Toshihiko Toki

Levels of oxidative stress and redox-related molecules in the placenta in preeclampsia and fetal growth restriction.

Recent evidence suggests that oxidative stress is involved in the pathophysiology of preeclampsia. Using immunohistochemistry and Western blotting, we investigated the oxidative stress- and redox-related molecules, such as 8-hydroxy-2′-deoxyguanosine (8-OHdG), 4-hydroxynonenal (4-HNE), thioredoxin (TRX) and redox factor-1 (ref-1) in the placenta in preeclampsia, intrauterine growth restriction (IUGR), preeclampsia + IUGR and in normal pregnancy. Using immunohistochemistry, the level of 8-OHdG was significantly higher in IUGR (P=0.012) or preeclampsia + IUGR (P=0.0021) than in normal pregnancy, while TRX expression was significantly higher in preeclampsia (P=0.045), and ref-1 expression was significantly higher in preeclampsia (P=0.017), IUGR (P=0.016) and preeclampsia + IUGR (P=0.0038) than in normal pregnancy. The levels of 4-HNE did not differ significantly between either preeclampsia or IUGR and normal pregnancy. A significant positive correlation was observed between TRX and ref-1 expressions in both normal (ρ=0.52) and complicated (ρ=0.43) pregnancies. Using Western blotting, ref-1 expression tended to be higher in complicated pregnancies than in normal pregnancy (P=0.09). These results suggest that oxidative DNA damage is increased in IUGR and that redox function is enhanced in both preeclampsia and IUGR compared with normal pregnancy.

CD10 is a marker for normal and neoplastic endometrial stromal cells.

Using the immunohistochemical technique, we investigated the expression of CD10 in normal female genital tissues, chorionic villi and decidua of early gestation, endometriotic lesions, and uterine mesenchymal tumors. The cytoplasm of normal endometrial stromal cells was consistently positive for CD10. During early gestation, decidualized endometrial stromal cells were negative or only focally positive for CD10, whereas nondecidualized stromal cells were diffusely positive. Syncytiotrophoblast was positive for CD10 on the apical surface, whereas chorionic mesenchymal cells were diffusely positive within the cytoplasm. Cytotrophoblast and intermediate trophoblast were negative for CD10. Groups of stromal cells surrounding cervical glands were often positive for CD10. Myometrium, endometrial and cervical glands, cervical squamous epithelia, and tubal epithelia and stroma exhibited no reactivity for CD10. In endometriosis and adenomyosis, ectopic endometrial stromal cells were usually positive for CD10. Endometrial stromal tumors, including undifferentiated uterine sarcomas, mostly showed diffuse immunoreactivity for CD10. Leiomyomas and leiomyosarcomas were negative or focally (< 5% of cells staining) positive (8/12 leiomyomas and 4/8 leiomyosarcomas) for CD10, except for 1 myxoid leiomyosarcoma that showed CD10 staining in the myxoid areas. These data suggest that diffuse CD10 staining is characteristic of normal and neoplastic endometrial stromal cells, unless they are decidualized.

A new view of the so-called adenoma malignum of the uterine cervix

Abstract Adenoma malignum of the uterine cervix (mucinous type of minimal deviation adenocarcinoma, mucinous MDA), is a unique neoplasm that is difficult to diagnose owing to the deceptively benign appearance of the tumour cells. The present study was undertaken to explore the phenotypic expression of this tumour compared with those of non-neoplastic cervical tissues and of cervical carcinomas of various types. Ten cases of mucinous MDA, 50 cases with non-neoplastic cervical tissues, 13 of cervical adenocarcinoma including the mucinous (endocervical or intestinal type) and endometrioid types, and 2 of mucoepidermoid carcinoma were examined by various histochemical staining methods, including those for gastric mucins, pepsinogen, lysozyme, chromogranin A and carcinoembryonic antigen. The results revealed that mucinous MDA characteristically exhibited gastric phenotypes. The presence of gastric metaplasia was also demonstrated in 9 cases of mucinous MDA and in 5 of the other cases examined. The 7 endocervical-type adenocarcinomas also included 4 that expressed gastric phenotypes, and 2 of the 3 intestinal-type adenocarcinomas showed the same properties focally. These results indicate the presence of a group of lesions expressing gastric phenotypes in the uterine cervix and suggest a close relationship between these lesions. Cervical adenocarcinomas expressing gastric phenotypes are probably derived from MDA.

