Myelin-associated neurite growth-inhibitory proteins and suppression of regeneration of immature mammalian spinal cord in culture.

Abstract

Neurite outgrowth across spinal cord lesions in vitro is rapid in preparations isolated from the neonatal opossum Monodelphis domestica up to the age of 12 days. At this age oligodendrocytes, myelin, and astrocytes develop and regeneration ceases to occur. The role of myelin-associated neurite growth-inhibitory proteins, which increase in concentration at 10-13 days, was investigated in culture by applying the antibody IN-1, which blocks their effects. In the presence of IN-1, 22 out of 39 preparations from animals aged 13-17 days showed clear outgrowth of processes into crushes. When 34 preparations from 13-day-old animals were crushed and cultured without antibody, no axons grew into the lesion. The success rate with IN-1 was comparable to that seen in younger animals but the outgrowth was less profuse. IN-1 was shown by immunocytochemistry to penetrate the spinal cord. Other antibodies which penetrated the 13-day cord failed to promote fiber outgrowth. To distinguish between regeneration by cut neurites and outgrowth by developing uncut neurites, fibers in the ventral fasciculus were prelabeled with carbocyanine dyes and subsequently injured. The presence oflabeled fibers in the lesion indicated that IN-1 promoted regeneration. These results show that the development of myelin-associated growthinhibitory proteins contributes to the loss of regeneration as the mammalian central nervous system matures. The definition of a critical period for regeneration, coupled with the ability to apply trophic as well as inhibitory molecules to the culture, can permit quantitative assessment of molecular interactions that promote spinal cord regeneration.As the central nervous system (CNS) matures in embryonic and neonatal mammals, a sharp decrease occurs in the ability of nerve fibers to grow across spinal cord lesions, to effect repair, and to restore functions (1-5). The exact time at which regeneration ceases varies from species to species (1-4, 6) and within an animal, depending on the maturity of the tract that has been disrupted (7-9, 33). Eventually all CNS regeneration fails except in the olfactory system. Several new strategies have been developed with the aim of prolonging the capacity for regeneration and restoration of function after lesions in adult CNS. These include the use of grafts of peripheral nerve (10, 11), embryonic tissue (12-14), and Schwann cells (15, 16), as well as the application of trophic molecules such as neurotrophin 3 (17) to promote fiber outgrowth across CNS lesions or brain-derived neurotrophic factor to promote survival of axotomized cells (18).In pathways that become myelinated there is a correlation with the development of myelin and the appearance of myelinassociated inhibitory proteins known as NI-35/250 that prevent neurite outgrowth (19)(20)(21). After the effects of these molecules have been blocked by a specific monoclonal antibody (IN-1) neurons can once again grow across spinal cord lesions at stages when regeneration would normally be impossible (17,22,23). Thus, in r...

Increased corticofugal plasticity after unilateral cortical lesions combined with neutralization of the IN‐1 antigen in adult rats

“…The rat monoclonal antibody IN-1 was able to neutralize the human CNS myelin inhibitory property in vitro (Spillmann et al, 1997). Taking into account that mAb IN-1 directed against rat NI-250 could also neutralize the inhibitory activity of frog, opossum, and bovine CNS myelin (Lang et al, 1995;Varga et al, 1995;Spillmann et al, 1998), it is very likely that the IN-1 antigen is highly conserved among different species. A well-established observation is that the outcome of a CNS lesion in humans also depends on the age at which the lesion occurred (Kennard, 1936(Kennard, , 1938.…”

Nogo and Nogo‐66 receptor in human and chick: Implications for development and regeneration

“…These factors include chondroitin sulphate proteoglycans (Zuo et al, 1998;Bradbury et al, 2002), myelin-associated glycoprotein (McKerracher et al, 1994;Mukhopadhyay et al, 1994), and Nogo (Varga et al, 1995). Neutralization of Nogo function with specific antibodies appears to enhance axonal extension in vitro (Spillmann et al, 1997) and in vivo (Brosamle et al, 2000).…”

Identification and Characterization of a Bovine Neurite Growth Inhibitor (bNI-220)

“…Proteins eluted from gel slices containing molecules with an apparent molecular mass of 35 and 250 kDa (SDS-PAGE) showed a very potent inhibitory activity (23,24). A monoclonal antibody (mAb IN-1), which has been raised against rat , neutralizes the neurite growth inhibitory property of differentiated oligodendrocytes and [24][25][26][27][28][29][30]. Immunoprecipitation of CNS myelin proteins by the mAb IN-1 removed more than 50% of inhibitory substrate properties (25,31).…”