Objective:To study the influence of Wnt5a gene silence on the proliferation,migration and invasion of lung squamous carcinoma cells.Methods:A recombinant plasmid pHI-siRNA~(Wnt5a-)was constructed and used to deliver small interference RNA (siRNA)targeting Wnt5a in SK-MES-1 cells;the transfected cells were screened to establish a stable transgenic cell line.MTT,cell cycle and Transwell assays were employed to evaluate the effect of Wnt5a gene silence on the proliferation,migration and invasion of lung squamous carcinoma cells.Results:Western blotting assay revealed that Wnt5a was lowly expressed in SK-MES-1~(Wnt5a-)(13.6%).The proliferation index(PI)of transgenic cell line was slightly lower than that of the control cell line([28.3?3.8]% vs[30.5?5.2]%). The migration and invasion capabilities of SK-MES-1~(Wnt5a-)cells were(47.3?9.2)% and(39.7?11.7)% of the control cells, respectively.Conclusion:Low Wnt5a expression can significantly inhibit the migration and invasion capabilities of SK-MES-1 cells, indicating that Wnt5a might be a potential target for gene therapy of lung squamous carcinoma.

It is of great clinical significance to investigate noninvasive glucose detection. As one of the most potential methods, the noninvasive glucose detection based on near-infrared has become a hot research area recently. In this paper the principles and research methods of noninvasive glucose detection based on near-infrared spectrum are introduced. With the comparison between the research status at home and abroad in recent years, we summarize and discuss crucial issues in near-infrared noninvasive glucose detection, such as the selection of measurement method, selection of measurement position and choice of wavelength, and, furthermore, setting up models.
Blood Glucose ; analysis ; Diabetes Mellitus ; blood ; Humans ; Monitoring, Physiologic ; methods ; Spectroscopy, Near-Infrared ; instrumentation ; methods

Aim The effect of tetrandrine on TGF-?1 mRNA expression in glomerulosclerosis rat was observed. Methods The rats were randomly divided into four groups, such as the normal control group (sham operative rat), glomerulosclerosis model group,tetrandrine group and amlodipine group. The expression of TGF-?1 mRNA was analyzed by Northern blot hybridization. Results The expressions of TGF-?1 mRNA in two treating groups were much lower than untreated model group. There were no difference between these two treating groups. Conclusion Tetrandrine can decrease the expression of TGF-?1 mRNA in glomerulosclerosis rat induced by unilateral renctomy plus adriamycin.

BACKGROUND:Systemic erythematosus lupus (SLE) is closely associated with activity of dendritic cells (DCs) function.DCs derive from hematopoietic precursor cells (HPCs) in vivo and many factors might affect the differentiation and maturation of DCs.The cytokine network was in disturbance and the levels of various cytokines were anomalous in SLE serum.Previous studies mainly addressed effects of single factors.OBJECTIVE:To investigate the effects of joint action of serum interferon (IFN)-α and interieukin (IL)-6 in the serum of SLE patients on the differentiation and maturation of DCs derived from CD34~+ HPCs.DESIGN,TIME AND SETTING:The grouping,controlled and completely random designed study was performed at the Department of Microbiology and Immunology,Nanjing Medical University from May 2006 to October 2008.MATERIALS:Peripheral blood samples were collected from 50 SLE patients from the Department of Rheumatology,First Affiliated Hospital of Nanjing Medical University and 30 healthy volunteers from Nanjing Medical University.Cord blood samples were collected from 15 full-term normal delivery neonates from Nanjing Maternity and Children Health Care Hospital Affiliated to Nanjing Medical University.Informed consent was obtained from all patients and their family members.METHODS:Peripheral blood samples were collected from SLE patients and healthy volunteers.According to serum IFN-α and IL-6 concentrations of normal person,95% reference value range of serum IFN-α and IL-6 concentrations were calculated by the method of percentiles.Above this range represented abnormal increase.Cord blood mononuclear cells were harvested by Ficoll-Hypaque density gradient centrifugation.CD34~+HPCs were purified from cord blood by magnetic cell sorting system (MACS),and cultured to differentiate to DCs.There were 6 groups in this study.Normal serum,SLE serum with elevated levels of IFN-α,SLE serum with elevated levels of IL-6,SLE serum with elevated levels of IFN-α and IL-6,SLE serum with anti-IFN-α and anti-IL-6 neutralizing antibodies,or normal serum with exogenous IFN-α and IL-6 were respectively added to the culture medium in each group.10% volume fraction serum was used in each group,for totally 14 days incubation.MAIN OUTCOME MEASURES:The phenotype of DCs was analyzed by flow cytometry (FCM).Cytokine production was assessed by ELISA.The capacity of DCs to stimulate allogenic T lymphocyte proliferation and to regulate T cell differentiation was evaluated in a mixed lymphocyte reaction (MLR).RESULTS:Compared with normal serum,HLA-DR,CD80 and CD86 expression was significantly increased in the medium containing SLE serum with elevated levels of IFN-α,SLE serum with elevated levels of IL-6,SLE serum with elevated levels of IFN-α and IL-6 and normal serum with exogenous IFN-α and IL-6 (P＜0.05),whereas IL-12 levels were significantly decreased (P＜0.05),and capacity of DCs to stimulate allogenic T lymphocyte proliferation was significantly enhanced (P＜0.05).Above-mentioned indexes recovered to a normal level in the medium containing SLE serum with anti-IFN-α and anti-IL-6 neutralizing antibodies.Compared with normal serum,proportion of both CD3~+CD8~-IFN-γ~+ and CD3~+CD8~+IFN-γ~+ T cell subsets was significantly reduced in the medium containing SLE serum with elevated levels of IL-6 (P＜0.05).However,proportion of both CD3~+CD8~-IFN-γ~+ and CD3~+CD8~+IFN-γ~+ T cell subsets was significantly increased in the medium containing SLE serum with elevated levels Of IFN-α,SLE serum with elevated levels of IFN-α and IL-6,normal serum with exogenous IFN-α and IL-6 (P＜0.05).CONCLUSION:The joint action of IFN-α and IL-6 in SLE serum promotes the differentiation and maturation of DCs derived from CD34+HPCs and affects the DC-mediated T cell differentiation,which might contribute to the pathogenesis of SLE.

