Relationship between the angiotensin-converting enzyme genotype and the forearm vasodilator response to estrogen replacement therapy in postmenopausal women.

OBJECTIVES: We sought to evaluate the relationship between the angiotensin-converting enzyme (ACE) genotype and the change in forearm vasoreactivity in response to a three-month course of oral estrogen in postmenopausal women. BACKGROUND: The ACE genotype is a known predictor of the response to an ACE inhibitor drug; however, it is not clear whether it can modify the effect of estrogen replacement therapy (ERT) on endothelial function in postmenopausal women. METHODS: Fifty-five postmenopausal women received 0.625 mg of conjugated equine estrogen daily for three months. Forearm blood flow (FBF) was measured by strain-gauge plethysmography. RESULTS: Twenty-one, 25 and 9 patients had the insertion/deletion (ID), II and DD genotypes, respectively. Plasma ACE activity was significantly higher at baseline in patients with either the DD or ID genotype than in those with the II genotype (p < 0.05). A significant decrease in plasma ACE activity with ERT was seen in the ID and II genotypes (p < 0.05), but not in the DD genotype. There were no significant differences in the FBF responses to reactive hyperemia at baseline between the three groups. Estrogen replacement therapy did not alter the FBF response to reactive hyperemia in the DD genotype (4.0 +/- 1.3%), although ERT significantly increased the FBF response in the ID and II genotypes (32.6 +/- 7.5% and 30.6 +/- 6.5%, respectively; p < 0.05). Forearm blood flow after administration of sublingual nitroglycerin did not change over three months in any of the three groups. CONCLUSIONS: These findings suggest that the effect of ERT in postmenopausal women on forearm endothelial function may be determined in part by the genotype of the ACE gene.

Inhibition of telomerase activity and cell proliferation by a reverse transcriptase inhibitor in gynaecological cancer cell lines.

Telomerase is a ribonucleoprotein which has a RNA template to bind and extend telomere ends, so prolonging the life of tumour cells. The aim of this study was to determine whether transcriptase function of telomerase could be inhibited by the reverse transcriptase inhibitors (RTI); azydothymidine (AZT), dideoxyinosine (ddI) and AZT-5' triphosphate (AZT-TP). We examined their effects on the proliferation of cancer cells and the antitumour effects of cisplatin in vitro. The three agents did not cause major changes in telomerase activity or telomere length in MCAS cells. However, in HEC-1 cells changes in telomerase activity and telomere length were observed that were dependent on the RTI concentration and duration of exposure. ddI and AZT-TP reduced telomerase activity and shortened the length of the telomere. In the presence of RTI, the antitumour effects of cisplatin were enhanced. This was particularly evident in HEC-1 cells where there was a marked reduction in cell proliferation, appearance of morphological changes and senescent-like cells in the presence of ddI or AZT-TP. In MCAS cells, TP53 expression was increased by ddI and AZT-TP, while p21 expression was unchanged. In HEC-1 cells the expression of both TP53 and P21 was increased by ddI. Continuous administration of RTI enhanced the cell growth inhibition of cisplatin. RTI also inhibited the proliferation of some cells.

Macrophage-colony stimulating factor inhibits the growth of human ovarian cancer cells in vitro.

The effect of macrophage-colony stimulating factor (M-CSF), which regulates the growth and differentiation of haematopoietic progenitor cells on the growth of ovarian cancer cells was investigated in three ovarian cancer cell lines in vitro. The spontaneous growth of these cells was significantly inhibited by the addition of M-CSF in a concentration-dependent manner over 96 h of culturing. The maximum response was obtained with 10 ng/ml (3857 U/ml) of M-CSF by counting the viable cell number using the trypan blue exclusion assay. [(3)H]-thymidine incorporation by these cells was also suppressed following a 96-h incubation with M-CSF. The inhibitory effect of M-CSF was reversed by the addition of anti-M-CSF monoclonal antibody. Flow cytometric analysis revealed that the treated ovarian cancer cells arrested at the G0/G1 phase of the cell cycle. These cells expressed M-CSF receptors on their surface as detected by Scatchard plot analysis using (125)I-labelled M-CSF. These results indicate that M-CSF has an antitumour activity for ovarian cancer cells and suggest that it can be applied for the treatment of this disease.

Estrogen replacement therapy in postmenopausal women augments reactive hyperemia in the forearm by reducing angiotensin converting enzyme activity.

