Objective:To observe the effects of AFP gene silencing by siRNA on the Survivin mRNA of hepatocellular carcino-ma cell line HepG2. Methods:AFP gene expression was downregulated in HepG2 cell by RNAi, and the AFP content in the superna-tant was detected by ELISA. Survivin mRNA level was tested by RT-PCR. MTT was applied to evaluate cell proliferation. Flow cytom-etry was employed to observe cell apoptosis. Results:At 48 h after transfection, AFP expression was almost completely inhibited, cell proliferation activity was decreased by 43.1%, cell apoptosis rate was increased by 24.3%, and the Survivin mRNA expression was re-duced to 22.0%in the experimental group. No evident changes were observed in negative control and blank groups. Conclusion:AFP gene silenced by RNAi induces growth inhibition and apoptosis promotion of hepatocellular carcinoma cell line HepG2. This gene may be associated with the suppression of Survivin mRNA.

Objective To explore the relationship of the ratio of plasma adrenomedullin(AM)and endothelin-1(ET-1)with serum concentration of neuron-specific enolase(NSE)in full-term neonates with hypoxic-ischemic encephalopathy(HIE).Methods Plasma concentrations of AM,ET-1 and serum NSE from 32 full-term neonates with HIE were detected by radioimmunoassay(RIA)on the 1,3 and 7 d after parturition,30 neonates in the corresponding periods in our hospital were employed as controls.The infants with HIE were divided into mild,moderate or severe group in terms of diagnostic standard of HIE.Results 1.Plasma concentrations of AM and ET-1 in newborns with mild,moderate or severe HIE were significantly higher than that of control group at 1 d after life with a decline from 3-7 d(Pa

Objective To explore the effect of exogenous adrenomedullin(ADM) on expressions of glutathion in plasma and brain tissue inneonatal rats with hypoxic-ischemia reperfusion brain damage(HIRBD) and the mechanism of action.Methods Fifty-six cases of 7 d SD rats were randomly divided into 4 groups including normal control group(without any treatment),HIRBD group(the model with hypoxia for 2 h and ischemia for 1 h),primed group(abdomen infusion of ADM at 0.5 h before making model,the other was same to the HIRBD group)and treatment group(abdomen infusion of ADM at onces after making model,the other was same to the HIRBD group).The neonatal rats in 4 groups were derived blood and brain tissue after decapitation at either 4 h or 24 h after reperfusion.The levels of gtutathion(GSH) in plasma and brain tissue were determined by using chromatometry.Results In HIRBD group,the affection areas at 24 h after reperfusion enlarged compared with those at 4 h after reperfusion.Meanwhile,the affection of punctiform degeneration or necrosis at 4 h after reperfusion transformed into the affection of lamellar or diffuse degeneration or necrosis at 24 h after reperfusion.The levels of GSH in plasma and brain tissue in HIRBD group at either 4 h or 24 h after reperfusion were significantly lower than that in normal control group(Pa0.05).Meanwhile the pathology score of brain section in primed group and treatment group were significantly lower than that in HIRBD group at either 4 h or 24 h after reperfusion(Pa0.05).Conclusion Exogenous ADM can induce the neuroprotection in HIRBD by adjusting the expression of GSH.

OBJECTIVETo prepare monoclonal antibodies (McAbs) against human mesenchymal stem cells (hMSCs) and to study their biological characteristics.

METHODSBALB/C mice were immunized with pooled hMSCs. McAbs were prepared by hybridoma technique and their biological characteristics were analyzed by indirect immunofluorescence, immunohistochemistry and flow cytometry.

RESULTFive hybridoma cell lines were successfully established, which secret McAbs specifically against hMSCs. Investigations showed that all these McAb reacted only to hMSCs and had no cross-reaction to other human cells, the relative affinities of 5 McAbs were 1x10(6) (ZUB1), 1x10(5) (ZUB4), 1x10(6) (ZUC3), ZUE12 (1x10(5)) and 1x10(5) (ZUF10), respectively. Isotype analysis showed that ZUB1, ZUE12, ZUF10 against the same isotype, while ZUC3, ZUB4 against other two different isotypes alone. Flow cytometric analysis showed that the positive expression rate of cultured hMSCs was 87.39% (ZUB1), 88.07% (ZUB4), 88.12% (ZUC3), 69.89% (ZUE12) and 83.67% (ZUF10).

