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Large amounts of rice husk ash (RHA) are discharged from rice husk power plants, and the development of an effective system for recycling this RHA waste would be desirable. In the current study, the silica component of RHA obtained from a rice husk power plant in Myanmar was successfully used as raw material for the synthesis of the aluminosilicate nanotube (ASNT) material. The RHA used contained over 90 wt% of silica. The synthesized ASNT had a web-like structure formed of fi brous bundles, each containing many individual ASNT.

Nanosized Mg-Al layered double hydroxides (LDHs) with Cl−, NO3− or SO42− as the interlayer anion have been synthesized by a modified coprecipitation method. The obtained LDHs were confirmed to be composed of a single phase and to be highly substituted by Al (Mg/Al ratio ∼1.9). The abilities of the LDHs to adsorb several harmful anions (F−, CrO42−, HAsO42− and HSeO3−) in aqueous solution were studied. The LDHs exhibit high adsorption abilities. The amount of adsorption onto the LDHs differed between the starting interlayer anions, and decreased in the following order of the interlayer anions: NO3− > Cl− > SO42−. The NO3-formed Mg-Al LDH reached a CrO42− adsorption equilibrium state within only 30 min, much faster than those in previous reports. Thus, the nanocrystallized, highly Al substituted phase of the NO3-formed Mg-Al LDH is found to markedly enhance the anion adsorption ability.

We describe the prevalence of community-acquired M. pneumoniae pneumonia diagnosed by culture methods in a single institute in Japan from January 1979 to December 1999. Cultures were performed in 2971 pneumonia cases and yielded M. Pneumoniae in 508 cases. The epidemic peaks recurred regularly at 4-year intervals (1980, 84, 88 and 91–2). Although a large epidemic has not occurred since 1992, traces of epidemic periodicity have still persisted from 1992 to 1999 at 3-year intervals.

Methods HIV-specific proliferative CD4+ T cell responses were assessed by CFSE assays performed after stimulation with recombinant HIV p24 in the presence of a blocking antiIL10R antibody or an isotype control, with or without depletion of CD25+ cells. IL-10 production was measured in cell subsets by ICS or by luminex of culture supernatants from PBMCs depleted of CD14+, CD19+ or CD25+ cells. Finally, monocytes and regulatory T cells were sorted and cultured alone or together in transwell plates and secreted IL-10 was measured.

1. The spatial patterns of reflexes elicited by localized pressure stimulation of the foot skin were analysed by recording electromyographic activities of various hindlimb muscles or muscle nerve discharges in cats anaesthetized with sodium pentobarbitone. 2. Reflex discharges evoked by stimulation of the central pad occurred mainly in physiological toe extensors located in the foot. Stimulus-response relationships of single motor units revealed characteristically wide ranges of graded response and recruitment. 3. Within the central pad, the strongest excitation was evoked from the central lobe and was distributed to extensors of all four toes. Excitation from the medial and the lateral lobes was usually asymmetrical and weaker in toe muscles of the stimulated side. It is suggested that the weakness was in part due to concomitant inhibition. 4. Stimulation of a toe pad caused marked suppression of central pad-evoked activity of toe extensors with a highly specific spatial pattern. The inhibition was strongest in extensors of its own toe, and gradually weaker in the more distant toes. Weak excitation was occasionally evoked in extensors of the most medial or lateral toes, when the most lateral or the most medial toe-pad, respectively, was stimulated. 5. A similar pattern of relfex to that from the toe pad was evoked from the claw base and the hairy toe dorsum of each digit. 6. Reflex effects, both inhibitory and excitatory, from the central and toe pads, claw bases and toe dorsum were maintained during prolonged stimuli, indicating that slowly adapting receptors contributed to these reflexes. 7. It is concluded that stimulation of localized skin areas of the foot, particularly the pads, evokes highly specialized reflexes, which may be important in controlling movements of individual digits.

