干扰MITF表达对恶性黑素瘤细胞B16F10增殖及黑素生成的影响
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摘要
目的验证小眼畸形相关转录因子(MITF)高表达在恶性黑素瘤细胞中的关键作用。方法利用RNA干扰技术特异性抑制MITF基因及其蛋白表达水平,研究该基因对B16F10细胞增殖活性的影响,验证MITF对黑素瘤细胞内外黑素分泌水平的影响及对与该分泌水平密切相关的酪氨酸酶(TYR)活性的影响。结果 RNA干扰技术在B16F10细胞中能显著抑制MITF基因及蛋白表达水平,分别为82.7%、50.0%,干扰MITF表达能抑制B16F10增殖活性达49.5%,能抑制黑素瘤细胞内、外黑素分泌水平,分别为49.7%、43.8%,能抑制TYR活性达48.0%。结论 MITF对黑素瘤细胞增殖活性具有重要作用,对黑素瘤细胞中的TYR活性及黑素生成具有重要作用,与黑素瘤细胞中的黑素功能密切相关。
Objective To demonstrate that overexpression of microphthalmia-associated transcription factors(MITF)was associated with malignant melanoma cell proliferation and melanogenesis.Methods The effects of the gene on the prolifer-ation of B16 F10 cells and the effect of MITF on the secretion of melanin and the tyrosinase(TYR)activity closely related tothe secretory level of the melanoma cells were investigated by the specific inhibition of the expression of MITF gene and its pro-tein expression by RNA interference technique.Results By the RNA interfering technology,the MITF gene and protein ex-pression was significantly reduced compared to the negative control.Knockdown of MITF m RNA and protein expressions inhibit-ed B16 F10 cell proliferation,melanogenesis and TYR activity.Conclusion This study showed that MITF can be useful as abiomarker for prediction of malignant melanoma progression and melanogenesis and target of MITF as a novel strategy for malig-nant melanoma therapy.
Objective To demonstrate that overexpression of microphthalmia-associated transcription factors(MITF)was associated with malignant melanoma cell proliferation and melanogenesis.Methods The effects of the gene on the prolifer-ation of B16 F10 cells and the effect of MITF on the secretion of melanin and the tyrosinase(TYR)activity closely related tothe secretory level of the melanoma cells were investigated by the specific inhibition of the expression of MITF gene and its pro-tein expression by RNA interference technique.Results By the RNA interfering technology,the MITF gene and protein ex-pression was significantly reduced compared to the negative control.Knockdown of MITF m RNA and protein expressions inhibit-ed B16 F10 cell proliferation,melanogenesis and TYR activity.Conclusion This study showed that MITF can be useful as abiomarker for prediction of malignant melanoma progression and melanogenesis and target of MITF as a novel strategy for malig-nant melanoma therapy.
引文
[1]VERRAS M,SUN Z.Roles and regulation of Wnt signaling and beta-catenin in prostate cancer[J].Cancer Lett,2006,237(1):22-32.
[2]CHUNG ES,SABEL MS,SONDAK VK.Current state of treatment forprimary cutaneous melanoma[J].Clin Exp Med.2004.4(2):65-77.
[3]BALCH CM,BUZAID AC,ATKINS MB,et al.A new American Joint Com-mittee on Cancer staging system for cutaneous melanoma[J].Cancer,2000,88(6):1484-1491.
[4]BUSAM KJ,KUCUKG?L D,SATO E,et al.Immunohistochemical analy-sis of novel monoclonal antibody PNL2 and comparison with other mela-nocyte differentiation markers[J].Am J Surg Pathol,2005,29(3):400-406.
[5]O′REILLY FM,BRAT DJ,MCALPINE BE,et al.Microphthalmia tran-scription factor immunohistochemistry:a useful diagnostic marker in thediagnosis and detection of cutaneous melanoma,sentinel lymph nodemetastases,and extracutaneous melanocytic neoplasms[J].J Am AcadDermatol,2001,45(3):414-419.
