Cloning, expression and characterization of a murine-human chimeric antibody with specificity for pre-S2 surface antigen of hepatitis B virus

The cloning, expression and characterization of a murine-human chimeric antibody with specificity for the pre-S2 surface antigen (Ag) of hepatitis B virus (HBV) is described. The heavy and light chain variable region (VH and VL) genes encoding the murine monoclonal antibody (mAb) were isolated and combined with human γ 1 and κ constant region genes, respectively. The expression vectors containing the chimeric heavy and light chain genes were sequentially electroporated into murine Sp2/0 hybridoma cells and transfectomas secreting chimeric antibody were isolated. The chimeric antibody was purified and characterized by ELISA, Western analysis and competition immunoassay, demonstrating that the transfectoma functionally express and secrete murine-human chimeric antibody which retained the specificity and affinity of the parental murine mAb.     

The distribution of fetal nuchal translucency thickness in normal Korean fetuses

The aim of present study was to establish normative data for the distribution of nuchal translucency (NT) thickness in normal Korean fetuses. The data were collected from pregnant women with singleton pregnancies in whom fetal ultrasound was performed and the fetal NT thickness was measured between 11 and 14 weeks of gestation. Among them, a total of 2,577 fetuses with a known normal outcome were included in this study. The distribution of multiple of median (MoM) values of the NT thickness with crown-rump length (CRL) in 10-mm intervals and the 95th percentile of MoM were calculated with the linear regression method. The present study showed that NT measurements increase with increasing CRL and a false positive rate increases with increasing gestational age. Therefore, a fixed cut-off point through the first trimester was not appropriate and each NT measurement should be examined according to the gestational age. The present study offers normative data of the fetal NT thickness in a Korean population, which can be used as reference for screening chromosomal aberrations or other congenital abnormalities in the first trimester.     

The complete nucleotide sequence and genome organization of odontoglossum ringspot tobamovirus RNA

Summary The complete nucleotide sequence of the genomic RNA of odontoglossum ringspot tobamovirus (ORSV) was determined. The RNA genome of ORSV is 6618 nucleotides long and contains five open reading frames (ORFs 1 to 5) coding for proteins of Mr 126 K, 181 K, 34 K, 18 K and 52 K, respectively. This is the longest RNA of the known viruses of theTobamovirus genus. The sequences of the ORSV RNA encoded proteins exhibit high homology to the proteins of the members of theTobamovirus genus. The genomic organization and sequence analysis showed that ORSV is more closely related to tobacco mild green mosaic virus (TMGMV), pepper mild mottle virus (PMMV), tomato mosaic virus (ToMV) and TMV than to cucumber green mottle mosaic virus (CGMMV) and sunn-hemp mosaic virus (SHMV).The nucleotide sequence reported in this paper will appear in the EMBL, GenBank and DDBJ nucleotide sequence databases under the accession number X82130.     

Implantation of bone marrow mononuclear cells using injectable fibrin matrix enhances neovascularization in infarcted myocardium

Neovascularization may improve cardiac function and prevent further scar tissue formation in infarcted myocardium. A number of studies have demonstrated that bone marrow-derived cells have the potential to induce neovascularization in ischemic tissues. In this study, we hypothesized that implantation of bone marrow mononuclear cells (BMMNCs) using injectable fibrin matrix further enhances neovascularization in infarcted myocardium compared to BMMNC implantation without matrix. To test this hypothesis, infarction was induced in rat myocardium by cryoinjury. Three weeks later, rat BMMNCs were mixed with fibrin matrix and injected into the infarcted myocardium. Injection of either BMMNCs or medium alone into infarcted myocardium served as controls. Eight weeks after the treatments, histological analyses indicated that implantation of BMMNCs using fibrin matrix resulted in more extensive tissue regeneration in the infarcted myocardium compared to BMMNC implantation without matrix. Examination with fluorescence microscopy revealed that cells labeled with a fluorescent dye prior to implantation survived in the infarcted myocardium at 8 weeks of implantation. Importantly, implantation of BMMNCs using fibrin matrix resulted in much more extensive neovascularization in infarcted myocardium than BMMNC implantation without matrix. The microvessel density in infarcted myocardium was significantly higher (p < 0.05) when BMMNCs were implanted using fibrin matrix (350 +/- 22 microvessels/mm2) compared to BMMNC implantation without matrix (262 +/- 13 microvessels/mm2) and medium injection (76 +/- 9 microvessels/mm2). In addition, average internal diameter of microvessels was significantly larger (p < 0.05) in BMMNC implantation with fibrin matrix group (14.6 +/- 1.2 microm) than BMMNC implantation without matrix group (10.2 +/- 0.7 microm) and medium injection group (7.3 +/- 0.5 microm). These results suggest that fibrin matrix could serve as a cell implantation matrix that enhances neovascularization efficacy for myocardial infarction treatment.     

