人子宫内膜异位症的可示踪裸鼠模型构建
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摘要
目的运用异体移植动物模型及活体成像技术,构建人子宫内膜异位症的可示踪裸鼠模型。方法采集人子宫在位内膜组织,制成悬液。应用慢病毒载体将荧光素酶和红色荧光蛋白基因转入内膜组织并培养48h,将等体积的悬液(分成3组:A组已转染的内膜组织,B组未转染的内膜组织,C组等体积的DMEM/F12培养基)注射到BALB/C雌性裸鼠腹腔内,运用活体成像仪监测异位病灶的发展情况。28d后分离异位病灶,运用苏木素-伊红(HE)染色和免疫组织化学鉴定异位病灶的来源并分析成模情况。结果已成功转染内膜组织的裸鼠在活体成像仪监测下显示,异位病灶的荧光信号在种植后第7天至第14天减小(P<0.01);第14天到第28天增大(P<0.05)。HE染色和免疫组织化学显示异位病灶均来自于人子宫内膜组织。结论人子宫内膜异位症的可示踪裸鼠模型构建成功。
Objective To establish tracing nude mouse model of human endometriosis,which was transfected by lentivirus-RFP-luciferase and detected by in vivo bioluminescence imaging(BLI)system.Methods Human endometrial tissues were collected and made into suspension,which were successfully transfected by lentivirus-RFP-luciferase and cultured for 48 hin vitro.Isometric suspension,which was divided into three groups:the transfected group(group A),the untransfected group(group B)and the DMEM/F12 group(group C),was intraperitoneally injected into female BALB/C nude mice.The development of ectopic lesion was dynamically detected by BLI system.Twenty-eight days later,in order to learn the establishment of model,ectopic lesions were separated and determinated by biotechnology such as HE stain and immunohistochemical analysis.Results The BLI system showed that the ROI of ectopic lesions decreased from the 7 th to the14 th day(P<0.01),and increased from the 14 th to the 28 th day in group A(P<0.05).Both HE stain and immunohistochemical analysis showed that the ectopic lesions were derived from human endometrial tissues.Conclusion The tracing nude mouse model of human endometriosis is successfully established.
Objective To establish tracing nude mouse model of human endometriosis,which was transfected by lentivirus-RFP-luciferase and detected by in vivo bioluminescence imaging(BLI)system.Methods Human endometrial tissues were collected and made into suspension,which were successfully transfected by lentivirus-RFP-luciferase and cultured for 48 hin vitro.Isometric suspension,which was divided into three groups:the transfected group(group A),the untransfected group(group B)and the DMEM/F12 group(group C),was intraperitoneally injected into female BALB/C nude mice.The development of ectopic lesion was dynamically detected by BLI system.Twenty-eight days later,in order to learn the establishment of model,ectopic lesions were separated and determinated by biotechnology such as HE stain and immunohistochemical analysis.Results The BLI system showed that the ROI of ectopic lesions decreased from the 7 th to the14 th day(P<0.01),and increased from the 14 th to the 28 th day in group A(P<0.05).Both HE stain and immunohistochemical analysis showed that the ectopic lesions were derived from human endometrial tissues.Conclusion The tracing nude mouse model of human endometriosis is successfully established.
引文
[1]VERCELLINI P,VIGANO P,SOMIGLIANA E,et al.Endometriosis:pathogenesis and treatment[J].Nat Rev Endocrinol,2014,10(5):261-275.
[2]金济云,夏亲华.子宫内膜异位症动物模型研究进展[J].中国医药导报,2016,13(5):28-31.
[3]GRUMMER R,SCHWARZER F,BAINCZYK K,et al.Peritoneal endometriosis:validation of an in-vivo model[J].Hum Reprod,2001,16(8):1736-1743.
[4]EDWARDS A K,NAKAMURA D S,VIRANI S,et al.Animal models for anti-angiogenic therapy in endometriosis[J].J Reprod Immunol,2013,97(1):85-94.