The precise mechanism of the vasoprotective effect of estrogen replacement therapy in postmenopausal women is not fully understood. The present study sought to determine the role of nitric oxide (NO) and angiotensin-converting enzyme (ACE) in the vasodilator response of the forearm vessels induced by estrogen administration to postmenopausal women. Subjects were divided into two groups. One group received conjugated equine estrogen (0.625 mg daily) orally for 3 months (n=26), while the other received no treatment (control group, n=10). Forearm blood flow was measured by strain-gauge plethysmography. The concentrations of nitrite/nitrate (metabolites of NO), ACE activity, and lipid parameters were measured. Basal forearm blood flow, body weight, blood pressure, and heart rate were similar at baseline in both groups. After 3 months of estrogen administration, the maximal forearm blood flow response during reactive hyperemia and the serum level of nitrite/nitrate each showed a significant increase over baseline values: from 23.6+/-2.0 to 36.5+/-3.1 ml/min per 100 ml tissue (P<0.01), and from 24.8+/-2.3 to 38.6+/-3.6 micromol/l (P<0.01), respectively. Plasma levels of ACE activity were significantly reduced from baseline after 3 months of estrogen treatment (from 12.2+/-0.6 to 10.9+/-0.6 IU/l, P<0.01). No changes were seen in controls. The change in forearm blood flow after sublingual nitroglycerin was similar at baseline versus after 3 months of estrogen administration. The increase in the serum level of nitrite/nitrate after 3 months of estrogen therapy showed a significant inverse correlation (r=0.52, P<0.01) with the reduction in the plasma level of ACE activity. There was no significant correlation between the increase in serum nitrite/nitrate and any change in serum lipids, blood pressure, or other parameters. The administration of oral estrogen to postmenopausal women for 3 months increased the NO-mediated forearm endothelium-dependent vasodilatation. This was likely due, at least in part, to ACE inhibition. The latter may be one mechanism by which ERT provides its well-known cardiovascular benefit.

Lifespan of human memory T-cells in the absence of T-cell receptor expression.

To evaluate the intrinsic lifespan of human memory T-cells in the absence of T-cell receptor signaling, we used radiation-induced mutant CD4+ T-cells lacking surface expression of TCR/CD3 complex as an in vivo cell marker. We analyzed the long-term kinetics of TCR/CD3 - mutant T-cells among CD4+ CD45RA+ naive and CD4+ CD45RA- memory T-cell fractions in peripheral blood of gynecological cancer patients receiving radiotherapy. Both the proportion and number of these mutant T-cells decayed exponentially with time following radiotherapy. The estimated half-life of mutant memory T-cells was 2 to 3 years and did not differ from that of mutant naive T-cells. These results indicate that the lifespan of mature CD4+ T-cells is limited regardless of their memory or naive phenotype in the absence of TCR/CD3 expression. This finding may suggest that continued T-cell receptor signaling is required for lifetime maintenance of human memory T-cells.

Ring chromosome 6: case report and review.

A ring chromosome 6 was identified in an apparently healthy girl with short stature and microcephaly. Of 100 peripheral lymphocyte metaphases analyzed, chromosome 6 was replaced in 73% by a monocentric ring chromosome, in 10% by a dicentric, in 1% by a tricentric, and 3% by two rings. Thirteen other cells were 45,XX,-6, which may represent 46,XX,r(6)/45,XX,-6 mosaicism. The breakpoints were located on bands p24 or p25 and q26 or q27. Eight other patients with a ring chromosome 6 have been reported. The most characteristic findings in subjects with a ring chromosome 6 are mental retardation and eye and ear abnormalities, none of which were present in our patient.

Decreased superoxide dismutase-2 activity in a patient with ring chromosome 6.

Manganese superoxide dismutase (SOD-2) activity was measured in erythrocytes and leukocytes of a patient with ring chromosome 6. Compared to normal control individuals, SOD-2 activity in leukocytes of the patient was lower, whereas cuprozinc SOD (SOD-1) activity was normal. These findings suggest the cause of decreased enzyme activity is the effect of the ring form on production of the enzyme or inherent instability of ring chromosomes.

Expression of telomerase reverse transcriptase mRNA and its quantitative analysis in human endometrial cancer.

