Contribution of glucan-binding protein A to firm and stable biofilm formation by Streptococcus mutans.

Glucan-binding proteins (Gbps) of Streptococcus mutans, a major pathogen of dental caries, mediate the binding of glucans synthesized from sucrose by the action of glucosyltransferases (GTFs) encoded by gtfB, gtfC, and gtfD. Several stress proteins, including DnaK and GroEL encoded by dnaK and groEL, are related to environmental stress tolerance. The contribution of Gbp expression to biofilm formation was analyzed by focusing on the expression levels of genes encoding GTFs and stress proteins. Biofilm-forming assays were performed using GbpA-, GbpB-, and GbpC-deficient mutant strains and the parental strain MT8148. The expression levels of gtfB, gtfC, gtfD, dnaK, and groEL were evaluated by reverse transcription-quantitative polymerase chain reaction (RT-qPCR). Furthermore, the structure of biofilms formed by these Gbp-deficient mutant strains was observed using confocal laser scanning microscopy (CLSM). Biofilm-forming assay findings demonstrated that the amount formed by the GbpA-deficient mutant strain (AD1) was nearly the same as that by the parental strain, while the GbpB- and GbpC-deficient mutant strains produced lower amounts than MT8148. Furthermore, RT-qPCR assay results showed that the expressions of gtfB, dnaK, and groEL in AD1 were elevated compared with MT8148. CLSM also revealed that the structure of biofilm formed by AD1 was prominently different compared with that formed by the parental strain. These results suggest that a defect in GbpA influences the expression of genes controlling biofilm formation, indicating its importance as a protein for firm and stable biofilm formation.

Fluvoxamine alleviates ER stress via induction of Sigma-1 receptor.

We recently demonstrated that endoplasmic reticulum (ER) stress induces sigma-1 receptor (Sig-1R) expression through the PERK pathway, which is one of the cell's responses to ER stress. In addition, it has been demonstrated that induction of Sig-1R can repress cell death signaling. Fluvoxamine (Flv) is a selective serotonin reuptake inhibitor (SSRI) with a high affinity for Sig-1R. In the present study, we show that treatment of neuroblastoma cells with Flv induces Sig-1R expression by increasing ATF4 translation directly, through its own activation, without involvement of the PERK pathway. The Flv-mediated induction of Sig-1R prevents neuronal cell death resulting from ER stress. Moreover, Flv-induced ER stress resistance reduces the infarct area in mice after focal cerebral ischemia. Thus, Flv, which is used frequently in clinical practice, can alleviate ER stress. This suggests that Flv could be a feasible therapy for cerebral diseases caused by ER stress.

Absolute calibration of imaging plate for GeV electrons.

An imaging plate has been used as a useful detector of energetic electrons in laser electron acceleration and laser fusion studies. The absolute sensitivity of an imaging plate was calibrated at 1 GeV electron energy using the injector Linac of SPring-8. The sensitivity curve obtained up to 100 MeV in a previous study was extended successfully to GeV range.

Human round spermatids from azoospermic men exhibit oocyte-activation and Ca2+ oscillation-inducing activities.

During mammalian fertilization, intracellular Ca2+ oscillations are important for both oocyte activation and embryonic development. As the ability of round spermatids (ROS) to induce Ca2+ oscillations and oocyte activation is different between species, we examined Ca2+ oscillation- and oocyte activation-inducing abilities of human ROS originating from patients with non-obstructive azoospermia. Human ROS from 11 non-obstructive azoospermic patients were collected during their TESE-ICSI cycles. Following injection into mature unfertilized mouse oocytes, we examined the oocyte-activating and Ca2+ oscillation-inducing activities of ROS by using Ca2+ imaging and confocal laser scanning microscopy (mouse test). In these 11 cases, clinical TESE-ICSI using mature testicular spermatozoa was successful, with the exception of one case in which only one sperm-injected oocyte was not fertilized. The mean fertilization rate was 70.1% (40-100%); the mean cleavage rate was 97.9% (46/47). Two pregnancies were established from 10 transfer cycles (PR; 20%). When the ROS from these patients were injected into mouse oocytes, the ROS from all patients induced at least some intracellular Ca2+ oscillations (25-100%). In all patients, 40 out of 82 oocytes injected with ROS exhibited normal oscillation patterns of [Ca2+]i. Human spermatogenetic cells acquired oocyte-activating and Ca2+ oscillation-inducing abilities at the round spermatid stage, an earlier stage than found for rodent cells. These data indicate that human ROS might be useful for clinical treatments of non-obstructive azoospermic patients exhibiting mature spermatozoa in biopsied specimens.

