title: Restoration of the balanced alpha/beta-globin gene expression in beta654-thalassemia mice using combined RNAi and antisense RNA approach.
abstract: The beta-thalassemia is associated with abnormality in beta-globin gene, leading to imbalanced synthesis of alpha-/beta-globin chains. Consequently, the excessive free alpha-globin chains precipitate to the erythrocyte membrane, resulting in hemolytic anemia. We have explored post-transcriptional strategies aiming at alpha-globin reduction and beta-globin enrichment on beta(654) (Hbb(th-4)/Hbb(+)) mouse, carrying a human splicing-deficient beta-globin allele (Hbb(th-4)). Lentiviral vectors of short hairpin RNA (shRNA) targeting alpha-globin and/or antisense RNA facilitating beta-globin correct splicing were microinjected into beta(654) single-cell embryos. Three transgenic strains were generated, as alpha(i)-Hbb(th-4)/Hbb(+)(shRNA), beta(a)-Hbb(th-4)/Hbb(+)(antisense) and alpha(i)beta(a)-Hbb(th-4)/Hbb(+)(both shRNA and antisense). Without notable abnormalities, all the founders and their offsprings showed sustained amelioration of hematologic parameters, ineffective erythropoiesis and extramedullary hematopoiesis. Augmented effects appeared in alpha(i)beta(a)-Hbb(th-4)/Hbb(+), which correlated with a better-balanced alpha-/beta-globin mRNA level. Among the transgenic mice integrated with shRNA and antisense RNA, one homozygous mouse (Hbb(th-4)/Hbb(th-4)) had been viable, and the 3-week survival rate for heterozygotes (Hbb(th-4)/Hbb(+)) was 97%, compared with 45.4% for untreated. Our data have demonstrated the feasibility of techniques for beta-thalassemia therapy by balancing the synthesis of alpha-/beta-globin chains.
authors: Shanghai Institute of Medical Genetics, Shanghai Children's Hospital, Shanghai Jiao Tong University School of Medicine, Shanghai, People's Republic of China.
authors: Shu-Yang
authors: SY
authors: Xie
authors: Zhao-Rui
authors: ZR
authors: Ren
authors: Jing-Zhi
authors: JZ
authors: Zhang
authors: Xin-Bin
authors: XB
authors: Guo
authors: Qing-Xue
authors: QX
authors: Wang
authors: Shu
authors: S
authors: Wang
authors: Dan
authors: D
authors: Lin
authors: Xiu-Li
authors: XL
authors: Gong
authors: Wei
authors: W
authors: Li
authors: Shu-Zhen
authors: SZ
authors: Huang
authors: Fanyi
authors: F
authors: Zeng
authors: Yi-Tao
authors: YT
authors: Zeng
mesh_terms: D000740:Anemia / Q000628:therapy; D000818:Animals; D001853:Bone Marrow / Q000473:pathology; D004920:Erythropoiesis / Q000235:genetics; D015870:Gene Expression; D015316:Genetic Therapy / Q000379:methods*; D005822:Genetic Vectors / Q000235:genetics; D005914:Globins / Q000037:antagonists & inhibitors* / Q000235:genetics; D006454:Hemoglobins / Q000235:genetics; D006801:Humans; D016086:Lentivirus / Q000235:genetics; D051379:Mice; D008822:Mice, Transgenic; D010641:Phenotype; D012326:RNA Splicing / Q000235:genetics; D016372:RNA, Antisense / Q000235:genetics*; D034741:RNA, Small Interfering / Q000235:genetics*; D017086:beta-Thalassemia / Q000628:therapy*
chemical_list: D006454:Hemoglobins; D016372:RNA, Antisense; D034741:RNA, Small Interfering; D005914:Globins