New antitumor agents must be added to the current neuroblastoma treatment regimens to improve the clinical results. We investigated whether recombinant human endostatin (rhEndostatin), an antiangiogenic agent, is effective against human neuroblastoma in the human neuroblastoma xenograft model designated TNB9. When tumors on the back of nude mice grew to a weight of 90-95 mg, rhEndostatin 10 mg/kg/day was administered subcutaneously every day for 10 consecutive days. Mean relative tumor weight in mice administered rhEndostatin (n=5) was significantly less than that in controls (n=12) on days 2, 4, and 6 after the start of administration (p

The expression of Fas, a cell surface receptor directly responsible for triggering cell death by apoptosis, and its ligand (FasL) was investigated on both human colonic intraepithelial T lymphocytes (IELs) and peripheral blood mononuclear lymphocytes (PBMLs). FACS analysis indicated that IELs have increased expression of Fas compared with PBMLs, together with the progress activation marker, CD45RO. A discrete fraction of freshly isolated IELs also constitutively expressed FasL, perhaps as a result of recent in vivo activation. Using monoclonal antibody APO2.7, which detects mitochondrial 7A6 antigen specifically expressed by cells undergoing apoptosis, we further investigated the apoptosis-inducing effect of anti-Fas monoclonal antibody (CH11) on both IELs and PBMLs. FACS analysis revealed that CH11 increased the percentage of apoptotic cells, in IELs but not in PBMLs. Culture with anti-FasL monoclonal antibody (4H9) significantly recovered cell viability in IELs, but not in PBMLs. These results indicate that IELs constitutively express both Fas and FasL and that Fas crosslinking generates signals resulting in apoptosis, outlining a potential mechanism involved in intestinal tolerance.

In response to 1-methyladenine, a maturation-inducing substance, starfish oocytes undergo reinitiation of meiosis with germinal vesicle breakdown through activation of p34cdc2-cyclin B, which results in the dispersal of the nucleolus. Little information has been elucidated thus far on nucleolar proteins that are phosphorylated by p34cdc2-cyclin B during meiotic maturation. Here, we describe a novel nucleolar protein of the starfish Asterina pectinifera oocyte, which is designated ANO39 and which is phosphorylated during meiotic maturation. A full-length ANO39 cDNA of 2106 base pairs encodes a polypeptide of 346 amino acids having a calculated Mr of 39 005. The amount of ANO39 is kept nearly constant during oocyte maturation and embryogenesis up to the midgastrula stage. The transcript encoding ANO39 was present in growing oocytes but not in full-grown ones, as evidenced by Northern blot hybridization. Ser145 is specifically phosphorylated when ANO39 is incubated in vitro with purified starfish p34cdc2-cyclin B. This phosphorylation site corresponds to that is phosphorylated during meiotic maturation in vivo. Immunoblot analysis using phosphoserine145-specific antibody as a probe revealed that some populations of ANO39 of the immature oocytes at the G2 stage have been already phosphorylated on Ser145 and Ser145 is maximally phosphorylated during meiotic maturation.

SUMMARYIntestinal intraepithelial lymphocytes (IEL) constitute the first lymphoid compartment to encounter dietary antigens and intestinal pathogens. IEL are proposed to be involved in the defence against bacterial and viral invasion and to play an important role in mucosal immunity. Fas (CD95/APO-1) is a surface receptor that induces apoptotic cell death upon ligation with Fas ligand (FasL). The aim of this study was to examine the expression and function of Fas and FasL on freshly isolated normal human colonic IEL. The expression and function of Fas and FasL on IEL isolated from 40 normal colonic specimens were examined by flow cytometry, reverse transcriptase-polymerase chain reaction, immunohistochemistry, and DNA-release cytotoxicity assay. Virtually all CD3+ IEL (95.2 ± 4.3%) expressed Fas and were sensitive to agonistic anti-Fas antibody, whereas only 56.6 ± 8.4% of peripheral T lymphocytes expressed Fas and were resistant to the antibody. We also detected FasL mRNA and protein (40.1 ± 4.2%) on IEL, and found that IEL exerted FasL-mediated cytotoxicity against Fas-expressing target cells. These findings suggest that human IEL are activated in situ but are tightly regulated by the constitutive expression of functional Fas and FasL to maintain homeostasis of the mucosal immune system.

