In contrast, at similar time points following nerve damage, only minor fragments of axonal debris remained within the nerve and neurofilament protein was no longer detectable ( Figure S3B). Although the degree of demyelination was severe in the P0-RafTR mice, it was not complete, as some axons were still myelinated. To determine whether the incomplete phenotype was due to insufficient levels of tamoxifen throughout
the nerve, we performed intraneural injections of tamoxifen into several P0-RafTR and control mice ( Figure S3D). Consistent with this hypothesis, we found complete demyelination of nerves in the proximity of the injection site. Importantly, this occurred in the absence of observable axonal damage and the structure of the nerve buy LBH589 was normal in sections far from the injection site (
Figures S3E and S3F). Thus, activation of Raf-kinase activity in myelinating Schwann cells is sufficient to drive Schwann cell dedifferentiation in adult nerve without causing axonal damage. The ability to tightly regulate Raf-kinase activity in Schwann cells in the context of a normal nerve allows us to determine the role of this specific signaling pathway in Schwann cells in the broader inflammatory and regenerative response to injury. EM examination of the nerves Enzalutamide in vitro from P0-RafTR animals following tamoxifen injection, and revealed an increase in the size of the collagen-rich spaces between Schwann cell/axon units, which contained cells that were not present in control nerves (Figure 4A) and quantification confirmed this increase in cell number (Figure 4B). We also observed a large increase in p75-positive cells, presumably largely due to the dedifferentiation of myelinating cells to a progenitor-like state (Figure 4B). Moreover, proliferation markers showed there was considerably more proliferation in the nerves from injected P0-RafTR mice compared to controls and that a significant proportion
of these proliferating cells were Schwann cells (Figures 4C and S4). When peripheral nerves are injured, inflammatory cells are recruited to the injury site and throughout the distal stump where they aid in the clearance of myelin debris—a prerequisite for efficient nerve regeneration (Chen et al., 2007). Following a physical trauma, chemoattractants are released which attract inflammatory cells. Naked axons, myelin debris, and dedifferentiated Schwann cells have all been proposed as potential sources of such inflammatory signals (MacDonald et al., 2006 and Martini et al., 2008). As aberrant inflammatory responses have been linked both to peripheral neuropathies and the development of peripheral nerve tumors, it is important to determine the cellular and molecular basis of these responses (Martini et al., 2008, Meyer zu Hörste et al.