Yoshihiko Nanbu

Mitotic activity in uterine leiomyomas during the menstrual cycle.

To investigate the role of the menstrual cycle in the growth of uterine leiomyomas, the mitotic count per 100 high-power fields and the relation of this to the patients age (30 to 54 years) were examined in tissue sections of leiomyomas from 181 surgically removed myomatous uteri. The mean mitotic count in the secretory phase (12.7 per 100 high-power fields) was significantly higher than that of the proliferative phase (3.8 per 100 high-power fields) (p less than 0.01) or menses (8.3 per 100 high-power fields) (p less than 0.05). We found the highest mitotic count (54 per 100 high-power fields) in a leiomyoma at the early secretory phase. In the secretory phase, the younger age group exhibited a significantly higher mitotic count than the older group. Increased mitotic activity in leiomyomas under the hormonal milieu of the secretory phase of the menstrual cycle suggests that the growth of these tumors is affected by progesterone level. In addition, this study defined the range of mitotic counts occurring in the tissue sections of typical leiomyomas during the menstrual cycle.

Immunohistochemical localization of oestrogen receptors and progesterone receptors in the human ovary throughout the menstrual cycle

Immunohistochemistry was used to determine the distribution of oestrogen receptors (ER) and progesterone receptors (PR) in the human ovary during folliculogenesis. Primordial and preantral follicles did not contain ER or PR. The granulosa cells of antral follicles had ER, but negligible PR, before the LH surge. In contrast, at the time of LH surge, these cells of the dominant follicle contained PR, but not ER. On the other hand, granulosa cells of the non-dominant follicles had ER, but not PR. After ovulation, the PR persisted in the luteinized granulosa cells and in the corpus luteum during early pregnancy. The theca interna and surrounding stromal cells were ER-negative and PR-positive throughout the menstrual cycle. Thus, the results show that ER and PR are not expressed simultaneously in the granulosa cells, the thecal cells, or the stromal cells during folliculogenesis. Mechanisms controlling the expression of steroid receptors during the normal menstrual cycle and in early pregnancy are discussed.

Expression and growth-promoting effect of adult t-cell leukemia-derived factor a human thioredoxin homologue in hepatocellular carcinoma

Adult T‐cell leukemia‐derived factor (ADF), originally defined as an interleukin‐2 receptor inducer, is a human thioredoxin homologue. ADF is detected in many malignant tissues and has a growth‐promoting effect on transformed cells. In this study, ADF expression was examined immunohistochemically in human liver cell lines and liver tissues, and its growth‐promoting effect was tested on human hepatoma cells. On three liver cell lines—PLC/PRF/5, HepG2, and Chang liver cells—ADF stained positively and also was detected by immunoblotting. ADF had strong staining in the fetal liver (n = 8), although it was faint in the normal adult liver (n = 6). In hepatocellular carcinoma (n = 25), ADF expression generally was enhanced and was very strong in 52% (13 of 25) of the cases, although it was moderate in cases of chronic hepatitis or cirrhosis. ADF augmented the growth of PLC/PRF/5 cells and showed an additive effect with epidermal growth factor. These results indicate possible involvement of ADF in cell activation and growth of hepatocytes, as is the case with lymphocytes.

Coexpression of adult T-cell leukemia-derived factor, a human thioredoxin homologue, and human papillomavirus DNA in neoplastic cervical squamous epithelium

Adult T‐cell leukemia‐derived factor (ADF) is an autocrine interleukin‐2 receptor‐inducing factor produced by human T‐lymphotropic virus‐1 (HTLV‐1)‐transformed lymphocytes, which has a high structural homology with an endogenous dithiol reducing coenzyme, thioredoxin. Its localization was investigated immunohistochemically in the cervix, using normal tissue (27 samples) and squamous neoplastic tissue (three condylomas, 42 cervical intraepithelial neoplasia [CIN] samples, 34 invasive squamous cell carcinoma samples). The expression of human papillomavirus (HPV) DNA was also studied in serial sections of the same subjects. Normal squamous cells and glandular cells of the cervix were negative for ADF. However, intracytoplasmic and/or intranuclear ADF‐positive cells were usually found in the intermediate and superficial layers of the neoplastic squamous epithelium of condylomas (three of three cases) and CIN (35/42 cases). HPV DNA was detected in all condylomas and in 27 of 42 CIN specimens. HPV DNA‐positive cells were usually localized in the intermediate and superficial layers of the neoplastic squamous epithelium. These HPV DNA‐positive cells were also positive for ADF. Invasive squamous cell carcinoma was also positive for ADF (24/34 cases) and HPV DNA (11/34 cases). The coexpression of HPV DNA and ADF was observed in all HPV DNA‐positive cases. Coexistence of HPV DNA and ADF immunopositivity in neoplastic squamous cells of the cervix suggests that ADF expression closely reflects the intracellular event on HPV DNA replication.

