Chondroitin sulphate proteoglycans produced by astrocytes modulate neuron adhesion and neurite outgrowth

Abstract

(Uncorrected OCR) ABSTRACT

Abstract of thesis entitled

'Chondroitin sulphate proteoglycans produced by astrocytes modulate neuron adhesion and neurite outgrowth' submitted by HO Yeung

for the degree of Master of Philosophy at the University of Hong Kong in 2000

Several types of proteoglycans were known to be associated with the glial boundary during CNS development and glial scar following CNS injury. The aim of this study is to investigate the role of astroglial proteoglycans, particularly chondroitin sulfate proteoglycans (CSPGs), in modulating neuron adhesion and neurite growth.

Neonatal rat cerebral cortical astrocytes as well as an astrocyte cell line DI TNCI were studied for the expression of CSPGs and their effects on neuron adhesion and neurite outgrowth. Immunocytochemical staining with CS-56, a monoclonal antibody against epitopes in the CS moiety of CSPGs, demonstrated the localization of CSPGs on both primary astrocytes and DI TNCI cell line. Immunostaining with anti-neurocan (a neuronal CSPG originally thought) showed pericellular and intracellular distribution in confluent cultures of primary astrocytes but only intracellular distribution in the DI TNCI cells, both being positive for glial fibrillary acidic protein (GFAP). Neurocan was also found deposited by primary astrocytes on the culture substratum. Western blots of the conditioned medium of primary astrocytes indicated a distinct 220 kDa neurocan core protein band. Five immunoreactive bands were also found on Western blots detected with stub-sensitve antibody against C-6-S-PG. Furthermore, RT-PCR analysis indicated the expression of neurocan mRNA in the primary astrocytes.

Cortical neural cells (from E18 rats) selectively attached and extended neurites on the primary astrocytes but not on the non-cellular, neurocan immuoreactive gaps left by the astrocytes nor on the DI TNCI cells. Neuronal distribution on astrocytes remained similar even after digestion with chondroitinase or neutralization with antibody, CS-56 or antineurocan. Neurite length on the chondroitinase ABC (20mU/ml) treated astrocytes was however increased.

Taken together, these in vitro observations suggest that apart from neurons, astrocytes are another cellular source of brain-specific CSPG, neurocan. They also suggest that CS components expressed on the surface of astrocytes are supportive of neuronal attachment and survival but limiting towards neurite extension. Evidence is also provided for the selective development of embryonic neurons directly on astrocytes and not on neurocan immunoreactive non-cellular routes traversed by astrocytes.

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