BACKGROUND: Although phospholipase C (PLC)-B3 is thought to be a very important enzyme in intracellular signal transduction, the sophisticated and complicated purification steps make it difficult to obtain sufficient amount of protein to study regulation of its activity by G proteins or other proteins. In order to get large amount of PLC-B3 protein, I employed baculovirus expression system which is known to express large amount of functionally active proteins. METHODS: In order to make recombinant baculovirus which expresses PLC-B3 gene, partial cDNA of PLC-B3 which lacked 51 nucleotides was used to make full length PLC-B3 cDNA. By PCR, 5-end sequence of PLC-B3 was ligated into partial rat PLC-B3 cDNA and later cloned into pVL1393 transfer vector to make recombinant baculovirus. This recombinant baculovirus containing PLC-B3 sequence was used to infect Sf9 insect cells. RESULTS: Infection of Sf9 cells with recombinant baculovirus rendered expression of 152 kDa-PLC-B3 protein, which was confirmed by immunoblot assay and PLC activity assay. CONCLUSION: The whole length PLC-B3 cDNA was expressed in Sf9 cells using baculovirus expression system. By using it, homogeneous enzyme is expected to be purified to study precise activation and regulation mechanisms of PLC-B3.
Animals ; Baculoviridae* ; Clone Cells ; DNA, Complementary ; GTP-Binding Proteins ; Insects ; Nucleotides ; Phospholipases* ; Polymerase Chain Reaction ; Rats ; Sf9 Cells ; Signal Transduction ; Type C Phospholipases

No Abstract Available.
GTP-Binding Proteins* ; Receptors, Cell Surface*

BACKGROUND: The Chinese hamster ovary cells transfected with human TSH receptor cDNA (hTSHR-CHO), expressing functional human TSH receptors, are known to be more sensitive in detection of thyroid stimulating antibodies than FRTL-5 cells. There has been no report on the usefulness of these cells to measure thyroid stimulation blocking antibody (TSBAb) activity which is frequently found in patients with primary myxedema, METHODS: We established the optimal assay condition of measurement of TSBAb using hTSHR-CHO cells, and simultaneously measured TSBAb activities with FRTL-5 cells and with hTSHR-CHO cells in 49 patients with primary myxedema, compared them with their thyrotropin binding inhibitor immunoglobulin (TBII) activities. RESULTS: 1) hTSHR-CHO cells specifically bound bTSH and were stimulated by bTSH in terms of cyclic AMP generation in a dose dependent manner. 2) Myxedema IgG suppressed TSH-stimulated cAMP production of hTSHR-CHO cells in a dose dependent manner reaching plateau at the concentration of I g/L. Normal pooled IgG has no suppressive action at the concentration of less than 1 g/L, but caused significant suppression at the concentration of greater than 1g/L. 3) TSBAb activities measured by hTSHR-CHO cells in 49 patients with primary myxedema were as follows: Four of 25 TBII-negative cases (16%) and 22 of 24 TBII-positive cases (92%) had TSBAb activities. Most of TSBAb positive patients (95%), especially in TBII positive cases, showed very high activities of more than 90%. 4) TSBAb activities measured by hTSHR-CHO cells and those by FRTL-5 cells were both positive in 24 patients (49%), both negative in 18 patients (37%), and were discrepant in 7 patients (14%). The TSBAb activities measured with hTSHR-CHO cells and those measured with FRTL-5 cells were significantly correlated (r=0.71, p< 0.01). 5) Forty five percent of patients with primary myxedema had all of 3 kinds of activities (TBII, hTSHR-CHO cell TSBAb, FRTL-5 cell TSBAb), 37% of them had none of 3 activities and 18% of them had 1 or 2 kinds of activities only. CONCLUSION: The usefulness of hTSHR-CHO cells in measurements of TSBAb activities were confirmed. The TSBAb activities of most patients with primary myxedema measured by hTSHR-CHO cells were concordant with those measured by FRTL-5 cells. However, a small subset of patients (18%) had discrepant results in assays of TSH receptor antibodies according to the differences in TSH receptors (rat, human and porcine) used in assay. Such discrepancy may be explained by heterogeneity in epitopes for blocking TSH receptor antibodies.
Animals ; Antibodies ; Asian Continental Ancestry Group* ; Cricetinae ; Cricetulus* ; Cyclic AMP ; DNA, Complementary ; Epitopes ; Female ; Humans ; Humans* ; Hypothyroidism* ; Immunoglobulin G ; Immunoglobulins ; Immunoglobulins, Thyroid-Stimulating ; Myxedema ; Ovary* ; Population Characteristics ; Receptors, Thyrotropin* ; Thyroid Gland* ; Thyrotropin

