Child ; Child, Preschool ; Female ; Humans ; Infant ; Infant, Newborn ; Intracranial Hemorrhages ; etiology ; pathology ; Magnetic Resonance Imaging ; Prognosis ; Tomography, X-Ray Computed

In order to provide substantial scientific information for exploring the mechanism of porcine liver injury caused by Taenia asiatica (T.asiatica),the expression of Cysteine aspartyl proteinase 3 (Caspase-3) from liver tissues of porkets that were experimentally infected by T.asiatica was examined.The T.asiatica adults were collected from the taeniasis patients in Duyun,Guizhou Province and identified biologically.The eggs were harvested from gravid proglottids and prepared by repeated washing and centrifugation.Twelve 20-days old Yorkshire and Seghers hybrid porkets were randomly divided into experimental and control groups as six pigs per group.The experimental group was orally administrated with 1.5 × 106 eggs per porket at day 0 post-infection.The porkets of both groups were sacrificed on the day 15 and day 75 post-infection (three pigs per time point) respectively,and liver samples were collected for further experiments.Quantitative real-time polymerase chain reaction method was employed to detect the mRNA levels of Caspase-3,and western blotting and immunohistochemistry methods were performed to detect the level of Caspase-3 expression in both groups.At the day 15 post-infection,the mRNA level and expression level of Caspase-3 of the experimental group were significantly decreased,comparison with the control group (P =0.011,P=0.008 and P=0.004 respectively).It was positive with Caspase-3 when yellow or brown signal appeared in the cytoplasm of liver cells by immunohistochemistry.However,at the day 75 post-infection,the mRNA level and expression level of Caspase-3 of the experimental group were dramatically similar to the control group.Furthermore,in the experimental group,the mRNA level and expression level of Caspase-3 were significantly increased at day 75 post-infection than day 15 post-infection (P--0.018,P=0.003 and P=0.002 respectively).These results suggested that Caspase-3 might be involved into the regulation of the damage of porcine liver induced by T.asiatica challenge at the early infection stage and have on effect to the hepatic injury because of the dramatic recovery of Caspase-3 at the consequent infection stage.

This study was purpose to investigate the frequency of HLA-DRB1*15 expression in children with aplastic anemia (AA) and its relation to effect of immunosuppressive therapy. HLA-DR genotypes were detected by SSP-PCR in 40 patients with acquired aplastic anemia and 107 normal controls, and the expressions of HLA-DR gene in AA patients and normal controls were compared. 32 out of 40 patients were treated with immunosuppressive therapy, which included antilymphocyte globulin combining with cyclosporine or cyclosporine alone, the relation of HLA-DRB1*15 expression to efficacy of immunosuppressive therapy and relapse of AA was explored. The results showed that the mean age of the patients was 9.0 years with a ratio of male to female 1.5:1. The frequency of HLA-DRB1*15 genotype expression in patients with idiopathic aplastic anemia was 51.5% (17/33), which was markedly higher than that of healthy controls (20.6%, p<0.01). All of 7 patients with second acquired aplastic anemia showed negative expression of HLA-DRB1*15. The rates of all responses, including complete remission and partial remission (CR+PR), and CR to immunosuppressive therapy in 16 patients who bared HLA-DRB1*15 were 93.8% and 87.5% respectively, which were higher significantly than those of patients without bearing HLA-DRB1*15 (56.3% and 31.3%, p<0.01). Relapse occurred in 5 patients who bared HLA-DRB1*15 genotype. It is concluded that the frequency of HLA-DRB1*15 genotype expression in children with AA is significantly higher than that in normal controls, and the immunosuppressive therapy for patients bared HLA-DRB1*15 shows favourable effect with high incidence of complete remission.
Adolescent ; Anemia, Aplastic ; drug therapy ; genetics ; immunology ; Antilymphocyte Serum ; therapeutic use ; Child ; Child, Preschool ; Cyclosporine ; therapeutic use ; Female ; Genotype ; HLA-DR Antigens ; metabolism ; HLA-DRB1 Chains ; Humans ; Immunosuppressive Agents ; therapeutic use ; Male

OBJECTIVETo study the role of cyclinD1 and CDK4 in malignant transformation of human fetal lung diploid fibroblast cell line (2BS) induced by silica.

METHODSRecombination vectors with sense and antisense pXJ41-cyclinD1 and pXJ41-CDK4 were constructed, and then transfected into the malignant transformed cells induced by silica, respectively. At the same time, pXJ41-neo was used as the control.

