何首乌对衰老小鼠脑组织凋亡相关基因表达的影响
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摘要
目的应用转录组测序技术观察何首乌提取液对衰老小鼠关凋亡基因表达的影响,了解何首乌延缓衰老的机制。方法首先腹腔注射D-半乳糖制备C57BL/6小鼠衰老模型,然后把衰老小鼠随机分为衰老组(MC)、何首乌提取液作用高剂量组(MH)和何首乌作用低剂量组(ML),每组10只,MH灌胃何首乌提取液1 g/mL/kg,ML灌胃何首乌提取液0.3 g/mL/kg,MC灌胃等量生理盐水,每天1次,连续45 d。实验末摘除眼球取血,检测血清中的丙二醛(MDA)含量、超氧化物歧化酶(SOD)和谷胱甘肽过氧化物酶(GSH-Px)的活力来判断衰老模型和何首乌延缓衰老小鼠的作用;安乐处死小鼠,取脑组织,采用RNA-seq技术对MC和MH进行转录组测序,通过文献和数据库查找凋亡相关基因(共744个),分析相关基因的差异表达。实时荧光定量PCR(Q-PCR)验证相关凋亡基因的表达。结果相对于衰老组,对照组和何首乌提取液作用高剂量组中的SOD、GSH-Px明显升高(P<0.05),MDA明显下降(P<0.05)。转录组测序方法发现了12个差异较大基因,其中Cdh1、Ido1、Pou4f1、Foxb1、Lhx3等基因转录变化在何首乌作用后尤为显著。相关基因表达的Q-PCR验证与RNA-seq检测结果趋势一致。结论何首乌能改善因衰老引起的酶活性降低;并能延缓D-半乳糖引起的小鼠衰老,其机制可能是与凋亡相关基因的表达有关。
Objective To observe the effects of Polygonum Multiflorum Thunb(PMT) extract on the expression of apoptosis genes in senescent mice using transcription group sequencing technique,and further understand the mechanisms of delaying senescence in PMT.Methods First,the aging model of C57 BL/6 mice was prepared by intraperitoneal injection of D-galactose,and then the aging mice were randomly divided into senescence Group(MC),PMT extract solution high Dose Group(MH) and PMT extract solution Low dose Group(ML) with 10 mice in each group.The MH was daily intragastrically given PMT(1 g/ml/kg),and the ML was intragastric administration(0.3 g/ml/kg) and the MC receiced intragastric administration of equivalent saline for 45 d.The effects of Malondialdehyde(MDA) content,superoxide dismutase(SOD) and glutathione peroxidase(GSH-Px) in serum were detected at the end of the experiment to determine the role of PMT in delaying aging mice.Mice were sacrificed in euthanasia and the brain tissue was removed.Transcriptome sequencing was performed using RNA-seq technology.The apoptosis related genes were searched through literature and database(total 744),and the differential expression of related genes was analyzed.Real-time fluorescence quantitative PCR(Q-PCR) was used to verifies the expressions of related apoptotic genes.Results Compared with the senescence group,the increased SOD and GSH-Px levels and the decreased MDA content were indicated in the control group and the high dose group of extract solution of PMT.In addition,twelve genes were expressed differently,among which the transcriptional changes of Cdh1,Ido1,Pou4f1,Foxb1 and Lhx3 were particularly significant after PMT administration.The Q-PCR verification of related gene expression was consistent with the trend of RNA-seq test results.Conclusion PMT could delay the aging of mice caused by D-galactose,and its mechanism might be related to regulating the expressions of apoptosis-related genes.
Objective To observe the effects of Polygonum Multiflorum Thunb(PMT) extract on the expression of apoptosis genes in senescent mice using transcription group sequencing technique,and further understand the mechanisms of delaying senescence in PMT.Methods First,the aging model of C57 BL/6 mice was prepared by intraperitoneal injection of D-galactose,and then the aging mice were randomly divided into senescence Group(MC),PMT extract solution high Dose Group(MH) and PMT extract solution Low dose Group(ML) with 10 mice in each group.The MH was daily intragastrically given PMT(1 g/ml/kg),and the ML was intragastric administration(0.3 g/ml/kg) and the MC receiced intragastric administration of equivalent saline for 45 d.The effects of Malondialdehyde(MDA) content,superoxide dismutase(SOD) and glutathione peroxidase(GSH-Px) in serum were detected at the end of the experiment to determine the role of PMT in delaying aging mice.Mice were sacrificed in euthanasia and the brain tissue was removed.Transcriptome sequencing was performed using RNA-seq technology.The apoptosis related genes were searched through literature and database(total 744),and the differential expression of related genes was analyzed.Real-time fluorescence quantitative PCR(Q-PCR) was used to verifies the expressions of related apoptotic genes.Results Compared with the senescence group,the increased SOD and GSH-Px levels and the decreased MDA content were indicated in the control group and the high dose group of extract solution of PMT.In addition,twelve genes were expressed differently,among which the transcriptional changes of Cdh1,Ido1,Pou4f1,Foxb1 and Lhx3 were particularly significant after PMT administration.The Q-PCR verification of related gene expression was consistent with the trend of RNA-seq test results.Conclusion PMT could delay the aging of mice caused by D-galactose,and its mechanism might be related to regulating the expressions of apoptosis-related genes.