BACKGROUND: Blood stem cells (BSCs) are the most primitive cells in the immune system and can be differentiated into many kinds of cells. As the regulatory cells of immune response, dendritic cells (DCs) have been attracted more and more attention in the field of autoimmune diseases. Due to different resources of precursor cells, DCs have different cytokines, ideal cytokine matching,applying orders, and experimental cultured conditions. Furthermore, development, phenotypic expression, and mature degree are still different.OBJECTIVE: To investigate the influence of tumor necrosis factor- α (TNF- a ) and interleukin-4 (IL-4) on the culture system and provide improved method for inducing functional DCs in vitro, derived from cord blood CD34+ hematopoietic precursor cells (HPC).DESIGN, TIME AND SETTING: Observational study, which was performed in the Department of Microbiology and Immunology, Nanjing Medical University from March 2005 to November 2005.MATERIALS: The cord blood was collected from neonatal umbilical cord in the 81 Hospital of Nanjing City. CD34 monoclonal antibody coated magnetic bead system (MACS) was provided by Miltenyi Biotec Company, Germany; human recombinant granulocyte-macrophage colony-stimulating factor (rhGM-CSF), human recombinant interleukin-4 (rhlL-4), and human recombinant tumor necrosis factor-α (rhTNF-α) by Pepro Tech Company, USA.METHODS: CD34+HPC were isolated and purified from umbilical cord blood by using a high-gradient magnetic cell sorting system (MACS). Then the cells were cultured with different culture medium which contained different combinations of cytokines:GM-CSF and TNF-α (GT) or GM-CSF, TNF-α and IL-α (GTI) in order to be induced to differentiate into dendritic cells (DCs).The DCs derived from CD34+HPC were identified for their morphology and phcnotype by FACS and laser scanning con_focal fluorescence microscopy, and also for their abilities of inducing proliferation of allogenic T cells by 3H-TdR incorporation assay.RESULTS: The purity of selected CD34+ cells with MACS was more than 90%. DCs could be obtained from CD34+HPC by the culture in presence of GM-CSF and TNF-α or GM-CSF, TNF- o and IL-4. With the time of culture lasting, the cells expressed lower level surface antigen of CD34 and HLA-DR (P ＜ 0.05), and possessed the phenotypes of DCs characterized by higher expression of CD80, CD86, CD83 and CDIa. At 13-15 days, the cells possessed higher level of phenotypes of DCs compared to 7-9 days and 10-12 days. DCs induced with GTI culture system expressed higher levels of surface antigen CD80, CD86, CD83,CD 1 a and a lower level of CD 14 than those induced with GT culture system. DCs displayed typical morphology and property and expressed higher level surface antigen of CD86 and CD80, especially expressed CD86 when they were induced with proper cytokines of GM-CSF and TNF- α added at 0 hour, and of IL-4 added at 48 hours.CONCLUSION: DCs can be generated from CD34+HPC by proper culture system, the design of GM-CSF + TNF- α + IL-4(GM-CSF and TNF- α were added at 0 hour, IL-4 was added at 48 hours) is preferred.

