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The molecular structure of plasmid-like DNAs (designated type-II) which were isolated from mitochondria in the ciliated protozoan Paramecium caudatum was characterized. These type-II DNAs are always detected as a set of four kinds with sizes of 8.2, 4.1, 2.8 and 1.4 kb. The DNAs of 8.2 and 2.8 kb exist as dimers consisting of 4.1- and 1.4-kb monomer molecules, respectively. Electron microscopic observations indicated configurations of a hairpin structure that had a protruding end of single-stranded DNA in one terminus and a loop in the other terminus. The monomers stick together with base-pairing in opposite directions at the protruding end to form the dimers, suggesting the presence of inverted repeats. These unusual dimers may have a role in replication of the DNAs in which the monomers can serve as a primer for each other.

Fetal and newborn lambs' hearts have limited capacity to adapt to increased afterload. In human preterm infants, however, this limited left ventricular pump function has not been investigated. To determine the normal left ventricular resting contractile state in preterm infants, we studied the relation of end-systolic wall stress to the rate-corrected velocity of circumferential fiber shortening (VCF) in 11 preterm infants of gestational ages 28 to 36 weeks. We performed the first (7 to 14 days of postnatal age) and the second (29 to 39 days of postnatal age) examinations. There was a significant inverse linear correlation between wall stress and VCF in the first and the second examinations (r = -0.616 and r = -0.715, respectively). Both regression lines had slopes that were steeper than those previously reported for older age groups. The preterm infants had high VCF values (mean 1.05 and 1.09 cir/sec, respectively) and low-wall stress values (mean 36.0 and 36.9 gm/cm2, respectively) in the first and the second examinations. We therefore suggest that the preterm infants have a high resting contractile state, but left ventricular performance may deteriorate in the face of increased afterload.

Effects of indeloxazine, a cerebral activator, on passive avoidance learning by disruption of cholinergic transmission were studied in rats. The shortened latency was prolonged when indeloxazine was administered before training in rats subjected to scopolamine-injection, ethylcholine aziridinium ion (AF64A)-treatment and nucleus basalis magnocellularis (NBM) lesion. Indeloxazine administered immediately after training also showed ameliorating effects on passive avoidance in NBM-lesioned rats. Indeloxazine increased extracellular acetylcholine concentration in the frontal cortex of rats. Indeloxazine possesses facilitatory effects on cerebral function in part due to activation of the central cholinergic system.

The antitumor effects of a monoclonal antibody against a human cancer cell line and a protein-bound polysaccharide, PSK, obtained from cultured mycelia of Coriolus versicolor in basidiomycetes were examined. The IgG2a monoclonal antibody against the human colon cancer cell line colo 205 induced in vitro antibody-dependent macrophage-mediated cytotoxicity against the cancer cells, but only slightly suppressed the in vivo growth of the cancer cells. Concurrent use of PSK with the antibody enhanced the in vitro antibody-dependent macrophage-mediated cytotoxicity as well as the in vivo antitumor activity. These findings suggest that the combined use of a monoclonal antibody and PSK, which have different modes of action, may be useful in the treatment of cancer.

Background and Purpose: We evaluated the utility of blood flow velocity measurements by transcranial Doppler ultrasonography as a tool to indirectly measure cerebral perfusion during cardiopulmonary bypass., Methods: We simultaneously measured blood flow velocity in the middle cerebral artery and physiological variables in 18 patients undergoing cardiac surgery under hypothermic cardiopulmonary bypass in which pH and PaCO2 were managed with the alpha-stat acid-base strategy. We expressed blood flow velocity as a relative value of control obtained under normothermia and normocarbia before bypass. We also developed an original index, modified cerebral metabolic rate for oxygen, to estimate cerebral metabolic rate for oxygen., Results: Relative velocity was significantly (P < .01) reduced during stable aortic cross-clamp compared with before bypass and was significantly (P < .01) increased during rewarming compared with at aortic cross-clamp. Modified cerebral metabolic rate for oxygen significantly correlated with nasopharyngeal temperature during cooling, aortic cross-clamp, and rewarming (r = .756, P < .0001; r = .4, P < .01; r = .725, P < .0005, respectively). Calculated temperature coefficient for modified cerebral metabolic rate of oxygen was 2.7 +/- 1.4 (mean +/- SD, n = 10) during cooling. Only nasopharyngeal temperature and PaCO2 were significant determinants of relative velocity during aortic cross-clamp., Conclusions: We can monitor cerebral perfusion and metabolism by measurements of relative velocity and modified cerebral metabolic rate for oxygen during hypothermic cardiopulmonary bypass.

