;54:831–62.
Blatt T, Lenz H, Koop U, et al. Stimulation of skin’s energy metabolism provides multiple benefits for mature human skin. Biofactors. 2005;25:179–85.
Wilken B, Ramirez JM, Probst I, Richter DW, Hanefeld F. Creatine protects the central respiratory network of mammals under anoxic conditions. Pediatr Res. 1998;43:8–14.
Bessman SP. The creatine phosphate energy shuttle--the molecular asymmetry of a “pool”. Anal Biochem. 1987;161:519–23.
Wallimann T, Wyss M, Brdiczka D, Nicolay K, Eppenberger HM. Intracellular compartmentation, structure and function of creatine kinase isoenzymes in tissues with high and fluctuating energy demands: the ‘phosphocreatine circuit’ for cellular energy homeostasis. Biochem J. 1992;281(Pt 1):21–40.
Wyss M, Smeitink J, Wevers RA, Wallimann T. Mitochondrial creatine kinase: a key enzyme of aerobic energy metabolism. Biochim Biophys Acta. 1992;1102:119–66.
Schlattner U, Mockli N, Speer O, Werner S, Wallimann T. Creatine kinase and creatine transporter in normal, wounded, and diseased skin. J Invest Dermatol. 2002;118:416–23.
Zemtsov A. Skin phosphocreatine. Skin Res Technol. 2007;13:115–8.
Snow RJ, Murphy RM. Creatine and the creatine transporter: a review. Mol Cell Biochem. 2001;224:169–81.
Speer O, Neukomm LJ, Murphy RM, et al. Creatine transporters: a reappraisal. Mol Cell Biochem. 2004;256–257:407–24.
McCully KK, Forciea MA, Hack LM, et al. Muscle metabolism in older subjects using 31P magnetic resonance spectroscopy. Can J Physiol Pharmacol. 1991;69:576–80.
Steinhagen-Thiessen E, Hilz H. The age-dependent decrease in creatine kinase and aldolase activities in human striated muscle is not caused by an accumulation of faulty proteins. Mech Ageing Dev. 1976;5:447–57.
Verzar F, Ermini M. Decrease of creatine-phosphate restitution of muscle in old age and the influence of glucose. Gerontologia. 1970;16:223–30.
Bogatskaia LN, Shegera VA. Creatine kinase activity and isoenzymic spectrum of myocardium creatine kinase in rats of different age. Ukr Biokhim Zh. 1981;53:71–4.
Ponticos M, Lu QL, Morgan JE, et al. Dual regulation of the AMP-activated protein kinase provides a novel mechanism for the control of creatine kinase in skeletal muscle. EMBO J. 1998;17:1688–99.
Stachowiak O, Schlattner U, Dolder M, Wallimann T. Oligomeric state and membrane binding behaviour of creatine kinase isoenzymes: implications for cellular function and mitochondrial structure. Mol Cell Biochem. 1998;184:141–51.
Chung JH, Eun HC. Angiogenesis in skin aging and photoaging. J Dermatol. 2007;34:593–600.
Ames BN, Shigenaga MK, Hagen TM. Oxidants, antioxidants, and the degenerative diseases of aging. Proc Natl Acad Sci U S A. 1993;90:7915–22.
Balaban RS, Nemoto S, Finkel T. Mitochondria, oxidants, and aging. Cell. 2005;120:483–95.
Shigenaga MK, Hagen TM, Ames BN. Oxidative damage and mitochondrial decay in aging. Proc Natl Acad Sci U S A. 1994;91:10771–8.
Wei YH, Lu CY, Wei CY, Ma YS, Lee HC. Oxidative stress in human aging and mitochondrial disease-consequences of defective mitochondrial respiration and impaired antioxidant enzyme system. Chin J Physiol. 2001;44:1–11.
Chance B, Sies H, Boveris A. Hydroperoxide metabolism in mammalian organs. Physiol Rev. 1979;59:527–605.
Cadenas E, Davies KJ. Mitochondrial free radical generation, oxidative stress, and aging. Free Radic Biol Med. 2000;29:222–30.
