Estudos funcionais e biológicos associados à interação de DUSP3 com a proteína HNRNPC1/C2 / Functional and biological studies associated to DUSP3 and HNRNPC1/C2 protein inte ractions

A regulação da fosforilação/desfosforilação das proteínas é o eixo central de muitas cascatas de sinalização. A fosfatase DUSP3, constituída apenas por um único domínio catalítico, desempenha papéis fundamentais na proliferação e senescência celular. Nas células HeLa, submetidas ao estresse genotóxico, o DUSP3 interage fisicamente com as proteínas HNRNPC, mas o efeito dessa função molecular ainda é desconhecido. Aqui demostramos que a ausência de DUPS3 mantem a proteína HNRNPC1/C2 num estado hiperfosforilado. Para entender melhor o envolvimento da interação DUSP3-HNRNPC nas funções biológicas da HNRNPC1/C2, foram estudadas células de fibroblasto deficientes de DUSP3. Foi analisado o efeito da deficiência de DUSP3 na biogênese dos ribossomos através do ensaio de perfil de polirribossomos e quantificação dos rRNAs com RT-qPCR. Os resultados mostraram que a deficiência de DUSP3 não afeta a maturação das subunidades ribossômicas, mas teria um impacto na transcrição dos pré-rRNAs e no acumulo das espécies 47S/45S. A expressado de genes contendo sequencias IRES foi analisado através do RT-qPCR e sua tradução ao longo do ciclo e em condições de estresse. Da expressão, não existe nenhuma diferença nos níveis de transcrição dos genes c-myc e xiap nas células normais e deficientes de DUSP3 em condições basais. Embora a síntese destas proteínas é maior nas células deficientes, mantendo um nível maior de tradução ao longo de todo o ciclo. Sob condições de estresse, esta duas proteínas sempre mantem uma maior expressão nas células Knockdown para DUSP3. Neste trabalho também foi estabelecido a presença de DUSP3 nos complexos da subunidade 40S, através do analise das frações obtidas do ensaio de polirribossomos e interação in vitro (Co-IP). A presença de DUSP3 nas subunidades 40S, os monossomas 80S e polissomos poderia ser através da interação direta com proteínas que possuem um domínio RRM e seria dependente dos complexos formados pelas proteínas e seus RNAs alvos. Aqui mostramos a interação in vitro de DUSP3 com a proteína PABP (com quatro domínios RRM), proteína que tem um papel importante na manutenção da taxa global de tradução, esta interação é enfraquecida na ausência de RNAs. A deficiência de DUSP3 também teria um impacto na interação das proteínas HNRNPC1/C2 e P53 in vitro. A ausência de DUSP3 diminui a interação HNRNPC-P53 através da hiperfosforilação da proteina HNRNPC1/C2. A perda desta interação, aumentaria os níveis da proteína P53 na célula deficiente de DUSP3 e poderia gerar parada no ciclo celular. Através de ensaios de imunofluorescência, se observo uma maior taxa de transcrição global na célula deficiente de DUSP3. Por fim, aqui demostramos que a interação direta de DUSP3 e HNRNPC1/C2 vai permitir a regulação das funções biológicas desta proteína, e a ausência de DUSP3 vai ter efeitos pleiotrópicos na homeostase da célula