We have investigated the frequency of p53 gene mutations in Ewing's sarcoma (ES) and neuroblastoma (NB) by using polymerase chain reaction-single strand conformation polymorphism analysis for genomic DNA or complementary DNA generated from total RNA. Mutations of the p53 gene were found in six of seven ES cell lines: a missense mutation of TGC (Cys)--TAC (Try) at codon 141 in one, a missense mutation of CGT (Arg)--TGT (Cys) at codon 273 in one, a missense mutation of TGC (Cys)--TTC (Phe) at codon 176 in three, and one base deletion of CGC--CG at codon 283 in one. Further analysis of 14 ES and related primary tumors showed mutations of the p53 gene in only two: one base insertion of CCG--CCCG at codon 152 in one and a missense mutation of GGC (Gly)--GTC (Val) at codon 154 in the other. Both of the two tumors were obtained from patients with an advanced stage disease. Three of the eight ESs with mutations of the p53 gene showed the same missense mutation at codon 176, suggesting the mutational hot spot of the p53 gene in ESs. In contrast to ES, none of 6 NB cell lines or 48 NB tumors including advanced-stage ones with or without N-myc amplification showed any aberration of the p53 gene. Our findings suggest that mutations of the p53 gene in ES might represent late genetic events related to tumor progression, and that aberrations of the p53 gene might not be involved in the development or the progression of NB.

Parathyroid hormone-related protein (PTHrP), which is responsible for producing hypercalcemia in patients with humoral hypercalcemia of malignancy, has recently been identified in several normal tissues. Because PTHrP, like parathyroid hormone (PTH), is known to exhibit vasodilatory properties, we investigated the expression and regulation of PTHrP mRNA in cultured rat aortic smooth muscle cells (SMC). We report here that PTHrP mRNA is expressed in SMC and is markedly induced by serum in a time- and concentration-dependent fashion. Addition of 10% fetal calf serum to serum-deprived, confluent cells, resulted in a marked induction of PTHrP mRNA by 2 h with a peak at 4-6 h. PTHrP was detected in SMC by immunocytochemistry and radioimmunoassay of conditioned medium, and was shown to be up-regulated within 24 h after the addition of serum. The serum induction of PTHrP mRNA was blocked by actinomycin D and by cycloheximide indicating the need for protein synthesis to evoke the serum effect on PTHrP gene transcription. In addition, treatment with dexamethasone, which has been previously shown to reduce the constitutive expression of PTHrP in human cancer cells, also blunted the serum induction of PTHrP mRNA in SMC. Treatment of quiescent cells with the serum mitogens platelet-derived growth factor or insulin-like growth factor-I had no effect on PTHrP, whereas the vasoactive peptides endothelin, norepinephrine and thrombin stimulated PTHrP expression. Exogenous addition of recombinant PTHrP-(1-141) had no significant effect on SMC DNA synthesis as measured by [3H]thymidine incorporation. In summary, the abundance of PTHrP mRNA and the characteristics of its regulation in SMC suggest a major role for PTHrP as a local modulator in vascular smooth muscle.

The capacity of the v-myc-transformed, chicken myelomonocytic cell line HD-11 to metabolize 25-hydroxyvitamin D3 (25-OHD3) was examined. HD-11 cells produced and secreted a metabolite of 25-OHD3 that was bound with high affinity by receptor for 1,25-dihydroxyvitamin D3 [1,25-(OH)2D3]. On normal-phase HPLC, this metabolite cochromatographed with authentic 1,25-(OH)2D3 in both hexane- and methylene chloride-based solvent systems. The 25-OHD3 1-hydroxylation reaction was substrate saturable with a Km of 73 nM 25-OHD3 and a maximal velocity of 167 fmol per 10(6) cells per h. This reaction was inhibited by ketoconazole, a recognized inhibitor of cytochrome P450 mixed-function oxidases including the authentic, renal 25-OHD3 1-hydroxylase. On the other hand, HD-11 cell 1,25-(OH)2D3 production was not affected by the antioxidant DPPD, a known inhibitor of free radical-generated 1,25-(OH)2D3. In addition to synthesizing 1,25-(OH)2D3, this monocyte-macrophage cell line also has the potential to be a target for the hormone; HD-11 cells express high-affinity receptor for 1,25-(OH)2D3 (Kin = 0.06 nM).

