Skip to main content
SpringerLink
Log in

High density of beta2-adrenoceptors in a human keratinocyte cell line with complete epidermal differentiation capacity (HaCaT)

  • Original Contributions
  • Published:
Archives of Dermatological Research Aims and scope Submit manuscript

Summary

A non-tumorigenic keratinocyte cell line with complete epidermal differentiation capacity (HaCaT) was used in radioligand binding experiments to determine the number of beta-adrenoceptors. Intact cells were saturated with 3H-labelled (−)CGP-12177 (CGP), a hydrophilic non-selective beta-adrenergic antagonist as radioligand. In order to investigate the beta-adrenergic subtype selectivity, displacement experiments were performed with different antagonists and agonists. Binding of CGP to keratinocytes has been shown to be reversible and saturable and to have high affintiy (B max=114.0±8.8 fmol/107 cells with 6866 receptors/cell, K D=0.095±0.017 nmol/l; n=11). Betaadrenergic antagonists inhibited binding yielding monophasic displacement curves. IC50-values (nmol/l) were: propranolol (non-selective) 1.68; CGP-12177 (non-selective) 1.08; ICI 118,551 (beta2-selective) 2.92; bisoprolol (beta1-selective) 1230; and CGP-20712 (beta1-selective) 24980. Agonists displaced CGP in the order isoprenaline> adrenaline>noradrenaline. We conclude that HaCaT cells express a high density of beta2-adrenoceptors providing a good model system to study adrenergic receptor mechanisms under reproducible experimental conditions in keratinocytes.

This is a preview of subscription content, log in via an institution to check access.

Access this article

Log in via an institution

Price excludes VAT (USA)
Tax calculation will be finalised during checkout.

Instant access to the full article PDF.

Institutional subscriptions

Similar content being viewed by others

References

  1. Affolter H, Hertel C, Jaeggi K, Portenier M, Staehelin M (1985) (−)-S[3H]CGP-12177 and its use to determine the rate constants of unlabeled Β-adrenergic antagonists. Proc Natl Acad Sci USA 82:925–929

    Google Scholar 

  2. Bilski A, Dorries S, Fitzgerald JD, Jessup R, Tucker H, Wale J (1980) ICI 118,551, a potent Β-adrenoceptor antagonist (abstract). Br J Pharmacol 69:292P-293P

    Google Scholar 

  3. Boukamp P, Petrussevska RT, Breitkreuz D, Hornung J, Markham A, Fusenig NE (1988) Normal keratinization in a spontaneously immortalized aneuploid human keratinocyte cell line. J Cell Biol 106:761–771

    Google Scholar 

  4. Boyce ST, Ham RG (1983) Calcium-regulated differentiation of normal human epidermal keratinocytes in chemically defined clonal culture and serum-free serial culture. J Invest Dermatol 81 [Suppl 1]:335–405

    Google Scholar 

  5. Brodde OE (1986) Bisoprolol (EMD 33512), a highly selective Β 1-adrenoceptor antagonist: in vitro and in vivo studies. J Cardiovasc Pharmacol 8 [Suppl 11]:S29-S35

    Google Scholar 

  6. Brodde OE, Kretsch R, Ikezono K, Zerkowski HR, Reidemeister JC (1986) Human Β-adrenoceptors: relation of myocardial and lymphocyte Β-adrenoceptor density. Science 231:1584–1585

    Google Scholar 

  7. Brodde OE, Beckeringh JJ, Michel MC (1987) Human heart Β-adrenoceptors: A fair comparison with lymphocyte Β-adrenoceptors? Trends Pharmacol Sci 8:403–407

    Google Scholar 

  8. Cavey MT, Cavey D, Shroot B, Reichert U, Gazith J (1986) Receptor linked adenylate cyclase in the membranes of cultured human epidermal keratinocytes. Arch Dermatol Res 278:293–297

    Google Scholar 

  9. Dooley DJ, Bittiger H, Reymann NC (1986) CGP 20712 A: a useful tool for quantitating Β 1- and Β 2-adrenoceptors. Eur J Pharmacol 130:137–139

    Google Scholar 

  10. Duell EA, Voorhees JJ, Kelsey WH, Hayes E (1971) Iso-proterenol-sensitive adenyl cyclase in a particulate fraction of epidermis. Arch Dermatol 104:601–610

    Google Scholar 

  11. Dulis BH, Wilson IB (1980) The beta-adrenergic receptor of live polymorphonuclear leucocytes. J Biol Chem 255:1043–1048

