The enhancement of cell adherence and inducement of neurite outgrowth of dorsal root ganglia co-cultured with hyaluronic acid hydrogels modified with Nogo-66 receptor antagonist in vitro

doi:10.1016/j.neuroscience.2005.09.029

Neuroscience

Volume 137, Issue 2, 2006, Pages 519-529

https://doi.org/10.1016/j.neuroscience.2005.09.029 Get rights and content

Abstract

Hyaluronic acid hydrogels modified with polyclonal anti-Nogo-66 receptor antibody were developed in order to promote regeneration in the injured CNS. These modified hydrogels were intended not only to deliver antibodies, but also to serve as a scaffold for neural regeneration following their implantation into injured tissue. Since unmodified hyaluronic acid-hydrogels do not support cell attachment, the gels were modified with polyclonal anti-Nogo-66 receptor with the aim of altering the surface properties of the gels in such a way as to improve neuronal adherence and survival. After evaluating the immobilization efficiency of the system, chicken dorsal root ganglia and dorsal root ganglia cells were planted on the surface of the modified gels to determine cell viability. Dorsal root ganglia were also cultured close to the gels in order to assay the inducement of neurite outgrowth. In dorsal root ganglia and cell viability assay, dorsal root ganglia and neuron cells could adhere to the modified hydrogels and survive well, but it did not happen to unmodified hydrogels. After 72 h, these attached cells were stained positively with immuno-staining for neurofilament. Neurite outgrowth inducement assay showed that the number and length of dorsal root ganglia neurites on the side toward modified hydrogels were significantly more than that on the opposite side (both P<0.01). The results reveal that hyaluronic acid-hydrogels modified with anti-Nogo-66 receptor can support neural cell attachment and survival in vitro. Furthermore, this system can greatly induce neurite outgrowth. The results also indicate that this modified hydrogels have potential to repair injury in the CNS.

Section snippets

Materials

HA sodium salt (MW 2.6,000,000–2.7,000,000 Da) was purchased from Shandong Fureda Biochem (Fureda, Jinan, Shandong, China) and ethyl N,N-dimethylaminopropyl carbodiimide (EDC), adipic dihydrazide (ADH), complete Freund’s adjuvant and incomplete Freund’s adjuvant were obtained from Sigma (St. Louis, MO, USA). Embryonic day 8 chicks were provided by Center of Experimental Animals, Capital University of Medical Sciences, China. All procedures were approved by the Capital University of Medical

The preparation of HA-hydrogels

The chemistry manipulation uses controlled, carbodiimide-mediated coupling of hydrazides to the carboxylic acid functions of a pre-defined number of the glucuronic acid moieties of HA. HA ranging in size from six disaccharide units (1200 Da) to high mass HA (M. 2,000,000 rDa) can be modified. Covalent attachment of ADH to the carboxylic acid groups of HA under mild conditions results in the availability of pendant hydrazide amino functionalities arrayed along the hyaluronate backbone.

The gels

Discussion

In the preparation of HA-hydrogels, this chemistry manipulation uses controlled, carbodiimide-mediated coupling of hydrazides to the carboxylic acid functions of a pre-defined number of the glucuronic acid moieties of HA. Modification of 0.1–50% of the available carboxylic functions can be achieved to meet different needs, such as biochemical probes, drug delivery agents, or biomolecular hydrogels scaffold (Pouyani and Prestwich, 1994). It clearly showed that free hydrazide groups had been

Conclusion

In conclusion, the surface properties of the HA-hydrogels can be altered as soon as anti-NgR links to the gels. The alteration extremely administers to neural cell adherence and survival on the hydrogels. The HA-hydrogels modified with anti-NgR could release antibody and induce neurite outgrowth in vitro. The results presented here might lead to a potential therapeutic approach for CNS injury in the future.

Acknowledgments

This work was supported by the National Natural Grant of China 30470616 and Beijing Scientific Committee Grant, 515038.

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Research paperNeuroanatomyThe enhancement of cell adherence and inducement of neurite outgrowth of dorsal root ganglia co-cultured with hyaluronic acid hydrogels modified with Nogo-66 receptor antagonist in vitro

Abstract

Section snippets

Materials

The preparation of HA-hydrogels

Discussion

Conclusion

Acknowledgments

Biomaterials

Neuron

Neurosci Lett

J Vasc Surg

Adv Immunol

J Biol Chem

Exp Neurol

J Control Release

Neuron

J Control Release

Biomaterials

J Control Release

Biomaterials

J Control Release

Anal Biochem

J Biol Chem

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Glia

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Research paper
Neuroanatomy
The enhancement of cell adherence and inducement of neurite outgrowth of dorsal root ganglia co-cultured with hyaluronic acid hydrogels modified with Nogo-66 receptor antagonist in vitro