Three-dimensional syndrome differentiation is a new mode of syndrome differentiation of exopathic febrile diseases, which is based upon the six meridian syndrome differentiation initiated in Treatise on Cold Attack and defence-qi-nutrient-blood and triple-Jiao syndrome differentiation in the art of warm disease of traditional Chinese medicine. Syndromes appeared in the course of cold pathogenic diseases and warm diseases were differentiated through the survey of clinical cases and observation on syndrome features of exopathic febrile diseases, by applying means of analyzing, reasoning, concluding and deducting, from the stadium, location and nature of the disease. Three-dimensional syndrome differentiation theory explains and generalizes pathological mechanism and types of syndrome on acute exopathic febrile diseases roundly, has constructed theoretical frame and synthetical model of syndrome differentiation of exopathic febrile diseases. However, any new method of syndrome differentiation can not be leave nothing to be desired in the beginning, so it is proposed that we must develop epidemiologic investigation on syndromes of exopathic febrile diseases sequentially, make models of clinical commonly seen syndromes of exopathic febrile diseases and dispose fuzzy phenomenon of syndrome by applying mathematical method. By above measures, three-dimensional syndrome differentiation will be perfected gradually, therefore, it can provide scientific theoretical basis for the treatment of exopathic febrile diseases.

Objective: To study the pharmacokinetics and brain/plasma concentration ratio of nortriptyline at multiple doses in mice which were pre-treated with physiological saline, piperine and verapamil. Methods: A total of 216 male Kun Ming mice[(25±3) g] were equally divided into 4 groups randomly. Each group was intragastrically administered physiological saline (B), piperine (170 μg/kg), piperine (5 mg/kg) and verapamil (5 mg/kg) for 8 days. On the 8th day, 1 h atfer giving the above drugs, each mice was intraperitoneally injected nortriptyline (13 mg/kg). The mice were sacriifced by picking off eyeballs at the time intervals of 5, 15, 30 min, and 1, 2, 4, 6, 8 and 12 h, andthe cerebra were collected and weighted. Nortriptyline in mouse plasma and brain was determined by HPLC-MS/MS. The pharmacokinetic properties of the plasma, brain and brain/plasma were calculated. Results: hTe AUC0-12 h of brain/plasma concentration ratio in the 170 μg/kg piperine group was significantly lower than that in the other groups (P<0.05), while the AUC0-12 h of brain/plasma concentration ratios in the 5 mg/kg piperine group and the verapamil group were not signiifcantly different from those of untreated mice. Conclusion: Piperine (170 μg/kg) may induce P-glycoprotein expression in the blood-brain barrier, while piperines at 5 mg/kg has no influence on P-glycoprotein expression in the blood-brain barrier.

Objective:To investigate the clinical efficacy of fluorescence in situ hybridization(FISH)in prenatal diagnosis of chromosomal aneuploidy.Methods:FISH and karyotyping analysis were done in 120 samples of amniotic fluid from pregnant women.Results:FISH analysis results were consistent with those of karyotyping,the dectecting rate can reach 100%for chromosomal aneuploidies.There was no significant difference in the preference for FISH or karyotying for prenatal diagnosis when facing high matemal age,multi-indications and other factors(P>0.05).FISH was better when biochemical datafor down's syndrome were positive (P=0.029),however karyotyping was better when there was abnormal fetal ultrasound scan(P=0.000).Conclusions:FISH can detect the chromosomal aneuploidy quickly and accurately,especially when biochemical data for down's syndrome were positive.FISH could be the first choice for prenatal diagnosis in the third trimester of high-risk pregnancy.

Objective Using an in vivo adeno-associated virus(AAV)-mediated gene transfer technique,this study was designed to evaluate the protective effects of human osteoprotegerin(OPG)transgene against joint destruction in collagen induced arthritis(CIA)model.MethodsAfter CIA was established in the Sprague-Dawley rats,the experimental animals were treated with PBS or rAAV-EGFP or rAAV-hOPG (100μl/d)intra-articular injection 25 days after arthritis induction for 10 days.Paraffin-embedded joints were then analyzed histologically.The joint destruction was evaluated by Larsen Score.The protein expression of OPG,IL-1,MMP-3 was identified by enzyme-linked immunosorbent assay(ELISA).Results Suecessful trans-gene expression was confirmed by the detection of OPG by ELISA and positive fluorescence of the frozen joint section. Image analysis revealed that the expression of OPG significantly protected against joint destruction by 30% compared with the CIA group. Conclusion OPG gene transfer mediated by rAAV effectively protects against bone destruction induced by CIA model. Those data suggest that gene transferring using rAAV-OPG may be a feasible and effective therapeutic approach to treat or prevent joint destruction in inflammatory arthritis.

