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Finasteride is the main inhibitor of 5α-reductase activity in dermal papillae of human hair follicles
ESDR I JSID I SID Abstracts
0337
0340
THYMIDINE DINUCLEOTIDE WCREASES THE LEVEL OF DNA REPAIR PROTEINS IN HUMAN CELLS. D. Goukassiuu, M. Eller, M.Yaar. and BA Gilchrest. Department of Dermatology, Boston University School of Medicine, Boston, MA. DNA damaging ultraviolet (W) irradiation upregulates cellular DNA repair mechanisms, at least m purl through actwation of ~53 aud p53-regulated proteins. Because thvmidrne dinucleotide (vTpTT),when added 1u cultured skin cells, mimics some UV respon?esmcluding enhancer&t of DNA repair capacity and activation of ,153, we compared Ihe effect of LN and pTpT un 953 and its regulated proleins ~21, proliferating cell nuclear antigen (PCNA) and MDM2. Newborn tibroblasts were exposed tu 5 or 30 mJ/cm’ UVB, or we= sham irradiated. Duplicate cultures were provided 100 uM pTpT or diluenf. RNA and proteins were collected up tu 5 days after treafment, analyzed by northern and western blotting and band intensity was determrned by deusitometric analysis. Both UV and pTpT increased ~21 mRNA within 8 hrs, but p53 mRNA was unchangedthrough 48 hrs. Compared to sham mndiated cultures, within 3 hours at 5 mJ/cu?. ~53 and p2, pm&s were induced 400% and 200% and remained elevated through 7 hours. At 30 mJ/crn’ ~53 and p2, were elevated 100% aud 1100% and remained elevated through 48 hrs, the Iat timepoint examined. PCNA pro&ins and MDMZ undetectablein sham or 5 ml/cm’ irradiated cultures. were strongly expressed 48 hours after 30 ml/cm’. Interestingly. pTpT also strongly induced ~53. pZ1 and PCNA (6CQ%,450% and 400%. respectively) above dilueut treated controls. pTpT induction was observed us early as 72 hours and was max,mal 5 days after stimulation, at (he termination of the experimem To determine if pTpT can enhance the cellular response tu WV irradiation, cultures were pretreated with pTpT or dduenf fur 5 days and then were UV or sham irradiated. At 5 and 30 mIlc/cm’UV irradiation, in cultures pre-treated wth pTpT as compared tu dilueut treated cultures, FCNA level was 500% and 200% higher. respectively and MDMZ level was 200% higher. ~21 was also slightly induced. Our studiessuggestfhat pTpT enhances cellular DNA repair capacity through upregulation
DIFFERENT FREQUENCY OF GENE !tA-DNA OLIGONiJCLEO~~DE,&K)a~G tam. S bralv .AdamE ntz. nn
of key protiens, in the absenceof actual DNA damage.
s57
TARGETING EVENTS BY THE EPITHELIAL CELLS. m
ntoandKvon~~eunYoon, Department of Dermat&gy ~~Cutaneons Biology, Jefferson Medical College, Phila, PA 19107 A unique hybrid oligonudeotide composed of both RNA and DNA wus shown to correct a point mutation inn site-specific and inheritable manner in extmchmmasomal and chrumcsomal targets. (Yam et al. Fmc. NatI. Acad. Sci._?&, 2071. 1996, ColeStrauss et al. Sciences 13&5,1966). The RNA stretches of the oligonwleotide was hypothesized IO fadlitate the homologous pecombinatim and a mismatch created tehveen the targeted strand and RNA-DNA digonudeotide may then be.r@nd by the DNA repair system. In order to investigate whether the fnquency d tbe gene targeting is dependent on cell types, we tested two digonncleotides which W&E shown to create a point mntatica in alkaline phosphatax and bglobin gene respectively, in several epithelinl cells. Highly ttansformed epithdial cells (&La) exhibited a much higher frequency in gene conversion by b&b RNA-DNA digonucleotides in mmparison to other innncalized epitbelial cells (HaCaT) or primary keratinocytes. However. the innacellular unrentradon of the digonwleotide in the nnclens of H&a cells was lower than that of HaCaT or primary kaatinocytes. measured by a radic&beled and a iluoresrein-ccmjugated digonnclwtide. Oligcmwleotide extracted from cells was shown to be stable for within the time period III which gene conversion event was measured. Thus, an uptake and inhacellular stability of the digonncleotide are not limiting facbxs in gene targeting events under our experimental conditions. These results suggest that endogenous cellular recombination and DNA repair activities may vary behveen different cdl types and are important for the gene conversion event mediated by the RNA-DNA digrmudwtide.
