title: RNA interference of PPARgamma using fiber-modified adenovirus vector efficiently suppresses preadipocyte-to-adipocyte differentiation in 3T3-L1 cells.
abstract: The peroxisome proliferator-activated receptor (PPAR) gamma is regarded as a "master regulator" of adipocyte differentiation and is abundantly expressed in adipose. To understand the biological role of PPARgamma in adipose, RNA interference (RNAi) of PPARgamma should be a powerful tool. 3T3-L1 cell line serves an excellent model to investigate the mechanism of preadipocyte-to-adipocyte differentiation. However, this cell line is difficult to transfect by plasmid vectors and viral vectors. We optimized the transduction of both 3T3-L1 preadipocytes and adipocytes by means of fiber-modified adenovirus (Ad) vectors. Among the various vectors tested, polylysine modification of the C-terminal of the fiber knob most markedly improved the transduction efficiency in both 3T3-L1 preadipocytes and adipocytes. Then, we examined whether fiber-modified Ad vectors with polylysine peptides expressing the small interfering RNA (siRNA) for PPARgamma inhibit the differentiation of 3T3-L1 preadipocytes into adipocytes. Oil red O staining and measurement of glycerol-3-phosphate dehydrogenase (GPDH) activity indicated that the vectors effectively suppressed the differentiation of 3T3-L1 preadipocytes to adipocytes. These results suggested that the combination of fiber-modified Ad vectors containing polylysine peptides and RNAi is an effective tool for the study of the biological and physiological mechanism of adipogenesis in adiposity and diabetes using 3T3-L1 models. Ad vector-mediated RNAi for PPARgamma should also be useful to clarify the biological role of the PPARgamma pathway in various tissues in addition to adipose and for therapeutic application to a variety of diseases, including adiposity and diabetes.
authors.affiliation: Division of Cellular and Gene Therapy Products, National Institute of Health Sciences, Tokyo 158-8501, Japan.
authors.forename: Tetsuji
authors.initials: T
authors.lastname: Hosono
authors.forename: Hiroyuki
authors.initials: H
authors.lastname: Mizuguchi
authors.forename: Kazufumi
authors.initials: K
authors.lastname: Katayama
authors.forename: Naoya
authors.initials: N
authors.lastname: Koizumi
authors.forename: Kenji
authors.initials: K
authors.lastname: Kawabata
authors.forename: Teruhide
authors.initials: T
authors.lastname: Yamaguchi
authors.forename: Shinsaku
authors.initials: S
authors.lastname: Nakagawa
authors.forename: Yoshiteru
authors.initials: Y
authors.lastname: Watanabe
authors.forename: Tadanori
authors.initials: T
authors.lastname: Mayumi
authors.forename: Takao
authors.initials: T
authors.lastname: Hayakawa
mesh_terms: D041721:3T3-L1 Cells; D000256:Adenoviridae / Q000235:genetics; D017667:Adipocytes / Q000166:cytology / Q000378:metabolism*; D000818:Animals; D036022:Capsid Proteins / Q000235:genetics; D002454:Cell Differentiation / Q000235:genetics / Q000502:physiology*; D002460:Cell Line; D062727:Coxsackie and Adenovirus Receptor-Like Membrane Protein; D015870:Gene Expression; D005822:Genetic Vectors / Q000235:genetics; D006801:Humans; D051379:Mice; D041681:NIH 3T3 Cells; D047495:PPAR gamma / Q000235:genetics / Q000378:metabolism*; D034622:RNA Interference / Q000502:physiology*; D012333:RNA, Messenger / Q000235:genetics / Q000378:metabolism; D034741:RNA, Small Interfering / Q000235:genetics / Q000378:metabolism; D011991:Receptors, Virus / Q000235:genetics; D020133:Reverse Transcriptase Polymerase Chain Reaction; D014162:Transfection
chemical_list: C568771:CLMP protein, human; C568772:CLMP protein, mouse; D036022:Capsid Proteins; D062727:Coxsackie and Adenovirus Receptor-Like Membrane Protein; D047495:PPAR gamma; D012333:RNA, Messenger; D034741:RNA, Small Interfering; D011991:Receptors, Virus; C018315:hexon capsid protein, Adenovirus