Expression of PLCgamma2 in a wild-type mouse model of chronic periapical periodontitis

doi:10.3969/j.issn.2095-4344.1023

Abstract

Abstract:

BACKGROUND: Inhibiting BTK-PLCγ2-Ca2+ signaling pathway has been shown to inhibit formation and function of osteoclasts, suggesting that PLCγ2 participates in cell formation and differentiation.
OBJECTIVE: To study the expression of PLCγ2 in periapical tissues by establishing a model of periapical periodontitis in wild-type mice.
METHODS: Twenty male wild-type C57BL/6L mice were selected. Bilateral mandibular first molar pulp cavities of 16 mice were exposed to the oral environment and the formation of apical periodontitis was induced (experimental group). The mice were randomly sacrificed at 1, 2, 3 and 4 weeks and the lower jaws were isolated. The remaining four mice were used as blank control group, and were sacrificed at 0 week to isolate the lower jaws. After being histologically processed, they were hematoxylin-eosin stained to observe the inflammation conditions in the apical area. Expression and distribution of PLCγ2 were analyzed by immunohistochemical staining. Enzyme-histochemical staining was used to detect the expression of osteoclasts in apical inflammatory tissue.
RESULTS AND CONCLUSION: According to the histological examination, there was almost no inflammatory cell infiltration in the periodontal tissue of the mandibular first molar in the blank control group. In the experimental group, at 1 to 4 weeks after operation, the infiltration of inflammatory cells in the periodontal tissues and the destruction of alveolar bone of mandibular first molar increased gradually, indicating that the mouse periapical periodontitis model was established successfully. Immunohistochemical analysis found that the expression of PLCγ2 was detected at 0 to 4 weeks after periapical periodontitis in mice. A small amount of PLCγ2 expressed in normal dental apical tissues. PLCγ2 in the experimental group was mainly expressed in the nucleus of lymphocytes, plasma cells and other cells, and there was a small amount of expression in the cell membrane and cytoplasm. At the1st, 2nd, 3rd, 4th weeks, the positive expression of PLCγ2 also increased due to the gradual expansion of inflammatory cell infiltration. The positive cell counts were all significantly different (P < 0.05). The results of TRAP staining showed that almost no osteoclasts were observed in periapical periodontal tissues of blank control group. In the experimental group, at 1 week after operation, a small amount of multinucleated osteoclasts were observed. The number of osteoclasts continued to increase at 2 weeks after operation. At 3 weeks, the number of osteoclasts reached the peak. At 4 weeks postoperatively, a decrease in the number of osteoclasts was observed. The difference was significant between experimental and control groups (P < 0.05). Correlation analysis between TRAP positive cell count and PLCγ2 positive cell count showed a significant correlation (r=0.627, P < 0.001). These results indicate that PLCγ2 is expressed in periapical tissues of mice, implying that it may be related to periapical inflammatory reaction and bone resorption.

中国组织工程研究杂志出版内容重点：组织构建；骨细胞；软骨细胞；细胞培养；成纤维细胞；血管内皮细胞；骨质疏松；组织工程

Key words: Phospholipase C gamma, Periapical Periodontitis, Osteoclasts, Tissue Engineering

CLC Number:

R466
R318

Yang Chunhe, Zhang Hong, Dong Ming, Wang Lina, Niu Weidong. Expression of PLCgamma2 in a wild-type mouse model of chronic periapical periodontitis[J]. Chinese Journal of Tissue Engineering Research, 2019, 23(7): 1057-1062.

Figures/Tables 2

2.1 实验动物数量分析实验选用小鼠20只，分为5组，实验过程无脱失，全部进入结果分析。 2.2 苏木精-伊红染色结果见图1所示，正常对照组(0 周)根尖周组织的牙周膜纤维未发生明显改变，偶有极少量炎性细胞浸润，未发现牙周膜的增宽；实验组1周可见根尖周组织出现小范围的炎性细胞浸润，牙周膜少量增宽，此时可见少量中性粒细胞；实验组2周，炎性细胞浸润范围逐渐增大，主要以中性粒细胞、淋巴细胞、巨噬细胞为主，开始出现牙槽骨的吸收；实验组3，4周，炎性细胞浸润进一步加重，多以淋巴细胞为主，同时伴有毛细血管和成纤维细胞的出现，牙槽骨吸收范围继续扩大。 2.3 免疫组织化学染色结果 PLCγ2的阳性细胞通过免疫组织化学反应染成深褐色，在实验性小鼠的根尖周炎的0-4周都有表达，并随着炎症细胞浸润范围和牙槽骨吸收范围的增大而增多，见图2。正常对照组根尖周组织中仅有少量PLCγ2的阳性表达。实验组1-4周，小鼠根尖周的炎症区域的中央和周边都可以可见到PLCγ2阳性表达，主要位于淋巴细胞、浆细胞等细胞的细胞核中，细胞膜和细胞质中也有少量表达。实验组2，3，4周时由于炎症细胞浸润范围的逐渐扩大，PLCγ2阳性表达也随之增长，阳性细胞计数差异均有显著性意义(P < 0.05)，见图3，与此同时可见根尖骨破坏血管周围及根分叉区域的PLCγ2阳性表达也有所增多。 2.4 酶组织化学染色结果小鼠根尖周组织病变发展过程中TRAP染色结果见图4，正常对照组根尖牙周组织中几乎不能观察破骨细胞。实验组1周，根尖周可见牙槽骨吸收部位附近可观察到少量的多核破骨细胞，经TRAP染色后的破骨细胞胞浆呈酒红色，胞核呈蓝色，分布于骨小梁附近的骨吸收陷窝中；实验组2周，破骨细胞的数量持续增多；实验组3周达到峰值；实验组4周时，可以观察到破骨细胞开始减少，与对照组相比差异均有显著性意义(P < 0.05)，见图5。"

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