Effect of an isoflavones-containing red clover preparation and alkaline supplementation on bone metabolism in ovariectomized rats.

The aim of this study was to test the combined effect of a quality-controlled red clover extract (RCE) standardized to contain 40% isoflavones by weight (genistein, daidzein, biochanin A, and formononetin present as hydrolyzed aglycones) together with a modified alkaline supplementation on bone metabolic and biomechanical parameters in an experimental model of surgically-induced menopause. Sprague-Dawley female rats were maintained under controlled standard conditions of light and fed with conventional food of standard calcium content and no alfalfa or soybean components. Rats were randomized into four groups: Group A represented normal rats (sham operated) while three other groups were ovariectomized (OVX) and fed for three months as follows: standard food (group B), 6 mg/kg/day food mixed with RCE (Group C), or given 6 mg/kg/day of RCE plus a modified alkaline supplementation (BP) through a nasogastric tube at a dose of 16 mg (group D). The animals were killed 90 days after surgery. As compared to group B, RCE or RCE + BP treatments brought about significantly higher level of estradiol and mitigated the weight loss of the uterus and improved maximum load of the femoral neck. Osteocalcin level showed an over 65% increase in group B but both RCE and RCE + BP treatments prevented such abnormality with a significantly better result in RCE + BP group which virtually normalized such parameter as well as urinary excretion of DPD. Group C and D reduced the over 20% loss of bone mineral density and bone mineral content/body weight ratio observed in untreated post-ovariectomy group. Untreated ovariectomy caused about 48% decrease of cancellous bone mass in the femoral neck while this abnormality was prevented at similar extent by both RCE and RCE + BP treatments. Ovariectomy determined an over 80% increase of bone alkaline phosphatase (BALP) level but both RCE and RCE + BP treatments significantly mitigated such variable. The BALP decrease yielded by the combined RCE + BP treatment was statistically lower than RCE alone. Taken together these data show that red clover preparation in dosages amenable to clinical practice do improve OVX-induced osteoporosis while a mild metabolic alkalosis might further synergize some therapeutic aspects.

The aging gut motility decay: may symbiotics be acting as "implantable" biologic pace-makers?

Motility recording of small and large intestine was performed in old Wistar rats divided into three groups: (a) standard diet, (b) standard diet plus a symbiotic preparation, and (c) standard diet plus a heat-inactivated symbiotic preparation. SCM-III. significantly increased the myoelectric activity of small intestine and colon (p < 0.01 versus [a] and [c]) paralleling "young" values of 4-month-old rats and increased the spike burst frequency in the proximal-distal colon (p < 0.05). SCM-III significantly increased the frequency and duration of spike bursts in the jejunum, transverse-distal colon, and defecation frequency, while decreasing the intervals of migrating motor complex in the colon (p < 0.01) to "young" values with an increased mRNA expression of VIP (p < 0.05). Gut flora manipulation aimed to modulate myoelectric activity can tentatively help reversing age-related motility decay.

Metal-induced oxidative damage in cultured hepatocytes and hepatic lysosomal fraction: beneficial effect of a curcumin/absinthium compound.

OBJECTIVE: Metals undergo redox cycling and there is increasing evidence of free radical generation and oxidative injury in the pathogenesis of liver injury and fibrosis in metal storage diseases. The aim of the present study was to test a natural hepatoprotective compound in metal-induced liver injury. METHODS: Hepatocytes were isolated from Wistar rats by collagenase perfusion method and cultured as such and also with alpha-linolenic acid (LNA)-bovine serum albumin (BSA). Hepatocytes were then cultured with a graded dilution of PN-M001 (100 microg/mL and 200 microg/mL), which is a curcuma/absinthium-containing compound, or sylibin (100 microg/mL) dissolved in dimethyl sulfoxide for 10 min before the addition of metallic salts (iron, copper and vanadium). Lysosomal fractions were prepared for lysosome fragility tests in which beta-galactosidase activity and lactate dehydrogenase (LDH) leakage were measured, as well as oxidative damage tests in the presence of hydrophilic and lipophilic free radical generators. Quenching activity by 1,1-diphenyl-2-picrylhydrazyl (DPPH) was also assessed. RESULTS: Malonildialdehyde accumulation in the medium showed a direct time-course increase with incubation time. Both PN-M001 and sylibin showed a significant protective effect against all challenge metal ions, as expressed by the half inhibition concentration (IC(50)) against lipid peroxidation. However, on a molar ratio, sylibin seemed to be more effective than PN-M001 in Fe-induced peroxidative damage (P < 0.05). Both test compounds, irrespective of the concentration, significantly reduced the LDH and beta-galactosidase concentration in the lysosomal fractions. As compared with untreated lysosomal fractions challenged with the two peroxide radicals generators, either PN-M001 or sylibin exerted significant protection However, PN-M001 was significantly better than sylibin in suppressing acid phosphatase enzyme activity. Both compounds showed comparable and significant DPPH radical-scavenging activity. CONCLUSION: These data support the potential clinical application of curcumin-containing compounds.