CONCLUSIONThe prepared five McAbs can specifically react against hMSCs, which can be used for selection and study of hMCSs.

Animals ; Antibodies, Monoclonal ; biosynthesis ; Antibody Specificity ; Bone Marrow Cells ; cytology ; immunology ; Fluorescent Antibody Technique ; methods ; HL-60 Cells ; Humans ; Hybridomas ; secretion ; Immunoglobulin G ; immunology ; Immunoglobulin M ; immunology ; Mesenchymal Stromal Cells ; cytology ; immunology ; Mice ; Mice, Inbred BALB C ; Rats

OBJECTIVETo study the effects of the superficial venous trunk (SVT) and its nourishing vessels on early revascularization of the reverse-flow random flap in order to provide theoretical evidences for clinical applications.

METHODSThe morphologic changes of micrangium of the reverse-flow random flap with SVT and without SVT were observed at different phases and compared by optical microscopy and stereological methods.

RESULTSThe micrangium density of the flaps with SVT and without SVT showed a tendency of increasing at 3 days after the operation. At 5 to 10 days, the micrangium density of the flap with SVT was much higher than the flap without SVT. There was significant difference between the two groups, which was proved by the optical microscopy observation.

CONCLUSIONSThis study suggests that SVT and its nourishing vessels could promote early revascularization of the flap after transplantation. The SVT can be of benefit to the survival of the reverse-flow random flap.

Animals ; Graft Survival ; Microcirculation ; Reconstructive Surgical Procedures ; methods ; Skin Transplantation ; Surgical Flaps ; blood supply ; Swine ; Swine, Miniature ; Veins ; transplantation

OBJECTIVESTo observe the detrusor ultrastructure in BPH patients and to investigate the relationship between detrusor instability and ultrastructure.

METHODSThe patients were divided into groups of detrusor instability(DI) and detrusor stability(DS) according to urodynamics examination. The structure of the detrusor were observed by light microscopy and transmission electron microscopy(TEM).

RESULTSThe intercellular intermediate junctions and cytoplasmic process junctions in DS were 11.34 +/- 3.23 and 4.26 +/- 1.78 respectively. The intercellular intermediate junctions decreased obviously (3.12 +/- 1.47, P < 0.01) instead of a large amount of cytoplasmic process junctions (26.37 +/- 7.14, P < 0.01) in DI.

CONCLUSIONSThere is a close relevance between intercellular junctions and DI. The observation of the ultrastructure of the detrusor is helpful for the diagnosis of BPH with DI and for the clinical treatment.

Aged ; Aged, 80 and over ; Humans ; Intercellular Junctions ; ultrastructure ; Male ; Microscopy, Electron ; Middle Aged ; Muscle, Smooth ; ultrastructure ; Prostatic Hyperplasia ; pathology ; Urinary Bladder ; ultrastructure

Epilepsy is one of the most common and debilitating neurological diseases that affects more than 40 million people worldwide. Genetic factors contribute to the pathogenesis of epilepsy. Molecular genetic studies have identified 15 disease-causing genes for epilepsy. The majority of the genes encode ion channels, including voltage-gated potassium channels KCNQ2 and KCNQ3, sodium channels SCN1A, SCN2A, and SCN1B, chloride channels CLCN2, and ligand-gated ion channels CHRNA4, CHRNB2, GABRG2, and GABRA1. Interestingly, non-ion channel genes have also been identified as epilepsy genes, and these genes include G-protein-coupled receptor MASS1/VLGR1, GM3 synthase, and proteins with unknown functions such as LGI1, NHLRC1, and EFHC1. These studies make genetic testing possible in some patients, and further characterization of the identified epilepsy genes may lead to the development of new drugs and new treatments for patients with epilepsy.
Chloride Channels ; genetics ; Epilepsies, Myoclonic ; genetics ; Epilepsy ; genetics ; Epilepsy, Absence ; genetics ; Humans ; KCNQ2 Potassium Channel ; genetics ; KCNQ3 Potassium Channel ; genetics ; NAV1.1 Voltage-Gated Sodium Channel ; NAV1.2 Voltage-Gated Sodium Channel ; Nerve Tissue Proteins ; genetics ; Sodium Channels ; genetics

OBJECTIVETo construct the eukaryotic expression plasmid containing mouse vasoactive intestinal peptide(VIP) gene with biological activities.

