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Thermosensitive quaternized chitosan hydrogel scaffolds promote neural differentiation in bone marrow mesenchymal stem cells and functional recovery in a rat spinal cord injury model

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Abstract

A thermosensitive quaternary ammonium chloride chitosan/β-glycerophosphate (HACC/β-GP) hydrogel scaffold combined with bone marrow mesenchymal stem cells (BMSCs) transfected with an adenovirus containing the glial cell-derived neurotrophic factor (GDNF) gene (Ad-rGDNF) was applied to spinal cord injury (SCI) repair. The BMSCs from rats were transfected with Ad-rGDNF, resulting in the expression of GDNF mRNA in the BMSCs increasing and their spontaneous differentiation into neural-like cells expressing neural markers such as NF-200 and GFAP. After incubation with HACC/β-GP hydrogel scaffolds for 2 weeks, neuronal differentiation of the BMSCs was confirmed using immunofluorescence (IF), and the expression of GDNF by the BMSCs was detected by Western blot at different time points. MTT assay and scanning electron microscopy confirmed that the HACC scaffold provides a non-cytotoxic microenvironment that supports cell adhesion and growth. Rats with SCI were treated with BMSCs, BMSCs carried by the HACC/β-GP hydrogel (HACC/BMSCs), Ad-rGDNF-BMSCs, or Ad-rGDNF-BMSCs carried by the hydrogel (HACC/GDNF-BMSCs). Animals were sacrificed at 2, 4, and 6 weeks of treatment. IF staining and Western blot were performed to detect the expression of NeuN, NF-200, GFAP, CS56, and Bax in the lesion sites of the injured spinal cord. Upon treatment with HACC/BMSCs, NF200 and GFAP were upregulated but CS56 and Bax were downregulated in the SCI lesion site. Furthermore, transplantation of HACC/GDNF-BMSCs into an SCI rat model significantly improved BBB scores and regeneration of the spinal cord. Thus, HACC/β-GP hydrogel scaffolds show promise for functional recovery in spinal cord injury patients.

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Funding

This study was supported by the National Natural Science Foundation of China, No. 81071466; the Science and Education Leading Talents Program for Invigorating Health of Yangzhou during 13th Five-Year Plan Period, No. LJRC20182; the Science and Technology Program of Nantong City, No. GJZ16044; and the Science and Technology Program of Rugao City, No. SRG(15) 3015.

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  1. Department of Orthopedics, Affiliated Rugao Hospital of Nantong University, Ninghai Road No. 278, Rugao, Jiangsu Province, China

    Cheng Huang, Yuanbing Liu, Jian Ding, Yongping Dai, Lixiang Le, Liangliang Wang & Erhu Ding

  2. Clinical Medical College, Yangzhou University , Jiangyang Middle Road No. 136, Yangzhou, Jiangsu Province, China

    Jiandong Yang

  3. Northern Jiangsu People’s Hospital Affiliated to Yangzhou University, Nantong West Road No. 98, Jiangsu Province, Yangzhou, China

    Jiandong Yang

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  1. Cheng Huang
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Correspondence to Jiandong Yang.

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Huang, C., Liu, Y., Ding, J. et al. Thermosensitive quaternized chitosan hydrogel scaffolds promote neural differentiation in bone marrow mesenchymal stem cells and functional recovery in a rat spinal cord injury model. Cell Tissue Res 385, 65–85 (2021). https://doi.org/10.1007/s00441-021-03430-x

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