人胎盘间充质干细胞移植对化疗所致卵巢早衰大鼠卵巢功能的影响
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摘要
目的研究人胎盘间充质干细胞(h PMSC)移植对环磷酰胺诱导的卵巢早衰(POF)模型大鼠卵巢功能修复过程中沉默信息调节因子1(SIRT1)、过氧化物酶体增殖物活化受体γ共激活因子1-α(PGC1-α)以及核转录因子E2相关因子2(Nrf2)活性的影响。方法将60只雌性SD大鼠随机分为空白对照组、POF模型组、h PMSC治疗组、治疗对照组,每组15只。通过大鼠腹腔注射环磷酰胺建立POF模型,治疗组于造模成功后给予h PMSC治疗。采用ELISA法测定血清中雌二醇(E2)、促卵泡生成素(FSH)、抗苗勒管激素(AMH)水平; HE染色法和Van Gie-son染色观察卵巢组织形态学和纤维化变化,Western blotting和免疫组织化学法检测卵巢组织SIRT1、PGC1-α、Nrf2的表达水平。结果 ELISA结果显示,POF模型组血清FSH水平明显高于空白对照组(P <0. 05),而E2、AMH水平则明显低于空白对照组,差异有统计学意义(P <0. 01);与POF模型组和治疗对照组相比,h PMSC移植治疗后FSH水平下降,AMH水平升高,差异有统计学意义(P <0. 01)。HE染色结果显示,POF模型组卵巢组织结构紊乱,原始卵泡和初级卵泡减少,闭锁卵泡增加,h PMSC治疗组中正常形态卵泡数目增多,闭锁卵泡数目减少。免疫组织化学结果显示,POF模型组SIRT1、PGC1-α、Nrf2蛋白表达相对于空白对照组降低,h PMSC治疗组蛋白表达相对于POF模型组升高。Western blotting结果显示,与空白对照组相比,POF模型组SIRT1、PGC1-α、Nrf2蛋白表达显著降低(P <0. 01);与POF模型组相比,h PMSC治疗组SIRT1、PGC1-α、Nrf2蛋白表达显著升高(P <0. 01)。结论 h PMSC移植对POF模型大鼠有抗氧化与修复卵巢功能的作用,其机制可能与激活SIRT1/PGC1-α/Nrf2信号通路有关。
Objective To investigate the antioxidant potential of human placenta mesenchymal stem cell( hPMSC)transplantation on the silent information regulator transcript-1( SIRT1) and nuclear factor-erythroid 2-related factor 2( Nrf2) and peroxisome proliferator-activated receptor gamma coactivator1-α( PGC1-α) in the treatment of premature ovarian failure( POF) in rats. Methods A total of 60 SD female rats with normal estrous cycle were randomly divided into 4 groups( n = 15) : blank control group,POF model group,hPMSC transplantation group,and treatment control group. The POF models were established by intraperitoneal injection of cyclophosphamide. The serum levels of estradiol( E2),follicle-stimulating hormone( FSH) and anti-müllerian hormone( AMH) were determined with ELISA. The fibrosis and histomorphological features of ovarian tissues were observed with Van Gieson( VG) staining and HE staining.The expressions of SIRT1,Nrf2 and PGC1-α were detected with Western blotting and immunohistochemical staining.Results Serum FSH level was significantly higher in the POF model group than in blank control group( P < 0. 05),while E2 and AMH levels were significantly lower( P < 0. 01). Compared with the treatment control group and POF model group,the hPMSC transplantation group had decreased FSH level and increased AMH level( P < 0. 01). HE staining showed that in the POF model group,the numbers of primordial follicles and primary follicles increased,while the number of atretic follicles decreased; in the hPMSC transplantation group,the number of normal follicles increased,while the number of atretic follicles decreased. Immunohistochemical staining results showed that the protein expressions of SIRT1,Nrf2 and PGC1-α were significantly lower in the POF model group than in the blank control group,and significantly higher in the hPMSC transplantation group than in the POF model group. Western blotting results showed the same tendency of changes( P < 0. 01). Conclusion The hPMSC transplantation may have antioxidant potential and protective effects on ovarian by activating the SIRT1/PGC1-α/Nrf2 signaling pathway.
