Promotion of Peripheral Nerve Regeneration by Hepatocyte Growth Factor through Activation of Schwann Cells

Abstract

During the peripheral nerve regeneration process, a variety of neurotrophic factors play roles in nerve repair by acting on neuronal or non-neuronal cells. In this thesis work, I investigated the role(s) of hepatocyte growth factor (HGF) and its receptor, c-met, in peripheral nerve regeneration. When mice were subjected to sciatic nerve injury, the HGF protein level was highly increased at the injured and distal sites. The levels of both total and phosphorylated c-met were also highly up-regulated, but almost exclusively in Schwann cells (SCs) distal from the injury site. When mice were treated with the c-met inhibitor PHA-665752, myelin thickness and axon regrowth were decreased, indicating that re-myelination was hindered. HGF promoted the migration and proliferation of cultured SCs, and also induced the expression of various genes such as GDNF and LIF, presumably by activating ERK pathways. Furthermore, the exogenous supply of HGF around the injury site via intramuscular injection of a plasmid DNA expressing human HGF enhanced the myelin thickness and axon diameter in injured nerves.<br/> During the above work, it was found that c-Fos is one of many cellular genes whose expressions are affected by the increased levels of HGF. c-Fos is an important component of the AP-1 heterodimer, but its role has not been clearly understood in the context of HGF and SCs. I investigated the relationship between HGF and c-Fos. First, it was confirmed that in the nerve crush model, both RNA and protein levels of c-Fos were increased, while this effect was abrogated by PHA-665752, an inhibitor of the c-met receptor. Increased expression of c-Fos was observed predominantly in distal SCs as measured by immunostaining. When primary SCs were treated with recombinant HGF protein, c-Fos expression was regulated in a typically quick and transient fashion at both RNA and proteins levels. HGF-mediated induction of c-Fos expression was highly suppressed by specific inhibitors of ERK and CREB, U0126 and CBPi, respectively. The knockdown of c-Fos expression by siRNA almost completely blocked various HGF-mediated effects in SCs—such as the induction of the gene expression of GDNF, LIF, and c-Myc, and the migration of SCs—indicating that c-Fos might play a key role in HGF effects. My thesis work revealed that HGF and c-met played important roles in Schwann cell-mediated nerve repair, that HGF gene transfer might provide a useful tool for treating peripheral neuropathy, and also that c-Fos played a key role in HGF-mediated effects on neuropathic gene and cell migration. These results show that HGF has great potential to be used as a platform for developing novel therapeutics for a variety of neurological diseases.