Droge W. Free radicals in the physiological control of cell function. Physiol Rev. 2002;82:47–95.
Stadtman ER. Protein oxidation and aging. Science. 1992;257:1220–4.
Huber LA, Xu QB, Jurgens G, et al. Correlation of lymphocyte lipid composition membrane microviscosity and mitogen response in the aged. Eur J Immunol. 1991;21:2761–5.
Nelson KK, Melendez JA. Mitochondrial redox control of matrix metalloproteinases. Free Radic Biol Med. 2004;37:768–84.
Harman D. Aging: a theory based on free radical and radiation chemistry. J Gerontol. 1956;11:298–300.
Sohal RS. Hydrogen peroxide production by mitochondria may be a biomarker of aging. Mech Ageing Dev. 1991;60:189–98.
Mori A, Utsumi K, Liu J, Hosokawa M. Oxidative damage in the senescence-accelerated mouse. Ann N Y Acad Sci. 1998;854:239–50.
Chiba Y, Yamashita Y, Ueno M, et al. Cultured murine dermal fibroblast-like cells from senescence-accelerated mice as in vitro models for higher oxidative stress due to mitochondrial alterations. J Gerontol A Biol Sci Med Sci. 2005;60:1087–98.
Trifunovic A, Wredenberg A, Falkenberg M, et al. Premature ageing in mice expressing defective mitochondrial DNA polymerase. Nature. 2004;429:417–23.
Hayakawa M, Torii K, Sugiyama S, Tanaka M, Ozawa T. Age-associated accumulation of 8-hydroxydeoxyguanosine in mitochondrial DNA of human diaphragm. Biochem Biophys Res Commun. 1991;179:1023–9.
Mecocci P, MacGarvey U, Kaufman AE, et al. Oxidative damage to mitochondrial DNA shows marked age-dependent increases in human brain. Ann Neurol. 1993;34:609–16.
Ames BN, Shigenaga MK, Gold LS. DNA lesions, inducible DNA repair, and cell division: three key factors in mutagenesis and carcinogenesis. Environ Health Perspect. 1993;5 Suppl 101:35–44.
Sohal RS, Dubey A. Mitochondrial oxidative damage, hydrogen peroxide release, and aging. Free Radic Biol Med. 1994;16:621–6.
Laganiere S, Yu BP. Modulation of membrane phospholipid fatty acid composition by age and food restriction. Gerontology. 1993;39:7–18.
Dumas M, Maftah A, Bonte F, et al. Flow cytometric analysis of human epidermal cell ageing using two fluorescent mitochondrial probes. C R Acad Sci III. 1995;318:191–7.
Paradies G, Ruggiero FM. Age-related changes in the activity of the pyruvate carrier and in the lipid composition in rat-heart mitochondria. Biochim Biophys Acta. 1990;1016:207–12.
Paradies G, Ruggiero FM. Effect of aging on the activity of the phosphate carrier and on the lipid composition in rat liver mitochondria. Arch Biochem Biophys. 1991;284:332–7.
Ruggiero FM, Cafagna F, Petruzzella V, Gadaleta MN, Quagliariello E. Lipid composition in synaptic and nonsynaptic mitochondria from rat brains and effect of aging. J Neurochem. 1992;59:487–91.
Orrenius S, Gogvadze V, Zhivotovsky B. Mitochondrial oxidative stress: implications for cell death. Annu Rev Pharmacol Toxicol. 2007;47:143–83.
Ha MK, Chung KY, Bang D, Park YK, Lee KH. Proteomic analysis of the proteins expressed by hydrogen peroxide treated cultured human dermal microvascular endothelial cells. Proteomics. 2005;5:1507–19.
Scharffetter-Kochanek K, Wlaschek M, Brenneisen P, et al. UV-induced reactive oxygen species in photocarcinogenesis and photoaging. Biol Chem. 1997;378:1247–57.
Suzuki YJ, Forman HJ, Sevanian A. Oxidants as stimulators of signal transduction. Free Radic Biol Med. 1997;22:269–85.
