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L-精氨酸对猪胎儿皮肤成纤维细胞免疫应激参数和β防御素基因mRNA表达的影响

DOI: 10.3969/j.issn.1006-267x.2013.05.019

Keywords: L-精氨酸,脂多糖,免疫应激,β防御素,基因表达

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Abstract:

本试验旨在研究L-精氨酸(L-Arg)对猪胎儿皮肤成纤维细胞(PFSF)免疫应激参数和β防御素基因mRNA表达的影响。采用培养至4~5代的PFSF,通过脂多糖诱导建立免疫应激模型,在DMEM/F12培养液中添加不同浓度(0、35、70、140、280和560μg/mL)的L-Arg,培养24h后,检测培养液一氧化氮(NO)含量及乳酸脱氢酶(LDH)、一氧化氮合酶(NOS)和溶菌酶(LSZ)活性;采用实时定量PCR法检测PFSFβ防御素1(pBD-1)、β防御素2(pBD-2)和β防御素3(pBD-3)基因mRNA表达量。结果表明:1)L-Arg能够降低培养液LDH活性,正常细胞和应激细胞分别在L-Arg浓度为70和140μg/mL时达到最低;2)L-Arg能够极显著提高培养液NO含量(P<0.01),正常细胞和应激细胞均在L-Arg浓度为140μg/mL达到最高;3)L-Arg能够显著或极显著增加正常细胞培养液NOS活性(P<0.05或P<0.01),而对应激细胞无显著影响(P>0.05),L-Arg浓度为35和70μg/mL时,应激细胞显著或极显著高于正常细胞(P<0.05或P<0.01);4)560μg/mLL-Arg可显著提高培养液LSZ活性(P<0.05);5)70μg/mLL-Arg可极显著提高正常细胞和应激细胞pBD-1和pBD-2基因mRNA表达量(P<0.01),140μg/mLL-Arg可极显著提高正常细胞和应激细胞pBD-3基因mRNA表达量(P<0.01)。结果提示,适宜浓度的L-Arg能够降低培养液LDH活性,提高NO含量以及NOS和LSZ活性,上调应激细胞和正常细胞β防御素基因mRNA表达;本试验条件下,L-Arg的适宜添加浓度为70~140μg/mL。

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