Expression of the virulence genes (vir) on the hairy-root-inducing plasmid pRiA4 is induced by plant signals in Agrobacterium cells through a two-component regulatory system, the VirA-VirG system. We constructed an in vitro transcription system that consisted of the purified VirG protein and the Agrobacterium RNA polymerase holoenzyme. Both versions of VirG, the non-phosphorylated form and the VirA-phosphorylated form, were active but showed different patterns of the pH-dependency for transcriptional activation.

This study is a survey of the overall clinical results achieved with our pain treatment method, percutaneous epidural low-frequency (1.6-8.0 Hz) spinal cord stimulation. It examines the relationship between the effectiveness of epidural spinal cord stimulation (ESCS) and diseases or sites of pain. Continuous indwelling of the catheter electrodes in the posterior epidural space ranged from 3 to 67 days, and the duration of percutaneous ESCS varied from less than 1 wk to more than 1 yr. Complete pain relief (100%) was achieved during stimulation in 11.5% of the patients (52 of 454). Complete (100%) to partial (more than 30%) pain relief occurred in 71.1% of the patients. In six (1.3%) patients pain was aggravated by stimulation. Analgesics and/or sedatives were discontinued completely after treatment in 52 patients (11.5%) and reduced in 263 patients (57.9%). The number of patients who rated pain relief better than 50% was significantly more in carcinoma/sarcoma and causalgia (P < 0.001), and significantly less in postherpetic neuralgia and thromboangitis obliterans/arterial sclerosis obliterans (P < 0.001) than the average in all diseases. There was a significantly high responsiveness to ESCS in female patients in comparison to male patients (P < 0.05). Pain in the head/face, neck/upper extremities, and trunk responded more to ESCS than pain in the lower extremities. Alleviation of pain by ESCS was lower when the verbal pain score was high. There were no major complications in percutaneous ESCS. Thus, we have demonstrated that pain-alleviating effects of ESCS varies significantly by disease and site of pain, and that this simple percutaneous method can be used for a relatively long period.

The effects of hemodilution on cerebral hemodynamics and oxygen consumption during nonpulsatile hypothermic cardiopulmonary bypass (CPB) were investigated in 16 patients undergoing elective open heart surgery under CPB. They were divided into 2 groups; high (25%) (H) and low hematocrit (15%) (L) groups according to the hematocrit values during CPB. Simultaneous measurements of blood flow velocity in the middle cerebral artery (VMCA) and oxygen tension in the jugular venous bulb (PjvbO2) were performed at prebypass, bypass, and post bypass periods. There were no significant differences in nasopharyngeal temperature (NPT), cerebral perfusion pressure, pump flow rate, and PaCO2 between the two groups at any period. VMCA changed in correlation with NPT in both groups, and VMCA in the L group was faster than that in the H group at any period. PjvbO2 increased during aortic cross clamp, and decreased on rewarming in both groups. However, there were no significant differences in VMCA and PjvbO2 between the two groups at any period. These findings suggest that increased cerebral blood flow may compensate the decreased oxygen content due to the hemodilution during CPB. Therefore, the profound hemodilution up to 15% of hematocrit is considered to be a safe technique during CPB.

A 6-year-old boy with nephrogenic diabetes insipidus (NDI) and intracranial calcification is reported. The calcifications were symmetrical and located in the basal ganglia and in the subcortical regions of the frontal, temporal, parietal and occipital lobes. Episodes of hyperosmolality during infancy are considered to be one of the causes of intracranial calcification in NDI. However, other unknown factors may be involved, because up to now there have been no reports of intracranial calcification in patients with central diabetes insipidus.

We determined the effects of cyclosporine on calcium, magnesium, and potassium metabolism in rats. Thirty Sprague-Dawley rats were randomized into three groups of ten animals each--control rats given olive oil, rats given cyclosporine at a dosage of 5 mg/kg daily, and rats given 15 mg/kg daily for four weeks. Urinary excretion of calcium, magnesium, and potassium was determined before and after 2 and 4 weeks of cyclosporine therapy. All rats were sacrificed after 4 weeks of therapy, and calcium, magnesium, and potassium concentrations in serum and tissues were determined. Serum magnesium levels were significantly lower in the cyclosporine-treated groups than in the control group, but there was no significant difference between the control and either of the cyclosporine-treated groups with regard to total urinary excretion of magnesium after four weeks of treatment. Magnesium content in the kidney, muscle, and liver was significantly higher in the 15 mg/kg group than in the control group. Calcium content in the kidney and liver was significantly higher as well. Potassium content in any type of tissue was similar in the three groups. We conclude that the intracellular migration of magnesium plays an important role--as does impaired renal conservation of magnesium--in the pathogenesis of cyclosporine-induced hypomagnesemia and that there is a discrepancy between magnesium and potassium metabolism in cyclosporine-treated rats.

