Anti-senescence effect and molecular mechanism of the major royal jelly proteins on human embryonic lung fibroblast(HFL-I) cell line

蜂王浆主蛋白对人胚肺成纤维细胞HFL-I的抗衰老作用及其作用机制研究（英文）

Royal jelly (R J)from honeybee has been widely used as a health promotion supplement.The major royal jelly proteins (MRJPs)have been identified as the functional component of RJ.However,the question of whether MRJPs have anti-senescence activity for human cells remains.Human embryonic lung fibroblast (HFL-I)cells were cultured in media containing no MRJPs (A),MRJPs at 0.1mg/ml (B),0.2mg/ml (C),or 0.3mg/ml (D),or bovine serum albumin (BSA)at 0.2mg/ml (E).The mean population doubling levels of cells in media B,C,D,and E were increased by 12.4%,31.2%,24.0%,and 10.4%,respectively,compared with that in medium A.The cells in medium C also exhibited the highest relative proliferation activity,the lowest senescence,and the longest telomeres.Moreover, MRJPs up-regulated the expression of superoxide dismutase-1(SOD1)and down-regulated the expression of mammalian target of rapamycin (MTOR),catenin beta like-1(CTNNB1),and tumor protein p53(TP53).Raman spectra analysis showed that there were two unique bands related to DNA synthesis materials,amide carbonyl group vibrations and aromatic hydrogens.These results suggest that MRJPs possess anti-senescence activity for the HFL-I cell line,and provide new knowledge illustrating the molecular mechanism of MRJPs as anti-senescence factors.

目的:探讨不同浓度蜂王浆主蛋白(MRJPs)对人胚肺成纤维细胞HFL-I的抗衰老作用,并对其可能的分子机理进行预测。创新点:本实验采用拉曼光谱法探究添加MRJPs对HFL-I细胞内成分的影响。实验发现添加MRJPs的HFL-I与对照组的HFL-I的拉曼光谱图中,与DNA和蛋白质合成相关的拉曼峰发生了变化。该发现在未来可作为鉴定衰老HFL-I的一种手段。方法:实验以蜂王浆为原料,利用透析法得到MRJPs。通过对不同浓度MRJPs处理的HFL-I进行细胞形态学观察,测定HFL-I生命周期以及检测其β-半乳糖苷酶活性,评价了MRJPs的抗衰老效果。并且通过MTT法考察了不同浓度MRJPs对HFL-I的促增殖作用。同时,实验一方面采用实时定量聚合酶链反应(q PCR)测量了不同浓度MRJPs处理的HFL-I的细胞端粒相对长度以及MTOR、SOD1、CTNNB1、TP53四种衰老相关基因的相对表达量;另一方面还通过拉曼光谱实验法,进一步探索了MRJPs抗衰老的分子机理。结论:本实验结果显示,添加MRJPs能降低HFL-I衰老细胞所占比例,显著延长HFL-I的生命周期,一定量的MRJPs还能够促进HFL-I的增殖。此外,MRJPs还能减缓细胞端粒长度的缩短,上调衰老相关基因SOD1的表达,下调抗衰老基因MTOR、CTNNB1和TP53的表达。通过MRJPs添加组细胞的拉曼光谱上出现的独特的碱基、酰胺羰基和芳香氢峰,推测MRJPs的抗衰老作用可能与促进DNA及蛋白质在细胞中的合成密切相关。