Matrix metalloproteinases as promising regulators of axonal regrowth in the injured adult zebrafish retinotectal system

Abstract

Overcoming the failure of axon regeneration in the mammalian central nervous system (CNS) after injury remains a major challenge, which makes the search for proregenerative molecules essential. Matrix metalloproteinases (MMPs) have been implicated in axonal outgrowth during CNS development and show increased expression levels during vertebrate CNS repair. In mammals, MMPs are believed to alter the suppressive extracellular matrix to become more permissive for axon regrowth. We investigated the role of MMPs in axonal regeneration following optic nerve crush (ONC) in adult zebrafish, which fully recover from such injuries due to a high intrinsic axon growth capacity and a less inhibitory environment. Lowering general retinal MMP activity through intravitreal injections of GM6001 after ONC strongly reduced retinal ganglion cell (RGC) axonal regrowth, without influencing RGC survival. Based on a recently performed transcriptome profiling study, the expression pattern of four MMPs after ONC was determined via combined use of western blotting and immunostainings. Mmp-2 and -13a were increasingly present in RGC somata during axonal regrowth. Moreover, Mmp-2 and -9 became upregulated in regrowing RGC axons and inner plexiform layer (IPL) synapses, respectively. In contrast, after an initial rise in IPL neurites and RGC axons during the injury response, Mmp-14 expression decreased during regeneration. Altogether, a phase-dependent expression pattern for each specific MMP was observed, implicating them in axonal regrowth and inner retina remodeling after injury. In conclusion, these data suggest a novel, neuron-intrinsic function for multiple MMPs in axon regrowth that is distinct from breaking down environmental barriers. J. Comp. Neurol. 524:1472-1493, 2016. © 2015 Wiley Periodicals, Inc.

Identification of target genes downstream of semaphorin6A/PlexinA2 signaling in zebrafish

“…mmp2 has a 1.33 logFC in sema6a and a 1.31 logFC in plxnaA2 morphants (Table 1). Recently, mmp2 was shown to be important for degrading nonpermissive ECM to allow for zebrafish retinal ganglion cell axon outgrowth during recovery from injury (Lemmens et al, 2016). Increases in MMP2 could lead to increased degradation of the ECM surrounding RPCs, and removal of key proteins required transverse 20 mm sections through 18 somite stage brain and eye vesicles processed for in situ hybridization with rasl11b and shtn-1 probes.…”

Complementary research models and methods to study axonal regeneration in the vertebrate retinofugal system

“…In the first model, the nerve is cut with scissors, thereby creating a proximal and distal nerve stump that are physically separated from each other. In contrast, in the ONC model, the nerve is crushed with forceps to separate the axons, but leaving the connective tissue around them intact, and thus holding the proximal and distal ends together (Becker et al 2000;McCurley and Callard 2010;Fleisch et al 2011;Lemmens et al 2015;Van Houcke et al 2017). As expected, the ONT model is somewhat more drastic and regeneration progresses slower as compared to ONC (Zou et al 2013).…”

“…Frequently used anterograde tracers are tagged dextrans (Elsaeidi et al 2014) and biocytin. The latter is transported very rapidly, and labeling axons in the adult zebrafish optic tectum takes only about 2.5-4 h. Making use of biocytin's high affinity for avidin, traced axons in the tectum can be easily visualized with common immunohistochemistry techniques (Becker et al 2000(Becker et al , 2004Munzel et al 2014;Bhumika et al 2015;Lemmens et al 2015Lemmens et al , 2016.…”

Axonal regeneration in zebrafish spinal cord

“…The secretion of MMPs can be seen in both mammalian and fish CNS injury. Dynamic expression of four specific MMPs (MMP‐2, MMP‐9, MMP‐13a, and MMP‐14) during different phases of retinotectal regeneration has been reported in zebrafish (Lemmens et al., ; McCurley & Callard, ). Our microarray analysis data demonstrate upregulation of several MMP genes in different phases after SCI in adult zebrafish.…”