The effect of a protein-bound polysaccharide (PSK) obtained from cultured mycelia of the Basidiomycetes Coriolus versicolor on activities involved in the host defence mechanism of C57BL/6 mice bearing adenocarcinoma 755 was compared with that of live bacille Calmette-Guérin (BCG). Delayed footpad reaction, the activity of splenic natural killer cells and interferon production induced by concanavalin A in splenic cells of healthy mice were little affected by PSK, but in mice bearing tumours PSK prevented the tumour-induced reduction in these activities. Live BCG augmented these activities in healthy mice but had little effect on the reduction of activities induced by a tumour. The immunosuppressive activity of the serum of tumour-bearing mice was reduced by PSK administration; live BCG did not have this effect. The combined use of live BCG and PSK improved these activities in the host, with synergistic increases in the antitumour effect. These results suggest that the combined use of live BCG and PSK, which have different modes of action, may be useful in the treatment of cancer.

We have previously shown by cDNA cloning that a higher plant, Arabidopsis thaliana, possesses at least two CDC2 genes (CDC2a and CDC2b) similar to the cell-cycle-controlling cdc2 gene of Schizosaccharomyces pombe. To understand the exon-intron organization of these genes, genomic clones were isolated and their nucleotide sequences determined. The coding and 5'-untranslated regions of CDC2a were interrupted by seven and one introns, respectively, whilst CDC2b contained three introns within the coding portion. These intron positions partly overlapped with each other and with those of the yeast cdc2 gene, nevertheless the lengths and sequences of the corresponding introns were diverse.

Drug release was controlled by a combination of prodrug and polymer matrix. Prodrugs of 5-fluoro-2'-deoxyuridine with different physicochemical properties were synthesized by esterification with aliphatic acids (propionate, n-butyrate, and n-pentanoate). Microspheres containing these ester prodrugs were prepared with poly(3-hydroxybutyrate) of three molecular weights (65,000, 135,000, and 450,000). The release rates from the spheres depended on both the lipophilicity of the prodrug and the molecular weight of the polymer. Regardless of the polymer, the relative release rates were propionate greater than butyrate greater than pentanoate. The release of butyrate and pentanoate from the spheres consisting of low-molecular-weight polymer (M(r), 65,000) was faster than that from the spheres of higher molecular weight (M(r), 135,000 or 450,000). A single intraperitoneal injection of spheres of the highest molecular weight polymer containing butyrate or pentanoate resulted in higher antitumor effects against P388 leukemia in mice than did free prodrugs given over a period of five consecutive days. The polymer sphere itself showed low toxicity to and good biocompatibility with mice and rats.

We studied the pathogenesis of cyclosporine-induced hypomagnesemia in five patients with nephrosis. Serum magnesium concentrations and urinary excretion of magnesium were reduced by the therapy. In contrast, the magnesium concentrations in mononuclear blood cells were increased. We conclude that short-term use of cyclosporine induces an intracellular shift of magnesium and causes hypomagnesemia.

Effects of hypoxia and/or glucose depletion on apparent intracellular pH, (pHi)app, were studied in hippocampal slices of mouse brain using the fluorescence pH indicator BCECF (2',7'-bis(carboxyethyl)carboxyfluorescein). In the presence of glucose (11 mM), (pHi)app of the CA1 pyramidal cell layer decreased with hypoxia but rapidly recovered with reoxygenation. The response in (pHi)app to hypoxia was consistent with changes in lactate content. By superfusion with a hypoxic and glucose-devoid solution, (pHi)app decreased in the first 5 min, but then increased in two phases: during a 5- to 7-min period and again during the latter period of superfusion. BCECF fluorescence rapidly diminished in this latter period. These changes in (pHi)app could not be explained only by the lactate content in the slices. Some other mechanisms may also regulate the (pHi)app in the slices during and after hypoxia and glucose depletion.