A (H+ + K+)-ATPase-enriched membrane fraction derived from the fundic portion of hog gastric mucosa was obtained by a combination of differential and repeated 7% Ficoll gradient centrifugation. The microsomal membrane fraction isolated by repeated 7% Ficoll gradient centrifugation was free of ouabain-sensitive (Na+ + K+)-ATPase, 5'-nucleotidase and succinate dehydrogenase; and it was highly enriched in (H+ + K+)-ATPase and K(+)-stimulated p-nitrophenylphosphatase (p-NPPase). The (H+ + K+)-ATPase had a pH optimum of 7.4 and was stimulated by Tl+, K+, Rb+ and NH4+ with Ka values of 0.0667, 0.526, 0.667 and 3.03 mM, respectively, at this pH. On the other hand, monovalent cations such as Na+, Li+ and (CH3)4N+ as well as divalent cations such as Cu2+, Ca2+, Ba2+, Sr2+ and Cd2+ inhibited this enzyme activity concentration-dependently. Ouabain and oligomycin had no effect, whereas omeprazole, a specific (H+ + K+)-ATPase inhibitor, inhibited this enzyme activity in a pH-dependent manner. Sodium dodecyl sulfate (SDS)-polyacrylamide gel electrophoresis showed a major band (greater than or equal to 90% of protein) at 97,400 daltons, which was phosphorylated in the presence of Mg2+ and [gamma-32P]-ATP and dephosphorylated in the presence of K+. The present method was very simple, and the (H+ + K+)-ATPase activity of the microsomal fraction obtained by this method was much higher compared with those obtained by other methods such as free-flow electrophoresis.

The pattern of la-connections to motor nuclei of 17 hindlimb muscles (or groups of muscles) has been investigated in baboons by intracellular recording of Ia-e.ps.p.s evoked in motoneurons from different muscle nerves. The amplitudes are normalized to 70 mV resting potential and compared with similarly normalized Ia-e.ps.p.s in cats. As in the cat, la-excitation is drawn from a restricted number of muscles and the homonymous effect is usually dominating. Heteronymous connections to many motor nuclei are different in the two species. For example, hip extensors are generally more la-isolated from each other in baboons than in cats, and also knee flexors have fewer Iainterconnections than in cats. A unidirectional Ia-synergism between some hip extensors and knee flexors in cats has changed to a bidirectional one in baboons, with a tendency to lateralization of the connections. Among ankle extensors, soleus has smaller heteronymous I a-connections from its synergic ankle extensors than in cats. In baboons, plantaris is heteronymously I a-excited from gastrocnemius-soleus but not from the intrinsic plantar muscles; whereas in cats there exists a considerable la-projection from the intrinsic plantar muscles but not from gastrocnemius-soleus. There is a corresponding difference in the insertion of the plantaris tendon, which shows that this muscle acts as toe extensor in cats but as ankle extensor in baboons. For most of the motor nuclei, the homonymous as well as the total aggregate of Ia-e.ps.p.s is smaller in the baboon than in the cat; but the amplitude range between different motor nuclei is larger in the baboon. Reciprocal Ia-i.ps.p.s are evoked only after spinal transection or when brain function is depressed. It is postulated that baboons, contrary to cats, have descending tonic inhibition of transmission in the reciprocal la-inhibitory pathway. The phylogenetic flexibility of I a-connections is discussed and contrasted with their ontogenetic stability.