[6]CHELI Y,OHANNA M,BALLOTTI R,et al.Fifteen-year quest for microphthalmia-associated transcription factor target genes[J].PigmentCell Melanoma Res,2010,23(1):27-40.
[7]KING R,GOOGE PB,WEILBAECHER KN,et al.Microphthalmia tran-scription factor expression in cutaneous benign,malignant melanocytic,and nonmelanocytic tumors[J].Am J Surg Pathol,2001,25(1):51-57.
[8]KIM HJ,KIM JS,WOO JT,et al.Hyperpigmentation mechanism ofmethyl 3,5-di-caffeoylquinate through activation of p38 and MITF induc-tion of tyrosinase[J].Acta Biochim Biophys Sin(Shanghai),2015,47(7):548-456.
[9]DENAT L,LARUE L.Malignant melanoma and the role of the paradoxalprotein Microphthalmia transcription factor[J].Bull Cancer,2007,94(1):81-92.
[10]BERTOLOTTO C,BUSCàR,ABBE P,et al.Different cis-acting ele-ments are involved in the regulation of TRP1 and TRP2 promoter activi-ties by cyclic AMP:pivotal role of M boxes(GTCATGTGCT)and of mi-crophthalmia[J].Mol Cell Biol,1998,18(2):694-702.
[11]VACHTENHEIM J,ONDRU?OVáL.Microphthalmia-associated tran-scription factor expression levels in melanoma cells contribute to cell in-vasion and proliferation[J].Exp Dermatol,2015,24(7):481-484.
[2]CHUNG ES,SABEL MS,SONDAK VK.Current state of treatment forprimary cutaneous melanoma[J].Clin Exp Med.2004.4(2):65-77.
[3]BALCH CM,BUZAID AC,ATKINS MB,et al.A new American Joint Com-mittee on Cancer staging system for cutaneous melanoma[J].Cancer,2000,88(6):1484-1491.
[4]BUSAM KJ,KUCUKG?L D,SATO E,et al.Immunohistochemical analy-sis of novel monoclonal antibody PNL2 and comparison with other mela-nocyte differentiation markers[J].Am J Surg Pathol,2005,29(3):400-406.
[5]O′REILLY FM,BRAT DJ,MCALPINE BE,et al.Microphthalmia tran-scription factor immunohistochemistry:a useful diagnostic marker in thediagnosis and detection of cutaneous melanoma,sentinel lymph nodemetastases,and extracutaneous melanocytic neoplasms[J].J Am AcadDermatol,2001,45(3):414-419.
[6]CHELI Y,OHANNA M,BALLOTTI R,et al.Fifteen-year quest for microphthalmia-associated transcription factor target genes[J].PigmentCell Melanoma Res,2010,23(1):27-40.
[7]KING R,GOOGE PB,WEILBAECHER KN,et al.Microphthalmia tran-scription factor expression in cutaneous benign,malignant melanocytic,and nonmelanocytic tumors[J].Am J Surg Pathol,2001,25(1):51-57.
[8]KIM HJ,KIM JS,WOO JT,et al.Hyperpigmentation mechanism ofmethyl 3,5-di-caffeoylquinate through activation of p38 and MITF induc-tion of tyrosinase[J].Acta Biochim Biophys Sin(Shanghai),2015,47(7):548-456.
[9]DENAT L,LARUE L.Malignant melanoma and the role of the paradoxalprotein Microphthalmia transcription factor[J].Bull Cancer,2007,94(1):81-92.
[10]BERTOLOTTO C,BUSCàR,ABBE P,et al.Different cis-acting ele-ments are involved in the regulation of TRP1 and TRP2 promoter activi-ties by cyclic AMP:pivotal role of M boxes(GTCATGTGCT)and of mi-crophthalmia[J].Mol Cell Biol,1998,18(2):694-702.
[11]VACHTENHEIM J,ONDRU?OVáL.Microphthalmia-associated tran-scription factor expression levels in melanoma cells contribute to cell in-vasion and proliferation[J].Exp Dermatol,2015,24(7):481-484.