Adolescent ; Adult ; Blood Pressure ; Gravity Suits ; Heart Function Tests ; Humans ; Male ; Positive-Pressure Respiration ; Young Adult

Child, Preschool ; Chromosomes, Human, Pair 22 ; Chromosomes, Human, Pair 8 ; Chromosomes, Human, Pair 9 ; Humans ; In Situ Hybridization, Fluorescence ; Karyotyping ; Male ; Precursor Cell Lymphoblastic Leukemia-Lymphoma ; genetics ; Translocation, Genetic

Objective To evaluate the feasibility of monitoring the gene expression of VEGF165 via the diglycylcysteine (GGC) reporter gene system by reporter probe of 99Tcm-GH. Methods DNA fragments encoding GGC binding motifs were prepared by PCR and positioned at the C end of VEGF165 gene after the linearization of pcDNA3-VEGF165 plasmid. A replication-defective adenovirus vector Ad5-VEGF165GGC motif-internal ribosomal entry site(IRES) -enhanced green fluorescent protein (EGFP) (Ad5-VIE)was constructed, with a cytomegalovirus (CMV) early promoter driving the expression of VEGF165 gene,GGC motif and EGFP, under the aid of an IRFS. A replication-defective adenovirus carrying the Ad5-EGFP was used as the control. Mensenchymal stem cells (MSC) were infected with the recombinant adenovirus at a multiplicity of infection (MOI) from 0 to 100 infectious units (0,10,25,50,100). The cellular uptake of 99Tcm-GH in infected MSC were then studied at 30, 60, 90 and 120 min. VEGF165 was detected by quantitative reverse transcriptase real-time PCR (RT-PCR), Western-blot, and immunohistochemisty. EGFP was observed by RT-PCR and fluorescence microscopy. The correlation analysis was studied between the cellular uptake of 99Tcm-GH and the expression of VEGF165. SPSS 13.0 was applied for statistical analysis. Independent samples t-test, q-test and Pearson correlation coefficient were used. Results After infected with different viral titer of Ad-VIE, the cellular uptake of 99Tcm-GH increased with the increasing virus titer(r2 =0.86, P ＜0.05), with the peak rate (7.94 ±0.75) % at MOI = 100. In time-dependent uptake study, the cellular uptake rates increased rapidly with the time extension, and the highest uptake occurred at 120 min with the peak uptake rate (7.72 ±0.22)%. The uptake rates of 99Tcm-GH in Ad5-VIE-infected cells were significantly higher than those of Ad5 -EGFP-transfected cells at all time points (t = 15.10- 54.92, all P ＜0.05). The VEGF165 and EGFP mRNA levels increased with increasing virus titer, and the VEGF165 mRNA correlated well with the EGFP mRNA(r2 = 0. 99, P ＜ 0.05). After infected with different MOI of Ad5-VIE, good relationship was found between the cellular uptake of 99Tcm-GH and the expression of VEGF165protein in MSC(r2 =0.90, P ＜0.05). Inmunohistochemisty showed VEGF165 protein expressed obviously at Ad5-VIE-infected MSC, and the EGFP was observed by fluorescence microscopy. Conclusions The cellular uptake of 99Tcm-GH in Ad5-VIE-infected MSC are well correlated with the expression of VEGF165 in vitro. The expression of therapeutic gene VEGF165 can be monitored by the GGC peptide expression.

OBJECTIVETo analyze the genetic relations of Shigella isolated from Shenzhen in 2001-2006 and develop primary molecular subtyping surveillance network of Shigella.

METHODSChromosomal DNAs from 55 isolated in agarose were digested with the restriction enzyme Xba I, and then were analyzed by pulsed-field gel electrophoresis. Pulsed-field gel electrophoresis (PFGE) patterns were clustered using BioNumerics software.

RESULTSAll 41 distinctive PFGE patterns were identified among 55 strains. 32 strains belonged to one cluster. Differences were observed in other strains.

CONCLUSIONBoth genetic-related clones and non-related clones of Shigella existed in Shenzhen. The development of PFGE molecular subtyping surveillance network would contribute to the active surveillance, outbreak investigation and source tracking for Shigellosis.

Bacterial Typing Techniques ; China ; Electrophoresis, Gel, Pulsed-Field ; methods ; Feces ; microbiology ; Humans ; Shigella ; classification ; isolation & purification

OBJECTIVETo characterize clinicopathological features of allergic fungal sinusitis (AFS).

METHODSThirty-six cases of AFS were retrieved from the department archival files of Beijing Tongren Hospital from 2002 to 2006. AB-PAS, GMS and MUC5B stain were performed using paraffin-embedded tissues of the cases. Ten cases with available fresh diagnostic tissue were investigated by electron microscopy.