Effects of dorsal root entry zone lesions (DREZLs) on cerebrospinal fluid (CSF) and plasma concentrations of neuropeptides, catecholamines, and cyclic nucleotides were studied in 9 patients with intractable chronic pain. Contents of beta-endorphin-like-material in CSF decreased in all patients 12-17 days following DREZLs during which complete to good pain relief was achieved. Contents of beta-endorphin-like-material in CSF increased again about one month after DREZLs in two and remained unchanged in one of three patients tested, who complained of partial reappearance of pain. Contents of beta-endorphin-like-materials in plasma showed no significant changes after DREZLs. Substance P, noradrenaline, adrenaline, and cyclic nucleotide levels in both CSF and plasma were variable among the subjects and did not change significantly following the operations. Thus, the results suggest that production of beta-endorphin-like-material in the central nervous system is decreased by DREZL, though the increase in its turn-over might not be neglected. The mechanisms of the decrease in contents of beta-endorphin-like-material in CSF after DREZLs were discussed in terms of our current knowledge of pain and pain inhibitory systems.

A previous report from this laboratory showed that in vitro transcription of total genomic DNA of the newt Cynopus pyrrhogaster resulted in a discrete sized 8 S RNA, which represented highly repetitive and transcribable sequences with a glutamic acid tRNA-like structure in the newt genome. We isolated four independent clones from a newt genomic library and determined the complete sequences of three 2000 to 2400 base-pair PstI fragments spanning the 8 S RNA gene. The glutamic acid tRNA-related segment in the 8 S RNA gene contains the CCA sequence expected as the 3' terminus of a tRNA molecule. Further, the 11 nucleotides located 13 nucleotides upstream from one of the two transcription initiation sites of the 8 S RNA were found to be repeated in the region upstream from the termination site, suggesting that the original unit, which is shorter than the 8 S RNA, was retrotransposed via cDNA intermediates from the PolIII transcript. In the upstream region of the 8 S RNA gene, a 360 nucleotide unit containing the glutamic acid tRNA-related segment was found to be duplicated (clones NE1 and NE10) or triplicated (clone NE3). Except for the difference in the number of the 360 nucleotide unit, the three sequences of the 2000 to 2400 base-pair PstI fragment were essentially the same with only a few mutations and minor deletions. Inverse polymerase chain reaction and sequence determination of the products, together with a Southern hybridization experiment, demonstrated that the family consists of a tandemly repeated unit of 3300, 3700 or 4100 base-pairs. Thus during evolution, this family in the newt was created by retroposition via cDNA intermediates, followed by duplication or triplication of the 360 nucleotide unit and multiplication of the 3300 to 4100 base-pair region at the DNA level.

Poly(L-lactic acid) (L-PLA) microspheres containing 5-fluoro-2'-deoxyuridine (FUdR) or its ester prodrugs with saturated aliphatic acids (FUdR-Cn, n = 2, 3, 4, 5, 6, 8, 10 and 12) were prepared. The physicochemical and biological properties and antitumor activity of the L-PLA microspheres were studied. The lipophilicity of FUdR-Cn was increased by prolonging its acyl-promoieties. FUdR-C5, FUdR-C6, FUdR-C8, FUdR-C10 and FUdR-C12 showed almost complete incorporation into the microspheres, while incorporation of hydrophilic FUdR and FUdR-C2 was poor. The sustained release of FUdR from the microspheres containing FUdR-C4, FUdR-C5 and FUdR-C6 was obtained in the presence of esterase, and higher antitumor activity against P388 leukemia was observed in vivo. On the other hand, the release rates of FUdR from the microspheres containing FUdR-C10 and FUdR-C12 were very small, and their antitumor activity was much smaller than that of the free prodrug suspension. Effects of the susceptibility to enzymatic hydrolysis and the physiocochemical properties of prodrugs on the release profiles of FUdR from spheres were discussed.

Poly-L-lactic acid (MW 6000) microspheres (PLA-MS) containing 5-fluoro-2'-deoxyuridine (FUdR) or four ester prodrugs of FUdR were prepared and examined with regard to the in vitro release kinetics. The incorporation efficiency of the lipophilic prodrugs into the PLA-MS was higher than that of FUdR or the hydrophilic prodrugs. The release of the lipophilic FUdR prodrugs from PLA-MS was sustained as compared with that of FUdR from PLA-MS. The release kinetics of the FUdR prodrugs appears to fit the Higuchi, Baker, and Lonsdale model in the early stage of the release process. The slope of the Baker and Lonsdale plots of the release of divaleryl-FUdR from PLA-MS decreased as the initial drug loading was decreased. The order of the release rates of FUdR prodrugs from PLA-MS with the same prodrug content was similar to that of the water solubilities of the prodrugs. These results suggest that the ester prodrugs could be released from PLA-MS by diffusion through water-filled capillaries or a series of pores rather than by diffusion through the PLA matrix.