Immunohistochemical analysis of oestrogen receptors, progesterone receptors and Ki-67 in leiomyoma and myometrium during the menstrual cycle and pregnancy

Immunohistochemical distribution of oestrogen receptors (ER), progesterone receptors (PR), and the cell proliferation-associated antigen Ki-67 was investigated in leiomyomas and the myometrium during the menstrual cycle and pregnancy. In the myometrium, ER expression was observed in the proliferative phase, but was suppressed in the secretory phase and during pregnancy. In leiomyomas, ER expression was observed throughout the menstrual cycle, but was suppressed during pregnancy. However, PR was expressed both in the myometrium and leiomyomas throughout the menstrual cycle and pregnancy. In both the myometrium and leiomyomas, a higher number of Ki-67-positive cells was observed during pregnancy than in the secretory phase, and Ki-67 was negative during menopause. The Ki-67-positive cell count in leiomyomas was significantly higher than that in the myometrium throughout the menstrual cycle and pregnancy. Thus both myometrium and leiomyomas have high growth activity under the hormonal milieu of high progesterone levels. The growth potential of leiomyomas is apparently higher than that of myometrium throughout the menstrual cycle and during pregnancy.

Immunohistochemical analysis of estrogen receptors, progesterone receptors, Ki‐67 antigen, and human papillomavirus DNA in normal and neoplastic epithelium of the uterine cervix

To investigate the relationship between the sex steroid receptor (estrogen receptor [ER] and progesterone receptor [PR]) status and the cell proliferation kinetics during the menstrual cycle in normal and neoplastic epithelium of the uterine cervix, immunohistochemical localization of ER, PR, and cell proliferation‐associated antigen, Ki‐67, was investigated in 35 normal cervical specimens, 3 condylomas, 26 cervical intraepithelial neoplasia (CIN) samples, and 22 invasive squamous carcinoma samples. The presence of human papillomavirus (HPV) DNA was also studied. In the normal cervix, basal cells were usually ER positive, PR negative, and Ki‐67 negative throughout the menstrual cycle. Parabasal cells were ER positive and PR negative in the follicular phase, but ER negative and PR positive, and Ki‐67 positive in the luteal phase, and Ki‐67‐positive cells increased in number in the luteal phase. In contrast, PR positivity was observed in the cells of condyloma (2 of 2 cases), CIN (19 of 26 cases), and invasive squamous carcinoma (13 of 22 cases) irrespective of the menstrual phase, Moreover, most neoplastic cells containing HPV DNA type 16/18 were ER negative, whereas several lesions containing HPV DNA type 31/33/35 were weakly ER positive. Many Ki‐67‐labeled cells were observed in the neoplastic lesions. These results suggest that reduced ER expression and increased PR expression are associated with the proliferation of normal cervical squamous epithelium, and this proliferation‐related receptor status, which is probably induced by HPV infection, is usually expressed in neoplastic cervical squamous cells.

Immunohistochemical localization of c-erbB-2 protein and epidermal growth factor receptor in normal surface epithelium, surface inclusion cysts, and common epithelial tumours of the ovary

The c-erbB-2 (HER-2/neu) protein is a membrane glycoprotein growth factor receptor showing molecular homology with the epidermal growth factor receptor (EGFR). We examined the immunohistochemical reactivity of monoclonal antibodies against both of these proteins in normal surface epithelium, surface inclusion cysts, and common epithelial tumours of the ovary. The ovarian tumours were classified as benign (16), borderline malignant (2), and malignant (19). Normal surface ovarian epithelium was weakly positve for both c-erbB-2 protein and EGFR. In surface inclusion cysts, however, the epithelial cells lining the lumen exhibited stronger staining for c-erbB-2 protein, but no staining for EGFR. All 16 benign ovarian tumours and the 2 borderline malignant ovarian tumours were positive for c-erbB-2 protein and negative for EGFR. Of the ovarian carcinomas, 13 of the 19 (68.4%) were positive for c-erbB-2 protein and negative for EGFR, while 4 showed positivity for both c-erbB-2 protein and EGFR. Two cases were negative for both proteins. Expression of both c-erbB-2 protein and EGFR was found in endometrioid carcinoma with squamous differentiation and in clinically advanced poorly differentiated serous carcinomas. Expression of c-erbB-2 protein appears to be increased and that of EGFR is reduced in the early stage of epithelial ovarian oncogenesis. The expression of EGFR with c-erbB-2 protein in ovarian carcinoma is related both to histological differentiation and/or advanced clinical stage.

