[Objective] To evaluate the therapeutic effect of Oleum Curcumae Aromaticae (OCA) on leukemia cell strain HL-60 and K562 by observing its sensitivity in inhibiting HL-60 and K562. [Methods] Subculture of HL-60 and K562 cells was performed on the 96-hole culture plate and OCA 100?L at the final concentration of 400, 200, 100 and 50mg/L respectively was added. The blank control was added with RPMI-1640 culture fluid. After 24, 48 and 72 hours of treatment, MTF 10?L (5mg/mL) was added and the inhibitory rate on the cells was assayed by detecting the absorption value at 570nm wavelength. [Results] The inhibitory effect of OCA on HL-60 and K562 increased with the treatment time. The difference of inhibitory rates 24, 48 and 72 hours after treatment were significant (P

Objective By in vitro culture of mouse macrophage cell line RAW264. 7 and primary mouse bone marrow macrophages, the expression of Siglec-1 when stimulated by ox-LDL was observed. Meanwhile, Siglec-1 was up-regulated by M-CSF and down-regulated by small interference RNA targeting Siglec-1 ( si-RNA-Siglec-1) , and the expression of chemokines and lipid uptake ability by macrophages were observed, to explore the role of Siglec-1 on macrophages in atherosclerosis. Methods LDL was oxidized by copper. According to preliminary experiment results, ox-LDL 100 μg/ml was selected as a stimulus. There were 6 experimental groups:normal control group,ox-LDL 100 μg/ml group, ox-LDL 100 μg/ml + si-RNA 2509 2 ng/ml group,ox-LDL 100 μg/ml + si-RNA 3618 2 ng/ml group,ox-LDL 100 μg/ml + M-CSF 5 ng/ml group and ox-LDL 100 μg/ml + M-CSF 10 ng/ml group. si-RNA-Siglec-1 was transfected into macrophage to inhibit the expression of Siglec-1, whereas M-CSF 10 ng/ml or 5 ng/ml were added into the culture medium to enhance the expression of Siglec-1. Quantitative real-time polymerase chain reaction ( qRT-PCR) was used to determine the interfere efficiency of si-RNA-Siglec-1 or M-CSF. After stimulation with ox-LDL for 48 h, cell culture supernatants were collected to determine MIP-1 alpha, MCP-1 and IL-8 concentration by ELISA (n =3 for each group) to evaluate the activation of macrophages. Internalization of lipid particles by macrophages was analyzed by oil red 0 staining. Results Observed by fluorescence microscope, si-RNA-Siglec-1 could be effectively transfected into macrophages with a transfection efficiency about 90% ;PCR results showed that si-RNA 2509 and si-RNA 3618 in a concentration of 40 pmol/L had an inhibition rate of 0. 54 ±0. 11 or 0. 52 ±0. 16 vs 1. 00 ±0. 24 (control group) , t =5. 227 and 4. 992, respectively, all P < 0.01, while M-CSF 10 ng/ml could increase Siglec-1 mRNA expression approximately 4-fold (4. 16 ± 1. 25 vs 1.00 ±0. 24, t =7. 448, P<0. 01). The secretion of MCP-1, MIP-1 alpha, and MIP-2 in si-RNA3618-Siglec-1 group [(359. 28±47. 80) pg/ml, (33. 76 ± 14. 28) ng/ml and (7.87±1.55) ng/ml for MCP-1,MIP-1 alpha, and MIP-2, respectively] was significantly reduced in compare with ox-LDL 100 μg/ml group [ (577. 89 ± 35. 95 ) pg/ml, (69. 17 ± 11. 82) ng/ml and (12.28 ± 1.19) ng/ml for MCP-1, MIP-1 alpha, and MIP-2, respectively], with P value of 0.01, 0.05 and 0.01. In contrary, ox-LDL 100 μg/ml plus M-CSF 10 ng/ml group could significantly promote macrophage chemokine secretion [ (672. 89 ± 43.80) pg/ml, (101.31 ±24.17) ng/ml and (14.81 ±0.54) ng/ml for MCP-1, MIP-1 alpha, and MIP-2, respectively], with P < 0.05 compared with ox-LDL 100 μg/ml group. Meanwhile, lipid intemalization and foam cell formation was inhibited in si-RNA3618-Siglec-l group while ox-LDL 100 μg/ml plus M-CSF 10 ng/ml group could enhance the phagocytosis of ox-LDL by macrophage. Conclusions Siglec-1 may served as a potential phagocytic receptor for ox-LDL involving in macrophage uptake of lipid and turn into foam cells. Furthermore, it can active macrophages and enhance the secretion of MIP-1 alpha, MCP-1 and IL-8, attracting more macrophages and lymphocytes to the site of inflammatory plaque. Targeted inhibition of Siglec-1 reduces macrophage uptake of lipid and secretion of chemokines. Siglec-1 may possibly serve as a potential target of treatment or delay the development of atherosclerosis.