Successful pregnancy after ICSI with strontium oocyte activation in low rates of fertilization.

Fertilization failure (complete fertilization failure or low fertilization rates) after intracytoplasmic sperm injection (ICSI) can occur in rare cases. In the majority of these cases, the unfertilized oocytes are inactivated. Assisted oocyte activation was applied as a treatment option for a case of low fertilization rate as a clinical trial. A patient with a low fertilization rate (ranging from 0% to 33.3%; mean = 17.0%) after eight previous ICSI cycles at another hospital, was diagnosed with fertilization failure. The most likely cause of fertilization failure was failure of oocyte activation. Therefore, artificial oocyte activation by strontium treatment was combined with ICSI to achieve viable fertilized oocytes. Oocytes were stimulated with strontium (10 mM SrCl(2), 60 min) approximately 30 min after ICSl. Six injected oocytes were stimulated and all were then successfully fertilized. Two blastocysts were transferred into the uterus, resulting in a pregnancy and birth. A second pregnancy was achieved following implantation of two cryopreserved embryos (one blastocyst and one morula). In conclusion, strontium treatment was found to be an effective method for artificial oocyte activation in a case with a low fertilization rate after ICSI.

Use of sublingual isosorbide dinitrate tablet for manual extraction of a retained placenta.

We present the first report of the use of an isosorbide dinitrate tablet for the purpose of uterine relaxation for manual extraction of a retained placenta. The tablet administered sublingually proved to be a rapid and effective uterine muscle relaxant for manual removal of the placenta without overt adverse effects.

Angiosarcoma of vagina successfully treated with interleukin-2 therapy and chemotherapy: a case report.

We report a case of angiosarcoma of the vagina in a 61-year-old woman who had undergone radical hysterectomy and pelvic irradiation for uterine cervical adenocarcinoma 14 years previously. Combination chemotherapy (cyclophosphamide, vincristine, doxorubicin and dacarbazine) and interleukin-2 induced complete remission of the tumor. The patient remained free from disease for 15 months.

Oocyte activation and Ca(2+) oscillation-inducing abilities of mouse round/elongated spermatids and the developmental capacities of embryos from spermatid injection.

To investigate differences in fertilization mechanisms and the potential clinical use of round/elongated spermatid, we conducted detailed studies of oocyte activation and Ca(2+) oscillation-inducing abilities in these immature sperm cells and compared these abilities against those of mature spermatozoa. When round spermatids from B(6)D(2)F(1) mice were injected, none of the oocytes was activated and no intracellular Ca(2+) ([Ca(2+)](i)) increases were observed. Elongated spermatids could induce activation normally in 87% of injected oocytes, but Ca(2+) oscillation could not be induced at all and most of the oocytes (94%) exhibited only several transient [Ca(2+)](i) rises (transient patterns). Because normal offspring could be obtained when embryos through elongated spermatid injection were transferred to foster mothers, it seems that a normal oscillation pattern of [Ca(2+)](i) is not essential for normal fertilization and embryo development. [Ca(2+)](i) patterns of injected oocytes changed from transient patterns to oscillation patterns while the injected immature sperm cells were maturing to spermatozoa. Dissociations were seen between the timing of appearance of oocyte activation and that of Ca(2+) oscillation-inducing abilities in maturing sperm cells. These dissociations may be due to differences in the thresholds to oocyte activation and Ca(2+) oscillation-inducing factor for inducing oocyte activation and Ca(2+) oscillation.

