Development of enzyme immunoassay for Platonin (NK 19)

A sensitive and specific enzyme immunoassay for an immunomodulator PLATONIN (NK 19) has been developed and applied to the evaluation of pharmacokinetics in human and rats. NK 19 analog, succinyl NK 19 has been synthesized and coupled to bovine serum albumin. This conjugate is immunogenic and when injected into rabbits antibodies of high affinity for NK 19 were obtained. Some light decomposed products of Platonin were of lower cross-reactivity with these antibodies. For competitive reactions, these antibodies were incubated with a mixture of succinyl NK 2900-beta-D-galactosidase conjugate and standard NK 19 or sample. Then, antibody-bound enzyme-hapten were separated from free one using anti-rabbit IgG immobilized on polystyrene balls. Activity of the enzyme on the solid phase was fluorometrically determined. The present immunoassay allows detection of as low as 10 pg of NK 19 in the biological fluids using 50 microliters of the samples. The accuracy and reproducibility were also quite satisfactory. By use of this assay, plasma levels of NK 19 after a single oral administration of 5 mg or 10 mg/body to human volunteers or intravenous injection of 2 micrograms/kg to rats could be determined.     

Second meiotic nondisjunction of the rearranged chromosome in a familial reciprocal 5/13 translocation

We describe a 20-week-gestation male fetus with partial dup(5p) and proximal dup(13q), 47,XY,t(5;13)(p15;q21), + der(13)t(5;13)(p15;q21) mat. This finding is attributable to second meiotic nondisjunction of the rearranged chromosome in a maternal balanced reciprocal translocation. To the best of our knowledge, there have been only 3 previous reports of a similar error in the segregation of the rearranged chromosomes. For the first time evidence has been given that this unusual segregation is due to maternal second meiotic nondisjunction, using QFQ banding heteromorphisms. Second meiotic malsegregation should be taken into account in the consideration of reproductive problems in carriers of balanced translocations.     

Metabolic fate of bufalin in rats]

In an attempt to evaluate the metabolic fate of "Kyushin", which is a traditional medicine containing Toad venom, a 3H-labeled compound of bufalin, which is one of the main active components, has been synthesized. The metabolic fate of the 3H-bufalin was studied after its single and repeated oral administration in rats. After a single administration of the 3H-bufalin (20 micrograms/kg), the radioactivity in the blood reached a maximum level at 15 min. The radioactivity in the blood declined in a triphasic manner with half-life times of 18 min, 2.6 and 86 h. Within 24 h after the single administration, the excretion of the radioactivity into the urine and feces amounted to 1.3% and 81% of the administered dose, respectively. Tissue radioactivity was higher in the stomach, small intestine, liver, lung, kidney and pancreas, while the radioactivity in other tissues was lower than that in blood. Radioactivity disappeared rapidly from any tissues. In case of repeated administration for 14 d, the disposition of radioactivity was almost same as the result after a single dosing, and radioactivity scarcely remained in any tissues after the last administration.     

Hepatocyte growth factor gene therapy reduces ventricular arrhythmia in animal models of myocardial ischemia

It was recently reported that gene therapy using hepatocyte growth factor (HGF) has the potential to preserve cardiac function after myocardial ischemia. We speculated that this HGF gene therapy could also prevent ventricular arrhythmia. To investigate this possibility, we examined the antiarrhythmic effect of HGF gene therapy in rat acute and old myocardial infarction models. Myocardial ischemia was induced by ligation of the left descending coronary artery. Hemagglutinating virus of Japan (HVJ)-coated liposome containing HGF genes were injected directly into the myocardium fourteen days before programmed pacing. Ventricular fibrillation (VF)was induced by programmed pacing. The VF duration was reduced and the VF threshold increased after HGF gene therapy ( p< 0.01). Histological analyses revealed that the number of vessels in the ischemic border zone was greatly increased after HGF gene injection. These findings revealed that HGF gene therapy has an anti-arrhythmic effect after myocardial ischemia.     

Detection of human cytomegalovirus DNA in perilymph of patients with sensorineural hearing loss using real-time PCR

Although cytomegalovirus (CMV) has been detected in the inner ear fluid of patients who succumbed to the complications of symptomatic congenital CMV infection, it has not been detected in the inner ear fluid of living patients. In this study, real-time polymerase chain reaction (PCR) was used to measure CMV DNA in clinical samples (including perilymph) collected from five patients with deafness. In case 1, diagnosed as a symptomatic congenital CMV infection, 3 copies/microl of CMV DNA were detected in perilymph, although no viral DNA was detected in peripheral blood mononuclear cells (PBMCs) or urine samples. In case 4, a suspected asymptomatic congenital CMV infection, 36 copies/microg of CMV DNA were detected in PBMCs, but neither perilymph nor urine contained viral DNA. Likewise, in case 5, a case of deafness of unknown origin, 48 copies/microg of CMV DNA were detected in the PBMCs, but none in the perilymph or urine. CMV DNA was not detected in the samples obtained from the remaining two cases with deafness of unknown etiology. To our knowledge, this is the first report to detect CMV DNA in an inner ear sample obtained from a living human subject.     