引文
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[18]陈玺龙,陈晓春,李永民.和营安心方对衰老大鼠血清和脑组织白细胞介素-6和肿瘤坏死因子-α的影响[J].河北北方学院学报:自然科学版,2016,32(12):1-3.
[19]胡兵,安红梅,沈克平.细胞衰老与肿瘤发生[J].生命科学,2008,20(3):447-449.
[20]García-Ayuso D,Di P J,Esquiva G,et al.Inherited photoreceptor degeneration causes the death of melanopsin-positive retinal ganglion cells and increases their coexpression of Brn3a[J].Investigative Ophthalmology & Visual Science,2015,56(8):4592-4604.
[21]王立敏,李桂玲.LIM同源结构域转录因子LHX3与LHX4的研究进展[J].国际儿科学杂志,2013,40(5):518-521.
[22]葛学铭,廖杰,廖维靖,等.同源盒基因Lhx3和Lhx4在成年大鼠缺血性脑损伤后的表达变化[J].解剖学报,2004,35(1):1-4.
[2] Fusco D,Colloca G,Lo Monaco M R,et al.Effects of antioxidant supplementation on the aging process[J].Clinical Interventions in Aging,2007,2(3):377-387.
[3] Keller J N.Age-related neuropathology,cognitive decline,and Alzheimer's disease[J].Ageing Res Rev,2006,5(1):1-13.
[4] 江振华,马赛,陈江炜,等.白藜芦醇通过抑制细胞凋亡延缓内皮细胞衰老[J].心脏杂志,2016,28(6):638-641.
[5] Ling S,Xu J W.Biological activities of 2,3,5,4′-tetrahydroxystilbene-2-O-β-D-Glucoside in antiaging and antiaging-related disease treatments[J].Oxidative Medicine and Cellular Longevity,2016,2016(1):1-14.
[6] 杨红莉,葛珍珍,孙震晓.何首乌药理研究新进展[J].中药材,2013,36(10):1713-1717.
[7] Zhang P X,Tang X L,Piao J H,et al.Effect of polygonum multiflorumthunb on antiaging and enhancing cognition function in D-gal-induced-rats[J].Chinese Journal of Rehabilitation Medicine,2005,20(4):251-253.
[8] Ryu G,Ju J H,Yong J P,et al.The radical scavenging effects of stilbeneglucosides from Polygonummultiflorum[J].Archives of Pharmacal Research,2002,25(5):636-639.
[9] 陈泰华,黄雪涛,王俊山,等.何首乌苷对奥沙利铂诱导大鼠星形胶质细胞凋亡的影响[J].中华临床医师杂志:电子版,2015,9(12):2349-2354.
[10] 高宁,刘博,杨德强,等.转录组学在中药研究中的应用现状[J].化学工程师,2017,31(6):50-53.
[11]陈惠,辛丽丽,龚婕宁.基于全转录组测序技术的转录组学在中医药领域的应用前景分析[J].环球中医药,2013,6(10):759-763.
[12]Shen Y,Zhang H ,Cheng L ,et al.In vitro and in vivo antioxidant activity of polyphenols extracted from black highland barley[J].Food Chemistry,2016,194:1003-1012.
[13]Barrera G,Pizzimenti S,Daga M,et al.Lipid peroxidation-derived aldehydes,4-Hydroxynonenal and malondialdehyde in aging-related disorders[J].Antioxidants (Basel,Switzerland),2018,7(8):102.
[14]Arenas-Rios E,Rosado Garcia A,Cortes-Barberena E,et al.Reactive oxygen species production and antioxidant enzyme activity during epididymal sperm maturation in Corynorhinusmexicanus bats[J].Reprod Biol,2016,16(1):78-86.
[15]Lei L,Ou L,Yu X.The antioxidant effect of Asparagus cochinchinensis (Lour.) Merr.shoot in d-galactose induced mice aging model and in vitro[J].Journal of the Chinese Medical Association,2016,79(4):205-211.
[16]Pan L,Chen S,Weng C,et al.Stem cell aging is controlled both intrinsically and extrinsically in the drosophila ovary[J].Cell Stem Cell,2007,1(4):458-469.
[17]Schr?cksnadel K,Wirleitner B,Winkler C,et al.Monitoring tryptophan metabolism in chronic immune activation[J].Clinica Chimica Acta,2006,364(1):82-90.
[18]陈玺龙,陈晓春,李永民.和营安心方对衰老大鼠血清和脑组织白细胞介素-6和肿瘤坏死因子-α的影响[J].河北北方学院学报:自然科学版,2016,32(12):1-3.
[19]胡兵,安红梅,沈克平.细胞衰老与肿瘤发生[J].生命科学,2008,20(3):447-449.
[20]García-Ayuso D,Di P J,Esquiva G,et al.Inherited photoreceptor degeneration causes the death of melanopsin-positive retinal ganglion cells and increases their coexpression of Brn3a[J].Investigative Ophthalmology & Visual Science,2015,56(8):4592-4604.
[21]王立敏,李桂玲.LIM同源结构域转录因子LHX3与LHX4的研究进展[J].国际儿科学杂志,2013,40(5):518-521.
[22]葛学铭,廖杰,廖维靖,等.同源盒基因Lhx3和Lhx4在成年大鼠缺血性脑损伤后的表达变化[J].解剖学报,2004,35(1):1-4.