Magnesium sulphate and perinatal mortality and morbidity in very-low-birthweight infants born between 24 and 32 weeks of gestation in Japan.

OBJECTIVE: Maternal exposure to magnesium sulphate has a neuroprotective effect in premature infants. This study aimed to examine this neuroprotective effect and the dose-response relationship in very-low-birthweight infants born between 24 and 32 weeks of gestation. STUDY DESIGN: A retrospective cohort study compared the rates of mortality and brain damage between three groups: no magnesium sulphate, low-dose (<50g) magnesium sulphate and high-dose (≥50g) magnesium sulphate. RESULTS: Japanese maternal and neonatal databases were linked using six key parameters from 2003 to 2007. Of 298,514 deliveries, 9101 were very-low-birthweight infants. Among these, full matching was possible for 5562 infants. Of the fully-matched infants, 3763 were born between 24 and 32 weeks of gestation, and 1813 (48%) were followed-up beyond 18 months. A multivariate analysis of the data, including gestational age, sex, fetal growth restriction, antenatal steroids and low pH (<7.1), showed that the low-dose group had no beneficial effects in terms of a reduction in mortality or incidence of brain damage (cerebral palsy or mental retardation). The high-dose group showed a significantly higher mortality rate [odds ratio (OR) 1.9, 95% confidence interval (CI) 1.2-2.9]. A stratified subgroup analysis of infants born between 28 and 32 weeks of gestation showed that survivors in the low-dose group had significantly lower rates of cerebral palsy (OR 0.4, 95% CI 0.2-0.98) and brain damage (OR 0.2, 95% CI 0.1-0.9), while the high-dose group did not show any significant changes. CONCLUSION: This study found that antepartum exposure to magnesium sulphate did not reduce the infant mortality rate or influence neurological outcomes. However, among infants born between 28 and 32 weeks of gestation, rates of cerebral palsy and brain damage were found to be significantly lower among survivors in the low-dose group.

Epigallocatechin-3-gallate (EGCG) affects the antioxidant and immune defense of the rainbow trout, Oncorhynchus mykiss.

Epigallocatechin-3-gallate (EGCG), a very potent antioxidant derived from green tea, was compared with vitamin E in terms of its effects on antioxidant defense and immune response of rainbow trout, by means of a feeding trial of eight weeks. Two of the experimental diets were supplemented with EGCG at either 20 or 100 mg kg(-1) diet (which contained only 30% of the intended levels) and the third was provided with 100 mg kg(-1) vitamin E but not EGCG. The control diet was not supplemented with the test components. Observation of tissue levels indicated that the high amount of EGCG helped to increase the availability of the lipid-soluble antioxidant vitamin E. The lower levels of lipid hydroperoxide in the liver of fish fed the higher amount of EGCG suggested that it was an effective antioxidant. Considering the immune indices, EGCG and vitamin E at 100 mg (actual amounts 31.9 and 94.1 mg kg(-1) diet, respectively) had identical capabilities in improving phagocytic activity and controlling hydrogen peroxide production by leucocytes. However, EGCG could possibly be more effective at enhancing serum lysozyme activity and the alternative complement activity. This work revealed the potential of EGCG as an antioxidant and an immunostimulant for rainbow trout, at least at the inclusion level of 32 mg kg(-1) diet.

Enhancement of innate immunity in rainbow trout (Oncorhynchus mykiss Walbaum) associated with dietary intake of carotenoids from natural products.

The effects of orally administered carotenoids from natural sources on the non-specific defense mechanisms of rainbow trout were evaluated in a nine-week feeding trial. Fish were fed four diets containing either beta-carotene or astaxanthin at 100 and 200 mg kg-1 from the marine algae Dunaliella salina and red yeast Phaffia rhodozyma, respectively, and a control diet containing no supplemented carotenoids. Specific growth rate and feed:gain ratio were not affected by dietary carotenoid supplementation. Among the humoral factors, serum alternative complement activity increased significantly in all carotenoid supplemented groups when compared to the control. On the other hand, serum lysozyme activity increased in the Dunaliella group but not in the Phaffia group, whereas plasma total immunoglobulin levels were not altered by the feeding treatments. As for the cellular responses, the superoxide anion production from the head kidney remained unchanged while the phagocytic rate and index in all supplemented groups were significantly higher than those of the control. These findings demonstrate that dietary carotenoids from both D. salina and P. rhodozyma can modulate some of the innate defense mechanisms in rainbow trout.