Three major components of telomerase, i.e., human telomerase RNA (hTERC), telomerase-associated protein (TEP1), and the human telomerase reverse transcriptase (hTERT), have been identified. Among them, TERT expression is very closely related to telomerase activity. The purpose of this study was to evaluate the implications of TERT expression and telomerase activity in endometrial cancer. Fresh surgical specimens of 36 endometrial carcinomas (CA group) and 9 samples of postmenopausal endometrial tissue without malignancy (NP group) were obtained at operation in our hospital. These specimens were analyzed for telomerase activity and TERT expression by TRAP assay and RT-PCR, respectively, and the detection and quantitative analysis were made. The results for endometrial cancer were compared with those for normal endometrium and with the clinical data. In the CA group, TERT expression was detected in 35/36 subjects (97.2%), whereas in 1/9 subject (11.1%) from the NP group. Relative TERT mRNA expression was 0.50 in the CA group, and this was significantly higher compared with the level of 0.10 in the NP group (p<0.05). Telomerase activity was detected in 34/36 subjects (94.4%) from the CA group and in 3/9 subjects (33.3%) from the NP group (p<0.05), while the RTA was 30.9 and 0.2, respectively (p<0.05). There was a significant correlation between the relative TERT expression and RTA (n=45, R=0.413, p<0.05). RTA was significantly higher at an advanced surgical stage (FIGO II, III or IV) than at an early stage (FIGO 0 or I) (52.4 vs. 20.4, p<0.05), but other clinical factors showed no relationship with TERT and RTA values. The detection and quantitative analysis of telomerase activity and TERT expression is helpful for distinguishing malignant from normal endometrium when the patient is postmenopausal, even if the tumor is very small or of low malignancy.

p27, cyclin E, and CDK2 expression in normal and cancerous endometrium.

The objective was to investigate the immunohistochemical expression of p27, cyclin E, and CDK2 in normal and cancerous endometrium. Expression of p27 in premenopausal normal endometrium was significantly higher than that in postmenopausal normal endometrium (p=0.019). A significantly lower amount of p27 staining was observed in endometrial cancer tissues from premenopausal women than in normal premenopausal endometrium (p=0.015). Cyclin E expression in premenopausal normal endometrium was significantly higher than that in postmenopausal normal endometrium (p=0.003). A significantly higher amount of cyclin E staining was observed in endometrial cancer tissues from postmenopausal women than in normal postmenopausal endometrium (p=0.017). Regarding menopausal status, no significant difference in CDK2 staining was observed between cancerous and normal endometrium. There was a positive significant correlation between cyclin E and CDK2 expression levels in endometrial cancers (p<0.05). Western blot analysis confirmed elevated p27 protein levels in samples with positive p27 immunostaining. Considerable levels of p27 mRNA were detected in all normal and cancerous samples examined by semi-quantitative PCR. No significant relationship was found between telomerase activity and its association with p27 and cyclin E expression in endometrial cancers. These findings suggested that the decreased expression of p27 caused by post-translational mechanism might play an important role in endometrial cancer development in premenopausal women. In addition, increased cyclin E expression may play an important role in endometrial cancer development in postmenopausal women.

Expression of matrix metalloproteinases (MMP-2, MMP-9, MT1-MMP) and their inhibitors (TIMP-1, TIMP-2) in common epithelial tumors of the ovary.

Matrix metalloproteinases (MMPs) are known to play an important role in cancer cell invasion by mediating the degradation of extracellular matrix proteins. The activity of such MMPs are regulated by tissue inhibitors of metalloproteinases (TIMPs). In this study, we investigated the immunohistochemical expression of MMP-2, MT1-MMP, TIMP-2, MMP-9, and TIMP-1 in 114 epithelial ovarian tumors (14 adenomas, 22 borderline tumors, and 78 adenocarcinomas). mRNA expression of MMP-2, MT1-MMP, and TIMP-2 was determined by RT-PCR in selected samples. The diffuse positive rates of MMP-2, MT1-MMP, TIMP-2, and MMP-9 in ovarian carcinomas were significantly higher than those in the borderline and in benign tumors. Conversely, the diffuse positive rate of TIMP-1 was higher in the benign and borderline ovarian tumors than that in ovarian carcinomas. The percentages of the cases with triple diffuse positive expression for MMP-2, MT1-MMP, and TIMP-2 within the same tumor was significantly higher in malignant tumors than those in borderline and in benign tumors. With respect to clinical stage, the triple diffuse positive rate in advanced-stage (stage II/III/IV) carcinomas was significantly higher than that in early-stage (stage I) carcinomas. A significantly higher triple diffuse positive rate was also observed in high-grade (grade 2/3) disease than in low-grade (grade 1) disease. Considerable levels of mRNA expression of MMP-2, MT1-MMP and TIMP-2 were detected in all selected samples that showed triple diffuse positive immunostaining, confirming the co-expression of MMP-2, MT1-MMP, and TIMP-2 at the transcriptional level within the same tumor. All cases with diffuse positive expression for MMP-9 showed regional or negative TIMP-1 expression. The diffuse positive rate of MMP-9 was significantly higher in ovarian carcinomas with lymph node metastasis than in those without lymph node metastasis. Our results suggest that the overexpression of MMP-2, MT1-MMP, TIMP-2, and MMP-9 and down-regulation of TIMP-1 may contribute to the development or enhanced growth capacity of ovarian tumors. Co-expression of MMP-2, MT1-MMP, and TIMP-2 within the same tumor seems to play an important role in the progression of ovarian cancer. Elevated MMP-9 expression together with low expression of TIMP-1 may also contribute to the lymph node metastasis of ovarian carcinoma cells.