0338
0341
EKFRESSIONOF VBGF IN CHRONICNON-HElALINGWOUNDS.s
EXPRESSION OF A PUTATIVE PROOPIOMELANOCORTlNmRNA VARIANT IN .a.1 HUMAN EPIDERMIS AND EPIDERMAL CELLS IN CULTURE. m Bshiib Bud&. StevenBovcd. 4 James Nordlund’.Deparnnent of Dermatology’, University of Cincinnati College of Medicine 4 Shrlners Bums Institu&, Cincinnati, OH, USA
*Department of Dermatolo8y. Urdverslty of K(lln. Kdln, Oermany; ‘*Department of Molecular Cell Biology, Max Planck InaUtute.Bad Nauheim,Gummy. The patbngenesls of chronic wounds Is largely unknown. An hypothesis for the mecba&ms undexlyln8 wxmd frdlure suggestr that the buxeaed protedytlc actwty in chronic wounda leads to the de8wwion Of 8rcwth factma ogzssaryf‘xmccesstwrepllr. To tat tbls hypothesis we evaluated the expression of vwxlar en&tbellal groMh factur (VEQF) in chronic mm-be&ng leg u1cea-sand anllyzed the mechanism of VBGF ~~th&r&r& pmteolysls In chmnlc wounds. VBGF 1s a potem an8iogerd&medIatcedIatc play an lmpxtant rolS ln tissue nzpair. Immunobl~ of VEGF expreaslon ln the hyperplastIc epldmnls and damal mononuclear cells of cbronlc wounds. when compared to uninjured control skin. The epidermat sl8nat lo chrnrdc arounds was s&rdEcantly weaka us compared tu epldermal VBGF stalnlng in psoriatic kslons. arhlch saved as. positive WntmI. IntawIngty. the in sin4 hyb&l7.mlon of cbronlc wumls showed a feIaUve stmn8 VEGF mRNA expression in the epldumis and ckmnal mcmonuclearcells, comparable to that of pwrlatJc lesions. Explanatlotts for the dlszre+cy bdueen the strnog mRNA expression and relative low protein sl@al In the e@dennls of cbmnlc woumis suggest eltber a lransltionnl d&a or that VEOF proWn Is unstable in the cbrnnlc wound environment. ‘Ilxrefore. reamblnant VBGF165 was incubated wltb cbronk wound flnld Tbls resulted In the degradation of rVBGl+.. whereas lncnbe.tlonin acute wound fluid did not. Pmteaselnblbltnr &wiles suggested that serineprotepaes are at least *ally responsible for rVBGFl65 degradation in cbrouic wound fluid. Our data demorx?&atefor the first time that VBGF mRNA expression is upregulated In cllronlc wounds and that chronic wound fluid can &grade VEGF. Therefore.our data support the hypomenisthat ti lxreased paeOlytIc a&‘@ in cltmnlc wounds leads to the degradntlon of growth factors and &ruby may impsir their cnntibutlcm to wnund b?aIlng. Funher studlea are needed to a&ate r&zslgnlflcance of this observation for other cyWkloes bnportattt in tissue re@r and our data propose 0 develop strategies hew to mntrd pmtxolytlc acavity in chrnnlc wounds.
FWmpiomeIauocnrtin(PGMC) is a -30,ooO d&n prmein precursor for various bioactive peptidessuch as ACTH, a, p, 7, MSH’s, l&mdorphin and l3-lipotropin. Butb ACTH and a-
MSH have recently been implicatediu cutaueoushyperpigmentation. Previous sludies from our labaraturyhave shownthe presenceof POMC mRNA in uorrnalhuman skin and cultures of epidermal melanocytesand keratiwcytea by in-situ hybridization. In addition, srmog signals of immuuoreactiveACTH and a-MSH were detectedin normal human skin and the epidermal cells. In tie present study, we utilized a combination of immunobistochemistq and in stiu hybridiiion to identify the cell type expressiug POMC mRNA in human epidermis. A melanocyte specific antibody (m&J) and a keratirocyte specific antibody (anticymkeratin)were employed to identify melanocytesand keratirwcytesrespectively in human skin. By immunochemicalstaining, the melanocytes and keratinocytesthat were positively stained with their respective antibodies also exhibited strung signals for POMC mRNA by in-situ hybridization. In order tu identify POMC mRNA iu human skin, reverse transcription-pdytnerasechain reaction (RT-PCR) was performed. An -300 hp POMC product was detected in normal human skin, cultured melanocyteSand keratinocytes. By Southern blot analysis this product was hybridized specifically m the PGMC cDNA. RTPCR productsobtainedfrom normal skin and culturesof melanocytesand keratiwcytes, gave au identical sequence. Comparison of RT-PCR sequenceby Genebauk/Blastrevealed 83% homology to POMC cDNA from human, bovine, pig and monkey sources. This data suggests the presenceof a putative iwform or variant of POMC mRNA in human skin. The detection of POMC mRNA variant in skin may aid in understandingthe mechanism of cutaneoushyperpigmaatiun.