Nonspecific immune response of rainbow trout (Oncorhynchus mykiss Walbaum) in relation to different status of vitamin E and highly unsaturated fatty acids.

This study was designed to examine the effects of dietary vitamin E (VE) on modulation of immune responses when supplied with two levels of n-3 highly unsaturated fatty acids (n-3 HUFA) in rainbow trout, Oncorhynchus mykiss. Six semipurified diets were prepared containing three levels of dietary VE (0, 100 or 1000 mg alpha-tocopheryl acetate kg(-1) diet) and n-3 HUFA either at 20 or 48% of dietary lipid provided from fish oil or docosahexaenoic acid (DHA) concentrated fish oil respectively. The diets were fed to rainbow trout (100 g initial mean weight) for 15 weeks. The VE, vitamin C (VC) content in plasma and tissues and the nonspecific immune responses, both humoral (alternative complement activity, total immunoglobulin) and cellular (phagocytosis, nonspecific cytotoxicity) were examined. VE contents in the kidney reflected the dietary input but were lower in fish fed 48% n-3 HUFA diets, and could have impaired some of immune responses compared to fish fed 20% n-3 HUFA. VC contents in kidney followed the same pattern as VE. Both humoral and cellular immune functions deteriorated in fish fed VE deficient diets whereas improvement in most of the parameters corresponded to its supplementation. However, the higher dose of dietary VE did not substantially enhance the responses assayed compared to the 100 mg dose. Besides clearly indicating the role of VE in maintaining the immune functions in fish in relation to dietary n-3 HUFA, this study has revealed that optimum health benefits could be achieved when VE is maintained slightly above the levels generally recommended for normal growth.

Absence of arabinan in the side chains of the pectic polysaccharides strongly associated with cell walls of Nicotiana plumbaginifolia non-organogenic callus with loosely attached constituent cells.

When leaf disks from haploid plants of Nicotiana plumbaginifolia Viv. were transformed with T-DNA and cultured on shoot-inducing medium, nonorganogenic callus. designated nolac (for non-organogenic callus with loosely attached cells), appeared on approximately 7% of leaf disks. In contrast, normal callus was generated on T-DNA-transformed leaf disks from diploid plants and on non-transformed leaf disks from haploid and diploid plants. Transmission electron microscopy revealed that the middle lamellae and the cell walls of one line of mutant callus (nolac-H14) were barely stained by ruthenium red. even after demethylesterification with NaOH, whereas the entire cell wall and the middle lamella were strongly stained in normal callus. In cultures of nolac-H14 callus, the level of sugar components of pectic polysaccharides in the hemicellulose fraction was reduced and that in the culture medium was elevated, as compared with cultures of normal callus. These results indicate that pectic polysaccharides are not retained in the cell walls and middle lamellae of nolac-H14 callus. In nolac-H14, the ratio of arabinose to galactose was low in the pectic polysaccharides purified from all cell wall fractions and from the medium, in particular, in the hemicellulose fractions. The low levels of arabinofuranosyl (T-Araf, 5-Araf, 2,5-Araf, and 3,5-Araf) residues in the pectic polysaccharides of the hemicellulosic fraction of nolac-H,14 indicated that no neutral-sugar side chains, composed mainly of linear arabinan. were present in nolac-H14. Arabinose-rich pectins. which are strongly associated with cellulose-hemicellulose complexes, might play an important role in intercellular attachment in the architecture of the cell wall.

[Effects of cernitin pollen-extract (Cernilton) on inflammatory cytokines in sex-hormone-induced nonbacterial prostatitis rats].