inglêsProtein phosphorylation/dephosphorylation regulation is a central axis of many signaling cascades. DUSP3 phosphatase, consisting only of a single catalytic domain, plays key roles in cell proliferation and senescence. In HeLa cells subjected to genotoxic stress, DUSP3 physically interacts with HNRNPC proteins, but the effect of this molecular function is still unknown. Here we demonstrate that the absence of DUPS3 keeps the HNRNPC1/C2 proteins in a hyperphosphorylated state. To better understand the involvement of DUSP3- HNRNPC interaction on the biological functions of HNRNPC1/C2, DUSP3 deficient fibroblast cells were studied. The effect of DUSP3 deficiency on ribosome biogenesis was analyzed by polyribosome profile assay and RT-qPCR for rRNA quantification. The results showed that DUSP3 deficiency does not affect ribosomal subunit maturation, but would have an impact on transcription of pre-rRNAs and accumulation of 47S / 45S species. The expression of genes containing IRES sequences was analyzed by RT-qPCR and their translation throughout the cycle and under stress conditions. From expression, there is no difference in transcriptional levels of c-myc and xiap genes in normal and DUSP3 deficient cells under basal conditions. Although, the synthesis of these proteins is higher in deficient cells and these maintain a higher level of translation throughout the cell cycle. Under stress conditions, these two proteins always maintain higher expression in Knockdown cells for DUSP3. In this work, the presence of DUSP3 in the 40S ribosomal subunit complexes was also established by analyzing the fractions obtained from the polyribosome assay and in vitro interaction (CoIP). The presence of DUSP3 in the 40S subunits, 80S monosomes and polysomes could be through direct interaction with proteins that have an RRM domain and would be dependent on the complexes formed by the proteins and their target RNAs. Here we show the in vitro interaction of DUSP3 with PABP protein (with four RRM domains), a protein that plays an important role in maintaining the overall translation rate, this interaction is weakened in the absence of RNAs. DUSP3 deficiency would also have an impact on the interaction of HNRNPC1/C2 and P53 proteins in vitro. The absence of DUSP3 decreases HNRNPC-P53 interaction through hyperphosphorylation of the HNRNPC1/C2 proteins. Loss of this interaction would increase P53 protein levels in the DUSP3 deficient cell and could lead to cell cycle arrest. Through immunofluorescence assays, a higher overall transcription rate is observed in the DUSP3 deficient cell. Finally, we demonstrate that the direct interaction of DUSP3 and HNRNPC1/C2 will allow the regulation of the biological functions of this protein, and the absence of DUSP3 will have pleiotropic effects on cell homeostasis

Effects of tumor biobank storage on polysome stability

Background: Human biological material has become an important resource for biomedical research. Tumor Biobanks are facilities that collect, store and distribute samples of tumor and normal tissue for further use in basic and translational cancer research. mRNA-translation has been demonstrated to modulate protein levels and is considered a fundamental post-transcriptional mechanism of gene expression regulation. Thus, determining translation efficiencies of individual mRNAs in human tumors may add another layer of information that contributes to the understanding of tumorigenic pathways. To analyze the RNAs actively engaged in translation, RNAs associated with ribosomes (polysomes) are isolated, identified and compared to total RNA. However, the application of this technique in human tumors depends on the stability of the polysomal structure under Biobank storage conditions that usually consists of ultra-low temperature. Since the effect of freezing on the stability of the polysomal structure in stored tumor samples is not known, it is essential to evaluate this factor in the frozen samples, validating the use of biobank samples in studies of translational efficiency. Methods: Xenograft tumors were divided in two parts, half was subject to immediate processing, and half was frozen for posterior analysis. Both parts were subject to polysomal separation, RNA extraction and identification through RNAseq. Results: It was possible to successfully extract and identify total and polysomal RNA from both fresh and frozen tumoral tissue. The quantification of the polysome profile indicated no difference in the translational efficiency estimated in fresh versus frozen tissue. Gene expression data from the fresh versus frozen tissues were compared and the correlation between the polysome associated fresh x frozen (R = 0,89) and total fresh x frozen (0,90) mRNAs was calculated. No difference was identified between the two conditions. Conclusions: We demonstrated that tissue freezing does not affect the polysomal structure, consequently validating the viability of the use of biobank stored tissue for polysome associated RNA analysis (AU)

Ultrastructural changes of corpus luteum after ovarian stimulation at implantation period

To achieve multiple oocytes for in vitro fertilization, ovulation induction is induced by gonadotropins; however, it has several effects on oocytes and embryo quality and endometrium receptivity. The aim of this study was to assess ultrastructural changes of corpus luteum after ovarian induction using human menopausal gonadotropin [HMG] and human chorionic gonadotropin [HCG] during luteal phase at implantation period. Female NMRI mice [6-8 weeks] were divided into control and stimulated groups. In the control group, the mice were rendered pseudopregnant and in the ovarian induction group, the mice were rendered pseudopregnant after the ovarian induction. The samples were obtained from the ovary in each group at the same time during luteal phase at implantation period. Ultrastructural changes were assessed using electron microscopy study. Our results displayed some identifiable changes in ultrastructure of corpus luteum in ovarian induction group. These changes included enhancement of the apoptosis and intercellular space, whereas the angiogenesis was decreased. The findings indicated a decline in organelle density in the cytoplasm of ovarian induction, such as mitochondria, endoplasmic reticulum and polyribosome. Furthermore, chromatin condensation of nuclei was observed in some cells. The ovarian induction using HMG and HCG resulted in some ultrastructural changes on the corpus luteum at implantation period, which could affect on the pregnancy rate

