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. 2003 Oct 18;3:29. doi: 10.1186/1471-230X-3-29

Figure 1.

Figure 1

Adenoviral expression and antifibrotic capacity of an AS-TGF-β1 mRNA in culture-activated HSC. (A) Schematic representation of Ad5-CMV-AS-TGF-β1 [17] and Ad5-CMV-EGFP [22] expressing their transgene under regulatory control of the human cytomegalovirus immediate-early gene 1 promoter (CMV). A SV40 polyadenylation signal (pA) downstream of each transgene directs the proper processing of the 3' end of the corresponding mRNA. (B) Northern blot analysis from untreated HSC (1), and HSC infected with Ad5-CMV-EGFP (2) or Ad5-CMV-AS-TGF-β1 (3). The blot was subsequently hybridized with 32P-labelled probes specific for TGF-β1, colα1(I), and GAPDH. (C) Strategy for relative quantification of transcripts specific for TGF-β1 or AS-TGF-β1. Primer P1 and P2 specific for endogenous TGF-β1, or primer P3 and P4 specific for both, endogenous TGF-β1 and AS-TGF-β1, were used in RT-PCR. The assay was conducted in a LightCycler system. (D, E) RNA isolated from untreated HSC (1), and HSC infected with Ad5-CMV-EGFP (2) or Ad5-CMV-AS-TGF-β1 (3) was assayed in duplicate for endogenous TGF-β1 transcripts (D) or alternatively for TGF-β1 and AS-TGF-β1 (E) using primer combinations P1 and P2 (D) or P3 and P4 (E), respectively. Note, that small amounts of artificially synthesized DNA at higher cycle numbers occurring in the no-template control are due to primer dimer formation (4).