Heejeong Lee

Cancer-Associated Expression of Minichromosome Maintenance 3 Gene in Several Human Cancers and Its Involvement in Tumorigenesis

Purpose: The purpose of our study was to identify an unique gene that shows cancer-associated expression, evaluates its potential usefulness in cancer diagnosis, and characterizes its function related to human carcinogenesis. Experimental Design: We used the differential display reverse transcription-PCR method with normal cervical, cervical cancer and metastatic tissues, and cervical cancer cell line to identify genes overexpressed in cancers. Results: We identified a minichromosome maintenance 3 (MCM3) gene that was overexpressed in various human cancers, including leukemia, lymphoma, and carcinomas of the uterine cervix, colon, lung, stomach, kidney and breast, and malignant melanoma. Western blot and immunohistochemical analyses also revealed that MCM3 protein was elevated in most of human cancer tissues tested. We compared the MCM3 protein expression levels in human cancers with conventional proliferation markers, Ki-67 and proliferating cell nuclear antigen. MCM3 antibody was the most specific for multiple human cancers, whereas proliferating cell nuclear antigen was relatively less effective in specificity, and Ki-67 failed to detect several human cancers. The down-regulation of MCM3 protein level was examined under serum starvation in both normal and cancer cells. Interestingly, MCM3 protein was stable in MCF-7 breast cancer cells even up to 96 hours after serum starvation, whereas it was gradually degraded in normal BJ fibroblast cells. Nude mice who received injections of HEK 293 cells stably transfected with MCM3 formed tumors in 6 weeks. Conclusions: Our study indicates that determination of MCM3 expression level will facilitate the assessment of many different human malignancies in tumor diagnosis, and MCM3 is involved in multiple types of human carcino-genesis.

Expression of miRNAs and PTEN in endometrial specimens ranging from histologically normal to hyperplasia and endometrial adenocarcinoma

We investigated the relationship between frequently deregulated microRNAs (miRNAs) and enodometrial pathology in an attempt to find the most dependable miRNA or combination of miRNAs to identify normal, hyperplastic and malignant endometrial tissues. We also investigated the association between those miRNAs and PTEN status. We measured the expression of six miRNAs (miR-21, 182, 183, 200a, 200c and 205) in 75 formalin-fixed, paraffin-embedded normal, hyperplastic, and malignant endometrial tissue blocks using Taqman-based real-time PCR assays. PTEN loss of expression was assessed in the same endometrial tissues by immunohistochemistry. Expression of five miRNAs (miR-182, 183, 200a, 200c and 205) was significantly higher in endometrial carcinoma (CA) when compared with complex atypical hyperplasia (CAH), simple hyperplasia (SH) and normal endometrial tissue (P<0.05, respectively). Considering the likelihood ratio and number of parameters, the composite panel of six miRNAs was the best marker, revealing a sensitivity of 91% and a specificity of 94% in differentiating endometrial CA from endometrial hyperplasia or normal endometrium while the individual miRNAs exhibited 64–77% sensitivity and 66–91% specificity. Interestingly, in distinguishing endometrial CA from CAH, the composite panel of four miRNAs (miR-182, 183, 200a, 200c) was the best marker, producing 95% sensitivity and 91% specificity. The percentage of PTEN loss was significantly higher in endometrial CA compared with SH (68% vs 24%, P<0.05), and it was also higher in CAH compared with SH (71% vs 24%, P<005). Aberrant expression of miRNAs and loss of PTEN expression are common in endometrial hyperplasia and CA. They might serve to increase the diagnostic reproducibility and improve discrimination, especially, between CAH and CA by miRNA expression profiles and between simple and complex hyperplasia through PTEN expression patterns. Those expression profiles of biomarkers also might be used to predict the potential for progression from endometrial hyperplasia to invasive CA.

Expression of HPV L1 capsid protein in cervical specimens with HPV infection

We tried to investigate the expression rate of human papillomavirus (HPV) L1 capsid protein in uterine cervical specimens and correlate it with the grade of dysplasia, HPV genotype and age of the patients. Among uterine cervical specimens proved to have HPV by DNA genotyping test, eighty cytology‐biopsy matched cases and 22 unmatched cytology specimens were selected. Immunostaining for L1 capsid protein was performed on both cervical smears and tissue sections. The L1 capsid protein was expressed mainly in the nuclei, but occasionally in the cytoplasm of cells located in the superficial layer of squamous epithelium. The immunostaining for L1 capsid protein showed positive reaction in 47 cases (46.1%) of cervical smears and in 10 cases (12.5%) of tissue sections (P = 0.001). Cytologic diagnosis revealed a higher expression rate in LSILs (25/33; 75.8%) than in HSILs and cervical cancers (8/20; 40.0% and 2/5; 40%, respectively) (P = 0.006). In LSILs, cases with low‐risk type HPV showed a higher L1 capsid expression rate than those with the high‐risk type HPV (88.9% vs. 70.8%). The L1 capsid expression rate decreased in the over‐40‐year‐old age group compared to the younger age (49.2% vs. 50.8%). Cytology smears were superior to tissue sections for the detection of L1 capsid protein expression. LSILs and HPV low‐risk group showed higher L1 capsid expression rate than HSILs and HPV high‐risk group, which suggests that L1 capsid expression might be related to a favorable disease biology. Diagn. Cytopathol. 2008.

MicroRNA expression profiling and Notch1 and Notch2 expression in minimal deviation adenocarcinoma of uterine cervix.

