Particle-Mediated Gene Transfer of PDGF Isoforms Promotes Wound Repair

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Particle-Mediated Gene Transfer of PDGF Isoforms Promotes Wound Repair Sabine A. Eming, Jeffrey S. Whitsitt, Lan He, Thomas Krieg, Jeffrey R. Morgan, Jeffrey M. Davidson Journal of Investigative Dermatology Volume 112, Issue 3, Pages 297-302 (March 1999) DOI: 10.1046/j.1523-1747.1999.00522.x Copyright © 1999 The Society for Investigative Dermatology, Inc Terms and Conditions

Figure 1 Plasmid constructs. MFG-PDGF-A, MFG-PDGF-B, MFG-PDGF-B211, and the cytomegalovirus promoter luciferase construct. LTR, long-terminal repeat: SD, splice donor; SA, splice acceptor;ψ+, packaging signal. Journal of Investigative Dermatology 1999 112, 297-302DOI: (10.1046/j.1523-1747.1999.00522.x) Copyright © 1999 The Society for Investigative Dermatology, Inc Terms and Conditions

Figure 2 Transgenes are expressed in vivo after particle bombardment. The unwounded dorsal skin of three rats was transfected with MFG-PDGF-A and cytomegalovirus-luciferase (–) expression constructs. To follow transgene expression total RNA was isolated at days 1, 3, and 5, after transfection, and subjected to reverse transcriptase-PCR using PDGF-A specific oligonucleotide primers. One sample was analyzed at each time point. (A) The PCR product of both species is 510 bp. In the human this product is cleaved by Sal1 into 316 bp and 194 bp products. (B) The digestion products of the reverse transcriptase-PCR reaction were analyzed by gel electrophoresis and detected by Southern blotting with a 32P-labeled human PDGF-A cDNA. The endogenous rat PDGF-A reaction product is detected at 510 bp, whereas the cleaved human products migrate at 316 bp and 194 bp. (C) The intensity of the human-specific transgene signal was quantitated by scanning densitometry, and is presented as a function of days after transfection.[TAB] Journal of Investigative Dermatology 1999 112, 297-302DOI: (10.1046/j.1523-1747.1999.00522.x) Copyright © 1999 The Society for Investigative Dermatology, Inc Terms and Conditions

Figure 3 Wound tensile strength increases after particle mediated transfection with PDGF-A, PDGF-B, and PDGF-B211 expression constructs. The dorsal skin of rats (5.5 mo at day 7 and 3.5 mo at day 14) was transfected with PDGF-A, PDGF-B, PDGF-B211 (experimental) and luciferase (control) expression constructs, after which full thickness incisional wounds were made across the targeted tissue sites. Individual sets of wounds compared either PDGF-A (day 7, n = 7; day 14, n = 10) and PDGF-B (day 7, n = 8; day 14, n = 10) or PDGF-B and PDGF-B211 with controls (n = 8 in all groups). The animals were sacrificed on the day indicated, and wound tensile strength was measured as described in Materials and Methods. The results are presented as ratios (experimental/control) of tensile strengths measured in the wounds transfected with PDGF expression constructs to those in incisional wounds transfected with a cytomegalovirus-luciferase construct for each animal and each location (anterior or posterior). The horizontal bars represent the means of the experimental/control ratios within each group of animals. The Student’s t-test was used to calculate significant differences of paired values. Journal of Investigative Dermatology 1999 112, 297-302DOI: (10.1046/j.1523-1747.1999.00522.x) Copyright © 1999 The Society for Investigative Dermatology, Inc Terms and Conditions