Davies KJ. The broad spectrum of responses to oxidants in proliferating cells: a new paradigm for oxidative stress. IUBMB Life. 1999;48:41–7.
Bladier C, Wolvetang EJ, Hutchinson P, de Haan JB, Kola I. Response of a primary human fibroblast cell line to H2O2: senescence-like growth arrest or apoptosis? Cell Growth Differ. 1997;8:589–98.
Chen Q, Ames BN. Senescence-like growth arrest induced by hydrogen peroxide in human diploid fibroblast F65 cells. Proc Natl Acad Sci U S A. 1994;91:4130–4.
Campisi J. The role of cellular senescence in skin aging. J Investig Dermatol Symp Proc. 1998;3:1–5.
Wallace DC. Mitochondrial genetics: a paradigm for aging and degenerative diseases? Science. 1992;256:628–32.
Pang CY, Lee HC, Yang JH, Wei YH. Human skin mitochondrial DNA deletions associated with light exposure. Arch Biochem Biophys. 1994;312:534–8.
Linnane AW, Marzuki S, Ozawa T, Tanaka M. Mitochondrial DNA mutations as an important contributor to ageing and degenerative diseases. Lancet. 1989;1:642–5.
Richter C. Oxidative damage to mitochondrial DNA and its relationship to ageing. Int J Biochem Cell Biol. 1995;27:647–53.
Piko L, Hougham AJ, Bulpitt KJ. Studies of sequence heterogeneity of mitochondrial DNA from rat and mouse tissues: evidence for an increased frequency of deletions/additions with aging. Mech Ageing Dev. 1988;43:279–93.
Cortopassi GA, Arnheim N. Detection of a specific mitochondrial DNA deletion in tissues of older humans. Nucleic Acids Res. 1990;18:6927–33.
Eshaghian A, Vleugels RA, Canter JA, et al. Mitochondrial DNA deletions serve as biomarkers of aging in the skin, but are typically absent in nonmelanoma skin cancers. J Invest Dermatol. 2006;126:336–44.
Porteous WK, James AM, Sheard PW, et al. Bioenergetic consequences of accumulating the common 4977-bp mitochondrial DNA deletion. Eur J Biochem. 1998;257:192–201.
Shoffner JM, Lott MT, Voljavec AS, et al. Spontaneous Kearns-Sayre/chronic external ophthalmoplegia plus syndrome associated with a mitochondrial DNA deletion: a slip-replication model and metabolic therapy. Proc Natl Acad Sci U S A. 1989;86:7952–6.
Schroeder P, Gremmel T, Berneburg M, Krutmann J. Partial depletion of mitochondrial DNA from human skin fibroblasts induces a gene expression profile reminiscent of photoaged skin. J Invest Dermatol. 2008;128:2297–303.
Berneburg M, Plettenberg H, Medve-Konig K, et al. Induction of the photoaging-associated mitochondrial common deletion in vivo in normal human skin. J Invest Dermatol. 2004;122:1277–83.
Yang JH, Lee HC, Wei YH. Photoageing-associated mitochondrial DNA length mutations in human skin. Arch Dermatol Res. 1995;287:641–8.
Birket MJ, Passos JF, von Zglinicki T, Birch-Machin MA. The relationship between the aging- and photo-dependent T414G mitochondrial DNA mutation with cellular senescence and reactive oxygen species production in cultured skin fibroblasts. J Invest Dermatol. 2009;129(6):1361–6.
Bandy B, Davison AJ. Mitochondrial mutations may increase oxidative stress: implications for carcinogenesis and aging? Free Radic Biol Med. 1990;8:523–39.
Wei YH, Lee CF, Lee HC, et al. Increases of mitochondrial mass and mitochondrial genome in association with enhanced oxidative stress in human cells harboring 4,977 BP-deleted mitochondrial DNA. Ann N Y Acad Sci. 2001;928:97–112.
Lu CY, Lee HC, Fahn HJ, Wei YH. Oxidative damage elicited by imbalance of free radical scavenging enzymes is associated with large-scale mtDNA deletions in aging human skin. Mutat Res. 1999;423:11–21.