Single-blind follow-up study on the effectiveness of a symbiotic preparation in irritable bowel syndrome.

OBJECTIVE: Experimental and clinical studies have shown that a novel symbiotic (known as SCM-III) exerts a beneficial effect on gut translocation and local and systemic inflammatory and microbial metabolic parameters. The present investigation was a preliminary trial on the effectiveness of SCM-III for irritable bowel syndrome (IBS). METHODS: Sixty-eight consecutive adult patients with IBS who were free from lactose malabsorption, abdominal surgery, overt psychiatric disorders and ongoing psychotropic drug therapy or ethanol abuse were studied prospectively and divided into 2 groups that were comparable for age, gender, body size, education and pattern of presenting symptoms. The 2 groups were blindly given for 12 weeks either SCM-III 10 mL t.i.d or the same dosage of heat-inactivated symbiotic. RESULTS: Treatment with SCM-III was 'effective' or 'very effective' in more than 80% of the patients (P < 0.01 vs baseline values and control). Less than 5% reported 'not effective' as the final evaluation compared with over 40% of patients in the control group. After 6 weeks of treatment, a significant improvement of pain and bloating was reported in the treatment group compared with control and baseline values. There was also a benefit for bowel habits, mostly for patients with constipation or alternating bowel habits. No overt clinical or biochemical adverse side-effects were recorded. CONCLUSION: Compared with baseline values and the control group, SCM-III resulted in a significant increase in lactobacilla, eubacteria and bifidobacteria, which suggests that some selected IBS patients could benefit substantially from symbiotics, but the treatment may need to be given on a cyclic schedule because of the temporary modification of the fecal flora.

Clinical benefits of lenograstim in patients with neutropenia due to chemotherapy for multiple myeloma (MM).

The object of this study was to determine the efficacy and safety of glycosylated recombinant human granulocyte colony-stimulating factor (rHuG-CSF; lenograstim) after combination chemotherapy consisting of ranimustine, vindesine, melphalan and prednisolone (MCNU-VMP). One hundred thirty-nine consecutive patients with newly diagnosed multiple myeloma (MM) were allocated at random to a lenograstim group (n = 70) or a placebo group (n = 69). Patients were treated with two cycles of MCNU-VMP, and either lenograstim (2 microg/kg daily, s.c.) or placebo was administered from the day neutrophils decreased to less than 1.000/microl and was discontinued when neutrophils exceeded 5,000/microl. The median duration of neutropenia (neutrophils under 1,000/microl) was significantly shorter for the lenograstim group than the placebo group (2 days vs 9 days in the first cycle; 1 day vs 13 days in the second cycle). The incidence of febrile neutropenia in the first cycle was significantly lower in the lenograstim group than in the placebo group (9.2% vs 30.4%). No life-threatening infections were observed in either group. The two cycles of MCNU-VMP therapy were completed in 90.8% of the patients, and a higher average relative dose intensity (ARDI; 0.94) was achieved in the lenograstim group. The tumor response rate of the lenograstim group (57.8%) was higher than that of the placebo group (43.1%), but the difference was not statistically significant (chi2 = 2.634, df = 1, P = 0.105). Lenograstim was well tolerated, and no unexpected adverse events occurred. Lenograstim proved effective in controlling chemotherapy-induced neutropenia in MM patients under MCNU-VMP therapy.

Enhanced expression of vsmNOS mRNA in glomeruli from rats with unilateral ureteral obstruction.