RESULTSPatients included 21 males and 15 females. The age of patients ranged from 11 to 53 years. Atopy was very common in these patients. On plain CT scans, the affected nasal sinuses were filled with soft tissue shadow with patchy hyperdensity. The bony sinus wall showed areas of pressure erosion. Skin antigen tests showed fungal positivity in 31 of 36 cases. Serum levels of the total IgE and/or the specific fungal IgE were elevated in 20 cases. The eosinophil quantity was elevated in 23 cases. Fungal culture was positive in 10 cases. Gross examination showed thick putty secretions within the lesions. Light microscopy showed typical "eosinophilic mucin". Fungal elements were seen with AB-PAS, GMS and MUC5B stains. Electron microscopy demonstrated degranulation by the eosinophils.

CONCLUSIONS"Eosinophilic mucin" is the typical histopathological feature of AFS. AB-PAS, GMS and MUC5B staining methods can used to detect fungal species in mucin. Accurate diagnosis of AFS requires correlations among clinical findings, radiologic examinations, laboratory tests and histopathologic features. However, the ultimate diagnosis requires a histopathologic confirmation.

Adolescent ; Adult ; Child ; Eosinophils ; microbiology ; ultrastructure ; Female ; Fungi ; isolation & purification ; ultrastructure ; Humans ; Hypersensitivity ; blood ; immunology ; pathology ; Immunoglobulin E ; blood ; Leukocyte Count ; Male ; Middle Aged ; Paranasal Sinuses ; diagnostic imaging ; microbiology ; pathology ; Radiography ; Sinusitis ; blood ; immunology ; microbiology ; pathology ; Young Adult

OBJECTIVETo report a rare variant of t(15;17), ins(17;15)(q21;q14q22) in an acute promyelocytic leukemia (APL) patient and the results of cytogenetic and molecular genetic studies.

METHODSChromosomes were prepared after 24 hours culture of bone marrow cells and peripheral blood cells. R-banding technique was used to analyze karyotypes. Chromosome painting analysis was performed using whole chromosome paints for chromosomes 15 and 17. PML-RAR alpha and RAR alpha-PML fusion transcripts were detected by reverse transcription-polymerase chain reaction (RT-PCR).

RESULTSKaryotypic analysis using both specimens from bone marrow and peripheral blood leukemic cells revealed 15q- and 17q+. Chromosome painting analysis confirmed that the karyotypic abnormality was ins(17;15). PML-RAR alpha fusion transcript (S type) was detected by RT-PCR, while RAR alpha-PML fusion transcript was not detected.

CONCLUSIONChromosome painting and RT-PCR are reliable methods for characterization of the insertion involving chromosomes 15 and 17 in APL patients.

Adult ; Chromosome Painting ; methods ; Chromosomes, Human, Pair 15 ; genetics ; Chromosomes, Human, Pair 17 ; genetics ; Humans ; Leukemia, Promyelocytic, Acute ; diagnosis ; genetics ; Male ; Neoplasm Proteins ; genetics ; Oncogene Proteins, Fusion ; genetics ; Reverse Transcriptase Polymerase Chain Reaction ; Transcription, Genetic ; genetics ; Translocation, Genetic

OBJECTIVETo explore the value of dual-color fluorescence in situ hybridization (D-FISH) in the detection of inv(16) in acute myeloid leukemia (AML).

METHODSEleven AML patients were investigated by D-FISH with two-color break apart probe for MYH11 labeled directly by fluorescein isocyanate (FITC) and a Texas Red. The results were associated or compared with those of cell morphology, cytogenetics, single color fluorescence in situ hybridization (FISH) and reverse transcription-polymerase chain reaction (RT-PCR).

RESULTSFour cases (M4Eo three cases, M2a one case) had inv(16), of which one had trisomy 22 in addition to inv(16), while the other seven cases had no inv(16), of which, five cases (M4Eo three cases, M4 two cases)had a normal karyotype, one (M2a) had 5p+ and trisomy 22, one (M4Eo) had a translocation t(9;22) on G-banded karyotypic analysis. All 11 cases of AML were positive for the rearrangement of inv(16) detected by D-FISH. The average positive cell rate for these 11 AML patients was 93.45% (range 86.6%-98.7%). Of them, four had a minimal deletion of 16p13 in addition to inv(16). The results of D-FISH coincided with those of RT-PCR or single color FISH.

CONCLUSIOND-FISH is a powerful tool for the detection of inv(16) due to its sensitivity and specificity. For raising the detecting rate of inv(16), it is necessary to screen inv(16) rearrangement by D-FISH in all M4- and M2-AML cases or the cases with trisomy 22, no matter whether they are accompanied by bone marrow eosinophilia.

