Yi Cui1, Yanyun Yin2, Yunlong Zou3, Yannan Zhao2, Jin Han2, Bai Xu2, Bing Chen2, Zhifeng Xiao2, Hongwei Song4, Ya Shi2, Weiwei Xue2, Xu Ma5, Jianwu Dai6. 1. Reproductive and Genetic Center of National Research Institute for Family Planning, Beijing, 100081, China. 2. Key Laboratory of Molecular Developmental Biology, Institute of Genetics and Developmental Biology, Chinese Academy of Sciences, 3 Nanyitiao, Zhongguancun, Beijing, 100190, China. 3. Orthopaedics Surgery Department, China-Japan Union Hospital of Jilin University, No. 126, Xiantai Street, Changchun, 130033, Jilin Province, China. 4. EHBIO gene technology, No. 46, Jiugulou Street, Beijing, 100100, China. 5. Reproductive and Genetic Center of National Research Institute for Family Planning, Beijing, 100081, China. genetic88@126.com. 6. Key Laboratory of Molecular Developmental Biology, Institute of Genetics and Developmental Biology, Chinese Academy of Sciences, 3 Nanyitiao, Zhongguancun, Beijing, 100190, China. jwdai@genentics.ac.cn.
Abstract
BACKGROUND: Recently, neural stem cell (NSC) therapy has shown promise for the treatment of many neurological diseases. Enhancing the quality of implanted cells and improving therapeutic efficacy are currently research hotspots. It has been reported that collagen sponge material provided sufficient room for cell growth in all directions and promoted the absorption of nutrients and removal of wastes. And also, the Rotary Cell Culture System (RCCS), which mimics the microgravity environment, can be used to culture cells for tissue engineering. MATERIALS AND METHODS: We performed the mRNA and miRNA sequencing to elucidate the regulatory mechanism of NSCs cultured on the collagen sponge in the RCCS system. The luciferase assay and Western blot revealed a direct regulatory role between let-7i-5p and neurotrophic receptor tyrosine kinase 3 (NTRK3; also called TrkC). And then, the neural differentiation markers Tuj1 and Map2 were detected by immunofluorescence staining. In the meantime, the migratory ability of NSCs was detected both in vitro and in spinal cord injury animals. RESULTS: In this study, we demonstrated that the expression of NTRK3 was elevated in NSCs cultured on collagen sponge in the RCCS system. Furthermore, increased NTRK3 expression was regulated by the downregulation of let-7i-5p. Compared to traditionally cultured NSCs, the NSCs cultured on collagen sponge in the RCCS system exhibited better neuronal differentiation and migratory ability, especially in the presence of NT-3. CONCLUSIONS: As the biological properties and quality of transplanted cells are critical for therapeutic success, the RCCS system combined with the collagen sponge culture system shows promise for applications in clinical practice in the future.
BACKGROUND: Recently, neural stem cell (NSC) therapy has shown promise for the treatment of many neurological diseases. Enhancing the quality of implanted cells and improving therapeutic efficacy are currently research hotspots. It has been reported that collagen sponge material provided sufficient room for cell growth in all directions and promoted the absorption of nutrients and removal of wastes. And also, the Rotary Cell Culture System (RCCS), which mimics the microgravity environment, can be used to culture cells for tissue engineering. MATERIALS AND METHODS: We performed the mRNA and miRNA sequencing to elucidate the regulatory mechanism of NSCs cultured on the collagen sponge in the RCCS system. The luciferase assay and Western blot revealed a direct regulatory role between let-7i-5p and neurotrophic receptor tyrosine kinase 3 (NTRK3; also called TrkC). And then, the neural differentiation markers Tuj1 and Map2 were detected by immunofluorescence staining. In the meantime, the migratory ability of NSCs was detected both in vitro and in spinal cord injury animals. RESULTS: In this study, we demonstrated that the expression of NTRK3 was elevated in NSCs cultured on collagen sponge in the RCCS system. Furthermore, increased NTRK3 expression was regulated by the downregulation of let-7i-5p. Compared to traditionally cultured NSCs, the NSCs cultured on collagen sponge in the RCCS system exhibited better neuronal differentiation and migratory ability, especially in the presence of NT-3. CONCLUSIONS: As the biological properties and quality of transplanted cells are critical for therapeutic success, the RCCS system combined with the collagen sponge culture system shows promise for applications in clinical practice in the future.
Authors: Stephanie Szobota; Pranav D Mathur; Sairey Siegel; KristenAnn Black; H Uri Saragovi; Alan C Foster Journal: PLoS One Date: 2019-10-31 Impact factor: 3.240