DNA ; DNA Fingerprinting ; Homicide ; Humans

OBJECTIVE To investigate the hepato-protective mechanism of thymoquinone (TQ) onthe development of acetaminophen (APAP)- induced liver injury. METHODS In vivo, male kunming mice were injected with a single dose of 300 mg·kg-1 APAP. Some mice were pretreated with TQ (5 or 20 mg·kg-1) and N-acetylcysteine (NAC, 300 mg·kg-1) 2 h before APAP injection. Mice were euthanized at 2 h, 6 h, 12 h after APAP treatment. In vitro, human Chang liver cells were incubated with 3.125, 6.25 or 12.5 μmol·L-1 TQ, 10 μmol·L-1 SP600125 and 500 μmol·L-1 AICAR in the presence of APAP for 24 h. Cell viability were analyzed by MTT assay, protein expressions were assessed by Western blot. RESULTS TQ pretreatment significantly reduced serum aminotransferase and increased hepatic gluta?thione (GSH) and glutathione peroxidase (GSH-PX) activities, while significantly inhibited interleukin-1β(IL-1β) levels. TQ significantly inhibited c-Jun N-terminal kinase (JNK), extracellular signal regulated kinase (ERK) and P38 phosphorylation induced by APAP. Moreover, TQ inhibited phosphatidylinositol 3- kinase (PI3K)/mammalian target of rapamycin (mTOR) signaling activation and activated AMPK phosphorylation induced by APAP. In addition, TQ inhibited signal transducer and activator of transcription 3 (STAT3) phosphorylation on APAP-induced liver injury. In vitro, APAP enhanced JNK phosphorylation and attenuated AMPK phosphorylation in Chang liver cells, and these effects were blocked by pretreatment with TQ, SP600125 (JNK inhibitor) and AICAR (AMPK activator). CONCLUSION Our findings suggest that TQ may actively prevent APAP-induced liver injury, and this effect may be mediated by JNK and AMPK signaling pathways.

OBJECTIVE To investigate the mechanism of SIRT1/AMPK signaling pathway between hepatocytes and hepatic stellate cells (HSCs). METHODS Normal human Chang liver cells and human hepatic stellate cell line, LX-2 cells were treated with SRT1720 (10 μmol·L-1) and AICAR (500 μmol·L-1) prior to ethanol (50 mmol·L-1) for 24 and 48 h. Cell viability was analyzed by methyl thiazolyl tetrazolium assay. SIRT1, AMPK and p-AMPK mRNA levels for 24 h and 48 h were analyzed by RT-PCR, SIRT1, AMPK and p-AMPK protein expressions in the supernatant at 24 and 48 h was detected by Western blot. RESULTS SRT1720 and AICAR effectively decreased LX-2 cell viabilities and exhibited scarcely little toxicity in human Chang liver cells. SRT1720 and AICAR attenuated collagen-I, α-smooth muscle actin (α-SMA) levels, activated liver kinase B-1 (LKB1) and AMPK phosphorylation in ethanol treated LX-2 cells. Meanwhile, SRT1720 and AICAR enhanced SIRT1 expression mediated by ethanol both in Chang liver cells and LX-2 cells. Furthermore, SRT1720 and AICAR suppressed the expression of sterol regulatory element-binding protein-1 (SREBP-1) to regulate fatty acid synthesis. CONCLUSION SIRT1 agonist and AMPK agonist blocked the crosstalk between hepatocytes and HSCs via SIRT1/AMPK signaling pathway to modulate hepatocytes accumulation of lipid and HSCs activation.

s:The chemical constituents of gliocladium roseum(called Y)accelerating the growth of famous medicin al plant Anoectochilus roxburghiiwas studied.Five comp ounds were separated by silica gel column chromatograph from this fungal mycelia and their structures were elu cidated by the data of IR,NMR,UV and MS.Compound I was 6,22-diene-3-hydroxy- 5,8-epidioxy ergosta,compound 2 is ergosterol,compound 3 is D-arabitol and com pound 4 is mannitol.

The medium and process parameters were optimized in batchaminoglycoside antibiotic JI-20A fennentation. The optimal medium consists mainly of comstarch 60g/L, peanut meal 30g/L, com slurry 8g/L, maltose 10g/L, inorganic compound and amylase moderate , methionin 1g/L and cobalt chloride 6?g/mL. It was significant to adjust medium pH after autoclaving and oxygenate the fennentation medium for the cell growth and JI-20A product.