Objective To investigate the antioxidant potential of human placenta mesenchymal stem cell( hPMSC)transplantation on the silent information regulator transcript-1( SIRT1) and nuclear factor-erythroid 2-related factor 2( Nrf2) and peroxisome proliferator-activated receptor gamma coactivator1-α( PGC1-α) in the treatment of premature ovarian failure( POF) in rats. Methods A total of 60 SD female rats with normal estrous cycle were randomly divided into 4 groups( n = 15) : blank control group,POF model group,hPMSC transplantation group,and treatment control group. The POF models were established by intraperitoneal injection of cyclophosphamide. The serum levels of estradiol( E2),follicle-stimulating hormone( FSH) and anti-müllerian hormone( AMH) were determined with ELISA. The fibrosis and histomorphological features of ovarian tissues were observed with Van Gieson( VG) staining and HE staining.The expressions of SIRT1,Nrf2 and PGC1-α were detected with Western blotting and immunohistochemical staining.Results Serum FSH level was significantly higher in the POF model group than in blank control group( P < 0. 05),while E2 and AMH levels were significantly lower( P < 0. 01). Compared with the treatment control group and POF model group,the hPMSC transplantation group had decreased FSH level and increased AMH level( P < 0. 01). HE staining showed that in the POF model group,the numbers of primordial follicles and primary follicles increased,while the number of atretic follicles decreased; in the hPMSC transplantation group,the number of normal follicles increased,while the number of atretic follicles decreased. Immunohistochemical staining results showed that the protein expressions of SIRT1,Nrf2 and PGC1-α were significantly lower in the POF model group than in the blank control group,and significantly higher in the hPMSC transplantation group than in the POF model group. Western blotting results showed the same tendency of changes( P < 0. 01). Conclusion The hPMSC transplantation may have antioxidant potential and protective effects on ovarian by activating the SIRT1/PGC1-α/Nrf2 signaling pathway.
引文
[1]佟超.线粒体异常与卵巢早衰[J].山东大学学报(医学版),2018,56(4):23-27.TONG Chao. Mitochondrial defects and premature ovarianfailure[J]. Journal of Shandong University(Health Sci-ences),2018,56(4):23-27.
[2]Luo QQ,Yin N,Zhang LS,et al. Role of SDF-1/CXCR4and cytokines in the development of ovary injury in chemo-therapy drug induced premature ovarian failure mice[J].Life Sci,2017,179:103-109. doi:10. 1016/j. lfs. 2017.05. 001.
[3]Sheikhansari G,Aghebati-Maleki L,Nouri M,et al. Currentapproaches for the treatment of premature ovarian failure withstem cell therapy[J]. Biomed Pharmacother,2018,102:254-262. doi:10. 1016/j. biopha. 2018. 03. 056.
[4]相丽,马会明,何艳桃,等.人胎盘间充质干细胞移植通过降低超氧化物歧化酶1和解耦联蛋白-2的表达提高卵巢功能[J].中华生殖与避孕杂志,2018,38(2):101-108.XIANG Li,MA Huiming,HE Yantao,et al. Effects of hu-man placenta mesenchymal stem cell transplantation on su-peroxide dismutase 1 and uncoupling protein-2 in the treat-ment of premature ovarian failure in rats[J]. Reproductionand Contraception,2018,38(2):101-108.
[5]Di Emidio G,Rossi G,Bonomo I,et al. The natural carot-enoid crocetin and the synthetic tellurium compound AS101protect the ovary against cyclophosphamide by modulatingSIRT1 and mitochondrial markers[J]. Oxid Med CellLongev, 2017, 2017:8928604. doi:10. 1155/2017/8928604.
[6] Wang ZH,Yuan S,Li YB,et al. Regulation on SIRT1-PGC-1α/Nrf2 pathway together with selective inhibition ofaldose reductase makes compound hr5F a potential agentfor the treatment of diabetic complications[J]. BiochemPharmacol,2018, 150:54-63. doi:10. 1016/j. bcp.2018. 01. 034.
[7]王永峰,马会明,相丽. BMSCs移植对大鼠卵巢早衰的修复及TGF-βRⅡ表达的影响[J].宁夏医科大学学报,2017,39(6):627-630.WANG Yongfeng,MA Huiming,XIANG Li,et al. Ex-pression of TGF-beta Receptor II in Ovarian Premature Ag-ing Rats with the Treatment of BMSCs Transplantation[J].Journal of Ningxia Medical University,2018,39(6):627-630.
