Pou6f2基因对泌乳素腺瘤MMQ细胞增殖和分泌能力的影响
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摘要
目的观察Pou class 6同源框2(Pou6f2)基因对泌乳素腺瘤MMQ细胞增殖和分泌能力的影响。方法敲低Pou6f2实验:将细胞分为对照组和实验1、2组,按Lipofectamine~? 3000转染试剂盒说明书分别转染control-siRNA、Pou6f2-siRNA-1、Pou6f2-siRNA-2,72 h后收集细胞。过表达Pou6f2实验:将细胞分为对照组和实验组,按Lipofectamine~? 3000转染试剂盒说明书分别转染空载质粒、野生型Pou6f2质粒,72 h后收集细胞。采用Western blotting法检测细胞中Pou6f2表达量,MTS法测定细胞增殖活性(A_(490)值),ELISA法测定细胞泌乳素(PRL)分泌水平。结果敲低Pou6f2实验中,与对照组比较,实验1、2组MMQ细胞Pou6f2表达量降低、增殖的A_(490)值增加、分泌PRL水平升高(P均<0. 01),实验1、2组各指标差异均无统计学意义。过表达Pou6f2实验中,与对照组比较,实验组MMQ细胞Pou6f2表达量增加、增殖的A_(490)值降低、分泌PRL水平下降(P均<0. 01)。结论 Pou6f2可以抑制泌乳素腺瘤MMQ细胞的增殖及PRL分泌能力,有望成为泌乳素腺瘤治疗的新靶点。
Objective To observe the effects of Pou class 6 homeobox 2 (Pou6f2) on the proliferation and secretion of prolactin of rat prolactinoma MMQ cells. Methods The experiment of knocking down Pou6 f2: The MMQ cells were divided into the control group and the experimental groups 1 and 2,and the control-siRNA,Pou6f2-siRNA-1,Pou6 f2-siRNA-2 were transfected into cells respectively according to the instructions of Lipofectamine~? 3000 transfection kit,and the cells were collected 72 hours later. The experiment of overexpressing Pou6f2: The MMQ cells were divided into the control group and experimental group,and the empty plasmid and wild-type Pou6f2 plasmid were transfected into cells respectively according to the instructions of Lipofectamine~? 3000 transfection kit,and the cells were collected 72 hours later. Western blotting was used to detect the expression of Pou6f2. MTS was used to determine the cell proliferation activity( A_(490) value),and ELISA was used to determine the level of prolactin( PRL) secretion. Results In the experiment of knocking down Pou6f2,compared with the control group,the expression of Pou6f2 decreased( P < 0. 01),the value of A_(490) increased( P < 0. 01),and the level of prolactin secretion increased( P < 0. 01) in the experimental group 1 and group 2.There were no significant differences between the experimental group 1 and experimental group 2. In the experiment of overexpressing Pou6f2,compared with the control group,the expression of Pou6f2 increased( P < 0. 01),the value of A_(490) decreased( P < 0. 01),and the level of prolactin secretion decreased in the experimental group( P < 0. 01). Conclusion Pou6f2 can inhibit the proliferation of MMQ cells and reduce the ability of PRL secretion,which provides a new target for the treatment of prolactinoma.
Objective To observe the effects of Pou class 6 homeobox 2 (Pou6f2) on the proliferation and secretion of prolactin of rat prolactinoma MMQ cells. Methods The experiment of knocking down Pou6 f2: The MMQ cells were divided into the control group and the experimental groups 1 and 2,and the control-siRNA,Pou6f2-siRNA-1,Pou6 f2-siRNA-2 were transfected into cells respectively according to the instructions of Lipofectamine~? 3000 transfection kit,and the cells were collected 72 hours later. The experiment of overexpressing Pou6f2: The MMQ cells were divided into the control group and experimental group,and the empty plasmid and wild-type Pou6f2 plasmid were transfected into cells respectively according to the instructions of Lipofectamine~? 3000 transfection kit,and the cells were collected 72 hours later. Western blotting was used to detect the expression of Pou6f2. MTS was used to determine the cell proliferation activity( A_(490) value),and ELISA was used to determine the level of prolactin( PRL) secretion. Results In the experiment of knocking down Pou6f2,compared with the control group,the expression of Pou6f2 decreased( P < 0. 01),the value of A_(490) increased( P < 0. 01),and the level of prolactin secretion increased( P < 0. 01) in the experimental group 1 and group 2.There were no significant differences between the experimental group 1 and experimental group 2. In the experiment of overexpressing Pou6f2,compared with the control group,the expression of Pou6f2 increased( P < 0. 01),the value of A_(490) decreased( P < 0. 01),and the level of prolactin secretion decreased in the experimental group( P < 0. 01). Conclusion Pou6f2 can inhibit the proliferation of MMQ cells and reduce the ability of PRL secretion,which provides a new target for the treatment of prolactinoma.