Objective This study was designed to investigate the expression changes of osteopro-tegerin (OPG), tartrate-resistant acid phosphatase (TRAP) and vascular endothelial growth factor (VEGF) mRNA in collagen induced arthritis(CIA) rats. Methods After CIA was induced in Sprague-Dawley rats, the experimental animals were treated with PBS or rAAV-EGFP or rAAV-hOPG (100 μl/day) intra-articular injection for 10 days. Messenger RNAs (mRNAs) were obtained from CIA synovium 40days after first immun-ization. Reverse transcriptase-polymerase chain reactions (RT-PCR) were carried out to detect the mRNA encoding OPG, TRAP, VEGF and β-actin, which acted as inner control. The genes detected clearly by RT-PCR were quantified using real-time PCR. Results The expression of all genes was confirmed by specific single bands in RT-PCR. Real-time PCR showed that the expression levels of TRAP and VEGF were increased, whereas those of OPG mRNA were decreased in CIA group compared with normal controls. The intra-articular gene transduction markedly increased the gene copies of OPG by 128.21% (P<0.01). The expression change of OPG in synovium also caused the decrease of the expression levels of TRAP and VEGF by 58.79% (P<0.01)and 17.85% (P>0.05) respectively, however, the expression change of VEGF was not statistically significant. Conclusion OPG gene mediated by rAAV can be successfully tranfered to knee joint synovium in vivo. The results of this study suggest that gene transfer using rAAV-OPG may be a feasible and effective therapeutic approach to treat or prevent joint destruction in inflammatory arthritis.

OBJECTIVETo study the expression of bcr/abl hybridized gene in chronic myeloid leukemia (CML), acute lymphatic leukemia (ALL) and polycythemia vera (PV), and its clinical significance.

METHODSThe bcr/abl hybridized gene of interphase metaphase cells of bone marrow in 67 such patients were investigated with a probe of dual color-dual fusion translocation fluorescence in situ hybridization (D-FISH).

RESULTSIn 38 CML patients, 34 (89.5%) were positive, with one having a typical t (9; 22) at first, which changed into negative after having been treated with interferon for 38 months. In another patient, 60 days after post-allogeneic peripheral blood stem cell transplantation (PBSCT), the cytomorphology and cytogenetics were in completely remission. But 3% cells were bcr/abl positive as detected by D-FISH. Six (25%) of 24 ALL patients were positive for Bcr/abl fusion gene, which was negative in 2 PV patients. Three patients suspected of having CML were also negative and one of these three was finally diagnosed as suffering from primary thrombocythemia and one, acute myeloid leukemia (M(2a)) as detected by ETO/AML(1) gene, though the other one was still not confirmed. Two (67%) of the 3 bcr/abl negative CML patients and 5 (87%) of the 6 bcr/abl positive ALL patients had refractory leukemia.

CONCLUSIONbcr/abl hybridized gene is accurately detected by a probe of dual color-dual fusion translocation fluorescence in situ hybridization, which can serve as an effective index for clinical diagnosis, estimation of prognosis and monitor of minimal residual disease in some hematopathies.

Adolescent ; Adult ; Aged ; Child ; Child, Preschool ; Female ; Fusion Proteins, bcr-abl ; genetics ; Humans ; In Situ Hybridization, Fluorescence ; methods ; K562 Cells ; Leukemia, Myelogenous, Chronic, BCR-ABL Positive ; drug therapy ; genetics ; physiopathology ; Male ; Middle Aged ; Polycythemia Vera ; genetics ; physiopathology ; Precursor Cell Lymphoblastic Leukemia-Lymphoma ; genetics ; physiopathology

OBJECTIVETo optimize pre-coated multiple-probe fluorescence in situ hybridization (FISH) to improve its efficiency in cytogenetic diagnosis of acute leukemia.

METHODSThe original multiple-probe FISH techniques were optimized by adjusting the cell density and adding a process of protease digestion. Cytogenetic anomalies were detected in 141 patients with acute lymphocytic leukemia (ALL) or acute myeloid leukemia/ myelodysplastic syndromes (AML/MDS) using the modified technique, and 35 of the patients were also examined using the original technique. The successful detection rate and positive site detection rate were compared between the modified and original techniques.

