Each probe was then labeled with [35S]-UTP using the Riboprobe Transcription System (Promega, Madison, WI, USA). P7.0, both mRNAs were weakly expressed but maintained their spatial expression patterns. Immunostaining showed that biglycan was localized in the dental papillae and pulp. In addition, all four SLRPs showed obvious immunostaining in predentin, even Givinostat though expressions of Edg3 and mRNAs were not recognized in the tooth germs examined. The organ culture data obtained supported the histological findings that biglycan is usually more predominant than decorin at the apposition stage. These results were used to identify biglycan as the principal molecule among the SLRPs investigated. Our findings show that decorin and biglycan show spatial and temporal differential expressions and play their own tissue-specific functions in tooth development. hybridization, organ culture Introduction Small leucine-rich proteoglycans (SLRPs) have Givinostat been identified as important components in the extracellular matrix (ECM) and are involved in several biological and pathological processes in various tissues. They are proteoglycans (PGs) with small core proteins of tandem leucine-rich repeats that carry one or more glycosamino – glycan (GAG) chains, such as chondroitin sulfate (CS), dermatan sulfate (DS), or keratan sulfate (KS). Eighteen SLRP genes have been identified to date and are classified into five groups based on their characteristics at both the genomic and protein levels.1-3 It has been reported that these SLRPs play significant functions in regulating the ECM by collagen formation and ECM arrangement, matrix mineralization, transmission transduction, and epithelialmesenchymal interactions, as well as in immunity and tumor growth.4 The most commonly identified SLRPs in teeth are of class I and class II.5 Class I SLRPs include decorin and biglycan, which are the most analyzed and are relatively abundant. They are found in various tissues, Givinostat such as skin, tendon, bone, cartilage, and muscle mass.6 They contain CS/DS chains; one in decorin and two in biglycan. Decorin plays a significant role in collagen fibrillogenesis and in preventing premature mineralization,7 whereas biglycan is usually more significant in osteoblast differentiation Givinostat and matrix mineralization.8 In geneknockout studies, the absence of gives rise to skin fragility and dysregulation of lateral fibril growth.9 Likewise, the absence of results in osteoporosis-like phenotypes.10 Class II SLRPs include fibromodulin and lumican, which are close relatives of decorin and biglycan in terms of their function. They have KS chains and are found in tissues such as tendons, cartilage, sclera, and the cornea.6 Goldberg has been found to affect dentin mineralization, whereas biglycan has been shown to have a multifunctional involvement, the knockout of which affects both amelogenesis and dentinogenesis, together with collagen fibrillogenesis in predentin.25 deficiency has been found to impact enamel formation and increases the collagen diameter in predentin, as well as causing dentin hypomineralization in newborn mice, in contrast to the self-repair observed in adult mice.11,12 deficiency results in periodontal ligament phenotypes.26 Many immunohistochemical studies in tooth germs have also been performed in rodents21,27 and humans.13,20 Several hybridization studies in mice have also been carried out. 28-30 These studies, however, have not examined the temporal changes in expression during the process of identical tooth germ formation. Thus, our hypothesis that SLRPs are closely involved in tooth formation led us to perform an hybridization study to elucidate their specific functions by investigating the formation of identical tooth.
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