Objective To investigate the expression and distribution of aquaporin 3 (AQP3) in mouse early embryos at different stages.Methods Controlled ovarian hyperstimulation model of Kunming mouse was used to collect four-cell embryos,eight-cell embryos,morula stage,and early blastocysts.Immunofluorescence microscopy and laser confocal microscopy were used to detect expression and distribution of AQP3 channels in these stages.Results Fluorescence signal of AQP3 was found in four embryonic stages of mice.Distribution within embryo was different at different embryonic stages.AQP3 was mainly expressed on the karyotheca of blastomeres at four-cell and eight-cell stage.In morula stage,AQP3 was mainly expressed on cell membrane of each blastomere.In early blastocysts,AQP3 was predominantly expressed on the cell membrane and cytoplasm of trophoblastic cell.Conclusions AQP3 trans-membrane channel might have potential regulation function on mouse embryonic development.

Objective To compare the stress distributions on the anterior horn,body part and posterior horn of menisci between the normal and the injured knees with anterior cruciate ligament (ACL) deficiency using the three-dimensional finite element analysis.Methods A three-dimensional finite element model of tibiofemoral joint was created to simulate the motion states of the normal and ACL-deficiency knees at extension,as well as 15° and 30° flexions.Meanwhile,700N axial load and 134N posterior load were applied to the femur.Then,the stress on the anterior horn,body part and posterior horn of medial and lateral menisci were compared between the normal and ACL-deficient knees.Results With ACL deficiency,when stretching the knees straightly,the stress on the anterior horn of medial meniscus increased to 100.7％ of the normal knees,bigger than that of the affected lateral meniscus (30.7％).At 15° and 30° flexions,the stress on the posterior horn of the medial meniscus in ACL-deficiency knees increased by 36.4％ and 59.7％ respectively when compared to normal knees,while the stress on that of the lateral meniscus did not increase significantly.Apart from the stress on the body part of the lateral meniscus increasing by 39.5％ at extension in ACL-deficiency knees,no obvious changes were observed in the stress on the body part of the medial and lateral menisci.Conclusion ACL deficiency has different effect on the stress of different parts of the meniscus.It mainly increases the stress on the anterior horn of the medial meniscus at extension and that of the posterior horn of the medial meniscus at flexion.

Objective To study the clinical effect and application value of micro-plasma beam joint radiofrequency treatment for the striae of pregnancy.Methods 21 female patients with the striae of pregnancy were included in this study,treated from the July 2012 to March 2014,aged 25-37 years;and time of the striae was from 3 months to 7 years.Micro-plasma radiofrequency technology was used to treat the striae,with interval of 30 days each time for total seven months.The total effective rate,satisfaction,and the adverse reaction were evaluated after the treatment.Results 21 patients included grade 4 in 6 cases,grade 6 in 10 cases,grade 2 in 4 cases and grade 1 in 1 case;the total effective rate was 95.2％ (20/21).Satisfactory degree was for the level C in 6 cases,B in 14 cases,and A in 1 case,with total satisfactory rate of 95.2％ (20/21).Adverse reactions included mild pigmentation in 2 patients after scab skin falling off,and disappeared at the end of the treatment course.Conclusions Micro-plasma beam combined with radio frequency in treating the striae of pregnancy has clear curative effect and good clinical application value.

Objective To establish a HPLC method for determination of naringenin and apigenin in Premna fulva. Methods The SHISEIDO￣SPOLAR C18(250 mm×4.6 mm,5μm) was used as analytical column.The mobile phase consisted of methanol￣0.2% phosphoric acid (42:58) with isocratic elution at a flow rate of 1.0 mL.min-1 .The detection wavelength of naringenin and apigenin was 288 nm and 340 nm, respectively.Column temperature was set at 35 ℃ , the injection Volume was 10 μL. Results Naringenin and apigenin had a good linear relationship in the concentration range of 0.180 ~ 3.60 μg (r =0.999 9) and 0. 0052 ~ 0. 1040 μg ( r = 0. 999 9), respectively. Conclusion The method is accurate and reliable. It is appropriate for the quantitative determination of naringenin and apigenin in Premna fulva and its preparations.

0.05). Conclusion The labeling of iTRAQ in HK-2 cells is successful with favourable reproducibi-lity,which lays a foundation for the further research of proteomics in renal diseases.

To explore the feasibility of real-time quantitative PCR (QRT-PCR) for selecting cell strains which overexpress a certain transgene, expression level of RbAp46 was detected in transfected cell strains by using optimal real-time PCR with SYBR Green I. Meanwhile, semi-quantitative RT-PCR and Western blot were performed to compare with the QRT-PCR. The results showed that values of RbAp46(N) were 2064.42 +/- 253.47, 860.94 +/- 291.07, 234.456 +/- 31.08, 18.17 +/- 5.14 and 1.46 +/- 0.54 in K562/RbAp46, K562/CMV, SHG44/RbAp46 monoclone, SHG44/RbAp46 multiclone and SHG44/CMV, respectively. The results were consistent with that determined by semi-quantitative RT-PCR and Western blot. It is concluded that QRT-PCR provides a highly efficient and reproducible method for selection of transfected cell subclones at different level of transgene expression.
Blotting, Western ; Carrier Proteins ; genetics ; metabolism ; Gene Expression Regulation, Neoplastic ; Humans ; K562 Cells ; Nuclear Proteins ; genetics ; metabolism ; Organic Chemicals ; chemistry ; RNA, Neoplasm ; metabolism ; Retinoblastoma-Binding Protein 7 ; Reverse Transcriptase Polymerase Chain Reaction ; methods ; Transfection ; Transgenes ; genetics