Immunohistochemical localization of adult T-cell leukaemia-derived factor, a human thioredoxin homologue, in human fetal tissues

An immunohistochemical study of the expression of adult T-cell leukaemia-derived factor (ADF), a human thioredoxin homologue, was performed using a rabbit antibody against the C-terminal peptides of ADF. Tissues were obtained from human fetuses between 9 and 23 weeks of gestation. It was revealed that ADF was widely distributed in different organs and tissues during the fetal period. The ADF antibody reacted selectively with medullary cells of the thymus, lung epithelium, the epithelium of the digestive tract, hepatocytes, bladder epithelium, peripheral nerve cells, hair follicles, sebaceous glands, osteoblasts and the proximal tubules of the kidney. It also reacted with cells destined to differentiate into ciliated cells in the fallopian tube and efferent ductules of the testis, interstitial cells in the ovary, Leydig cells of the testis, and dendritic cells in the spleen and lymph nodes. This is the first report on the thioredoxin system in human cells during the early fetal period. The selectivity of ADF staining in fetal tissues suggests that, during early fetal life, ADF expression correlates well with the cellular function of certain tissues.

Expression of c-erbB-2 protein and epidermal growth factor receptor in normal tissues of the female genital tract and in the placenta

The c-erbB-2 (HER-2/neu) protein is a membrane glycoprotein growth factor receptor that has molecular homology with the epidermal growth factor receptor (EGFR). To investigate the relationship between the expression of c-erbB-2 protein and EGFR in the tissues of the human female genital tract and in the placenta, we examined the immunohistochemical reactivity of monoclonal antibodies against both of these proteins. In the müllerian-derived genital tract, epithelial cells of the fallopian tube, endometrium, and endocervix showed reactivity for c-erbB-2 protein, whereas reactivity for EGFR was distributed mainly in the stromal cells throughout the menstrual cycle and during pregnancy. In addition, the staining intensity for EGFR in the endometrial stroma increased with its decidualization. In the exocervical squamous epithelium, basal cells were cerbB-2 protein-negative and EGFR-positive, but the more differentiated squamous cells of the intermediate layer were c-erbB-2 protein-positive and EGFR-negative. In the placental tissues, cytotrophoblasts and syncytiotrophoblasts of the chorionic villi were c-erbB-2 protein-negative and EGFR-positive. In contrast, intermediate trophoblasts in the extravillous space were c-erbB-2 protein-positive and EGFR-negative. Thus, there is an inverse relationship between the expression of c-erbB-2 protein and EGFR in the tissues of the female genital tract and in the placenta. This suggests that there may be a regulatory mechanism(s) for the expression of both proteins that is associated with the differentiation and/or function of cells in the female genital tract and the placenta.

Molecular interaction during pregnancy: Biochemical and topological analysis of adult T-cell leukaemia-derived factor, homologous to thioredoxin, in the pregnant human uterus

Adult T-cell leukaemia-derived factor (ADF), homologous to thioredoxin, displays various biological activities, such as radical scavenging action and the reduction of protein disulphide bonds. We examined the biochemical and immunohistochemical localization of ADF in the pregnant human uterus, using two heteroantibodies to ADF, antibody C and W. Immunohistochemically, decidua and trophoblast cells were intensely stained by antibody C. The concentration of ADF-like substance in the decidua was 95.9 ng/mg protein, determined by enzyme-linked immunosorbent assay. The molecular weight of ADF-like substance in these tissues was determined by gel electrophoresis to be 13 kDa, the same as that of recombinant ADF. These findings indicate that abundant ADF is present in decidua and trophoblast cells ; the localization of such a potent dithiol reducing substance may be beneficial in protecting the fertilized egg and placental trophoblasts from the cytotoxic effects of oxygen radicals.