Wide use of skin-lightening soap may cause mercury poisoning in Kenya.

In a previous study, we speculated that some of the high mercury levels observed in head hair from a total of 14 subjects who resided around Lake Victoria, Tanzania, might be attributable to the habitual use of toilet soap containing considerable amounts of mercury (Harada et al. Sci Total Environ 1999;227:249-256). In August 1998, the current study was conducted to investigate if such mercury-containing soap was also available in the surroundings of Lake Victoria, Kenya, and if so, its toxic effects. A total of nine goldminers, 44 fishermen and their families, and 12 residents of Kisumu City, Kenya, volunteered for the study. Fourteen types of toilet soap were collected in Kisumu. Total mercury content was very significantly higher than in European-made soap (0.47-1.7%, as mercury iodide) compared with Kenya-made soap (0.41 x 10(-4)-6.2 x 10(-4)%). Indeed, all the subjects with a high hair mercury level (> 36.1 ppm) had made habitual use of European-made soap, accompanied by various symptoms, such as tremor, lassitude, vertigo, neurosthenia, and black and white blots, suggesting inorganic-mercury poisoning. On the other hand, any subject who had used soap other than the European-made soap, did not exceed a mercury level of 10 ppm in hair that is well within normal limits (Harada et al. Sci Total Environ 1999:227:249-256). The findings obtained suggest that the mercury-containing soap must be barred from circulation without delay, and that the residents' health in addition to the environmental pollution in Lake Victoria (Kenya as well as Tanzania) should be kept under close observation.

Influence of sperm immobilization on onset of Ca(2+) oscillations after ICSI.

Sperm immobilization prior to intracytoplasmic sperm injection (ICSI) is thought to be necessary for efficient fertilization. A variety of methods of sperm immobilization (pipetting, squeezing and piezo application) are currently employed in ICSI. The effect of differences in immobilization method on the timing of initial Ca(2+) oscillations of oocytes in ICSI was investigated. Motile spermatozoa were immobilized in eosin Y solution using pipetting, squeezing and piezo application. Complete staining of the sperm head was achieved after 220.7, 42.2 and 5.0 s respectively. Oscillations after ICSI were measured fluorometrically for each method. The onset of Ca(2+) oscillations was observed at 4.8 to 80.4 min after ICSI. Ca(2+) oscillations developed earlier with the piezo method (14.4 +/- 6.4 min) than other methods (pipetting, 43.1 +/- 20.2 min, P < 0.01; squeezing, 18.4 +/- 3.8 min, P = NS). The piezo method produced the earliest staining of the sperm head and may have caused the most damage to the sperm membrane. A more rapid onset of Ca(2+) oscillations was also observed with the piezo method. The method of sperm immobilization may be important for the rapid release of sperm factors that initiate oocyte activation. This study also showed that Ca(2+) oscillations develop earlier in human oocytes treated by ICSI than indicated in previous reports.

A new fetal therapy for chylothorax: pleurodesis with OK-432.

We present a case of pleurodesis by intrapleural injection of OK-432 for the treatment of fetal chylothorax at an early gestational age. OK-432 injection achieved rapid and effective control of pleural effusion with no adverse effects.

Comparison of oocyte activation and Ca2+ oscillation-inducing abilities of round/elongated spermatids of mouse, hamster, rat, rabbit and human assessed by mouse oocyte activation assay.

Oocyte activation and Ca2+ oscillation-inducing abilities of round spermatid (ROS) and elongated spermatid (ELS) of some rodents and human were assessed by their injection into mouse (B6D2F1) oocytes (mouse test). With mice (B6D2F1, ICR) and rat, ROS displayed no oocyte activation or Ca2+ oscillation-inducing abilities. Although ELS could induce activation at 87, 86 and 31% of injected oocytes respectively, most of the intracellular calcium concentration ([Ca2+]i) responses of ELS-injected oocytes did not show oscillation patterns; only several transient [Ca2+]i rises (transient pattern) were seen. Similarly, with hamster, rabbit and human, while ROS could induce oocyte activation efficiently (70, 71 and 52% respectively), most of the [Ca2+]i patterns of injected oocytes were transient patterns, and not oscillation patterns. When ROS nuclei only from these latter species were injected into mouse oocytes, most of the oocytes could not be activated. [Ca2+]i patterns of oocytes injected with immature sperm cells changed from transient pattern to oscillation pattern while the cells were maturing into spermatozoa. With hamster ROS, oocyte-activating factor was found to be distributed mainly in the cytoplasm. It was interesting that there is a dissociation between the timings of appearance of oocyte activation and that of Ca2+ oscillation of oocytes injected with developing immature sperm cells.

