Effects of Altering Magnesium Metal Surfaces on Degradation In Vitro and In Vivo during Peripheral Nerve Regeneration

Abstract

In vivo use of biodegradable magnesium (Mg) metal can be plagued by too rapid a degradation rate that removes metal support before physiological function is repaired. To advance the use of Mg biomedical implants, the degradation rate may need to be adjusted. We previously demonstrated that pure Mg filaments used in a nerve repair scaffold were compatible with regenerating peripheral nerve tissues, reduced inflammation, and improved axonal numbers across a short—but not long—gap in sciatic nerves in rats. To determine if the repair of longer gaps would be improved by a slower Mg degradation rate, we tested, in vitro and in vivo, the effects of Mg filament polishing followed by anodization using plasma electrolytic oxidation (PEO) with non-toxic electrolytes. Polishing removed oxidation products from the surface of as-received (unpolished) filaments, exposed more Mg on the surface, produced a smoother surface, slowed in vitro Mg degradation over four weeks after immersion in a physiological solution, and improved attachment of cultured epithelial cells. In vivo, treated Mg filaments were used to repair longer (15 mm) injury gaps in adult rat sciatic nerves after placement inside hollow poly (caprolactone) nerve conduits. The addition of single Mg or control titanium filaments was compared to empty conduits (negative control) and isografts (nerves from donor rats, positive control). After six weeks in vivo, live animal imaging with micro computed tomography (micro-CT) showed that Mg metal degradation rates were slowed by polishing vs. as-received Mg, but not by anodization, which introduced greater variability. After 14 weeks in vivo, functional return was seen only with isograft controls. However, within Mg filament groups, the amount of axonal growth across the injury site was improved with slower Mg degradation rates. Thus, anodization slowed degradation in vitro but not in vivo, and degradation rates do affect nerve regeneration.

Volume loss during muscle reinnervation surgery is correlated with reduced CMAP amplitude but not reduced force output in a rat hindlimb model

“…Our results also suggest that the muscle fiber cross-sectional area could be positively correlated with CMAP amplitude ( R 2 = 0.208 and p = 0.001) ( Supplementary Figure S5C ), which is supported by other studies in rat hindlimb models of peripheral nerve injury; when transecting a nerve and coaptating it back together, leg circumference is an often-used indicator of the progress in nerve regeneration ( Vennemeyer et al, 2015 ; Tatu et al, 2023 ). Our results are also similar to those studies in that very high intra-group and inter-animal variability among all the measurements prevents us from drawing strong statistical conclusions with the variables that we have analyzed ( Schmalbruch, 1986 ; Tatu et al, 2023 ). With regards to muscle force production after reinnervation, our results do not demonstrate any clear patterns.…”

“…Both the muscle mass and CMAP amplitudes of reinnervated DMTs are lower than naïve positive control muscles, but in this model, removing tissue during MR surgery did not significantly impact muscle mass 100 days later (Figure 3B). Our results also suggest that the muscle fiber crosssectional area could be positively correlated with CMAP amplitude (R 2 = 0.208 and p = 0.001) (Supplementary Figure S5C), which is supported by other studies in rat hindlimb models of peripheral nerve injury; when transecting a nerve and coaptating it back together, leg circumference is an often-used indicator of the progress in nerve regeneration (Vennemeyer et al, 2015;Tatu et al, 2023). Our results are also similar to those studies in that very high intra-group and inter-animal variability among all the measurements prevents us from drawing strong statistical conclusions with the variables that we have analyzed (Schmalbruch, 1986;Tatu et al, 2023).…”

Nerve Regeneration with a Scaffold Incorporating an Absorbable Zinc-2% Iron Alloy Filament to Improve Axonal Guidance

“…The potential advantages of Zn include a slower in vivo degradation rate (better for support across longer nerve gaps, an issue that we encountered with Mg), less reactivity to water (Mg degradation produces hydrogen bubbles that can disrupt tissue attachment), and that there are greater anti-inflammatory and antisepsis effects of Zn versus Mg ions [18][19][20][21][22][23][24]. Our studies on Mg filaments for nerve regeneration showed that the Mg filaments developed gaps by six weeks in vivo, which we speculated would be too soon to provide sufficient physical support for longer injury gaps [12][13][14]. We saw that Mg filaments improved regeneration across short gaps (6 mm), but not 15 mm gaps, which is a critical size for gaps in rats and requires the metal to remain intact longer [13].…”

“…Because these metals release beneficial metal ions during degradation, they are also efficacious for soft tissue repairs. Our lab has previously shown that single Mg filaments (250-300 µm diameter) placed inside nerve conduits provided physical support, allowed cellular attachment, improved aspects of short-gap nerve growth, left no scar after degradation, and reduced tissue inflammation [12][13][14]. Others have shown that increasing systemic or local tissue levels of Mg ions improved nerve regeneration after a nerve crush [15][16][17].…”

Influence of Magnesium Degradation on Schwannoma Cell Responses to Nerve Injury Using an In Vitro Injury Model

“…Several in vitro and in vivo studies have shown that Mg improves nerve regeneration through reduced inflammation [14], enhanced remyelination and growth factor expression [15][16][17], improved neurite outgrowth [16][17][18][19], axon number [20], and improved functional recovery [15]. In particular, Li et al and Pixley et al have used Mg metal wires in their studies, with positive effects on nerve repair [15,[20][21][22]. However, while many of the in vivo studies have targeted the effects on axons and inflammation, observing the nerve as a whole, few studies have specifically studied the effects on SCs [14,23], and the in vitro studies focused mainly on cytocompatibility [24,25] or did not consider the injury microenvironment [17,19].…”