BACKGROUND: The vasodilatory/cytotoxic gas, nitric oxide (NO), is associated with an alteration in glomerular hemodynamics seen after the induction of ureteral ligation. As yet the type of nitric oxide synthase (NOS) protein involved in the mechanism has not been clearly established in the unilateral ureteral obstruction (UUO) model. METHODS: Using reverse transcription (RT)-polymerase chain reaction (PCR), the expression and localization of vascular smooth muscle-derived nitric oxide synthase (vsmNOS) mRNA were examined in glomeruli from sham-operated control (SOC) rats and rats with UUO of three hours duration. Moreover, the effect of endogenous angiotensin II on the expression of vsmNOS mRNA in glomeruli was explored using SOC rats and rats with UUO that were pretreated or not with enalapril, an angiotensin-converting enzyme inhibitor. RESULTS: The expression of vsmNOS mRNA was significantly greater in glomeruli of rats with UUO than in those of SOC rats. In rats with UUO, the expression of vsmNOS mRNA was substantially increased in glomeruli of the obstructed kidney (OK) compared to the contralateral, nonobstructed kidney (CLK). Suppression of angiotensin II production in vivo with enalapril restored the expression of vsmNOS mRNA in glomeruli of the CLK and OK from rats with UUO to levels comparable to that seen in glomeruli from SOC rats. In addition, the in situ RT-PCR analysis, a novel method for mRNA identification in cells and tissue, revealed that vsmNOS mRNA was expressed in the cytoplasm of glomerular mesangial and epithelial cells in SOC rats and rats with UUO. CONCLUSIONS: An increase in vsmNOS mRNA expression in glomeruli of the CLK and OK from rats with UUO may be mediated by increased action of endogenous angiotensin II that occurs after the onset of ureteral obstruction. Enhanced expression of vsmNOS mRNA in glomeruli of the OK compared to the CLK may be due to differences in levels of angiotensin II acting on the two kidneys in vivo. Additionally, the expression of vsmNOS mRNA in glomeruli originates in mesangial and epithelial cells in SOC rats and rats with UUO.

[Marked thrombocytopenia after high-dose intravenous gamma globulin in a pregnant woman with idiopathic thrombocytopenic purpura].

A 35-year-old pregnant woman had thrombocytopenia with a platelet count of 6.3 x 10(4)/microliter. After her third normal delivery, peripheral blood studies revealed that the patient had a normal Hb concentration and leukocyte count, with mild thrombocytopenia. A diagnosis of idiopathic thrombocytopenic purpura (ITP) was made based on the high megakaryocyte count of 338/microliter and PAIgG of 40.8 ng/10(7) cells in January 1995. The patient was followed without treatment. She was 9 weeks pregnant on June 7, 1996, and desired an abortion. Her platelet count was 6.3 x 10(4)/microliter, leukocyte count 8,600/microliter, and Hb 13.7 g/dl at the time. She was given high-dose intravenous gammaglobulin (Globenin-I) at 400 mg/kg/day for 5 consecutive days. The platelet count was found to have decreased markedly, to 0.9 x 10(4)/microliter on June 11. The percentage reduction in the Hb concentration, leukocyte count, and platelet count after gammaglobulin treatment was 11.7%, 46.6%, and 85.8%, respectively. The PAIgG titer had increased to 181.2 ng/10(7) cells on June 17, but hypergammaglobulinemia was suspected. The patient was started on prednisolone on June 24, and an abortion was performed on July 29. The mechanism of thrombocytopenia after infusion of Globenin-I was unknown. We suspect that Globenin-I treated with polyethylene glycol was one of the possible causes of myelosuppression in this case.

Myelodysplastic syndromes with nephrotic syndrome.