In order to study the potential effects of exogenous WT1 gene isoform on apoptosis in leukemia cell line NB4 and its possible molecular mechanisms, the eukaryotic expression recombinant vector (pCB6(+)/WTA) containing full-length human WT1 isoform (WTA: -17aa/-KTS) cDNA and the vacant vector-alone were introduced into the leukemia cell line NB4 respectively by electroporation. The WTA mRNA and protein in cells were detected by RT-PCR and Western blot. Binding of Annexin V were tested by flow cytometry and agarose gel electrophoresis to verify whether exogenous WTA could induce apoptosis of NB4 cells. Expressions of p21, p53, bcl-2, bcl-XL and c-myc genes were determined by semi-quantitative RT-PCR after introducing recombinant vectors into the NB4 cells. The results showed that in exposure to As(2)O(3) at 0.8 micromol/L for 48 hours, the NB4/WTA cells exhibited the morphological hallmarks of apoptosis, the marked DNA ladder shown by gel electrophoresis, and the enhanced apoptosis rate marked by Annexin V. RT-PCR showed an increase in p21 and c-myc genes expression, a decrease in bcl-2 and a relative constant expression of p53, bcl-XL in NB4/WTA cells. It is concluded that the introduction and expression of exogenous WTA gene can lead to apoptosis of NB4/WTA cells by down-regulating the Bcl-2 gene expression and up-regulating the p21 and c-myc genes expression.
Apoptosis ; genetics ; physiology ; Blotting, Western ; Cell Line, Tumor ; Cyclin-Dependent Kinase Inhibitor p21 ; genetics ; Gene Expression Regulation, Neoplastic ; Humans ; Leukemia ; genetics ; metabolism ; pathology ; Proto-Oncogene Proteins c-bcl-2 ; genetics ; Proto-Oncogene Proteins c-myc ; genetics ; RNA, Messenger ; biosynthesis ; genetics ; Reverse Transcriptase Polymerase Chain Reaction ; Transfection ; Tumor Suppressor Protein p53 ; genetics ; WT1 Proteins ; genetics ; metabolism ; physiology ; bcl-X Protein ; genetics

Acute Disease ; Blast Crisis ; genetics ; Genes, Retinoblastoma ; Genes, Wilms Tumor ; Humans ; Leukemia ; genetics ; Leukemia, Myelogenous, Chronic, BCR-ABL Positive ; pathology ; Polymerase Chain Reaction ; U937 Cells

OBJECTIVETo study the potential effects of exogenous WT1 gene isoform on the differentiation of leukemia cell line NB4 and its possible molecular mechanisms.

METHODSThe recombinant eukaryotic expression vector (pCB6 + /WTA) containing full-length human WT1 isoform (WTA: -17AA/ -KTS) cDNA and the blank pCB6 + vector were transfected into leukemia cell line NB4 by electroporation. The WTA mRNA and protein in cells were detected by RT-PCR and Western blot. Cell morphology, NBT reduction and CD11b antigen expression in NB4 cells were assayed to evaluate cell differentiation. Expression of PML/RARalpha, p21 and c-myc genes was determined by semi-quantitative RT-PCR after transfection.

RESULTSCompared with NB4/WTA cells, NB4 and NB4/CMV (NB4 cells transfected with pCB6 + vector) cells had higher morphological differentiation rates and higher CD11b expression levels after exposure to ATRA for 48 hours. The percentage of NBT reduction in NB4/WTA cells was lower than that in control groups. The difference in NBT reduction rate between NB4/WTA and control cells was gradually increased after treated with ATRA for three days. The expression levels of PML/RARalpha, p21 and c-myc genes in NB4/WTA cells were notably increased.

CONCLUSIONOverexpression of exogenous WTA gene could partially inhibit the differentiation of NB4 cells by up-regulating the expression of PML/RARalpha, p21 and c-myc genes.

Cell Cycle ; Cell Differentiation ; genetics ; Cell Line, Tumor ; Cyclin-Dependent Kinase Inhibitor p21 ; metabolism ; Gene Expression Regulation, Neoplastic ; Genetic Vectors ; Humans ; Leukemia, Promyelocytic, Acute ; genetics ; metabolism ; pathology ; Proto-Oncogene Proteins c-myc ; metabolism ; RNA, Messenger ; genetics ; Transfection ; WT1 Proteins ; genetics ; metabolism

OBJECTIVETo study the DNA methylation status of WT1 gene promoter region in hematologic malignancy cell lines and its correlation with WT1 gene expression.

METHODS1. RT-PCR and methylation-specific PCR were performed for detecting WT1 gene expression and DNA methylation status in its promoter region in 8226, HL-60, Jurkat, K562, KG-1, NB4, Raji, SHI-1, U266 and U937 cell lines. 2. Treatment of U937 cells with 5-aza-CdR, a demethylation inducing agent and the changes in WT1 gene expression level and its promoter region methylation status were determined.

RESULTS1. HL-60, K562, KG-1, NB4 and SHI-1 cells showed higher levels while 8226, Jurkat, Raji, U266 and U937 cells showed extremely low levels of WT1 expression. DNA hypermethylation in WT1 gene promoter region was identified in 8226, Jurkat, Raji, U266 and U937 cells. 2. The WT1 gene expression in U937 was enhanced after treatment with 5-aza-CdR accompanied with the decrease of methylated and the increase of unmethylated levels in its promoter region.

CONCLUSIONModulation of the DNA methylation status in WT1 promoter region is one of the epigenetic mechanisms for regulating its expression.

Cell Line, Tumor ; DNA Methylation ; Gene Expression Regulation, Neoplastic ; Hematologic Neoplasms ; genetics ; Humans ; Promoter Regions, Genetic ; genetics ; WT1 Proteins ; genetics