Isolation and characterization of human and rabbit sperm tail fibrous sheath

Using mechanical and chemical dissection methods, fibrous sheath was isolated both from normal ejaculated human spermatozoa and from rabbit cauda epididymal spermatozoa. The same techniques did not produce a pure preparation of fibrous sheath from ejaculated rabbit spermatozoa, suggesting that further cross-linking and stabilization of sperm structures occurs in response to components of the seminal plasma. The isolation procedures were monitored by phase contrast microscopy and the purity of the fibrous sheath was verified by electron microscopy. Sodium dodecyl sulphate-polyacrylamide gel electrophoresis (SDS-PAGE) of isolated human fibrous sheath revealed at least 14 protein bands of which the most intensely stained were of molecular weight 84, 72, 66.2, 57, 32 and 28.5 kDa. The rabbit fibrous sheath revealed at least 10 protein bands, of which the most intensely stained were 35.2, 32.7 and 28.5 kDa. The amino acid composition of the purified fibrous sheath from human and rabbit spermatozoa was similar, being high in aspartic acid and/or asparagine and glutamic acid and/or glutamine, serine, alanine, leucine, lysine and glycine, but low in histidine, tyrosine and isoleucine. This composition is similar to that reported for the rat and suggests that mammalian sperm tail fibrous sheaths are composed of similar types of proteins, although there are apparent differences in protein components between species.      

Characterization of erythromycin-resistant Streptococcus pneumoniae in Korea, and In Vitro activity of telithromycin against erythromycin-resistant Streptococcus pneumoniae

To characterize the phenotypes and genotypes of erythromycin-resistant clinical isolates of Streptococcus pneumoniae in Korea and to evaluate the in vitro activity of telithromycin against these erythromycin-resistant isolates, we tested a total of 676 isolates of S. pneumoniae collected from 1997 to 2002 in a tertiary hospital in Seoul, Republic of Korea. MICs for erythromycin and telithromycin were determined by the agar dilution method. The macrolide resistance phenotypes of erythromycin-resistant isolates were determined by the erythromycin- clindamycin-rokitamycin triple disk (ECRTD) and MIC induction tests, whereas their macrolide resistance genotypes were determined by PCR for the erm(B), erm(A), subclass erm(TR), and mef genes. To discriminate between mef(A) and mef(E), PCR-restriction fragment length polymorphism (RFLP) analyses were performed. Of the 676 S. pneumoniae isolates, 459 (67.9%) were resistant to erythromycin. Of the 459 erythromycin-resistant isolates, 343 (74.7%) were assigned to the cMLS phenotype, 48 (10.4%) to the iMcLS phenotype, 4 (0.9%) to the iMLS phenotype, and 64 (14.0%) to the M phenotype. The erm(B) gene was detected in 251 (54.6%) isolates, the mef gene was detected in 64 (14.0%), and both the erm(B) and mef genes were detected in 144 (31.4%) isolates. All of the mef genes detected were identified as mef(E). Of the 459 erythromycin- resistant isolates, all but one were susceptible to telithromycin. The MIC(50)/MIC(90) to telithromycin of isolates carrying erm(B), mef(E), and both genes was 0.06/0.5 microg/ml, 0.03/0.125 microg/ml, and 0.5/1.0 microg/ml, respectively. Although the MICs of telithromycin for the erythromycin-resistant isolates varied according to genotype, telithromycin was very active against these erythromycin-resistant S. pneumoniae.     

Ultrastructural localization of phosphoglycerate kinase in adult<Emphasis Type="Italic"> Clonorchis sinensis</Emphasis>

Phosphoglycerate kinase (PGK) is an enzyme that produces one ATP molecule in the glycolytic pathway. Clonorchis sinensis is largely dependent on glycolysis for energy production. We performed immunoelectron microscopy on adult C. sinensis by using mouse immune serum raised against recombinant C. sinensis PGK. A high density of gold particles was found in the microvilli of the intestinal epithelium and in lamellae of the sperm duct. PGK was common in the somatic cells of intra-uterine eggs and in excreted products. It was localized with moderate intensity in muscular fibers of the subtegumental muscle layer, and in the myoepithelia of the intestine and excretory bladder. We suggest that PGK plays an essential role in C. sinensis energy production for movement via muscle contraction.     

Acute and Subacute Toxicity of Aspilia Africana Leaves

This study was designed to evaluate the toxicity of the aqueous extract of Aspilia africana leaves. Oral doses of 500 mg/kg and 1000 mg/kg were administered for 28 days to rats after every 2 days for sub-acute toxicity. For acute toxicity, 5 doses of 2, 4, 8, 12 and 16g/Kg body weight were investigated in mice. The control groups consisted of mice or rats administered with distilled water. The signs of toxicity fluctuated lightly from one mammal to another throughout the experiment. The liver, kidneys and heart weight of rats revealed no significant differences between the test groups and the control. The results indicated that the medium lethal dose (LD₅₀) was found to be greater in females than males with an average of 6.6g/Kg body weight for both sexes. Regardless of the significant differences observed at certain points in some biochemical parameters (ALT, AST, ALP, Creatinine and Glutathione); none showed any linear dose responsiveness. On the other hand, most of the parameters investigated were found to be gender dependent. These results suggested that A Africana can be classified among substances with low toxicity.