[8]黄平,张坤,李芳,等.脐带和脂肪源性间充质干细胞生物学特性比较[J].山东大学学报(医学版),2018,56(3):72-78.HUANG Ping,ZHANG Kun,LI Fang,et al. Comparativestudy on biological characteristics of mesenchymal stemcells from human umbilical cord and adipose[J]. Journalof Shandong University(Health Sciences),2018,56(3):72-78.
[9]Koippallil Gopalakrishnan AR,Kishore U,Madan T. Mes-enchymal stem cells:a promising tool for targeted genetherapy of endometriosis[J]. Regen Med,2017,12(1):69-76.
[10]Domnina A,Novikova P,Obidina J,et al. Human mes-enchymal stem cells in spheroids improve fertility in modelanimals with damaged endometrium[J]. Stem Cell ResTher,2018,9(1):50. doi:10. 1186/s13287-018-0801-9.
[11] Nie XW,Dai YJ,Zheng Y,et al. Establishment of amouse model of premature ovarian failure using consecu-tive superovulation[J]. Cell Physiol Biochem,2018,51(5):2341-2358.
[12]Liang QX,Wang ZB,Lin F,et al. Ablation of beta sub-unit of protein kinase CK2 in mouse oocytes causes follicleatresia and premature ovarian failure[J]. Cell Death Dis,2018,9(5):508. doi:10. 1038/s41419-018-0505-1.
[13]秦莹莹,张茜蒻.卵巢早衰的免疫学病因及免疫干预研究进展[J].山东大学学报(医学版),2018,56(4):33-37.QIN Yingying,ZHANG Xiruo. Research advances on theimmunological pathogenesis and immunotherapy of prema-ture ovarian failure[J]. Journal of Shandong University(Health Sciences),2018,56(4):33-37.
[14]Kim TH,Choi JH,Jun Y,et al. 3D-cultured human pla-centa-derived mesenchymal stem cell spheroids enhance o-vary function by inducing folliculogenesis[J]. Sci Rep,2018,8(1):15313. doi:10. 1038/s41598-018-33575-9.
[15] Kadam S,Muthyala S,Nair P,et al. Human placenta-derived mesenchymal stem cells and islet-like cell clustersgenerated from these cells as a novel source for stem celltherapy in diabetes[J]. Rev Diabet Stud,2010,7(2):168-182.
[16]Wang WW,Li H,Xu FH,et al. IFN-γup-regulates PD-L1 expression in human placenta mesenchymal stem cellsand enhances cell ability to induce the differentiation ofIL-10+T cells from cord blood-and peripheral blood-de-rived T cells[J]. Xi bao yu fen zi mian yi xue za zhi,2016,32(2):191-195.
[17]Xu K,Guo LQ,Bu HX,et al. Daphnetin inhibits highglucose-induced extracellular matrix accumulation,oxida-tive stress and inflammation in human glomerular mesang-ial cells[J]. J Pharmacol Sci,2018. doi:10. 1016/j.jphs. 2018. 11. 013
[18]王凯民,谭娟娟,严志强,等. Sirtuin在低氧诱导人脐带间充质干细胞增殖中的作用[J].山东大学学报(医学版),2017,55(7):23-30.WANG Kaimin,TAN Juanjuan,YAN Zhiqiang,et al.Effects of Sirtuin on hypoxia-induced proliferation of hu-man umbilical cord-derived mesenchymal stem cells[J].Journal of Shandong University(Health Science),2017,55(7):23-30.
[19]Inflammation MO. Retracted:treadmill training increasesSIRT-1 and PGC-1αprotein levels and AMPK phosphoryl-ation in quadriceps of middle-aged rats in an intensity-de-pendent manner[J]. Mediators Inflamm,2017,2017:8287646. doi:10. 1155/2017/8287646.
[20]崔春英,申超,洪艳,等.柿叶黄酮提取物对氧糖剥夺/复糖复氧损伤的HT22细胞氧化应激的保护作用[J].山东大学学报(医学版),2018,56(6):6-12.CUI Chunying,SHEN Chao,HONG Yan,et al. Protec-tive effect of flavonoid components extracted from DiopyrosKaki on oxidative stress induced by OGD/R in HT22 cells[J]. Journal of Shandong University(Health Sciences),2018,56(6):6-12.
[21]Sova M,Saso L. Design and development of Nrf2 modula-tors for cancer chemoprevention and therapy:a review[J]. Drug Des Devel Ther,2018,12:3181-3197. doi:10. 2147/DDDT. S172612
[22] Yamamoto M,Kensler TW,Motohashi H. The KEAP1-NRF2system:a thiol-based sensor-effector apparatus formaintaining redox homeostasis[J]. Physiol Rev,2018,98(3):1169-1203.