0339
0342
HUMAN PAPILLOMAVIRUS E6 PROTEIN FROM BOTH CERVICAL AND C”TANEOUS TYPE.9INHIBIT p53-DEPENDENT AND p53 -INDBPENDENTAPOPTOSIS S.Ja&ou. I.M.Leil andASturq Centrefur Cutmeow Research. St.Bartholomew’s andthe Royal Schoolof MedicineandDe&try, 2. Newark SW&. London. The two maw transfurmiug pr&ns of oncogemc HumanPapdlomavimses (HPVs) are encodedby the E6 and W ancogcnes and their transfomung abdity can be explainedby their asracianonwiti tumuursuppressor proteins.The E6 protan formscomplexeswuh p53 causinginactivationof p53 fuuctmnincludingcell cyclecheckpoint cuutmlandapoptasis andtheFI prutembindstic Rb protein. The aim of the studywas to invesrigate whetherIhe E-5proteinfrom lug+ and low risk HPVs could mhlbit apaptosisand to detenuineif this E6 Rnctlonwas mediatedby inachvatwnof ~53. Apoprosrs was inducedusmg ultravioletradia6on (WR) and measuredusing terruinalrmusferax ruck end labelbugWNEL) andcolonyassaysfallowingtransfcc6on of HPV genesintoa vanelyof cell types. Our resultsfmm TUNEL stainingof HT1080 cells.wixchcontainwild-type~53, showedthatfbe ~6 prutemfrom bothhigh andlow risk typesinhibitedp53-dependem apoptosis following DNA damage. To investiaate if theE6 urutemscouldalsoiohibito53-iudcoendem aou~tusrs. 10.1lo53 null) cel,l WHIP
FINASTERIDE IS THE MAIN INHIBITOR OF 5u-REDUCTASE ACTIVITY IN DERMAL PAPILLAE OF HL?fAN HAIR FOLLICLES. Rolf Ho@ne*, Wolf ang ,I ?f Derrnatdop Ph$pps U~ver@t, Ma&g ~v~o~$~~p& rhooncology, echmsche Unwers~t& Dresden, T&e Sa-reductase isoeazyma type 1 and 2 and 1713-hydroxysteroid dehydrogenase (170-HSD) are man enzymes in the intracellular androgen metabolism, but at present the knowledge about the ph siolo ‘c role of 5a-reductase isoenzymea is onl incompletely understood Several antK!’ nrs we tned to localii both &enzymes wit&n the human hair follicle, but several m or immnnohistochanical studies gave
were pooled, incubated with SOnMtestosterone for 18 hours and Sa-reduced steroids were analysed by thin-layer chromate phy. Our results show that the volume activity of 5a-reductase in the DP exceeded $. ose tn KS or CTS at least by a factor I4 in the scalp and a &or 80 in the beard. The RS expressed low Sa-reductase, but high 17l3HSD levels, with nndrostenedione as major rnetabdite. The CTS expressed both Sareductase and 17B-HSD, resultin in androstenedione, Sa-androsterone, and 5aandrostanedione. To our surprise f nastende (I&,: 10nrnolil) but not MK-386 (max. cone: Ipmolil) inhibited approx. 80% of 5u-reductase activity in DP and CTS, su esting the pr.+n~ .of type 2 Sa-rednctase, whereas 5a-rednctase acti& within the% S was mamly mbtbrted by MK-386 (IC,: 2Cnmovl) and not by fmasten,Be (I&,. 2COnmoI!l), suggesting the presence of type 1 5a-reductase. Coincubations with fmasteride and hlK-386 resulted in 95-100% inhibition of 5a-reductase activity in DP (It&: approx. Smnov1).Therefore we conclude that type 2 is the main Sa-reductase present m DP.