We sequenced a 4.2-kb DNA region encompassing the vir A locus of the hairy-root-inducing plasmid pRiA4, and compared its sequence with the published vir A region sequences of four tumor-inducing plasmids. An open reading frame capable of coding for 829 amino acids was identified for vir A. Deletion mutants of vir A constructed by fusing to lacZ, but not the wild-type game itself, were efficiently expressed in Escherichia coli when they were put downstream front the lac promoter. These fused gene products became soluble or insoluble depending on the length of their lacZ moieties.

On in vitro transcription of total genomic DNA of the tortoise (Geoclemys reevessi), a discrete-sized RNA of 6.5S was obtained that represented a highly repetitive and transcribable sequence in the tortoise genome. Three sequences of the 6.5S RNA gene were sequenced, and a consensus sequence was deduced from these three sequences and one reported previously [Endoh, H & Okada, N. (1986) Proc. Natl Acad. Sci. USA 83, 251-255]. The 5' part of the gene showed close similaries to lysine (rabbit) and threonine (mouse) tRNAs (overall similarity 68-70%), so this tortoise sequence may have evolved from one of these tRNAs. The consensus sequence retained the expected CCA triplet at the 3' end of tRNA, but not at the 3' end of tDNA, supporting the idea that the tRNA-related region of the gene was generated via an RNA intermediate. The 5' and 3' flanking sequences of the four genes were found to be completely different from each other. Fingerprint analysis and S1 nuclease mapping analysis also showed that sequence boundaries of tortoise repetitive units exactly corresponded to RNA species. These results, together with data obtained by Southern blot hybridization, indicated that the 6.5S RNA genes are dispersed in the tortoise genome. Therefore, generation of the tRNA-related region of the gene and amplification of the whole unit of the gene are both RNA-mediated events. The existence of this tortoise sequence suggests that short interspersed sequences are more common in eukaryotic genomes than had previously been thought.

Total DNAs from various animals were transcribed in vitro in a HeLa cell extract, and it was found that one to several discrete RNAs were transcribed by RNA polymerase III. With tortoise (Geoclemys reevessi) and newt (Cynops pyrrhogaster), distinct 6.5S and 8S RNAs were transcribed from these respective DNAs. Representative phage clones carrying the 6.5S and 8S RNA genes were isolated from genomic libraries of these animals, and the sequences of these genes were determined. The 5' parts of highly repetitive and transcribable sequences of tortoise and newt were found to have close resemblance to tRNALys1 (rabbit) gene (78% homology) and a tRNAGlu (Drosophila) gene (74% homology, not counting the aminoacyl stem region), respectively. The homologies extended to secondary structures, homologous nucleotides being located on similar secondary structures. It is proposed that many, if not all, highly repetitive and transcribable sequences detected by total DNA transcription have specific tRNA genes as their progenitors.

A study was done to investigate the involvement of interferon (IFN) in immunological acceptance of fetal allograft. IFN was induced by stimulating OK-432 from lymphocyte of pregnant and nonpregnant women as well as men. There was no difference in the IFN titer in these groups. In pregnant women there was no difference between trimesters, either. In addition, we examined the influence of sex steroid hormones such as progesterone and estrogen on IFN production by the lymphocytes of women and men. Generally, each steroid hormone suppressed the production of IFN. However, the extent of suppression varied with the concentration of hormones, resulting in different suppression curves for men and women in a physiological concentration. Differences in response by men and women were also noticed when interleukin 2 was added to examine whether it could reduce the suppression by steroid hormone. This unique sensitivity of female lymphocyte to steroid hormones may play a role in successfully taking a fetal allograft in pregnancy.

Antiserum against a protein-bound polysaccharide preparation (PSK) was produced by immunizing New Zealand White rabbits with PSK. The intestinal absorption of PSK in mice was visualized by indirect immunofluorescent staining with anti-PSK serum. The change in blood levels of 14C after oral administration of 14C-PSK and the recovery of 14C by antiserum were determined. The results indicated that antigenic epitopes in PSK are not completely destroyed during the process of digestion, absorption and distribution, but the changes of serum levels of 14C radioactivity differ from those of immunoreactive radioactivity. These results suggest that multiple processes are involved in the fate of PSK administered orally.

