Comparison of the effects of controlled-rate cryopreservation and vitrification on 2-cell mouse embryos and their subsequent development.

Effects of two cryopreservation procedures (conventional slow controlled-rate freezing using a programmable freezer and vitrification by direct plunging into liquid nitrogen) were compared on 2-cell embryos and their subsequent development to blastocysts, fresh or cryopreserved 2-cell mouse embryos were developed into blastocysts in vitro. The percentage of vitrified embryos which developed into blastocysts was significantly lower than that of fresh and slow controlled-rate frozen embryos. Although blastocysts from each cryopreservation procedure appeared morphologically normal and neither number of cells in the blastocysts nor in-vitro trophoblast spreading differed significantly, there were significant differences in their functional viability. First, the glucose incorporation activity in terms of [(3)H]2-deoxyglucose (2-DG) uptake in vitrified and thawed 2-cell embryos significantly decreased compared with fresh or slow controlled-rate frozen and thawed 2-cell embryos. Second, 2-DG uptake by blastocysts developed in vitro from fresh 2-cell embryos and from slow controlled-rate frozen or vitrified 2-cell embryos was 105 +/- 75, 43.0 +/- 28.3 and 22.0 +/- 11.4 fmol/embryo/h respectively. Third, the implantation rate of blastocysts developed in vitro from vitrified 2-cell embryos (10.2%) was significantly lower than that from fresh 2-cell embryos (30.8%) or slow controlled-rate frozen 2-cell embryos (22.1%). Since these data suggest that cryopreservation may have ulterior consequences on the functional development of embryos and that vitrification may exert a more harmful effect than slow controlled-rate freezing, more attention should be paid to its safety before vitrification is used routinely in a clinical programme.

The circadian variation of urinary N-acetyl-beta-D-glucosaminidase and gamma-glutamyl transpeptidase in clinically healthy cats.

The circadian variation of urinary N-acetyl-beta-D-glucosaminidase (NAG, EC 3.2.1.30) and gamma-glutamyl transpeptidase (gamma-GTP, EC 2.3.2.2) was evaluated in cats. Urine and blood were collected at 4-hr intervals from adult cats (3 males, 9 females) weighing between 2.6 and 5.0 kg. There was no circadian variation in the urine volume, creatinine clearance, creatinine excretion, NAG excretion or gamma-GTP excretion. The average NAG and gamma-GTP indices in the 4-hr urine were similar to those for the 24-hr urine. However, the variance for the 4-hr urine samples was higher than that of 24-hr urine. In conclusion, although 4-hr urine samples can be used to estimate 24-hr urinary enzyme excretion, short-term spot urine samples may cause increased variation in the enzyme index.

Presence of dioxins in human follicular fluid: their possible stage-specific action on the development of preimplantation mouse embryos.

Examination of human follicular fluid revealed the presence of polychlorinated dibenzodioxins (PCDDs) and dibenzofurans (PCDFs) at concentrations of approximately 1 pg/ml (0.01 pg TEQ/ml). To study their possible action, two-cell mouse embryos were cultured in the presence of 2,3,7,8-tetrachlorodibenzo-p-dioxin (TCDD) at concentrations between 0.5 and 100 pM and evaluated at 24-h intervals for their development to the eight-cell and blastocyst stages. The percentage of eight-cell embryos exposed to TCDD at 1, 2, and 5 pM concentrations was significantly lower than that of controls. However, blastocyst formation of the surviving eight-cell embryos was accelerated, with the number of cells in the blastocysts increased in a dose-dependent manner. Findings suggest that PCDDs and PCDFs may be present in human reproductive fluid and may exert some stage-specific effects on early embryonic development.

Cryopreservation of mouse embryos affects later embryonic development possibly through reduced expression of the glucose transporter GLUT1.