Grânulos de processamento de mRNAs em Trypanosoma cruzi / Granules of processing of mRNAs in Trypanosoma cruzi

Em tripanossomatídeos a regulação da expressão gênica ocorre principalmente em nível pós-transcricional por mecanismos que envolvem mudanças na estabilidade e no acesso dos mRNAs aos polissomos. Estes mecanismos permitem uma rápida adaptação destes parasitas a diferentes condições as quais são expostos durante o ciclo de vida. Estudos recentes têm demonstrado que grânulos de mRNA, presentes em diversos eucariotos têm papel fundamental na regulação da expressão gênica em nível pós-transcricional. Estes grânulos são divididos em diferentes classes (entre elas os P-bodies e os grânulos de estresse), contém mRNPs não comprometidas com a tradução, compartilham algumas proteínas e podem utilizar mecanismos semelhantes para regular o metabolismo de mRNAs. Os P-bodies são sítios de estocagem e/ou degradação de mRNAs, enquanto os grânulos de estresse estão envolvidos na triagem e estocagem de diversos transcritos. Como existe evidência de que moléculas de mRNA podem ser mantidas estáveis e não associadas à polissomos em T. cruzi, nós conjecturamos a possibilidade de que estas estruturas estejam presentes e sejam funcionais neste parasita. Os genes que codificam alguns componentes de P-bodies estão presentes em T. cruzi e embora algumas proteínas que constituem o cerne destas estruturas pareçam estar ausentes neste parasita, a maioria das funções inerentes aos P-bodies estão representadas. Nós identificamos e clonamos a proteína TcDhh1, uma DEAD box RNA helicase altamente conservada entre os eucariotos e considerada marcadora de P-bodies em mamíferos e leveduras. Esta proteína é constitutivamente expressa durante todo o ciclo de vida do parasita, está presente em complexos independentes de polissomos e está localizada em foci citoplasmáticos que variam em número quando as células são submetidas a condições de estresse ou ao tratamento com as drogas cicloheximida e puromicina. Para determinar a composição destes grânulos, nós realizamos ensaios de...

Growth Inhibitory Effects of Various Herbal Extracts on Metronidazole Resistant Strain of Trichomonas vaginalis / 감염과화학요법

BACKGROUND: Metronidazole has been known as the most effective drug for treatment of Trichomonas vaginalis-related diseases. However, it has been reported that metronidazole has adverse effects and incidence of metronidazole-resistant T. vaginalis (CDC085) has increased. Development of new drug, which is effective against metronidazole-resistant T. vaginalis and showing no adverse effects, has been required. METHODS: The purpose of this study was to investigate effects of various extracts from herbs such as Quisqualis indica, Gleditsia sinensis, Prunus armeniaca, Morus alba, Platycodon grandiflorum, Ailanthus altissima, Stemona japonica, Biota orientalis, Dryobalanops aromatica, and Cimicifuga heracleifolia on metronidazole resistant strain of T. vaginalis in vitro (CDC085). RESULTS: Anti-Trichomonas activities were observed in T. vaginalis treated with G. sinensis, P. armeniaca, and P. grandiflorum on the growth and fine structure of metronidazole resistant strain of T. vaginalis. Of the three standard extracts that showed the most effective anti-trichomonas activity, G. sinensis was the most effective. The inhibitory effects of fraction extracts of this drug were shown on the growth of T. vaginalis. The fine structure of the cytoplasm was changed after application of G. sinensis extract. The number of polyribosome and hydrogenosome decreased whereas the number of food vacuole and vacuole in the cytoplasm increased, compared with that of untreated control group. CONCLUSION: The results of our study indicate that G. sinensis may induce the inhibition of cell multiplication as well as impairment of protein synthesis of metronidazole resistant strain of T. vaginalis in vitro.