DNA MUTATION INDUCED IN THE SEQUENCE UPSTREAM OF THE SECRETED MYU C-TERMINAL CODING SEQUENCE BY ULTRAVIOLET IRRADIATION IN THE CELL LINE OF BLOOM'S SYNDROME

Selective IgM deficiency is commonly found in Bloom's syndrome (BS). We reported that membrane-bound μ (μ m) mRNA was well transcribed but secreted μ (μ s) mRNA was not, although there was no mutation or deletion in the sequence including the (is C-terminal coding sequence in the patients with BS. Furthermore, we have shown previously, preferential damage to IgM production by ultraviolet (UV) irradiation of the cells of the patient. In the study described here, mutation in the sequence which is upstream of the 5’ end of the μ s C-terminal coding sequence was induced by UV irradiation in the lymphoblastoid cell line (LCL) of BS patient. These results suggest that abnormal repair of DNA damage is present in this LCL, and that preferential damage to IgM production by UV irradiation in this LCL may be due to the abnormal repair of DNA damage.     

Diffuse pulmonary hamartoma: a case report

Pulmonary hamartoma usually occurs as a benign, well-circumscribed single nodule in the lung parenchyma. We report on a unique case of hamartoma that extended along the bronchial tree, formed endobronchial polypoid lesions, and expanded into the lung parenchyma. The patient, a 48-year-old man, was admitted with dyspnea and chest pain. A transbronchial biopsy was performed on a tumorous lesion that was diagnosed histologically as a hamartoma. As this lesion was found to be growing diffusely along the bronchial tree, a left pneumonectomy was performed. Gross examination showed that yellowish soft tissue had surrounded the bronchial tree and extended into the lung parenchyma. Histologically, the lesion contained mainly mature adipose tissue, cartilage, and muscle tissue with a minor component of short spindle cells in a myxomatous matrix. The patient was diagnosed as having diffuse pulmonary hamartoma.     

Functional analysis of four naturally occurring variants of human constitutive androstane receptor

The human constitutive androstane receptor (CAR, NR1I3) is a member of the orphan nuclear receptor superfamily that plays an important role in the control of drug metabolism and disposition. In this study, we sequenced all the coding exons of the NR1I3 gene for 334 Japanese subjects. We identified three novel single nucleotide polymorphisms (SNPs) that induce non-synonymous alterations of amino acids (His246Arg, Leu308Pro, and Asn323Ser) residing in the ligand-binding domain of CAR, in addition to the Val133Gly variant, which was another CAR variant identified in our previous study. We performed functional analysis of these four naturally occurring CAR variants in COS-7 cells using a CYP3A4 promoter/enhancer reporter gene that includes the CAR responsive elements. The His246Arg variant caused marked reductions in both transactivation of the reporter gene and in the response to 6-(4-chlorophenyl)imidazo[2,1-b][1,3]thiazole-5-carbaldehyde O-(3,4-dichlorobenzyl)oxime (CITCO), which is a human CAR-specific agonist. The transactivation ability of the Leu308Pro variant was also significantly decreased, but its responsiveness to CITCO was not abrogated. The transactivation ability and CITCO response of the Val133Gly and Asn323Ser variants did not change as compared to the wild-type CAR. These data suggest that the His246Arg and Leu308Pro variants, especially His246Arg, may influence the expression of drug-metabolizing enzymes and transporters that are transactivated by CAR.     

Basic imaging properties of a computed radiographic system with photostimulable phosphors

We measured the characteristic curve, modulation transfer function (MTF), and the Wiener spectrum of a commercially available computed radiographic (CR) system with photostimulable phosphor plate (imaging plate, IP). The characteristic curve (system response) obtained by an inverse-square x-ray sensitometry showed a wide dynamic range (order of 10(3) in maximum). The slit technique was employed to determine the MTF's, such as IP MTF, presampling MTF including the unsharpness of the detector (IP) and the blurring effect of the sampling aperture, and laser-printer MTF. It was found that the MTF of the standard type of IP was comparable to that of medium-speed screen/film systems. The noticeable degradation of resolution in our CR system, however, occurred at the stage of image data sampling: the presampling MTF was inferior to the IP MTF due to the effect of the scattering and resultant spreading of the incidence laser beam and the emitted luminescence. The noise was characterized by means of digital Wiener spectrum using uniformly exposed noise data. Exposure ranges could be separated into different sections depending upon the noise sources, such as quantum mottle at low exposure and system structure noise at high exposure.