In order to study the effects of cryopreservation on later embryonic development, two-cell mouse embryos were frozen, thawed, and then allowed to develop into blastocysts. The percentage of cryopreserved embryos which developed into blastocysts was significantly lower than that of fresh two-cell embryos. The amount of glucose incorporation in terms of 3H-2-deoxyglucose uptake in blastocysts developed in vivo, and in vitro from fresh or frozen-thawed two-cell embryos, was 473 +/- 108, 105 +/- 75, and 43.0 +/- 28.3 fmol per embryo per hour, respectively. Quantification of glucose transporter GLUT1 in these embryos by Western blotting was reflective of the degree of glucose incorporation. The implantation rate of blastocysts developed in vitro from frozen-thawed two-cell embryos (22.0%) was significantly lower than that developed in vivo (41.1%). These data suggest that cryopreservation may have later consequences on embryonic development through a mechanism that involves altered GLUT1 expression.

The variation in excretory urinary glycyl-prolyl dipeptidyl aminopeptidase in dogs.

We studied the excretory variation of urinary glycyl-prolyl dipeptidyl aminopeptidase (GP-DAP, EC 3.4.14.5) in dogs. Adult domestic mongrel dogs (seven males and nine females, 7.5 to 13 kg bodyweight) which were considered to be healthy by laboratory tests were used. Urine and blood samples were obtained every four hours. Urine volume was measured for each sample and urine GP-DAP activity, and creatinine levels were determined. Creatinine clearance, creatinine excretion, GP-DAP activity and GP-DAP index (GP-DAP/Cr ratio) did not show any significant variation between the time periods. However, urine volume and urinary GP-DAP excretion significantly increased from 8:00 am to 12:00 pm. The GP-DAP index in spot urine samples showed low correlation with 24 hour GP-DAP excretion. In addition, a sex difference was observed in GP-DAP excretion. In conclusion, urinary GP-DAP excretion showed a circadian variation and sex difference. Therefore, GP-DAP in spot urine is not of use for the detection of renal disorders, and the 24-hour excretion value of GP-DAP should be determined.

[Clinical study on symptoms in patients with strongyloidiasis].

We treated strongyloidiasis patients and obtained the following results: Of the 299 patients (184 males and 115 females), 81 patients (27.1%) had no complaints before treatment, 218 patients complaints of some symptoms, including arthralgia and/or lumbago (28.4%), abdominal pain and/or borborygmus (19.3%), numbness of extremities (18.1%), constipation (16.3%) and itching (15.7). We treated 219 patients with mebendazole and symptoms improved after treatment described below: Thirty-seven of the 63 patients (58.7%) with arthralgia and/or lumbago improved. Twenty-seven of the 36 patients (75.0%) with numbness of extremities improved. Thirty-one of the 32 patients (96.9%) with heartburn improved. We treated 26 patients with mebendazole plus thiabendazole and twelve of 14 patients (85.7%) with abdominal pain and/or borborygmus were improved after treatment. We treated 54 patients with ivermectin and five of 18 patients (27.8%) with arthralgia and/or lumbago were improved after treatment.

[Clinical study of albendazole therapy for strongyloidiasis].

We reported the efficacy of albendazole (ABZ) for the treatment of 27 patients with strongyloidiasis. Twenty-seven patients, 23 males and 4 females, received 200 mg of ABZ one hour before breakfast and supper for 3 days and this treatment was repeated 2 weeks later. The following results were obtained: 1) The eradication rate at 2 weeks after the initial treatment was 70.4% (19 of 27 patients) and 2 weeks after the second course was 66.7% (16 of 24 patients). 2) One patients (3.7%) complained of abdominal pain after the first treatment. Four patients (14.8%) complained of headache (n = 2), nausea (n = 1) and exanthema (n = 1) after the second treatment. But all symptoms were mild and required no treatment and subsided in a few days. 3) Positive rate of HTLV-1 antibody was 45.8% in the patients. As described above, side effects occurred in some cases, although they were mild and transient. From these results, it can be concluded that on increased dose of ABZ could be much more favorable for the treatment of strongyloidiasis.

[Clinical study of eradicated and resistant patients to treatment with ivermectin for strongyloidiasis].

