A histochemical study of the cementum mineralization in unfixed, undemineralized sections

固定的脱矿切片通常用于骨质的组织学和组织化学研究。然而，固定和脱矿可能导致基质的结构变化和/或损失。钙尤其不能在脱矿切片中检测到。为了克服这一困难，不固定、不脱矿的部分是必要的。采用粘膜法制备未固定、未脱矿的大鼠磨牙冰冻切片，偶氮染色检测碱性磷酸酶（ALP）的定位，乙二醇双（2-羟基苯胺）（GBHA）染色检测钙的定位，免疫组化检测骨涎蛋白（BSP）和骨桥蛋白（OPN）的定位。结果表明：1)矿化发生后，矿化后的牙本质上出现初始脱细胞牙骨质；2)初始脱细胞骨质缺乏原纤维，富含BSP和OPN；3)在脱细胞成水泥初期，成水泥细胞表现出强烈的ALP反应，根表面出现了小的GBHA反应颗粒（= 1μm，直径约1μm）；4)随着主要纤维附着在初始骨质上，ALP反应减弱，GBHA反应颗粒数量减少。结果表明，在脱细胞胶凝发生初期，根表面出现基质囊泡和钙化球粒，但在主纤维附着后，这两种结构中的任何一种都看不到。综上所述，我们得出结论：1)在初始的脱细胞骨水泥形成过程中，成骨水泥细胞释放基质囊泡，形成与GBHA活性颗粒相对应的钙化小球体；钙化球粒诱导初始脱细胞牙骨质矿化；3)主纤维附着后，矿化沿主纤维推进，无基质囊泡。此外，本研究表明，与固定、脱矿切片相比，未固定、未脱矿切片的组织化学和免疫组织化学反应保存得更好。少

Fixed, demineralized sections have been generally used for histological and histcochemical studies of the cementum. The fixation and demineralization, however, may cause the structural change and/or loss of matrices. Calcium especially can not be detected in demineralized sections. To overcome the difficulty, unfixed,undemineralized sections are necessary.We prepared unfixed, undemineralized frozen sections of rat molars by an adhesive film method, and examined the localization of alkaline phosphatase (ALP) by Azo-dye staining, calcium by glycol bis(2-hydroxyanil)(GBHA) staining, and bone sialoprotein(BSP) and osteopontin(OPN) by immunohistochemistry. We found the following : 1) with the onset of dentin mineralization, the initial acellular cementum appears on the mineralized dentin ; 2) the initial acellular cementum is fibril-poor, and rich in BSP and OPN ; 3) at the initial acellular cementogenesis, cementoblasts show the intense ALP reaction, and small GBHA reactive particles (=cal … More cium-GBHA comprex, about 1μm in diameter) appear on the root surface; 4) as principal fibers attach to the initial cementum, the ALP reaction became low and GBHA reactive particles decrease in number.It has been shown that matrix vesicles and calciferous spherules appear on the root surface at the initial acellular cementogenesis, but after principal fiber attachment, either of the two structures is not seen. From the above findings, we concluded that: 1) at the initial acellular cementogenesis, cementoblasts release matrix vesicles, which result in calciferous spherules corresponding to the GBHA reactive particles; 2) calciferous spherules induce the mineralization of the initial acellular cementum; 3) after principal fiber attachment, the mineralization advances along principal fibers without matrix vesicles. In addition, this study showed that the histochemical and immunohistochemical reactions are preserved better in unfixed, undemineralized sections than in fixed, demineralized sections. Less