RESUMO
BACKGROUND: Many studies have proven the beneficial effects of regular exercise on psychiatric conditions. This study was set to explore the therapeutic effects and the mechanisms of treadmill exercise on a time-dependent sensitization (TDS) model which is a classical animal model for mimicking posttraumatic stress disorder (PTSD). METHODS: Forty-seven rats were randomly assigned to one of four groups: CON (control), TDS (model), EX (treadmill), or SER (sertraline). TDS model was developed to evaluate the anti-PTSD-like effects of moderate treadmill exercise with 4-week running program. High-performance liquid chromatography technology was used to determine the levels of monoamine neurotransmitters in TDS rats. The expression of key proteins in BDNF/PI3K/Akt/CREB signaling pathway were assayed by the Western blot method. RESULTS: The TDS procedures induced behavioral deficiencies. These deficiencies were reversed by treadmill exercise. Subsequent monoamine assays revealed that the treadmill exercise significantly increased serotonin levels in the hippocampus and decreased dopamine levels in the prefrontal cortex. Data from Western blot experiment demonstrated that exercise could normalize the decreased BDNF/TrkB/pAkt/pCREB levels in the hippocampus. CONCLUSIONS: This study deduced that treadmill exercise ameliorated contextual fear conditioning and anxiety-like behavior in TDS model. According to the study, the mechanism involved in alleviating PTSD symptoms by treadmill exercise was due to increased 5-HT levels in the hippocampus and decreased DA levels in the prefrontal cortex. It also involved the upregulation of BDNF and the related PI3K/AKT/CREB signaling pathway.
Assuntos
Terapia por Exercício , Transtornos de Estresse Pós-Traumáticos/psicologia , Transtornos de Estresse Pós-Traumáticos/terapia , Animais , Ansiedade , Fator Neurotrófico Derivado do Encéfalo/genética , Fator Neurotrófico Derivado do Encéfalo/metabolismo , Modelos Animais de Doenças , Medo , Hipocampo/metabolismo , Masculino , Neurotransmissores/metabolismo , Fosfatidilinositol 3-Quinases/genética , Fosfatidilinositol 3-Quinases/metabolismo , Condicionamento Físico Animal , Ratos , Ratos Sprague-Dawley , Serotonina/metabolismo , Transdução de Sinais , Transtornos de Estresse Pós-Traumáticos/metabolismoRESUMO
<p><b>OBJECTIVE</b>To construct, express and purify human fusion proteins composed of a single-chain antibody fragment scFv that recognizes the prostate specific membrane antigen (PSMA) protein, Fdt, HA2 and tp, and to analyze the binding activity of the expressed fusion proteins.</p><p><b>METHODS</b>The fusion protein genes scFv, scFv-tp, and scFv-Fdt-HA2-tp were amplified by PCR, and the genes obtained were then cloned into the expression vector pET28 and expressed in E. coli BL21. The expressed products were identified by SDS-PAGE and Western blot and purified with Ni(2+)-NTA chelating agarose. The antigen-binding activity of the fusion proteins was determined by ELISA.</p><p><b>RESULTS</b>The human anti-PSMA fusion gene was successfully constructed and expressed in M15 as the inclusion body after induced with IPTG. All the target proteins expressed could bind the PSMA antigen.</p><p><b>CONCLUSION</b>Fusion proteins can specifically bind the PSMA antigen. This finding contributes to the study of the targeted delivery of siRNA.</p>
Assuntos
Humanos , Masculino , Antígenos de Superfície , Alergia e Imunologia , Clonagem Molecular , Eletroforese em Gel de Poliacrilamida , Escherichia coli , Genética , Alergia e Imunologia , Glutamato Carboxipeptidase II , Alergia e Imunologia , Reação em Cadeia da Polimerase , RNA Interferente Pequeno , Alergia e Imunologia , Proteínas Recombinantes de Fusão , Genética , Alergia e Imunologia , Anticorpos de Cadeia Única , Genética , Alergia e ImunologiaRESUMO
Objective: To construct, express and purify ScFvl4/EGFP fusion proteins which containing Arg9, and to study their binding activities and internalization functions. Methods: Arg9 gene was recombined into 5' terminal, 3' terminal of ScFv/EGFP gene and between them respectively before they were cloned into the expression vector pET32a. After induced in E. coli BL21 and purified, their binding activities and internalization were respectively analyzed by indirect ELISA and indirect immunofluorescence analysis. Results: DNA sequencing and restriction endonuclease digestion proved that the four fusion genes were correctly constructed. SDS-PAGE analysis and Western blot showed that they were successfully expressed and purified. Indirect ELISA confirmed that the expressed products had antigen specific binding activities. Indirect immunofluorescence analysis revealed the fusion protein containing Arg9 at its N terminal had much better internalization function, but never internalized into the cells which do not express HBsAg. Conclusion: The four fusion genes were constructed, expressed and purified successfully. The purified fusion proteins maintained the binding activities to HBsAg and the fusion protein containing Arg9 at its N terminal had much better internalization effect.