Cernitin pollen-extract (Cernilton, CN) is a preparation made from eight kinds of pollen and has been used for various prostatic diseases in Japan and Europe. We reported previously that CN possessed a recovery action on the sex-hormone-induced nonbacterial prostatitis in rats. To clarify the possible mechanism of action of CN, we investigated the effects of CN on inflammatory cytokines (IL-1 beta, IL-6 and TNF-alpha) in the same model. Aged Wistar rats were castrated and injected 17 beta-estradiol (0.25 mg/kg/day, s.c.) for 30 days. CN (630 and 1,260 mg/kg, p.o.) or testosterone (2.5 mg/kg, s.c.) was administered for the last 14 days of the treatment of 17 beta-estradiol. In control rats, prostatic IL-6 and TNF-alpha contents were increased approximately 2-3 fold, and acinar glandular inflammation and stromal proliferation were found histopathologically, as compared with those of intact rats. On the other hand, CN decreased the increased contents of cytokines in a dose-dependent manner. The histopathological changes mentioned above were restored in rats treated with 1,260 mg/kg. Testosterone also ameliorated them significantly. These results indicate that CN has an anti-inflammatory action, and that the inhibitory effect of CN on the prostatic inflammatory cytokine is an important factor in its action.

Identification, characterization, and functional analysis of heart-specific myosin light chain phosphatase small subunit.

Myosin light chain phosphatase consists of three subunits, a 38-kDa catalytic subunit, a large 110-130-kDa myosin binding subunit, and a small subunit of 20-21 kDa. The catalytic subunit and the large subunit have been well characterized. The small subunit has been cloned and studied from smooth muscle, but little is known about its function and specificity in the other muscles such as cardiac muscle. In this study, cDNAs for heart-specific small subunit isoforms, hHS-M(21), were isolated and characterized. Evidence was obtained from an analysis of genome to suggest that the small subunit was the product of the same gene as the large subunit. Using permeabilized renal artery preparation and permeabilized cardiac myocytes, it was shown that the small subunit increased sensitivity to Ca(2+) in muscle contraction. It was also shown using an overlay assay that hHS-M(21) bound the large subunit. Mapping experiments demonstrated that the binding domain and the domain involved in the increasing Ca(2+) sensitivity mapped to the same N-terminal region of hHS-M(21). These observations suggest that the heart-specific small subunit hHS-M(21) plays a regulatory role in cardiac muscle contraction by its binding to the large subunit.

A new model of cerebral microthrombosis in rats and the neuroprotective effect of a Rho-kinase inhibitor.

BACKGROUND AND PURPOSE: The aim of this study was to develop a new model of stroke based on endothelial damage and thrombotic occlusion in a perforating artery, leading to small cerebral infarcts and neurological deficits in rats. Moreover, the neuroprotective efficacy of fasudil, a rho-kinase inhibitor, was investigated in this model. METHODS: Fifty-six male Sprague-Dawley rats were used in the present study. Rats were anesthetized with sodium pentobarbital, and 100 microg of sodium laurate was injected into the left internal carotid artery on days 1 and 3. The thrombus induction and consequent of ischemic brain damage were examined by histopathological analyses and neurological deficit scoring in a posture reflex test. To investigate the neuroprotective effects of fasudil, 1 or 10 mg/kg was administered intraperitoneally 5 minutes after the first injection of sodium laurate and once daily thereafter on the following 2 days. RESULTS: One hour after the injection of sodium laurate, microscopic examination of phosphotungstic acid hematoxylin-stained sections (n=5) revealed that microthrombi containing fibrin strands obstructed the perforating arteries in the ipsilateral hemisphere. Under a transmission electron microscope (n=6), endothelial cells appeared exfoliated and the vascular lumen was obstructed by a thrombus composed of degranulated platelets, fibrin, leukocytes, and erythrocytes. No evidence of endothelial cell damage or thrombus could be found in the ipsilateral side of the pial artery (middle cerebral artery). Twenty-four hours after the second injection of sodium laurate (day 4), 13 of 15 rats (86.6%) showed mild to severe neurological deficits. Multiple small cerebral infarcts were observed in the hippocampus, cortex, and thalamus. Treatment with fasudil (1 and 10 mg/kg, n=15 each) resulted in a significant improvement in neurological deficits. Fasudil also significantly reduced the area of cerebral infarction. CONCLUSIONS: We present a new model of stroke in rats, in which the perforating arteries are selectively occluded by microthrombi. This model is useful to investigate the pathophysiology and treatment of small cerebral infarction, which is caused by perforating arterial occlusive diseases such as lacunar infarcts. Fasudil may be beneficial in the treatment of acute ischemic stroke.

Vascular tissue-specific gene expression of xylem sap glycine-rich proteins in root and their localization in the walls of metaxylem vessels in cucumber.

