Glioma/glioblastoma-specific adenoviral gene expression using the nestin gene regulator.
For glioma- and glioblastoma-specific gene expression, we utilized a nestin regulatory element whose activity was evaluated by the reporter gene lacZ. Nestin is a 38-kDa intermediate filament protein, and is expressed specifically in the neuroepithelial stem cells. Nestin is detected in gliomas and glioblastomas, but not in normal brain tissue. We constructed a nestin gene regulator by placing nestin's second intron before the 5' upstream region (2iNP). To obtain enhanced expression of this tissue-specific regulator, we utilized the adenovirus double-infection method with a Cre-loxP on/off switching system. We constructed a 'regulator' vector, Ax2iNPNCre, which expresses Cre recombinase under the control of the nestin regulatory element, 2iNP. A 'reporter' vector, AxCALNLNZK, expresses lacZ under the control of a strong CAG promoter when the stuffer sequence has been removed by Cre recombinase at a pair of loxP sites. We used seven human glioma/glioblastoma cell lines: U251, KG-1C, NGM5, U87 MG, LN-Z308, NP-2 and T98G. Of these, nestin was expressed highly in U251 and KG-1C, less in NGM5, and undetectably in the other four lines. With the use of the two adenovirus vectors, we found X-gal staining and high nestin regulator-promoted beta-galactosidase activities in four of the seven glioma/glioblastoma cell lines. Staining was strong in U251, KG-1C and NGM5, and less in U87 MG. LacZ expression was nearly undetectable in the non-glioma cell line, HeLa, but a little in COS-7. The adenovirus double-infection method, which uses a nestin regulator, is applicable for glioma/glioblastoma-specific expression.
Kurihara
H
H
Department of Neurosurgery, Gunma University School of Medicine, Japan.
Zama
A
A
Tamura
M
M
Takeda
J
J
Sasaki
T
T
Takeuchi
T
T
D000256:Adenoviridae / Q000235:genetics*; D001932:Brain Neoplasms / Q000628:therapy*; D015870:Gene Expression; D015316:Genetic Therapy / Q000379:methods*; D005822:Genetic Vectors / Q000008:administration & dosage*; D005909:Glioblastoma / Q000628:therapy; D005910:Glioma / Q000628:therapy*; D006801:Humans; D007381:Intermediate Filament Proteins / Q000235:genetics*; D007763:Lac Operon; D009419:Nerve Tissue Proteins; D064231:Nestin; D014407:Tumor Cells, Cultured
D007381:Intermediate Filament Proteins; C577484:NES protein, human; D009419:Nerve Tissue Proteins; D064231:Nestin