RESULTSModification of the pre-coated multiple-probe FISH technique resulted in an significant increase of the successful detection rate (from 85.3% to 100%) and the positive site detection rate (from 5.1% to 8.6%) in ALL patients; in AML/MDS patients, the successful detection rate was significantly improved from 67.4% to 99.8% and the positive site detection rate from 3.5% to 6.0% (P<0.01).

CONCLUSIONThe modified pre-coated multiple-probe FISH technique can significantly increase the diagnostic efficiency of cytogenetic abnormalities in leukemic patients.

Chromosome Aberrations ; Humans ; In Situ Hybridization, Fluorescence ; methods ; Karyotyping ; Leukemia, Myeloid, Acute ; diagnosis ; genetics ; Myelodysplastic Syndromes ; diagnosis ; genetics

OBJECTIVETo explore the association between chimerism, minimal residual disease (MRD) and relapse after sex-mismatched allogeneic hematopoietic stem cell transplantation (allo-HSCT) for leukemia.

METHODSFifty-seven patients with leukemia received allogeneic hematopoietic stem cell grafts from HLA-matched or partially matched, but sex-mismatched donors. Chimeric status and MRD were detected by dual-color interphase fluorescence in situ hybridization (I-FISH) using X/Y sex chromosome centromere DNA probe and bcr/abl dual fusion DNA probe, respectively, at different time points after transplantation. SPSS software was used to analyse the correlation between chimeric status, MRD and relapse.

RESULTSIn comparison with karyotype analysis, I-FISH was of higher sensitivity in detecting sex chromosome and bcr/abl fusion gene. Chimeric status was negatively correlated with MRD (r=-0.9690, P<0.01). In the early times of transplantation (within 3 months), mixed chimerism had higher relapse rate than did complete chimerism. Chimeric status and MRD were correlated with leukemic relapse (r=-8240, P<0.01; r=-0.9040, P<0.01). The decrease in chimeric status occurred before leukemic relapse in hematology.

CONCLUSIONI-FISH is a more specific and sensitive test for monitoring MRD after transplantation. The clinical value of sex chromosome is identical to that of the special tumor gene for monitoring MRD after transplantation. Chimeric status is negatively correlated with MRD. Chimeric status and MRD are associated with leukemic relapse. The decrease in chimeric status is considered a mark of leukemic relapse after transplantation.

Adolescent ; Adult ; Child ; Chromosomes, Human, X ; genetics ; Chromosomes, Human, Y ; genetics ; DNA Probes ; genetics ; Female ; Fusion Proteins, bcr-abl ; genetics ; Hematopoietic Stem Cell Transplantation ; methods ; Humans ; In Situ Hybridization, Fluorescence ; methods ; Leukemia ; genetics ; surgery ; Male ; Middle Aged ; Transplantation Chimera ; genetics ; Transplantation, Homologous ; Young Adult

OBJECTIVETo detect copy number changes of α-globin gene, and analyze molecular mechanism of the impacts of fetal hemoglobin (HbF) levels for α-globin gene copy numbers loss or increase.

METHODSA total of 15 cases with combined increased levels of fetal hemoglobin with β-thalassemia were collected. Firstly, three common α-thalassemia deletions were validated by Gap-PCR. Secondly, the largest deletions of the β-globin gene cluster were detected by multiplex ligation-dependent probe amplification (MLPA).

RESULTSAmong the 15 cases, there was 1 case with duplication of the α-globin gene cluster, 3 cases of SEA heterozygote deletion of the α-globin gene, 1 cases of α 3.7 deletion heterozygote of the α-globin gene, 1 case of alpha 4.2 deletion homozygote of the α-globin gene, 1 case of deletion homozygote in the like α-globin gene. A compound heterozygous for SEA and α 3.7 of the α-globin gene was also detected. However, 7 cases showed no copy numbers loss and increase of the the α-globin gene cluster.

CONCLUSIONAdditional α-globin gene can produce excessive α-chain, which can aggravate imbalance for α and β-chain, and cause clinical symptoms in patients with β-thalassemia. Yet, copy number loss or mutation in α-globin gene will cause a milder clinical phenotype.

Adult ; Asian Continental Ancestry Group ; genetics ; China ; DNA Copy Number Variations ; Female ; Fetal Hemoglobin ; metabolism ; Humans ; Infant ; Male ; Mutation ; Pedigree ; alpha-Globins ; genetics ; beta-Thalassemia ; genetics ; metabolism