Biomedical subjects
Publications and source records attributed to T Maruyama.
A benzoyl group was introduced onto the 3'-hydroxyl group of 6-chloropurine riboside by treatment with benzoic anhydride in the presence of a base in aqueous solution. The product (3b) was converted to 9-(2,3-Di-deoxy-2-fluoro-beta-D-threo-pentofuranosyl)adenine (1, FddA) in 6 steps, including radical deoxygenation.
Idiopathic left ventricular (LV) tachycardia usually exhibits right bundle branch block morphology. There are only a few sporadic cases that exhibit left bundle branch block (LBBB) morphology. We report a patient whose QRS complex during ventricular tachycardia (VT) was relatively narrow (100 msec) and exhibited LBBB (precordial R wave transition between V3 and V4) and a normal frontal plane axis. This VT was ablated successfully by radiofrequency current applied to the LV upper septum, where the earliest endocardial activation was recorded.
OBJECTIVE: The reason that precore negative mutants (e-minus DNA) gradually become predominant in some patients during chronic hepatitis B virus infection is not clear. Theoretically, as long as both e-plus and e-minus DNA share the same epitopes in the core region, HBcAg-specific cytotoxic T lymphocytes (CTLs) cannot distinguish between the target peptides expressed by e-plus and e-minus DNA. Therefore, e-minus DNA may be accompanied by additional mutations in the core region, which may affect cytotoxic T lymphocyte recognition. To examine this possibility, the sequences of the precore and the entire core region of the hepatitis B virus genome were analyzed from paired serum samples in CH-B patients who experienced HBeAg to anti-HBe seroconversion (SC). METHODS: Patients were divided into two groups. Group A patients (n = 17) genome-converted to e-minus DNA in the precore region, which abolished HBeAg secretion within 3-4 yr after SC. Group B patients (n = 16) retained precore wild-type DNA for more than 3-4 yr after SC. To investigate the impact of the emergence of precore mutant type DNA on liver injury, alanine aminotransferase (ALT) levels were also examined. RESULTS: ALT flares were more severe among patients in group A than in group B. The average mean ALT level during the HBeAg negative phase of chronic infection was 54 +/- 38 in group A and 28 +/- 24 in group B. The average maximal ALT level during the HBeAg negative phase was 235 +/- 249 in group A and 83 +/- 106 in group B. Furthermore, all 17 patients in group A developed new core mutants during genome conversion. The average number of mutations in the core gene was 0.9 +/- 1.2 before genome conversion (e-plus DNA dominant phase) and increased to 2.8 +/- 1.3 for the 3-4 yr during genome conversion (e-minus DNA dominant phase). In contrast, only 56% (nine of 16) of patients in group B developed new core mutations after the loss of HBeAg. The average number of mutations in the core gene was 1.8 +/- 1.3 before SC (HBeAg-positive and e-plus DNA dominant phase), and decreased to 1.1 +/- 1.1 for 3-4 yr after seroconversion (anti-HBe-positive and e-plus DNA dominant phase). CONCLUSIONS: These data indicate that the emergence of a predominant precore negative genotype late in chronic hepatitis B virus infection is associated with the selection of additional mutations in the core gene, as well as with liver injury.
PROBLEM: There is no reliable laboratory test available to diagnose immunologically mediated miscarriages. We investigated the clinical significance of maternal anti-paternal leukocyte antibodies by flow cytometry after alloimmunization. METHOD OF STUDY: The flow cytometry crossmatch (FCXM) was performed in 158 patients with a history of three or more unexplained first-trimester miscarriages without live birth. After negative FCXM patients were immunized, subsequent pregnancy outcomes and FCXM results were followed. RESULTS: Of 112 subsequent pregnancies, 83 of 100 (83.0%) FCXM-positive patients after immunotherapy had successful pregnancy outcomes, whereas seven of 10 (70.0%) FCXM-negative patients had miscarriages (P = 0.0001). The percent live birth ratio was 2.77 (CI, 1.07-7.16; P=0.0001) for FCXM-positive patients compared to FCXM-negative patients. The calculated predictive value showed that 75.6% of FCXM-negative patients would have subsequent miscarriages. CONCLUSIONS: Positive FCXM is closely associated with successful pregnancy outcome following immunotherapy. We propose that FCXM might be included in the routine laboratory tests for the management of recurrent miscarriages.