Dimeric inhibin A as a fourth marker for Down's syndrome maternal serum screening in native Japanese women.

OBJECTIVE: This study was conducted to assess the usefulness of dimeric inhibin A as a fourth marker for Down's syndrome screening in addition to AFP, hCG and uE3 markers for native Japanese women. METHODS: Serum specimens from 367 native Japanese women in the second trimester were assayed for dimeric inhibin A levels. Day specific dimeric inhibin A medians were established for gestational ages 15.0-21.9. Weekly median values for the native Japanese were compared with those of a U.S. population. Selected Japanese specimens from 15 diagnosed Down's syndrome and 3 trisomy 18 cases were also assayed for dimeric inhibin A. RESULTS: Dimeric inhibin A levels did not vary greatly over the gestational age range as expected. Median value comparison showed that native Japanese dimeric inhibin A medians are higher than the U.S. population medians by an average of 7.95%. Native Japanese dimeric inhibin A median values in this study are 1.77 times higher in Down's syndrome cases than in unaffected pregnancies. Trisomy 18 dimeric inhibin A levels show no significant difference from the unaffected pregnancies. CONCLUSIONS: This report shows for the first time that dimeric inhibin A can be informative as a fourth marker for Down's syndrome screening in native Japanese women. We expect the addition of dimeric inhibin A to a triple marker protocol will increase the accuracy of predicted risk for all pregnancies screened and increase the detection rate of Down's syndrome affected pregnancies.

Oocyte activation induced by spermatids and the spermatozoa.

It has been reported that a sperm factor (SF) found in spermatozoa plays a critical role in fertilization. However, particulars of the oocyte-activating and Ca2+ oscillation (Ca-Os)-inducing abilities of this SF remain unknown. We examined these abilities of spermatids in mouse, hamster and human by a mouse test (injection of spermatids into mouse oocytes). In mice, the round spermatids (ROS), elongated spermatids (ELS) and spermatozoa activated 0%, 93% and 92% of the oocytes, respectively. ROS injection resulted in no Ca-Os (type C). ELS induced a normal oscillation (type A) at 0% and an abnormal oscillation (type B) at 94%. Mouse spermatozoa induced type A Ca-Os at 90%. For mice, oocyte-activating and Ca2+ oscillation-inducing ability arose in different phases of spermiogenesis. We also observed this differential timing for hamster spermatids. Hamster ROS activated 74% of oocyte (ELS: 90%, sperm: 86%). Human ROS activated 64% of oocytes (sperm: 100%), but only 35% of the oocytes showed type A Ca-Os. These results indicate that oocyte activation generally occurs between the ROS and ELS phases, although these phases differ among species. They also indicate that oocyte activation is not necessarily accompanied by Ca-Os. These findings suggest the existence of different thresholds at which the SF induces oocyte activation and Ca2+ oscillation, or of different factors that induce oocyte activation and Ca-Os. We found SF to be clinically impaired in 0.9% of ICSI patients. A combination of artificial oocyte activation and ICSI proved effective with such patients.

Precise measurements of the gamma-ray emission probabilities of 186Re and 188Re.

Gamma-ray emission probabilities of 186Re and 188Re are important in nuclear medicine, and have been precisely measured by 4pibeta-gamma coincidence using a live-timed two-dimensional data-acquisition system. The gamma-ray emission probabilities of the 137.2 keV for 186Re and 155.1 keV for 188Re were 0.09487+/-0.00029 and 0.15425+/-0.00072, respectively.

