Determination of the parent of origin in nine cases of prenatally detected chromosome aberrations found after intracytoplasmic sperm injection

Prenatal cytogenetic analysis of 71 fetuses conceived by intracytoplasmic sperm injection (ICSI) resulted in the detection of nine (12.7%) chromosome aberrations including two cases of 47,XXY, four cases involving a 45,X cell line and three autosomal trisomies. Molecular analysis of the parental origin of the deleted or supernumerary chromosome was performed by using polymorphic microsatellite markers. Six cases involving a sex chromosome abnormality were found to be of paternal origin while the two trisomic cases that could be analysed were of maternal origin. Two cases involved the same infertile couple who had two consecutive ICSI pregnancies terminated because of a chromosome abnormality. The replaced embryos in both cases originated from a single batch of ICSI fertilized oocytes of which part was used to initiate the first pregnancy and part was cryopreserved and used to initiate the second pregnancy.      

High-performance liquid chromatographic determination of trimebutine and its major metabolite, N-monodesmethyl trimebutine, in rat and human plasma

A rapid, selective and very sensitive ion-pairing reversed-phase HPLC method was developed for the simultaneous determination of trimebutine (TMB) and its major metabolite, N-monodesmethyltrimebutine (NDTMB), in rat and human plasma. Heptanesulfonate was employed as the ion-pairing agent and verapamil was used as the internal standard. The method involved the extraction with a n-hexane-isopropylalcohol (IPA) mixture (99:1, v/v) followed by back-extraction into 0.1 M hydrochloric acid and evaporation to dryness. HPLC analysis was carried out using a 4-microm particle size, C18-bonded silica column and water-sodium acetate-heptanesulfonate-acetonitrile as the mobile phase and UV detection at 267 nm. The chromatograms showed good resolution and sensitivity and no interference of plasma. The mean recoveries for human plasma were 95.4+/-3.1% for TMB and 89.4+/-4.1% for NDTMB. The detection limits of TMB and its metabolite, NDTMB, in human plasma were 1 and 5 ng/ml, respectively. The calibration curves were linear over the concentration range 10-5000 ng/ml for TMB and 25-25000 ng/ml for NDTMB with correlation coefficients greater than 0.999 and with within-day or between-day coefficients of variation not exceeding 9.4%. This assay procedure was applied to the study of metabolite pharmacokinetics of TMB in rat and the human.     

Histamine synthesis by non-mast cells through mitogen-dependent induction of histidine decarboxylase.

Culture of spleen cells of C57BL/6 mice led to a spontaneous increase in activity of histidine decarboxylase (HDC) in the cell and the medium. Concomitantly histamine increased in the cells and, especially, in the medium. Addition of concanavalin A (Con A) or lipopolysaccharide (LPS) to the culture enhanced these processes. There was also a significant Con A-dependent increase in HDC activity and histamine biosynthesis in the culture of spleen cells of genetically mast-cell deficient W/Wv mice. Peritoneal macrophages of C57BL/6J mice had constitutively 11-19-fold as much HDC as T and B lymphocytes when compared on the basis of same number of cells. Con A had no effect on HDC activity when the macrophages were cultured alone. However, co-culture with T lymphocytes, separated from macrophages by a millipore filter membrane (pore size, 0.45 micron), rendered the macrophages responsive to Con A, leading to a notable increase in HDC activity in the cell. Addition of LPS caused a small but significant increase in HDC activity in macrophages, even when the cells were cultured alone. Co-culture with T or B cells enhanced the LPS-dependent increase in HDC activity in macrophages. In contrast, the HDC activity in T and B lymphocytes did not change essentially in the presence of any of these mitogens, even when they were co-cultured with macrophages. These results suggest that histamine is synthesized by non-mast cells through HDC.     

Clinical prognostic values of vascular endothelial growth factor, microvessel density,and p53 expression in esophageal carcinomas

Vascular endothelial growth factor (VEGF) is known to play a key role in tumor angiogenesis. The tumor-suppressor gene p53 has been thought to regulate VEGF. We investigated the effect of VEGF on esophageal carcinoma and the correlation between VEGF and p53. Tissue samples were taken from 81 patients with esophageal carcinoma after surgery. VEGF and p53 expressions were examined by immunohistochemical staining. Microvessels in the tumor stained for CD34 antigen were also counted. VEGF and p53 expressions were observed in 51.3% (41/80) and 51.9% (41/79), respectively. The microvessel density was 70.9+/-6.7 (mean+/-SE) in VEGF-positive group and 68.7+/-5.1 in VEGF-negative group. However, no correlation was noted between VEGF and p53 expression. Whereas the tumor size, nodal status, depth of invasions, and tumor stage were associated with poor overall survival, VEGF expression or p53 expression was not. These results indicate that VEGF and p53 are highly expressed in esophageal carcinomas. Since the VEGF expression is not correlated with the p53 expression, microvessel density or clinicopathological findings, further studies with other angiogenic molecules are needed to determine the role in esophageal carcinomas.     

