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ras p21 expression in proliferating skin diseases
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PRESENCE AND LOCALIZATION OF PROTEINS IMMUNOLOGICALLY RELATED TO ERYTHROCYTE SPECTRIN AND PROTEIN
-ras ~21 EXPRESSIONIN PROLIFERATINGSKIN DISEASES
4.1 IN SKIN HIROKO KOIZUMI, AND AKIRA Department of Dermatology, Hokkaido University School of Medicine, Sapporo
TADAMICHI OHKAWARA.
SHIMIZU,
Analogues of human erythrocyte spectrin and protein 4.1 have been examined in the skin by immunochemical techniques using anti-human erythrocyte spectrin, anti- pig brain 8-fodrin and anti-human erythrosyte protein 4.1 antibodies. Immunoblot analysis revealed that pig epidermis contains spectrin-like proteins of 240kDa and 235kDa as well as 4.1-like protein of 68kDa. In addition, pig epidermis also contains higher molecular weight proteins. Immunofluorescence microscopy using these antibodies revealed that both spectrin-like proteins and 4.1-like proteins localize at the cell peripheral cytoplasma of the basal cells and eccrine sweat gland cells. These results suggest the presence of membrane skeletal protein lattice in these cells.
FIBRONECTIONAND FIBRIN IN EXPERIMENTALMOUSE SKIN STAPHYLOCOCCUSAUREUS INFECTION
ARATA KIKUCAI,MASAYUKIAMAGAI*, KOICHI SAKURAOKA, AND TAKEJI NISHIRAWA* Departmentsof Dermatology, SaiseikaiYokohamashiNanbu Hosp., Yokohama,*Keio UniversitySchool of Medicine,Tokyo We reportedthe localizationof ras oncogeneproduct~21 (p21ras)PositiveCells in various-&n tumors and indicated that p21ras may play s role in the differentiation of cells in various skin conditionspreviously. In this study, we have examinedp2lrss positivecells not only in skin carcinomasbut also in other proliferatingskin diseasessuch as psoriasiswlgaris, lichen planus,verruca vulgaris,vsrruca planae juvenilisand verruca senilis to elucidatethe role of p2lrss in the epidermis. p2lrss positivecells were detectedby anti-rasp21 monoclonalantibodyNCC-RAS-001. We concludxhat in both tumorousand inflammatoryskin conditionsp21ras may play a role in the differentiation of epidermalcells.
DETMITION OF ORNITHINE DEXXRBOXYL?!SEGRdE EXPRESSION IN MCUSESKIN'IWA~WITHPH9REOLFSTER'IUMORPFX2GTERAND UL'll7AVIOLEX LIGHT USING IN SITU HYPRIDIZATICN. MXXIO FQKUDA, TAKESHI KC&O, MASAhlITSU ISHII, NOBLMJKI MIZUiW, ISPD MXI'SIJI-YUASA#, SWZO OU?NI#, AND ?DSHIO HAMADA. Departments of Dermatology and Bicchemistry#,Osaka City
HISANORIAKIYAMA, YOSHIKOABE, AKIKO KANAMOTO,HIROKO KANZAKI,JIROARATA Departmentof Dermatology, Okayama Univ. Medical School,Okayama Fibronectin(FN)wss studiedby immunohistochemistry in experimentalstaphylococcussureus (SA)skin infectionin cyclophosphamide-treated mice. FN began to deposit in a granularpattern in the dermis and musculus cutaneuslayer at b hours after the inoculationand increasedwith time. No deposit of FN was observedin s linear patternuntil 7 days after the inoculation, By electronmicroscopy, SA was observedin edematousdermal tissue associatedlooselywith fibrin and collagenfiber until 24 hours, encircledby fibrin during 2 to 6 days, and began to conglomeratein fibrin mass at 7 days after the inoculation. These results demonstrate that solubleFN is main FN in experimentalmouse ski" SA infection,and indicatethat both FN and fibrin play a" importantrole in the developementand eliminationof ski" SA infection,
UniversityMedical School,Osaka Ornithinedecarbcxylase(ODC), a rate-limitingenzyme in polytine biosynthesis,is a well-established marker of skin tumor pramticn. In the present study, we examined the lccalizaticnof OCC gene expression in the mouse skin treatedwith ohotil ester /TPA)and ultravioletliaht >. (WL) using in situ-hybridization.techniw. In the skin treatedwith TPA, the rum&r of positivecells (containingmxe than 5 grains) increasedby 52-fold in the follicular epidermis, by 19-fold in the interfollicular epidermis, and by I-fold in the dermis, as campared with controls. In the skin treated with UVL, each increasedby 25-fold, by la-fold, and by 3.5-fold,respectively.These results indicate that TPA and UVL increase ODC gene expression in the -se epidermis, especially in the follicularregion.
