Clinical applications of analysis of plasma circulating complete hydatidiform mole pregnancy-associated miRNAs in gestational trophoblastic neoplasia: a preliminary investigation.

The aim of this study was to investigate the clinical application of plasma complete hydatidiform mole pregnancy-associated microRNAs (CHM-miRNAs: hsa-miR-520b, hsa-miR-520f and hsa-miR-520c-3p). We measured plasma CHM-miRNA concentration by real-time quantitative reverse transcriptase polymerase chain reaction in two cases of CHM resulting in gestational trophoblastic neoplasia later. As progress of treatments in both cases, the plasma concentrations of CHM-miRNAs showed a decreasing tendency similar to the pattern for serum hCG concentration, but exhibited a transient increasing tendency after each course of chemotherapy, suggesting that the plasma CHM-miRNAs could be an additional follow-up marker for malignant changes of CHM.

Mutant p53 gain-of-function induces epithelial-mesenchymal transition through modulation of the miR-130b-ZEB1 axis.

The tumor suppressor gene p53 has been implicated in the regulation of epithelial-mesenchymal transition (EMT) and tumor metastasis by regulating microRNA (miRNA) expression. Here, we report that mutant p53 exerts oncogenic functions and promotes EMT in endometrial cancer (EC) by directly binding to the promoter of miR-130b (a negative regulator of ZEB1) and inhibiting its transcription. We transduced p53 mutants into p53-null EC cells, profiled the miRNA expression by miRNA microarray and identified miR-130b as a potential target of mutant p53. Ectopic expression of p53 mutants repressed the expression of miR-130b and triggered ZEB1-dependent EMT and cancer cell invasion. Loss of an endogenous p53 mutation increased the expression of miR-130b, which resulted in reduced ZEB1 expression and attenuation of the EMT phenotype. Furthermore, re-expression of miR-130b suppressed mutant p53-induced EMT and ZEB1 expression. Importantly, the expression of miR-130 was significantly reduced in EC tissues, and patients with higher expression levels of miR-130b survived longer. These data provide a novel understanding of the roles of p53 gain-of-function mutations in accelerating tumor progression and metastasis through modulation of the miR-130b-ZEB1 axis.

Study of edema reduction patterns during the treatment phase of complex decongestive physiotherapy for extremity lymphedema.

Shortening the treatment phase of complex decongestive physiotherapy (CDP) is extremely important both for individual patients and medical economics. In 83 patients with stage II unilateral secondary extremity lymphedema (31 upper extremities and 52 lower extremities), the daily changes in the volume of affected extremities during the treatment phase of CDP were prospectively investigated. For the upper extremity lymphedemas, the biggest change was seen between days 1 (100% residual edema rate) and 2 (46.0 +/- 2.7%; mean +/- SD) of therapy with a 54.0% reduction (p < 0.0001). Between days 2 and 3 (38.0 +/- 2.6%) of therapy, there was an 8.0% reduction (p < 0.05). From days 3 to 6 of therapy, slight changes ranging from 0.2 to 3.2%/day were seen. For the lower extremity lymphedemas, the biggest change was seen between days 1 (100%) and 2 (44.5 +/- 2.1%) of therapy with a 55.5% reduction (p < 0.0001). Between days 2 and 3 (33.5 +/- 2.6%) of therapy, there was an 11.0% reduction (p < 0.001). The daily volume changes from days 4 to 6 were slight, ranging from 0.1 to 1.0%/day. During the treatment phase of CDP, the largest volume changes were seen soon after the start of therapy.

High-grade broad ligament cancer of müllerian origin: immunohistochemical analysis of a case and review of the literature.

Malignant neoplasms arising in the broad ligament are quite unusual. Although tens of broad ligament cancers of low potential malignancy have been documented, there have been only two reports of high-grade cases. Furthermore, it remains controversial whether broad ligament tumors are mesonephric (wolffian) or paramesonephric (müllerian) in origin. This case study reports a 49-year-old nulliparous woman who, during her follow-up for breast cancer, demonstrated a 39- x 32-mm solid and poorly differentiated adenocarcinoma within the broad ligament of the right adnexa. Because of its rarity and poor differentiation, immunohistochemical analysis was performed to identify the primary malignancy and determine its origin. Positive expression of WT1, CA125, and ER and negative expression of GCDFP-15, HER-2, and PgR in broad ligament cancer, in conjunction with positive expression of GCDFP-15 and ER and negative expression of WT1, HER-2 and PgR in the previous breast cancer, suggested broad ligament cancer to be a gynecological primary malignancy with serous characteristics rather than a metastasis from the previous breast cancer. In addition, EMA-positive and CK7-negative expression indicated a müllerian origin. Broad ligament cancers of müllerian origin occur predominantly in women of reproductive age, most of whom are infertile or nulliparous. This feature, together with the positive expression of ER in this case, may raise the possible hormonal influence on the etiology of broad ligament cancer of müllerian origin.