Triple marker screening for trisomy 21, trisomy 18 and open neural tube defects in singleton pregnancies of native Japanese pregnant women.

OBJECTIVE: To report the results of prenatal triple marker screening on a population of Japanese pregnant women. METHODS: From April 1994 through March 1999, a total of 32,925 native Japanese women with singleton pregnancies requested a triple marker-screening test. Multiples of the median values for 3 markers and individual risks for each patient were calculated following adjustment for the Japanese weight correction factor. The risk cut-off values used for Down syndrome (T21), open spina bifida (OSB) and trisomy 18 (T18) were 1: 295, 1: 290, and 1: 100, respectively. Follow-up information was collected postpartum and statistically analyzed. RESULTS: Detection rates (DR) of T21 for women less than 35 years, over 35 years and overall were 58, 94, and 83%, respectively. DR of T18 for women less than 35 years, over 35 years and overall were 75, 79, and 79%, respectively. DR of open neural tube defects (ONTD) was 100%. CONCLUSIONS: The first cumulative data of an intervention program and prospective follow-up studies in Japan have proven to be similar to other published reports. Individual risk values were calculated for each pregnancy for T21, T18 and ONTD. This screening program is more effective than age-dependent screening for detecting T21, T18 and ONTD pregnancies.

The value of pre-operative diagnostic procedures for cervical involvement in uterine corpus carcinoma.

OBJECTIVE: To assess the value of procedures for pre-operative diagnosis of cervical involvement of uterine corpus carcinoma. MATERIALS AND METHODS: Four diagnostic procedures, including cervical cytology, endocervical curettage (ECC), magnetic resonance imaging (MRI), and hysteroscopy, were performed for diagnosis of cervical involvement in 60 patients with uterine corpus carcinoma. The preoperative diagnosis based on results obtained using by each procedure was retrospectively compared with the diagnosis based on histological examination of surgical specimens. Data were analyzed according to the standard definition of sensitivity, specificity, positive predictive value and negative predictive value. RESULTS: Cervical involvement was confirmed in 18 patients (30%). ECC showed high sensitivity (90.9%) and specificity (88.9%). Cervical cytology showed high specificity (88.6%). MRI showed very high specificity (99.2%) and high sensitivity (88.5%) in cases with cervical stromal invasion. CONCLUSION: Cervical cytology and MRI are useful for excluding cervical involvement. ECC is useful for positive diagnosis. MRI may be useful for cases with stromal invasion. The use of a combination of several procedures is essential for obtaining an accurate diagnosis of cervical involvement in cases of uterine corpus carcinoma.

Myxoid leiomyosarcoma of the uterus.

A 70-year-old, menopausal Japanese woman with large uterine tumor underwent total hysterectomy. Microscopic examination revealed a myxoid leiomyosarcoma with 5 to 6 mitoses/10 high power field (HPF). Immunohistochemically, the tumor cells reacted with desmin, suggesting that they are derived from uterine smooth muscle cells. The patient was treated with the combination chemotherapy consisting of cisplatin, adriamycin and cyclophosphamide, and she has been free from disease for 70 months after the operation.

A pyridylamination method aimed at automatic oligosaccharide analysis of N-linked sugar chains.

The procedure for preparation of pyridylaminated sugar chains from glycoproteins was improved with a view to its eventual automation. Following on the coupling reaction improvement already reported [N. Kuraya and S. Hase (1992) J. Biochem. 112, 122-126], two further aspects were improved in this study. Instead of sodium bicarbonate-acetic anhydride, volatile reagents were adopted for the re-N-acetylation of hexosamine residues after hydrazinolysis to give rapid removal of excess reagents. Subsequent to the pyridylamination reaction, excess reagents were removed by cation-exchange to isolate the pyridylaminated oligosaccharides in place of gel filtration. These alterations rendered a one-pot reaction possible and resulted in a large reduction in the amount of time needed compared with other methods so far reported. The procedure was successfully applied to the detection of sugar chains from Taka-amylase A and human erythrocyte membranes.