BackgroundMicroRNA (miRNA) expression is known to be deregulated in cervical carcinomas. However, no data is available about the miRNA expression pattern for the minimal deviation adenocarcinoma (MDA) of uterine cervix. We sought to detect deregulated miRNAs in MDA in an attempt to find the most dependable miRNA or their combinations to understand their tumorigenesis pathway and to identify diagnostic or prognostic biomarkers. We also investigated the association between those miRNAs and their target genes, especially Notch1 and Notch2.MethodsWe evaluated miRNA expression profiles via miRNA microarray and validated them using.real-time PCR assays with 24 formalin-fixed, paraffin-embedded tissue blocks of MDA and 11 normal proliferative endocervical tissues as control. Expression for Notch1 and 2 was assessed by immunohistochemistry.ResultsMiRNA-135a-3p, 192-5p, 194-5p, and 494 were up-regulated, whereas miR-34b-5p, 204-5p, 299-5p, 424-5p, and 136-3p were down-regulated in MDA compared with normal proliferative endocervical tissues (all P <0.05). Considering the second-order Akaike Information Criterion consisting of likelihood ratio and number of parameters, miR-34b-5p showed the best discrimination power among the nine candidate miRNAs. A combined panel of miR-34b-5p and 194-5p was the best fit model to discriminate between MDA and control, revealing 100% sensitivity and specificity. Notch1 and Notch2, respective target genes of miR-34b-5p and miR-204-5p, were more frequently expressed in MDA than in control (63% vs. 18%; 52% vs. 18%, respectively, P <0.05). MiR-34b-5p expression level was higher in Notch1-negative samples compared with Notch1-positive ones (P <0.05). Down-regulated miR-494 was associated with poor patient survival (P = 0.036).ConclusionsMDA showed distinctive expression profiles of miRNAs, Notch1, and Notch2 from normal proliferative endocervical tissues. In particular, miR-34b-5p and 194-5p might be used as diagnostic biomarkers and miR-494 as a prognostic predictor for MDA. The miR-34b-5p/Notch1 pathway as well as Notch2 might be important oncogenic contributors to MDA.

HCCRBP‐1 directly interacting with HCCR‐1 induces tumorigenesis through P53 stabilization

Oncogene HCCR‐1 functions as a negative regulator of the p53 and contributes to tumorigenesis of various human tissues. HCCR transgenic mice developed breast cancers but it is unknown how HCCR‐1 contributes to human tumorigenesis. This study identified a HCCR‐1‐binding protein 1 (HCCRBP‐1) as an HCCR binding partner by performing yeast two hybrid screening. Their endogenous interaction was further confirmed by coimmunoprecipitation experiments. These two proteins colocalized in the mitochondria. HCCRBP‐1 was overexpressed in various human tumors. In addition, HCCRBP‐1 alone converted NIH/3T3 cells into tumor cells in combination with no other oncogenes. HCCRBP‐1 induced tumorigenesis by markedly activating PKC activities but decreasing the pro‐apoptotic PKCα and PKCδ isoform levels. We observed that p53 stabilization also occurred with functional impairment in HCCRBP‐1‐transfected 293 cells, as indicated by defective induction of p21, MDM2 and bax. Indeed, HCCRBP‐1 decreased p21 promoter activity probably via p53 stabilization leading to the defective function. These results indicate that HCCRBP‐1 oncogene induces p53 stabilization and thereby contributes to tumorigenesis.

De Novo Transcriptome Analysis of Cucumis melo L. var. makuwa

Oriental melon (Cucumis melo L. var. makuwa) is one of six subspecies of melon and is cultivated widely in East Asia, including China, Japan, and Korea. Although oriental melon is economically valuable in Asia and is genetically distinct from other subspecies, few reports of genome-scale research on oriental melon have been published. We generated 30.5 and 36.8 Gb of raw RNA sequence data from the female and male flowers, leaves, roots, and fruit of two oriental melon varieties, Korean landrace (KM) and Breeding line of NongWoo Bio Co. (NW), respectively. From the raw reads, 64,998 transcripts from KM and 100,234 transcripts from NW were de novo assembled. The assembled transcripts were used to identify molecular markers (e.g., single-nucleotide polymorphisms and simple sequence repeats), detect tissue-specific expressed genes, and construct a genetic linkage map. In total, 234 single-nucleotide polymorphisms and 25 simple sequence repeats were screened from 7,871 and 8,052 candidates, respectively, between the KM and NW varieties and used for construction of a genetic map with 94 F2 population specimens. The genetic linkage map consisted of 12 linkage groups, and 248 markers were assigned. These transcriptome and molecular marker data provide information useful for molecular breeding of oriental melon and further comparative studies of the Cucurbitaceae family.

Cytologic Features of Hydatidiform Mole

BACKGROUND Manifestation of a hydatidiform mole in a cervical cytologic specimen is extremely rare. CASE A 52-year-old woman presented with heavy vaginal bleeding. Transvaginal ultrasound scan showed a 2.5 x 2.2 x 2.0-cm highly vascular mass-like lesion, containing multiple cystic areas in the lower part of the uterus and partly extending into the cervix and vagina. Cervical cytology revealed much obscuring fresh blood and low cellularity. Most of the cells were large and pleomorphic with orangeophilic cytoplasms and hyperchromatic nuclei and had been misdiagnosed as squamous cell carcinoma of the uterine cervix. Histologic examination of endometrial curettage revealed a partial hydatidiform mole with involvement of the cervix. CONCLUSION Although correct interpretation of the cytologic findings of a hydatidiform mole is difficult, our careful search revealed 3 types of trophoblastic cells, especially syncytiotrophoblastic cells, making possible the accurate diagnosis of a hydatidiform mole in a cervical specimen, together with the clinical findings. To the best of our knowledge, this is the first case report in the literature of a hydatidiform mole in a cervical cytologic specimen.