Cloning and expression of equine insulin-like growth factor binding proteins in normal equine tendon.
Abstract: To define a portion of the nucleotide sequences of each of the 6 insulin-like growth factor (IGF) binding proteins (IGFBPs) in horses and describe patterns of messenger RNA (mRNA) and protein expression for IGFBPs in normal equine tendons. Methods: 7 horses. Methods: Total RNA was extracted from the tensile region of normal superficial digital flexor tendons and reverse transcribed into complimentary DNA (cDNA). The cDNA was amplified via PCR, and products representing portions of each IGFBP were cloned and sequenced. Nucleotide sequences were used to deduce the amino acid sequences, and both nucleotide and predicted amino acid sequences were compared with those published for bovine, human, mouse, and ovine IGFBPs. Gene expression was quantitated by real-time PCR assay, and protein expression was evaluated by western ligand blot (WLB). Results: Clones ranged in size from 262 to 522 bp and had high degrees of sequence homology with other mammalian species. Sequence homology was highest between bovine and equine IGFBPs (86% to 95%) and amongst the IGFBP-5 sequences from the various species (92% to 95%). Message for IGFBP-2 to -6, but not IGFBP-1, was expressed in normal tendon. Protein expression for IGFBP-2, -3, and -4 was detected byWLB in normal tendon and markedly increased in damaged tendons. Conclusions: Results provide basic information and tools needed for further characterization of the role of the IGF system in tendon healing and may lead to the ability to potentiate the response of healing tendon to exogenous IGF-I via concurrent manipulation of IGFBPs.
Publication Date: 2005-03-11 PubMed ID: 15757131DOI: 10.2460/ajvr.2005.66.300Google Scholar: Lookup
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- Journal Article
- Research Support
- U.S. Gov't
- P.H.S.
Summary
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The abstract details a study in which researchers cloned and analyzed sequences of insulin-like growth factor binding proteins (IGFBPs) in horses to understand their contribution to normal tendon function. They discovered high sequential similarities with other species, specifically for IGFBP-5, and detected protein expression for IGFBP-2, -3, and -4 in damaged tendons, implying relevance in the healing process of tendons.
Cloning and Sequencing of IGFBPs
- The researchers extracted total RNA from the tense region of healthy superficial digital flexor tendons in horses.
- The RNA was transformed into complementary DNA (cDNA) using reverse transcription.
- The cDNA was amplified through Polymerase Chain Reaction (PCR). This yielded products representing portions of each IGFBP, which were then cloned and sequenced.
- The nucleotide sequences from the cloning process were used to deduce amino acid sequences.
- Both the nucleotide and deduced amino acid sequences were compared with published IGFBPs sequences from various mammal species, including bovines, humans, mice, and sheep.
Analysis of Homology and Expression
- Clones ranged in size from 262 to 522 base pairs and showed high degrees of sequence homology (similarity) with the published IGFBPs of other mammalian species.
- The highest sequence homology was found between the horse and bovine IGFBPs (86% to 95%) and specifically amongst the IGFBP-5 sequences from the various species (92% to 95%).
- The research confirmed the expression of the message for IGFBP-2 to -6, but not for IGFBP-1, in normal tendon tissues.
- Protein expression for IGFBP-2, -3, and -4 was detected in normal tendons using a method called western ligand blot (WLB) and was found to be markedly increased in damaged tendons.
Conclusions and Future Implications
- The results provide essential understanding of the insulin-like growth factor (IGF) system in relation to tendon healing.
- This can potentially lead to the development of a method for enhancing the response of a healing tendon to exogenous IGF-I (a type of growth factor) by concurrently manipulating IGFBPs.
Cite This Article
APA
Dahlgren LA, Nixon AJ.
(2005).
Cloning and expression of equine insulin-like growth factor binding proteins in normal equine tendon.
Am J Vet Res, 66(2), 300-306.
https://doi.org/10.2460/ajvr.2005.66.300 Publication
Researcher Affiliations
- Comparative Orthopaedics Laboratory, Department of Clinical Sciences, College of Veterinary Medicine, Cornell University, Ithaca, NY 14853-6401, USA.
MeSH Terms
- Amino Acid Sequence
- Animals
- Base Sequence
- Blotting, Western / veterinary
- Cloning, Molecular
- DNA Primers / chemistry
- DNA, Complementary / genetics
- Gene Expression
- Horses
- Insulin-Like Growth Factor Binding Proteins / genetics
- Insulin-Like Growth Factor Binding Proteins / metabolism
- Molecular Sequence Data
- RNA, Messenger / metabolism
- Reverse Transcriptase Polymerase Chain Reaction / veterinary
- Sequence Homology, Nucleic Acid
- Tendons / metabolism
Grant Funding
- AR08587 / NIAMS NIH HHS
Citations
This article has been cited 2 times.- Miescher I, Rieber J, Calcagni M, Buschmann J. In Vitro and In Vivo Effects of IGF-1 Delivery Strategies on Tendon Healing: A Review. Int J Mol Sci 2023 Jan 25;24(3).
- Chen Y, Zhu Y, Song S, Hu Y. Contribution of insulin‑like growth factor‑1 to tendon repair (Review). Int J Mol Med 2025 Nov;56(5).
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