A study of risk factors for ovarian metastases in stage Ib-IIIb cervical carcinoma and analysis of ovarian function after a transposition.

OBJECTIVE: The objective of the present study was to examine the incidence and risk factors of ovarian metastases in cervical carcinoma. The function of transposed ovaries was also studied. METHODS: In order to analyze the risk factors of ovarian metastases, 255 slides of pathological specimens were reassessed by multivariate logistic regression analysis. Fifty-six patients were studied prospectively on the basis of the function of transposed ovaries. Basal body temperature and serum hormone levels were analyzed. RESULTS: Ovarian metastasis was identified in 2 of 485 (0.4%) patients with squamous cell carcinoma and in 12 of 146 (8.2%) patients with nonsquamous tumors of the cervix. Histologic type (P = 0.0014) and blood vessel invasion (P = 0.0433) were significant independent risk factors for ovarian metastases, as revealed by multivariate logistic regression analysis. Cumulative survival curves of preserved ovaries showed a significant (P < 0.005) decline in the group with postoperative radiotherapy. CONCLUSION: Preservation of ovarian function should be pursued in patients with squamous cell carcinoma of the cervix, provided that the patient has no other risk factor (blood vessel invasion) for ovarian metastases. Moreover, sufficient attention should be paid to the proper handling of ovarian blood vessels during surgery, in order to shield and protect them from exposure to scattered radiation administered during postoperative radiotherapy.

Prognostic significance of combined conventional and immunocytochemical cytology for peritoneal washings in endometrial carcinoma.

BACKGROUND: Noncancerous cells simulating adenocarcinoma cells may interfere with the analysis of peritoneal cytology (PC) in patients with endometrial carcinoma. Immunocytochemistry (ICC) may improve the diagnosis of PC. METHODS: PC slides from 115 patients with endometrial carcinoma were reviewed. Suspicious or positive cell clusters were recovered with a cell transfer method and were subjected to ICC for MOC-31, cytokeratin 5/6, and p53. Conventional Papanicolaou staining and ICC results were compared directly on the same cells. RESULTS: By combined conventional and immunocytochemical PC (CONV-ICC-PC), cytodiagnosis was positive in 18 of 115 patients (15.7%) and suspicious in 3 of 115 patients (2.6%). According to a multivariate Cox regression analysis of patients with tumors confined to the uterus that included grade, myometrial invasion, cervical involvement, and CONV-ICC-PC, only CONV-ICC-PC was an independent prognostic factor (P < 0.05). A multivariate analysis for all of the patients studied that compared CONV-ICC-PC with staging variables revealed that only peritoneal metastasis (P < 0.0001) and lymph node metastasis (P < 0.01) were independent prognostic factors. When peritoneal metastases were excluded, CONV-ICC-PC (P < 0.01) and lymph node metastasis (P < 0.0025) were the independent prognostic factors. By cell transfer and p53 immunostaining in samples from 14 patients with malignant cells in their peritoneal washings, no deaths occurred among 5 patients with negative p53, whereas 5 of 9 patients with positive p53 died of disease at the time of data analysis. CONCLUSIONS: MOC-31 immunostaining improves the diagnosis of PC in endometrial carcinoma. Positive PC is an important prognostic factor for patients with endometrial carcinoma confined to the uterus. The p53 positive cells in PC have possible prognostic significance.

Prevention of lymphocyst formation following systematic lymphadenectomy.

BACKGROUND: The occurrence of pelvic lymphocysts is an important complication following systematic lymphadenectomy for gynecological malignancies. We employed a procedure to prevent vaginal shortening following radical hysterectomy and we examined whether this procedure could be effective in preventing pelvic lymphocyst formation. METHODS: We studied the incidence of lymphocysts in 190 patients with 84 cervical cancers, 74 endometrial cancers and 32 ovarian cancers, using computed tomographic examination at 3 and 6 months subsequent to the surgery. The surgery included radical hysterectomy and a procedure to prevent vaginal shortening (101), modified radical hysterectomy (79) and simple hysterectomy (7), with systematic lymphadenectomy. RESULTS: There was a significant difference in the incidence of pelvic lymphocysts between cervical cancer (4.8%) and ovarian cancer (18.8%). The postoperative incidence of lymphocyst formation in patients undergoing radical hysterectomy with the procedure to prevent vaginal shortening (5.9%) was significantly lower than in those who underwent modified radical hysterectomy (15.2%). CONCLUSION: Our procedure to prevent vaginal shortening could be effective in preventing not only the shortening of the vagina but also the occurrence of pelvic lymphocysts in patients undergoing radical hysterectomy with systematic lymphadenectomy for gynecological malignancies.