Underexpression of cyclin-dependent kinase inhibitor p27 is associated with poor prognosis in serous ovarian carcinomas.

Reduced expression of a cyclin-dependent kinase inhibitor, p27, has been reported to be associated with poor prognosis in several human cancers. The aim of this study was to investigate the potential role of p27 in ovarian cancer development and progression. Immunohistochemical expression of p27 was determined using 117 epithelial ovarian tumor tissues and 8 normal ovaries. p27 mRNA expression was examined by semi-quantitative PCR amplification using 26 ovarian cancer samples. Nuclear staining of p27 was commonly observed in the normal ovarian surface epithelium and the epithelial cells of germinal inclusion cysts. Positive p27 staining rates were significantly higher in serous adenomas (p=0.006) and in serous LMP tumors (p=0.013) than that in serous carcinomas (Fisher's exact test). In serous ovarian cancers, positive p27 staining rate was significantly higher in early stage (stage1/2) than that in advanced stage (stage 3/4) diseases (p=0.030, Fisher's exact test). Log-rank testing showed that negative p27 expression significantly correlates with poor survival in serous ovarian cancer patients (p=0.041). Considerable levels of p27 mRNA were detected in all ovarian cancer samples examined. These results suggest that the underexpression of p27 caused by post-translational mechanism may contribute to the development and progression and result in poor prognosis of serous ovarian cancers.

Prevalence of uterine myoma detected by ultrasound examination in the atomic bomb survivors.

Benign tumors of several organs have been demonstrated to occur as late effects of atomic bomb exposure, and a recent addition to the list of affected organs is the uterus. The increased incidence of uterine myoma noted in Radiation Effects Research Foundation (RERF) Adult Health Study Report 7 (Wong et al., Radiat, Res. 135, 418-430, 1993), however, was based on self-reported information, optional gynecological examination and patient-requested ultrasound examination. Thus the possibility of dose-related bias in case detection was a serious concern. Therefore, the relationship between the prevalence of uterine myoma and dose to the uterus was examined after excluding as much bias as possible by asking all women who had undergone biennial examinations from December 1991 through December 1993 to undergo ultrasound examinations. Among 2506 female participants in Hiroshima, the uterus was visualized by ultrasound examination in 1190, and 238 were found to have uterine nodules. Multiple logistic analysis using Dosimetry System 1986 uterine doses revealed a significant dose response for the prevalence of uterine nodules. The odds ratio at 1 Gy was 1.61 (95% confidence interval: 1.12-2.31). It is unlikely that the observed relationship after adjusting for bladder filling, volume of the uterus, age and menopause status was the result of dose-related bias. These results support previous findings at RERF and provide further evidence that radiation exposure is one of the factors associated with uterine myoma.

Three-year study of estrogen alone versus combined with progestin in postmenopausal women with or without hypercholesterolemia.