A NEW METHOD FOR QUANTIFYINGKERATINOCYTEDIFFERENTIATION USING IMMUNOFLUORESCENT STAININGOF INVPLL'CRIN AND CYTOFLUOROGRAPHY.
EFFECTS OF BUTYLATED PHENOLIC ANTIOXIDANTS ON EPIDERMAL OIWITHINE DECARESJW AmIVITY AND ITS GENE EXPRESSION INCUCEDBYpHoRBoL Es~lvMxPRC#YrER.
HIDENOBUOKUMURA*,**,KOJI HASHIMOTO*,KUNIO MATSUMOTO*, KUNIHIKOYOSHIF.AWA*. *OSAKA UNIVERSITY SCHOOL OF MEDICINE,**NOEVIRCo.Ltd.,SHIGA INVOLUCRIN IS A PRECURSOR OF DETERGENT-INSOLUBLECORNIFIED ENVELOPE AND A MARKER OF TERMINAL DIFFERENTIATIONOF EPIDERMAL KERATINOCYTES. TO QUANTIFY DIFFERENTIATIONOF CULTURED HUMAN KERATINOCYTES, THE POPULATION OF INVOLUCRINPOSITIVE CELLS WAS ESTIMATED BY IMMUNOFLUORESCENTSTAINING USING ANTI-INVOLUCRIN ANTIBODY AND FLOW CYTOMETRY. NORMAL HUMAN KERATINOCYTES WERE CULTURED UNDER THREE CONDITIONS FOR INDUCTION OF DIFFERENTIATION:O.ItiCaZ+,l.&nM CaZ+,AND l'.'BmM Ca2+ WITH 10% FETAL CALF SERUM. THE POPULATION OF INVOLUCRIN-POSITIVECELLS INCREASED FROM 8.0% TO 22.1%(2.8 FOLD) BY ELEVATING Ca2+ CONCENTRATION,AND TO 37.1%(4.6 FOLD) BY ADDITION OF FCS. THE CYTOFLUOROGRAPHICANALYSIS OF INVOLUCRIN EXPRESSION MADE IT POSSIBLE TO DETERMINE THE NUMBER OF DIFFERENTIATED CELLS IN /4 LARGE NUMBER OF SAMPLES PRECISELY AND RELIABLY. THUS, IT IS A USEFUL METHOD FOR QUANTIFYING KERATINOCYTE DIFFERENTIATION.
HI KCNO, NCWYUKI MIZIJN3,1S.W MATSUIsHc*TITANIm, T 9?S AND 'ICSHIOHAMADn. YUASA#, sHu20oTAN1,
Departments of Dermstolcxqy and Biochemist@, Osaka City UniversityMedical School,Osaka It has been reportedthat free-radicalscavengers,such as butylated hydroxyanisole (BHA) or hydroxytoluene (BFT), suppress skin tumor promotion. In the present study, we examined the effect of BHA and BHT on the activity of ornithinedecxboxylase (One, a well-establishedindicator of tumor promotion) and its gene expression induced by phorhol ester tumor prater (TPA) in the -se skin. TPA-induced ODC activity was inhibited remarkably by topical application of 55 pmol of BHA (inhibition rate; about SO%, measured at 6h). In Northern and dot-blot analysis,.TPA-caused
ODC mRNA level was also markedly
inhibit4 hy the sas~edose of BH?+ (inhibitionrate; a!xut 60%, measured at 4h). On the other hand, BHT failed to <..&
PRESENCE AND LOCALIZATION OF PROTEINS IMMUNOLOGICALLY RELATED TO ERYTHROCYTE SPECTRIN AND PROTEIN
-ras ~21 EXPRESSIONIN PROLIFERATINGSKIN DISEASES
4.1 IN SKIN HIROKO KOIZUMI, AND AKIRA Department of Dermatology, Hokkaido University School of Medicine, Sapporo
TADAMICHI OHKAWARA.