It is sometimes reported that the immunological abnormalities in myelodysplastic syndromes (MDS) induce autoimmune disease (i.e., acute systemic vasculitic syndrome, chronic cutaneous vasculitis, polyneuropathy, relapsing polychondritis, and steroid-responsive pulmonary disorders). We investigated the clinical features of patients with MDS accompanied by nephrotic syndrome. We enrolled 125 patients with MDS who were admitted between January 1979 and May 1996 in this study. The renal function was assessed based on the laboratory data and the findings at the physical examination. The diagnoses of nephrotic syndrome and glomerular disease were established when 24-hr urinary excretion was more than 3.5 g and serum total protein was less than 6.0 g/dl, and when the 24-hr protein excretion was more than 1.5 g. Five patients (4%) had glomerular disease, and three (2.4%) had nephrotic syndrome. Of the five patients with glomerular disease, two had refractory anemia (RA), and three had chronic myelomonocytic leukemia (CMMOL). Three of the total 11 patients with CMMOL were diagnosed as having nephrotic syndrome. Among the CMMOL patients, those with nephrotic syndrome showed higher absolute monocyte numbers than did those without nephrotic syndrome (8830 +/- 4677/microl vs. 3061 +/- 2887/microl, P = 0.03). One CMMOL patient was treated with VP-16 and hydroxyurea. As the white blood cell count in this patient decreased, the 24-hr urine protein excretion and the serum tumor necrosis factor alpha level decreased. The relationship between nephrotic syndrome and CMMOL was not clear. High monocyte count and the serum cytokines in MDS patients may play a partial role in the evolution of glomerulonephritis, and CMMOL may be closely related to nephrotic syndrome.

Prognostic value of morphological classifications and clinical variables in elderly and young patients with multiple myeloma.

The prognostic value of morphological classifications and clinical variables was compared between 31 elderly (> or = 65 years) and 43 young (< 65 years) patients with myeloma. Prognostic factors were divided into three groups: factors useful in elderly patients, e.g., calcium, albumin; factors useful in young patients, e.g., platelet, creatinine, light-chain type; and factors useful in both patients, e.g., clinical stage, hemoglobin, LDH, CRP, bone marrow plasma cell and plasmablast percentages, light- and electron-microscopic classifications. The 5-year survival rates of elderly patients with calcium < 12 and > or = 12 mg/dl were 66.2 and < 11.1%, respectively (p<0.01). Those of the young patients were 64.1 and 33.3%, respectively. The 5-year survival rates of elderly patients with platelets > or = 200 x 10(9)/l and < 100 x 10(9)/l were 59.7 and 50.0%, respectively. Those of the young patients were 68.9 and 33.3%, respectively (p<0.05). The 5-year survival rates of elderly patients with few and numerous electron-microscopic abnormalities were 90 and 0%, respectively (p<0.01), those of young patients were 92.9 and < 14.3%, respectively (p<0.01). These findings suggest that individual clinical variables may differ in prognostic importance in elderly and young patients.

A staging system for multiple myeloma based on the morphology of myeloma cells.

The morphology of myeloma cells is reported to be one of the prognostic factors in multiple myeloma (MM) patients. We analyzed the prognostic factors, including morphological classification, in 292 patients with MM in order to select poor-risk patients who should be considered candidates for early intensive chemotherapy, including stem cell transplantation. Multivariate analysis was applied to 90 patients diagnosed between 1989 and 1996, because serum beta-2-microglobulin (beta2M) has been measured regularly since 1989, and showed that serum albumin, serum beta2M, and the morphology of myeloma cells predicted survival. According to these factors, patients were divided into 3 risk groups; a high-risk group (14%), a intermediate-risk group (46%) and a low-risk group (40%). There were significant differences between survival times in these 3 groups (median survival: high-risk, 16; intermediate-risk, 22; and low-risk, 44 months).

Ultrastructural abnormalities of bone marrow erythroblasts in refractory anemia.

Ultrastructural abnormalities of erythroblasts in 30 patients with refractory anemia (RA) according to FAB classification were studied. Nuclear clefts, iron-laden mitochondria, and myelin figures in cytoplasm were most frequently observed. Eleven patients (36.7%) with nuclear clefts of erythroblasts had a higher platelet count and higher incidence of cytogenetic abnormalities linked to poor prognosis than the patients without nuclear clefts. They also had a higher frequency of leukemic transformation and shorter survival than the patients without nuclear clefts. The nuclear clefts of erythroblasts, which are the result of impairment of DNA metabolism or nuclear formation and fusion during mitosis, may be one of the adverse prognostic factors for patients with RA.

Coincidental occurrence of pernicious anemia and mycosis fungoides in two elderly males.

