CD. {:em)· R. liwiii!WrSiriim L, Slal11 • /. Sc'qumlialnpret\1011 oj an amelin gene in ;lle\·cnch) 'lllal and epithelial cells during odmrtogel/c. \i\ in rtlls. Eur J Oral Sci /99H; f(}fJ (suppl /): 324 330. < ur J Oral ci, 1998 'ovcl mR 1 oforms cncodmg the enamel matrix proteins amclin-1. amclm-2 and ameloblastm have been recently des(;nbed . We ha\e applied <.!<.:tailed immunohistochemical as well as non-radioactive in .1itu hybriditatwn anal)~s to I oliO\\ am lin-I expression m developing rat incisors and molar ... \\ e constructed an expression vector overproduced recombinant amchn in Ellhcrtrhia coli and prcp;tred ttn ant1bod>. In addition to the pre\iou ly reported <~mel in mR NA cxpre~~10n patterns 111 umelobla-.b. the amclin mess,tgc \HIS also detected in pulpal mesenchymal cells mcludmg preodontoblasts and young odontohla.ts. 1 he signal il1 these cells r ·rsisted until deposition or mantle dentin bccam~: evidenl. rhe immunolocalmttion of amt:lin-1 in pr l>dontobla~ts und mn~:lnblasts cssent1,tll> followed the path:rn of mR. cxpre am. 1 he most intense sta1n1n • was f'ound 111 the enamel matrix atiJ&H:cnt to crctor> amcloblasts. Focal accumtdalion~ of 1mmu1wrcact1ve material \\ere found ill the dcnt•no~:namcl JUI1Ctlon durinv th~: muturatwn tag~:. I o, usin • 5'-R I~ (Rapid Amplification of d)NA ntis) w~: could confirm only amclin-1 and amcloblu~tin mc~sage~ 1t1 the total R pool from r

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Since 1974, when Slavkin and his collaborators proposed the epithelial origin of cementum, many experiments have been carried out to provide evidence for deposition of enamel-related proteins along the root surface. However, neither amelogenin nor other proteins have fully satisfied expectations. In previous studies, we have identified a novel mRNA coding for an extracellular-like protein which we called amelin. It was expressed at high levels in secretory and postsecretory ameloblasts in rat molars and incisors. In situ hybridization experiments described in the present study also localized the amelin message to epithelial cells adjacent to the peripheral surface of newly deposited dentin in the root end and to cells embedded in cellular cementum in molars. In incisors, the amelin RNA positive cells were detected in the area where cementum formation had been initiated. No amelogenin RNA signal was found in the cells at the root surface. We postulate that the epithelial cells of the root sheath as well as the ameloblasts are synthesizing amelin which might be one of the key proteins coupled to the process of cementogenesis.

We have recently identified a novel RNA sequence in ameloblasts, coding for amelin (Cerny et al., 1996). In the present paper, its expression has been compared with that of amelogenin in developing incisors and molars of rats, by means of in situ hybridization of paraffin sections. The RNAs for both amelin and amelogenin were highly expressed in secretory ameloblasts. The expression of RNA for amelogenin gradually decreased in the post-secretory ameloblasts. In contrast, the RNA expression for amelin remained high in post-secretory ameloblasts up to the stage of fusion between dental and oral epithelia at the time of tooth eruption. We suggest that amelin might be involved in the mineralization of enamel or in the attachment of ameloblasts to the enamel surface. The whole-mount in situ hybridization procedure is described for the first time in dental research. It proved to be a useful method and confirmed the results of the conventional in situ hybridization.