SHIMIZU,
Analogues of human erythrocyte spectrin and protein 4.1 have been examined in the skin by immunochemical techniques using anti-human erythrocyte spectrin, anti- pig brain 8-fodrin and anti-human erythrosyte protein 4.1 antibodies. Immunoblot analysis revealed that pig epidermis contains spectrin-like proteins of 240kDa and 235kDa as well as 4.1-like protein of 68kDa. In addition, pig epidermis also contains higher molecular weight proteins. Immunofluorescence microscopy using these antibodies revealed that both spectrin-like proteins and 4.1-like proteins localize at the cell peripheral cytoplasma of the basal cells and eccrine sweat gland cells. These results suggest the presence of membrane skeletal protein lattice in these cells.
FIBRONECTIONAND FIBRIN IN EXPERIMENTALMOUSE SKIN STAPHYLOCOCCUSAUREUS INFECTION
ARATA KIKUCAI,MASAYUKIAMAGAI*, KOICHI SAKURAOKA, AND TAKEJI NISHIRAWA* Departmentsof Dermatology, SaiseikaiYokohamashiNanbu Hosp., Yokohama,*Keio UniversitySchool of Medicine,Tokyo We reportedthe localizationof ras oncogeneproduct~21 (p21ras)PositiveCells in various-&n tumors and indicated that p21ras may play s role in the differentiation of cells in various skin conditionspreviously. In this study, we have examinedp2lrss positivecells not only in skin carcinomasbut also in other proliferatingskin diseasessuch as psoriasiswlgaris, lichen planus,verruca vulgaris,vsrruca planae juvenilisand verruca senilis to elucidatethe role of p2lrss in the epidermis. p2lrss positivecells were detectedby anti-rasp21 monoclonalantibodyNCC-RAS-001. We concludxhat in both tumorousand inflammatoryskin conditionsp21ras may play a role in the differentiation of epidermalcells.
DETMITION OF ORNITHINE DEXXRBOXYL?!SEGRdE EXPRESSION IN MCUSESKIN'IWA~WITHPH9REOLFSTER'IUMORPFX2GTERAND UL'll7AVIOLEX LIGHT USING IN SITU HYPRIDIZATICN. MXXIO FQKUDA, TAKESHI KC&O, MASAhlITSU ISHII, NOBLMJKI MIZUiW, ISPD MXI'SIJI-YUASA#, SWZO OU?NI#, AND ?DSHIO HAMADA. Departments of Dermatology and Bicchemistry#,Osaka City
HISANORIAKIYAMA, YOSHIKOABE, AKIKO KANAMOTO,HIROKO KANZAKI,JIROARATA Departmentof Dermatology, Okayama Univ. Medical School,Okayama Fibronectin(FN)wss studiedby immunohistochemistry in experimentalstaphylococcussureus (SA)skin infectionin cyclophosphamide-treated mice. FN began to deposit in a granularpattern in the dermis and musculus cutaneuslayer at b hours after the inoculationand increasedwith time. No deposit of FN was observedin s linear patternuntil 7 days after the inoculation, By electronmicroscopy, SA was observedin edematousdermal tissue associatedlooselywith fibrin and collagenfiber until 24 hours, encircledby fibrin during 2 to 6 days, and began to conglomeratein fibrin mass at 7 days after the inoculation. These results demonstrate that solubleFN is main FN in experimentalmouse ski" SA infection,and indicatethat both FN and fibrin play a" importantrole in the developementand eliminationof ski" SA infection,
UniversityMedical School,Osaka Ornithinedecarbcxylase(ODC), a rate-limitingenzyme in polytine biosynthesis,is a well-established marker of skin tumor pramticn. In the present study, we examined the lccalizaticnof OCC gene expression in the mouse skin treatedwith ohotil ester /TPA)and ultravioletliaht >. (WL) using in situ-hybridization.techniw. In the skin treatedwith TPA, the rum&r of positivecells (containingmxe than 5 grains) increasedby 52-fold in the follicular epidermis, by 19-fold in the interfollicular epidermis, and by I-fold in the dermis, as campared with controls. In the skin treated with UVL, each increasedby 25-fold, by la-fold, and by 3.5-fold,respectively.These results indicate that TPA and UVL increase ODC gene expression in the -se epidermis, especially in the follicularregion.
