Uracil-tegafur and tamoxifen vs cyclophosphamide, methotrexate, fluorouracil, and tamoxifen in post-operative adjuvant therapy for stage I, II, or IIIA lymph node-positive breast cancer: a comparative study.

BACKGROUND: It has been reported that treatment with uracil-tegafur (UFT) has shown significantly better survival and relapse-free survival (RFS) than surgery alone. Therefore, we compared UFT with a combination therapy of cyclophosphamide, methotrexate, and fluorouracil (CMF) in patients who had undergone curative surgery for axillary lymph node-positive breast cancer. METHODS: A total of 377 node-positive patients with stage I, II, or IIIA disease were registered from September 1996 through July 2000 and were randomly assigned to either 6 cycles of CMF or 2 years of UFT. In both arms, tamoxifen (TAM) was concurrently administered for 2 years. The primary end point in this study was the non-inferiority of UFT to CMF. RESULTS: No statistically significant difference between the two groups was observed with regard to the 5-year RFS rate (72.2% in the UFT and 76.3% in the CMF). Adverse event profiles differed between the two groups, with a significantly lower incidence of leukopenia and anaemia in the UFT group, as well as anorexia, nausea/vomiting, stomatitis, and alopecia, which have implications for quality of life. CONCLUSION: UFT administered in combination with TAM holds promise in the treatment of lymph node-positive early breast cancer. On stratified analysis, the recurrence rate in the UFT group was found to be better in oestrogen receptor (ER)-positive patients. Tegafur-based treatment should be evaluated by a prospective randomised trial conducted in ER-positive patients.

Prokinetic effect of a Kampo medicine, Hange-koboku-to (Banxia-houpo-tang), on patients with functional dyspepsia.

Limited evidence is available as to whether Kampo medicine modifies gastrointestinal function in humans. We investigated the effect of a Kampo medicine, Hange-koboku-to (Banxia-houpo-tang, HKT), on patients with functional dyspepsia (FD) and on healthy volunteers with regard to gastric motility. The gastric emptying rate (GER) in FD patients was significantly lower than in the healthy subjects. GER in FD patients and in healthy volunteers showed a significant increase after 2 weeks of medication with HKT. Furthermore, gastrointestinal symptoms improved significantly in the FD patients after the administration of HKT. These results suggest that HKT improves delayed gastric emptying and acts as a prokinetic agent.

Extraction of total RNA from leaves of Eucalyptus and other woody and herbaceous plants using sodium isoascorbate.

Rapid extraction of total RNA from Eucalyptus leaves is difficult due to the high content of polyphenolics and polysaccharides. A rapid and simple method was developed by using an extraction buffer containing sodium isoascorbate at a concentration of 500 mM. This method consisted of one or two chloroform extractions, one acid guanidium-phenol-chloroform extraction, and isopropanol precipitation alone. The yields of the RNA fractions were 246-1750 micrograms/g fresh weight when leaves of Eucalyptus, five other woody plants, and four herbaceous plants were used as samples. The contamination of the RNA fractions by proteins and polysaccharides was very limited as judged spectrophotometrically. When the RNA fractions were subjected to agarose gel electrophoresis, intact rRNA bands were detected. The RNA fractions could be used for RT-PCR. These results indicate that our new method achieves a simple and rapid preparation of high-quality RNA from leaves of Eucalyptus and other plant species.

[Nutritional consideration for changes in dietary habit and health promotion practices in community health care; from the view point of selenium].