Mutations in the STK11 gene characterize minimal deviation adenocarcinoma of the uterine cervix.

Minimal deviation adenocarcinoma (MDA) is a well-differentiated variant of mucinous adenocarcinoma of the uterine cervix and is found relatively infrequently in the general population. However, MDA is strongly associated with Peutz-Jeghers syndrome (PJS), a rare hereditary autosomal disorder characterized by benign hamartomatous polyposis in the gastrointestinal tract and mucocutaneous pigmentation. A serine threonine kinase gene, STK11, has been identified as the tumor suppressor gene responsible for the PJS. In this study we investigated the possible direct role of STK11 in the development of MDA of the uterine cervix. Eleven rare cases of mucinous MDA, not known to be associated with PJS, were screened for the presence of mutations in the STK11 gene by single-strand conformation polymorphism analysis of PCR-amplified DNA fragments. Subsequently our findings were confirmed with cloning and sequencing. As a control, 24 cases of endocervical adenocarcinomas of other histologic subtypes, with no family history of PJS (19 mucinous adenocarcinomas, 4 endometrioid adenocarcinomas, and 1 clear cell adenocarcinoma), 15 cases of squamous cell carcinomas of the uterine cervix, 5 cases of endocervical glands with pyloric gland metaplasia, and 2 deeply situated nabothian cysts were investigated. Somatic mutations of the STK11 gene were confirmed in 6 (55%) of the 11 mucinous MDAs and 1 (5%) of the 19 mucinous adenocarcinomas, but not in the 5 nonmucinous adenocarcinomas, the 15 squamous cell carcinomas, nor the 5 endocervical glands with gastric metaplasia. MDAs with the STK11 mutation had a significantly poorer prognosis than MDAs without the STK11 mutation (p = 0.039). A germline mutation of STK11 was detected in one PJS patient with mucinous adenocarcinoma of the uterine cervix. These results suggest that mutations in the STK11 gene may play an important role in the etiology of MDA of the uterine cervix and may distinguish this rare tumor from other common types of adenocarcinoma of the uterine cervix.

A comparative study of pre-operative procedures to assess cervical invasion by endometrial carcinoma

Objective To compare the accuracy of different diagnostic procedures currently used to assess cervical involvement in endometrial carcinoma.

Minimal deviation adenocarcinoma of the uterine cervix has abnormal expression of sex steroid receptors, CA125, and gastric mucin

SummaryTo provide clues to the histological differentiation between minimal deviation adenocarcinoma (MDA) of the uterine cervix and normal cervical glands, we analyzed the histochemical expression of ovarian steroid receptors [estrogen receptor (ER) and progesterone receptor (PR)], mucosubstances such as gastric mucin, CA125, and carcinoembryonic antigen (CEA) in normal cervical glands (10 cases) and MDA (seven cases). Mucin histochemistry showed that gastric mucin was focally demonstrated in all the cases of MDA but in none of the normal cervical glands. ER and PR were not expressed in MDA, whereas both receptors were invariably expressed in normal cervices. Expression of CA125 was significantly decreased in MDA, but was diffusely positive in normal endocervical glands. CEA was focally positive in all cases of MDA, but it was consistently negative in normal cervical glands. These results suggest that MDA lacks expression of the characteristic müllerian-type substances such as ER, PR, and CA125, and that a proportion of its cells contain gastric epithelial substances, comprising gastric mucin and CEA. In conclusion, loss of the expression of ER and PR, decreased expression of CA125, and staining for gastric mucin and CEA could be used for histologic discrimination of MDA from benign cervical glands.