Regulation of the kinetics of intracellular Ca(2+) signals with a novel, membrane-penetrable, inositol 1,4,5-trisphosphate (InsP(3)) receptor/Ca(2+) channel modulator, 2-amino-ethoxydiphenyl borate (2APB), has been investigated using patch-clamp, whole-cell recording to monitor Ca(2+)-activated Cl(-) currents in single isolated pancreatic acinar cells. 2APB itself fails to evoke a detectable current response but it dramatically changes the kinetics of agonist-induced Ca(2+) release from pulsatile spikes to long-lasting, huge Ca(2+) waves, suggesting that 2APB coordinates local Ca(2+) release to generate global Ca(2+) signals. The regulation by 2APB can be elicited by internal perfusion of InsP(3) in a concentration-dependent manner, indicating that this regulation is not mediated through membrane receptors or G protein signal transduction. The InsP(3) receptor blocker heparin, but not the ryanodine-sensitive receptor blockers ruthenium red or ryanodine, abolishes 2APB-mediated regulation of Ca(2+) release. This results also suggest that 2APB effects are mediated through InsP(3) receptors. 2APB substantially modifies single inward Cl(-) current pulse evoked by the photolytic release of caged InsP(3) but not by caged Ca(2+). These data indicate that 2APB-induced regulation is mediated neither by Ca(2+)-induced Ca(2+) release nor by affecting Cl(-) channel activity directly. We conclude that 2APB regulates the kinetics of intracellular Ca(2+) signals, represented as the change in the Ca(2+) oscillation patterns from brief pulsatile spikes to huge, long-lasting Ca(2+) waves. Moreover, this regulation seems to be mediated through InsP(3)-sensitive Ca(2+) pools. 2APB may act as a novel, useful pharmacological tool to study the genesis of intracellular Ca(2+) signals.
The distribution of culturable hyperthermophiles was studied in relation to environmental conditions in the Kubiki oil reservoir in Japan, where the temperature was between 50 and 58 degrees C. Dominant hyperthermophilic cocci and rods were isolated and shown to belong to the genera Thermococcus and Thermotoga, respectively, by 16S rDNA analyses. Using the most-probable-number method, we found that hyperthermophilic cocci were widely distributed in several unconnected fault blocks in the Kubiki oil reservoir. In 1996 to 1997, their populations in the production waters from oil wells were 9.2 x 10(3) to 4.6 x 10(4) cells/ml, or 10 to 42% of total cocci. On the other hand, hyperthermophilic rods were found in only one fault block of the reservoir with populations less than 10 cells/ml. Dominant Thermococcus and Thermotoga spp. grew at reservoir temperatures and utilized amino acids and sugars, respectively, as sole carbon sources. While organic carbon was plentiful in the environment, these hyperthermophiles were unable to grow in the formation water due to lack of essential nutrients. Concentrations of some organic and inorganic substances differed among fault blocks, indicating that the movement of formation water between fault blocks was restricted. This finding suggests that the supply of nutrients via fluid current is limited in this subterranean environment and that the organisms are starved in the oil reservoir. Under starved conditions at 50 degrees C, culturable cells of Thermococcus sp. remained around the initial cell density for about 200 days, while those of Thermotoga sp. decreased exponentially to 0. 01% of the initial cell density after incubation for the same period. The difference in survivability between these two hyperthermophiles seems to reflect their populations in the fault blocks. These results indicate that hyperthermophilic cocci and rods adapt to the subterranean environment of the Kubiki oil reservoir by developing an ability to survive under starved conditions.
