IFN-β基因修饰的人骨髓间充质干细胞抗前列腺癌细胞系PC-3效应的实验研究
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摘要
目的:观察人类β干扰素(Interferon-beta, IFN-β)基因转染的人骨髓间充质干细胞(Human mesenchymal stem cells, hMSCs)对前列腺癌细胞PC-3生长情况的影响,初步探讨人骨髓间充质干细胞作为基因运载细胞治疗前列腺癌的可行性。
方法:应用人前列腺癌细胞株PC-3异种皮下种植建立前列腺癌SCID鼠模型,通过密度梯度离心法分离、培养hMSCs。将体外CM-DiI标记的4～6代hMSCs经尾静脉和肿瘤周围,注射入荷瘤SCID鼠体内,将肿瘤和肝、肺、脾、肾等脏器作冰冻切片和石蜡切片,在荧光显微镜下观察肿瘤及各脏器中hMSCs的分布情况。将已构建的人类β干扰素基因腺病毒表达载体Ad-IFN-β,体外转染人骨髓间充质干细胞。将成功表达β干扰素的IFN-β-hMSCs经尾静脉和肿瘤瘤内注射两种途径将IFN-β-hMSCs注射入小鼠模型体内,观察动物模型中肿瘤大小及生存期,评价IFN-β-hMSCs治疗前列腺癌的实验效果。
结果:前列腺癌SCID鼠模型成瘤率91.7%,经病理学证实为均为恶性组织。hMSC经尾静脉注射后,可以向前列腺癌组织趋向和聚集,而在正常组织中未发现hMSC的分布(如,肺脏,肝脏,脾脏,肾脏,肌肉)。经尾静脉和肿瘤原位两种方式途径注射IFN-β-hMSCs都可延长荷瘤鼠的生存期,抑制肿瘤的生长,实验组与各对照组之间比较差异存在统计学意义(P<0.001)。
结论:hMSC可以向肿瘤微环境趋向转移,经β干扰素基因修饰的hMSCs体内实验可以有效抑制前列腺癌细胞生长,延长荷瘤鼠的生存期。研究结果可能为前列腺癌的基因靶向治疗提供新的策略。
Objective: To observe the effects of human interferon-beta gene cDNA engineered human bone marrow mesenchymal stem cells(hMSCs) on the growth of prostate cancer cell line PC-3 in vivo,and explore the feasibility of hMSCs as vehicles for interferon-beta delivery into prostate cancer.
Methods: The human prostate cancer cell line PC-3 were injected into the axillary of SCID mice subcutaneously to establish human prostate cancer xenograft models. hMSCs were harvested from donor’s ribs of human cadaver renal transplantation and separated by density gradient centrifuge. hMSCs between passages 4 to 6 were labeled with CM-DiI. CM-DiI-labeled hMSCs were injected into the bearing cancer SCID mice by tail vein of labeled MSCs or tumor. The mice were killed and their tumors、livers、lungs、spleens and kidneys were harvested.Frozen sections and paraffin sections were used to observe the distribution of exogenous CM-DiI-labeled hMSCs in vivo by fluorescence microscope.The adenoviral expression vector containing huIFN-βgene(Ad-IFN-β) was constructed and transfected into hMSCs.The human prostate cancer cell line PC-3 were injected into severe combined immunodeficiency mouse(SCID) subcutaneously to establish human prostate cancer xenograft models.IFN-β-hMSCs were injected into the bearing cancer SCID mice by tail vein or intra-tumor.We observed the influence of IFN-β-hMSCs on the tumor or mice’survival and evaluated the effects of prostate cancer in response to IFN-β-hMSCs in vivo.
Results: In SCID mice injected with PC-3 subcutaneously,the tumor take rate was91.7%.The tumors were identified by pathology After injection IFN-β-hMSCs into bearing cancer SCID mice by tail vein,IFN-β-hMSCs could migrate to prostate cancer microenviroment in vivo.But no IFN-β-hMSCs was seen in the lungs、livers、spleens、kidneys and muscles.Compared with control mice,injection of IFN-β-hMSCs by tail vein or intra-tumor could prolong mice’survival and inhibit the growth of prostate cancer. There was significant difference between experiment group and control group(P<0.001).
