B18R--mRNA重编程诱导iPS的必备利器

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作者:联科生物 发布日期:2014-01-26 07:00

B18R--mRNA重编程诱导iPS的必备利器
1.B18R的组成及作用

8R蛋白是由痘苗病毒WesternReserve株编码的受体,以可溶性和膜型两种形式存在。该蛋白可作为I型干扰素(IFN)受体与其结合,并具有广泛的种属特异性。B18R蛋白对人IFN-α具有高亲和力,也能与兔、牛、小鼠、猪和小鼠的IFN-α和IFN-β结合[1,2]。由于该蛋白的两种存在形式,因此可阻断IFN的自分泌和旁分泌功能,从而抑制人IFN-α1、IFN-α2、IFN-α8/1/8和IFN-ω的抗病毒潜能。另有报道发现该蛋白也能高效中和猪IFN-δ[3]。分泌的B18R可与感染和未感染细胞结合,阻止IFN-α和IFN-β诱导细胞进入抗病毒状态,从而使痘苗病毒复制[4]。

2.B18R无需病毒载体,无需反复转染,效率更高

近年来,研究者在培养基中添加B18R蛋白,抑制IFN诱导的凋亡,提高细胞活力,转染修饰合成的mRNA重编程因子(OCT4, SOX2, KLF4, c-MYC和Lin28),从而使体细胞发生高效的mRNA重编程,诱导产生iPS [6-9]。该方法较蛋白/多肽转入细胞更加有效,这是由于后者在细胞中的半衰期较短[10,11]。相对于RNA转染引起先天免疫应答激活,产生严重的毒性,该方法更加安全可靠[12,13]。此外这种方法与利用病毒载体插入基因的方法相比,消除了基因整合和插入诱变的风险,也无需病毒载体,无需反复转染,效率更高,更为临床应用扫除了一大障碍。

图2 利用修饰mRNA对人成纤维细胞进行重编程[14]。

3.产品列表

厂商 货号 产品名称 规格 目录价(¥)
eBioscience 14-8185-62 B18R Recombinant Protein 5 μg 4548.00
eBioscience 34-8185-81 B18R Recombinant Protein Carrier-Free 50μg 6588.00
eBioscience 34-8185-85 B18R Recombinant Protein Carrier-Free 500μg 34188.00
eBioscience 34-8185-86 B18R Recombinant Protein Carrier-Free 1 mg 53988.00
Miltenyi 130-096-524 Stemgent Oct4 mRNA, human 20 μg 6250.00
Miltenyi 130-096-527 Stemgent Sox2 mRNA, human 20 μg 6250.00
Miltenyi 130-096-526 Stemgent Klf4 mRNA, human 20 μg 6250.00
Miltenyi 130-096-523 Stemgent c-Myc mRNA, human 20 μg 6250.00
Miltenyi 130-096-525 Stemgent Lin-28 mRNA, human 20 μg 6250.00

4.参考文献

[1]Colamonici OR, Domanski P, Sweitzer SM, et al. Vaccinia virus B18R gene encodes a type I interferon-binding protein that blocks interferon alpha transmembrane signaling.J Biol Chem. 1995, 270(27):15974-8.

[2]Vancová I, La Bonnardiere C, Kontsek P. Vaccinia virus protein B18R inhibits the activity and cellular binding of the novel type interferon-delta.J Gen Virol.1998, 79:1647-9.

[3] Liptáková H, Kontseková E, Alcamí A, et al. Analysis of an interaction between the soluble vaccinia virus-coded type I interferon (IFN)-receptor and human IFN-alpha1 and IFN-alpha2.Virology. 1997, 232(1):86-90.

[4] Alcamí A, Symons JA, Smith GL. The vaccinia virus soluble alpha/beta interferon (IFN) receptor binds to the cell surface and protects cells from the antiviral effects of IFN.J Virol. 2000, 74(23):11230-9.

[5] Elde NC, Malik HS.The evolutionary conundrum of pathogen mimicry.Nat Rev Microbiol. 2009, 7(11):787-97.

[6] Heng BC, Heinimann K, Miny P, et al. mRNA transfection-based, feeder-free, induced pluripotent stem cells derived from adipose tissue of a 50-year-old patient.Metab Eng. 2013, 18:9-24.

[7] Drews K, Tavernier G, Demeester J, et al. The cytotoxic and immunogenic hurdles associated with non-viral mRNA-mediated reprogramming of human fibroblasts.Biomaterials. 2012, 33(16):4059-68.

[8] Warren L, Manos PD, Ahfeldt T, et al. Highly efficient reprogramming to pluripotency and directed differentiation of human cells with synthetic modified mRNA.Cell Stem Cell. 2010, 7(5):618-30.

[9] Gore A, Li Z, Fung HL, et al. Somatic coding mutations in human induced pluripotent stem cells.Nature. 2011, 471(7336):63-7.

[10] Warren L, Ni Y, Wang J, et al. Feeder-free derivation of human induced pluripotent stem cells with messenger RNA.Sci Rep. 2012, 2:657-63.

[11] Tavernier G, Wolfrum K, Demeester J, et al. Activation of pluripotency-associated genes in mouse embryonic fibroblasts by non-viral transfection with in vitro-derived mRNAs encoding Oct4, Sox2, Klf4 and cMyc.Biomaterials. 2012, 33(2):412-7.

[12] Angel M, Yanik MF. Innate immune suppression enables frequent transfection with RNA encoding reprogramming proteins.PLoS One. 2010, 5(7):e11756.

[13] Anderson BR, Muramatsu H, Nallagatla SR,Incorporation of pseudouridine into mRNA enhances translation by diminishing PKR activation.Nucleic Acids Res. 2010, 38(17):5884-92.

[14] Mandal PK, Rossi DJ.Reprogramming human fibroblasts to pluripotency using modified mRNA.Nat Protoc. 2013, 8(3):568-82.