Objective To investigate whether the hyper-reactivity of monocytes from the patients with primary biliary cirrhosis (PBC) to LPS and Poly I ∶ C was associated with miR-146a.Methods Peripheral blood mononuclear cells from PBC patients and healthy controls were stimulated with 10 μg/ml of LPS or Poly I ∶ C.The levels of IL-1 β,IL-6,TNF-α and IFN-α in the culture medium were detected by ELISA.The relative expressions of miR-146a,miR-21,let-7e,miR-155 and miR-125b were detected by RT-PCR.The regulatory role of miR-146a on the production of inflammatory factors in LPS or Poly I ∶ C stimulated THP-1 cells was studied by gain-and-loss function assay.Results The up-regulation of miR146a,which was induced by both LPS or Poly I ∶ C,was impaired in the monocytes from PBC patients.The production of LPS or Poly I ∶ C induced inflammatory factor,could be enhanced by miR-146a down-regulation.Conclusion The hyper-reactivity of monocytes from PBC patients to LPS and Poly I ∶ C was associated with miR-146a.

Objective To construct lentiviral vectors containing small interfering RNA (siRNA) sequence of Siglec-1 and to screen the effective vector.Methods Three fragments of Siglec-1 siRNA were designed and cloned into pGCSIL-GFP lentiviral plasmid.And then the plasmid was cotransfected into 293T cells with pHelper 1.0 and pHelper 2.0 plasmids.Forty-eight hours later,culture supernatant with virus particles was collected and concentrated.Virus titer was determined by 10-fold serial dilution method and virus was transduced into primary cultured mouse bone marrow-derived macrophages (BMM).Flow cytometry and QRT-PCR were used to screen effective vector with inhibition ability.Results Three vshRNA lentiviral plasmids and a control plasmid were constructed successfully and verified by DNA sequencing.Virus titer was between 1×10s TU/ml and 1×109 TU/ml,which was suitable for in vitro and in vivo experiments.The Lv-1 could inhibit Siglec-1 expression effectively in vitro transduction of BMM.Conclusion Lentiviral vectors containing siRNA sequence of Siglec-1 were constructed successfully and an effective vector was screened,which may lay the foundation for using the vector in gene knockdown experiment in vivo.

Objective To investigate the effect of H.pylori infection and eradication on gastric parietal cell and H + K +ATPase mRNA expression in a murine model. Methods Twenty 7 week old SPF BALB/C mice (10 males and 10 females) each were fed by H.pylori strain (Sydney Strain 1,SS1) at a dose of 0.4 ml (10 9CFU) per day for consecutive 5 days. Two months after infection of H. pylori, all mice were divided into two groups, the eradication group (10 mice) and the infection group (10 mice). Mice in the eradication group were administered clarithromycin ( 13.5 mg?kg -1 ?d -1 ) twice per day for one week (one mouse was died).Meanwhile, mice in the infection group were given the same amount placebo. All mice were killed at one month after the administration.The gastric mucosa was removed for rapid urease testing (RUT) and Giemsa stainning. The expression of H + K +ATPase mRNA was detected by RT PCR. Morphological changes in parietal cells were assessed by electron microscope. Results The animals in infection group were 100% infected by H.pylori, and RUT and Giemsa staining were all positive. Meanwhile , all but one mouse in the eradcation group were negative to RUT and Giemsa staining. In the infection group, the average ratio A C to A T (A C means the area of the canaliculi, A T means the area of the parietal cells ) was ( 2.20 ? 0.06 )/10 4, significant lower than that in the eradication group [(3.20 ? 0.06 )/10 4, P