Berneburg M, Grether-Beck S, Kurten V, et al. Singlet oxygen mediates the UVA-induced generation of the photoaging-associated mitochondrial common deletion. J Biol Chem. 1999;274:15345–9.
Kueper T, Grune T, Prahl S, et al. Vimentin is the specific target in skin glycation. Structural prerequisites, functional consequences, and role in skin aging. J Biol Chem. 2007;282:23427–36.
Hipkiss AR. Does chronic glycolysis accelerate aging? Could this explain how dietary restriction works? Ann N Y Acad Sci. 2006;1067:361–8.
Alikhani Z, Alikhani M, Boyd CM, et al. Advanced glycation end products enhance expression of pro-apoptotic genes and stimulate fibroblast apoptosis through cytoplasmic and mitochondrial pathways. J Biol Chem. 2005;280:12087–95.
Kasper M, Funk RH. Age-related changes in cells and tissues due to advanced glycation end products (AGEs). Arch Gerontol Geriatr. 2001;32:233–43.
Rugolo M, Lenaz G. Monitoring of the mitochondrial and plasma membrane potentials in human fibroblasts by tetraphenylphosphonium ion distribution. J Bioenerg Biomembr. 1987;19:705–18.
Scaduto Jr RC, Grotyohann LW. Measurement of mitochondrial membrane potential using fluorescent rhodamine derivatives. Biophys J. 1999;76:469–77.
Koopman WJ, Visch HJ, Smeitink JA, Willems PH. Simultaneous quantitative measurement and automated analysis of mitochondrial morphology, mass, potential, and motility in living human skin fibroblasts. Cytometry A. 2006;69:1–12.
Plasek J, Vojtiskova A, Houstek J. Flow-cytometric monitoring of mitochondrial depolarisation: from fluorescence intensities to millivolts. J Photochem Photobiol B. 2005;78:99–108.
Distelmaier F, Koopman WJ, Testa ER, et al. Life cell quantification of mitochondrial membrane potential at the single organelle level. Cytometry A. 2008;73:129–38.
Cossarizza A, Baccarani-Contri M, Kalashnikova G, Franceschi C. A new method for the cytofluorimetric analysis of mitochondrial membrane potential using the J-aggregate forming lipophilic cation 5,5′6,6′-tetrachloro-1,1′3,3′-tetraethylbenzimidazolcarbocyanine iodide (JC-1). Biochem Biophys Res Commun. 1993;197:40–5.
Hagens R, Khabiri F, Schreiner V, et al. Non-invasive monitoring of oxidative skin stress by ultraweak photon emission measurement. II: biological validation on ultraviolet A-stressed skin. Skin Res Technol. 2008;14:112–20.
Khabiri F, Hagens R, Smuda C, et al. Non-invasive monitoring of oxidative skin stress by ultraweak photon emission (UPE)-measurement. I: mechanisms of UPE of biological materials. Skin Res Technol. 2008;14:103–11.
Vandenberghe K, Goris M, Van Hecke P, et al. Long-term creatine intake is beneficial to muscle performance during resistance training. J Appl Physiol. 1997;83:2055–63.
Daly MM, Seifter S. Uptake of creatine by cultured cells. Arch Biochem Biophys. 1980;203:317–24.
Meyer LE, Machado LB, Santiago AP, et al. Mitochondrial creatine kinase activity prevents reactive oxygen species generation: antioxidant role of mitochondrial kinase-dependent ADP re-cycling activity. J Biol Chem. 2006;281:37361–71.
Lenz H, Schmidt M, Welge V, et al. Inhibition of cytosolic and mitochondrial creatine kinase by siRNA in HaCaT- and HeLaS3-cells affects cell viability and mitochondrial morphology. Mol Cell Biochem. 2007;306:153–62.
O’Gorman E, Beutner G, Dolder M, et al. The role of creatine kinase in inhibition of mitochondrial permeability transition. FEBS Lett. 1997;414:253–7.
Brdiczka D, Beutner G, Ruck A, Dolder M, Wallimann T. The molecular structure of mitochondrial contact sites. Their role in regulation of energy metabolism and permeability transition. Biofactors. 1998;8:235–42.
