Hirofumi Nonogaki

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.

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.

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.

Estrogen receptor localization in normal and neoplastic epithelium of the uterine cervix

To investigate the estrogen receptor (ER) status of cells during carcinogenesis of the uterine cervix, the immunohistochemical reactivity for a monoclonal anti‐ER antibody (H 222) was studied in 26 normal cervical specimens, 21 cases of cervical intraepithelial neoplasia (CIN), and 21 cases of invasive cervical carcinoma. in addition, the presence of human papillomavirus (HPV) DNA (types 6/11, 16/18, or 31/33/35) was analyzed by in situ hybridization. in the normal cervix, basal cells of the squamous epithelium, metaplastic cells, and endocervical glandular cells were ER positive. in contrast, neoplastic cells of CIN (17 of 21 cases) and invasive carcinoma (19 of 21 cases) were ER negative. the remaining four cases of CIN and two cases of invasive carcinoma were focally ER positive. the HPV DNA analysis revealed that HPV DNA in ER‐negative cases was either types 16/18 or undetectable, but all ER‐positive neoplasms contained HPV DNA types 31/33/35. These results suggest that most neoplastic cells in CIN and invasive cervical carcinoma lose their ER expression and that this may be related to the HPV DNA types which they possess.

Immunohistochemical localizations of CA 125, carcinoembryonic antigen, and CA 19‐9 in normal and neoplastic glandular cells of the uterine cervix

Immunohistochemical reactivities with CA 125, carcinoembryonic antigen (CEA), and CA 19‐9 were studied in 15 cervixes without histologic abnormalities, ten with microglandular hyperplasia, nine adenocarcinomas in situ, ten adenocarcinomas with an invasion of less than 5 mm in depth, and 25 frankly invasive adenocarcinomas of the uterine cervix. The glandular cells of the normal cervix and microglandular hyperplasia always exhibited reactivity with anti‐CA 125 in the luminal surfaces and the secretory products within the lumina, but there was usually no reactivity with either anti‐CEA or anti‐CA 19‐9. In the neoplastic glandular cells (adenocarcinoma in situ and invasive adenocarcinoma), the reactivity with anti‐CA 125 was negative or was not in the luminal surface, but in the perinuclear region of the cytoplasm as accumulations of atypical coarse granules. In addition, the positive reaction with anti‐CEA and anti‐CA 19‐9 was seen in 78% and 0% of adenocarcinomas in situ, in 80% and 40% of adenocarcinomas less than 5 mm in depth, and in 84% and 56% of frankly invasive adenocarcinomas, respectively. Therefore, neoplastic cervical glandular cells show an unusual staining pattern with anti‐CA 125, which may be associated with impaired production and/or the intracytoplasmic transport of CA 125, and they begin to produce antigens recognized by anti‐CEA and/or anti‐CA 9‐9, which are usually negative in the normal tissue.

CA 125 in the epithelium closely related to the embryonic ectoderm: The periderm and the amnion

The amnion is believed to be derived from either cytotrophoblastic cells or embryonic ectoderm. However, it produces and secretes CA 125, which is considered a differentiation antigen of fetal coelomic epithelium derived from the mesoderm of germ cells. To verify this, the immunohistochemical localization of CA 125 in human fetal tissues (between 7 and 23 weeks of gestation) derived from the ectoderm, endoderm, or mesoderm, and in the fetal membranes and placenta was studied. Among the mesoderm-derived tissues, only the fetal coelomic epithelium-related tissues were positive for anti-CA 125 from 15 weeks of gestation. The endoderm-derived tissues did not react with anti-CA 125. However, among the ectoderm-derived tissues, only the periderm reacted with anti-CA 125 from 7 weeks until it sloughed from the stratum intermedium by 23 weeks of gestation. Among the fetal membranes and placenta, only the amnion reacted with anti-CA 125 from 9 weeks to term. These findings indicate that the amnion and the periderm, both of which constitute the epithelia covering the amniotic cavity, in addition to the fetal coelomic epithelium-related tissues, produce CA 125.

Analysis of the levels of CA125, carcinoembryonic antigen, and CA19-9 in the cervical mucus for a detection of cervical adenocarcinoma

To verify whether analysis of the levels of CA125, carcinoembryonic antigen (CEA), and CA19‐9 in the cervical mucus is effective for a detection of cervical adenocarcinomas or not, simultaneous measurement of these three tumor markers in the cervical mucus samples from women without gynecologic disorders, with leiomyoma, with cervical squamous cell carcinomas, and with cervical adenocarcinomas was performed. Extremely high levels of CA125 with low levels of both CEA and CA19‐9 were demonstrated in the cervical mucus samples from women without gynecologic disorders and with leiomyoma. The cervical mucus samples from cervical adenocarcinomas showed low CA125 levels with extremely high CEA and/or CA19‐9 levels. Therefore, analysis of the levels of these three tumor markers in the cervical mucus possibly helps in the diagnosis of cervical adenocarcinomas if CEA and/or CA19‐9 show extremely high levels. When a ratio of (CEA + CA19–9)/CA125 was calculated, all women without gynecologic disorders and with leiomyoma showed a ratio <0.5, whereas ten of 11 cases of cervical adenocarcinomas had a ratio ≥0.5. Only one case of microinvasive adenocarcinoma showed a ratio <0.5. Accordingly, the ratio ≥0.5 strongly suggested an existence of cervical adenocarcinoma, although it included some cases of squamous cell carcinomas (four of 17 cases).