Catecholamines in Human Keratinocyte Differentiation

Karin U. Schallreuter, K Regina Lemke, Mark R. Pittelkow, John M. Wood, Clinsta Körner, Ruth Malik
1995 Journal of Investigative Dermatology  
and i"Depart1l1ent of Biomedical Sc iences. Univ ersiry of Bradfo rd. Un.ited Kingdom Human keratinocytes have the capacity to synthesize catecholamines from L-tyrosine, which in turn is produced from L-phenylalanine Ilia phenylalanine hydroxylase. This enzyme activity is controlled by the supply of the essential cofactor/electron donor (6R)5,6, 7 ,8 tetrahydrobiopterin (6-BH4)' Undifferentiated keratinocytes express high levels of the rlttelimiting enzymes for the de 110110 synthesis of 6-BH4'
more » ... i.e., GTP-cyclohydrolase-l, and for its recycling, Le., 4a-hydroxytetrahydrobiopterin dehydratase. As a consequence of 6-BH. synthesis, phenylalanine hydroxylase is activated, yielding L-tyrosine, which in the presence of excess 6-BH 4 turns on the biosynthesis of catecholamines Ilia the rate-limiting enzynJe tyrosine hydroxylase. Therefore, undifferentiated keratinocytes contain high levels of the catecholamine R ecently it h as bee n shown tha t the hum an epide rmis h as the cap acity for total ca techolamine biosynthesis (Fig 1) . Tyrosine h ydroxylase (T H) , th e key e n zym e, and phenylethanolamin e-N-meth y l tran sferase (PNMT), the e n zym e for con ve rsio n o f norepine phrine to epine phrine, h ave b een identified in keratinocytes in epide rm al suction blister roofs and in full-thickn ess skin [1]. Monoclonal antibo di es aga.inst T H and PNMT co nfimled tb e locatio n of b oth en zym es in keratinocytes (A.J. Thody. p e rson a l communicatio n) . T h e biosynth esis of th e catecholamin es d ep e nds o n the availa bility of L-tyrosin e synth es ized fr o m the essenti al amino acid Lphen ylalan ine. This reaction is controlled by pheny lalanine h ydl:"o ;'{y lase (PAH) and its essenti al cofacto r/e lectron donor (6R)5, 6, 7 ,8 tetrahydro biopterin (6-BH.,). 6-BH 4 is pro du ced de 11 0110 from GYP Il in the rate-limiting en zym e GTP-cyclohydrolase 1 (GTP-CH-l ) and recycled IIin the activities of 4a-h ydroxytetrahydro biop terU1 d e hydra tase (DH) together with dih ydroptel;dine re du ctase. DH is th e rate-1.imiting en zym e for the recyding of 6-BH 4 [2-4] (Fi~ 2) . On ly rece ntl y, it has b een reported that hum an undifFe re ntiated keratinocytes and ceIl extracts fro m epidermal su ction blisters express con stituti ve levels of PAH with sp ecifi c activities con.,p a-Manu script receivcd February 3, 1994; final revision received Decclbbc r 29, '1994; accepted for publi cation Janu ary 18, 1995. Reprint req uests to: Dr. K.U. SchaUreuter, Dcpa rtment of DerInatology, Universiry of Hamburg. Martinistra sse 52. 20246 Hamburg. Germany. Abbrev iations : 6-BH.I' (6R.)5,6,7,8 tetrahydrobiopteri n; DH, 4a-hYdroxytetrahydrobioptcrin dehydratase; GTP-CH-:I, GTP cyclohydrolase_ 1; PAH, phenylalanine hydroxylase; PNMT, phenylethanolamine-N-metbyl transfcrase; T H, tyrosin c hydro"-ylase. system yielding sufficient levels of norepinephrine and epinephrine, required for the induction of beta-2-adrenoceptors. Stimulation of beta-2-adrenoceptors by epinephrine causes a rise in intracellular calcium Ilia extracellular influx. This event corresponds with keratinocyte differentiation. In differentiated keratinocytes, all enzyme activities involved in 6-BH4' L-tyrosine, and epinephrine biosynthesis are decreased, resulting in significantly lower levels of epinephrine and a concomitant decrease in the expression of beta-2-adrenoceptors. These data strongly suggest a connection between catecholamine biosynthesis, beta-2-adrenoceptor expression, calcium flux, and the differentiation ofkeratinocytes in human epidermis. K ey IIIol·d: beta-2-advelloceptovs. ",est Del'11latol :1,04:953-957, 1995 [able to those of PAH found in liver and n e uro nal tissues, w h e reas difFe re ntiate d keratinocytes h ave barely d etectable levels [3, 4] . In addition, it h as b een shown that keratinocytes and epid e m 131 cell e xtracts h ave the cap acity for de 11 0110 synth esis and recycling of 6-BH4' th e rate-limiting cofactor/electron donor for PAH and TH [3, 4] (Fig 2) . T h e biosyn tll es is of epinephrine in th e e pidermis lead s to ill villo express ion of a h.i gh d en sity of b eta-2-adren oceptors in keratinocytes, as d etermine d by radio li gand binding and au toradiograph.ic techniques [5-7] . Upon adre n ergic stim ulati o n witl1 epinephrine, a signifi can t increase in both intracellular cal cium and cAMP occurs [7-9J. Cyclic A MP h as b een shown to induce the TH gen e, leading to an in crease in TH mRNA . It h as b een su ggested that the b eta-2adrenoceptor sys tem in ke ratinocytes primarily controls calcium hom eostasis during difFer entiation . The purpose of this study was to exa mine the relation between th e biosynth esis of e pinep hrin e togetller wi th the expression of b eta-2-adre n oceptors and the calcium flu x during th e difFerentiation of k e ratino cytes und er ill vitro conditi o n s [8 -1 2]. M.ATERIALS AND METH ODS 6-BH'I' 6,7 dimcthyl-BH •. and 6-biopterin were obtained from Schircks Oona, Switzerland). All othcr enzym es and reagents were purchased from Sigma (St. Louis, MO). I·C UL L-phenylaJanin c (51. 3 m Ci/ mm( 1) was fro m IC N isotopes (CA) . and (1.06 mC i/ mmol) was from Amersham Radiochemicals (England). (-)-["'H) CG P 12177 (55 mCi/mmol) was purchascd from Amersham Buchl er (Braunschweig, Germany). 45Ca.lcium was fro m New England Nuclear (Dupont, Boston, MA) (200 mCi / ml) . Dowex AG50 (WX8) was from Biorad. Monoclonal mouse anti-human cytokeratin antibody (CK 10) was obtained fro m DAKO A / S (Denmark). Monoclonal mouse anti-human cyto keratin 5/ 6 antibody was fro m Boehringer (Mannheim, Germany). 0022-202X/95/S09.50 • SSD I0022-202X (95)00169-L •
doi:10.1111/1523-1747.ep12606218 pmid:7769265 fatcat:7jd5qtxwbrcbxenf23ri2pikqi