Retrovirus-mediated gene transfer of transthyretin and transthyretin-methionine 30: a potential tool for the study of amyloidogenesis.
Familial amyloidotic polyneuropathy (FAP) is a genetic disease characterized by systemic amyloid deposition particularly in the peripheral nervous system. These deposits are composed mainly of a mutant form of the serum protein transthyretin (TTR) having a methionine for valine substitution at position 30-TTR Met 30. The factors involved in the formation of these deposits are unknown. The existence of animal models for FAP should allow elucidation of these factors. As one approach to the development of animal models for amyloidogenesis in FAP, we have constructed recombinant retrovirus vectors, carrying the full length human cDNA for either TTR or TTR Met 30 under the control of the Moloney murine leukemia virus (MoMLV) LTR element. After transfection of the packaging cell line, psi 2, viral stocks were used to infect a rat hepatoma cell line, H56, mouse fibroblast cell line, NIH3T3, and mouse primary fibroblasts. H56 cells efficiently secreted both TTR and TTR Met 30 as assessed by immunoprecipitation and ELISA, whereas NIH3T3 fibroblasts appeared not to release these proteins under the conditions tested. Primary fibroblasts secreted the mutant protein as assessed by ELISA. These genetically modified cells can be grafted into animals for in vivo study of amyloidogenesis, as well as be used in culture to investigate factors that might regulate the rate of amyloid deposition.
Bonifácio
M J
MJ
Centro de Estudos de Paramiloidose, Hospital de Santo António, Porto, Portugal.
Ezzeddine
D
D
Sakaki
Y
Y
Breakfield
X O
XO
Saraiva
M J
MJ
D016475:3T3 Cells; D000682:Amyloid / Q000096:biosynthesis*; D000686:Amyloidosis / Q000235:genetics / Q000378:metabolism*; D000818:Animals; D001483:Base Sequence; D006528:Carcinoma, Hepatocellular; D002460:Cell Line; D004247:DNA / Q000302:isolation & purification / Q000378:metabolism; D018014:Gene Transfer Techniques; D005822:Genetic Vectors; D006801:Humans; D008113:Liver Neoplasms; D008715:Methionine; D051379:Mice; D008969:Molecular Sequence Data; D008970:Molecular Weight; D008979:Moloney murine leukemia virus; D009838:Oligodeoxyribonucleotides; D010957:Plasmids; D017354:Point Mutation; D016133:Polymerase Chain Reaction; D011228:Prealbumin / Q000096:biosynthesis* / Q000235:genetics; D051381:Rats; D012091:Repetitive Sequences, Nucleic Acid; D012190:Retroviridae; D014407:Tumor Cells, Cultured
D000682:Amyloid; D009838:Oligodeoxyribonucleotides; D011228:Prealbumin; D004247:DNA; D008715:Methionine