Root-specific cDNAs of glycine-rich protein (cucumber root glycine rich protein-1 and -2; CRGRP-1 and CRGRP-2) were cloned previously by use of an antiserum raised against whole xylem sap of Cucumis sativus. The accumulation of the corresponding mRNA at high levels was detected in the root-hair zone of cucumber tap root [Sakuta et al. (1998) Plant Cell Physiol. 39: 1330]. The RNA gel blot analysis with the CRGRP-1- and -2-specific probes revealed that the CRGRP genes expressed only in root but not at all in aboveground organs. When the localization of these mRNAs were examined by in situ hybridization, CRGRP mRNAs were found only in the parenchyma cells in the central cylinder of young lateral roots and it was most abundant in the cells that surrounded xylem vessels in the root-hair zone of the tap root. In immunoblotting of xylem sap collected from cucumber stem with an antiserum raised against CRGRP-1 that had been produced in an E. coli expression system, the antibodies, which did not cross-react with GRP1.8 of kidney bean, reacted with two proteins, whose mobilities corresponded to those of proteins deduced from the CRGRP-1 and -2 cDNAs. Immunohistochemical staining revealed that the CRGRPs accumulated specifically in the lignified walls of metaxylem vessels in the root, stem and leaf and in the lignified cell walls of perivascular fibers in cucumber stems. Immunostaining was also detected in the walls of metaxylem vessels and in the cell walls of adjacent sclerenchyma in the hypocotyl of kidney bean. These data clearly indicate that the novel glycine-rich proteins were produced in the vascular tissue of the root, transported systemically over a long distance via the xylem sap and immobilized in the walls of metaxylem vessels and sclerechyma cells in aboveground organs.

[Consultation and survey for drug interaction in outpatients taking the medicines potentially interact with St. John's Wort].

Ministry Health Welfare of Japan announced the caution for drug interaction of St. John's Wort (SJW), a herbal supplement occasionally used for depression, on May, 2000. Immediately after the announcement, we conducted drug consultation for outpatients prescribed the medicines potentially interacting with SJW. We provided information concerning possible drug interaction with SJW for 741 outpatients (except for pediatrics) during the period of May 22-June 16, 2000. The potential drugs prescribed frequently were warfarin (28.0%), theophylline (19.7%), digitalis (18.4%), carbamazepine (7.2%), disopyramide (6.9%) and cyclosporin (6.3%). Of the patients, 401 subjects were surveyed by collecting the questionnaires to clarify the background of SJW drug interaction. Twenty-two subjects (5.5%) have known commercially available SJW products, 5 subjects (1.2%) have ever taken SJW products before and 2 subjects (0.5%) have taken SJW products concomitant with prescribed medicines. Gender difference was observed in paying attention to SJW products; female subjects (8.6%) tended to have more interest in SJW products than male subjects (2.8%). Two subjects taking SJW have realized for the first time that the supplements they took were SJW products when their package photographs were shown at the consultation. Showing the package photographs might be helpful for making the patients easy to identify the SJW products, because most patients do not pay attention to whether the supplements contain SJW or not. It is recommended that drug consultation should be provided to avoid serious drug interaction with SJW while the outpatients are taking potential medicines prescribed.

A novel class of orally active non-peptide bradykinin B2 receptor antagonists. 2. Overcoming the species difference between guinea pig and man.

Recently we reported the identification of a series of 8-[[3-(N-acylglycyl-N-methylamino)-2, 6-dichlorobenzyl]oxy]-3-halo-2-methylimidazo[1,2-a]pyridines as the first orally active non-peptide bradykinin (BK) B2 receptor antagonists (1-3). These compounds inhibited the specific binding of [3H]BK to guinea pig ileum membrane preparations expressing B2 receptors with nanomolar IC50's and also displayed in vivo functional antagonistic activities against BK-induced bronchoconstriction in guinea pigs at 1 mg/kg by oral administration. However, it was found that their affinities for the B2 receptors in human A-431 cells (human epidermoid carcinoma) were much lower. Intensive modifications of the terminal substituents at the glycine moiety elucidated the structure-activity relationships (SAR) for human B2 receptors, leading to an extended basic framework which incorporated a novel key pharmacophore. Thus, we overcame the species difference and identified the first clinical candidate 18c (FR167344) with IC50's of 0.66 and 1.4 nM for guinea pig ileum and human A-431 cells, respectively. This compound displayed in vivo functional antagonistic activity against BK-induced bronchoconstriction in guinea pigs with an ED50 value of 0.17 mg/kg by oral administration. This novel non-peptide B2 antagonist is extremely potent both in vitro and in vivo by oral administration and is expected to be the first member of a new class of drug for the treatment of various inflammatory diseases.