A NEW METHOD FOR QUANTIFYINGKERATINOCYTEDIFFERENTIATION USING IMMUNOFLUORESCENT STAININGOF INVPLL'CRIN AND CYTOFLUOROGRAPHY.
EFFECTS OF BUTYLATED PHENOLIC ANTIOXIDANTS ON EPIDERMAL OIWITHINE DECARESJW AmIVITY AND ITS GENE EXPRESSION INCUCEDBYpHoRBoL Es~lvMxPRC#YrER.
HIDENOBUOKUMURA*,**,KOJI HASHIMOTO*,KUNIO MATSUMOTO*, KUNIHIKOYOSHIF.AWA*. *OSAKA UNIVERSITY SCHOOL OF MEDICINE,**NOEVIRCo.Ltd.,SHIGA INVOLUCRIN IS A PRECURSOR OF DETERGENT-INSOLUBLECORNIFIED ENVELOPE AND A MARKER OF TERMINAL DIFFERENTIATIONOF EPIDERMAL KERATINOCYTES. TO QUANTIFY DIFFERENTIATIONOF CULTURED HUMAN KERATINOCYTES, THE POPULATION OF INVOLUCRINPOSITIVE CELLS WAS ESTIMATED BY IMMUNOFLUORESCENTSTAINING USING ANTI-INVOLUCRIN ANTIBODY AND FLOW CYTOMETRY. NORMAL HUMAN KERATINOCYTES WERE CULTURED UNDER THREE CONDITIONS FOR INDUCTION OF DIFFERENTIATION:O.ItiCaZ+,l.&nM CaZ+,AND l'.'BmM Ca2+ WITH 10% FETAL CALF SERUM. THE POPULATION OF INVOLUCRIN-POSITIVECELLS INCREASED FROM 8.0% TO 22.1%(2.8 FOLD) BY ELEVATING Ca2+ CONCENTRATION,AND TO 37.1%(4.6 FOLD) BY ADDITION OF FCS. THE CYTOFLUOROGRAPHICANALYSIS OF INVOLUCRIN EXPRESSION MADE IT POSSIBLE TO DETERMINE THE NUMBER OF DIFFERENTIATED CELLS IN /4 LARGE NUMBER OF SAMPLES PRECISELY AND RELIABLY. THUS, IT IS A USEFUL METHOD FOR QUANTIFYING KERATINOCYTE DIFFERENTIATION.
HI KCNO, NCWYUKI MIZIJN3,1S.W MATSUIsHc*TITANIm, T 9?S AND 'ICSHIOHAMADn. YUASA#, sHu20oTAN1,
Departments of Dermstolcxqy and Biochemist@, Osaka City UniversityMedical School,Osaka It has been reportedthat free-radicalscavengers,such as butylated hydroxyanisole (BHA) or hydroxytoluene (BFT), suppress skin tumor promotion. In the present study, we examined the effect of BHA and BHT on the activity of ornithinedecxboxylase (One, a well-establishedindicator of tumor promotion) and its gene expression induced by phorhol ester tumor prater (TPA) in the -se skin. TPA-induced ODC activity was inhibited remarkably by topical application of 55 pmol of BHA (inhibition rate; about SO%, measured at 6h). In Northern and dot-blot analysis,.TPA-caused
ODC mRNA level was also markedly
inhibit4 hy the sas~edose of BH?+ (inhibitionrate; a!xut 60%, measured at 4h). On the other hand, BHT failed to <..&