Growth Inhibitory Effects of Various Herbal Extracts on Metronidazole Resistant Strain of Trichomonas vaginalis / 감염과화학요법

BACKGROUND: Metronidazole has been known as the most effective drug for treatment of Trichomonas vaginalis-related diseases. However, it has been reported that metronidazole has adverse effects and incidence of metronidazole-resistant T. vaginalis (CDC085) has increased. Development of new drug, which is effective against metronidazole-resistant T. vaginalis and showing no adverse effects, has been required. METHODS: The purpose of this study was to investigate effects of various extracts from herbs such as Quisqualis indica, Gleditsia sinensis, Prunus armeniaca, Morus alba, Platycodon grandiflorum, Ailanthus altissima, Stemona japonica, Biota orientalis, Dryobalanops aromatica, and Cimicifuga heracleifolia on metronidazole resistant strain of T. vaginalis in vitro (CDC085). RESULTS: Anti-Trichomonas activities were observed in T. vaginalis treated with G. sinensis, P. armeniaca, and P. grandiflorum on the growth and fine structure of metronidazole resistant strain of T. vaginalis. Of the three standard extracts that showed the most effective anti-trichomonas activity, G. sinensis was the most effective. The inhibitory effects of fraction extracts of this drug were shown on the growth of T. vaginalis. The fine structure of the cytoplasm was changed after application of G. sinensis extract. The number of polyribosome and hydrogenosome decreased whereas the number of food vacuole and vacuole in the cytoplasm increased, compared with that of untreated control group. CONCLUSION: The results of our study indicate that G. sinensis may induce the inhibition of cell multiplication as well as impairment of protein synthesis of metronidazole resistant strain of T. vaginalis in vitro.

TNF-alphaUp-regulated the Expression of HuR, a Prognostic Marker for Ovarian Cancer and Hu Syndrome, in BJAB Cells

BACKGROUND: Hu syndrome, a neurological disorder, is characterized by the remote effect of small cell lung cancer on the neural degeneration. The suspicious effectors for this disease are anti-Hu autoantibodies or Hu-related CD8+ T lymphocytes. Interestingly, the same effectors have been suggested to act against tumor growth and this phenomenon may represent natural tumor immunity. For these diagnostic and therapeutic reasons, the demand for antibodies against Hu protein is rapidly growing. METHODS: Polyclonal and monoclonal antibodies were generated using recombinant HuR protein. Western blot analyses were performed to check the specificity of generated antibodies using various recombinant proteins and cell lysates. Extracellular stimuli for HuR expression had been searched and HuR-associated proteins were isolated from polysome lysates and then separated in a 2-dimensional gel. RESULTS: Polyclonal and monoclonal antibodies against HuR protein were generated and these antibodies showed HuR specificity. Antibodies were also useful to detect and immunoprecipitate endogenous HuR protein in Jurkat and BJAB. This report also revealed that TNF-alphatreatment in BJAB up-regulated HuR expression. Lastly, protein profile in HuR-associated mRNA- protein complexes was mapped by 2-dimensional gel electrophoresis. CONCLUSION: This study reported that new antibodies against HuR protein were successfully generated. Currently, project to develop a diagnostic kit is in process. Also, this report showed that TNF-alphaup-regulated HuR expression in BJAB and protein profile associated with HuR protein was mapped.

In vitro Effect of Kalopanaxsaponin A on the Ultrastructure of Trichomonas vaginalis / 감염과화학요법

BACKGROUND: T. vaginalis is a pathogenic protozoa infecting genitourinary tract. Metronidazole is a drug of choice to treat T. vaginalis infection. However, because of appearance of metronidazole- resistant T. vaginalis, it is needed to develop an alternative drug. So, we evaluated the effects of various concentration of kalopanaxsaponin A against T. vaginalis. METHODS: The antiprotozoal effect of kalopanaxsaponin A on the growth and fine structure of T. vaginalis was examined by using trypan blue exclusion assay and electron microscopy. RESULTS: The viability test showed markedly decreased number of T. vaginalis, treated with increasing concentration of kalopanaxsaponin A. We evaluated the electron microscopic findings for antiprotozoan effects against T. vaginalis. SEM showed that in T. vaginalis treated with 4 microgram/mL for 1 hour, axostyle was shrinked and flagella began to disappear. With gradual increase of the concentration of kalopanaxsaponin A, destruction of cell membrane was getting more severe, and degenerative change was observed in T. vaginalis treated with 8 microgram/mL for 2 hours. TEM showed that in T. vaginalis treated with 4 microgram/mL for 2 hours, the vacuoles in cytoplasm were larger and hydrogenosomes were smaller than those in control group. The number of vacuole was increased, the nucleus was destroyed, the number of polyribosome and free ribosome was also decreased in trichomonads treated with kalopanaxsaponin A (8 microgram/mL), which indicated the occurrence of degenerative changes. CONCLUSION: These results indicated that kalopanaxsaponin A had the antiprotozoal effect on T. vaginalis by inhibition of cell multiplication as well as an impairment of protein synthesis.