This study compares the effects of long-term hormone replacement therapy on the lipid profile of postmenopausal women with or without hypercholesterolemia, with a comparison of 2 different regimens over a 3-year period. A total of 209 women were enrolled in this prospective, nonrandomized trial. They were classified into 2 groups according to baseline serum levels of total cholesterol and low-density lipoprotein (LDL) cholesterol. The hypercholesterolemic group consisted of 83 subjects with a total cholesterol level of 220 mg/dL or higher and LDL cholesterol 140 mg/dL or higher. The normocholesterolemic group consisted of 126 subjects with normal total and LDL cholesterol levels. Therapy was assigned as follows: 44 subjects in the hypercholesterolemic group and 67 in the normal cholesterol group with a total hysterectomy received conjugated equine estrogen (CEE) 0.625 mg/d, while 39 subjects in the hypercholesterolemic group and 59 in the normal cholesterol group with a physiological menopause received CEE 0.625 mg/d plus medroxyprogesterone acetate 2.5 mg/d. Fasting blood samples were monitored periodically for 3 years. Nine women withdrew from the study. Hormone replacement therapy had a more favorable effect in the hypercholesterolemic group versus the normal cholesterol group by decreasing total and LDL cholesterol, 7.0% and 16.6%, versus the normal cholesterol group, 0.8% and 3.9%. Serum levels of high-density lipoprotein (HDL) cholesterol were increased in both groups (hypercholesterolemic, 14.4%; normal cholesterol group, 26.5%), with the increase being larger in the normal cholesterol group. These changes were similar with both treatments and were maintained over 3 years. Serum levels of triglyceride were also increased in both groups, with the increase being statistically significant only in the group with normal cholesterol levels at baseline. There were no consistently reported side effects of therapy. The effects of postmenopausal hormone replacement therapy, estrogen with or without progestin, on the lipid profile appear to be related to the subject's baseline lipid values. Thus, such therapy may have a more favorable effect on LDL cholesterol in postmenopausal women with hypercholesterolemia, with the beneficial effect being maintained over 3 years.

Cytogenetic and clinicopathologic studies of partial moles.

Fifty-six partial moles from 55 patients were karyotyped and analyzed clinicopathologically. Forty-seven were triploid and nine diploid. Triploid partial moles were characterized histologically by focal mild-to-moderate trophoblastic hyperplasia with stromal trophoblastic inclusion. Diploid partial moles lacked marked trophoblastic hyperplasia and stromal trophoblastic inclusions. Four of the nine diploid moles were analyzed for chromosome heteromorphisms, enzyme polymorphisms, and histocompatibility leukocyte antigen specificities to determine their origin, and two were found to be normal. In the follow-up study, none of the partial moles developed into invasive mole or choriocarcinoma. These results indicate that most partial moles are triploid but a minority are normal diploid; regardless of the karyotype, however, partial moles do not have a high propensity for malignancy.

Precise diagnosis by gene analysis and successful management of delivery in three patients with type IIB von Willebrand disease.

Type IIB is a rare variant of von Willebrand disease (vWD). An affected individual's bleeding tendency and thrombocytopenia are exacerbated with pregnancy, and the proper management of these patients during delivery has not been well established. Since it is important to distinguish this disease from the platelet type vWD in order to administer the appropriate therapy, gene analysis is necessary to make the precise diagnosis. We now report the successful management of delivery in three patients, who were diagnosed as having type IIB vWD by the detection of missense mutations in the von Willebrand factor (vWF) gene (C3916-->T and C3922-->T). These changes cause Arg543-->Trp and Arg545-->Cys substitutions in the A1 domain of vWF. We were able to manage the bleeding tendency of these patients at delivery mainly with vWF concentrates to supply normal vWF.

Increased spontaneous and mitomycin C-induced sister chromatid exchanges in patients with cancer of the cervix uteri, with special reference to stage of cancer.

The frequencies of spontaneous and mitomycin C (MMC)-induced sister chromatid exchange (SCE) were examined in 35 patients with cancer of the cervix uteri (stage 0, eight cases; stage I, nine cases; stage II, nine cases, and stage III, nine cases) before they had undergone cancer treatment, as well as in seven patients with uterine myoma and 18 healthy women as controls. The frequency of SCE was analyzed in reference to the stage of cancer in the cancer group and in reference to chromosome group in the cancer and normal groups. The frequencies of spontaneous and MMC-induced SCE in the cancer group were 10.0 +/- 1.8 and 20.7 +/- 2.6, respectively, and both were significantly higher than in the myoma (8.1 +/- 0.8 and 17.6 +/- 1.8) and normal (7.6 +/- 0.8 and 17.6 +/- 2.3) groups. Furthermore, the frequency of SCE in the cancer group increased with cancer stage. All chromosome groups contributed equally to the increase in SCE in the cancer group. These results indicate that an increase in the frequency of SCE in patients with cervical cancer is related to the presence of cancer, but is not related to a predisposition to cancer.

Detection of chromosomal abnormalities in uterine leiomyoma using conventional cytogenetic method and interphase fluorescence in situ hybridization.