Immunogenicity and Safety of Two Different Haemophilus influenzae Type b Conjugate Vaccines in Korean Infants

The incidence of invasive diseases, including meningitis caused by Haemophilus influenzae type b (Hib) was markedly decreased after routine immunization of Hib vaccine through diverse schedules in many countries. The purpose of this study was to evaluate the immunogenicity and safety of Hib conjugate vaccines in Korean children before the implementation of a national immunization program against Hib in Korea. A multicenter controlled trial was performed on two different Hib vaccines in Korean children. A total of 319 infants were enrolled: 199 infants were immunized with the Hib polysaccharide conjugated to the tetanus toxoid (PRP-T) and 120 infants with the Hib polysaccharide conjugated to the outer-membrane protein of Neisseria meningitides (PRP-OMP). Immunogenicity was evaluated by enzyme-linked immunosorbent assay (ELISA) and serum bactericidal assay. Both vaccines showed good immunologic responses after primary immunization. After 2 doses of PRP-T or PRP-OMP, 78.9% and 91.7% of infants achieved an antibody level of ≥1.0 µg/mL, respectively. Both vaccines were safe and well-tolerated. No serious adverse events were observed. Thus, Hib conjugate vaccines appear to be safe and show good immunogenicity in Korean infants. These results will be important reference data for the implementation of Hib vaccine in the national immunization program of Korea.     

Effect of fentanyl on TNF-alpha and IL-1beta levels during global ischemia/reperfusion in rats

To reduce surgical stress, fentanyl is frequently used for neurosurgical procedures in which focal and/or global ischemia may occur. However, the effect of fentanyl on cytokine levels during ischemia/reperfusion is still uncertain. The goal of this study was to evaluate the effect of fentanyl infusion on levels of the proinflammatory cytokines, tumor necrosis factor (TNF)-alpha and interleukin (IL)-1beta, during global cerebral ischemia/reperfusion in rats using the intracerebral microdialysis technique. Forty male Sprague-Dawley rats weighing 280-320 g were randomly assigned to each of four groups: group 1 (no fentanyl infusion and only ischemia/reperfusion); group 2 (1.5 ng/ml of fentanyl infusion during ischemia/reperfusion) and group 3 (3 ng/ml of fentanyl infusion during ischemia/reperfusion) (n=5 in each group). The rats were anesthetized with an intraperitoneal injection of pentobarbital (50 mg/kg). They were then intubated and ventilated with room air using an animal ventilator. A CMA-12 probe was inserted into the left hippocampal CA-1 region according to the guidelines. Artificial cerebrospinal fluid was run from the inserted microdialysis probe and infused with or without fentanyl at 3 microl/min using a microinjection syringe pump during ischemia/reperfusion. Ischemia was induced by clamping the carotid arteries. Hemorrhagic hypotension was induced for 17 min via the femoral artery, and reperfusion was accomplished by unclamping the sling and reinfusing the blood via the femoral artery. After 2 h of stabilization, the microdialysate was collected 10 times every 17 min, just before ischemia (control), after ischemia (I) and after reperfusion (R1-R8), and stored at -80 degrees C until analysis using high-performance liquid chromatography During global ischemia/reperfusion, TNF-alpha and IL-1beta significantly increased at reperfusion (R5) compared with the control value (p < 0.05). However, in both cases of fentanyl infusion, TNF-alpha and IL-1beta showed no increase compared with the control value. Fentanyl inhibited an increase of the proinflammatory cytokines, TNF-alpha and IL-1beta levels, during global cerebral ischemia/reperfusion in rats.     

