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Characterization of Thrombospondin Type 1 Repeat in Haliotis diversicolor and Its Possible Role in Osteoinduction

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Abstract

Thrombospondin repeats (TSR) are important peptide domains present in the sequences of many extracellular and transmembrane proteins with which a variety of ligands interact. In this study, we characterized HdTSR domains in the ADAMTS3 protein of Thai abalone, Haliotis diversicolor, based on the transcriptomic analysis of its mantle tissues. PCR amplification and localization studies demonstrated the existence of HdTSR transcript and protein in H. diversicolor tissues, particularly in both the inner and outer mantle epithelial folds. We, therefore, generated a short recombinant protein, termed HdTSR1/2, based on the existence of the WxxWxxW or WxxxxW motif (which binds to TGF-β, a known signaling in bone formation/repair) in HdTSR1 and HdTSR2 sequences and used it to test the osteoinduction function in the pre-osteoblastic cell line, MC3T3-E1. This recombinant protein demonstrated the ability to induce the differentiation of MC3T3-E1 cells by the concentration- and time-dependent upregulation of many known osteogenic markers, including RUNX2, COL1A1, OCN, and OPN. We also demonstrated the upregulation of the SMAD2 gene after cell treatment with HdTSR1/2 proteinindicating its possible interaction through TGF-β, which thus activates its downstream signaling cascade and triggers the biomineralization process in the differentiated osteoblastic cells. Together, HdTSR domains existed in an extracellular ADAMTS3 protein in the mantle epithelium of H. diversicolor and played a role in osteoinduction as similar to the other nacreous proteins, opening up its possibility to be developed as an inducing agent of bone repair.

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Acknowledgements

We thank the Center of Nanoimaging (CNI) and the Central Instrument Facility (CIF), Faculty of Science, Mahidol University, for providing instrumental support throughout this work.

Funding

This research project is supported by Mahidol University (to SA).

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Author notes

  1. Valainipha Habuddha and Chanyatip Suwannasing are co-first authors of this paper.

Authors and Affiliations

  1. Department of Anatomy, Faculty of Science, Mahidol University, Bangkok, Thailand

    Valainipha Habuddha, Aticha Buddawong, Chanyarak Sombutkayasith, Wattana Weerachatyanukul & Somluk Asuvapongpatana

  2. School of Allied Health Science, Walailak University, Nakhon Si Thammarat, Thailand

    Valainipha Habuddha

  3. Department of Radiological Technology, Faculty of Allied Health Sciences, Naresuan University, Phitsanulok, Thailand

    Chanyatip Suwannasing

  4. Chulabhorn International College of Medicine, Thammasat University, Khlong Nueng Pathumthani 12121, Rangsit Campus, Thailand

    Aticha Buddawong

  5. Department of Clinical Microscopy, Faculty of Medical Technology, Mahidol University (Salaya Campus), Salaya, Nakhonpathom, Thailand

    Kanokwan Seenprachawong, Thitinat Duangchan & Aungkura Supokawej

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  1. Valainipha Habuddha
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Correspondence to Somluk Asuvapongpatana.

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The study was carried out in accordance with the protocol approved by the Animal Care Committee, Faculty of Science, Mahidol University (Protocol # MUSC-60–040-390).

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Habuddha, V., Suwannasing, ., Buddawong, A. et al. Characterization of Thrombospondin Type 1 Repeat in Haliotis diversicolor and Its Possible Role in Osteoinduction. Mar Biotechnol 23, 641–652 (2021). https://doi.org/10.1007/s10126-021-10054-3

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