[23]Wang XJ,Chen Q,Zhang ZY,et al. Novel enteromorphaprolifera based carbon dots:probing the radical scaven-ging of natural phenolic compounds[J]. Colloids Surf BBiointerfaces,2018,174:161-167. doi:10. 1016/j. col-surfb. 2018. 11. 019
[24]Corsello T,Komaravelli N,Casola A. Role of hydrogensulfide in NRF2-and sirtuin-dependent maintenance ofcellular redox balance[J]. Antioxidants(Basel),2018,7(10):E129. doi:10. 3390/antiox7100129
[25] Nowak WN,Taha H,Kachamakova-Trojanowska N,etal. Murine bone marrow mesenchymal stromal cells re-spond efficiently to oxidative stress despite the low level ofheme oxygenases 1 and 2[J]. Antioxid Redox Signal,2018,29(2):111-127.
[2]Luo QQ,Yin N,Zhang LS,et al. Role of SDF-1/CXCR4and cytokines in the development of ovary injury in chemo-therapy drug induced premature ovarian failure mice[J].Life Sci,2017,179:103-109. doi:10. 1016/j. lfs. 2017.05. 001.
[3]Sheikhansari G,Aghebati-Maleki L,Nouri M,et al. Currentapproaches for the treatment of premature ovarian failure withstem cell therapy[J]. Biomed Pharmacother,2018,102:254-262. doi:10. 1016/j. biopha. 2018. 03. 056.
[4]相丽,马会明,何艳桃,等.人胎盘间充质干细胞移植通过降低超氧化物歧化酶1和解耦联蛋白-2的表达提高卵巢功能[J].中华生殖与避孕杂志,2018,38(2):101-108.XIANG Li,MA Huiming,HE Yantao,et al. Effects of hu-man placenta mesenchymal stem cell transplantation on su-peroxide dismutase 1 and uncoupling protein-2 in the treat-ment of premature ovarian failure in rats[J]. Reproductionand Contraception,2018,38(2):101-108.
[5]Di Emidio G,Rossi G,Bonomo I,et al. The natural carot-enoid crocetin and the synthetic tellurium compound AS101protect the ovary against cyclophosphamide by modulatingSIRT1 and mitochondrial markers[J]. Oxid Med CellLongev, 2017, 2017:8928604. doi:10. 1155/2017/8928604.
[6] Wang ZH,Yuan S,Li YB,et al. Regulation on SIRT1-PGC-1α/Nrf2 pathway together with selective inhibition ofaldose reductase makes compound hr5F a potential agentfor the treatment of diabetic complications[J]. BiochemPharmacol,2018, 150:54-63. doi:10. 1016/j. bcp.2018. 01. 034.
[7]王永峰,马会明,相丽. BMSCs移植对大鼠卵巢早衰的修复及TGF-βRⅡ表达的影响[J].宁夏医科大学学报,2017,39(6):627-630.WANG Yongfeng,MA Huiming,XIANG Li,et al. Ex-pression of TGF-beta Receptor II in Ovarian Premature Ag-ing Rats with the Treatment of BMSCs Transplantation[J].Journal of Ningxia Medical University,2018,39(6):627-630.
[8]黄平,张坤,李芳,等.脐带和脂肪源性间充质干细胞生物学特性比较[J].山东大学学报(医学版),2018,56(3):72-78.HUANG Ping,ZHANG Kun,LI Fang,et al. Comparativestudy on biological characteristics of mesenchymal stemcells from human umbilical cord and adipose[J]. Journalof Shandong University(Health Sciences),2018,56(3):72-78.
[9]Koippallil Gopalakrishnan AR,Kishore U,Madan T. Mes-enchymal stem cells:a promising tool for targeted genetherapy of endometriosis[J]. Regen Med,2017,12(1):69-76.
[10]Domnina A,Novikova P,Obidina J,et al. Human mes-enchymal stem cells in spheroids improve fertility in modelanimals with damaged endometrium[J]. Stem Cell ResTher,2018,9(1):50. doi:10. 1186/s13287-018-0801-9.
[11] Nie XW,Dai YJ,Zheng Y,et al. Establishment of amouse model of premature ovarian failure using consecu-tive superovulation[J]. Cell Physiol Biochem,2018,51(5):2341-2358.