RESULTSDuring the progress of the malignant transformation of 2BS cells induced by silica, cyclinD1 and CDK4 were overexpressed. Antisense RNA suppressed cyclinD1 and CDK4 gene expression in the antisense pXJ41-cyclinD1 and pXJ41-CDK4 transfected cells. Antisense RNA led to cell cycle arrest, resulting in lengthened G1 phase (the percentages of cells in the G1 phase changed from 45.1% to 52.7% and 58.0% for cyclinD1 and CDK4 transfected cells, respectively), and eventually attenuated the increase of the proliferation of malignant transformed cells induced by silica. Compared with malignant transformed cells induced by silica, cells transfected with antisense pXJ41-cyclinD1 and pXJ41-CDK4 showed obviously reduced growth rates. On the 8th day, the suppression rates were 58.69 and 77.43% (the growth rate of malignant transformed cells induced by silica was 100%), doubling time changed from 21.0 h to 31.4 h and 21.0 h to 42.7 h, respectively, the growth capacities on soft agar of cells transfected by antisense pXJ41-cyclinD1 and pXJ41-CDK4 decreased obviously.

CONCLUSIONCyclinD1 and CDK4 play an important role in maintaining transformed phenotype of the cancer cells.

Carcinogens, Environmental ; toxicity ; Cell Line ; Cell Proliferation ; Cell Transformation, Neoplastic ; chemically induced ; Cyclin D1 ; genetics ; metabolism ; physiology ; Cyclin-Dependent Kinase 4 ; genetics ; metabolism ; physiology ; Humans ; Plasmids ; RNA, Antisense ; metabolism ; RNA, Messenger ; analysis ; metabolism ; Silicon Dioxide ; toxicity

OBJECTIVETo analyze the non-invasive indexes for predicting esophageal varices (EV) in liver cirrhosis, and to establish a model for predicting the degree of EV.

METHODSA total of 294 patients with liver cirrhosis and portal hypertension were divided into the following groups according to EV grade as assessed by endoscopy: non-EV and grade I EV, grade II EV and grade III EV. The non-invasive EV predictive measures of liver stiffness (LS), platelet (PLT) count, spleen thickness (ST), PLT/ST ratio, portal vein diameter, portal vein flow velocity and Child-Pugh score (CPS) were assessed by univariate analysis and multivariate logistic regression analysis, and used to generate a predictive model. The t-test, chi-square test, logistic analysis and receiver operating characteristic (ROC) curve were used in statistical analyses.

RESULTSThe area under the ROC for the new model was 0.990. The best cutoff value for the score was 0.898, as defined from the ROC. The sensitivity of the model was 96.5%, and the specificity was 99.2%.

CONCLUSIONSThe model for predicting EV was composed of LS, PLT count, ST, PLT/ST and CPS, which was accurate and sensitive, and could be used to predict EV in clinic.

Endoscopy, Digestive System ; Esophageal and Gastric Varices ; Humans ; Hypertension, Portal ; Liver Cirrhosis ; Platelet Count ; ROC Curve ; Spleen

OBJECTIVETo study the role of cyclin D1 in malignant transformation of human embryonic lung diploid fibroblasts (HELF) induced by quartz.

METHODSpXJ41-cyclin D1 expressing sense and antisense cyclin D1 RNA were transinfected into malignant transformed HELF induced by quartz with DNA recombination and gene transduction. The expression of cyclin D1 was detected with hybridization in situ and immunohistochemistry methods to analyze changes in cell growth, double multiplication time, distribution of cell cycles, colony forming ability on soft agar, etc., before and after cyclin D1 transduction.

RESULTSDuring the process of malignant transformation of HELF induced by quartz, cyclin D1 gene was overexpressed. Antisense pXJ41-cyclin D1 RNA could suppress the growth and proliferation of malignant transformed cells induced by quartz. Growth speed of antisense pXJ41-cyclin D1 transinfected cells decreased by 58.69% on the 8th day in culture, as compared to malignant transformed cells induced by quartz, and its double multiplication time prolonged from 21.0 h to 31.4 h. Antisense cyclin D1 RNA led to cell cycle arrest, resulting in lengthened G1 phase (proportion of cells in phase G1 increased to 52.7% from 45.1% and that of cells in phase S decreased to 33.1% from 40.3%). Colony forming rate reduced significantly and size of colony became smaller.

CONCLUSIONSAbnormal expression of cyclin D1 in cells related to their malignant transformation induced by quartz. Highly expressed cyclin D1 could play an important role in maintaining the transformed phenotype of malignant cells.

Cell Transformation, Neoplastic ; Cells, Cultured ; Cyclin D1 ; biosynthesis ; genetics ; Embryo, Mammalian ; Fibroblasts ; cytology ; Humans ; Lung ; cytology ; Quartz ; toxicity

OBJECTIVETo determine distribution and mutation pattern of type P ATP7B gene mutation and to explore genotype and phenotype correlation in patients with Wilson's disease (WD).