In vitro Effect of Kalopanaxsaponin A on the Ultrastructure of Trichomonas vaginalis / 감염과화학요법

BACKGROUND: T. vaginalis is a pathogenic protozoa infecting genitourinary tract. Metronidazole is a drug of choice to treat T. vaginalis infection. However, because of appearance of metronidazole- resistant T. vaginalis, it is needed to develop an alternative drug. So, we evaluated the effects of various concentration of kalopanaxsaponin A against T. vaginalis. METHODS: The antiprotozoal effect of kalopanaxsaponin A on the growth and fine structure of T. vaginalis was examined by using trypan blue exclusion assay and electron microscopy. RESULTS: The viability test showed markedly decreased number of T. vaginalis, treated with increasing concentration of kalopanaxsaponin A. We evaluated the electron microscopic findings for antiprotozoan effects against T. vaginalis. SEM showed that in T. vaginalis treated with 4 microgram/mL for 1 hour, axostyle was shrinked and flagella began to disappear. With gradual increase of the concentration of kalopanaxsaponin A, destruction of cell membrane was getting more severe, and degenerative change was observed in T. vaginalis treated with 8 microgram/mL for 2 hours. TEM showed that in T. vaginalis treated with 4 microgram/mL for 2 hours, the vacuoles in cytoplasm were larger and hydrogenosomes were smaller than those in control group. The number of vacuole was increased, the nucleus was destroyed, the number of polyribosome and free ribosome was also decreased in trichomonads treated with kalopanaxsaponin A (8 microgram/mL), which indicated the occurrence of degenerative changes. CONCLUSION: These results indicated that kalopanaxsaponin A had the antiprotozoal effect on T. vaginalis by inhibition of cell multiplication as well as an impairment of protein synthesis.

Development and Migration of Megakaryocyte during Hepatic Hemopoiesis in Human Fetuses / 대한해부학회지

Liver tissuses obtained from 5 human fetuses between 11 weeks and 23 weeks of gestation during the high activity of hepatic hemopoiesis were observed with transmission electron microscope using continuous series of thin sections. The objective of present study was to evaluate ultrastructures of megakaryopoietic cells, the migration of extravascular megakaryocyte into the sinusoidal lumen and the relevence between a migrated megakaryocyte and a Kupffer cell. Immature megakaryocytes were usually observed between growing hepatic laminae and within hepatic sinusoids. A megakaryoblast contained numerous polyribosomes, rather large mitochondria, short tubular elements of rough endoplasmic reticulum and small granules. Moreover, demarcation tubules and a few small specific granules were observed in immature megakaryocytes. The nucleus was mononuclear but frequently indented. With maturation, the nuclei were multilobulated. In the cytoplasm, in contrast to the decrease in polyribosomes and rough endoplasmic reticulum, the numerous specific granules and well -developed demarcation membrane system were predominant. Thereafter cytoplasmic zonation was observed clearly in maturing and mature megakaryocytes. Some megakaryocytes passed through the sinusoidal lining epithelium and into the hepatic sinusoids. The cell to cell interaction was often found as adhesion between migrated megakaryocyte and Kupffer cell, and erythroblasts within megakaryocyte (emperipolesis). These results suggest that intravascular megakaryopoiesis in addition to extravascular megakaryopoiesis occurs to produce platelet during the human fetal liver.

In vitro Effect of Sophora flavescens on the Ultrastructure of Trichomonas vaginalis Donne / 감염

BACKGROUND: Trichomonas vaginalis is a pathogenic protozoa infecting human genitourinary tract. Metronidazole is currently the drug of choice to treat T. vaginalis infection. However, because of the side effects and the occurrence of resistant strains of metronidazole, it is needed to investigate alternatives. METHODS: The antiprotozoal effect of aquatic extract from Sophora flavescens on the growth and fine structure of T. vaginalis was examined by using trypan blue exclusion assay and electron microscopy. RESULTS: One hour after the addition of 4 mg/mL extract and half hour after the addition of 5 mg/mL showed antiprotozoal effect. One to two hours after the addition of 3 mg/mL extract, the movement of flagella and axostyle had disappeared, but death of the cells had not occurred until two hours after the addition. The fine structure of the cytoplasm was also changed half an hour to two hours after addition. The number of polyribosome decreased when that of single ribosomes in the cytoplasm increased. CONCLUSION: These results indicated that S. flavescens had the antiprotozoal effect on T. vaginalis by inhibition of cell multiplication as well as an impairment of protein synthesis.