Induction of transformation and p53-dependent apoptosis by adenovirus type 5 E4orf6/7 cDNA.

Adenovirus (Ad) E4orf6/7, one of the early gene products of human Ads, forms a stable complex with the cellular transcription factor E2F to activate transcription from the Ad E2 promoter. E2F cDNAs have growth-promoting and apoptosis-inducing activities when overexpressed in cells. We cloned Ad5 E4orf6/7 cDNA in both simian virus 40- and human cytomegalovirus-based expression vectors to examine its transforming and apoptotic activities. The cloned E4orf6/7 collaborated with a retinoblastoma protein (RB)-nonbinding and therefore E2F-nonreleasing mutant of Ad5 E1A (dl922/947) to morphologically transform primary rat cells, suggesting that E2F is an important cellular protein functioning downstream of E1A for transformation. In a G418 colony formation assay, E4orf6/7 was shown to suppress growth of untransformed rat cells. Moreover, a recombinant Ad expressing Ad5 E4orf6/7 induced apoptosis in rat cells when coinfected with wild-type p53-expressing Ad. Mutational analysis of E4orf6/7 revealed that both of the domains required for growth inhibition and transformation by E4orf6/7 lay in the C-terminal region, which is essential for transactivation from the upstream sequence of an E2a promoter containing E2F-binding sites. However, the smallest mutant of E4orf6/7, encoding the C-terminal 59 amino acids, failed to complement the RB-nonbinding dl922/947 mutant despite showing growth inhibition and E2F transactivation activities. Thus, it is suggested that a subregion of E4orf6/7 which is required for growth inhibition and transformation in collaboration with dl922/947 overlaps the transactivation domain of E4orf6/7.

Induction of apoptosis by the p53-273L (Arg --> Leu) mutant in HSC3 cells without transactivation of p21Waf1/Cip1/Sdi1 and bax.

Codon 273 is one of the hot spots of missense mutation of the p53 tumor suppressor gene found in human cancers. We have previously reported that a mutation at codon 273, p53-273L (Arg --> Leu), suppresses cell growth despite its having no p53-specific transactivation activity. To further elucidate the mechanism of growth suppression caused by p53-273L, we used squamous cell carcinoma cell line HSC3 to isolate subclones containing Zn2+-inducible wild-type (wt) p53, p53-175H, and p53-273L. Northern blot hybridization of the HSC3 cells possessing an inducible function of p53 as well as a luciferase assay for the p21Waf1/Cip1/Sdi1 promoter showed that only wt p53 could induce p21Waf1/Cip1/Sdi1 transcription. Meanwhile, the expression of bax remained unchanged between, before, and after the induction of any analyzed p53s. When wt p53 was induced in HSC3 cells cultured in medium containing 5% fetal bovine serum, cell growth was suppressed through G1 arrest. On the other hand, in medium with 0.1% fetal bovine serum, the growth of HSC3 cells expressing p53-273L was suppressed to a greater degree than that of cells expressing wt p53. Flow cytometric analysis and DNA ladder formation revealed that, unlike wt p53-SN3- and p53-175H-expressing HSC3 cells, p53-273L-expressing cells contained a larger sub-G1 fraction under this culture condition. These findings suggest that p53-273L can induce apoptosis in HSC3 cells without transactivation of p21Waf1/Cip1/Sdi1 and bax.

Sonodynamically induced cell damage with 4'-O-tetrahydropyranyladriamycin, THP.

BACKGROUND: 4'-O-tetrahydropyranyladriamycin (THP) is a novel anthracycline derivative with cardiotoxicity significantly lower than ADM. In this study, the ultrasonically induced in vitro cell damaging effect of THP was investigated. MATERIALS AND METHODS: Sarcoma 180 cells suspended in air-saturated PBS were exposed to ultrasound for up to 60 s in the presence and absence of THP. The viability of the isolated cells was determined by staining of the cells with Trypan Blue dye. RESULTS: The rate of inducing cell damage with ultrasound was doubled with 80 microM THP, while no cell damage was observed with THP alone. CONCLUSION: The enhancement of ultrasonically induced in vitro cell damage was demonstrated with THP. This enhancement was significantly inhibited by histidine, which may suggest a sonochemical mechanism.

Passive immune thrombocytopenia in neonates of mothers with idiopathic thrombocytopenic purpura: incidence and risk factors.