Enhancement of antitumor effect of bleomycin by low‐voltage in vivo electroporation: A study of human uterine leiomyosarcomas in nude mice

Uterine leiomyosarcoma is an extremely malignant neoplasm with high rates of distant metastasis, and systemic chemotherapy is not particularly effective. Thus, the introduction of more active anticancer agents, or of a new drug delivery system, is urgently needed. Recently, electrochemotherapy has been introduced as a way of enhancing the cytotoxic effects of chemotherapeutic agents. This involves administering the drug in combination with electric pulses (which permeabilize tumor cell membranes and allow the drug to enter the cells). In particular, bleomycin (BLM) cannot cross the plasma membrane efficiently, but its cytotoxicity can be enhanced by electropermeabilization. The aim of the present study was to investigate the effect of low‐voltage electroporation (EP) in combination with local BLM injection on the growth of uterine leiomyosarcoma in nude mice. Human uterine leiomyosarcoma cells (SK‐LMS‐1) were implanted subcutaneously into nude mice. Tumor growth in mice treated with EP (100 V/cm) plus BLM was compared with that in mice receiving BLM alone, EP alone, or no treatment (controls). Tissue BLM concentrations and histological analysis (including mitotic counts) were evaluated in tumor tissues. There was a significant reduction in tumor growth in mice that received EP with BLM. One hour after the treatment, the local BLM concentration was 10 times higher in the tumors that received EP with BLM than in those receiving only BLM. Moreover, the mitotic count was lower in the tumors that received EP plus BLM than in the controls. These results demonstrate the possible therapeutic value of low‐voltage EP with BLM in human uterine leiomyosarcoma. Int. J. Cancer 88:640–644, 2000.

Immunohistochemical analysis of cell cycle regulatory gene products in normal trophoblast and placental site trophoblastic tumor

SummaryIntermediate trophoblast (IT) rarely gives rise to a placental site trophoblastic tumor (PSTT). To examine the different growth mechanisms present in normal and neoplastic IT, the expression of cell cycle regulatory molecules was compared at normal implantation sites and in PSTTs. Normal implantation sites in early gestation (19 patients) and PSTTs (6 patients) were immunohistochemically studied using antibodies against cytokeratin, human chorionic gonadotropin, and human placental lac-togen to identify IT, and antibodies against Ki-67, cyclins (A, B, D1, and E), cyclin-dependent kinases (cdks), and p53 to investigate the proliferative activity of the trophoblast. Marked proliferative activity was observed in the trophoblast of the cell columns. Normal IT exhibited a very low labeling index for Ki-67, with negative expression for cdks and cyclins, except for cyclins B and E. The tumor cells of PSTT exhibited a high labeling index for Ki-67 with positive expression for all the cyclins and cdks examined. Expression of p53 was identified in tumor cells of PSTTs and the distribution of p53-positive cells correlated topographically with that of the cyclin A-positive cells. The transformed IT of PSTT has high proliferative activity with an abnormal expression of cell cycle regulatory molecules, which is not observed in normal IT.

Epithelioid trophoblastic tumor metastatic to the vagina: an immunohistochemical and ultrastructural study.

We describe an epithelioid trophoblastic tumor (ETT) metastatic to the vagina in a 30-year-old Japanese woman. A polypoid tumor in the vaginal orifice was composed of nests of intermediate trophoblastic cells that showed a striking epithelioid appearance. In the hysterectomy specimen, a tumor infiltrated through the myometrium and showed histologic findings similar to those of the vaginal tumor. The tumor cells were positive for cytokeratin, inhibin-alpha, and melanoma cell adhesion molecule (Mel-CAM, CD146) but were only focally positive for human placental lactogen. Electron microscopic examination revealed bundles of well-developed, intermediate-type filaments surrounding the nuclei.

Infrequent occurrence of high-risk human papillomavirus and of p53 mutation in minimal deviation adenocarcinoma of the cervix.

To assess the occurrence of high-risk human papillomavirus (HPV) infection and p53 alterations in minimal deviation adenocarcinoma (MDA) of the uterine cervix, paraffin sections were used to investigate the presence of HPVs 16 and 18 and p53 expression and mutation in six cases of MDA. By polymerase chain reaction, only one case was positive for HPV 16 and none was positive for HPV 18. By in situ polymerase chain reaction in the case positive for HPV 16, HPV 16 was detected in a low-grade squamous intraepithelial lesion overlying the MDA but not in the MDA itself. All of the MDAs were negative or only focally positive for p53 by immunohistochemistry. Our polymerase chain reaction DNA sequencing study failed to detect p53 mutation in exons 5 to 8 in those cases focally positive for p53 expression. These results suggest that MDA may be associated only occasionally with the high-risk HPVs or with p53 gene alterations.