The Japanese recommended dietary allowances (RDA) for major and some minor nutrients were revised in 1999, and included those for trace elements such as selenium. The requirement of selenium in animals was first recognized in 1957. It has been shown that cellular glutathione peroxidase (GPx) contains selenium but it was subsequently revealed that selenium has diverse biochemical effects, rather than simply functioning in the enzyme. At least twelve different selenoproteins have been identified. The role of selenium has been known as antioxidant, and non-antioxidant mediated through these enzymes. Now, selenium is well recognized as a preventive factor for cancer and cardiovascular diseases. Several dietary studies have shown that the selenium intake in Japan is adequate. One study estimated daily selenium intake to be 104.2 micrograms/day for adults. This value was 2 or 3 times higher than the lower limit of the safe range of dietary selenium (40 micrograms/day for men and 30 micrograms/day for women) estimated by WHO, and also exceeded the newly established RDA of 55-60 micrograms/day for men and 45 micrograms/day for women by the Japanese Public Health Council. However, the established RDA for selenium is tentative because of a lack of information on the 1) chemical forms of selenium in food, 2) differences in absorption rate and bio-availability in the chemical forms, and 3) interactions with other metals and trace elements. There are two potential problems concerning selenium nutrition in Japan. The first problem is that rice, which is the Japanese staple food, contains less than 0.05 microgram/g selenium whereas U.S. rice contains more than 0.3 microgram/g, probably due to differences in soil chemistry. The second problem is that although studies have shown that seafood, fish, shellfish and oysters, contain high levels of selenium (0.4-0.5 microgram/g), these being the main selenium source for Japanese, the bio-availability in fish is low. Thus, it is likely that the selenium status of those Japanese who eat an imbalanced diet is not sufficient or is not optimal even if the intake exceeds the RDA. Further studies are needed so that community health care specialists have available appropriate knowledge on the role of trace nutrients, including selenium, in human nutrition and health, to promote proper nutritional practices in the community.

Brief exposure to low-pH stress causes irreversible damage to the growing root in Arabidopsis thaliana: pectin-Ca interaction may play an important role in proton rhizotoxicity.

The viability of Arabidopsis thaliana (strain Landsberg) roots exposed to a low pH (4.5 or 4.7) solution that contained 100 microM CaCl(2) was examined by staining with fluorescein diacetate-propidium iodide. The elongation zone of growing roots lost viability within 1-2 h following exposure to low pH, but non-growing roots showed no damage under the same treatment. Low-pH damage in growing roots was irreversible after 1 h incubation at pH 4.5 as judged by regrowth in growing medium at pH 5.6. Growing lateral roots also lost viability in the same treatment, whereas non-growing lateral roots remained viable during and after the treatment. The low-pH damage was ameliorated by the simultaneous application of calcium, indicating the involvement of a calcium-requiring process in overcoming proton toxicity. At pH 5.0, growing roots required 25 microM of calcium to maintain elongation, and at pH 4.8 and pH 4.5 more than 250 microM and 750 microM, respectively. The low-pH damage was ameliorated by divalent cations in the order of Ba2+, approximately Sr2+>/=Ca2+>Mg2+. The monovalent cation K+ showed no ameliorative effect, but borate showed a strong ameliorative effect with Ca2+. These results indicate that the primary target of proton toxicity may be linked to a disturbance of the stability in the pectic polysaccharide network, where calcium plays a key role in plant roots.

Melatonin stimulates proliferation and type I collagen synthesis in human bone cells in vitro.

The pineal secretory product melatonin reportedly regulates release of growth hormone in humans and prevents phototherapy-induced hypocalcemia in newborn rats, suggesting that melatonin affects bone metabolism. Little is known about the effects of melatonin on bone in vitro or in vivo. The present study was undertaken to examine whether melatonin acts directly on normal human bone cells (HOB-M cells) and human osteoblastic cell line (SV-HFO cells) to affect osteogenic action in vitro. The effect of melatonin on bone cell proliferation was determined using the 2,3-bis[2-methoxy-4-nitro-5-sulfophenyl]-2H-tetrazolium-5-carbo xanilide (XTT) assay after a 24 hr incubation with melatonin. Melatonin significantly and dose-dependently increased the proliferation in HOB-M cells and SV-HFO cells by 215 +/- 22.1%, and 193 +/- 6.4%), respectively, with a maximal effect at a concentration of 50 microM. To evaluate the effect of melatonin on bone cell differentiation, alkaline phosphatase (ALP) activity, osteocalcin secretion and procollagen type I c-peptide (PICP) production (a measure of type I collagen synthesis) were measured after a 48 hr treatment. While melatonin at micromolar concentrations did not significantly affect either the ALP activity or the osteocalcin secretion, it significantly and dose-dependently increased the PICP production in HOB-M cells and SV-HFO cells by 983 +/- 42.2%, and 139 +/- 4.2%, respectively, with the maximal stimulatory doses between 50 and 100 microM. These results provide new evidence that melatonin stimulates the proliferation and type I collagen synthesis in human bone cells in vitro, suggesting that melatonin may act to stimulate bone formation.