0337
0340
THYMIDINE DINUCLEOTIDE WCREASES THE LEVEL OF DNA REPAIR PROTEINS IN HUMAN CELLS. D. Goukassiuu, M. Eller, M.Yaar. and BA Gilchrest. Department of Dermatology, Boston University School of Medicine, Boston, MA. DNA damaging ultraviolet (W) irradiation upregulates cellular DNA repair mechanisms, at least m purl through actwation of ~53 aud p53-regulated proteins. Because thvmidrne dinucleotide (vTpTT),when added 1u cultured skin cells, mimics some UV respon?esmcluding enhancer&t of DNA repair capacity and activation of ,153, we compared Ihe effect of LN and pTpT un 953 and its regulated proleins ~21, proliferating cell nuclear antigen (PCNA) and MDM2. Newborn tibroblasts were exposed tu 5 or 30 mJ/cm’ UVB, or we= sham irradiated. Duplicate cultures were provided 100 uM pTpT or diluenf. RNA and proteins were collected up tu 5 days after treafment, analyzed by northern and western blotting and band intensity was determrned by deusitometric analysis. Both UV and pTpT increased ~21 mRNA within 8 hrs, but p53 mRNA was unchangedthrough 48 hrs. Compared to sham mndiated cultures, within 3 hours at 5 mJ/cu?. ~53 and p2, pm&s were induced 400% and 200% and remained elevated through 7 hours. At 30 mJ/crn’ ~53 and p2, were elevated 100% aud 1100% and remained elevated through 48 hrs, the Iat timepoint examined. PCNA pro&ins and MDMZ undetectablein sham or 5 ml/cm’ irradiated cultures. were strongly expressed 48 hours after 30 ml/cm’. Interestingly. pTpT also strongly induced ~53. pZ1 and PCNA (6CQ%,450% and 400%. respectively) above dilueut treated controls. pTpT induction was observed us early as 72 hours and was max,mal 5 days after stimulation, at (he termination of the experimem To determine if pTpT can enhance the cellular response tu WV irradiation, cultures were pretreated with pTpT or dduenf fur 5 days and then were UV or sham irradiated. At 5 and 30 mIlc/cm’UV irradiation, in cultures pre-treated wth pTpT as compared tu dilueut treated cultures, FCNA level was 500% and 200% higher. respectively and MDMZ level was 200% higher. ~21 was also slightly induced. Our studiessuggestfhat pTpT enhances cellular DNA repair capacity through upregulation
DIFFERENT FREQUENCY OF GENE !tA-DNA OLIGONiJCLEO~~DE,&K)a~G tam. S bralv .AdamE ntz. nn
of key protiens, in the absenceof actual DNA damage.
s57
TARGETING EVENTS BY THE EPITHELIAL CELLS. m
ntoandKvon~~eunYoon, Department of Dermat&gy ~~Cutaneons Biology, Jefferson Medical College, Phila, PA 19107 A unique hybrid oligonudeotide composed of both RNA and DNA wus shown to correct a point mutation inn site-specific and inheritable manner in extmchmmasomal and chrumcsomal targets. (Yam et al. Fmc. NatI. Acad. Sci._?&, 2071. 1996, ColeStrauss et al. Sciences 13&5,1966). The RNA stretches of the oligonwleotide was hypothesized IO fadlitate the homologous pecombinatim and a mismatch created tehveen the targeted strand and RNA-DNA digonudeotide may then be.r@nd by the DNA repair system. In order to investigate whether the fnquency d tbe gene targeting is dependent on cell types, we tested two digonncleotides which W&E shown to create a point mntatica in alkaline phosphatax and bglobin gene respectively, in several epithelinl cells. Highly ttansformed epithdial cells (&La) exhibited a much higher frequency in gene conversion by b&b RNA-DNA digonucleotides in mmparison to other innncalized epitbelial cells (HaCaT) or primary keratinocytes. However. the innacellular unrentradon of the digonwleotide in the nnclens of H&a cells was lower than that of HaCaT or primary kaatinocytes. measured by a radic&beled and a iluoresrein-ccmjugated digonnclwtide. Oligcmwleotide extracted from cells was shown to be stable for within the time period III which gene conversion event was measured. Thus, an uptake and inhacellular stability of the digonncleotide are not limiting facbxs in gene targeting events under our experimental conditions. These results suggest that endogenous cellular recombination and DNA repair activities may vary behveen different cdl types and are important for the gene conversion event mediated by the RNA-DNA digrmudwtide.