We experienced two rare cases of pernicious anemia that presented in the course of mycosis fungoides in elderly males. Pernicious anemia has recently been reported to be caused by autoimmune gastritis that produces autoantibodies to gastric parietal cells and intrinsic factor. Immunological abnormalities in mycosis fungoides are reported to induce autoimmune diseases (i.e., autoimmune hemolytic anemia, anti-phospholipid antibody syndrome, arthritis, myasthenia gravis, necrotizing vasculitis, and vitiligo); the pernicious anemia in our two patients may have been closely related to the mycosis fungoides.

Action of 21-aminosteroid U74006F as an antioxidant against lipid peroxidation.

The dynamics of the action of the 21-aminosteroid U74006F as an antioxidant against lipid peroxidation were studied in organic solution and membranes. It was confirmed that the reactivities of this compound toward stable phenoxyl radical and peroxyl radical were quite low. In fact, U74006F did not exert appreciable antioxidant effect against the free radical-driven oxidation of methyl linoleate in acetonitrile solution. However, it suppressed the oxidation of phosphatidylcholine liposomal membranes into which it was incorporated in a concentration-dependent manner. The 21-aminosteroid U74006F did not exert any sparing effect on the rate of alpha-tocopherol consumption in the oxidation of methyl linoleate in solution, but when they were simultaneously incorporated into the membrane, U74006F spared alpha-tocopherol and exerted a synergistic effect against the oxidation of liposomal membranes. This suggests that lipophilic U74006F acts as an antioxidant against lipid peroxidation through a physicochemical and not a pure chemical mechanism, and that a physical interaction with the liposomal membrane may facilitate the inhibition of lipid peroxidation with U74006F.

Clinical analysis of the prognostic value of ultrastructural abnormalities of plasma cells in patients with multiple myeloma.

The prognostic value of ultrastructural abnormalities of plasma, cells was analyzed in 61 patients with multiple myeloma treated in our hospitals from 1985 to 1996. The survival curves demonstrated significant differences by stratification of ultrastructural abnormalities, such as irregular contour of the nuclear membrane, nuclear body, budding, disarrangement of organelles, fibrillar formation, intramitochondrial granules, multilamellar body, single-sac loop-like structure, and vacuole. However, categorizing by nuclear inclusion body, cytoplasmic inclusion body and dense body showed no significant difference in survival curves. Abnormalities, as well as maturity, may be useful in predicting the prognosis of myeloma patients.

Preoperative Diagnosis of Breast Diseases by Dynamic MR Mammography:Cut-off Point Establishment for Signal Intensity Ratio.

We have demonstrated that the dynamic study for breast lesions by magnetic resonance imaging(MRI)can differentiate benign from malignant lesions objectively.The cases were 57 histopathologically appraised breast lesions, including 20 cases of breast cancer, 28 cases of mastopathy, 8 cases of fibroadenoma and 1 caseof intraductal papillomatosis. We plotted time-signal intensity ratio curves and then determined 95% confidence intervals, plotting the signal intensity ratio for both breast cancer and mastopathy every 30 seconds during dynamic magnetic resonance mammography(MRM)after gadolinium-diethylenetriamine pentaacetic acid(Gd-DTPA)administration, and further established cut-off points to differentiate between them. We then tried of estimate objectively the benign-malignant differentiation to breast lesions by confirming their signal intensity ratio to be more or less than the cut-off points. We advocate this procedure, and call it the " dynamic ratio method. " As a result, we found highly significant differences between breast cancer and mastopathy at 30 and 60 seconds after Gd-DTPA administration(P < 0.0001). We also confirmed that the cut-off point for the dynamic ratio method was equivalent to 1.4 and 1.8 times the precontrast signal intensity value at30 and 60 seconds after administration of Gd-DTPA respectively. By performing this dynamic ratio method preoperatively we can assess objectively not only the malignancy of breast lesions, but also neighboring infiltration, extending intraductal component, and lymph node metastasis. Furthermore, the dynamic ratio method provides detailed information for selecting the appropriate region for breast conserving surgery preoperatively, and can be expected to reduce unnecessary biopsies of benign cases. The dynamic ratio method had a sensitivity of 95.0%, a specificity of 81.1% and a positive predictive value of 73.1%. Also, for detectinginvasive ductal carcinoma, the sensitivity of the dynamic ratio method was 100.0%.

Local structure study of dilute Er in III-V semiconductors by fluorescence EXAFS.