METHODSVIP cDNA including the sequences of signal peptide was cloned from mouse thymus by RT-PCR, and then inserted into the mammalian expression vector pcDNA3.1 between Hind III and EcoR I restriction sites. COS-7 cells were transfected with pcDNA3. 1-VIP using liposome, the expression of VIP was identified by Western blot and ELISA. Supernatant of transfected cell culture was added to LPS-stimulated macrophages and the TNF-alpha production in cell medium was observed by ELISA.

RESULTSThe cloned VIP cDNA was confirmed by enzyme digestion and DNA sequencing. The expression of VIP was detected in the pcDNA3. 1-VIP transfected COS-7 cells by Western blot and ELISA. The VIP in culture supernatant potently inhibited TNF-alpha production by LPS-induced Macrophages in vitro.

CONCLUSIONThe eukaryotic expression plasmid that expresses biological active murine VIP has been constructed successfully.

Animals ; Base Sequence ; COS Cells ; Cercopithecus aethiops ; Cloning, Molecular ; DNA, Complementary ; genetics ; Eukaryotic Cells ; metabolism ; Mice ; Molecular Sequence Data ; Plasmids ; Recombinant Proteins ; biosynthesis ; genetics ; Recombination, Genetic ; Reverse Transcriptase Polymerase Chain Reaction ; Vasoactive Intestinal Peptide ; biosynthesis ; genetics

A rapid and sensitive liquid chromatography-electrospray ionization-mass spectrometry (LC-ESI/ MS) method for quantification of pentoxyverine citrate in human plasma has been developed and applied for the bioequivalence and pharmacokinetics study. After extracted from plasma samples with ethyl acetate, analysis was performed in selected ion monitoring (SIM) mode with a positive electrospray ionization (ESI) interface with a mobile phase consisted of methanol and water (0.4% glacial acetic acid and 4 mmol x L(-1) ammonium acetate, 43 : 57, v/v). The linear concentration range of the calibration curves was 1.0-160.0 ng x mL(-1) for pentoxyverine citrate, inter- and intra-precision (RSD) was less than 12.5%, accuracy (RE) was in +/- 13.5% and absolute recovery was more than 80%. The method was proved simple, rapid, sensitive, specific and suitable for pharmacokinetic and bioequivalence study of Yufenweilin capsule containing pentoxyverine citrate.
Chromatography, Liquid ; methods ; Cyclopentanes ; blood ; pharmacokinetics ; Humans ; Male ; Sensitivity and Specificity ; Spectrometry, Mass, Electrospray Ionization ; methods ; Therapeutic Equivalency

OBJECTIVETo investegate the role of calcineurin (CaN) and its downstream nuclear factor of activated T-cells (NFATc3) in abdominal aorta restenosis following balloon dilatation in rats.

METHODSSD rats were randomly divided into sham-operated group, balloon group and cyclosporine A (CsA) group. The rats in the latter two groups were subjected to abdominal aorta injury with balloon dilatation, and those in CsA group were treated with CsA at the daily dose of 12.5 mg/kg from 3 days before the surgery to the end of the experiment. Thirty days afer the injury, histological analysis of the arterial wall was carried out with HE staining and immunohistochemistry. The expressions of CaN and NFATc3 in the abdominal aortas were detected with rea1-time PCR, and serum concentration of MCP-1 was determined using enzyme-linked immunosorbent assay.

RESULTSIntimal hyperplasia with irregular thickness of the neointima was observed in the aorta of rats with ballon injury. In rats with CsA treatment, the area of the intimal layers and the ratio of the intimal to the medial layers were obviously lower than those in the balloon injury group (P<0.05). Compared to those in the sham-operated group, the expressions of calcineurin protein and mRNA and NFATc3 mRNA in the arterial wall and serum level of MCP-1 increased significantly in the ballon injury group (P<0.05). CsA treatment significantly suppressed aorta restenosis and the alterations of CaN, NFATc3 and serum MCP-1 induced by ballon dilatation (P<0.05).

CONCLUSIONSCaN-NFATc3 signal transduction pathway mediates restenosis of rat abdominal aorta following ballon dilatation, and CsA can attenuate the restenosis by suppressing this pathway.