METHODSSixty patients with WD from 57 no kinship families, 37 male and 23 female, were enrolled in this study. The age of onset ranged from 4.6 - 39 years, < or = 16 years in 55 patients. Some exons of ATP7B gene mutation were analyzed in patients with WD by using biochemical methods, polymerase chain reaction-single strand configuration polymorphism (PCR-SSCP), restriction fragment and DNA sequence analysis. Totally 778 coding regions were identified with restriction enzyme Msp I. The activity of Cu-ATPase was assessed by measuring inorganic phosphorus in 3 patients with known genotype.

RESULTSFifty-two of 60 patients (86%) had presented with hepatic manifestations, 30 of them had only hepatic manifestations, 12/52 patients had hepatic and neurological manifestations at the same time; 10/52 patients had hepatic and other symptom; 7/60 patients had only neurological symptom, one patient had no symptom. Eleven mutations were detected by DNA sequencing, including five missense mutations (R778L, V1140A,G943S, V1106I and V1216M), one deletion (1384del17) and five polymorphisms (IVS4-5T/C, A2495G, C2310G, IVS18 + 6C/T and IVS20 + 5A/G) were identified. R778L mutation was identified 52/114 alleles (45.6%). R778L occurred in 38/52 patients with hepatic manifestation (73%), homozygosis of R778L was demonstrated in 14 patients and heterozygosity of R778L in 24 patients. V1106I mutation was 1.7%, G943S, V1140A, and V1216M was 0.86% respectively in this study. Two patients with delayed onset of neurological symptoms occurred V1106I mutation of ATP7B. Cu-ATPase activity of 3 patients with known mutation (R778L/V1106I, R778L/V1216M and R778L/R778L) declined by 44.55%, 88.23% and 69.49%, respectively, compared with normal control.

CONCLUSIONThe 1384del17bp and V1106I are two novel mutations found in patients with WD. R778L was common mutation of ATP7B gene with frequency of 45.6% in this study. The mutation in exon 8 of WD gene may play an important role in pathogenesis of WD in Chinese. Carriage of R778L mutation seems to be correlated with hepatic manifestation.

Adenosine Triphosphatases ; genetics ; Adolescent ; Adult ; Cation Transport Proteins ; genetics ; Child ; Child, Preschool ; Copper-transporting ATPases ; DNA Mutational Analysis ; Exons ; Female ; Gene Frequency ; Genotype ; Hepatolenticular Degeneration ; enzymology ; genetics ; pathology ; Humans ; Male ; Mutation ; Phenotype ; Polymerase Chain Reaction ; Polymorphism, Single-Stranded Conformational ; Sequence Analysis, DNA

OBJECTIVETo explore whether the co-transplantation of mesenchymal stem cells (MSC) from human umbilical cord blood (UCB) with UCB-derived CD34(+) cells in NOD/SCID mice could promote engraftment and accelerate hematopoiesis recovery.

METHODSAfter sublethal irradiated ((60)Co 2.5 Gy), NOD/SCID mice received within 24 hours UCB CD34(+) cells (1 x 10(5) per mouse for low dosage group or 1 x 10(6) per mouse for high dosage group) with or without human UCB-derived MSC (1 x 10(6) per mouse) transplantation by lateral tail vein injection. Peripheral blood cells of transplanted mice were measured for white blood cell count, hemoglobin and platelet count at 10th, 20th, 30th, 40th and 56th day. At the end of 8th week after transplantation, all the alive mice were sacrificed and human derived CD45(+), CD45(+)CD3(+), CD45(+)CD19(+), CD45(+)CD33(+) cells in the bone marrow (BM) were assayed by flow cytometry.

RESULTS(1) In the low dosage group, co-transplantation of MSC significantly raised the engraftment rate (26.02% vs 16.52%) (P < 0.05). (2) The survival rate in high dosage group was 80% for co-transplantation mice and 70% for CD34(+) cells alone transplantation mice. The survival rate in low dosage group was 70% for co-transplantation mice and 50% in CD34(+) cells transplantation mice. (3) In both dosages groups co-transplantation accelerated the hematopoiesis recovery. (4) At the end of 8 weeks after transplantation, in low dosage group, CD45(+)CD33(+) and CD45(+)CD19(+) cells were more in co-transplantation mice than in CD34(+) cells alone transplantation mice, but in high dosage group, the percentage of these two kinds of cells had no difference. In both dosage groups the percentage of CD45(+)CD41a(+) cells were higher in co-transplantation than in transplantation alone mice. CD45(+)CD3(+) cells were low in all groups.

CONCLUSIONS(1) In low dosage transplantation, human UCB MSC could promote human CD34(+) cells engraftment in transplanted mice. (2) Co-transplantation of human UCB MSC and human UCB CD34(+) cells could significantly promote the hematopoiesis reconstitution and improve the survival rate of NOD/SCID mice. (3) MSC could promote human UCB CD34(+) cells to differentiated into B-lymphocytes, granulocyte and megakaryocyte in vivo.

