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目的:研究小鼠下颌下腺细胞的培养方法,探讨下颌下腺细胞培养条件及细胞生长特性,为研究干细胞转分化涎腺腺泡细胞以及涎腺再生研究奠定理论基础和技术支持。方法:取2周龄的小鼠,组织块法和消化法分别进行培养,用活细胞观察法和HE染色法记录细胞形态学特征;免疫荧光染色法鉴定;通过生长曲线和细胞倍增时间来比较两种方法对细胞增殖能力的影响。结果:组织块法和消化法均可以成功的培养下颌下腺细胞,(1)组织块法培养的细胞呈卵圆形或多边形,10天左右成铺路石样;消化法培养细胞亦为上皮样细胞,呈多边形,胞质丰富;(2)HE染色下颌下腺细胞呈多边形,胞核明显;(3)Cytokeratin13和AQP5表达阳性,Wimentin表达阴性;(4)组织块法培养细胞增殖相对较平缓,消化法培养细胞增长较迅速;(5)消化法培养倍增时间(1.3471±0.6071)天比组织块法倍增时间(2.1887±1.1503)明显缩短(P<0.05);结论:体外可以成功的培养下颌下腺细胞,但是同时证明下颌下腺细胞长期传代较困难,这为研究干细胞转分化涎腺细胞和涎腺再生体内实验提供了理论和实验基础。
OBJECTIVE: To study the culture method of submandibular gland cells in mice and to explore the culture conditions and cell growth characteristics of submandibular gland cells, and to lay a theoretical foundation and technical support for the study on stem cell transdifferentiation of salivary gland acinar cells and salivary gland regeneration. Methods: Two-week-old mice were cultured by tissue block method and digestion method. Morphological characteristics were recorded by living cell observation and HE staining, respectively. Immunofluorescence staining was used to identify the growth curve and cell doubling time Effects of Two Methods on Cell Proliferation. Results: The submandibular gland cells could be successfully cultured by tissue block method and digestion method. (1) The cells cultured by tissue block method were oval or polygonal, and became paving stone samples in about 10 days. The cultured cells in digestion method were also epithelial-like cells (3) Cytokeratin 13 and AQP5 expression was positive, and Wimentin expression was negative; (4) Tissue block cultured cells were relatively flat and digested, with polygons rich in cytoplasm; (2) HE staining showed that the submandibular gland cells were polygonal with obvious nuclei; (5) digestive culture doubling time (1.3471 ± 0.6071) than tissue block doubling time (2.1887 ± 1.1503) was significantly shorter (P <0.05); Conclusion: in vitro can be successfully cultured submandibular gland cells , But at the same time it is proved that it is more difficult to passage the submandibular gland cells for a long time. This provides a theoretical and experimental basis for studying the in vivo experiments of stem cell transdifferentiation salivary gland cells and salivary gland regeneration.