We describe a novel nuclear factor called mitotic chromosome-associated protein (MCAP), which belongs to the poorly understood BET subgroup of the bromodomain superfamily. Expression of the 200-kDa MCAP was linked to cell division, as it was induced by growth stimulation and repressed by growth inhibition. The most notable feature of MCAP was its association with chromosomes during mitosis, observed at a time when the majority of nuclear regulatory factors were released into the cytoplasm, coinciding with global cessation of transcription. Indicative of its predominant interaction with euchromatin, MCAP localized on mitotic chromosomes with exquisite specificity: (i) MCAP-chromosome association became evident subsequent to the initiation of histone H3 phosphorylation and early chromosomal condensation; and (ii) MCAP was absent from centromeres, the sites of heterochromatin. Supporting a role for MCAP in G(2)/M transition, microinjection of anti-MCAP antibody into HeLa cell nuclei completely inhibited the entry into mitosis, without abrogating the ongoing DNA replication. These results suggest that MCAP plays a role in a process governing chromosomal dynamics during mitosis.
In chronic renal failure, substances that are effectively excreted in healthy subjects accumulate in serum. These substances, uremic toxins, include a variety of organic acids. It has been reported that a decrease in the bilirubin (BR) binding capacity occurs in the serum of renal failure patients. 3-Carboxy-4-methyl-5-propyl-2-furanpropanoic acid (CMPF) has a high affinity for human serum albumin (HSA) and is a potent inhibitor of the serum protein binding of many drugs. We recently reported that CMPF and BR share the binding site for dicarboxylate molecules on the HSA molecule [Pharm Res 1999;16:916-923]. In this study, in order to confirm whether CMPF is involved in the decrease of BR serum binding capacity in chronic renal failure patients, the total concentrations of uremic toxins, CMPF, and indoxyl sulfate (IS) and the free BR concentration in serum from healthy volunteers and renal failure patients were determined. Both total CMPF and IS concentrations correlate with the free BR concentration. However, results from the peroxidase method reveal that IS cannot displace BR under the physiological condition [IS]/[HSA] <1. We, therefore, conclude that CMPF is one of the substances which contribute to the decreased binding capacity of BR in uremic serum.
PGE2 functions as a potent stimulator of bone resorption. The action of PGE2 is thought to be mediated by some PGE receptor subtypes present in osteoblastic cells. In this study, we examined the involvement of PGE receptor subtypes, EP1, EP2, EP3, and EP4, in PGE2-induced bone resorption using specific agonists for the respective EPs. In mouse calvaria cultures, EP4 agonist markedly stimulated bone resorption, but its maximal stimulation was less than that induced by PGE2. EP2 agonist also stimulated bone resorption, but only slightly. EP1 and EP3 agonists did not stimulate it at all. RT-PCR showed that osteoblastic cells isolated from newborn mouse calvaria expressed all of the EPs messenger RNA (mRNA). Both EP2 agonist and EP4 agonist induced cAMP production and the expression of osteoclast differentiation factor (ODF) mRNA in osteoblastic cells. Simultaneous addition of EP2 and EP4 agonists cooperatively induced cAMP production and ODF mRNA expression. In mouse bone marrow cultures, EP2 and EP4 agonists moderately induced osteoclast formation, but the simultaneous addition of the two agonists cooperatively induced it, similar to that by PGE2. In calvaria culture from EP4 knockout mice, a marked reduction in bone resorption to PGE2 was found. In EP4 knockout mice, EP4 agonist failed to induce bone resorption, but EP2 agonist slightly, but significantly, induced bone resorption. These findings suggest that PGE2 stimulates bone resorption by a mechanism involving cAMP and ODF, which is mediated mainly by EP4 and partially by EP2.