Temporal and spatial patterns of accumulation of the transcript of Myo-inositol-1-phosphate synthase and phytin-containing particles during seed development in rice.

Myo-inositol-1-phosphate (I[1]P) synthase (EC 5.5.1.4) catalyzes the reaction from glucose 6-phosphate to I(1)P, the first step of myo-inositol biosynthesis. Among the metabolites of I(1)P is inositol hexakisphosphate, which forms a mixed salt called phytin or phytate, a storage form of phosphate and cations in seeds. We have isolated a rice (Oryza sativa L.) cDNA clone, pRINO1, that is highly homologous to the I(1)P synthase from yeast and plants. Northern analysis of total RNA showed that the transcript accumulated to high levels in embryos but was undetectable in shoots, roots, and flowers. In situ hybridization of developing seeds showed that the transcript first appeared in the apical region of globular-stage embryos 2 d after anthesis (DAA). Strong signals were detected in the scutellum and aleurone layer after 4 DAA. The level of the transcript in these cells increased until 7 DAA, after which time it gradually decreased. Phytin-containing particles called globoids appeared 4 DAA in the scutellum and aleurone layer, coinciding with the localization of the RINO1 transcript. The temporal and spatial patterns of accumulation of the RINO1 transcript and globoids suggest that I(1)P synthase directs phytin biosynthesis in rice seeds.

Separation of selenium-containing proteins in human and mouse plasma using tandem high-performance liquid chromatography columns coupled with inductively coupled plasma-mass spectrometry.

An analytical method that uses two different high-performance liquid chromatography (HPLC) columns in tandem has been developed that separates three major selenium-containing proteins (albumin, glutathione peroxidase, and selenoprotein P) found in human blood plasma. The first column was a heparin affinity column and the second was a gel filtration column whose outlet was directly connected to an inductively coupled plasma-mass spectrometer. The method successfully separated plasma selenium into the three selenium-containing proteins and revealed the preferential retention of selenium in the form of selenoprotein P in a selenium-deficient human and in selenium-deficient mice. Our results also confirm the results of previous studies that showed a preference for supplemented selenium to be taken up as selenoprotein P in rats. Advantages of the tandem column method are that it allows rapid and convenient analyses of the distribution of plasma selenium, and that it is suitable for stable isotope tracer studies and metal interaction studies.

Cyclophosphamide, adriamycin, 5-fluorouracil and high-dose toremifene for patients with advanced/recurrent breast cancer. The Japan Toremifene Cooperative Study Group.

BACKGROUND: Multi-combination chemotherapy consisting of anthracyclines has been effective but has not invariably prolonged the survival period in advanced/recurrent breast cancer. The possibility has been discussed that chemoendocrine therapy combined with endocrine agents is more effective. METHODS: In order to evaluate the toxicity and efficacy of a new endocrine therapy for advanced/recurrent breast cancer, we ran a pilot study during the period from July 1994 to July 1996. RESULTS: Twenty-two patients with advanced/recurrent breast cancer were treated with chemoendocrine therapy consisting of cyclophosphamide (100 mg/body) p.o. daily for 14 days, with adriamycin (40 mg/m2) i.v. and 5-fluorouracil (500 mg/body) i.v. on day 1 (repeated every 3 weeks for 9 weeks) (CAF therapy), and high-dose toremifene (120 mg/body) p.o. daily. Of 20 evaluable patients, two showed complete response (10%), eight partial response (40%), six no change (30%) and four progressive disease (20%). The overall response rate was 50%, and the median duration of response was 69.5 days (28-133+ days). The major toxicities were drug-induced alopecia, gastrointestinal toxicity and hematological toxicity, but these were clinically well tolerated. No serious cardiac, liver or renal symptom was seen. CONCLUSIONS: Based on these results, we consider the addition of high-dose toremifene to the CAF therapy to be useful in the treatment of advanced and recurrent breast cancer.