    Google Scholar 

  12. Durham AC, Walton JM (1982) Calcium ions and the control of proliferation in normal and cancer cells. Biosci Rep 2:15–30

    Google Scholar 

  13. Eedy DJ, Canavan JP, Shaw C, Trimble ER (1990) Beta-adrenergic stimulation of cyclic AMP is defective in cultured dermal fibroblasts of psoriatic subjects. Br J Dermatol 122:477–483

    Google Scholar 

  14. Fraser CM, Venter JC (1980) The synthesis of Β-adrenergic receptors in cultured human lung cells: induction by glucocorticoids. Biochem Biophys Res Commun 94:390–397

    Google Scholar 

  15. Gazith J, Reichert U (1982) High affinity membrane receptors in cultured human keratinocytes. I. The Β-adrenergic receptors. Br J Dermatol 107 [Suppl 23]:125–133

    Google Scholar 

  16. Gazith J, Reichert U (1987) Adrenergic receptors in the skin. In: Fitzpatrick TB, Eisen AZ, Wolff K, Freedberg IM, Austen KF (eds) Dermatology in general medicine, vol I. McGraw-Hill, New York, pp 375–380

    Google Scholar 

  17. Gazith J, Cavey MT, Cavey D, Braham S, Reichert U (1983) Characterization of the beta-adrenergic receptors of cultured human epidermal keratinocytes. Biochem Pharmacol 32:3397–3403

    Google Scholar 

  18. Haeusler G, Schliep HJ, Schelling P, Becker KH, Klockow M, Minck KO, Enenkel HJ, Schulze E, Bergmann R, Schmittges CJ, Seyfried C, Harting J (1986) High Β 1-selectivity and favourable pharmacokinetics as the outstanding properties of bisoprolol. J Cardiovasc Pharmacol 8 [Suppl 11]:S2-S15

    Google Scholar 

  19. Heinsimer JA, Lefkowitz RJ (1982) Adrenergic receptors: Biochemistry, regulation, molecular mechanism, and clinical implications. J Lab Clin Med 100:641–658

    Google Scholar 

  20. Honegger UE, Disler B, Wiesmann UN (1986) Chronic exposure of human cells in culture to the tricyclic antidepressant desipramine reduces the number of beta-adrenoceptors. Biochem Pharmacol 35:1899–1902

    Google Scholar 

  21. Iizuka H, Adachi K, Halprin KM, Levine V (1978) Cyclic AMP accumulation in psoriatic skin: differential responses to histamine, AMP, and epinephrine by the uninvolved and involved epidermis. J Invest Dermatol 70:250–253

    Google Scholar 

  22. Kajita S, Iizuka H, Hirokawa M, Tsutsui M, Mizumoto T (1986) Topical application of potent glucocorticoids augments epidermal beta-adrenergic adenylate cyclase response in vivo. Acta Derm Venereol (Stockh) 66:491–496

    Google Scholar 

  23. Kaumann AJ, Birnbaumer L, Wittmann R (1978) Heart Β-adrenoceptors. In: O'Malley BW, Birnbaumer L (eds) Receptors and hormone action, vol. III. Academic Press, London New York San Francisco, pp 133–177

    Google Scholar 

  24. Koch-Weser J (1970) Effects of beta-adrenergic stimulation and blockade on myocardial mechanics. In: Kattus AA, Ross G, Hall VE (eds) Cardiovascular beta-adrenergic responses. University of California Press, Berkeley Los Angeles London, pp 45–67

    Google Scholar 

  25. Koizumi H, Yasui C, Fukaya T, Ohkawara A, Ueda T (1991) Beta-adrenergic stimulation induces intracellular Ca++ increase in human epidermal keratinocytes. J Invest Dermatol 96:234–237

    Google Scholar 

  26. Laduron PM (1984) Criteria for receptor sites in binding studies. Biochem Pharmacol 33:833–839

    Google Scholar 

  27. Lands AM, Arnold A, McAuliff JP, Luduena FP, Brown TG (1967) Differentiation of receptor systems activated by sympathomimetic amines. Nature 214:597–598

    Google Scholar 

  28. Lands AM, Luduena FP, Buzzo HJ (1967) Differentiation of receptors responsive to isoproterenol. Life Sci 6:2241–2249

    Google Scholar 

  29. Lefkowitz RJ, Caron MG, Stiles GL (1984) Mechanisms of membrane-receptor regulation. Biochemical, physiological, and clinical insights derived from studies of the adrenergic receptors. N Engl J Med 310:1570–1579