[12]Liang QX,Wang ZB,Lin F,et al. Ablation of beta sub-unit of protein kinase CK2 in mouse oocytes causes follicleatresia and premature ovarian failure[J]. Cell Death Dis,2018,9(5):508. doi:10. 1038/s41419-018-0505-1.
[13]秦莹莹,张茜蒻.卵巢早衰的免疫学病因及免疫干预研究进展[J].山东大学学报(医学版),2018,56(4):33-37.QIN Yingying,ZHANG Xiruo. Research advances on theimmunological pathogenesis and immunotherapy of prema-ture ovarian failure[J]. Journal of Shandong University(Health Sciences),2018,56(4):33-37.
[14]Kim TH,Choi JH,Jun Y,et al. 3D-cultured human pla-centa-derived mesenchymal stem cell spheroids enhance o-vary function by inducing folliculogenesis[J]. Sci Rep,2018,8(1):15313. doi:10. 1038/s41598-018-33575-9.
[15] Kadam S,Muthyala S,Nair P,et al. Human placenta-derived mesenchymal stem cells and islet-like cell clustersgenerated from these cells as a novel source for stem celltherapy in diabetes[J]. Rev Diabet Stud,2010,7(2):168-182.
[16]Wang WW,Li H,Xu FH,et al. IFN-γup-regulates PD-L1 expression in human placenta mesenchymal stem cellsand enhances cell ability to induce the differentiation ofIL-10+T cells from cord blood-and peripheral blood-de-rived T cells[J]. Xi bao yu fen zi mian yi xue za zhi,2016,32(2):191-195.
[17]Xu K,Guo LQ,Bu HX,et al. Daphnetin inhibits highglucose-induced extracellular matrix accumulation,oxida-tive stress and inflammation in human glomerular mesang-ial cells[J]. J Pharmacol Sci,2018. doi:10. 1016/j.jphs. 2018. 11. 013
[18]王凯民,谭娟娟,严志强,等. Sirtuin在低氧诱导人脐带间充质干细胞增殖中的作用[J].山东大学学报(医学版),2017,55(7):23-30.WANG Kaimin,TAN Juanjuan,YAN Zhiqiang,et al.Effects of Sirtuin on hypoxia-induced proliferation of hu-man umbilical cord-derived mesenchymal stem cells[J].Journal of Shandong University(Health Science),2017,55(7):23-30.
[19]Inflammation MO. Retracted:treadmill training increasesSIRT-1 and PGC-1αprotein levels and AMPK phosphoryl-ation in quadriceps of middle-aged rats in an intensity-de-pendent manner[J]. Mediators Inflamm,2017,2017:8287646. doi:10. 1155/2017/8287646.
[20]崔春英,申超,洪艳,等.柿叶黄酮提取物对氧糖剥夺/复糖复氧损伤的HT22细胞氧化应激的保护作用[J].山东大学学报(医学版),2018,56(6):6-12.CUI Chunying,SHEN Chao,HONG Yan,et al. Protec-tive effect of flavonoid components extracted from DiopyrosKaki on oxidative stress induced by OGD/R in HT22 cells[J]. Journal of Shandong University(Health Sciences),2018,56(6):6-12.
[21]Sova M,Saso L. Design and development of Nrf2 modula-tors for cancer chemoprevention and therapy:a review[J]. Drug Des Devel Ther,2018,12:3181-3197. doi:10. 2147/DDDT. S172612
[22] Yamamoto M,Kensler TW,Motohashi H. The KEAP1-NRF2system:a thiol-based sensor-effector apparatus formaintaining redox homeostasis[J]. Physiol Rev,2018,98(3):1169-1203.
[23]Wang XJ,Chen Q,Zhang ZY,et al. Novel enteromorphaprolifera based carbon dots:probing the radical scaven-ging of natural phenolic compounds[J]. Colloids Surf BBiointerfaces,2018,174:161-167. doi:10. 1016/j. col-surfb. 2018. 11. 019
[24]Corsello T,Komaravelli N,Casola A. Role of hydrogensulfide in NRF2-and sirtuin-dependent maintenance ofcellular redox balance[J]. Antioxidants(Basel),2018,7(10):E129. doi:10. 3390/antiox7100129
[25] Nowak WN,Taha H,Kachamakova-Trojanowska N,etal. Murine bone marrow mesenchymal stromal cells re-spond efficiently to oxidative stress despite the low level ofheme oxygenases 1 and 2[J]. Antioxid Redox Signal,2018,29(2):111-127.