Purpose: Malignant gliomas are aggressive spinal cord tumors. In this study, we hypothesized that combination therapy using an anti-angiogenic agent, bevacizumab, and hypoxia-inducible glioblastoma-specific suicide gene could reduce tumor growth. Materials and Methods: In the present study, we evaluated the effect of combination therapy using bevacizumab and pEpo-NI2-SV-TK in reducing the proliferation of C6 cells and tumor growth in the spinal cord. Spinal cord tumor was generated by the injection of C6 cells into the T5 level of the spinal cord. Complexes of branched polyethylenimine (bPEI)/pEpo-NI2-SV-TK were injected into the spinal cord tumor. Bevacizumab was then administered by an intraperitoneal injection at a dose of 7 mg/kg. The anti-cancer effects of combination therapy were analyzed by histological analyses and magnetic resonance imaging (MRI). The Basso, Beattie and Bresnahan scale scores for all of the treatment groups were recorded every other day for 15 days to assess the rat hindlimb strength. Results: The complexes of bPEI/pEpo-NI2-SV-TK inhibited the viability of C6 cells in the hypoxia condition at 5 days after treatment with ganciclovir. Bevacizumab was decreased in the cell viability of human umbilical vein endothelial cells. Combination therapy reduced the tumor size by histological analyses and MRI. The combination therapy group showed improved hind-limb function compared to the other groups that were administered pEpo-NI2-SV-TK alone or bevacizumab alone. Conclusion This study suggests that combination therapy using bevacizumab with the pEpo-NI2-SV-TK therapeutic gene could be useful for increasing its therapeutic benefits for intramedullary spinal cord tumors.

BACKGROUND: ABO genotyping is being used increasingly when the results of serologic typing are unclear or there is some suspicion of rare ABO subtypes. Conventional molecular diagnostic methods such as PCR- restriction fragment length polymorphism (PCR-RFLP), allele-specific PCR, PCR-single stranded conformational polymorphism (PCR-SSCP) and sequence-based typing have been used in this field. Recently, a pyrosequencing technique was introduced into clinical laboratories. This study evaluated the possibility of applying pyrosequencing to ABO genotyping. METHODS: A total of 36 samples, which had previously been analyzed by PCR-RFLP and serological method in the Blood Genetics Clinic of Seoul National University Hospital between August 2001 and September 2004 and shown to have the A/A, A/B, A/O, B/B, B/O, O/O, cis-AB/O, cis-AB/A, or cis-AB/B genotypes, were analyzed by pyrosequencing analysis. Briefly, two PCR reactions were carried out separately for one region including nucleotide 261, and for another region including nucleotides 796 and 803. Pyrosequencing was then performed, and the pyrograms were interpreted using an automated interpretation program from the manufacturer and by researchers independently to determine the nucleotides 261, 796 and 803 for ABO genotyping. RESULTS: The ABO genotypes from pyrosequencing and the interpretation of the pyrograms according to the researcher on 36 samples were in complete concordance with the results obtained by PCR-RFLP. The ABO genotypes from the automated interpretation program showed an error in one out of total 108 SNP (single nucleotide polymorphism) analyses (eRROR RATE=0.9%) OF 36 SAMPLES. CONCLUSION: ABO genotyping for A, B, O, cis-AB alleles by pyrosequencing of nucleotides 261, 796 and 803 was relatively simple and accurate and could be an another field we can use in clinical laboratories.
Alleles ; Genetics ; Genotype ; Nucleotides ; Pathology, Molecular ; Polymerase Chain Reaction ; Polymorphism, Restriction Fragment Length ; Seoul