Morphological Change with the Induction of Cisplatin Resistance from the Bladder Cell Lines / 대한비뇨기과학회지

PURPOSE: When Managing metastatic bladder tumors, to overcome the resistance mechanism of cisplatin is a main problem to be solved. The objective is to confirm the changes of general and ultrastructural morph ology with the induction of cisplatin resistance from the bladder cell line. MATERIALS AND METHODS: The samples of this investigation are 2ng/ml-cisplatin resistant human bladder cell lines T24R2 established by SNUH Urology and the drug resistant bladder cell lines T24 was obtained from ATCC, as a control group. We cultured the resistant cell line on the slide and observed it using light microscopy to see the general morphology. For the ultrastructural morphology, we fixed cultured cells, made an epon block, sliced an ultrathin section and observed it using H-71000 EM. RESULTS: Under light microscopy, the cytoplasm of the resistant cell line shows a plumper pattern than that of the parent cell. Under electronmicroscopy, the chromatin of the resistant cell line has a relatively finely dispersed chromatin pattern when compared to the parent cell line, which shows a coarse and aggregated chromatin pattern. Within the cytoplasm, the mitochondrial volume, dilated rough endoplasmic reticulum, polyribosomes and ribosomes are moderately increased in the resistant cell line when compared to the parent cell line. In particular, we found a great amount of double membrane vesicle near the cell surface and pinocytic vesicles on the surface, which are seldom observed within the parent cells. CONCLUSIONS: We concluded that the cisplatin resistant human bladder cell lines (T24R2) underwent a morphological change with the induction of cisplatin resistance, and we hypothesize that the resistant cell's ultrastructure, which shows morphological change, will be involved in the drug resistance mechanism. Regarding this matter, further research will be needed.

Polypeptide pattern of mRNA isolated from polysomal and non-polysomal mRNA-protein(mRNP) fractions in rat myoblast.

Polypeptides synthesized from polysome-bound and unbound fractions of polyA mRNA were studied by in-vivo cells labelling and cDNA-mRNA hybrid-arrest translation. The polypeptides synthesized from unbound(free) fraction appeared to be significantly reduced in the absence of cycloheximide in L6 myoblast. The cDNA excess-mRNA hybrid-arrest translation exhibited rare polypeptide sequences in the unbound fractions.

Vaccinia core transcription prevents in vitro depolymerization of polyribosomes in rabbit reticulocyte lysate

When a rabbit reticulocyte lysate is incubated in the presence of vaccina cores, protein synthesis is umpaired at the level of the initiation step and the polyribosomes are depolymerized. However, when the same system is coupled with virus transcription: a) protein synthesis is restored, b) the initiation step is not inhibited, and c) the polyribosomes are not disaggregated. A viral factor activated in the absence of virus transcription and not activated when RNA synthesis occurs may be involved in the early mechanism of protein synthesis inhibition by vaccinia virus

A direct method for cDNA cloning employing translation released mRNA

1. A new cloning procedure is described for cDNA synthesis from mRNA released by in vitro translation of polysomes in a cell-free amino acid incorporating system. The usefulness of the method lies in the feasibility of employing nanogram amounts of mRNA. 2. Complementary DNA is synthesized derectly in the translation mixture simply by adjusting the concentration of some components and removing ribosomes by boiling and centrifugation. 3. As an example, we report here the construction and characterization of cDNA clone corresponding to chick alfa (1) procollagen starting from a collagen-synthesizing polysome fraction obtained from chick embryos

A Case of Lymphomatoid Papulosis / 대한피부과학회지

We report herein a case of lymphomatoid papulosis in a 23-year-old female who had recurrent erythematous pinhead to pea sized papules and nodules on the both inner thighs and forearms for 4 years. Some lesions showed central hemorrhagic necrosis or scale formation. Individual lesions persisted for several months and showed spontaneous regression leaving pigmentation or depigmented atrophic scar. Histopathologically, there was marked cell infiltration, especially in the dermoepiderrnal junction and perivascular area. Infiltraled cells consisted of sorne ncutrophils and numerous atypical cells that had variable sized and irregular shaped nuclei. Electronmicroscopically, some atypical cells had cerebriform nuclei with marked peripheral condensation of chromatin and cytoplasm contained a few organelles, many polyribosomes and some dense bodies.