Concentrations of cyclic AMP and cyclic GMP in arterial and internal jugular venous (IJV) blood were determined at the resting wakeful state in thirty surgical patients without neurological deficits. The levels of cyclic AMP in artery and IJV were 32.1 +/- 3.0 and 40.0 +/- 4.1 pmol/ml (means +/- standard errors), respectively, while those of cyclic GMP in artery and IJV were 12.2 +/- 2.7 and 14.4 +/- 3.0 pmol/ml, respectively. Concentrations of both cyclic nucleotides in IJV were significantly higher (P less than 0.001) than those in artery. IJV-arterial differences for cAMP and cGMP were 7.9 +/- 1.7 and 2.1 +/- 0.5 pmol/ml, respectively. The results indicate that both cyclic nucleotides are constantly produced and released from the normal human brain.

Survival rates following incomplete brain ischemia induced during pentobarbital anesthesia were significantly higher in mice with a minor brain injury, inflicted one week before, than in those given a sham operation. Improvement of the survival rates in mice with brain injury, however, became insignificant when brain ischemia was imposed during ketamine anesthesia, suggesting that the actions of certain factors or protective mechanisms against brain ischemia, developed by brain injury, are antagonized by ketamine and/or potentiated by barbiturate anesthesia.

Actin is ubiquitous in eukaryotes, nevertheless its existence has not yet been clearly proven in Tetrahymena. Here we report the cloning and sequencing of an actin gene from the genomic library of Tetrahymena pyriformis using a Dictyostelium actin gene as a probe. The Tetrahymena actin gene has no intron. The predicted actin is composed of 375 amino acids like other actins and its molecular weight is estimated as 41,906. Both T. pyriformis and T. thermophila possess a single species of actin genes which differ in their restriction patterns. Northern hybridization analysis revealed that the actin gene was actively transcribed in vivo. To detect the gene product, we synthesized an N-terminal peptide of the deduced sequence and prepared its antibody. Using an immunoblotting technique, we identified Tetrahymena actin on a two-dimensional gel electrophoretic plate. The actin spot migrated near an added spot of rabbit skeletal muscle actin, but clearly differed from the latter in its isoelectric point and apparent molecular weight. The primary structure of Tetrahymena actin shares about 75% homology equally with those of other representative actins. This value is extremely low as a homology rate between known actins. Tetrahymena actin diverges not only in relatively variable regions of other actins, but also in relatively constant regions. The hydrophilicity levels of two regions (residues 190 to 200 and residues 225 to 235) are also quite different between the Tetrahymena actin and skeletal muscle actin. Thus, we conclude that actin is present in Tetrahymena, but it is one of the most unique actins among the actins known hereto.

The effect of the opioid antagonist naloxone on hypoxia-induced blockade of synaptic transmission in the superior cervical ganglion (SCG) of the rat was studied in vitro. Naloxone (6 X 10(-6) M or more) attenuated the hypoxia-induced blockade of synaptic transmission in the SCG. In addition, in the concentrations studied, naloxone itself had a blocking effect on ganglionic transmission that involved terminal sites of the preganglionic axons. These data suggest that the protective effect of higher doses of naloxone on the hemodynamic responses to hypoxemia or ischemia may originate at least in part from the attenuating effect of naloxone on the hypoxia-induced blockade of ganglionic transmission.

Several aspects of total genomic DNA transcription in a HeLa cell extract were described. (1) By using in vitro transcripts from total genomic DNA as probes, we elucidated several structures of short interspersed elements which are transcribed by RNA polymerase III (designated as Pol III/SINEs. (2) To know whether the repetitive sequence we cloned and sequenced comprises a major or minor family, we developed in vitro run-off transcription assay of total genomic DNA. (3) Unlike in vitro transcription of total genomic DNA from vertebrates, major in vitro transcripts from macronuclear total DNA of Tetrahymena were found to be tRNA themselves.

Serum and renal tissue concentrations of ceftizoxime (CZX) after intravenous dosing with 1 g were determined in 15 patients with renal disease. The highest serum concentration of CZX was 105 micrograms/ml 6 minutes after dosing and the lowest concentration was 10.5 micrograms/ml at 170 minutes. The highest renal tissue concentration was 155 micrograms/g and the lowest concentration was 7.46 micrograms/g. There were no correlation between the tissue concentrations and sampling time. Tissue to serum concentration ratios ranged from 0.28 to 5.73. These patients were divided into 3 groups; hydronephrosis, normal kidneys and renal atrophy according to pyelographic findings. Their renal tissue concentrations were compared with serum concentrations in each group. The tissue concentrations were lower than the serum concentrations in the first group, and were almost the same or higher in the other groups. CZX concentrations in the medulla and the cortex were determined in 5 patients; and were higher in the medulla in 4 of these patients. Urinary tract infection due to E. coli, K. pneumoniae, P. mirabilis, S. pyogenes or S. epidermidis was present in 5 of the 25 patients. Intravenous injection of 1 g of CZX provided therapeutically effective concentrations against these urinary tract infections.