0338
0341
EKFRESSIONOF VBGF IN CHRONICNON-HElALINGWOUNDS.s
EXPRESSION OF A PUTATIVE PROOPIOMELANOCORTlNmRNA VARIANT IN .a.1 HUMAN EPIDERMIS AND EPIDERMAL CELLS IN CULTURE. m Bshiib Bud&. StevenBovcd. 4 James Nordlund’.Deparnnent of Dermatology’, University of Cincinnati College of Medicine 4 Shrlners Bums Institu&, Cincinnati, OH, USA
*Department of Dermatolo8y. Urdverslty of K(lln. Kdln, Oermany; ‘*Department of Molecular Cell Biology, Max Planck InaUtute.Bad Nauheim,Gummy. The patbngenesls of chronic wounds Is largely unknown. An hypothesis for the mecba&ms undexlyln8 wxmd frdlure suggestr that the buxeaed protedytlc actwty in chronic wounda leads to the de8wwion Of 8rcwth factma ogzssaryf‘xmccesstwrepllr. To tat tbls hypothesis we evaluated the expression of vwxlar en&tbellal groMh factur (VEQF) in chronic mm-be&ng leg u1cea-sand anllyzed the mechanism of VBGF ~~th&r&r& pmteolysls In chmnlc wounds. VBGF 1s a potem an8iogerd&medIatcedIatc play an lmpxtant rolS ln tissue nzpair. Immunobl~ of VEGF expreaslon ln the hyperplastIc epldmnls and damal mononuclear cells of cbronlc wounds. when compared to uninjured control skin. The epidermat sl8nat lo chrnrdc arounds was s&rdEcantly weaka us compared tu epldermal VBGF stalnlng in psoriatic kslons. arhlch saved as. positive WntmI. IntawIngty. the in sin4 hyb&l7.mlon of cbronlc wumls showed a feIaUve stmn8 VEGF mRNA expression in the epldumis and ckmnal mcmonuclearcells, comparable to that of pwrlatJc lesions. Explanatlotts for the dlszre+cy bdueen the strnog mRNA expression and relative low protein sl@al In the e@dennls of cbmnlc woumis suggest eltber a lransltionnl d&a or that VEOF proWn Is unstable in the cbrnnlc wound environment. ‘Ilxrefore. reamblnant VBGF165 was incubated wltb cbronk wound flnld Tbls resulted In the degradation of rVBGl+.. whereas lncnbe.tlonin acute wound fluid did not. Pmteaselnblbltnr &wiles suggested that serineprotepaes are at least *ally responsible for rVBGFl65 degradation in cbrouic wound fluid. Our data demorx?&atefor the first time that VBGF mRNA expression is upregulated In cllronlc wounds and that chronic wound fluid can &grade VEGF. Therefore.our data support the hypomenisthat ti lxreased paeOlytIc a&‘@ in cltmnlc wounds leads to the degradntlon of growth factors and &ruby may impsir their cnntibutlcm to wnund b?aIlng. Funher studlea are needed to a&ate r&zslgnlflcance of this observation for other cyWkloes bnportattt in tissue re@r and our data propose 0 develop strategies hew to mntrd pmtxolytlc acavity in chrnnlc wounds.
FWmpiomeIauocnrtin(PGMC) is a -30,ooO d&n prmein precursor for various bioactive peptidessuch as ACTH, a, p, 7, MSH’s, l&mdorphin and l3-lipotropin. Butb ACTH and a-
MSH have recently been implicatediu cutaueoushyperpigmentation. Previous sludies from our labaraturyhave shownthe presenceof POMC mRNA in uorrnalhuman skin and cultures of epidermal melanocytesand keratiwcytea by in-situ hybridization. In addition, srmog signals of immuuoreactiveACTH and a-MSH were detectedin normal human skin and the epidermal cells. In tie present study, we utilized a combination of immunobistochemistq and in stiu hybridiiion to identify the cell type expressiug POMC mRNA in human epidermis. A melanocyte specific antibody (m&J) and a keratirocyte specific antibody (anticymkeratin)were employed to identify melanocytesand keratirwcytesrespectively in human skin. By immunochemicalstaining, the melanocytes and keratinocytesthat were positively stained with their respective antibodies also exhibited strung signals for POMC mRNA by in-situ hybridization. In order tu identify POMC mRNA iu human skin, reverse transcription-pdytnerasechain reaction (RT-PCR) was performed. An -300 hp POMC product was detected in normal human skin, cultured melanocyteSand keratinocytes. By Southern blot analysis this product was hybridized specifically m the PGMC cDNA. RTPCR productsobtainedfrom normal skin and culturesof melanocytesand keratiwcytes, gave au identical sequence. Comparison of RT-PCR sequenceby Genebauk/Blastrevealed 83% homology to POMC cDNA from human, bovine, pig and monkey sources. This data suggests the presenceof a putative iwform or variant of POMC mRNA in human skin. The detection of POMC mRNA variant in skin may aid in understandingthe mechanism of cutaneoushyperpigmaatiun.
