Objective To investigate the properties of proliferation and differentiation of neonatal rat retinal progenitor cells (RPCs) in vitro. Methods RPCs were isolated from neonatal SD rats neural retina and cultured in DMEM/F12+N2 with EGF and bFGF (suspension medium )or 10%FBS without EGF and bFGF (differentiation medium). The cells grew as suspended spheres or adherent monolayers, depending on different culture conditions. The neural stem cells or retinal progenitors, neurons, astrocytes, retinal ganglion cells, rod photoreceptors and the proliferating cells were evaluated with immunofluorescence analysis by Nestin or Pax6, Map2, GFAP, Thy-1, Rhodopsin and BrdU antibodies respectively. Results RPCs could propagate and differentiate in suspension or differentiation medium and express the markers of Nestin (92.86%) or Pax6 (86.75%), Map2 (38.54%), GFAP (20.93%), Thy-1 (27.66%) and Rhodopsin(13.33%)in suspension medium; however, Nestin (60.27%), Pax6 (52%), Map2 (34.94%), GFAP (38.17%), Thy-1(30.84%) and Rhodopsin (34.67%) in differentiation medium. 96.4% of the population in the neurospheres was BrdU-positive cells. The cells could spontaneously adherent forming some subspheres and retinal specific cell types. Conclusion Neonatal rat RPCs possess the high degree of proliferation and can differentiate into neurons, astrocytes, retinal ganglion cells and rod photoreceptors in vitro. There are different proportions for RPCs to differentiate into specific cell types.

Child ; Child, Preschool ; Female ; Follicle Stimulating Hormone ; blood ; Gonadotropins ; blood ; Humans ; Infant ; Infant, Newborn ; Male ; Sex Factors ; Time Factors

Adolescent ; Child ; Collagen Type I ; metabolism ; Extracellular Matrix ; metabolism ; Female ; Fibrillin-1 ; Fibrillins ; Heart Defects, Congenital ; metabolism ; pathology ; Humans ; Male ; Microfilament Proteins ; metabolism

Antiviral Agents ; adverse effects ; therapeutic use ; Child ; Hepatitis B, Chronic ; complications ; drug therapy ; Humans ; Interferon Type I ; adverse effects ; therapeutic use ; Male ; Vitiligo ; chemically induced ; etiology

OBJECTIVETo investigate the relationship between polymorphisms of genes (CYP17, CYP19 and SULT1A1) involved in estrogen metabolism and susceptibility to breast cancer in Chinese women.

METHODSA case-control study was performed. PCR-base restriction fragment length polymorphism (PCR-RFLP) and short tandem repeat polymorphism (STRP) assays were used to detect the polymorphism distribution of CYP17, CYP19 and SULT1A1 in 213 breast cancer cases and 430 matched controls. Logistic regression analyses were used to determine the OR, multivariate adjusted OR and 95% CI of each and all three genes and estrogen exposure factors on the risk of breast cancer. Relationship between polymorphisms and clinic-pathological features was also assessed.

RESULTSThe frequency of A2 allele of CYP17 was 49.8% in cases and 49.1% in controls (P > 0.05). The frequency His allele of SULT1A1 in cases (13.6%) was significant higher than that of controls (9.5%) (P = 0.03). There was also significant difference in the frequencies of (TTTA)10 allele CYP19 which was 12.4% in cases and 8.2% in controls (P = 0.02). Multigenic model indicated that there was an increased risk of breast cancer with more numbers of high-risk genotypes in a dose-response effect (trend P = 0.05). Data from multivariate analysis showed that the allele of SULT1A1 His and CYP19 (TTTA)10 was positively associated with the risk of breast cancer. Other well-established risk factors as higher estrogen exposure including total years of menstrual, early menarche etc, and women with a higher BMI and WHR were all served as independent risks.

CONCLUSIONThis study indicated that the polymorphisms of estrogen-metabolizing genes were related to breast cancer.