We treated 70 patients with strongyloidiasis (41 males and 29 females) with ivermectin (IVM), and obtained the following results: 1. The eradication rates at 1-2 months, 3-4 months and 5-6 months after treatment were 90.7% (49 of 54 patients), 100.0% (47 of 47 patients) and 95.7% (45 of 47 patients), responsively. Twelve patients were resistant (non-responsive) to treatment. 2. When compared to patients whose parasites were completely eradicated, the resistant patients showed the following results: 1) Incidence of symptoms observed before treatment was significantly lower (50.0% vs. 84.5%). 2) Positive rate of anti-HTLV-I antibody was significantly higher (66.7% vs. 20.7%). 3) Blood eosinophil counts before treatment were significantly lower (266.6 +/- 117.2/mm3 vs. 533.2 +/- 429.7/mm3). 4) Serum IgE levels before treatment were significantly lower (217.2 +/- 442.9 IU/ml vs. 1,076.8 +/- 2,108.0 IU/ml). 5) There were no significant differences in age, sex and dosage of ivermectin. 3. Comparing anti-HTLV-I antibody positive and negative patients, the following results were obtained: 1) Eradicated patients; a) Eosinophils and IgE levels before and after the first administration of medicine in anti-HTLV-I antibody positive patients were significantly lower than those of negative patients. b) Gammaglobulin levels before treatment and after both administrations of the drug, IgG before therapy and OKT4/OKT8 after therapy were significantly higher than in anti-HTLV-I antibody positive patients. 2) Resistant patients; Eosinophils after treatment were significantly lower in anti-HTLV-I antibody negative patients than in positive patients.(ABSTRACT TRUNCATED AT 250 WORDS)

Hereditary angioedema: a case with ascites yet no symptoms in the family.

An interesting case of hereditary angioedema in a 26-year-old female is reported, with a finding of transient effusion of fluid into the peritoneal cavity during the attacks. The patient suffered from recurrent abdominal pain for several years, but no family members had any similar symptoms. In spite of repeated hospital admissions and many examinations, accurate diagnosis was not made until the most recent admission. The recognition of hereditary angioedema as a cause of acute and/or recurrent abdominal pain may avoid useless invasive procedures and lead to adequate treatment in other similar cases.

[Long term eradication rate of mebendazole therapy for strongyloidiasis].

We treated 245 strongyloidiasis patients with 7 schedules of mebendazole (MBZ) and obtained the eradication rates at 8 months to 2 years after the final treatment as described follows; MBZ (100 mg) was given twice a day orally. [Square bracket shows incidence of liver disfunction.] 1) The eradication rates at 2 years after single course of MBZ therapy for 28 days and a combination therapy (thiabendazole 500 mg powder form three times daily for 5 days followed by MBZ in powder form for 9 days, repeated once) were 93.8% (15 of 16 patients), [71.4%] and 100.0% (16/16), [50.0%]. 2) The eradication rates at 8-15 months after using MBZ alone in varying dosages were as follows: a) MBZ powder was administered for 5 days and was then repeated 1, 3 and 4 weeks later: 87.1 (27/31), [51.1%]. b) MBZ powder was given for 5 days and repeated 1 and 3 weeks later: 100.0% (7/7), [30.8%]. c) MBZ powder was administered for 4 days and repeated 1, 3 and 4 weeks later: 96.3% (26/27), [57.8%]. d) MBZ in tablet form was given for 4 days and repeated 1, 3 and 4 weeks later: 89.6% (43/48). [66.2%]. e) MBZ in tablet form was administered for 4 days and repeated 1 week later: 69.2% (9/13), [25.0%]. As described above, although the incidence of liver disfunction in the 4-day with 2-course therapy was lower than the other schedules, the eradication rate was lower. From these results, MBZ should be given to strongylodiasis patient for 4 days and repeated once, or for 3 days and repeated two or three times with 1 to 2 weeks intervals.

Efficacy of ivermectin against Strongyloides stercoralis in humans.

Okinawa Prefecture is an endemic area of Strongyloides stercoralis infection. Since treatment of this infection remains unsatisfactory, we evaluated the efficacy of ivermectin. Twenty-three patients were treated with a single oral dose of ivermectin (mean +/- SD, 105.5 +/- 20.8 mcg/kg of body weight), followed by a second dose two weeks later. The rate of cure was 85.7% at 2 weeks after the first treatment, and 90.5% at 2 weeks after the second treatment. Side effects occurred in 2 patients (8.7%), but they were mild and transient. The results indicate that ivermectin might be useful and relatively safe for the therapy of Strongyloides stercoralis infection as an alternative to thiabendazole or mebendazole.