The aim of this study was to evaluate risk factors for occurrence of neonatal passive immune thrombocytopenia (PIT) in pregnancy complicated by idiopathic thrombocytopenic purpura (ITP). We studied 63 pregnant women with ITP and the 66 neonates retrospectively. Neonatal platelet counts were compared with maternal platelet counts, platelet-associated gamma G immunoglobulin (PAIgG) values, and the presence of antiplatelet antibody in the maternal circulation, history of previous PIT, maternal treatments for ITP, and other maternal or neonatal factors. PIT (platelet counts <100 x 10(3)/microL) was observed in 9 (14.3%) of 63 pregnancies. Presence of circulating antiplatelet antibody in maternal blood, splenectomy prior to pregnancy, and history of previous PIT were observed more frequently with statistical significance in patients giving birth to neonates who developed PIT. No effect on occurrence of PIT was found by the administration of corticosteroids or immunoglobulin. Splenectomy prior to pregnancy was found by logistic regression analysis to be a single significant variable (p = 0.021, odds ratio 7.20, confidence intervals: 1.35 to 38.3) among the risk factors for PIT.

Sonodynamically-induced cell damage with fluorinated anthracycline derivative, FAD104.

The ultrasonically-induced in vitro cell damaging effect of fluorine-containing anthracycline derivative (FAD104) was investigated. Sarcoma 180 cells suspended in air-saturated PBS were exposed to ultrasound for up to 60 s in the presence and absence of FAD104. The rate of inducing cell damage with ultrasound was doubled with 80 microM FAD104, while no cell damage was observed with FAD104 alone. This enhancement was significantly inhibited by histidine, which may suggest a sonochemical mechanism.

Sonodynamically-induced in vitro cell damage enhanced by adriamycin.

Sonodynamically-induced cell damage and active oxygen generation enhanced by adriamycin (ADM) were compared in the same in vitro insonation set-up. Significant enhancement of the rates of both ultrasonically-induced cell damage and nitroxide generation was demonstrated with 40-160 microM ADM. Both rates correlated very well resulting in a correlation coefficient of more than 0.99. The enhancement of both rates was suppressed by 10 mM histidine. These results are consistent with the hypothesis that ultrasonically-generated active oxygen plays a major role in the sonodynamically-induced cell damage enhanced by ADM.

The 273rd codon mutants of p53 show growth modulation activities not correlated with p53-specific transactivation activity.

Cancer-related mutations of the p53 tumor suppressor gene are clustered in the four so-called 'hot spots', codons 175, 248, 273 and 281/282. By using recombination PCR in vitro mutagenesis, we introduced point mutations into the codon 273 of wild-type (wt) p53 (pC53-SN3) from Arg to His (pC53-273H [273H]), Asp (273D), Pro (273P), Lys (273K), Leu (273L) or Thr (273T), and compared their biological and biochemical activities with wt p53 and cancer-derived 175H, 248W and 273H/309S. Among them, 273H/309S, 273H and 273D as well as wt p53 transactivated the chloramphenicol acetyltransferase (CAT) gene placed downstream of the p53 binding consensus, while none of the other mutants including 273L did. Transcriptions from human c-fos and rat PCNA promoters were suppressed by wt p53 and 273D, while they were enhanced variously by all other mutants in Saos-2 and/or NIH3T3 cells. On the other hand, growth of human squamous carcinoma cell lines measured by the plating efficiency of G418-resistant colonies was enhanced by transfection of 175H, 248W, 273H/309S and 273P, while suppressed by not only wt p53, 273D and 273H but also 273L. Thus, 273H/309S enhanced cell growth in spite of its p53-specific transactivation activity, while 273L suppressed cell growth in spite of its complete loss of the p53-specific transactivation. We concluded that the sequence-specific transactivation of p53 is not always correlated with its growth inhibitory activity.

[Influence of nicardipine on the renin-angiotensin system ina hypertensive crisis during operation in elderly patients].

We studied the influence of nicardipine on renin-angiotensin (RA) system in twelve elderly patients, because the suppression of RA system, the decrease in reactivity of its feedback mechanism, and a high probability of arteriosclerosis and hypertension are well recognized in elderly patients. In hypertensive crisis during operation, intravenous bolus injection of nicardipine was done promptly by anesthesiologists in charge. Plasma renin activity (PRA), angiotensin I (A I), and angiotensin II (A II) were measured with RIA methods before administration of nicardipine and after hemodynamics were stabilized. Changes of systemic arterial pressure and heart rate were simultaneously recorded. Concentrations of PRA, A I, and A II were not significantly elevated, whereas systemic and diastolic blood pressures decreased significantly at 5 min, 15 min, 30 min and 60 min after administration of nicardipine. Heart rate was unchanged. But in a few cases there was an increase in PRA and A I. Average total dose of nicardipine was 26.8 micrograms.kg-1 and average fall of blood pressure was 23.9%. In conclusion, we can use intravenous nicardipine safely and stabilize hemodynamics easily without activation of RA system and rebounding of its feedback mechanism.