RESUMO
OBJECTIVE: To investigate the regulatory effects of rat mesenchymal stem cell (MSC) on T lymphocyte proliferation by examining the early activated markers such as CD25 and CD69. METHODS: MSC had been isolated and expanded in vitro. Then it was identified by cell morphology, membrane phenotype, and differentiation potential. Nylon wool column was applied to purify T-lymphocytes. MSCs and T-lymphocytes were cultured together and were stimulated by phytohaemagglutinin (PHA), and then the expressions of CD25 and CD69 were assessed. The levels of TGF-beta1 and IL-10 in the supernatants of MSC cultures were detected by using ELISA. RESULTS: (1) The expression of CD25 is suppressed in a dose-dependent manner when the T-lymphocytes are co-cultured with 10,000 MSCs or more, while 100 MSCs have no detectable effect; (2) The suppression of CD25 can be lasted more than 96 hrs; (3) The down regulation of CD25 is mediated by some soluble factors; (4) The reduced expression of CD25 caused by MSC inhibition is not mediated by TGF-beta1 and IL-10. CONCLUSION: MSCs have significant immune regulatory effects on PHA-stimulated T-lymphocyte culture. It might provide a remarkable immune suppression in organ-transplantation to achieve better outcome in the near future.
Assuntos
Subunidade alfa de Receptor de Interleucina-2/biossíntese , Células-Tronco Mesenquimais/citologia , Linfócitos T/metabolismo , Animais , Antígenos CD/biossíntese , Antígenos de Diferenciação de Linfócitos T/biossíntese , Células Cultivadas , Técnicas de Cocultura , Meios de Cultivo Condicionados/química , Citometria de Fluxo , Interleucina-10/análise , Lectinas Tipo C , Ativação Linfocitária/efeitos dos fármacos , Células Progenitoras Mieloides/citologia , Células Progenitoras Mieloides/efeitos dos fármacos , Células Progenitoras Mieloides/metabolismo , Fito-Hemaglutininas/farmacologia , Ratos , Ratos Sprague-Dawley , Ratos Wistar , Linfócitos T/citologia , Linfócitos T/efeitos dos fármacos , Fator de Crescimento Transformador beta1/análiseRESUMO
OBJECTIVE: To investigate the effects of vascular endothelial growth factor (VEGF) antisense oligodeoxynucleotide (ASODN) on the mRNA and protein expression of VEGF, Flt-1, and kinase insert domain containing receptor (KDR) and VEGF excretion in human gallbladder carcinoma cells. METHODS: Human gallbladder carcinoma cells of the line GBC-SD were cultured and transfected with VEGF ASODN and sense oligodeoxynucleotide (SODN) mediated by Oligofectamine. The toxicity of SODN and Oligofectamine to the GBC-SD cells was examined by MTT method. RT-PCR was used to detect the mRNA and expression of VEGF, Flt-1, and KDR, and ELISA was used to detect the protein expression of VEGF. RESULTS: MTT method showed that SODN and Oligofectamine were not toxic to the GBC-SD cells. The mRNA expression levels of VEGF, Flt-1, and KDR of the ASODN and ASODN + Oligofectamine groups were all significantly lower than those of the control group (all P < 0.05), and were the lowest 72 hours after transfection, and then gradually increased. ELISA showed that there were not significant differences in the VEGF protein concentration in the supernatant of the GBC-SD cells among the SODN, SODN + Oligofectamine, and control groups (all P < 0.05), however, the VEGF protein concentration in the supernatant of the GBC-SD cells of the ASDN and ASDN + Oligofectamine groups were significantly lower than that of the control group (both P < 0.05). CONCLUSION: VEGF ASODN inhibits the mRNA and protein expression of VEGF, Flt-1, and KDR and VEGF excretion in human gallbladder carcinoma cells. Oligofectamine strengthens the effect of ASODN.