A simple and effective method for the duplex PCR detection was developed by using sequences of exogenous gene(RDV MP-)and endogenous gene(Zein)as templates for PCR amplification.The results of routine PCR amplification for RDV MP-gene in transgenic maize suggested that RDV MP-gene can stably inheritate in transgenic plants and their progenies;The duplex PCR detection of all negative and part positive samples that obtained by routine PCR amplification confirmed that above negative results were exact,also showed that the quality of extracted DNA can meet the need of PCR amplification.The error ratio of negative samples was 1.4%.The method used in this study was simple and credible and can be used to detect transgenic plants and their products.

Objective To explore the auxiliary method of urethral reunion operation on bulbous urethral complete rupture by en-doscope .Methods 11 cases of bulbous urethral complete rupture were inserted urethral probe from suprapubic puncture cystosto-my ,when it was difficult to find near end of disrupt urethra only by endoscopic realignment .Results 9 cases were reconstructed in one-stage endoscopic urethral realignment with the aid of urethral probe from suprapubic puncture cystostomy .Conclusion Ure-thral probe from suprapubic puncture cystostomy will improve the success rate of one-stage endoscopic urethral realignment for the cases of bulbous urethral complete rupture .

The chromatographic fingerprint was established by eluting with the mobile phase consisted of acetonitrile and 0.2% formic acid water on an Agilent TC-C18 (2) column (4.6 mm x 250 mm, 5 microm). Six chromatographic peaks were identified by HPLC-MS/MS method. Ten batches of Glycyrrhizea Radix et Rhizoma Praeparata Cum Melle were determined, and the similarity was arranged from 0.72 to 0.99. Good precision, stability and repeatability were obtained, and this study provides a reference for the quality control of Glycyrrhizea Radix et Rhizoma Praeparata Cum Melle.
China ; Chromatography, High Pressure Liquid ; methods ; Drugs, Chinese Herbal ; chemistry ; Glycyrrhiza uralensis ; chemistry ; Mass Spectrometry ; Quality Control ; Rhizome ; chemistry

Adult ; Cutis Laxa ; complications ; diagnosis ; Emphysema ; diagnosis ; etiology ; Female ; Humans ; Male ; Middle Aged

Objective To investigate the increased expression of microRNA-146a(miR-146a) in peripheral blood mononuclear cells (PBMC) of patients with chronic immune thrombocytopenic purpura (ITP) and its clinical significance. Methods Twenty-eight patients with chronic ITP and 28 healthy controls matched with age and gender were enrolled in this study. Fluorescent quantitative PCR reaction was used to detect the relative expression of miR-146a in their PBMC. The serum concentration of TNF-α, IL-2,IL-1 β and IFN-γ were measured by ELISA. CCK-8 method was used to detect the proliferation ability of PBMC , which transfected with miR-146a mimics or inhibitor and then stimulated with platelet . Results The relative expression of miR-146a in ITP patients was higher than that of healthy controls. The increased expression of miR-146a was negatively correlated with the serum TNF-α, IL-2 and IFN-γ. The PBMC transfected with miR-146a mimics had reduced expression of IL-2 and proliferation when stimulated with platelet.In contrast, the opposite effect was observed with the miR-146a inhibitors transfection. Conclusion MiR146a was involved in the pathogenesis of chronic ITP by controlling IL-2 production and PBMC proliferation.Thus, it may be a potential therapy target for chronic ITP.