Lolita Singh

Abstract

Damage to the peripheral nerves causes a series of molecular, cellular, and structural changes critical to initiating a successful regenerative process involving degeneration, inflammation, axon re-growth and innervation of the denervated target, and finally functional recovery. In this thesis I sought to study the expression and role of an axon guidance molecule - Netrin-1 in peripheral nerve repair. Using a Schwann cell-specific Netrin-1 knockout mouse model, I showed an increased disorganisation and misdirection of re-growing axons within the nerve bridge following transection injury. However, this did not affect the number of axons innervating the distal nerve. Moreover, functional recovery was not impacted after a crush injury. Additional studies on axon outgrowth following crush injury, and analysis of Schwann cell proliferation, migration and remyelination of axons following injury, showed that Schwann cell-derived Netrin-1 is expendable for these functions. Furthermore, on analysing recently compiled single-cell RNA sequencing data for the sciatic nerve following injury, I report the highest expression of Netrin-1 in perineurial cells with low expression of Netrin-1 in Schwann cells both before and after injury. Therefore, I attempted to generate a global knockout of Netrin-1 in adult mice to study the effect of the overall loss of Netrin-1 on peripheral nerve regeneration, however this attempt unfortunately proved to be unsuccessful. Netrin-1 has been shown to mediate attraction on growing axons during development by binding to its receptor - deleted in colorectal carcinoma (DCC). To study the role of DCC in peripheral nerve injury and repair, I generated a global knockout of DCC in adult mice using a Tamoxifen inducible Cre recombinase system. A transection injury in DCC global knockout mice revealed aberrant trajectory of axons in the nerve bridge, however innervation of the distal nerve was not affected. Similar to the Schwann cell-specific Netrin-1 mouse model, functional recovery was comparable to the control following a crush injury. My data suggested that DCC might not be important for successful end-target innervation of axons as well as functional recovery following peripheral nerve injury. The final part of my thesis focused on studying the role of runt related transcription factor-2 (Runx2) in peripheral nerve regeneration. While the published literature and my in-silico analysis showed significant upregulation of Runx2 in Schwann cells following peripheral nerve injury, Runx2 has no previously established role in peripheral nerve repair. Therefore in collaboration with other members of my research group we established a Schwann-cell specific Runx2 knockout mouse model and investigated the effect of loss of Runx2 on the events of peripheral nerve repair. My work on the Runx2 mouse model showed normal myelination of axons by Schwann cells during development, however remyelination was impaired following a crush injury in the sciatic nerve. I also attempted to validate three predicted injury associated Runx2 targets; however, my results showed that they were not apparently regulated by Runx2. My results for this project, along with that of my research group identified Runx2 as a novel regulator of Schwann cell plasticity with a role in the regulation of peripheral nerve regeneration biology.

Keywords

Peripheral nerve regeneration, Netrin-1, Deleted in colorectal carcinoma (DCC), Runt-related transcription factor 2 (Runx2)

Document Type

Thesis

Publication Date

2023

Embargo Period

2024-07-21

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