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Effect of bFGF on fibroblasts derived from the golden snub-nosed monkey

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Abstract

Golden snub-nosed monkeys are endangered animals in China, and their cells have been demonstrated to be important as genetic resources and in applications for advancing biological research. Moreover, in primary research, basic fibroblast growth factor (bFGF) is used to promote the proliferation of fibroblasts to create abundant cells for cryopreservation. To further investigate the effect of bFGF on the efficiency of preservation of fibroblasts obtained from an endangered species, a fibroblast cell line was isolated from a dead golden snub-nosed monkey. Cell viability and mitochondrial membrane potential were assessed using CCK8 and JC-1 assay kits. The karyotype was analyzed by chromosomal microarray analysis, while RNA sequencing and gene expression analyses were performed to assess molecular changes in response to bFGF. Flow cytometry was used to characterize changes in cell surface markers in response to bFGF treatment. The results showed that cells maintained typical fibroblast morphology, while cell viability and mitochondrial membrane potential were not significantly affected between three and eight passages (p > 0.05). We also observed that the addition of bFGF promoted fibroblast proliferation and increased mitochondrial membrane potential. In addition, the bFGF treatment did not alter the normal karyotype of cells, downregulating fibroblast-associated genes and upregulating those associated with cell regulation, including those of the WNT, PI3K and MAPK pathways. The addition of bFGF also increased CD29, CD90, CD105, CD34 and CD44 expression while decreasing that of CD14 and HLA-DR at the protein level. Taken together, these results demonstrate that bFGF may upregulate the WNT, PI3K and MAPK pathways to promote cell proliferation while also increasing the expression of genes and surface markers associated with mesenchymal and hematopoietic cell linages.

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Data availability

The datasets generated for this study can be found in the NCBI/BioProject/PRJNA640274.

Abbreviations

ESCs:

Embryonic stem cells

iPS cells:

Induced pluripotent stem cells

MSCs:

Mesenchymal stem cells

HSCs:

Hematopoietic stem cells

bFGF:

Basic fibroblast growth factor

PBS:

Phosphate-buffered saline

FBS:

Fetal bovine serum

PFA:

Polyoxymethylene

SD:

Standard deviation

TDF-β:

Transforming growth factor-beta

EMT:

Epithelial-mesenchymal transition

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Funding

This work was supported by the National Key Program of Research and Development, Ministry of Science and Technology (2016YFC0503200), the National Natural Science Foundation of China (Grant No. 30900155), the Natural Science Foundation of Shaanxi Province, China (Grant No. 2014JM3062) and the Opening Foundation of Shaanxi Key Laboratory for Animal Conservation (Northwest University, Grant No. AC2019004).

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Authors and Affiliations

  1. Shaanxi Key Laboratory for Animal Conservation, Key Laboratory of Resource Biology and Biotechnology in Western China, Ministry of Education, College of Life Science, Northwest University, Xi’an, 710069, China

    Xin Liu, Hao Wei, Jing Yang, Pei Zhang, Yuli Li, Rong Hou, Xingrong Yan & Baoguo Li

  2. The School of Life Science and Technology, Xi’an Jiaotong University, Xi’an, 710049, China

    Jingjing Li

  3. College of Veterinary Medicine, Northwest Agriculture and Forestry University, Yangling, 712100, China

    Jinlian Hua

  4. Center for Excellence in Animal Evolution and Genetics, Chinese Academy of Science, Kumming, 650223, China

    Baoguo Li

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  1. Xin Liu
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Correspondence to Xingrong Yan or Baoguo Li.

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10329_2020_875_MOESM1_ESM.tif

Fig. 1 PCA and species distribution. The distributions of the top ten RNA species of the bFGF− and bFGF+ groups are shown in Fig. 1A and 1B, with distinctive colors and areas showing different species and proportions. PCA of fibroblasts is presented in Fig. 1C. Vertical and horizontal coordinates indicate PC2 and PC1, with blue and green dots indicating the control and treatment groups, respectively (TIF 37587 KB)

10329_2020_875_MOESM2_ESM.tif

Fig. 2 Expression levels of ITGβ1 mRNA in fibroblasts from a golden snub-nosed monkey. The expression levels of ITGβ1 (CD29) are shown in Fig. 2A (S. Table 2), and the basal protein fluorescence presented as the relative optical density is shown in Fig. 2B (S. Table 2) (TIF 87411 KB)

Supplementary file3 (DOC 33 KB)

Supplementary file4 (DOCX 27 KB)

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Liu, X., Wei, H., Yang, J. et al. Effect of bFGF on fibroblasts derived from the golden snub-nosed monkey. Primates 62, 369–378 (2021). https://doi.org/10.1007/s10329-020-00875-6

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