Aromatase ; genetics ; Arylsulfotransferase ; genetics ; Breast Neoplasms ; genetics ; Case-Control Studies ; China ; Female ; Genetic Predisposition to Disease ; Humans ; Polymerase Chain Reaction ; Polymorphism, Restriction Fragment Length ; Steroid 17-alpha-Hydroxylase ; genetics

This study was aimed to detect the methylation status of FHIT gene promoter region in the DNA from plasma of patients with myelodysplastic syndrome (MDS), and to investigate the demethylating effect of decitabine. Methylation-specific PCR method was used to detect the methylation status of FHIT gene promoter region in the DNA from plasma of 4 patients with MDS before and after treatment with decitabine plus semis CAG therapy (among them, 1 case of newly diagnosed MDS, 3 cases progressed into acute leukemia). The results indicated that 3 cases were found to have an increased methylation in the promoter region. After treatment with decitabine plus semis CAG, increased methylation was reversed in 2 cases. In 4 cases, 2 cases displayed clinical response. It is concluded that FHIT gene hypermethylation is associated with MDS pathogenesis. Decitabine has demethylating effect on the FHIT gene hypermethylation of plasma from MDS patients. Detecting the methylation status of FHIT gene in DNA from plasma may play a role in MDS auxiliary diagnosis or prognosis.
Acid Anhydride Hydrolases ; genetics ; Adult ; Aged ; Azacitidine ; analogs & derivatives ; therapeutic use ; DNA ; blood ; DNA Methylation ; Female ; Humans ; Male ; Middle Aged ; Myelodysplastic Syndromes ; blood ; drug therapy ; Neoplasm Proteins ; genetics ; Promoter Regions, Genetic

OBJECTIVETo study the effects of dexamethasone on intracellular expression of Th17 cytokine interleukin 17 and the mechanisms in asthmatic mice.

METHODSExperimental asthma was induced by ovalbumin (OVA) sensitization in 20 in female Balb/c mice with (dexamethasone group, n=10) or without dexamethasone treatment (model group, n=10), with another 10 serving as the control group. The levels of IL-17 in the bronchoalveolar lavage fluid (BALF) and serum of the mice were measured by enzyme-linked immunosorbent assay (ELISA), and the airway inflammation was evaluated by HE staining. The expressions of IL-17 and RORgammat mRNA were measured by reverse transcription-polymerase chain reaction (RT-PCR), and the expression of RORgammat protein was measured by immunohistochemical staining.

RESULTSThe levels of RORgammat and IL-17 mRNA and protein in the asthmatic model group were significantly higher than those in the control group (P<0.01), and the increased expressions of RORgammat and IL-17 mRNA and protein in the asthmatic mice were significantly reduced by dexamethasone treatment (P<0.05).

CONCLUSIONDexamethasone can inhibit the release of IL-17 probably by inhibiting RORgammat expression and blocking Th17 differentiation in asthmatic mice.

Animals ; Asthma ; chemically induced ; immunology ; metabolism ; Dexamethasone ; pharmacology ; Female ; Interleukin-17 ; genetics ; metabolism ; Mice ; Mice, Inbred BALB C ; Nuclear Receptor Subfamily 1, Group F, Member 3 ; metabolism ; Ovalbumin ; RNA, Messenger ; biosynthesis ; genetics ; T-Lymphocyte Subsets ; immunology ; metabolism ; T-Lymphocytes, Helper-Inducer ; immunology ; metabolism

This study was aimed to investigate the effects of xenogeneic antigen neu-Fc in combination with the recombinant human granulocyte-macrophage colony stimulating factor (GM-CSF) and Bacillus Calmette-Guerin (BCG) on the regulation of Th1 and Th2 immune response in vitro. The rat neu L2-S2 domain was engineered as a chimeric protein with human IgG Fc. The eukaryotic expression vector was constructed. The recombinant protein was stably expressed in CHO cells and purified by rProtein A Sepharose Fast Flow column. The recombinant protein was identified by SDS-PAGE and Western blot. Peripheral blood mononuclear cells (PBMNCs) were obtained by means of standard Ficoll separation from the blood of healthy donors. Neu-Fc-induced PBMNC proliferation was tested by MTT. The production of IL-12 and IL-10 was measured by ELISA. The results showed that the level of IL-12 decreased and IL-10 increased after PBMNCs were incubated with MCF-7 cultural supernatant. 10 nmol/L neu-Fc strongly induced the cell proliferation. Compared with neu-Fc or GM-CSF or BCG treatment alone, neu-Fc in combination with GM-CSF and BCG significantly stimulated IL-12 production and inhibited IL-10 production (p < 0.01). It is concluded that the neu-Fc can stimulate the proliferation activity of PBMNCs. neu-Fc, GM-CSF and BCG costimulation efficiently induces Th1 immune response.
Animals ; BCG Vaccine ; immunology ; CHO Cells ; Cricetinae ; Cricetulus ; Granulocyte-Macrophage Colony-Stimulating Factor ; immunology ; Humans ; Interleukin-10 ; metabolism ; Interleukin-12 ; metabolism ; Rats ; Recombinant Proteins ; immunology ; Th1 Cells ; immunology ; Th2 Cells ; immunology

