Ritsuto Fujiwaki

Expression of Angiopoietin-1, Angiopoietin-2, and Tie2 Genes in Normal Ovary with Corpus luteum and in Ovarian Cancer

Objective: The recent discovery of angiopoietin-1 (Ang1) and angiopoietin-2 (Ang2) has provided novel and important insights into the molecular mechanisms of blood vessel formation. Ang1 and Ang2 bind with similar affinity to the endothelial cell tyrosine kinase receptor Tie2. Our purpose was to assess the potential role of the Ang/Tie2 system in physiological and pathological angiogenesis in the ovary. Methods:Ang1, Ang2, and Tie2 gene expression in 14 normal ovaries with corpus luteum (CL) and in 19 cases of ovarian cancer were analyzed by polymerase chain reaction of RNA after reverse transcription. The level of each gene expression was presented by the relative yield of each gene to the β2-microglobulin gene, respectively. Furthermore, cellular distribution of Ang1 and Ang2 mRNA was examined by in situ hybridization, and localization of Tie2 was studied by immunochemistry. Results: The Ang1, Ang2, and Tie2 gene expression in normal ovary with CL ranged from 0.18 to 1.06 (median 0.54), 0.31–2.64 (median 1.01), and 0.10–0.47 (median 0.20), respectively. The expression of these same genes in ovarian cancer ranged from 0.06 to 0.75 (median 0.14), 0.69–1.59 (median 1.12), and 0.04–0.35 (median 0.15), respectively. Ang1 gene expression in normal ovary with CL was significantly higher than that in ovarian cancer (p = 0.0004). The gene expression levels of Ang2 and Tie2 were statistically the same in both groups. There was a significant correlation between Ang1 gene expression and Tie2 gene expression in normal ovary with CL (r = 0.619, p = 0.018). No such significant correlation was found in ovarian cancer. Moreover, Ang2 gene expression showed no significant correlation with the Tie2 gene expression either in normal ovary with CL or in ovarian cancer. Transcripts for Ang1 were observed in CL cells and endothelial cells around CL, and in tumor cells and endothelial cells at the periphery of tumor invasion. Ang2 transcripts were expressed in the same patterns. Tie2 expression was positive primarily in the endothelial cells around CL and in those at the periphery of tumor invasion. Conclusion: Our results indicate that there is a difference in the Ang/Tie2 gene expression between physiological and pathological angiogenesis in the ovary. This finding may aid in the development of new therapeutic interventions for ovarian cancer.

Gene Expression for Dihydropyrimidine Dehydrogenase and Thymidine Phosphorylase Influences Outcome in Epithelial Ovarian Cancer

PURPOSE Dihydropyrimidine dehydrogenase (DPD) is a pyrimidine salvage enzyme responsible for degradation of thymine, which is produced from thymidine by thymidine phosphorylase (TP). Our purpose was to determine whether DPD affects prognosis in patients with epithelial ovarian cancer and how the two enzymes may interact in such effects. PATIENTS AND METHODS DPD gene expression was analyzed by reverse transcription-polymerase chain reaction in 27 samples from normal ovaries and the 85 epithelial ovarian cancers previously studied with regard to TP gene expression. RESULTS DPD gene expression was significantly lower in epithelial ovarian cancers than in normal ovaries (P: <.0001), whereas TP gene expression and the ratio of TP to DPD gene expression (TP:DPD) were significantly higher in epithelial ovarian cancer (P: <.0001 for both). In patients with epithelial ovarian cancer, DPD gene expression and the TP:DPD ratio did not significantly correlate with any clinicopathologic factors. Patients with a high TP:DPD ratio (higher than the median) had significantly poorer outcomes than those with lower ratios (P: =.0002). The difference in survival between groups with high and low TP:DPD ratios was greater than the difference between groups with high and low TP gene expression. Multivariate analysis showed the TP:DPD ratio to be the independent prognostic factor (P: =.0002). In tumors with high TP gene expression, low DPD gene expression significantly correlated with poor survival (P: =. 04). CONCLUSION Downregulation of DPD gene expression may enhance the negative prognostic effect of high TP gene expression in patients with epithelial ovarian cancer. Certain newly available chemotherapeutic choices may take the TP:DPD ratio into consideration.

