Biomedical subjects

Lijia An

Publications and source records attributed to Lijia An.

31 records · Page 2Linked to original sources

PubMed2003

Neuroprotective effect of Alpinia oxyphylla Miq. fruits against glutamate-induced apoptosis in cortical neurons.

The protective effect of ethanol extract from the fruits of Alpinia oxyphylla on glutamate-induced neuronal apoptosis was examined in primary cultured mouse cortical neurons. After exposure of cortical neurons to 30 microM glutamate for 24 h, cortical neurons exhibited remarkable apoptotic-like death as evidenced by multi-indices including morphological features, cell viability assay, DNA fragmentation on agarose gel and flow cytometric analysis. Co-treatment of the neurons with A. oxyphylla fruits extract (AFEx) (80-200 microg/ml) in the presence of glutamate significantly elevated cell viability, reduced the number of apoptotic cells and decreased the intensity of glutamate-induced DNA fragmentation. These results suggest the neuroprotective potential of A. oxyphylla fruits against glutamate-induced neuronal apoptosis.

Alpinia↗

PubMed2003

The expression of GFP under the control of fibroin promotor in primary ovarian cells of Antheraea pernyi.

The fibroin promoter can stably express foreign gene in lepidopteran cells. Total RNA was extracted from the gland of silkworm, Antheraea pernyi and the transcription initiation site of fibroin gene of A. pernyi was identified by RNA ligase mediated rapid amplification of cDNA ends (RLM-RACE). The expression vector (pGFP-N2/Fib) was constructed by use of replacing the CMV promoter with the fibroin promoter. The results of visual screening under a fluorescent inverted microscope and Western blot analysis indicated that the GFP gene was expressed in the primary cells of ovary origins from A. pernyi.

Animals↗

PubMed2003

Expression of gloshedobin, a thrombin-like enzyme from the venom of Gloydius shedaoensis, in Escherichia coli.

Gloshedobin, a thrombin-like enzyme from the venom of Gloydius shedaoensis, was expressed in Escherichia coli using expression vector pET-32a(+). The gene was expressed under T7 promotor with a fusion partner of Thx.Tag and a 6xHis.Tag at its 5' terminal. After induction by IPTG for 6 h, the recombinant enzyme was expressed in the cytoplasm. Expression at 25 degrees C gave twice the amount of recombinant gloshedobin in cytoplasm than at 37 degrees C.

Animals↗

PubMed2003

High-level expression of a soluble snake venom enzyme, gloshedobin, in E. coli in the presence of metal ions.

The mature gene of gloshedobin, a snake venom thrombin-like enzyme from the snake, Gloydius shedaoensis, was cloned and expressed in strain E. coli BL21 (DE3). Having been induced by IPTG, the recombinant gloshedobin was in both soluble and insoluble forms. To avoid inclusion body formation, expression was optimized at 25 degrees C. Furthermore, a 50% increase in solubilization of the target protein was obtained by adding 0.1 mM Mg2+ to the medium. The purified recombinant gloshedobin gave a 44 kDa band on SDS-PAGE gel.

Animals↗

PubMed2003

Purification and characterization of Ulva pertusa Kjellm alkaline phosphatase.

The activity of alkaline phosphatase (ALP, EC 3.1.3.1.) was found in seaweeds, including five kinds of green alga, eighteen kinds of red alga, and six kinds of brown alga, collected from the seaside of Dalian in China. The enzyme was purified 1230-fold from Ulva pertusa Kjellm. It had a specific activity of 48.6 U/mg protein and was proven to be homogeneous by SDS-PAGE with a subunit molecular mass of 19.5 kDa. The activity of ALP peaked at pH9.8, and was completely inhibited by DTT and partly by NBS. The Michaelis-Menten constant Km and the maximum reaction velocity Vmax, at pH 9.8 and 37 degrees C were 0.950 mM and 5.00 microM/min, respectively.

Alkaline Phosphatase↗

PubMed2003

Construction of targeting vector and expression of green fluorescent protein in the silkworm, Antheraea pernyi.

The transcription initiation site of Anrheraea pernyi was identified, and a gene targeting vector was constructed. The green fluorescent protein (GFP) gene was inserted into this vector under the control of a fibroin promoter. This recombinant vector was used to target the GFP gene to the fibroin region of the silkworm genome. The DNA was injected into the testes during the pupae, and was also transferred into the egg via the sperm during fertilization. The analysis showed that the GFP gene was integrated into the fibroin gene on the genome by homologous recombination, and was expressed in the silk gland.

Animals↗

PubMed2003

Dextromethorphan protects dopaminergic neurons against inflammation-mediated degeneration through inhibition of microglial activation.