For understanding the luminescence of Er atoms in III-V semiconductors, OMVPE-grown InP doped with Er has been investigated by fluorescence EXAFS (extended X-ray absorption fine structure) in order to study the local structure around Er atoms. The local structures around the Er atoms doped in InP, with doping as dilute as 3 x 10(12) Er atoms in a 1.5 mm x 1.0 mm spot, were successfully measured by fluorescence EXAFS. The EXAFS analysis revealed that the Er atoms doped in InP above 853 K (which showed low luminescence) formed the rock-salt-structure ErP, while the Er atoms doped in InP below 823 K (which showed high luminescence) substituted on the In site of InP. The dependence of the local structure on growth temperature was observed for the samples doped with 3 x 10(12) atoms and 1.2 x 10(13) atoms of Er.

Severe thrombocytopenia suggesting immunological mechanisms in two cases of vivax malaria.

Case 1: A 27-year-old woman, referred to our hospital because of relapsing fever after travel to Thailand, was given a diagnosis of vivax malaria. Clinical investigation revealed thrombocytopenia, elevated platelet-associated IgG (PAIgG), and negative antibody against Plasmodium vivax antigen. After antimalarial treatment, the levels of both the platelets and PAIgG returned to normal. Case 2: A 28-year-old Sri Lankan man was admitted to our hospital with a complaint of fever. The patient had thrombocytopenia, elevated PAIgG, and positive antibody against Plasmodium vivax antigen. He contracted malaria in Sri Lanka about 6 months prior to this admission. After treatment, the platelet count and PAIgG level returned to normal. In these two cases, high levels of PAIgG may have been involved in the development of the thrombocytopenia. In the first patient, in particular, the thrombocytopenia was thought to be induced by some immunological mechanism prior to the detection of antimaralial antibodies in serum.

Inhibition of oxidation of low-density lipoprotein by a novel antioxidant, BO-653, prepared by theoretical design.

2,3-Dihydro-5-hydroxy-2,2-dipentyl-4,6-di-tert-butyl-benzofuran (BO-653) is a novel antioxidant synthesized by theoretical designing based on the previous experimental findings and consideration. The antioxidant activities of BO-653 against the oxidative modification of low-density lipoprotein (LDL) induced by free radicals were studied. BO-653 was consumed faster than endogenous alpha-tocopherol and inhibited the formation of lipid hydroperoxides, which was observed during the consumption of alpha-tocopherol. Doxyl stearic acids incorporated into LDL as spin probes competed with the antioxidants in scavenging radicals. It was found that the efficacy of radical scavenging by alpha-tocopherol became smaller as the radical went deeper into the interior of LDL particle, whereas that by BO-653 did not change. Ascorbic acid in the aqueous phase spared alpha-tocopherol efficiently during oxidation. On the other hand, the sparing effect of ascorbic acid for BO-653 was not remarkable, unlike that for alpha-tocopherol, which implied different locations of radicals derived from BO-653 and alpha-tocopherol within the LDL particle. It was concluded that BO-653 protected LDL from oxidative modification efficiently by scavenging peroxyl radicals and by reducing alpha-tocopheroxyl radicals and that this novel antioxidant might act as a potent inhibitor of development of atherosclerosis.

[Idiopathic CD4+ T-lymphocytopenia terminating in Burkitt's lymphoma].

A 33-year-old male was admitted because of severe neutropenia (55/microliter) in 1985. A diagnosis of autoimmune neutropenia was not made. CD4+ T-lymphocyte count was 128/microliter and CD4/CD8, 0.1, though the titers of immunoglobulins were in normal range. Nine times examinations of CD4+ T-lymphocyte count resulted in 5 times lower counts than 300/microliter between 1985 and 1993. His HIV (type 1 and type 2) studies were negative. He suffered from EB virus infection in 1987. The titers of VCA-IgG and EA-IgG had been elevated for half a year after that. He was operated polyp-like tumors in bilateral nasal cavities on February in 1994. The diagnosis was Burkitt's lymphoma that was also demonstrated by in situ hybridization using EB virus RNA. He was treated with a modification of COPBLAM III, but he died on June in 1994. Autopsy revealed tumor cell infiltrations in multiple organs. We suspect that he developed Burkitt's lymphoma 82 months later, when he had complicated EB virus infection in condition of idiopathic CD4+ T-lymphocytopenia caused by unknown origin.