BACKGROUND: The size reduction of low density lipoproteins(LDL) particles has been found associated with coronary artery disease(CAD), but the analysis of LDL subfractions is time consuming(over 24 hours) and technically demanding such as density gradient ultracentrifugation or gradient polyacrylamide gel electrophoresis(PAGE). In this study, the association between LDL subfractions and CAD was investigated using the new method for the separation of LDL subfractions, commercially available continuous disc PAGE. METHODS: LDL subfraction profiles were investigated in two groups(97 patients with CAD proven angiographically and 109 healthy controls) using Lipoprint LDL System(Quantimetrix Co., Redondo Beach, CA, USA). Seven LDL subfractions(from LDL-0 to LDL-6) can be identified with a specific electrophoretic mobility(Rf) relative to the HDL fraction. The relative percentage of the area under the curve(AUC) of each LDL band was multiplied by its band number and the sum of the relative AUC of all LDL bands percent was calculated to produce a final LDL score. RESULTS: Mean LDL score was significantly higher in CAD patients than in controls(2.09+/-0.77 and 1.01+/-0.38, respectively, P<0.0001). There was a significant association between the polydispersity of LDL subfractions and the presence of CAD(P=0.01). LDL score was highly correlated with triglyceride(r=0.330, P=0.0001). LDL score was the significant discriminators between control and CAD groups(F=168.3, P=0.0001). There was no significant association between LDL score and the severity of CAD. CONCLUSIONS: Lipoprint LDL System is an easy and quick method for routine assessment of LDL subfractions. LDL score was the best individual discriminant risk factor for CAD.
Area Under Curve ; Coronary Artery Disease* ; Coronary Vessels* ; Humans ; Lipoproteins* ; Risk Factors ; Triglycerides ; Ultracentrifugation

PURPOSE: Spinal cord injury (SCI) is associated with permanent neurological damage, and treatment thereof with a single modality often does not provide sufficient therapeutic outcomes. Therefore, a strategy that combines two or more techniques might show better therapeutic effects. MATERIALS AND METHODS: In this study, we designed a combined treatment strategy based on neural stem cells (NSCs) introduced via a neuronal cell type-inducible transgene expression system (NSE::) controlled by a neuron-specific enolase (NSE) promoter to maximize therapeutic efficiency and neuronal differentiation. The luciferase gene was chosen to confirm whether this combined system was working properly prior to using a therapeutic gene. The luciferase expression levels of NSCs introduced via the neuronal cell type-inducible luciferase expression system (NSE::Luci) or via a general luciferase expressing system (SV::Luci) were measured and compared in vitro and in vivo. RESULTS: NSCs introduced via the neuronal cell type-inducible luciferase expressing system (NSE::Luci-NSCs) showed a high level of luciferase expression, compared to NSCs introduced via a general luciferase expressing system (SV::Luci-NSCs). Interestingly, the luciferase expression level of NSE::Luci-NSCs increased greatly after differentiation into neurons. CONCLUSION: We demonstrated that a neuronal cell type-inducible gene expression system is suitable for introducing NSCs in combined treatment strategies. We suggest that the proposed strategy may be a promising tool for the treatment of neurodegenerative disorders, including SCI.
Cell Differentiation/genetics/physiology ; *Gene Expression ; Gene Regulatory Networks ; *Genetic Therapy ; Humans ; Luciferases/genetics/*metabolism ; *Neural Stem Cells ; Neurons/metabolism ; Phosphopyruvate Hydratase/metabolism ; Promoter Regions, Genetic ; Spinal Cord Injuries/*therapy ; Stem Cells/*metabolism

BACKGROUND: Rotavirus is the most common cause of childhood diarrhea worldwide. Although rotavirus is also the leading cause of infant and childhood diarrhea in Korea, much remains unknown about the trends of rotavirus infection by month and geographic region in Korea. To monitor epidemiologic trends of rotavirus infection, a laboratory-based rotavirus surveillance network was established in 2002. This is the first nationwide, multicenter evaluation of rotavirus epidemiology in Korea. METHODS: The rotavirus test results were collected retrospectively from eight network laboratories, from July 1999 to June 2002. Four laboratories used latex agglutination, three used immunochromatography, and one used enzyme-linked fluorescent assay for the detection of rotavirus antigen. RESULTS: Of 10, 441 stool specimens, 2, 496 (23.9%) were positive for rotavirus. During the 3-year period, the rotavirus season began in December-January, and ended in April-May. The rotaviruspositive percentage of summer, autumn, winter, and spring was 11.5%, 10.0%, 32.8%, and 30.0%, respectively. A few hospitals revealed summer epidemics. The rotavirus positive rate in each hospital varied from 15.3% to 44.2%. A common feature of the three hospitals showing the lowest rotavirus-positive percentage (i.e. <20%) was their large size (>800 beds). The secondary care hospitals showed a higher positive proportion (27.5%) compared with tertiary care hospitals (21.1%). CONCLUSIONS: Overall, the rotavirus-positive percentage among all diarrheal specimens was similar to that of other developed countries. The results of this study showed that the autumn epidemic of the rotavirus has declined or disappeared and the peak season for rotavirus has shifted to late winter/early spring in Korea.
Agglutination ; Developed Countries ; Diarrhea ; Epidemiology ; Humans ; Immunochromatography ; Infant ; Korea ; Latex ; Republic of Korea* ; Retrospective Studies ; Rotavirus ; Rotavirus Infections* ; Seasons ; Secondary Care ; Tertiary Healthcare

