獲得被膜形成に関する基礎的研究

獲得被膜モデルの形成と唾液糖蛋白シアル酸の関係についての走査型電顕による観察

抄録

It is generally established that the acquired pellicle is formed by selective adsorption of salivary glycoproteins to the tooth surface. However, this adsorption process has never been confirmed by morphological observation. The purpose of this study was to investigate the formation of acquired pellicle morphologically related to the behaviors of sialic acid of salivary glycoproteins. Whole saliva was collected and sterilized. Sublingual-submaxillary and parotid saliva were also collected from their ducts sterilizingly. Acquired pellicle model was made up on a polished enamel plate of extracted human tooth immersed in the experimental saliva. After incubation time from 1hr to 24hr at 37°C, enamel plate was taken out and observed its surface using a scanning electron microscope. Free and total sialic acids in the saliva were determinded with the lapse of time. Moreover, it was investigated whether the behaviors of sialic acid bound to glycoproteins effect on morphological changes of the enamel plate on which salivary components were formed.
The results were summarized as follows:
1) The formative substance on the enamel plate derived from sterilized whole saliva was observed morphologically. Within 3hrs, amorphous structures adsorbed onto enamel plate. After 4 hours, net-like formative substances were observed on the amorphous surfaces. From the estimation of free and total sialic acids in saliva, it was suggested that the formation of net-like substances was closely related to the behaviors of sialic acid of salivary glycoproteins.
2) Neuraminidase (NANase) and N-acetylneuraminic acid aldolase (aldolase) purified from C. perfringens were added to the sterilized whole saliva. The same net-like formative substances as those derived from sterilized whole saliva were observed on the enamel plate after 24hrs.
3) Sublingual-submaxillary saliva had not free sialic acid but bound to glycoproteins. It had also no NANase and aldolase activity. The enamel plate immersed in this saliva showed amorphous structures predominantly, but with the passage of time, rod-like formative substances (regarded as aggregated salivary glycoproteins) were adsorbed on over all the amorphous structures. When the NANase and aldolase from C. perfringens, were added, the release and break down of sialic acid from salivary glycoproteins occurred. In this case, rod-like substances were changed to indistinct structures in relation to the content of sialic acid released.
4) The enamel plate was then incubated with parotid saliva containing both the enzymes. in spite of treatment with enzymes, amorphous structures found in the non-treated parotid saliva were also observed on the enamel surface.
5) On the enamel plate immersed in mixed saliva (sublingual-submaxillary saliva+parotid saliva), amorphous structures alone were observed. However, with the addition of both the enzymes, net-like formative substances similar to those of whole saliva were seen.
6) When the inhibitor of NANase, 2-deoxy-2, 3-dehydro-N-acetylneuraminic acid was added in whole saliva, NANase activity was inhibited and free sialic acid was not released. Using this saliva, amorphous structures were clearly observed on the enamel plate.
7) The enamel plate was immersed in sterilized whole saliva and sublingual-submaxillary saliva which were previously incubated for 24hrs, in this case, only amorphous structures were found.