Prognostic value of overexpression of p53 in human ovarian carcinoma patients receiving cisplatin

A major obstacle to the treatment of ovarian carcinoma is intrinsic/acquired resistance to cisplatin-based chemotherapy. The clinical significance of p53 overexpression in patients with ovarian carcinoma is still controversial. The aim of this study was to investigate the independent prognostic significance of p53 overexpression in patients with ovarian carcinoma who are treated with cisplatin. We retrospectively examined the overexpression of p53 in primary ovarian carcinoma, and its association with chemotherapeutic efficacy. One hundred and thirty four ovarian carcinomas were surgically removed from patients who received adjuvant cisplatin-based chemotherapy. Immunohistochemical analysis of p53 was performed using a DO7 antibody against the p53 protein in 134 ovarian carcinomas. The significance of p53 in the prognosis of patients with ovarian carcinomas was also examined by a survival analysis of mortality follow-up data covering the period from 1988 to 2001. Thirty-three tumors (25%) exhibited p53 overexpression. Overexpression of p53 in grade 2/grade 3 tumors was significantly higher than that seen in grade 1 tumors (P=0.0088, 0.0229). Patients with tumors who also showed overexpression of p53 had a significantly inferior response to chemotherapy compared with the patients with p53-negative tumors (P=0.04). Cox regression analysis revealed that p53 overexpression was prognostic for poor disease outcome after adjustment for FIGO stage, grade and residual tumor. These findings suggest that overexpression of p53 in ovarian carcinoma is associated with unfavorable clinical outcome in patients treated with cisplatin-based chemotherapy. Therefore, detection of p53 overexpression using the DO7 antibody may be considered as a predictive marker of chemoresistance for cisplatin in patients with ovarian carcinoma.

Clinical significance of interleukin-1 receptor antagonist in patients with cervical carcinoma

OBJECTIVE Our purpose was to determine the clinical significance of interleukin-1 receptor antagonist (IL-1ra), which is an endogenous inhibitor cytokine of IL-1, in patients with cervical carcinoma. METHODS Tissue IL-1ra expression and serum IL-1ra level were examined by enzyme-linked immunosorbent assay (ELISA), Western blotting, and immunohistochemistry in normal controls and patients with cervical carcinoma. RESULTS Tissue IL-1ra protein level by ELISA was significantly higher in squamous cell carcinoma (n = 9) than in the normal cervix (n = 7) and adenocarcinoma (n = 3). Western blotting confirmed the main presence of intracellular IL-1ra type 1 in squamous cell carcinoma. Immunohistochemistry demonstrated significant IL-1ra expression only in tumor cells of squamous cell carcinoma. Elevation of serum IL-1ra level was found in patients with squamous cell carcinoma (n = 38) compared to normal women (n = 13), but not in patients with adenocarcinoma (n = 9). Although serum IL-1ra level did not correlate with clinical stage or any other tumor marker, high serum IL-1ra level was associated with pelvic lymph node metastasis and poor prognosis in patients with squamous cell carcinoma. On the other hand, these results were not obtained in patients with cervical adenocarcinoma. CONCLUSION IL-1ra may play important roles in local and general malignant behaviors in patients with cervical squamous cell carcinoma, and measurement of serum IL-1ra level may be useful in predicting patient survival.

Thymidine kinase in epithelial ovarian cancer: Relationship with the other pyrimidine pathway enzymes

TK is a pyrimidine metabolic pathway enzyme involved in salvage DNA synthesis. What roles TK may play in epithelial ovarian cancer and the relationships between TK and the other pyrimidine pathway enzymes remain unclear. We examined TK1 gene expression by RT‐PCR and related it to gene expression of TS, TP and DPD in 69 samples from epithelial ovarian cancer, 8 low‐malignant‐potential tumors, 16 benign ovarian tumors and 34 normal ovaries. Additionally, cytosolic and serum TK activities were determined by radioenzymatic assay. TK1 gene expression, the ratio of TK1 to TS gene expression, that of TK1 to TP and that of TK1 to DPD were significantly higher in epithelial ovarian cancer than in normal ovaries. In epithelial ovarian cancer, TK1 gene expression correlated with cytosolic and serum TK activities, TS and TP gene expression and the ratio of TP to DPD gene expression. Patients with high‐TK1 gene expression had a significantly poorer survival than those with low TK1 gene expression. Combined analysis demonstrated that the relative risk of cancer death for tumors with high TK1, high TS and high TP gene expression was greater than that for tumors with high TK1 gene expression alone. TK1 gene expression together with TS, TP and DPD gene expression may play important roles in influencing the malignant behavior of epithelial ovarian cancer. Combination therapy including TK inhibitor is a possible therapeutic intervention in patients with epithelial ovarian cancer.

An accurate antenatal diagnosis of vasa previa with transvaginal color Doppler ultrasonography

We describe a case of vasa previa diagnosed antenatally with transvaginal color Doppler ultrasonography. The diagnosis was confirmed at cesarean delivery. The benefits and advantages of the use of transvaginal color Doppler ultrasonography to diagnose vasa previa in utero are discussed.