A novel class of orally active non-peptide bradykinin B2 receptor antagonists. 3. Discovering bioisosteres of the imidazo[1,2-a] pyridine moiety.

Recently we reported on overcoming the species difference of our first orally active non-peptide bradykinin (BK) B2 receptor antagonists, incorporating an 8-[[3-(N-acylglycyl-N-methylamino)-2, 6-dichlorobenzyl]oxy]-3-halo-2-methylimidazo[1,2-a]pyridine skeleton, leading to identification of the first clinical candidate 4a (FR167344). With this potent new lead compound in hand, we then investigated further refinement of the basic framework by replacement of the imidazo[1,2-a]pyridine moiety and discovered several bioisosteric heterocycles. Extensive optimization of these new heteroaromatic derivatives revealed the detailed structure-activity relationships (SAR) around the imidazo[1, 2-a]pyridine ring and the 2,6-dichlorobenzyl moiety, leading to the discovery of our second clinical candidate 87b (FR173657) which inhibited the specific binding of [3H]BK to recombinant human B2 receptors expressed in Chinese hamster ovary (CHO) cells and guinea pig ileum membrane preparations expressing B2 receptors with IC50's of 1.4 and 0.46 nM, respectively. This compound also displayed excellent in vivo functional antagonistic activity against BK-induced bronchoconstriction in guinea pigs with an ED50 value of 0.075 mg/kg by oral administration. Further modifications of the terminal substituents on the pyridine moiety led to a novel pharmacophore and resulted in the identification of 99 (FR184280), whose IC50 value for human B2 receptors (0.51 nM) was comparable to that of the second-generation peptide B2 antagonist Icatibant.

Cervical spinal epidural abscess following acupuncture: successful treatment with antibiotics.

A 67-year-old man with poorly controlled diabetes mellitus (DM) had acupuncture several times a month for chronic shoulder muscle stiffness. A few days after acupuncture in the posterior nuchal region, a low-grade fever and backache developed, and subacutely progressed. Finally he complained of gait disturbance, and then respiratory distress appeared. Magnetic resonance imaging (MRI) demonstrated high cervical epidural abscess with massive soft tissue inflammation and vertebral osteomyelitis. Conservative treatment with antibiotics was effective and it was well documented by following serial MRIs. This case suggested that needle acupuncture should be avoided for immunocompromised subjects such as patients with poorly controlled DM.

[Arterial infusion chemotherapy with SMANCS-Lipiodol for multiple hepatocellular carcinoma].

Fifty-five patients with hepatocellular carcinoma were treated with oily anticancer agent SMANCS dissolved in Lipiodol (SMANCS-LPD). The local response rate after the first arterial infusion in all patients was 39%, against 63% in 27 patients with Lipiodol accumulation occupying more than two thirds of tumor areas. The infusion therapy with SMANCS-LPD is adapted for a vascular-rich hepatocellular carcinoma. An infusion of 4 mg of SMANCS was ineffective for patients with tumors distributing in bilateral lobes of liver. Thus, an increase of infusion dosage or repeated infusions were recommended for such cases.

Root-specific expression of genes for novel glycine-rich proteins cloned by use of an antiserum against xylem sap proteins of cucumber.

Candidate cDNAs for xylem sap proteins (XSPs) were isolated by screening of a cucumber cDNA library prepared from poly(A)+ RNA of cucumber roots with an antiserum raised against the XSPs of cucumber. The patterns of expression of corresponding transcripts and the sequences of the cDNAs were examined. Two cDNAs with a glycine-rich domain in each deduced amino acid sequence and whose transcripts were expressed predominantly in roots were selected for further analysis. Both cDNAs encoded proteins with a putative signal sequence, with similar non-glycine-rich domains at the amino terminal of the encoded polypeptides. The corresponding mRNAs accumulated concurrently with the formation of adventitious roots from cuttings of cucumber hypocotyls and the development of vascular tissues, and strong expression of the mRNAs was detected in the root-hair zone of tap roots. These data suggest that the production of the two novel glycine-rich proteins, which are putative XSPs, might be associated with the functions of the vascular tissues in roots.