A simple agglutination microassay is described for studying the presence of antigenic determinants on the surface of liposomal membranes. Antigen-sensitized radioactive liposomes, reacted with specific immune IgG in the wells of a microtest plate, were passed through a glass fiber filter in a multiple cell culture harvester. The amount of agglutinate trapped on the filter as determined by the radioactivity, was proportional to the amount of antigen in the liposomes and to the amount of specific immune IgG in the reaction mixture. The specificity of the agglutination was determined by inhibition with hapten and carrier antigens. Liposomes actively sensitized with hapten-conjugated lipid, and passively sensitized with protein, were used in the assay. A new method of preparing protein-bound liposomes is also described.

To evaluate early neonatal circulatory adaptation, left and right ventricular cardiac outputs were examined in 10 normal neonates using Doppler echocardiography. Serial examinations were performed until ductal closure was confirmed. Two-dimensional echocardiography and Doppler echocardiography were used to evaluate structures of the heart and great vessels and flow within them. Then, the diameters of the ascending aorta (dAo) and pulmonary artery (dPA) were measured using M-mode echocardiography. The flow velocity patterns of the ascending aorta and pulmonary artery were recorded, measuring mean aortic (VAo) and pulmonary artery velocities (VPA). Left (LVCO) and right ventricular cardiac outputs (RVCO) were calculated as follows: LVCO = (dAo)2/4 x pi x VAo x 60, RVCO = (dPA)2/4 x pi x VPA x 60. 1. Patency of the ductus arteriosus was confirmed by the shunt through it in all neonates initially examined. The initial velocity pattern of the ductal shunts was bidirectional (9/10) or continuous left-to-right (1/10). The flow velocity pattern changed to continuous left-to-right in most neonates, and spontaneous closure of the ductus was confirmed at the age of 13 to 64.5 (mean 31.4) hrs. 2. Left-to-right shunt through the stretched foramen ovale was noted in six neonates temporarily. 3. As for the arterial diameter, dAo did not change, but dPA decreased at the time of spontaneous closure of the ductus. 4. Concerning mean aortic velocity, VAo increased when a continuous ductal left-to-right shunt was initially suspected, while VPA increased when spontaneous closure of the ductus was confirmed. 5. For cardiac output, LVCO (ml/min) changed from 618.4----718.3----562.7, while RVCO showed no change (576.1----546.5----557.8) according to a ductal flow change from bidirectional to continuous left-to-right and finally to no shunt. The LVCO/RVCO ratio increased with increasing age and (in 7/9) the maximal rate was noted when the continuous left-to-right ductal shunt was confirmed. The increased LVCO contributed to the increased LVCO/RVCO ratio. Using this method, serial evaluations of two ventricular cardiac outputs could be made.

SH-bearing subunits of IgM antibody were coupled through the Fc portion to sonicated liposomes consisting of N-(m-maleimidobenzoyl)dipalmitoylphosphatidylethanolamine, dipalmitoylphosphatidylcholine and cholesterol. Liposomes coated with subunits of monoclonal IgM antibody against a mouse mammary tumor-associated antigen selectively bound to the antigenic target cells. Assays of the liposomes for complement dependent cytotoxicity and spleen cell binding revealed inactivation of the liposome-coupled antibodies in both assays.

In order to investigate the significance of overshoot acceleration in FHR with acidosis induced by serial cord compression, cerebral glucose metabolic rate (CMRglc) was measured in four chronically instrumented fetuses before cord compression and immediately after appearance of the overshoot acceleration in FHR. The investigation of CMRglc was carried out using positron emission tomography to assess accumulation of cyclotron-produced F18-FDG in the fetal brain. CMRglc in the fetal brain in the nonacidemic condition was 1.73 mg/100 g/min. In acidemia, with fetal arterial pH near 7.15 in which FHR showed the characteristic pattern with overshoot acceleration, CMRglc was significantly decreased to 0.73 mg/100 g/min. We concluded that, to avoid neuronal damage, it is necessary to intervene when the fetal condition deteriorates to the extent that overshoot acceleration in FHR occurs.