Seventy-nine uterine leiomyomas were examined using a conventional cytogenetic method and fluorescence in situ hybridization (FISH) for detection of chromosomal abnormalities of chromosome 12. Nine (17.6%) of 51 tumor samples examined showed chromosomal abnormalities by conventional cytogenetic analysis. Rearrangements of chromosome 12 were detected in two tumors. Other tumors showed abnormalities affecting chromosomes 1, 4, 6, 7, 10, 13, 14, and 22. For FISH, the whole-chromosome painting probe and the D12Z3 probe specific for the centromeric region were used to detect structural and numerical abnormalities of chromosome 12. Of forty-one tumor samples, six showed structural aberrations and four showed numerical aberrations of chromosome 12 by FISH analysis. Of the tumors with structural aberrations identified by FISH, two had normal karyotypes, two showed structural rearrangements of chromosome 12 cytogenetically, and two could not be analyzed because of an insufficient number of metaphases. There were no correlations between the cytogenetic data and clinical parameters. The results indicate that chromosomal abnormalities are important in the biology of at least some types of uterine leiomyomas, and that FISH is a useful complement to conventional cytogenetic analysis in the study of solid tumors.

Detection of vascular disease risk in women by panoramic radiography.

Low bone mineral density and rapid bone loss of the skeleton are associated with mortality risk from vascular diseases in post-menopausal women. Panoramic radiographic measurements are considered as indicators of skeletal bone mineral density or bone turnover. We hypothesize that such measurements may be associated with vascular disease risk in post-menopausal women. Associations of mandibular cortical shape and width on panoramic radiographs with skeletal bone mineral density and risk factors related to vascular diseases were investigated in 87 post-menopausal women. Cortical shape was associated with skeletal bone mineral density, low-density lipoprotein cholesterol, apolipoprotein B, resting heart rate, and endothelial dysfunction. Cortical width was associated with skeletal bone mineral density, low-density lipoprotein cholesterol, and apolipoprotein A1. Dentists may be able to refer women with increased risk of vascular diseases, as well as low bone mineral density, to medical professionals for further examination by panoramic findings.

Fetal diagnosis of galactosialidosis (protective protein/cathepsin A deficiency).

The fetal diagnosis of galactosialidosis is performed by measuring carboxypeptidase (cathepsin A) activity in cultured villous cells and by immunofluorescence analysis with an antibody against an oligopeptide corresponding to the N-terminal domain of the human mature protective protein. Neither carboxypeptidase activity nor immunofluorescence was detected in cultured villous cells derived from an at-risk fetus or in cultured fibroblasts derived from the sister with galactosialidosis. Neuraminidase and beta-galactosidase activities were also confirmed to be deficient or low. A direct assay system for protective protein/cathepsin A is useful for the accurate prenatal diagnosis of galactosialidosis.

Cyclin D1 overexpression and p53 mutation status in epithelial ovarian cancer.

OBJECTIVE: To examine the cyclin D1 mRNA expression level in ovarian tumor samples as compared with normal ovaries and to determine the relationship between cyclin D1 overexpression and p53 mutation status in ovarian tumors. METHODS: mRNA was isolated and cDNA was prepared from 27 epithelial ovarian tumors (3 tumors of low malignant potential (LMP) and 24 cancers) and 6 normal ovaries. The cyclin D1 sequences were amplified by using a thermal cycler in parallel with the beta-tubulin gene as an internal control. The cyclin D1 mRNA expression level relative to beta-tubulin was determined by 32P phosphoimager analysis. To confirm the overexpression of the cyclin D1 protein in ovarian tumor cells, immunostaining was performed. The p53 gene mutation status was examined by direct cDNA sequencing. RESULTS: mRNA levels of cyclin D1 were significantly higher in 21 (78%) of the 27 ovarian tumors than in normal ovaries. Cyclin D1 overexpression was detected in ovarian LMP tumors as well as in ovarian cancer cases. Positive immunostaining of cyclin D1 protein was observed in 10 of 18 (56%) ovarian tumors examined. p53 mutations were found in 11 (61%) of 18 ovarian tumors. Of 11 ovarian tumor cases with p53 mutations, 5 showed overexpression of cyclin D1. All 7 ovarian tumor cases without p53 mutations showed significant cyclin D1 mRNA overexpression. CONCLUSION: Cyclin D1 overexpression seems to be an early genetic event in ovarian tumor development. Although p53 may be one of the proteins whose function regulates the expression of G1 cyclins, ovarian tumors with no p53 mutation consistently showed cyclin D1 overexpression. Cyclin D1 overexpression may play an important role in the tumorigenesis of epithelial ovarian tumors.