A case of Cushing's syndrome due to ACTH-independent bilateral macronodular hyperplasia associated with excessive secretion of mineralocorticoids.

A 74-year-old man developed Cushing's syndrome and hypokalemia due to ACTH-independent bilateral macronodular adrenocortical hyperplasia (AIMAH) with excessive secretion of mineralocorticoid hormones. Plasma concentrations of weak mineralocorticoids were high. The increase in plasma cortisol did not have a diurnal rhythm, and was not suppressed by a high dose of dexamethasone. Plasma ACTH was undetectable, but plasma cortisol was increased by ACTH administration. The concentrations of mineralocorticoids, especially deoxycorticosterone and corticosterone were increased, and augmented the response to ACTH administration. Plasma renin activity and aldosterone concentrations were rather suppressed. Both adrenal glands, with a total weight of 110 g, were enlarged and contained several macronodules. These nodules were composed of hyperplasia of small cortical cells and usual clear cells. This is a rare case of ACTH-independent bilateral macronodular adrenocortical hyperplasia because there were excessive secretion of mineralocorticoid and hypokalemia.

[Benefits of adjuvant chemotherapy for breast cancer].

The main therapy for primary breast cancer is not surgery, but a systemic therapy involving administration of cytotoxic chemotherapy or the use of ablative or additive endocrine therapy to control disseminated micrometastasis. The results of randomized trials and meta-analysis show that CMF, the standard adjuvant chemotherapy, is effective regardless of axillar lymph node involvement or menopausal status. Effectiveness of adjuvant chemotherapy with an anthracyclin-based regimen remains controversial. The trial by CUBC and NSAS-BC comparing UFT, widely used in the management of patients with breast cancer in Japan, with CMF is on-going.

Angiogenesis inhibitor TNP-470 prevents implanted liver metastases after partial hepatectomy in an experimental model without impairing wound healing.

The ability of the angiogenesis inhibitor TNP-470 to prevent liver metastasis after partial hepatectomy, and whether TNP-470 impairs liver regeneration or skin wound healing, was evaluated. Following the injection of VX2 carcinoma cells into the portal vein of rabbits, half of the animals underwent resection of the middle hepatic lobe (hepatectomized group) and half did not (non-hepatectomized group). TNP-470 (50 mg) was infused continuously into the portal vein in both groups for 7 days, while controls received only water. The hepatectomized TNP-470-treated group had significantly fewer tumours (mean(s.e.m.) 23.3(12.3)) than the hepatectomized control group (123.7(24.4)). There was no significant difference in the 5-bromo-2'-deoxyuridine labelling index of regenerated hepatocytes between the TNP-470-treated and control groups. Wound healing in TNP-470-treated animals was not impaired. Intraportal infusion of TNP-470 prevents the recurrence of liver metastasis after partial hepatectomy without impairing healing or liver regeneration.

Determination of selenium in the human brain by graphite furnace atomic absorption spectrometry.

For the investigation of neurological disorders, a development of simple and accessible methods for determining selenium in human brain samples is required. We devised a method of determining selenium using graphite furnace atomic absorption spectrometry (GFAAS). An electrodeless discharge lamp provided the sufficient sensitivity to determine brain selenium. The matrix interferences were avoided by using high temperature, a prolonged pyrolysis step, and a palladium matrix modifier. The technique of standard addition was used to evaluate the sample concentrations. The accuracy of the method was confirmed by a bovine liver reference material. The detection limit of selenium was 0.04 ng. The determined selenium concentrations of human brain cortex and white matter were higher than those of putamen (115-155 and 206-222 ng/g wet wt, respectively). These GFAAS values agreed with those obtained by fluorometric analysis (r = 0.91, n = 10). Moreover, the GFAAS values were compatible to those reported by other researchers (99-274 ng/g wet wt), in which selenium concentrations in putamen also tended to be higher than the other two regions. We conclude that GFAAS is useful for selenium analysis in brain samples.