Eight full-length genes of an avian influenza virus Chinese isolate of H9N2 subtype, A/Chicken/Guangdong/HL/2006 (H9N2) (abbreviated as Ck/GD/HL/06), were amplified by RT-PCR, including the 5' and 3' non-coding region. All the genes were cloned and sequenced. The phylogenetic analysis results showed the HA gene of Ck/GD/HL/06 was located in the same phylogenetic clade as Dk/HK/Y280/97 (H9N2), while the Dk/HK/Y280/97-like viruses had been predominately isolated from chickens in mainland China. After the analysis of glycosylation sites and receptor-binding sites in the HA, it was shown that the HA of Ck/GD/HL/06 exhibited the common feature of H9 subtype avian influenza virus isolated from China, but the leucine (Leu) residue at the amino acid position 226 indicated the potential of binding with SA alpha,2-6 receptor. The three internal genes of Ck/GD/HL/06 (PB1, PA and NP) had the highest nucleotide identity with A/Viet Nam/1203/2004 (abbreviated A/VN/1203/04) isolate, which was shown to be transmitted from chickens to human and caused lethal infection in human. No analogous H9N2 strains was reported in previous studies. Based on the high similarity of Ck/GD/HL/06 three genes to A/VN/1203/04, it was suggested that the possibility of generating new highly pathogenic H5N1 AIVs by recombination was worthy of our attention. Further studies should be needed for molecular epidemiologic surveillance of H9N2 AIV in the south China for a long time.
Amino Acid Sequence ; Animals ; Chickens ; China ; Cloning, Molecular ; Evolution, Molecular ; Genes, Viral ; genetics ; Genomics ; Influenza A Virus, H9N2 Subtype ; genetics ; isolation & purification ; Influenza in Birds ; virology ; Phylogeny ; Polymerase Chain Reaction ; Sequence Alignment ; Sequence Analysis, DNA ; Sequence Homology, Nucleic Acid ; Viral Proteins ; chemistry ; genetics

Eukaryotic initiation factor 5A (eIF-5A) contains an unusual amino acid, hypusine, which is formed post-translationally. Although eIF-5A and its hypusine modification are essential for eukaryotic cell viability, the precise physiological function of it has remained elusive. The aim of the study is to investigate how hypusine formation modulate the proliferation, cell cycle and apoptosis in leukaemia cells. The effects of 1,7-diaminoheptane (DAH), a potent inhibitor of deoxyhypusine synthase, on proliferation and cell viability of leukemia cell lines (Mo7e, TF-1 and THP-1) and MCF-7 cells, were investigated. eIF-5A expression level was detected after cell synchronization. The results showed that inhibition of cell proliferation by DAH was in a concentration-dependent manner while apoptosis was also induced at the same time. Upon treatment of the cell lines with DAH, cell growth was inhibited. Cell cycle analysis showed that DAH induced cell growth arrest at the G(1)-S boundary of the cell cycle. In synchronized MCF-7 cells, the expression level of eIF-5A peaked at G(1) phase but very low at S and G(2)/M phases. It is concluded that hypusine formation of eIF-5A exits in the regulation of cell cycle and the results suggest that eIF-5A is involved in the expression of proteins regulating transition of G(1)-S phase of cell cycle.
Cell Line, Tumor ; Diamines ; pharmacology ; G1 Phase ; physiology ; Humans ; Lysine ; analogs & derivatives ; metabolism ; Peptide Initiation Factors ; physiology ; RNA-Binding Proteins ; S Phase ; physiology