Inflammation in the brain has increasingly been recognized to play an important role in the pathogenesis of several neurodegenerative disorders, including Parkinson's disease and Alzheimer's disease. Inflammation-mediated neurodegeneration involves activation of the brain's resident immune cells, the microglia, which produce proinflammatory and neurotoxic factors, including cytokines, reactive oxygen intermediates, nitric oxide, and eicosanoids that impact on neurons to induce neurodegeneration. Hence, identification of compounds that prevent microglial activation may be highly desirable in the search for therapeutic agents for inflammation-mediated neurodegenerative diseases. In this study, we report that dextromethorphan (DM), an ingredient widely used in antitussive remedies, reduced the inflammation-mediated degeneration of dopaminergic neurons through inhibition of microglial activation. Pretreatment (30 min) of rat mesencephalic neuron-glia cultures with DM (1-10 micro M) reduced, in a dose-dependent manner, the microglia-mediated degeneration of dopaminergic neurons induced by lipopolysaccharide (LPS, 10 ng/ml). Significant neuroprotection by DM was also evident when DM was applied to cultures up to 60 min after the addition of LPS. The neuroprotective effect of DM was attributed to inhibition of LPS-stimulated microglial activation because DM significantly inhibited the LPS-induced production of tumor necrosis factor-alpha, nitric oxide, and superoxide free radicals. This conclusion was further supported by the finding that DM failed to prevent 1-methyl-4-phenylpyridinium- or beta-amyloid peptide (1-42)-induced dopaminergic neurotoxicity in neuron-enriched cultures. In addition, because LPS did not produce any significant increase in the release of excitatory amino acids from neuron-glia cultures and N-methyl-D-aspartate antagonist dizocilpine maleate failed to afford significant neuroprotection, it is unlikely that the neuroprotective effect of DM is mediated through N-methyl-D-aspartate receptors. These results suggest that DM may be a promising therapeutic agent for the treatment of Parkinson's disease.

1-Methyl-4-phenylpyridinium↗

PubMed2003

[Study on the expression of telomerase RNA in leukocyte].

Total cDNA of human telomerase RNA(hTR) gene was cloned by means of RT-PCR and inverted into retroviral vector (pLNCX) to construct the mammalian cell expression plasmid. Then, by using lipofectin-mediated DNA transfection, the obtained expression plasmid was successfully transfected into human normal peripheral blood leukocyte. All data suggested that expression of transfected exogenous hTR gene can not reconstitute telomerase activity. Flow cytometry analysis and data from cell growth curve also indicated that expression of exogenous gene can not prolong the longevity of leukocyte, but rather inhibit the growth of leukocyte and induce its apoptosis. We conclude that expression of exogenous gene may block the coalition of telomerase RNA and its catalytic subunit(hTRT) and block the coalition of telomerase RNA template and telomere DNA, thus affecting telomerase activity and repressing cell proliferation.

Cells, Cultured↗

PubMed2003

Characterization of Sporothrix schenckii by random amplification of polymorphic DNA assay.

OBJECTIVES: To investigate the DNA polymorphism of Sporothrix schenckii (S. schenckii) and to find the relationship between DNA patterns and geographic areas and clinical manifestations. METHOD: The total DNA was extracted with hexadecyltrimethyl-ammonium bromide. Random Amplification of Polymorphic DNA (RAPD) assay was used to study DNA typing of 24 strains of S. schenckii collected from different areas and isolated from different clinical types. RESULTS: Of seven random primers used, three primers (OPAA11, OPD18 and OPB07) gave good reactions, the sequences of which were 5'-ACCCGACCTG-3', 5'-GAGAGCCAAC-3', 5'-GGTGAC~GCAG-3' respectively. The RAPD patterns of the 24 isolates were not completely identical, showing certain degrees of hereditary variability. Different isolates showed a common conserved DNA band with the same primer. Different clinical types showed different genotypes. CONCLUSION: RAPD analysis is useful in DNA typing of S. schenckii, the DNA band type of which is related to geographic origin and Clinical manifestation.

DNA, Fungal↗

PubMed2003

[Alzheimer's disease: advance in researches on Beta-amyloid protein].

The pathogenesis and treatment of Alzheimer's disease (AD) have developed into the frontier with the aging of people in the world. Meanwhile, they are the most difficult steps in the research on this degenerative disease of the nervous system. The over-deposition of beta-amyloid protein in nervous system is the most important feature. The formation and influencing factors of beta-amyloid protein are summarized in this paper. Furthermore, the methods and advance in treatment of AD are reviewed especially.

Aged↗

PubMed2002

A serum- and antioxidant-free primary culture model of mouse cortical neurons for pharmacological screen and studies of neurotrophic and neuroprotective agents.