0339
0342
HUMAN PAPILLOMAVIRUS E6 PROTEIN FROM BOTH CERVICAL AND C”TANEOUS TYPE.9INHIBIT p53-DEPENDENT AND p53 -INDBPENDENTAPOPTOSIS S.Ja&ou. I.M.Leil andASturq Centrefur Cutmeow Research. St.Bartholomew’s andthe Royal Schoolof MedicineandDe&try, 2. Newark SW&. London. The two maw transfurmiug pr&ns of oncogemc HumanPapdlomavimses (HPVs) are encodedby the E6 and W ancogcnes and their transfomung abdity can be explainedby their asracianonwiti tumuursuppressor proteins.The E6 protan formscomplexeswuh p53 causinginactivationof p53 fuuctmnincludingcell cyclecheckpoint cuutmlandapoptasis andtheFI prutembindstic Rb protein. The aim of the studywas to invesrigate whetherIhe E-5proteinfrom lug+ and low risk HPVs could mhlbit apaptosisand to detenuineif this E6 Rnctlonwas mediatedby inachvatwnof ~53. Apoprosrs was inducedusmg ultravioletradia6on (WR) and measuredusing terruinalrmusferax ruck end labelbugWNEL) andcolonyassaysfallowingtransfcc6on of HPV genesintoa vanelyof cell types. Our resultsfmm TUNEL stainingof HT1080 cells.wixchcontainwild-type~53, showedthatfbe ~6 prutemfrom bothhigh andlow risk typesinhibitedp53-dependem apoptosis following DNA damage. To investiaate if theE6 urutemscouldalsoiohibito53-iudcoendem aou~tusrs. 10.1lo53 null) cel,l WHIP
FINASTERIDE IS THE MAIN INHIBITOR OF 5u-REDUCTASE ACTIVITY IN DERMAL PAPILLAE OF HL?fAN HAIR FOLLICLES. Rolf Ho@ne*, Wolf ang ,I ?f Derrnatdop Ph$pps U~ver@t, Ma&g ~v~o~$~~p& rhooncology, echmsche Unwers~t& Dresden, T&e Sa-reductase isoeazyma type 1 and 2 and 1713-hydroxysteroid dehydrogenase (170-HSD) are man enzymes in the intracellular androgen metabolism, but at present the knowledge about the ph siolo ‘c role of 5a-reductase isoenzymea is onl incompletely understood Several antK!’ nrs we tned to localii both &enzymes wit&n the human hair follicle, but several m or immnnohistochanical studies gave
were pooled, incubated with SOnMtestosterone for 18 hours and Sa-reduced steroids were analysed by thin-layer chromate phy. Our results show that the volume activity of 5a-reductase in the DP exceeded $. ose tn KS or CTS at least by a factor I4 in the scalp and a &or 80 in the beard. The RS expressed low Sa-reductase, but high 17l3HSD levels, with nndrostenedione as major rnetabdite. The CTS expressed both Sareductase and 17B-HSD, resultin in androstenedione, Sa-androsterone, and 5aandrostanedione. To our surprise f nastende (I&,: 10nrnolil) but not MK-386 (max. cone: Ipmolil) inhibited approx. 80% of 5u-reductase activity in DP and CTS, su esting the pr.+n~ .of type 2 Sa-rednctase, whereas 5a-rednctase acti& within the% S was mamly mbtbrted by MK-386 (IC,: 2Cnmovl) and not by fmasten,Be (I&,. 2COnmoI!l), suggesting the presence of type 1 5a-reductase. Coincubations with fmasteride and hlK-386 resulted in 95-100% inhibition of 5a-reductase activity in DP (It&: approx. Smnov1).Therefore we conclude that type 2 is the main Sa-reductase present m DP.