Three pregnant women with fetal ascites were admitted to our hospital at 30, 32 and 33 weeks of gestation on diagnosis by sonogram. On sonogram, all the fetuses were initially with marked ascites but without any pleural or pericardial effusion or subcutaneous edema. Under observation by sonogram and cardiotocogram, obstetrical intervention was performed by cesarean section in all 3 cases; at 31 weeks of gestation due to notably increased ascites, at 32 weeks due to fetal distress and at 35 weeks due to development of subcutaneous edema, respectively, and the subsequent courses were uneventful. On the other hand, in our precious experience with 21 cases of NIHF over a period of 7 years, only 2 babies with NIHF have been born alive. We think fetal ascites should be considered an initial sign of NIHF and the successful outcome of these three cases were due to early diagnosis by frequent sonographic examination.

To test applicability of monitoring regional spinal cord blood flow by measuring regional epidural blood flow, both were simultaneously measured by the hydrogen clearance method in response to changes in PaCO2 and mean arterial blood pressure in rats anesthetized with pentobarbital. An excellent correlation was found between regional spinal and epidural blood flow over a physiological range of PaCO2 (27.8-66.7 torr) and blood pressure (30-130 torr), while a poor correlation was demonstrated between regional spinal or epidural blood flow and regional muscle blood flow in response to these same physiological parameters. Moreover, the rates of change in regional spinal and epidural blood flows were almost the same in response to both PaCO2 and blood pressure. These results suggest that both regional spinal and epidural blood flow are regulated by a similar mechanism and suggest that regional spinal cord blood flow can be monitored by regional epidural blood flow.

A hot-water extract from the seed of Plantago asiatica showed a potent inhibitory activity against jack bean alpha-mannosidase, and a flavanone glucoside, plantagoside, was isolated as the inhibitor. Plantagoside was a specific inhibitor for jack bean alpha-mannosidase (IC50 at 5 microM) and appeared to be a non-competitive inhibitor of the enzyme. Whereas, negligible or weak inhibitory activities were observed for beta-mannosidase, beta-glucosidase, and sialidase tested. Plantagoside also inhibited alpha-mannosidase activities in mouse liver lysosomal and microsomal fractions, and the enzyme inhibitory activity in microsomal fraction was enhanced in the presence of glucosidase inhibitor, castanospermine. Plantagoside suppressed antibody response to sheep red blood cells and concanavalin A induced lymphocyte proliferation which was measured by [3H]thymidine incorporation.

An autopsied case of the Lesch-Nyhan syndrome did not indicate the specific pathological features except delayed physical development. 2. Xanthine calculi caused by allopurinol administration scattered in the kidneys, brain, thymus, and thyroid glands, but its excretion into urine was not observed during his life. 3. Activities of hypoxanthine-guanine phosphoribosyltransferase (HGPRT) in various tissues indicate complete deficiency, but HGPRT in liver was normal.

Liposome surfaces were modified with normal rabbit IgG or rabbit anti-sheep erythrocyte IgG by adsorption or by coupling with a water soluble cross-linking reagent, 1-ethyl-3-(3-dimethylaminopropyl)carbodiimide (EDCI). Modification with normal IgG reduced the non-specific binding of the liposomes to erythrocytes. The immunological binding capacity of the immune IgG on liposomes was better for the chemically coupled preparation than for the adsorbed one. Complement dependent cytolytic activity of the immune IgG towards target erythrocytes was diminished as a consequence of liposome binding by either method, but to a lesser extent in the coupling method. Antibody coating by EDCI thus provides liposomes which can bind effectively to antigenic target cells.

Fetal blood sampling by cordocentesis was performed in 85 cases of fetuses with either nonimmune hydrops fetalis, anomalies, intrauterine growth retardation, maternal immunological disorders, Rh incompatibility, hydronephrosis, polyhydramnios, maternal preeclampsia or fetal tumor. Fetal heart rate monitoring was also recorded before cordocentesis. We evaluated the usefulness of the nonstress test (NST) with regard to fetal hematologic gas profiles including pH, pO2 and pCO2. When the acceleration was classified into reactive and nonreactive group, pH and pO2 were significantly higher in the reactive group than in the nonreactive group. pCO2 was significantly lower in the former than the latter. However, the sensitivity and specificity of the reactive NST predicting fetal umbilical venous pO2 over 20 mm Hg was 79.7 and 68.7%, respectively. The false-positive rate of nonreactive NST predicting hypoxemia below 20 mm Hg pO2 was 44%. As a consequence, we cannot always rely on the NST, and should perform fetal blood sampling to assess fetal condition for further management.