No abstract available.
Spondylolisthesis*

No abstract available.
Chorion* ; Chorionic Villi*

Extracorporeal shock wave lithotripsy (ESWL) has proved to be an effective method of treating upper urinary tract calculi in adults. But the application of ESWL in children was less clearly defined. We report on 28 pediatric patients who were treated by ESWL with the Northgate SD-3 lithotriptor between June, 1989 and May, 1994. The patient age ranged from 2 years to 17 years, with an average age of 11.1 years. The conditions known to be associated with stone formation were urinary tract infection in 6 patients (21.4%), vesicoureteral reflux in 1 patient (3.6%) and unknown in other patients. The total average success rate of treatment was 92.9%. ESWL complications required admission and surgical management were not observed. We concluded that ESWL with the Northgate SD-3 lithotriptor is a safe and effective method of treating upper urinary tract calculi in children.
Adult ; Calculi ; Child* ; Humans ; Lithotripsy* ; Shock* ; Urinary Tract ; Urinary Tract Infections ; Vesico-Ureteral Reflux

OBJECTIVE: The aim of our study was to compare the thickness and histopathologic changes in the fetal membrane between premature rupture of membranes (PROM) and intact membrane after delivery. METHODS: In a prospective study involving 31 patients who were divided into 4 groups such as <37 weeks without PROM, <37 weeks with PROM, >or=37 weeks without PROM, and >or=37 weeks with PROM, we measured the thickness of membrane and studied the histopathologic findings in vitro by light microscopy of histological sections. RESULTS: The membrane thickness of <37 weeks with PROM group was thinner (35.9 micrometer) than that (42.3 micrometer) of <37 weeks without PROM group, but there was no statistical significance. The membrane thickness of >or=37 weeks with PROM and >or=37 weeks without PROM were similar (25.6 micrometer, 26.0 micrometer). But the membrane thickness of >or=37 weeks with/without PROM was significantly thinner (25.8 micrometer) compared with that (38.9 micrometer) of <37 weeks with/without PROM. The histopathologic features of PROM positive group was amnionitis with neutrophilic infiltration, focally or diffusely necrotic change of amniotic membrane, separation of amniotic membrane and degeneration of chorionic villi. CONCLUSION: The thickness of fetal membrane between PROM group and intact membrane group was not different but the thickness of fetal membrane between <37 weeks and >or=37 weeks was statistically significant. The histopathologic change of PROM positive group was prominent as amnionitis. Further evaluation will be needed about the relationship between membrane thickness and PROM.
Amnion ; Chorioamnionitis ; Chorionic Villi ; Extraembryonic Membranes ; Female ; Humans ; Membranes* ; Microscopy ; Neutrophils ; Pregnancy ; Prospective Studies ; Rupture*

No abstract available.
Thyroid Gland*

BACKGROUND: An accurate and rapid method for specise identification of coagulase negative staphylococci(CNS) has been increasingly necessary for the clinical significance and planning the management of patients with staphylococcal infections. Recently, it has been reported that there is a highly conserved area on their 60KDa heat shock protein(HSP60) gene sequences between the interspecies of CNS and it can be amplified by a set of universal degenerate primer. This led us our attention to focus on whether the PCR-based RFLP method using Mse / restriction enzyme could be a useful tool for the species identification of CNS. METHODS: In the present study, we performed PCR-based RFLP analysis using a set of degenerate primers covering HSP60 and Mse / restriction enzyme on the reference strains and 25 clinical isolates(10 of S. epidermidis, 10 of S. haemolyticus, 4 of S. lugdunensis and 1 of S. warneri) which were previously identified by the API-STAPH, Vitek GPI card and/or with conventional biochemical test. RESULT: All the seven reference strains revealed that each strain has a distinct electrophoresed band patterns with combination of different number (up to 8) and size of fragments. And these distinct band patterns showed remarkable concordance with the seven reference strains and 25 clinical isolates. CONCLUSION: These result strongly suggest that the PCR-RFLP method using degenerate primers covering the HSP60 gene and Mse / digestion enzyme offer a convenient and accurate tool for species-specific identification of CNS.
Chaperonin 60* ; Coagulase* ; Digestion ; Heat-Shock Proteins* ; Hot Temperature* ; Humans ; Polymerase Chain Reaction ; Polymorphism, Restriction Fragment Length ; Shock ; Staphylococcal Infections

Five cases of histologically proven polymorphic reticulosis were examined with computed tomography(CT). CT findings were mucosal thickening along the septal and lateral walls of the nasal cavities(n=4), obliteration of the contour of the nasopharynx(n=4), involvement of the paranasal sinuses (n=2), destruction of the nasal septum and/or sinus walls(n=3) and mass in the palate, tonsil or neck (n=1). CT examination was helpful in determining the extent of the disease in the nasal cavity and paranasal sinuses. However, lesions in the palate and tonsils could not be easily evaluated with CT. CT findings of polymorphic reticulosis are nonspecific and granulomatous diseases may show similar CT findings.
Granuloma, Lethal Midline* ; Nasal Cavity ; Nasal Septum ; Neck ; Palate ; Palatine Tonsil ; Paranasal Sinuses