The Hermansky-Pudlak syndrome (HPS) protein is part of a high molecular weight complex involved in biogenesis of early melanosomes

Hermansky-Pudlak syndrome (HPS) is a rare autosomal recessive disorder in which oculocutaneous albinism, bleeding tendency and a ceroid-lipofuscin lysosomal storage disease result from defects of multiple cytoplasmic organelles: melanosomes, platelet dense granules and lysosomes. The HPS polypeptide, a 700 amino acid protein which is unrelated to any known proteins, is likely to be involved in the biogenesis of these different organelles. Here, we show that HPS is a non-glycosylated, non-membrane protein which is a component of two distinct high molecular weight complexes. In non-melanotic cells the HPS protein is contained almost entirely in an approximately 200 kDa complex that is widely distributed throughout the cytosol. In melanotic cells the HPS protein is partitioned between this cytosolic complex and a >500 kDa complex that appears to consist of the approximately 200 kDa complex in association with membranous components. Subcellular fractionation, immunofluorescence and immunoelectron microscopy studies indicate that the membrane-associated HPS complex of melanotic cells is associated with tubulovesicular structures, small non-coated vesicles, and nascent and early-stage melanosomes. These findings suggest that the HPS complex is involved in the biogenesis of early melanosomes.     

Cytokine responses in mice infected with<Emphasis Type="Italic"> Clonorchis sinensis</Emphasis>

FVB and BALB/c mice show different morbidity, development of Clonorchis sinensis, and pathological changes following C. sinensis infection. FVB mice are susceptible and BALB/c mice are relatively more resistant to C. sinensis infection. To investigate the relationship between cytokine reaction and susceptibility to C. sinensis infection in FVB and BALB/c mice, we described both the patterns and kinetics of Th1 cytokines and Th2 cytokines in spleen cell culture. Interleukin (IL)-4 and IL-10 cytokine production in the culture supernatants of the concanavalin-A-stimulated spleen cells increased at 2–3 weeks post-infection in both strains. IL-5 production increased between 2 and 5 weeks post-infection in both strains, and reached a peak level at 2 weeks post-infection in BALB/c mice and 4 weeks post-infection in FVB mice. In contrast, gamma interferon (IFN-) production decreased between 2 and 4 weeks in both strains. IL-2 production increased slightly in BALB/c mice following infection, but was unchanged in FVB mice. IL-4 production over preinfection levels was significantly higher in FVB mice, whereas IFN-, IL-2, and IL-10 production were significantly higher in BALB/c mice. The levels of serum immunoglobulin E (IgE) and blood eosinophils in both mouse strains significantly increased between 3 and 6 weeks postinfection. Serum IgE levels were significantly higher in FVB mice than in BALB/c mice. The results of this study suggest that susceptibility to C. sinensis infection is associated with Th2 cytokine production, especially IL-4 which is predominant in relatively susceptible FVB mice.     

MMA/MPEOMA/VSA copolymer as a novel blood-compatible material: effect of PEO and negatively charged side chains on protein adsorption and platelet adhesion

This study was designed to evaluate the effect of polyethylene oxide (PEO) and negatively charged side chains on blood compatibility. For this, novel copolymers (MMA/MPEOMA/VSA copolymers) with both PEO and negatively chargeable side chains were synthesized by random copolymerization of methyl methacrylate (MMA), methoxy PEO monomethacrylate (MPEOMA; PEO mol wt 1000), and vinyl sulfonic acid sodium salt (VSA) monomers of different compositions. MMA/MPEOMA copolymer (with PEO side chains) and MMA/VSA copolymer (with negatively chargeable side groups) also were synthesized for purposes of comparison. The synthesized copolymers were characterized by 1H-nuclear magnetic resonance spectroscopy and gel permeation chromatography. They were coated onto polyurethane (PU) or polymethyl methacrylate (PMMA) films by spin coating. The surface properties of MMA/MPEOMA/VSA copolymers were compared by water contact angle and zeta potential with those of MMA/MPEOMA and MMA/VSA copolymers of similar MPEOMA or VSA composition. Using electron spectroscopy for chemical analysis and scanning electron microscopy, respectively, the behaviors of the adsorption of blood proteins (albumin, gamma-globulin, fibrinogen, and plasma proteins) and the adhesion of platelets on the copolymer-coated surfaces also were compared. Among the copolymers, the MMA/MPEOMA/VSA copolymer with a monomer molar ratio 8:1:1 was observed to be particularly effective in preventing both protein adsorption and platelet adhesion on the surfaces, probably owing to the combined effects of highly mobile, hydrophilic PEO side chains and negatively charged side groups in aqueous solution.