Background: Gene therapy shows the ability to restore neuronal dysfunction via therapeutic gene expression. The efficiency of gene expression and delivery to hypoxic injury sites is important for successful gene therapy. Therefore, we established a gene/stem cell therapy system using neuron-specific enolase promoter and induced neural stem cells in combination with valproic acid to increase therapeutic gene expression in hypoxic spinal cord injury. Methods: To examine the effect of combined method on enhancing gene expression, we compared neuronal cell-inducible luciferase levels under normoxia or hypoxia conditions in induced neural stem cells with valproic acid. Therapeutic gene, vascular endothelial growth factor, expression with combined method was investigated in hypoxic spinal cord injury model. We verified gene expression levels and the effect of different methods of valproic acid administration in vivo. Results: The results showed that neuron-specific enolase promoter enhanced gene expression levels in induced neural stem cells compared to Simian Virus 40 promoter under hypoxic conditions. Valproic acid treatment showed higher gene expression of neuron-specific enolase promoter than without treatment. In addition, gene expression levels and cell viability were different depending on the various concentration of valproic acid. The gene expression levels were increased significantly when valproic acid was directly injected with induced neural stem cells in vivo. Conclusion In this study, we demonstrated that the combination of neuron-specific enolase promoter and valproic acid induced gene overexpression in induced neural stem cells under hypoxic conditions and also in spinal cord injury depending on valproic acid administration in vivo. Combination of valproic acid and neuron-specific enolase promoter in induced neural stem cells could be an effective gene therapy system for hypoxic spinal cord injury.

Background: Gene therapy shows the ability to restore neuronal dysfunction via therapeutic gene expression. The efficiency of gene expression and delivery to hypoxic injury sites is important for successful gene therapy. Therefore, we established a gene/stem cell therapy system using neuron-specific enolase promoter and induced neural stem cells in combination with valproic acid to increase therapeutic gene expression in hypoxic spinal cord injury. Methods: To examine the effect of combined method on enhancing gene expression, we compared neuronal cell-inducible luciferase levels under normoxia or hypoxia conditions in induced neural stem cells with valproic acid. Therapeutic gene, vascular endothelial growth factor, expression with combined method was investigated in hypoxic spinal cord injury model. We verified gene expression levels and the effect of different methods of valproic acid administration in vivo. Results: The results showed that neuron-specific enolase promoter enhanced gene expression levels in induced neural stem cells compared to Simian Virus 40 promoter under hypoxic conditions. Valproic acid treatment showed higher gene expression of neuron-specific enolase promoter than without treatment. In addition, gene expression levels and cell viability were different depending on the various concentration of valproic acid. The gene expression levels were increased significantly when valproic acid was directly injected with induced neural stem cells in vivo. Conclusion In this study, we demonstrated that the combination of neuron-specific enolase promoter and valproic acid induced gene overexpression in induced neural stem cells under hypoxic conditions and also in spinal cord injury depending on valproic acid administration in vivo. Combination of valproic acid and neuron-specific enolase promoter in induced neural stem cells could be an effective gene therapy system for hypoxic spinal cord injury.

Different subtypes of dendritic cells (DC) influence the differentiation of naive T lymphocytes into T helper type 1 (Th1) and Th2 effector cells. We evaluated the percentages of DC subtypes in peripheral blood from pregnant women (maternal blood) and their cord blood compared to the peripheral blood of healthy non pregnant women (control). Circulating DC were identified by flow cytometry as lineage (CD3, CD14, CD16, CD19, CD20, and CD56)-negative and HLA-DR-positive cells. Subtypes of DC were further characterized as myeloid DC (CD11c+/CD123+/-), lymphoid DC (CD11c-/CD123+++) and less differentiated DC (CD11c-/CD123+/-). The frequency of DC out of all nucleated cells was significantly lower in maternal blood than in control (P<0.001). The ratio of myeloid DC/lymphoid DC was significantly higher in maternal blood than in control (P<0.01). HLA-DR expressions of myeloid DC as mean fluorescence intensity (MFI) were significantly less in maternal blood and in cord blood than in control (P<0.001, respectively). The DC differentiation factors, TNF-alpha and GM-CSF, released from mononuclear cells after lipopolysaccharide stimulation were significantly lower in maternal blood than in control (P<0.01). The distribution of DC subtypes was different in maternal and cord blood from those of non-pregnant women. Their role during pregnancy remains to be determined.
Adult ; Cell Differentiation ; Dendritic Cells/*classification/cytology/immunology ; Female ; Fetal Blood/cytology/*immunology ; Flow Cytometry ; Granulocyte-Macrophage Colony-Stimulating Factor/metabolism ; HLA-DR Antigens/metabolism ; Humans ; Lipopolysaccharides/pharmacology ; Lymphocyte Activation ; Pregnancy ; T-Lymphocytes/cytology/immunology ; Th1 Cells/cytology/immunology ; Th2 Cells/cytology/immunology ; Tumor Necrosis Factor-alpha/metabolism