Animals ; Antigens, CD34 ; Cord Blood Stem Cell Transplantation ; Female ; Fetal Blood ; cytology ; Hematopoiesis ; Humans ; Mesenchymal Stem Cell Transplantation ; Mice ; Mice, Inbred NOD ; Mice, SCID ; Transplantation, Heterologous

OBJECTIVEChildhood absence epilepsy (CAE), a common form of idiopathic generalized epilepsy, accounts for 8% - 15% of all childhood epilepsies. A positive family history of epilepsy, a hereditary factor being one of the pathogeneses, is found in 15% - 44% of children with absence seizures. The phenotype of CAE is specific (including seizure forms and EEG), therefore it is suitable for genetic study. The purpose of this study was to confirm the linkage of childhood absence epilepsy to chromosome 8q24 in China.

METHODSTwenty-nine trios families (a patient and his/her parents) as patient group and 10 normal trios families as control group were investigated for chromosome 8q24 by haplotype analysis with 5 microsatellite DNA markers (D8S554, D8S534, D8S1100, D8S1783, D8S1753). Genomic DNA was isolated from 4 ml human peripheral blood by using the conventional procedure, and then was treated using the PCR method. PCR products were analyzed by gene scan. Statistical methodology included haplotype-based haplotype relative risk (HHRR) and transmission disequilibrium test (TDT).

RESULTSIn this study, the polymorphism information content (PIC) of 5 microsatellite DNA markers were: 0.519, 0.828, 0.528, 0.654 and 0.772. HHRR showed D8S554(4) (chi(2) = 5.939, P < 0.05), D8S1100(3) (chi(2) = 5.081, P < 0.05), D8S1783(6) (chi(2) = 4.308, P < 0.05). TDT showed D8S554(4) (chi(2) = 4.46, P < 0.05), D8S1783(6) (chi(2) = 4, P < 0.05). In order to exclude false association results, the authors analyzed every family in detail. Four trios families transmitted allele D8S1783(6) to their offspring, and the same allele hasn't been found in controls. The further work showed that locus D8S1783 had transmission disequilibrium with CAE, the other two loci were a false association.

CONCLUSION(1) Childhood absence epilepsy in the Chinese population may be linked to chromosome 8q24, the CAE gene is transmitted disequilibrium on locus D8S1783. Combined with other research results, we suppose that CAE gene may be in the ECA1 area on chromosome 8q24. (2) The CAE gene perhaps has a genetic heterogeneity in the population of different areas and different races. (3) HHRR and TDT seem to be the best statistical methods to do linkage disequilibrium study in the trios family.

Child ; Child, Preschool ; China ; Chromosomes, Human, Pair 8 ; genetics ; Epilepsy, Absence ; genetics ; Family Health ; Female ; Haplotypes ; genetics ; Humans ; Linkage Disequilibrium ; genetics ; Male ; Microsatellite Repeats ; Nuclear Family ; Risk Factors

OBJECTIVETo study the role of cycline dependent kinase 4 (CDK4) in the malignant transformation of human fetal lung diploid fibroblast cell (2BS) induced by silica.

METHODSRecombination vectors with antisense pXJ41-CDK4 were constructed, and then were transfected into the malignant transformed cells induced by silica. In situ hybridization and immunohistochemistry were used to analyze the expression of CDK4. Cell growth curve, doubling time, cell cycle distribution and the growth capacities on soft agar were analyzed before and after antisense CDK4 RNA was transferred into malignant transformed cells induced by silica.

RESULTSDuring the malignant transformation of 2BS cells induced by silica, CDK4 gene was overexpressed. Antisense pXJ41-CDK4 transduction suppressed CDK4 gene expression in the antisense pXJ41-CDK4 transfected cells. Antisense CDK4 RNA led to cell cycle arrest, resulting in lengthened G1 phase (the percentages of cells in the G1 phase increased from 45.1% to 58.0%), and eventually attenuated the proliferation of malignant transformed cells induced by silica. At the 8th day, the suppression rates decreased by 77.43%. The doubling time prolonged from 21.0 h to 42.7 h. The growth capacities on soft agar of cells transfected by antisense pXJ41-CDK4 were decreased.

CONCLUSIONCDK4 might play an important role in maintaining the transformed phenotype of the cancer cells.

Cell Transformation, Neoplastic ; drug effects ; Cells, Cultured ; Cyclin-Dependent Kinase 4 ; Cyclin-Dependent Kinases ; genetics ; physiology ; Embryo, Mammalian ; Fibroblasts ; cytology ; drug effects ; Humans ; Lung ; cytology ; Proto-Oncogene Proteins ; genetics ; physiology ; RNA, Antisense ; pharmacology ; RNA, Messenger ; genetics ; Silicon Dioxide ; toxicity