Dolder M, Walzel B, Speer O, Schlattner U, Wallimann T. Inhibition of the mitochondrial permeability transition by creatine kinase substrates. Requirement for microcompartmentation. J Biol Chem. 2003;278:17760–6.
Dolder M, Wendt S, Wallimann T. Mitochondrial creatine kinase in contact sites: interaction with porin and adenine nucleotide translocase, role in permeability transition and sensitivity to oxidative damage. Biol Signals Recept. 2001;10:93–111.
Berneburg M, Gremmel T, Kurten V, et al. Creatine supplementation normalizes mutagenesis of mitochondrial DNA as well as functional consequences. J Invest Dermatol. 2005;125:213–20.
Crane FL. Biochemical functions of coenzyme Q10. J Am Coll Nutr. 2001;20:591–8.
McLennan HR, Degli EM. The contribution of mitochondrial respiratory complexes to the production of reactive oxygen species. J Bioenerg Biomembr. 2000;32:153–62.
Lopez-Lluch G, Barroso MP, Martin SF, et al. Role of plasma membrane coenzyme Q on the regulation of apoptosis. Biofactors. 1999;9:171–7.
Mellors A, Tappel AL. The inhibition of mitochondrial peroxidation by ubiquinone and ubiquinol. J Biol Chem. 1966;241:4353–6.
Frei B, Kim MC, Ames BN. Ubiquinol-10 is an effective lipid-soluble antioxidant at physiological concentrations. Proc Natl Acad Sci U S A. 1990;87:4879–83.
Lass A, Kwong L, Sohal RS. Mitochondrial coenzyme Q content and aging. Biofactors. 1999;9:199–205.
Hoppe U, Bergemann J, Diembeck W, et al. Coenzyme Q10, a cutaneous antioxidant and energizer. Biofactors. 1999;9:371–8.
Podda M, Traber MG, Weber C, Yan LJ, Packer L. UV-irradiation depletes antioxidants and causes oxidative damage in a model of human skin. Free Radic Biol Med. 1998;24:55–65.
Kim DW, Hwang IK, Yoo KY, et al. Coenzyme Q_{10} effects on manganese superoxide dismutase and glutathione peroxidase in the hairless mouse skin induced by ultraviolet B irradiation. Biofactors. 2007;30:139–47.
Stab F, Wolber R, Blatt T, Keyhani R, Sauermann G. Topically applied antioxidants in skin protection. Methods Enzymol. 2000;319:465–78.
Hadshiew IM, Treder-Conrad C, v Bulow R, et al. Polymorphous light eruption (PLE) and a new potent antioxidant and UVA-protective formulation as prophylaxis. Photodermatol Photoimmunol Photomed. 2004;20:200–4.
Rippke F, Wendt G, Bohnsack K, et al. Results of photoprovocation and field studies on the efficacy of a novel topically applied antioxidant in polymorphous light eruption. J Dermatolog Treat. 2001;12:3–8.
Wolber R, Stab F, Max H, et al. Alpha-glucosylrutin, a highly effective flavonoid for protection against oxidative stress. J Dtsch Dermatol Ges. 2004;2:580–7.
Author information
Authors and Affiliations
Corresponding author
Editor information
Editors and Affiliations
Rights and permissions
Copyright information
© 2015 Springer-Verlag Berlin Heidelberg
About this entry
Cite this entry
Blatt, T., Wenck, H., Wittern, KP. (2015). Alterations of Energy Metabolism in Cutaneous Aging. In: Farage, M., Miller, K., Maibach, H. (eds) Textbook of Aging Skin. Springer, Berlin, Heidelberg. https://doi.org/10.1007/978-3-642-27814-3_29-2
Download citation
DOI: https://doi.org/10.1007/978-3-642-27814-3_29-2
Received:
Accepted:
Published:
Publisher Name: Springer, Berlin, Heidelberg
Online ISBN: 978-3-642-27814-3
eBook Packages: Springer Reference MedicineReference Module Medicine