Distribution of selenium in human plasma detected by high performance liquid chromatography-plasma ion source mass spectrometry.

The distribution of selenium in human plasma has been investigated by high performance liquid chromatography (HPLC) connected directly to inductively coupled plasma mass spectrometry (ICP-MS). Human plasma was loaded on to a size exclusion column and eluted with 0.01 M sodium phosphate buffer (pH 7.0) at a flow rate of 0.6 ml/min. Four peaks of selenium were detected in the chromatogram. The first selenium peak was obtained in the void volume. The retention time of the third peak was in accord with that of bovine serum albumin as a standard. The forth peak was thought to be a ghost. The method was applied to identify the chemical form of selenium in blood plasma immediately after intestinal absorption. The chromatographic pattern of selenium in postprandial human plasma was compared with that in fasting plasma. The first and third peaks in the postprandial plasma sample were slightly higher than those in the fasting plasma sample. This finding suggests that absorbed selenium is associated with the high molecular weight fraction and mercaptalbumin in blood plasma.

Effects of parathyroidectomy and calcium supplementation on the pressor response to angiotensin II in conscious rats.

OBJECTIVE: We evaluated the effect of parathyroidectomy (PTx) and of calcium supplementation on the pressor response to angiotensin II (Ang II) in conscious rats. STUDY DESIGN: PTx and sham surgery were performed on 10-week-old male Wistar rats. The mean arterial pressure (MAP), heart rate, and the effective pressor dose of Ang II (EPD) which defined as the dose of Ang II required to elicit a rise of 20 mmHg in MAP, were evaluated in PTx and sham operated rats. Intracellular free calcium in platelets was assessed with the fluorescent dye, fura-2 acetoxymethyl ester. In addition, after administering a dose of supplemental calcium chloride, a 10, 20, or 40 mg/rat, we determined the changes in the MAP, EPD, and serum calcium level. RESULTS: The EPD in the PTx rats was significantly lower than the sham operated rats. The serum concentration of calcium in PTx was also significantly lower than the sham operated rats. A statistically significant negative relationship was observed between the EPD and intracellular free calcium in PTx rats. Following administration of 20 mg of calcium chloride (7.4 mg of elemental calcium) to the PTx rats, the EPD returned to the level seen in sham operated rats. CONCLUSION: Results suggest that a depletion of parathyroid hormone is associated with the pressor response to Ang II, and is involved in the regulation of intracellular free calcium.

Isolation of human purH gene expressed in the rodent transformant cells by subtractive enrichment of 3'-untranslated region of human transcript.

A subtraction procedure was developed for identification and isolation of a human gene transcribed in mouse transformant cells. The procedure was based on subtractive enrichment of the products that were amplified by the combination of reverse transcription and polymerase chain reaction from the 3'-untranslated region (3'-UTR) of human poly(A)+ RNA expressed in the mouse transformant cells. To assess the ability and usefulness of the procedure, we attempted to recover the human purH gene from a mouse transformant cell line, which was originally established by functional complementation using the human metaphase chromosome-mediated gene transfer technique from a mouse purH-negative mutant cell line. Using our procedure, a part of the human transcript in the transformant cells was successfully identified and isolated. The full-length cDNA was isolated using the 3'-UTR clone as a probe, and its biological activity was confirmed by introducing it into the mouse purH-negative mutant cells.

Consistent relationship between selenium and apolipoprotein A-II concentrations in the sera of fasting middle-aged male abstainers and regular consumers of alcohol.

Several studies have suggested that selenium serum levels may be associated with serum lipids and apolipoproteins. In the present study, 99 clerical workers aged 40-49 yr were selected based on their drinking and smoking habits. The serum concentration of selenium was not affected by these lifestyle factors. The regular drinkers had raised serum high-density lipoprotein cholesterol, apo A-I, and apo A-II concentrations. Correlation analysis showed that serum selenium was positively and consistently associated with apo A-II regardless of alcohol consumption. Factor analysis revealed that serum selenium had no association with factors that represented each lipoprotein fraction (LDL, HDL, and VLDL). The present study indicates that serum selenium is positively correlated only with apo A-II levels.