Congestive heart failure (CHF) related to Sjögren's syndrome is extremely rare. This report concerns a patient who presented with CHF and severe thrombocytopenia (5,000/microl). Serum concentrations of K, Mg and digitoxin were 3.2mmol/L, 1.4mg/L and 57.2ng/ml, respectively. Digitoxin intoxication was evident, seemingly evoked by hypokalemia, hypomagnesemia, hepatorenal dysfunction and hypothyroidism. The severe thrombocytopenia was considered to have been caused by this intoxication, as it disappeared soon after the digitoxin was discontinued and potassium was supplemented.
Radiofrequency catheter ablation (RFCA) targeting the cavotricuspid isthmus is usually an effective treatment for common atrial flutter (AFL), except in a small subset of patients and the reason for this has yet to be elucidated. The present study investigated the relationship between the outcome of RFCA for common AFL and the anatomy of the right atrium as seen on angiography. Twenty consecutive patients who underwent RFCA for common AFL were divided into 2 groups according to the results of RFCA. Group A comprised 13 patients whose AFL was abolished, fulfilling the criteria of success by the conventional catheter approach, and group B comprised 7 patients whose AFL could not be abolished according to the criteria for success (n=4) or was abolished following an additional superior vena cava approach (n=3). On angiography, the cavotricuspid isthmus was longer (3.5+/-0.5 vs 2.2+/-0.6 cm) and deeper (0.94+/-0.35 vs 0.49+/-0.19 cm) in group B than in group A (both p<0.01). The height of the eustachian valve was also greater in group B than in group A (1.4+/-1.1 vs 0.48+/-0.48 cm, p<0.02). These results suggest that the anatomical structure of the cavotricuspid isthmus affects the outcome of RFCA for common AFL.
Heat shock proteins (HSPs) have been recognized as molecules that maintain cellular homeostasis during changes in the environment. Here we report that HSP90 functions not only in stress responses but also in certain aspects of cellular differentiation. We found that HSP90 showed remarkably high expression in undifferentiated human embryonal carcinoma (EC) cells, which were subsequently dramatically down-regulated during in vitro cellular differentiation, following retinoic acid (RA) treatment, at the protein level. Surprisingly, heat shock treatment also triggered the down-regulation of HSP90 within 48 h at the protein level. Furthermore, the heat treatment induced cellular differentiation into neural cells. This down-regulation of HSP90 by heat treatment was shifted to an up-regulation pattern after cellular differentiation in response to RA treatment. In order to clarify the functions of HSP90 in cellular differentiation, we conducted various experiments, including overexpression of HSP90 via gene transfer. We showed that the RA-induced differentiation of EC cells into a neural cell lineage was inhibited by overexpression of the HSP90alpha or -beta isoform via the gene transfer method. On the other hand, the overexpression of HSP90beta alone impaired cellular differentiation into trophoectoderm. These results show that down-regulation of HSP90 is a physiologically critical event in the differentiation of human EC cells and that specific HSP90 isoforms may be involved in differentiation into specific cell lineages.
Autoantibody against glutamic acid decarboxylase (GADA) is a highly sensitive predictor of insulin-dependency in adult diabetic patients as well as young individuals. A considerable number of diabetics who do not reach the insulin-dependent stage have this antibody. Recently, type 1 diabetes has been thought to be caused by T helper 1 (Thl)-type autoimmunity based on studies in non-obese diabetic mice, but it is still difficult to investigate antigen-specific T-cell function in human type 1 diabetes. We therefore assessed an IgG subclass assay for GADA, which should reflect T-helper function against GAD. Sera from 14 type 1 diabetic patients positive for GADA by radioligand binding assay were tested for the IgG subclass of GADA. The assay was based on an enzyme-linked immunosorbent assay, which showed a good correlation with radioligand binding assay. The sera of all but one of the 14 type 1 diabetic patients (93%) were positive for the IgG1 subclass of GADA. The IgG2 and IgG3 subclasses of GADA were also detected in one diabetic patient each who were also positive for IgG1. The IgG4 subclass was not detected in any of the sera we tested. We concluded that IgG1 is the dominant subclass of GADA in Japanese type 1 diabetic patients.
