Altered glutamate metabolism contributes to antiepileptogenic effects in the progression from focal seizure to generalized seizure by low-frequency stimulation in the ventral hippocampus.

As a promising method for treating intractable epilepsy, the inhibitory effect of low-frequency stimulation (LFS) is well known, although its mechanisms remain unclear. Excessive levels of cerebral glutamate are considered a crucial factor for epilepsy. Therefore, we designed experiments to investigate the crucial parts of the glutamate cycle.

A sub-threshold dose of pilocarpine increases glutamine synthetase in reactive astrocytes and enhances the progression of amygdaloid-kindling epilepsy in rats.

The prognosis of patients exposed to a sub-threshold dose of a proconvulsant is difficult to establish. In this study, we investigated the effect of a single sub-threshold dose of the proconvulsant pilocarpine (PILO) on the progression of seizures that were subsequently induced by daily electrical stimulation (kindling) of the amygdaloid formation. Male Sprague–Dawley rats were each implanted with an electrode in the right basolateral amygdala and an indwelling cannula in the right ventricle.

Simultaneous Determination of Glutamate, Glycine, and Alanine in Human Plasma Using Precolumn Derivatization with 6-Aminoquinolyl-N-hydroxysuccinimidyl Carbamate and High-Performance Liquid Chromatography.

A simple, sensitive and reproducible high-performance liquid chromatography (HPLC) method has been validated for determining concentrations of glutamate, glycine, and alanine in human plasma. Proteins in plasma were precipitated with perchloric acid, followed by derivatization with 6-aminoquinolyl-N-hydroxysuccinimidyl carbamate (AQC). Simultaneous analysis of glutamate, glycine, and alanine is achieved using reversed-phase HPLC conditions and ultraviolet detection.

Experimental investigation of HGF inhibiting glial scar in vitro.

To study the inhibitory effect of Hepatocyte growth factor (HGF) on the responsive hyperplasia of damaged astrocytes in vitro. We prepared damaged model of astrocytes to simulate the responsive hyperplasia of damaged astrocytes in vivo by culturing astrocytes in vitro; After the first day of Ad-HGF transfection, astrocytes were scratched, then after the first, the third, and the fifth day of scratch, we detect the expression amount of astrocytes specific glial fibrillary acidic protein (GFAP) and the ratio of S-phase cells with flow cytometry, both of which can reflect the proliferation status of damaged astrocytes; After HGF was added in scratched astrocytes, the activity of SPK and MAPK (P42/44) were detected by autoradiography and immunoblotting test; After adding different concentrations of HGF protein in astrocytes cultured in different serum concentrations and adding diverse concentrations of HGF protein, SPK and SPK inhibitor DMS in scratched astrocytes, we detect cell proliferation with 3H-TDR incorporation. The first day after Ad-HGF transfected astrocytes were scratched, the amount of GFAP secreted by astrocytes were decreased significantly (P < 0.

Human LINE1 endonuclease domain as a putative target of SARS-associated autoantibodies involved in the pathogenesis of severe acute respiratory syndrome.

Background: Severe acute respiratory syndrome (SARS) is a disease with a mortality of 9.56%. Although SARS is etiologically linked to a new coronavirus (SARS-CoV) and functional cell receptor has been identified, the pathogenesis of the virus infection is largely unclear.

[Screening of the genes of hepatitis B virus e antigen interacting proteins].

Objective: To screen and clone the genes in hepatocytes which encode protein that can interact with hepatitis B e antigen(HBeAg) by yeast-two hybridization.

Methods: Recombined HBeAg bait plasmid (pGBKT7-eAg) was transformed into yeast AH l09, followed by mating with yeast Yl87 containing liver cDNA library plasmid in 2 x YPDA medium. Diploid yeast was plated on synthetic dropout nutrient medium (SD/-Trp-Leu-Ade-His) which contains X-a-gal for selecting positive blue clones.

[Detection of autoimmune parameter of SARS patients].

Objective: To explore whether autoimmune phenomena exist in SARS patients, and to seek for unusual autoimmune antibodies in SARS patients.

Methods: Autoantibodies against cell nuclei (ANA), autoantibodies against smooth muscles (SMA), autoantibodies against parietal cells (PCA), autoantibodies against heart cells (HRA) were detected by using immunofluorescence, and autoantibodies against live-kidney microsomes (LKM) and anti-M2 antibodies were detected by ELISA in sera taken from 27 SARS patients and 18 healthy controls. Immunofluorescence was used to localize the targets antigens in slides with biochips of lung (monkey) of SARS associated antibodies.

[Study on binding of HBeAg to CD81].

Aim: To investigate the interaction between HBeAg and CD81.

Methods: The CD81 gene was amplified by RT-PCR from HepG2 cells. The recombinant expression vector pGADT7-CD81 was constructed by routine molecular biological method.

[Construction and expression of DNA-binding domain plasmid with hepatitis B virus e antigen in yeast double hybrid system].

Background: Using hepatitis B virus e antigen (HBeAg) gene to construct the DNA-binding domain vector, which can express HBeAg in yeast cell, and can be used in yeast double hybrid as "bait plasmid" to look for the gene from the cDNA library, which expresses the protein that can interact with HBeAg.

Methods: PCR was performed to amplify the HBeAg gene from a sera of hepatitis B patient. The product of the amplification was inserted into T-vector and was verified by sequencing.

[The expression and activity detection of a variant N protein of SARS-CoV].

Aim: To construct an expression vector pGEX-2T/N, and to express the fusion protein consisting of N protein of SARS-CoV and GST in E.coli.

Methods: The N region gene of SARS-CoV was cloned by RT-PCR.

[The cytokine secretion of peripheral blood mononucleocytes from patients infected with HCV].

Aim: To detect the levels of cytokines secreted by PBMC from patients infected with HCV after culture for 72 hours in-vitro, so as to reflect the immune state of the HCV-infected patients.

Methods: The levels of cytokines in the culture supernatant of PBMCs were detected by ELISA.

Results: (1)Compared with the cytokine level of normal control, the levels of IFN-gamma, TNF-alpha, and IL-10 of HCV patients notably increased while IL-2, IL-4, and IL-12 were not detected in the culture supernatant of PBMCs from both normal control and HCV patients after culture for 72 hours.

[The observation of the lymphocyte phenotype change in different lymphoid tissues of mice after intranasal immunization with bivalent Shigella vaccines].

Aim: To observe the changes of lymphocyte phenotype in different lymphoid tissues of mice at various time after intranasal immunization with bivalent Shigella vaccines.

Methods: BALB/c mice were randomly divided into three groups, 30 mice per group. Mice were intranasally immunized respectively with PBS, FSM-2117 or FS-5416 four times (bacterial number was sequentially 5x10(6), 1x10(7),4x10(7)and 4x10(7)CFU/mouse) with two week intervals.

[Expression and biological activities of TRAIL fusion protein gene].

Aim: To obtain and express the full-length of TRAIL gene in prokaryocytes.

Methods: The full-length of TRAIL gene was amplified from peripheral blood cells(PBCs)by RT-PCR and then cloned into the expression vector pGEX-2T. The TRAIL-GST was expressed in E.

[Cloning and expressing the E2 subunit of pyruvate dehydrogenase complex].

Objectives: To construct the expression vector of the pyruvate dehydrogenase complex E2 subunit gene (PDC-E2).

Methods: The PDC-E2 gene was amplified from human lymphocytes with RT-PCR, and was cloned into pExSecI vector to induce the PDC-E2 expression. The products were identified with western blot and ELISA.