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To explore the effect of antisense oligonucleotide on the production of IL-5 by mouse spleen T lymphocytes.Methods Based on the IL-5 cDNA sequence of mouse, a segment of antisense oligonucleotide was designed and synthesized. 5’-labeling of antisense oligonucleotide was signed by T_4 PNK in order that the efficiency of stearylamine liposome in transfecting antisense oligonucleotide can be evaluated. Asthma model was duplicated with ovalbumin(OVA) absorbed to aluminum hydroxide. T lymphocytes of mice were separated by nylon fiber method, then T lymphocytes transfected with different concentration of antisense oligonucleotide with cation stearylamine liposme were incubated respectively in order to observe the effect of antisense oligonucleotide on Il-5 production by T lymphocytes. IL-5 levels in the supernatants of T lymphocyte cultures were determined by ELISA.Results Stearylamine liposome could markedly increase the efficiency of antisense oligonucleotide transfection. The transfection efficiency of antisense oligouncleotide increased approximately 12 times at a ratio of 1: 15m/m (antisense oligonucleotide to SA liposome). In healthy and asthma Balb/c mice, IL-5 was not detectable in the supernatants of T lymphocyte cultures without stimulated with OVA; however, IL-5 was increased markedly in the supernatants of T lymphocyte cultures stimulated with OVA. After transfection with different concentrations of antisense oligonucleotide, IL-5 levels in the supernatants of T lymphocyte cultures were significantly lower than those in control cultured without antisense oligonucleotide transfection. IL-5 levels decreased from 44.60±6.23 pg/ml to 30.70±7.362 pg/ml, 17.20±6.181 pg/ml and 8.16±2.34 pg/ml respectively. And IL-5 synthesis was inhibited by 31.17%, 61.43% and 81.7% respectively.Conclusion IL-5 synthesis could be obviously inhibited by antisense oligonucleotide and showed a markedly correlation between dose and effectiveness. It suggests the production of IL-5 is inhibited by preventing the transcription of IL-5 from T lymphocytes. The study provides foundation for antisense gene therapy in asthma.
To explore the effect of antisense oligonucleotide on the production of IL-5 by mouse spleen T lymphocytes. Methods Based on the IL-5 cDNA sequence of mouse, a segment of antisense oligonucleotide was designed and synthesized. 5’-labeling of antisense oligonucleotide was signed by T_4 PNK in order that the efficiency of stearylamine liposome in transfecting antisense oligonucleotide can be evaluated. Asthma model was duplicated with ovalbumin (OVA) absorbed to aluminum hydroxide. T lymphocytes of mice were separated by nylon fiber method, then T lymphocytes transfected with different concentration of antisense oligonucleotide with cation stearylamine liposme were incubated respectively in order to observe the effect of antisense oligonucleotide on Il-5 production by T lymphocytes. IL-5 levels in the supernatants of T lymphocyte cultures were determined by ELISA. Results Stearylamine liposome could markedly increase the efficiency of antisense oligonucleotide transfection. The transfection efficiency of antisense oligouncleotide for about 12 times at a ratio of 1: 15 m / m (Antisense oligonucleotide to SA liposome). In healthy and asthma Balb / c mice, IL-5 was not detectable in the supernatants of T lymphocyte cultures without stimulated with OVA; however, IL-5 was increased markedly in the supernatants of T lymphocyte cultures stimulated with OVA. After transfection with different concentrations of antisense oligonucleotide, IL-5 levels in the supernatants of T lymphocyte cultures were significantly lower than those in control cultured without IL-5 levels decreased from 44.60 ± 6.23 pg / ml to 30.70 ± 7.362 pg / ml, 17.20 ± 6.181 pg / ml and 8.16 ± 2.34 pg / ml respectively. And IL-5 synthesis was inhibited by 31.17% 61.43% and 81.7% respectively.Conclusion IL-5 synthesis could be obviously inhibited by antisense oligonucleotide and showed a markedly correlation between dose and effectiveness. It suggests the production of IL-5 is inhibited by preventing the transcription of IL-5 from T lymphocytes. The study provides foundation for antisense gene therapy in asthma.