Vol.:(0123456789) 1 3 Anatomical Science International https://doi.org/10.1007/s12565-020-00588-2 ORIGINAL ARTICLE An in situ hybridization study of decorin and biglycan mRNA in mouse osteoblasts in vivo Angammana Randilini 1  · Kaoru Fujikawa 2  · Shunichi Shibata 1 Received: 26 August 2020 / Accepted: 4 November 2020 © Japanese Association of Anatomists 2020 Abstract In situ hybridization of decorin and biglycan mRNA, principal members of small leucine-rich proteoglycan, was performed using [ 35 S]-labeled RNA probes, in the context of the hypothesis that they show diferent expression patterns associated with osteoblast diferentiation in mice. We adopted two ossifying sites that can clearly follow the developmental process of bone formation: ossifying tympanic ring and developing bone collar of mandibular condylar cartilage. Decorin mRNA was expressed in osteoblasts of developing tympanic ring at E14.0, as well as of developing bone collar at E15.0, but biglycan mRNA was not, indicating decorin mRNA was expressed earlier in newly diferentiating osteoblasts than biglycan. With maturation of osteoblasts, biglycan mRNA became expressed and maintained its expression both in the outer region (peri- osteum) and in the interior region (endosteum) of bone. By contrast, decorin mRNA expression was maintained in the outer region but diminished in the interior region. These results indicate that decorin and biglycan show diferential expression patterns in diferentiating osteoblasts and play specifc roles in bone formation. Keywords Biglycan · Bone formation · Decorin · In situ hybridization · Osteoblasts Introduction Small leucine-rich proteoglycans (SLRPs) have been iden- tifed as important components in the extracellular matrix and are involved in several biological and pathological processes in various tissues. They are proteoglycans with small core proteins of tandem leucine-rich repeats that are usually attached to one or more glycosaminoglycan chains, such as chondroitin sulfate (CS), dermatan sulfate (DS), or keratan sulfate. Eighteen SLRP genes have been identifed to date, which are classifed into fve classes based on their characteristics at both genomic and protein levels (Schaefer and Iozzo 2008; Schaefer and Schaefer 2010; Nikitovic et al. 2012; Kram et al. 2020). Class I SLRPs are most commonly identifed and include decorin and biglycan, which are the most studied and relatively abundant. These SLRPs are found in various tissues, such as skin, tendon, bone, carti- lage, muscle, and teeth (Neame and Kay 2000). They contain CS/DS chains: one in decorin and two in biglycan. Decorin plays a signifcant role in collagen fbrillogenesis and pre- vention of premature mineralization (Mochida et al. 2009), whereas biglycan is more signifcant in osteoblast diferen- tiation and matrix mineralization (Parisuthiman et al. 2005). In gene-knockout studies, the absence of decorin results in skin fragility and dysregulation of lateral fbril growth (Corsi et al. 2002; Danielson et al. 1997), while the absence of biglycan induces osteoporosis-like phenotypes (Xu et al. 1998; Young et al. 2002). We have recently investigated expression, localization, and synthesis of decorin and biglycan in developing mouse molar tooth germ and demonstrated that decorin mRNA was expressed earlier in newly diferentiating odontoblasts than biglycan. Furthermore, with odontoblast maturation, decorin mRNA expression was diminished but biglycan mRNA extended its expression throughout new and mature odontoblasts (Randilini et al. 2020). These results led us to hypothesize that a similar phenomenon may occur in osteo- blasts, that is, decorin mRNA may be expressed earlier in newly diferentiating osteoblasts than biglycan mRNA. * Shunichi Shibata sshibata.mfa@tmd.ac.jp 1 Department of Maxillofacial Anatomy, Graduate School of Medical and Dental Sciences, Tokyo Medical and Dental University, 1-5-45, Bunkyo-ku, Yushima, Tokyo 113-8549, Japan 2 Department of Oral Anatomy and Developmental Biology, Showa University School of Dentistry, Tokyo, Japan