Chromosome Inversion ; Chromosomes, Human, Pair 16 ; genetics ; Female ; Humans ; In Situ Hybridization, Fluorescence ; methods ; Leukemia, Myeloid, Acute ; genetics ; Male ; Reverse Transcriptase Polymerase Chain Reaction

OBJECTIVETo report a case of myelodysplastic syndrome(MDS) with t(1;18)(p31;p11).

METHODSChromosome specimens were prepared by short-term culture of bone marrow cells. Karyotype analysis was made by R banding technique. Chromosome painting was performed using whole chromosome probes 1 and 18.

RESULTSConventional karyotype analysis revealed t(1;18)(p31;p11) in this patient. Chromosome painting analysis confirmed this result.

CONCLUSIONThe translocation of (1;18) was an unusual recurrent chromosome change and was reported on MDS for the first time.

Adult ; Chromosomes, Human, Pair 1 ; Chromosomes, Human, Pair 18 ; Humans ; In Situ Hybridization, Fluorescence ; Karyotyping ; Male ; Myelodysplastic Syndromes ; genetics ; Translocation, Genetic ; genetics

OBJECTIVETo determine the value of fluorescence in situ hybridization (FISH) to the diagnosis of chromosome abnormality in genetic diseases and prenatal diagnosis.

METHODSFISH was performed using appropriate probes, including alpha-satellite DNA probe, chromosome sequence specific probe and whole chromosome painting probe, to examine the blood samples from 36 patients who were suspected of having chromosome abnormality by conventional cytogenetics, and to examine the amniocytes from 45 pregnant women who were in need of prenatal diagnosis.

RESULTSAmong 36 patients, the following karyotypes 45, X; 45, X/46, XX; 45, X/46, Xr(X); 46, X, i(Xq); 47, XXY; 46, XX, t(4;7); 47, XYY; 47, XXX; 47, XXY, inv(7); 46, XY, inv(7); 47, XX, +21 were detected by FISH. Of the fetuses of the 45 pregnant women, two fetuses with chromosomal abnormalities were diagnosed by FISH; the karyotypes were 47, XX, +18 and 46, XY, der(15) t(Y;15) respectively.

CONCLUSIONFISH can precisely and rapidly detect the chromosome abnormalities. It is a complement to the conventional cytogenetics and can be widely used in the diagnosis of genetic diseases and prenatal diagnosis.

Adult ; Amniocentesis ; Chromosome Aberrations ; Female ; Humans ; In Situ Hybridization, Fluorescence ; methods ; Karyotyping ; Male ; Prenatal Diagnosis ; Sex Chromosome Aberrations ; Turner Syndrome ; diagnosis

To explore the value of detection of PML/RARalpha gene rearrangement on bone marrow smears (BMS) by dual-color fluorescence in situ hybridization (D-FISH) for the diagnosis of acute promyelocytic leukemia (APL), the locus-specific probes for PML and RARalpha genes labeled directly and respectively by Spectrum Green and Spectrum Orange and the D-FISH technique were used to detect the PML/RARalpha gene rearrangement on BMS in 27 suspected APL patients. The results were compared with that of conventional cytogentics (CCG) and reverse transcriptase polymerase chain reaction (RT-PCR). The results showed that out of 18 newly diagnosed patients 14 were found having t(15;17) translocation by CCG and PML/RARalpha gene rearrangement were confirmed by BMS-D-FISH and RT-PCR. Thus, their APL diagnosis was determined; out of 4 patients in whom t(15;17) translocation was not detected by CCG, one had positive BMS-D-FISH and RT-PCR results, thus, this case was considered as having a cryptic t(15;17) translocation, three had negative BMS-D-FISH and RT-PCR results, thus, they were diagnosed as having acute myeloid leukemia rather than APL. In 9 cases with remission, one case with partial remission was found having t(15;17) translocation by CCG, and he had positive BMS-D-FISH and RT-PCR results, the other 8 patients (6 cases with normal karyotype and 2 cases without CCG examination) displayed different BMS-D-FISH and RT-PCR results: negative in 6 cases, but positive in 2 cases. The 2 cases were believed that they survived with minimal residual disease (MRD). It is concluded that BMS-D-FISH is a sensitive and reliable method for the detection of PML/RARalpha rearrangement. It is helpful for diagnosing APL and monitoring its MRD, and especially fit to those patients presenting a cryptic translocation or with failed cytogenetics, lacking suitable material for RT-PCR, as well as needing retrospective study.
Adolescent ; Adult ; Bone Marrow Cells ; metabolism ; Female ; Gene Rearrangement ; Humans ; In Situ Hybridization, Fluorescence ; methods ; Leukemia, Promyelocytic, Acute ; diagnosis ; genetics ; Male ; Middle Aged ; Oncogene Proteins, Fusion ; genetics ; Receptors, Retinoic Acid ; genetics ; Reproducibility of Results ; Reverse Transcriptase Polymerase Chain Reaction ; Sensitivity and Specificity