[5]MEHEDINTU C,PLOTOGEA M N,IONESCU S,et al.Endometriosis still a challenge[J].J Med Life,2014,7(3):349-357.
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[7]曹芳,黄晓阳,葛素梅,等.子宫内膜异位症大鼠内膜窗口期模型的建立[J].重庆医学,2015,44(27):3805-3807.
[8]BRUNER-TRAN K L,MOKSHAGUNDAM S,HER-INGTON J L,et al.Rodent models of experimental endometriosis:identifying mechanisms of disease and therapeutic targets[J].Curr Women Health Rev,2018,14(2):173-188.
[9]GREAVES E,COUSINS F L,MURRAY A,et al.A novel mouse model of endometriosis mimics human phenotype and reveals insights into the inflammatory contribution of shed endometrium[J].Am J Pathol,2014,184(7):1930-1939.
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[12]HE H,LIU R,XIONG W,et al.Lentiviral vector-mediated down-regulation of Notch1in endometrial stem cells results in proliferation and migration in endometriosis[J].Mol Cell Endocrinol,2016,434:210-218.
[13]LU Z,ZHANG W,JIANG S,et al.Effect of oxygen tensions on the proliferation and angiogenesis of endometriosis heterograft in severe combined immunodeficiency mice[J].Fertil Steril,2014,101(2):568-576.
[14]ERTEN O U,ENSARI T A,DILBAZ B,et al.Vitamin Cis effective for the prevention and regression of endometriotic implants in an experimentally induced rat model of endometriosis[J].Taiwan J Obstet Gynecol,2016,55(2):251-257.
[15]刘斌,王宁宁.子宫内膜异位症动物模型活体无创观察研究[J].国际妇产科学杂志,2009,36(6):472-475.
[16]SIMITSIDELLIS I,GIBSON D A,SAUNDERS P.Animal models of endometriosis:Replicating the aetiology and symptoms of the human disorder[J].Best Pract Res Clin Endocrinol Metab,2018,32(3):257-269.
[17]FERRERO H,BUIGUES A,MARTINEZ J,et al.A novel homologous model for noninvasive monitoring of endometriosis progression[J].Biol Reprod,2017,96(2):302-312.
[18]WANG N,HONG S,TAN J,et al.A red fluorescent nude mouse model of human endometriosis:advantages of a non-invasive imaging method[J].Eur J Obstet Gynecol Reprod Biol,2014,176:25-30.
[19]BADR C E.Bioluminescence imaging:basics and practical limitations[J].Methods Mol Biol,2014,1098:1-18.
[20]ZHAO T,LIU X,ZHEN X,et al.Levo-tetrahydropalmatine retards the growth of ectopic endometrial implants and alleviates generalized hyperalgesia in experimentally induced endometriosis in rats[J].Reprod Sci,2011,18(1):28-45.
[21]SOHNGEN L,SCHMIDT M,WIMBERGER P,et al.Additional B-cell deficiency does not affect growth and angiogenesis of ectopic human endometrium in T-cell-deficient endometriosis mouse models during long-term culture[J].J Reprod Immunol,2014,106:50-57.
[22]GROOTHUIS P G.Angiogenesis and vascular remodelling in female reproductive organs[J].Angiogenesis,2005,8(2):87-88.
[2]金济云,夏亲华.子宫内膜异位症动物模型研究进展[J].中国医药导报,2016,13(5):28-31.
[3]GRUMMER R,SCHWARZER F,BAINCZYK K,et al.Peritoneal endometriosis:validation of an in-vivo model[J].Hum Reprod,2001,16(8):1736-1743.
[4]EDWARDS A K,NAKAMURA D S,VIRANI S,et al.Animal models for anti-angiogenic therapy in endometriosis[J].J Reprod Immunol,2013,97(1):85-94.