[Clinical study on ivermectin against Strongyloides stercoralis].

We previously reported the efficacy of ivermectin (IVM) for the treatment of 23 strongyloidiasis patients. We now reported the efficacy and safety of IVM therapy on 54 patients. Fifty-four patients, 28 males and 26 females, received a single oral dose of IVM one hour before breakfast and this treatment was repeated 2 weeks later. The following results were obtained: 1) The cure rate at 2 weeks after the initial treatment was 92.5% (49 of 53 patients) and 2 weeks after the second course was 96.0% (48 of 50 patients). 2) Four patients (7.4%) complained of diarrhea (n = 2 patients), constipation (n = 1), borborygmus (n = 1), dizziness (n = 1), diplopia (n = 1) and peri-anal itching (n = 1) after the first treatment. Three patients (5.6%) complained of borborygmus (n = 1), itching (n = 1) and exanthema (n = 1) after the second treatment. But all symptoms were mild and required no treatment and subsided in a few days. 3) Positive rate of HTLV-1 antibody was 25.9% in the patients. As described above, although side effects occurred in some cases, they were mild and transient. From these results, we concluded that IVM is an effective drug for strongyloidiasis.

[Comparison of efficacy of powder and tablet of mebendazole in the treatment of strongyloidiasis].

We previously treated 47 patients with 100 mg of mebendazole (MBZ) twice a day by oral use for 5 days and this treatment was repeated 1, 3 and 4 weeks later. Although the cure rate was 100%, liver injury was observed in 48.9% of the patients. On this study, we reduced the periods of administration of MBZ (powder; 100 mg twice a day) to 4 days, and repeated it once after 3 days interval, and this initial treatment was performed one more time after 10 days interval (group 1). As Strongyloides stercoralis is mainly located in upper digestive systems, we used the drug reduced to powder for the purpose of better contact with the parasites. We considered that the powder should be absorbed well and liver injury occurred in high incidence. As group 2, we used the tablet itself in the same schedules of group 1. The results obtained were as follows; 1) The eradication rates at 10 days after the initial treatment were 97.8% (44 of 45 patients) in group 1 (powder) and 93.0% (40/43) in group 2 (tablet). 2) At 3 days after the whole treatment, the eradication rates were 100.0% in group 1, and 97.7% (42/43) in group 2. 3) Slight side effects such as constipation (6.7% in the group 1), dizziness or vertigo (6.7% in the group 1) and itching (6.7% in the group 2) were observed. 4) Liver injury was observed at 11.1% (5/45) 10 days after the initial treatment in the group 1 and 13.3% (6/45) in the group 2.(ABSTRACT TRUNCATED AT 250 WORDS)

[Treatment of strongyloidiasis with mebendazole--long-term eradication and new trials].

We previously reported the short period cure rate of mebendazole (MBZ) treatment to strongyloidiasis. We are now reporting the long period cure rate of the treatment. The results were as follows: 1) The cure rate was 73.9% (17/23) in single use of MBZ (100 mg twice daily for 28 days). 2) The cure rate was 100.0% (22/22) in combination therapy (thiabendazole 500 mg three times a day for 5 days and after that, MBZ 100 mg twice daily for 9 days). Before we obtained the cure rate of 6 months after the treatment described above, we concluded that MBZ could be used for the treatment of strongyloides infection because of the lack of severe side effects and suitable intervals between courses would prevent liver injury. Thus, in this study, 47 patients were treated with 100 mg of MBZ twice a day for 5 days and this treatment was repeated 1, 3 and 4 weeks later on the same schedules (group 1). But because of liver injury, 13 patients were interrupted and moved to the 4th course (group 2). The following results were obtained: 1) Out of a total of 60 patients, the cure rate was 88.1% (52/59) after 2 courses, 92.3% (12/13) after 3 courses and 100.0% (46/46) after the 4 courses were finished. 2) Diarrhea (10.6%), arthralgia and lumbago (8.5%) were observed in group 1. No side effects were observed in group 2. 3) The incidence of liver injury occurred 48.9% (23/47) in group 1 and 30.8% (4/13) in group 2. 4) Positive rate of HTLV-I antibody was 40.0% (24/60).(ABSTRACT TRUNCATED AT 250 WORDS)