| Giant panda is the world’s flagship species of biodiversity conservation.The Qinling giant panda is an important subspecies of giant panda population,which with unique morphological characteristics and genetic characteristics.It’s also more precious and endangered compared with other areas.Mesenchymal stem cells are ideal seed cells in tissue engineering,regenerative medicine and cell therapy,which is the most valuable germplasm resources of giant pandas for conservation.In this study,bone marrow,dental pulp and adipose mesenchymal stem cells were isolated from qinling giant panda,and their morphological characteristics,stem cell characteristics and multidirectional differentiation potential were detected.At the same time,HE staining was used to observe the histological structure of fetal membrane of giant panda,and fibroblast-like cells from skin,liver,myocardium and umbilical cord tissue were isolated and cultured to obtain the germplasm resources of Qinling giant panda mesenchymal stem cells and histiocytes,which will provide necessary experimental materials for the protection and clinical application of genetic diversity of giant panda in China and related scientific research.The test results are as follows:1.In this experiment,the bone marrow,dental pulp and adipose tissue of Qinling giant panda were treated by tissue block method and collagenase digestion method,and the cells from each tissue were obtained.The morphology of these cells was long fusiform,and they all showed whirlpool growth state when the degree of cell fusion reached 90%.In the 10 th generation,the proliferation ability of all cells decreased and senescence was obvious.Bone marrow-derived cells expressed CD73,CD90,SOX-2 and SCFR,but not CD34.Dental pulp cells expressed SCFR,CD90 and CD73,but not CD34 and CD31.Adipose-derived cells were positive for CD73,CD90 and GNL3,but negative for CD34 and CD31.The results of induced differentiation showed that cells from various tissues could differentiate into adipocytes,osteoblasts and chondrocytes,indicating that mesenchymal stem cells from bone marrow,dental pulp and adipose tissue were successfully obtained in this experiment.2.The fetal membrane of Qinling giant panda is composed of chorionic membrane,amniotic membrane and allallic membrane.The chorionic membrane has the largest area and abundant blood vessels,which are composed of single ciliated columnar epithelium and reticular layer.The amniotic membrane is composed of the epithelium,middle layer and dense layer.The allantoic membrane is the thinnest and consists of a single layer of flat epithelium,loose connective tissue and blood vessels.3.In this study,fibroblasts from skin,liver,myocardium and umbilical cord of Qinling giant panda were obtained by tissue block method and collagenase digestion method.Fibroblasts from skin of giant panda were cultured for 24 h,and the degree of fusion was more than 90%.Fibroblasts from liver,myocardium and umbilical cord tissue of giant panda could reach more than 80% degree of fusion after 48 h culture.In conclusion,germplasm cells from different tissues of Qinling giant panda were successfully obtained and preserved in this study,which provided a material basis for the preservation of germplasm resources of Qinling giant panda,clinical disease treatment and scientific research. |