To observe cellular membranous systems under a light microscope, we modified Mayer's tannic acid-ferric chloride stain method by adding a treatment with hematoxylin after the original procedure. We used the modified tannic acid-ferric chloride (MTA-Fe) stain method to examine kidneys, liver, heart, trachea, epididymides and other organs of rats and dogs. The MTA-Fe stain clearly demonstrated the basement membrane, brush border, basolateral invaginations and cell processes in the kidneys which enabled easy differentiation of the S1 and S3 segments of proximal convoluted tubules. Our technique also demonstrated hepatic cell membranes and bile canaliculi in the liver, cross striations and longitudinal traveling of myofibrils in the heart, cilia of the epithelial cells in the trachea, and stereocilia and terminal bars in the epididymis. The MTA-Fe stain is a convenient method to visualize cellular membranous systems even for light microscopy. The stain has the advantages of using no toxic materials, simple and easy technique, little variation of staining results, and little fading for several months after staining.
We examined the relation ship between the charge of the amino or carboxylic function of a substrate and the substrate recognition by the large neutral amino acid (LNAA) transport carrier, using the in situ brain perfusion technique. Glucose-coupled L-tyrosine (GcpY), which has a free carboxylic function, and 2-(L-tyrosylamide)-2-deoxy-D-glucose (Y-2DG), which has a free amino function were synthesized. The inhibitory effect of GcpY on [3 H]L-tyrosine uptake was larger than that of N-methly-L-phenylalanine or N-acetyl-L-phenylalanine, whereas Y-2DG did not affect it. These results indicate that a free amino group is not required for recognition, provided that the modified amino group is able to take a positive charge. Steric factors appeared to be relatively unimportant.
The deformability of red blood cells (RBCs) is an important rheologic factor in the maintenance of normal blood flow in the microcirculation. Contrary to the well-known relationship between hyperlipidemia and atherosclerosis, the relationship between RBC rheology and the serum lipid profile has remained controversial and obscure. Moreover, the correlation of high-density lipoprotein (HDL)-cholesterol and RBC deformability has not been fully understood. In the present cross-sectional study of 139 apparently healthy subjects, we investigated whole-cell deformability (filterability) of RBCs in relation to the lipid profile, using a nickel mesh filter with 3.2-microm pores. RBC filterability was independent of gender, age and serum levels of low-density lipoprotein (LDL)-cholesterol. The filterability was significantly proportional to the HDL-cholesterol values (r = 0.382, p < 0.01), whereas it was inversely proportional to the triglyceride levels (r = -0.259, p < 0.01). These findings may provide new insight into the role of HDL-cholesterol not only in preventing atherosclerotic progression but also in improving RBC filterability.
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A 33-year-old pregnant woman developed respiratory difficulty with bilateral pleural effusion 31 weeks into gestation. On admission she had an elevated serum level of creatine kinase, but muscle weakness in the extremities was mild. After an immediate and successful Cesarean section, she developed respiratory failure and mechanical ventilation was required. The patient was diagnosed as having polymyositis from the limb muscle biopsy. She was treated with dexamethasone at a dose of 8 mg, methylprednisolone++, 1 g daily for three days, and then prednisolone 60 mg daily. One week later the serum level of creatine kinase was normalized and the patient was weaned from ventilator support. The dose of prednisolone was tapered 10 mg every week and she was discharged two months after delivery without prednisolone. A chest CT scan revealed no interstitial pneumonitis or aspiration pneumonia, so her respiratory failure seemed to be ascribable to polymyositis-related respiratory muscle weakness and pleural effusion. However, severe involvement of respiratory muscles without generalized marked muscle weakness is extremely rare, and pleural complications have usually been described in association with pulmonary parenchymal diseases. During the following two years polymyositis has not recurred in this patient and there have been no data indicative of other overlapping collagen diseases. Reports of polymyositis which occurred during pregnancy are rare, and the pathogenesis and clinical outcome of these patients need to be carefully investigated.