[5]MEHEDINTU C,PLOTOGEA M N,IONESCU S,et al.Endometriosis still a challenge[J].J Med Life,2014,7(3):349-357.
[6]KIM S M,YOO T,LEE S Y,et al.Effect of SKI2670,a novel,orally active,non-peptide GnRH antagonist,on hypothalamic-pituitary-gonadal axis[J].Life Sci,2015,139:166-174.
[7]曹芳,黄晓阳,葛素梅,等.子宫内膜异位症大鼠内膜窗口期模型的建立[J].重庆医学,2015,44(27):3805-3807.
[8]BRUNER-TRAN K L,MOKSHAGUNDAM S,HER-INGTON J L,et al.Rodent models of experimental endometriosis:identifying mechanisms of disease and therapeutic targets[J].Curr Women Health Rev,2018,14(2):173-188.
[9]GREAVES E,COUSINS F L,MURRAY A,et al.A novel mouse model of endometriosis mimics human phenotype and reveals insights into the inflammatory contribution of shed endometrium[J].Am J Pathol,2014,184(7):1930-1939.
[10]YAMAGATA Y,TAKAKI E,SHINAGAWA M,et al.Retinoic acid has the potential to suppress endometriosis development[J].J Ovarian Res,2015,8:49.
[11]AGOSTINIS C,ZORZET S,DE LEO R,et al.The combination of N-acetyl cysteine,alpha-lipoic acid,and bromelain shows high anti-inflammatory properties in novel in vivo and in vitro models of endometriosis[J].Mediators Inflamm,2015(2015):918089.
[12]HE H,LIU R,XIONG W,et al.Lentiviral vector-mediated down-regulation of Notch1in endometrial stem cells results in proliferation and migration in endometriosis[J].Mol Cell Endocrinol,2016,434:210-218.
[13]LU Z,ZHANG W,JIANG S,et al.Effect of oxygen tensions on the proliferation and angiogenesis of endometriosis heterograft in severe combined immunodeficiency mice[J].Fertil Steril,2014,101(2):568-576.
[14]ERTEN O U,ENSARI T A,DILBAZ B,et al.Vitamin Cis effective for the prevention and regression of endometriotic implants in an experimentally induced rat model of endometriosis[J].Taiwan J Obstet Gynecol,2016,55(2):251-257.
[15]刘斌,王宁宁.子宫内膜异位症动物模型活体无创观察研究[J].国际妇产科学杂志,2009,36(6):472-475.
[16]SIMITSIDELLIS I,GIBSON D A,SAUNDERS P.Animal models of endometriosis:Replicating the aetiology and symptoms of the human disorder[J].Best Pract Res Clin Endocrinol Metab,2018,32(3):257-269.
[17]FERRERO H,BUIGUES A,MARTINEZ J,et al.A novel homologous model for noninvasive monitoring of endometriosis progression[J].Biol Reprod,2017,96(2):302-312.
[18]WANG N,HONG S,TAN J,et al.A red fluorescent nude mouse model of human endometriosis:advantages of a non-invasive imaging method[J].Eur J Obstet Gynecol Reprod Biol,2014,176:25-30.
[19]BADR C E.Bioluminescence imaging:basics and practical limitations[J].Methods Mol Biol,2014,1098:1-18.
[20]ZHAO T,LIU X,ZHEN X,et al.Levo-tetrahydropalmatine retards the growth of ectopic endometrial implants and alleviates generalized hyperalgesia in experimentally induced endometriosis in rats[J].Reprod Sci,2011,18(1):28-45.
[21]SOHNGEN L,SCHMIDT M,WIMBERGER P,et al.Additional B-cell deficiency does not affect growth and angiogenesis of ectopic human endometrium in T-cell-deficient endometriosis mouse models during long-term culture[J].J Reprod Immunol,2014,106:50-57.
[22]GROOTHUIS P G.Angiogenesis and vascular remodelling in female reproductive organs[J].Angiogenesis,2005,8(2):87-88.