RNA interference in human foreskin fibroblasts within the three-dimensional collagen matrix.
The technique of RNA interference (RNAi) was trialed in primary human foreskin fibroblasts, both in monolayer culture and in the fibroblast-populated collagen matrix. Knockdown of lamin A/C, p53, and FAK was possible with low-confluency (<50%) monolayer fibroblasts, a transfection vehicle concentration of 1%, and an siRNA concentration of 25-50 nM. Knockdown also was possible in the collagen matrix using similar reagent concentrations and a cellular density of one million fibroblasts per ml of matrix. Optimization of transfection conditions appeared to be important to increase knockdown efficiency. Consistent with prediction, knockdown of FAK induced apoptosis in the fibroblast-populated collagen matrix.
Department of Surgery, VA Nebraska-Western Iowa Health Care System, Surgery 112, VA Medical Center, 4101 Woolworth Avenue, Omaha, NE 68105, USA. macarlso@unmc.edu
Mark A
MA
Carlson
Amy K
AK
Prall
Jeremiah J
JJ
Gums
D002478:Cells, Cultured; D003094:Collagen / Q000378:metabolism*; D005109:Extracellular Matrix / Q000378:metabolism*; D005347:Fibroblasts / Q000378:metabolism*; D005455:Fluorescent Antibody Technique; D051417:Focal Adhesion Kinase 1 / Q000037:antagonists & inhibitors / Q000235:genetics / Q000378:metabolism*; D052816:Foreskin / Q000166:cytology*; D006801:Humans; D007668:Kidney / Q000378:metabolism; D034904:Lamin Type A / Q000037:antagonists & inhibitors / Q000235:genetics / Q000378:metabolism*; D008297:Male; D034622:RNA Interference; D034741:RNA, Small Interfering / Q000494:pharmacology; D016159:Tumor Suppressor Protein p53 / Q000037:antagonists & inhibitors / Q000235:genetics / Q000378:metabolism*
D034904:Lamin Type A; D034741:RNA, Small Interfering; D016159:Tumor Suppressor Protein p53; C440308:lamin C; D003094:Collagen; D051417:Focal Adhesion Kinase 1