Effect of transforming growth factor-beta 1 and -beta 2 on Schwann cell proliferation on neurites.

Mechanisms regulating Schwann cell proliferation during development are unclear. Schwann cell division is known to be driven by an unidentified mitogen present on the surface of axons, but it is not known whether other molecules play a role in regulating this proliferation. Transforming growth factor-beta (TGF-beta) which is found in the developing peripheral nervous system (PNS) and is mitogenic for neuron-free Schwann cells in vitro could be involved. We have investigated the effects of TGF-beta 1, TGF-beta 2 and antibodies to TGF-beta 1 and TGF-beta 2 on axon driven Schwann cell proliferation. Rat embryonic dorsal root ganglion neurons (DRG) neurons and Schwann cells from the sciatic nerve were isolated, purified and recombined in vitro. Confirming earlier reports by others, we observed that TGF-beta 1 and TGF-beta 2 added to the culture medium stimulated the proliferation of Schwann cells in the absence of neurons. However, when added to neuron-Schwann cell co-cultures, TGF beta caused a variable response ranging from no effect to moderate inhibition of Schwann cell proliferation in different experiments. A stimulation of Schwann cell proliferation by TGF beta was never observed in neuron-Schwann cell co-cultures. Antibodies to TGF-beta 1 and TGF-beta 2 did not influence axon driven Schwann cell proliferation. To further determine the role of TGF-beta in Schwann cell proliferation and myelination, we studied Schwann cell proliferation in cultures from mice in which the TGF-beta 1 gene was delected by homologous recombination. Neuron-Schwann cell cultures from wild-type, heterozygous and homozygous mice were used. No differences were observed in either Schwann cell proliferation or myelination between cultures obtained from homozygous mutants and their heterozygous and wild-type controls. These findings suggest that TGF-beta does not function as a part of the mitogenic mechanism presented by neurons to Schwann cells, but that the presence of active TGF beta in the cellular environment might regulate the degree of proliferation induced by neuronal contact.

Transforming growth factor-beta blocks myelination but not ensheathment of axons by Schwann cells in vitro.

Mechanisms regulating Schwann cell differentiation into a myelinating or a mature nonmyelinating phenotype during development are poorly understood. Humoral factors such as members of the transforming growth factor-beta (TGF-beta) family, which are found in the developing and adult mammalian nervous system and are known to affect cell differentiation, could be involved. We tested the effects of TGF-beta isoforms on the ensheathment and myelination of dorsal root ganglion (DRG) neurons by Schwann cells in vitro. Rat embryonic DRG neurons and Schwann cells from the sciatic nerve were isolated, purified, and recombined. In serum-free conditions, TGF-beta blocked both Schwann cell myelination and the expression of the myelin-related molecules galactocerebroside, P0, myelin-associated glycoprotein, and myelin basic protein. In contrast, the expression of molecules characteristic of mature nonmyelinating Schwann cells, including neural-cell adhesion molecule, L1, and nerve growth factor receptor, was maintained when compared to Schwann cells in nondifferentiated cultures. Notably, the expression of glial fibrillary acidic protein, which is expressed only in mature nonmyelinating Schwann cells in vivo, was increased 10-fold in our cultures by TGF-beta. Electron microscopic analysis indicated that in the presence of TGF-beta, basal lamina deposition by Schwann cells was slightly increased. Most importantly, many axons in TGF-beta-treated cultures received ensheathment typical of mature nonmyelinated nerves. These effects of TGF-beta were partially reversed by specific neutralizing anti-TGF-beta antibodies. We interpret these results as evidence that TGF-beta regulates Schwann cell differentiation in vitro by blocking the expression of the myelinating phenotype and promoting the development of the nonmyelinating phenotype.

Astrocytes inhibit Schwann cell proliferation and myelination of dorsal root ganglion neurons in vitro.

Schwann cells promote the regrowth of nerve fibers in both the PNS and CNS and might thus be of value in strategies to promote repair following injury or demyelination in the CNS. The effectiveness of Schwann cells in promoting repair could, however, be limited by interactions with reactive astrocytes that are prominent at lesioned and demyelinated sites. To investigate this possibility, experiments were performed to determine the influence of cortical astrocytes on Schwann cell proliferation and myelination of dorsal root ganglion (DRG) neurons in vitro. DRG neurons from embryonic rats and Schwann cells, astrocytes, and fibroblasts isolated from the sciatic nerve, cerebral cortex, and cranial periosteum, respectively, of neonatal rats were purified and then recombined to provide neuron-Schwann cell, neuron-Schwann cell-astrocyte, and neuron-Schwann cell-fibroblast cultures. Astrocytes inhibited both neuron-dependent Schwann cell proliferation and the myelination of axons by Schwann cells. The expression of galactocerebroside, but not of the O4 antigen, was inhibited by astrocytes, suggesting that astrocytes blocked Schwann cell differentiation prior to the onset of myelination. Ultrastructural analysis of the cultures also indicated that both axonal ensheathment and the segregation of large axons into 1:1 relationships were decreased in the presence of astrocytes. Astrocytes did not affect the expression of the basal lamina components type IV collagen and laminin, and basal lamina formation assessed by electron microscopy was only slightly decreased. Some of these inhibitory effects appear to be mediated by diffusible factors since astrocyte-conditioned medium also reduced Schwann cell myelination. Fibroblasts or fibroblast-conditioned medium did not induce such inhibitory effects, indicating that the effects were astrocyte specific. We conclude that cortical astrocytes release a soluble factor(s) that inhibits specific aspects of neuron-Schwann cell interactions leading to myelination.