1. Sensory inputs to and locations of uncrossed spinocerebellar tract neurones in the lower lumbar cord were studied in chloralose-anaesthetized cats. 2. Neurones with axons ascending in the ipsilateral thoracic funiculi and projecting to the cerebellum were found mainly dorsal to the central canal (laminae V and VI) in the L5-L6 segments, i.e. at levels caudal to Clarke's column. Axons considered to originate from these cells were located in the dorsal half of the lateral funiculus at the level of L2, intermingled with axons of the dorsal spinocerebellar tract originating at the levels of Clarke's column. 3. Synaptic actions of primary afferents on neurones with antidromic invasion following stimuli applied to ipsilateral thoracic funiculi or to the cerebellum were investigated using intracellular or extracellular recording in the caudal lumbar segments. 4. Monosynaptic excitatory effects were evoked by electrical stimulation of group I muscle afferents of the hindlimb ipsilateral to the cell body. The majority of neurones received monosynaptic excitation from two or more muscles, predominantly extensors. They were frequently co-excited by group Ia muscle spindle and group Ib tendon organ afferents. 5. Volleys in cutaneous afferents produced excitation with short central latencies. In addition to the monosynaptic and disynaptic excitation from low-threshold cutaneous afferents, there were indications of monosynaptic effects from slightly slower conducting fibres. The majority of these neurones also received monosynaptic excitation from group I muscle afferents. Neurones with cutaneous input tended to be located more dorsally compared with those responding only to muscle afferents. 6. Volleys in joint afferents produced monosynaptic excitatory postsynaptic potentials (EPSPs) in the neurones with EPSPs from group I or group I and cutaneous afferents. 7. Some neurones were disynaptically inhibited from group I muscle afferents. Convergence of monosynaptic group I excitation and disynaptic group I inhibition occurred in varieties of patterns. 8. Polysynaptic excitation, inhibition or mixed effects of both were evoked from ipsilateral cutaneous afferents and high-threshold muscle and joint afferents, whereas effects from the controlateral afferents were feeble.(ABSTRACT TRUNCATED AT 400 WORDS)

The topographical distribution of interneurones mediating disynaptic inhibition of dorsal spinocerebellar tract (d.s.c.t.) cells from group I muscle afferents in the cat was investigated using both physiological and morphological techniques. Lesions of either the dorsal funiculi or of the lateral and ventral funiculi were made between L4 and L5 segments in two groups of cats. I.p.s.p.s. evoked from group I afferents were seen after both these lesions, showing that the i.p.s.p.s were evoked by interneurones located more caudally as well as by interneurones in the same segments as Clarke's column. Distribution of the caudally located interneurones in the lower lumbar segments was investigated after marking these interneurones with horseradish peroxidase retrogradely transported from Clarke's column. The horseradish peroxidase was injected along L3-L4 segments of Clarke's column in two cats with transected dorsal funiculi. The marked cells were found in L5, L6, L7 and S1 segments, with a highest density in L6 and L7. They were seen in laminae V, VI and VII. A search was made for interneurones which could be antidromically invaded following stimuli applied in Clarke's column and were monosynaptically excited by group I afferents. Such interneurones were found at locations corresponding to laminae V-VI of Rexed. The latencies of antidromic and orthodromic responses were within ranges allowing them to mediate disynaptic inhibition of d.s.c.t. cells.

1. Intra- and extracellular recordings were made from cells of the spinocervical tract in the lumbosacral spinal cord. A convergence of monosynaptic excitatory post-synaptic potentials (EPSPs) and disynaptic inhibitory post-synaptic potentials (IPSPs) was a general pattern of effects from the low threshold cutaneous fibres. Unitary IPSPs, probably mediated via the same disynaptic path, were evoked by light touch of hairs, which was also the adequate stimulus for exciting the cells. The receptive field for unitary IPSPs was closely related to the excitatory receptive field but was eccentric, not of a surround type. 2. EPSPs, IPSPs, or both, were evoked from the flexor reflex afferents in the great majority of neurones. Disynaptic IPSPs may be evoked from the interosseous nerve. No effects were produced by volleys in group I muscle afferents. 3. It is suggested, on the basis of the spatial organization of the excitatory and inhibitory receptive skin fields, that the spinocervical tract may give information regarding the direction of tactile stimuli.