[Effects of angiogenesis inhibitor TNP-470 to suppress the recurrence in the liver remnant after partial hepatectomy for liver metastases in rabbits].

We investigated the effects of an angiogenesis inhibitor, TNP-470, in suppressing the recurrence in the liver remnant after partial hepatectomy for VX2 liver metastases model of rabbits. After inoculation of 1 x 10(6) VX2 tumor cells via the portal vein, the medial lobe of the liver was resected. Then, TNP-470 (7 mg/body/day) was infused (Group 1), or distilled water (Group 2), continuously via the mesenteric vein for a week. The control group received only infused distilled water via the mesenteric vein. Fourteen days after the tumor inoculation, there were 66.0 +/- 20.2 (mean +/- SD) metastatic colonies on the liver surface of control group, against 23.3 +/- 24.0 and 123.7 +/- 47.9 colonies in Group 1 and Group 2, respectively. There were significant differences between control group and Group 2, and between Group 1 and Group 2. We consider that the liver regeneration following hepatectomy might provoke tumor growth of occult metastases in the liver remnants. TNP-470 suppressed these growth by its anti-neovascular activity. And this effect reduced the number of metastatic colonies. Fourteen days after hepatectomy, the weights of the liver were no different among 3 groups. The BrdU Labeling Index was also no different between Group 1 and Group 2. TNP-470 did not suppress the regeneration of the liver remnant. We consider that the systemic side effects of TNP-470 might be made minimal, because this agent affects only on endothelial cells. These results suggest that the angiogenesis inhibitor TNP-470 is beneficial to suppress hepatic recurrence after partial hepatectomy for liver metastases without suppression of liver regeneration.

Intra-arterial administration of the angiogenesis inhibitor TNP-470 blocks liver metastasis in a rabbit model.

We evaluated the best route of administration of TNP-470, an angiogenesis inhibitor, by comparing the anti-tumour effects and toxicity following injection via the hepatic artery, the portal vein, or the jugular vein in a rabbit model of liver metastases. Following the injections of 1 x 10(6) VX2 carcinoma cells into the portal vein of rabbits, 50 mg of TNP-470 was injected continuously into the hepatic artery, portal vein, or jugular vein for 7 days. The number of tumours on the surface of the liver was counted 14 days following the start of the infusion, and the serum glutamic-oxaloacetic transamine (GOT), glutamic-pyruvic transaminase (GPT) and total bilirubin concentrations were examined. In addition, a coloured silicon rubber was injected into the vessels of the liver to visualise the capillary networks around the tumours and assess the degree of suppression of angiogenesis by TNP-470. The mean number of tumours following intra-arterial injection (17.5 +/- 2.9) was significantly less than the control (237.0 +/- 34.0) (P < 0.05). The mean numbers of the tumours following intraportal (89.1 +/- 16.0) and intravenous (140.6 +/- 31.2) injection were both less than the controls (215.3 +/- 45.5, 284.8 +/- 55.4 respectively), but the differences were not significant. We conclude that intra-arterial injection of TNP-470 is the most effective method for preventing liver metastases in this model.

Prevention of preeclampsia with calcium supplementation and vitamin D3 in an antenatal protocol.

OBJECTIVES: Using an angiotensin sensitivity test we carried out a prospective study in an attempt to predict the possible onset of preeclampsia and to prevent it by calcium supplementation (elemental calcium 156 or 312 mg/day per os) and treatment with vitamin D3 (0.5 micrograms/3 day per os). METHOD: We used a study design in which 666 singleton pregnant women were managed with conventional antenatal care and 210 singleton pregnant women were managed with a protocol, together with conventional antenatal care. RESULT: Of the 666 women managed conventionally, 113 (16.9%) developed preeclampsia. However, the incidence of preeclampsia in the 210 women managed on the protocol was lower, at 10.9%. CONCLUSION: Our findings indicate that this protocol for the prediction and prevention of preeclampsia is useful for pregnant women at high risk of developing preeclampsia.