[Chemoradiation for rectal cancer].

Based on the results of the Study Group for Surgical Adjuvant Radiochemotherapy for Rectal Cancer and the Study Group for Surgical Therapy and Combined Irradiation in Rectal Cancer, we examined the clinical aspects of the relatively new field of chemoradiation for rectal cancer and examined improvements in the therapeutic results. Furthermore, we examined the future outlook with particular regard to selection of therapeutic drugs and autonomic nerve preserving operation.

[Arterial infusion chemotherapy with SMAN CS-Lipiodol for hepatocellular carcinoma evaluation of infusion method].

Forty-four patients with hepatocellular carcinoma were treated with oily anticancer agent SMANCS dissolved in Lipiodol (SMANCS-LPD). The local response rate after the first arterial infusion in all patients was 39%, against 63% in 27 patients with Lipiodol accumulation occupying more than two third of tumor areas. Repeated arterial infusion of SMANCS-LPD did not enhance the therapeutic effect. An infusion of 4mg of SMANCS was ineffective for patients with tumors distributing in bilateral lobes of liver, and 6 mg was recommended for such cases.

Antimutagenicity of Tochu tea (an aqueous extract of Eucommia ulmoides leaves): 1. The clastogen-suppressing effects of Tochu tea in CHO cells and mice.

The suppressing effect of crude extracts of Tochu tea, an aqueous extract of Eucommia ulmoides leaves and a popular beverage in Japan, on the induction of chromosome aberrations in CHO cells and mice was studied. When CHO cells were treated with Tochu tea crude extract after MMC treatment, the frequency of chromosome aberrations was reduced. Out of 17 Tochu tea components, 5 irridoids (geniposidic acid, geniposide, asperulosidic acid, deacetyl asperulosidic acid, and asperuloside) and 3 phenols (pyrogallol, protocatechuic acid, and p-trans-coumaric acid) were found to have anticlastogenic activity. Since the anticlastogenic irridoids had an alpha-unsaturated carbonyl group, this structure was considered to play an important role in the anticlastogenicity. The anticlastogenic effect of Tochu tea extracts was examined in mice using a micronucleus assay. When mice received 1.0 ml 4% Tochu tea extract by oral gavage 6 h before intraperitoneal injection of MMC, a decrease in the frequency of micronuclei was observed. This decrease was not due to a delay in the maturation of micronucleated reticulocytes.

Effect of isosorbide on distortion-product otoacoustic emissions and endocochlear DC potential in experimentally induced hydropic ears.

The effects of isosorbide on distortion-product otoacoustic emissions (DPOAEs) and endocochlear DC potential (EP) were examined in experimentally induced endolymphatic hydropic ears and untreated control ears using 20 albino guinea pigs. DPOAEs and EP in the hydropic ears decreased after obliteration of the endolymphatic duct and sac. The administration of isosorbide restored DPOAEs to the normal level, whereas EP was further reduced. In control ears there was no significant change in DPOAEs and EP after the administration of isosorbide. The results obtained in the present study indicate that the effect of isosorbide on DPOAEs is different from that on EP, and the recovery of DPOAEs in hydropic ears is not a secondary phenomenon due to the recovery of EP.

A case of possessive state with onset influenced by 'door-to-door' sales.

Recently in Japan, 'door-to-door sales' has become of concern because it has created numerous legal and social problems. In this paper, a 47 year old dissociative trance disorder case who presented with possession by God is discussed. Specific types of door-to-door sales is known to use superstition and folk beliefs as tools to lure customers. In this particular case, these religious factors seemed to have played an important role in the precipitation of the disorder and its presentation. In addition, the brain-washing environment observed in video lectures used in door-to-door sales seemed to play an important role in the development of the possessive state. We also performed social psychiatric analysis of the occurrence of the possessive state in a city area, which has been considered to develop within traditional culture. Phenomenological classification by one of the authors was useful for diagnosing underlying disorders in the possessive state.