The aim of the study was to investigate whether inhibition of dorsal spinocerebellar tract (d.s.c.t.) cells evoked from group I afferents is mediated by the same interneurones which mediate the non-reciprocal inhibition of hind-limb motoneurones. The origin of inhibition of d.s.c.t. cells from group I afferents was compared in intact preparations, after lesions of the dorsal funiculi (when it could only be mediated by lower lumbar interneurones) and after lesions of the lateral and ventral funiculi (when it would be expected to be evoked by upper lumbar interneurones). In all three preparations extensors were the most common source of inhibition, as in motoneurones. Lower lumbar interneurones inhibiting d.s.c.t. cells were found to be co-excited by group I (Ia and/or Ib) and cutaneous and joint afferents, and by rubrospinal tract fibres, as are interneurones mediating inhibition of motoneurones. Co-excitation by group I and rubrospinal fibres was also found for upper lumbar interneurones. I.p.s.p.s were evoked in hind-limb motoneurones from within Clarke's column in cats with the dorsal funiculi cut between L4 and L5 segments; they were evoked at thresholds as low as 2 microA, i.e. by stimuli with very local actions. The latencies of these i.p.s.p.s were short enough to allow them to be evoked monosynaptically via axonal branches of the same interneurones which projected to Clarke's column. Correspondingly, i.p.s.p.s were evoked in d.s.c.t. cells from within motor nuclei in L7 segments; they were evoked at similarly low thresholds and with similar latencies. In confirmation of previous reports i.p.s.p.s of Ia origin evoked in d.s.c.t. cells were not found to be depressed by Renshaw cells, which excludes their mediation by interneurones responsible for Ia reciprocal inhibition. The study leads to the conclusion that the inhibition of d.s.c.t. cells from group I afferents is, at least in part, collateral to the non-reciprocal inhibition of lumbar motoneurones.

1. Glass microelectrodes were inserted into the lateral vestibular nucleus of Deiters, and potential changes were recorded both extracellularly and intracellularly under stimulation of the spinal cord.2. When the ipsilateral C3 segment was stimulated at its ventrolateral surface, negative field potentials of several millivolts were recorded from the area which was histologically identified as Deiters' nucleus. These field potentials were presumed to be caused by the antidromic activation of Deiters' neurones through the vestibulospinal tract.3. The antidromic field potential of Deiters' nucleus showed different distributions in the ventrodorsal direction for activation at the C3 and L1 levels, which is in keeping with the histologically determined somatotopical arrange-ment.4. Within the nucleus of Deiters specified by the field potentials, a total of 134 units were impaled with microelectrode. They were identified as Deiters' neurones by the characteristics of the antidromic spikes, their afterhyperpolarization and by occurrence of synaptic noise and PSPs.5. In sixteen selected neurones the resting potential was -55 to -75 mV and the spike height 60 to 100 mV (mean, 80.8 mV). The duration of the spike was 0.4 to 0.6 msec (mean, 0.47 msec).6. The conduction velocity along the vestibulospinal fibres was determined in fifty Deiters' neurones which were activated antidromically both from the C3 and L1 segmental levels. It ranged from 24 to 140 m/sec, the peak frequency being at 90 to 100 m/sec.7. The falling phase of the spike, after it crossed the base line, reversed to an afterhyperpolarization which reached its summit at 0.7 to 8.8 msec (mean, 2.4 msec), and diminished within 33 to 100 msec (mean, 49 msec). These values are smaller than those for cat spinal motoneurones. As a unique feature in Deiters' neurones the afterhyperpolarization did not show a temporal summation when it was evoked successively at short intervals.8. The stimulation of the ipsilateral C3 segment just subthreshold for the axon of the impaled cells induced EPSPs monosynaptically in 15.4% and IPSPs polysynaptically in 7.7% of the examined neurones. The recurrent axon collaterals might be responsible for these IPSPs, but the possibility was not excluded for the other pathways.9. Stimulation at the C3 segment supramaximal for the antidromic field potential of Deiters' nucleus produced monosynaptic EPSP in 50% and polysynaptic IPSP in 28.6%. It was suggested that the monosynaptic EPSP was produced through the ascending tract fibres in the spinal cord.10. In view of the fast time course of the action potential and of rarity of the possible recurrent inhibition, it was postulated that Deiters' neurones would be grouped with the fast type of lumbosacral motoneurones.

Chez le chat anesthesie au Nembutal, les reponses evoquees dans les neurones de Deiters par les stimulations antidromiques ont ete etudiees au moyen de microelectrodes intracellulaires.

La stimulation du Noyau Rouge a pour effet une facilitation des interneurons intercales entre des afferences primaires et les motoneurons et provoque dans les motoneurons des potentiels postsynaptiques bisynaptiques.

Wahrend Reizung des caudalen Hirnstammes wurden durch intrazellulare Ableitung mit Mikroelektroden monosynaptische EPSPs in lumbosakralen γ-Motoneuronen registriert.