Conclusions: The hMSCs can express IFN-βsuccessfully after huIFN-βgene transfection. IFN-β-hMSCs can migrate to prostate cancer microenviroment in vivo. IFN-β-hMSCs can inhibit the growth of prostate cancer significantly and prolong mice’survival in vivo, which may develop a new strategy about gene therapy in prostate cancer.
方法:应用人前列腺癌细胞株PC-3异种皮下种植建立前列腺癌SCID鼠模型,通过密度梯度离心法分离、培养hMSCs。将体外CM-DiI标记的4～6代hMSCs经尾静脉和肿瘤周围,注射入荷瘤SCID鼠体内,将肿瘤和肝、肺、脾、肾等脏器作冰冻切片和石蜡切片,在荧光显微镜下观察肿瘤及各脏器中hMSCs的分布情况。将已构建的人类β干扰素基因腺病毒表达载体Ad-IFN-β,体外转染人骨髓间充质干细胞。将成功表达β干扰素的IFN-β-hMSCs经尾静脉和肿瘤瘤内注射两种途径将IFN-β-hMSCs注射入小鼠模型体内,观察动物模型中肿瘤大小及生存期,评价IFN-β-hMSCs治疗前列腺癌的实验效果。
结果:前列腺癌SCID鼠模型成瘤率91.7%,经病理学证实为均为恶性组织。hMSC经尾静脉注射后,可以向前列腺癌组织趋向和聚集,而在正常组织中未发现hMSC的分布(如,肺脏,肝脏,脾脏,肾脏,肌肉)。经尾静脉和肿瘤原位两种方式途径注射IFN-β-hMSCs都可延长荷瘤鼠的生存期,抑制肿瘤的生长,实验组与各对照组之间比较差异存在统计学意义(P<0.001)。
结论:hMSC可以向肿瘤微环境趋向转移,经β干扰素基因修饰的hMSCs体内实验可以有效抑制前列腺癌细胞生长,延长荷瘤鼠的生存期。研究结果可能为前列腺癌的基因靶向治疗提供新的策略。
Objective: To observe the effects of human interferon-beta gene cDNA engineered human bone marrow mesenchymal stem cells(hMSCs) on the growth of prostate cancer cell line PC-3 in vivo,and explore the feasibility of hMSCs as vehicles for interferon-beta delivery into prostate cancer.
Methods: The human prostate cancer cell line PC-3 were injected into the axillary of SCID mice subcutaneously to establish human prostate cancer xenograft models. hMSCs were harvested from donor’s ribs of human cadaver renal transplantation and separated by density gradient centrifuge. hMSCs between passages 4 to 6 were labeled with CM-DiI. CM-DiI-labeled hMSCs were injected into the bearing cancer SCID mice by tail vein of labeled MSCs or tumor. The mice were killed and their tumors、livers、lungs、spleens and kidneys were harvested.Frozen sections and paraffin sections were used to observe the distribution of exogenous CM-DiI-labeled hMSCs in vivo by fluorescence microscope.The adenoviral expression vector containing huIFN-βgene(Ad-IFN-β) was constructed and transfected into hMSCs.The human prostate cancer cell line PC-3 were injected into severe combined immunodeficiency mouse(SCID) subcutaneously to establish human prostate cancer xenograft models.IFN-β-hMSCs were injected into the bearing cancer SCID mice by tail vein or intra-tumor.We observed the influence of IFN-β-hMSCs on the tumor or mice’survival and evaluated the effects of prostate cancer in response to IFN-β-hMSCs in vivo.
Results: In SCID mice injected with PC-3 subcutaneously,the tumor take rate was91.7%.The tumors were identified by pathology After injection IFN-β-hMSCs into bearing cancer SCID mice by tail vein,IFN-β-hMSCs could migrate to prostate cancer microenviroment in vivo.But no IFN-β-hMSCs was seen in the lungs、livers、spleens、kidneys and muscles.Compared with control mice,injection of IFN-β-hMSCs by tail vein or intra-tumor could prolong mice’survival and inhibit the growth of prostate cancer. There was significant difference between experiment group and control group(P<0.001).
Conclusions: The hMSCs can express IFN-βsuccessfully after huIFN-βgene transfection. IFN-β-hMSCs can migrate to prostate cancer microenviroment in vivo. IFN-β-hMSCs can inhibit the growth of prostate cancer significantly and prolong mice’survival in vivo, which may develop a new strategy about gene therapy in prostate cancer.
引文
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