    Google Scholar 

  30. Ludbrook PA, Sobel BE (1975) Catecholamines, cyclic nucleotides, and cardiac function. In: Vasalle M (ed) Cardiac physiology for the clinician. Academic Press, New York London San Francisco, pp 117–140

    Google Scholar 

  31. Molinoff PB, Wolfe BB, Weiland GA (1981) Quantitative analysis of drug receptor interactions: II. Determination of the properties of receptor subtypes. Life Sci 29:427–443

    Google Scholar 

  32. Motulsky HJ, Insel PA (1982) Adrenergic receptors in man. Direct identification, physiologic regulation, and clinical alterations. N Engl J Med 29:18–29

    Google Scholar 

  33. Mui MM, Hsia SL, Halprin KM (1975) Further studies on adenyl cyclase in psoriasis. Br J Dermatol 92:255–262

    Google Scholar 

  34. Ohkawara A, Iizuka H (1985) Glucocorticoid-induced alteration of beta-adrenergic adenylate cyclase response of epidermis. Arch Dermatol Res 277:88–92

    Google Scholar 

  35. Orenberg EK, Pfendt EA, Wilkinson DI (1983) Characterization of α- and Β-adrenergic agonist stimulation of adenylate cyclase activity in human epidermal keratinocytes in vitro. J Invest Dermatol 80:503–507

    Google Scholar 

  36. Powell JA, Duell EA, Voorhees JJ (1971) Beta-adrenergic stimulation of endogenous epidermal cyclic AMP formation. Arch Dermatol 104:359–365

    Google Scholar 

  37. Staehelin M, Simons P, Jaeggi K, Wigger N (1983) CGP-12177. A hydrophilic Β-adrenergic receptor radioligand reveals high affinity binding of agonists to intact cells. J Biol Chem 258:3496–3502

    Google Scholar 

  38. Wang XL, Brinkmann M, Brodde OE (1985) Selective labelling of Β 1-adrenoceptors in rabbit lung membranes by (−)[3H]-bisoprolol. Eur J Pharmacol 114:157–165

    Google Scholar 

  39. Watt FM, Mattey DL, Garrod DR (1984) Calcium-induced reorganization of desmosomal components in cultured keratinocytes. J Cell Biol 99:2211–2215

    Google Scholar 

  40. Weiland GA, Molinoff PB (1981) Quantitative analysis of drugreceptor interactions: I. Determination of kinetic and equilibrium properties. Life Sci 29:313–330

    Google Scholar 

  41. Winek R, Bhalla R (1979) [3H]-dihyroalprenolol binding sites in rat myocardium: relationship between a single binding site population and the concentration of the radioligand. Biochem Biophys Res Commun 91:200–206

    Google Scholar 

  42. Yoshikawa K, Adachi K, Halprin KM, Levine V (1975) On the lack of response to catecholamine stimulation by the adenyl cyclase system in psoriatic lesions. Br J Dermatol 92:619–624

    Google Scholar 

Download references

Author information

Authors and Affiliations

  1. Department of Dermatology, University of Hamburg, UniversitÄts-Krankenhaus Eppendorf, Martinistrasse 52, W-2000, Hamburg 20, Federal Republic of Germany

    V. Steinkraus, C. Körner & H. Mensing

  2. Department of Pharmacology, University of Hamburg, UniversitÄts-Krankenhaus Eppendorf, Martinistrasse 52, W-2000, Hamburg 20, Federal Republic of Germany

    M. Steinfath

Authors
  1. V. Steinkraus
    View author publications

    You can also search for this author in PubMed Google Scholar

  2. C. Körner
    View author publications

    You can also search for this author in PubMed Google Scholar

  3. M. Steinfath
    View author publications

    You can also search for this author in PubMed Google Scholar

  4. H. Mensing
    View author publications

    You can also search for this author in PubMed Google Scholar

Rights and permissions

Reprints and permissions

About this article

Cite this article

Steinkraus, V., Körner, C., Steinfath, M. et al. High density of beta2-adrenoceptors in a human keratinocyte cell line with complete epidermal differentiation capacity (HaCaT). Arch Dermatol Res 283, 328–332 (1991). https://doi.org/10.1007/BF00376622

Download citation

  • Received:

  • Issue Date:

  • DOI: https://doi.org/10.1007/BF00376622

Key words

Search

Navigation