Elevation of serum interleukin‐1 receptor antagonist levels in women with gynaecological cancers

This study included 15 patients with gynaecological cancers (7 with cervical cancer, 6 with endometrial cancer, and 12 with ovarian cancer); 7 with benign gynaecological disorders (5 with benign ovarian tumour and 2 with uterine myoma); and 10 healthy women as a control group. Serum interleukin‐1 receptor antagonist (IL‐1 ra) levels in patients with gynaecological cancer were significantly higher than those in patients with benign gynaecological disorders (P= 0.04) and in healthy controls (P= 0.0009). IL‐1 ra may play an important role in host immune responses in local and general environments against gynaecological cancers.

Interleukin-4, interleukin-10, and soluble tumor necrosis factor receptors in cord blood.

OBJECTIVE We assessed cord blood for levels of interleukin-4 (IL-4), interleukin-10 (IL-10), and p55 (sTNFR-I) and p75 (sTNFR-II) soluble tumor necrosis factor receptors. STUDY DESIGN Umbilical cord IL-4, IL-10, tumor necrosis factor alpha (TNF alpha), sTNFR-I, and sTNFR-II were measured in 21 normal appropriately grown newborns delivered vaginally (normal pregnancy), and 3 abnormal pregnancies (1 preterm delivery, 1 premature rupture of membranes with chorioamnionitis, and 1 abruptio placentae with fetal and neonatal distress). Umbilical cord arterial blood pH and PO2 were also measured. RESULTS The TNF alpha, sTNFR-I, and sTNFR-II were detectable in all cord blood samples in normal pregnancies. IL-4 was detected in 10 of 21 samples (47.6%), and IL-10 was undetectable in normal pregnancies. IL-10 could be detected in the cases with chorioamnionitis and abruptio placentae. Soluble tumor necrosis factor receptors in the case with preterm delivery and the case with abruptio placentae were elevated compared with the levels in control samples. CONCLUSION Both the p55 and p75 soluble tumor necrosis factor receptors are physiologic constituents of term cord blood. An immunosuppressive role of IL-10 and a protective role of soluble tumor necrosis factor receptors are suggested in abnormal pregnancies. However, in view of the small number of abnormal pregnancies, these observations must be considered preliminary.

lnterleukin-6, lnterleukin-8, and Granulocyte Elastase in Newborns with Fetal Distress

OBJECTIVE Our purpose was to investigate the participation of cytokines and neutrophils in fetal distress. STUDY DESIGN Umbilical cord serum interleukin-6 (IL-6), interleukin-8 (IL-8), and plasma granulocyte elastase (GEL) were measured in 30 normally grown newborns without fetal distress (Group A), 10 growth-retarded newborns without acute fetal distress (Group B), 5 normally grown newborns with fetal distress (Group C), and 5 growth-retarded newborns with fetal distress (Group D). Umbilical arterial blood pH and PO2 were also measured. RESULTS Umbilical arterial blood pH and PO2 in either Group C or Group D were significantly lower than those in either Group A or Group B. The concentration of IL-6 in Group D was significantly higher than that in either Group A, B, or C. The level of IL-8 in either Group C or Group D was significantly higher than that in either Group A or Group B. The concentrations of GEL in Group D was significantly higher than that in either Group A or Group B. CONCLUSION This study suggests that fetal distress in utero causes an elevation of immune factors such as IL-6, IL-8 and GEL.

Cord blood cytokines and soluble adhesion molecules in vaginal and cesarean delivered neonates.

OBJECTIVE Our purpose was to evaluate the effect of labor pain on the concentrations of cytokines and soluble adhesion molecules in cord blood. METHODS Umbilical cord interleukin-6 (IL-6), interleukin-8 (IL-8), tumor necrosis factor-alpha (TNF-alpha), granulocyte elastase (GEL), intercellular adhesion molecule-1 (ICAM-1), and endothelial lymphocyte adhesion molecule-1 (ELAM-1) were measured in 21 normal appropriately grown newborns delivered vaginally (VD group), and 20 normal appropriately grown newborns delivered by elective cesarean section (ECS group). Umbilical cord arterial blood pH and PO2 were also measured. RESULTS Umbilical artery blood pH and PO2 in the VD group were not significantly different from those in the ECS group. There were no significant differences for concentrations of IL-6, IL-8, TNF-alpha, GEL, ICAM-1, and ELAM-1 in cord blood between VD and ECS groups. CONCLUSION These results suggests that labor pains do not affect the concentrations of cytokines and soluble adhesion molecules in cord blood.