引文
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[18]King R,Struebing FL,Li Y,et al.Genomic locus modulating corneal thickness in the mouse identifies POU6F2 as a potential risk of developing glaucoma[J].PLo S Genet,2018,14(1):e1007145.
[19]Di Renzo F,Doneda L,Menegola E,et al.The murine Pou6f2 gene is temporally and spatially regulated during kidney embryogenesis and its human homolog is overexpressed in a subset of Wilms tumors[J].J Pediatr Hematol Oncol,2006,28:791-797.
[20]Kang H,Tan M,Bishop JA,et al.Whole-exome sequencing of salivary gland mucoepidermoid carcinoma[J].Clin Cancer Res,2017,23(1):283-288.
[21]Yoshida S,Ueharu H,Higuchi M,et al.Molecular cloning of rat and porcine retina-derived POU domain factor 1(POU6F2)from a pituitary c DNA library[J].J Reprod Dev,2014,60(4):288-294.
[2]Caimari F,Korbonits M.Novel genetic causes of pituitary adenomas[J].Clin Cancer Res,2016,22(20):5030-5042.
[3]刘阳,连伟,王任直,等.垂体泌乳素腺瘤诊疗中常见问题及解决方案[J].中国临床神经外科杂志,2015,20(11):698-703.
[4]Perotti D,De Vecchi G,Testi MA,et al.Germline mutations of the POU6F2 gene in Wilms tumors with loss of heterozygosity on chromosome 7p14[J].Hum Mutat,2004,24(5):400-407.
[5]Xie W,Wang H,He Y,et al.CDK5 and its activator p35 in normal pituitary and in pituitary adenomas:relationship to VEGF expression[J].Int J Biol Sci,2014,10(2):192-199.
[6]Asa SL,Ezzat S.The pathogenesis of pituitary tumors[J].Annu Rev Pathol,2009,4:97-126.
[7]Valimaki N,Demir H,Pitkanen E,et al.Whole-genome sequencing of growth hormone(GH)-secreting pituitary adenomas[J].JClin Endocrinol Metab,2015,100(10):3918-3927.
[8]Reincke M,Sbiera S,Hayakawa A,et al.Mutations in the deubiquitinase gene USP8 cause Cushing's disease[J].Nat Genet,2015,47(1):31-38.
[9]Ma ZY,Song ZJ,Chen JH,et al.Recurrent gain-of-function USP8 mutations in Cushing's disease[J].Cell Res,2015,25(3):306-317.
[10]Bi WL,Horowitz P,Greenwald NF,et al.Landscape of genomic alterations in pituitary adenomas[J].Clin Cancer Res,2017,23(7):1841-1851.
[11]Song ZJ,Reitman ZJ,Ma ZY,et al.The genome-wide mutational landscape of pituitary adenomas[J].Cell Res,2016,26(11):1255-1259.
[12]Cook AL,Sturm RA.POU domain transcription factors:BRN2 as a regulator of melanocytic growth and tumourigenesis[J].Pigment Cell Melanoma Res,2008,21(6):611-626.
[13]Takagi M,Kamasaki H,Yagi H,et al.A novel heterozygous intronic mutation in POU1F1 is associated with combined pituitary hormone deficiency[J].Endocr J,2017,64(2):229-234.
[14]Bademci G,Lasisi A,Yariz KO,et al.Novel domain-specific POU3F4 mutations are associated with X-linked deafness:examples from different populations[J].BMC Med Genet,2015,16:9.
[15]Du W,Han MK,Wang DY,et al.A POU3F4 mutation causes nonsyndromic hearing loss in a chinese X-linked recessive family[J].Chin Med J(Engl),2017,130(1):88-92.
[16]Abate-Shen C.Homeobox genes and cancer:new OCTaves for an old tune[J].Cancer Cell,2003,4(5):329-330.
[17]Gidekel S,Pizov G,Bergman Y,et al.Oct-3/4 is a dose-dependent oncogenic fate determinant[J].Cancer Cell,2003,4(5):361-370.
[18]King R,Struebing FL,Li Y,et al.Genomic locus modulating corneal thickness in the mouse identifies POU6F2 as a potential risk of developing glaucoma[J].PLo S Genet,2018,14(1):e1007145.
[19]Di Renzo F,Doneda L,Menegola E,et al.The murine Pou6f2 gene is temporally and spatially regulated during kidney embryogenesis and its human homolog is overexpressed in a subset of Wilms tumors[J].J Pediatr Hematol Oncol,2006,28:791-797.
[20]Kang H,Tan M,Bishop JA,et al.Whole-exome sequencing of salivary gland mucoepidermoid carcinoma[J].Clin Cancer Res,2017,23(1):283-288.
[21]Yoshida S,Ueharu H,Higuchi M,et al.Molecular cloning of rat and porcine retina-derived POU domain factor 1(POU6F2)from a pituitary c DNA library[J].J Reprod Dev,2014,60(4):288-294.