Two trials of external quality assessment for Therapeutic Drug Monitoring (TDM) subcommittee of Korean Association of Quality Assurance for Clinical Pathology (KAQACP) were performed in 2003. Number of participating laboratories were 80 which is similar to those of the previous year. Response rates were elevated to 100.0% for both trials. Two kinds of control materials were requested to be tested in each trial so that each institution could know the possible systematic error. In both trials, 20 test items were responded at least from one laboratory. The average drug item was 7.0 per institution, which was elevated slightly from 6.5 in recent 5 years. The most common test items were valproic acid, digoxin, phenytoin, carbamazepine, theophylline, and phenobarbital, which were peformed in more than 75% of participating laboratories, followed by cyclosporine, lithium, methotrexate, tacrolimus, vancomycin, amikacin, gentamycin, salicylate, tobramycin, acetaminophen, primidone, free phenytoin, and amitryptyline. The most widely used TDM analyzer was Abbott TDx/TDxFLx (52%), but its proportion were decreased slightly comparing with the previous years. The interlaboratory coefficients of variations were not greatly improved comparing with previous years. In conclusion, the TDM external quality assessment of 2003 showed grossly similar pattern comparing with those of previous year, except that the response rate was elevated to 100% and two levels of control material were used in each trials.
Acetaminophen ; Amikacin ; Carbamazepine ; Cyclosporine ; Digoxin ; Drug Monitoring* ; Gentamicins ; Korea* ; Lithium ; Methotrexate ; Pathology, Clinical ; Phenobarbital ; Phenytoin ; Primidone ; Tacrolimus ; Theophylline ; Tobramycin ; Valproic Acid ; Vancomycin

Two trials of external quality assessment for Therapeutic Drug Monitoring (TDM) Subcommittee of Korean Association of Quality Assurance for Clinical Pathology (KAQACP) were performed in 2004. Number of participating laboratories were increased to 89, by 11.3% increase comparing with the previous year. Response rates were 100.0% for both trials just like the previous year. Two kinds of control materials were requested to be tested in each trial so that each institution could know the possible systematic error. In both trials, 20 test items were responded at least from one laboratory. The average drug item was 6.8 per institution, which was elevated slightly from 6.5 in recent 5 years. The most common test items were valproic acid, digoxin, phenytoin, carbamazepine, theophylline, and phenobarbital which were peformed in more than 71% of participating laboratories, followed by cyclosporine, lithium, tacrolimus, vancomycin, methotrexate, amikacin, gentamycin, salicylate, tobramycin, acetaminophen, primidone, free phenytoin, and amitryptyline. The most widely used TDM analyzer was Abbott TDx/TDxFLx (41.6%), followed by Roche Cobas Integra (21.3%), and Abbott AxSym (20.2%). The inter-laboratory coefficients of variations were not greatly improved comparing with previous years. We also determined cyclosporine and tacrolimus with reference method using liquid chromatography-tandem mass spectrometry. In conclusion, the TDM external quality assessment of 2004 showed grossly similar pattern comparing with those of previous year, except the 11.3% increase of participating laboratories.
Acetaminophen ; Amikacin ; Carbamazepine ; Cyclosporine ; Digoxin ; Drug Monitoring* ; Gentamicins ; Korea* ; Lithium ; Mass Spectrometry ; Methotrexate ; Pathology, Clinical ; Phenobarbital ; Phenytoin ; Primidone ; Tacrolimus ; Theophylline ; Tobramycin ; Valproic Acid ; Vancomycin