1. Morphologically developmental properties of fetal mouse cortical neurons in the chemically defined serum- and antioxidant-free culture condition were observed. Also, cellular composition in cultures was identified by immunostaining with anti-NSE and anti-GFAP. 2. Various cell densities ranging from 1 x 10(3) to 1 x 10(6) cells/cm2 were prepared to further assess the effect of cell density on time-course of neuronal survival by counting the number of remaining attached neurons after 3 and 7 days in culture. 3. Neuronal responses to neurotrophic effect of NGF on neurite outgrowth and neuroprotective effect of MK-801 against glutamate-induced excitotoxity were evaluated by image analysis and MTT assay, respectively. 4. Results showed that this culture system was neuronal-enriched with a neuronal lifetime more than 35 days. Neurons survived best when seeded at a density > or =1.5 x 10(5) cells/cm2. Cultured neurons were capable of exhibiting sensitive responses to the effects of NGF and MK-801. 5. These findings suggest that this primary culture system provides a sensitive and powerful in vitro model for pharmacological screen and studies of neurotrophic and neuroprotective agents.

Animals↗

PubMed2002

Inhibition by naloxone stereoisomers of beta-amyloid peptide (1-42)-induced superoxide production in microglia and degeneration of cortical and mesencephalic neurons.

Previously we reported that naloxone stereoisomers, in an opioid receptor-independent manner, attenuated the inflammation-mediated degeneration of dopaminergic neurons by inhibition of the activation of microglia, the resident immune cells in the brain. Recently we discovered that beta-amyloid peptide Abeta (1-42) exhibited enhanced neurotoxicity toward both cortical and mesencephalic neurons through the activation of microglia and production of superoxide. The purpose of this study was to determine whether naloxone isomers had any effect on Abeta (1-42)-induced neurodegeneration. Pretreatment of either cortical or mesencephalic neuron-glia cultures with 1 to 10 microM (-)-naloxone, prior to treatment for up to 11 days with 0.1 to 3 microM Abeta (1-42), afforded significant neuroprotection as judged by neurotransmitter uptake, immunocytochemical analysis, and cell counting. More importantly, (+)-naloxone, the ineffective enantiomer of (-)-naloxone in binding opioid receptors, was equally effective in affording neuroprotection. Mechanistically, inhibition of Abeta (1-42)-induced production of superoxide in microglia underlay the neuroprotective effect of naloxone stereoisomers. Moreover, neuroprotection and inhibition of Abeta (1-42)-induced superoxide production was also achieved with naloxone methiodide, a charged analog with quaternary amine, suggesting that the site of action for naloxone isomers is at the cell surface of microglia. These results demonstrated that naloxone isomers, through mechanisms unrelated to the opioid receptors, were capable of inhibiting Abeta (1-42)-induced microglial activation and degeneration of both cortical and mesencephalic neurons. Combined with our previous observations with inflammagen-induced neurodegeneration, naloxone analogs, especially (+)-naloxone, may have potential therapeutic efficacy for the treatment of Alzheimer's and Parkinson's disease.

Amyloid beta-Peptides↗

PubMed2002

Expression of endoplasmic reticulum molecular chaperon GRP94 in human lung cancer tissues and its clinical significance.

OBJECTIVE: To investigate the relationship between the expression of glucose regulated protein 94 (GRP94) at the level of mRNA and protein in vivo and in human lung cancer. METHODS: RT-PCR, immunohistochemistry and/or Western blot were used in 54 cases of lung cancer tissues and corresponding normal lung tissues. RESULTS: There was a significant overexpression of GRP94 mRNA and protein in lung cancer tissues as compared with lung normal tissues. In lung cancer tissue, the relative level of GRP94 mRNA as evaluated by RT-PCR was 3.48 +/- 2.06, the level of GRP94 protein as evaluated by immunohistochemistry was + + to + + +, and by Western blot was 256.7 +/- 80.6. In lung normal tissue, the relative level of GRP94 mRNA was 2.01 +/- 1.83, the level of GRP94 protein was + to + + and 108.1 +/- 42.3. The differences in expression of GRP94 between the two tissues were significant (P < 0.05). Furthermore, the over-expression of GRP94 in the lung cancer tissues was correlated to grade of differentiation and stage of tumors. There was stronger expression in poor-differentiated tumors than in mild-to-high differentiated tumors (P < 0.05). There was also a stronger expression in stage III than in stage I and II tumors (P < 0.05). No statistically significant difference was found among various pathological types of tumors. CONCLUSION: GRP94 was related with the occurrence, differentiation and progress of human lung cancer. Ascertaining the levels of GRP94 mRNA and protein may be valuable in evaluating the grade of differentiation and clinical stage of human lung cancer.

Adult↗