Antisense oligonucleotides reduce synthesis of procollagen alpha1 (V) chain in human patellar tendon fibroblasts: potential application in healing ligaments and tendons.
Many ligaments and tendons will heal after injury. However, they heal with poor mechanical properties when compared with the native tissue and show no improvement of these properties with time. Although the mechanisms that lead to this process are poorly understood, the presence of uniformly smaller collagen fibrils is believed to play a major role. Quantitatively minor when compared with type I collagen, type V collagen was found to be significantly elevated in healing medial collateral ligament of the rabbit knee. Previous studies have shown that type V collagen plays a role in regulating the diameter of type I collagen fibrils and reducing its level may lead to the formation of larger collagen fibrils in healing ligaments. Hence, type V collagen antisense gene therapy may be an approach to obtain this goal. In this study, our objective was to find specific antisense oligonucleotide sequences for type V procollagen alpha1 chain to elucidate the feasibility of type V collagen antisense gene therapy in ligaments or tendons. We hypothesized that antisense oligonucleotides that selectively target the type V procollagen alpha1 chain mRNA could partially reduce the synthesis of type V procollagen alpha1 chain in human tendon fibroblasts. Western blotting analysis showed that antisense oligonucleotides (AS-V1 and AS-V2) significantly reduced synthesis of type V procollagen alpha1 chain. In addition, reverse transcription polymerase chain reaction revealed that both antisense oligonucleotides partially reduced type V procollagen alpha1 chain mRNA expression. This experiment identified two sequences within the type V procollagen coding region that are susceptible to antisense suppression, and thus provide the basis to explore the effects of antisense oligonucleotides on type V collagen synthesis, collagen fibril diameter, and mechanical properties of healing tendons and ligaments.
D001483:Base Sequence / Q000235:genetics; D024042:Collagen Type I / Q000378:metabolism; D024062:Collagen Type V / Q000037:antagonists & inhibitors* / Q000096:biosynthesis / Q000235:genetics; D015536:Down-Regulation / Q000187:drug effects / Q000235:genetics; D005240:Feasibility Studies; D015316:Genetic Therapy / Q000379:methods; D006801:Humans; D016376:Oligonucleotides, Antisense / Q000235:genetics / Q000494:pharmacology* / Q000627:therapeutic use; D016366:Open Reading Frames / Q000235:genetics; D017847:Patellar Ligament / Q000166:cytology / Q000254:growth & development* / Q000378:metabolism; D011347:Procollagen / Q000037:antagonists & inhibitors* / Q000096:biosynthesis / Q000235:genetics; D014176:Protein Biosynthesis / Q000235:genetics; D012333:RNA, Messenger / Q000037:antagonists & inhibitors / Q000235:genetics / Q000378:metabolism; D012038:Regeneration / Q000235:genetics*; D013710:Tendons / Q000166:cytology / Q000254:growth & development* / Q000378:metabolism; D014945:Wound Healing / Q000235:genetics*
D024042:Collagen Type I; D024062:Collagen Type V; D016376:Oligonucleotides, Antisense; D011347:Procollagen; D012333:RNA, Messenger