In Vivo Evaluation of Polysialic Acid as Part of Tissue-Engineered Nerve Transplants

Abstract

With the aim to develop new biomaterials for peripheral nerve grafts, the current study used bioidentical polysialic acid (polySia) as complement in synthetic conduits. polySia provides an important guidance cue during nervous system development and regeneration. First in vivo results on the use of cell-free and Schwann cell-containing synthetic peripheral nerve grafts complemented with soluble exogenous K1-polySia are presented. Reconstructing 10 mm rat sciatic nerve gaps, K1-polySia complementation significantly improved structural nerve regeneration in comparison to cell-free and K1-polySia-free grafts. Subsequently, long nerve gaps (13 mm) were reconstructed by Schwann cell transplants plus K1-polySia and compared to nerve autotransplantation. Structural but also functional regeneration could be observed using K1-polySia transplants; however, autotransplantation was still significantly more successful. Overall, the current study demonstrates that exogenous K1-polySia has no negative but rather regeneration promoting effects. This is important novel evidence on the applicability of exogenous polySia in vivo. Further studies are required to develop solid three-dimensional polySia-based scaffolds for nerve tissue engineering. Biocompatible and assessable biodegrading materials will ensure long-lasting presence of polySia to allow its applicability and prolonged efficacy in the slow regenerating scenario of human peripheral nerve reconstruction.

Evaluation of periodic electrodiagnostic measurements to monitor motor recovery after different peripheral nerve lesions in the rat

“…Foot withdrawal and vocalization were recorded as positive responses indicative of recovery. 11,35 The right intact hindpaw of the same animal served as control.…”

“…Stimulation of the sciatic nerve was carried out using a bipolar steel hook electrode. 11,19 The nerve was stimulated twice, proximal and distal to the lesion/suture site/autograft, with gradually increasing intensity from 0.1 mA until maximal augmentation (pulse duration 0.1 ms, not exceeding 8 mA). CMAPs were recorded by the electrodiagnostic device to determine amplitude, latency, and intensity of current for the first measurable CMAP and for the CMAP with the highest amplitude.…”

“…The left sciatic nerves were explanted and processed as described elsewhere. 11,19 Briefly, the nerves were fixed in Karnovsky solution (2% paraformaldehyde, 2.5% glutaraldehyde in 0.2 M sodium cacodylate buffer, pH 7.3) for 24 h. After rinsing with 0.1 M sodium cacodylate buffer containing 7.5% sucrose, postfixation was performed in 1% OsO 4 for 1.5 h. Staining of myelin sheaths was done in 1% potassium dichromate (24 h), ethanol (25%, 24 h), and hematoxylin (0.5% in 70% ethanol, 24 h). After dehydration, tissue was Epon embedded.…”

“…Quantification of the number of regenerated myelinated axons within the sections was performed using a computer macro based on AnalySIS Pro, as described elsewhere. 11,19 Because of the large size of the cross-sections, only a small part of each section was analyzed, as described by Geuna et al, 36 followed by measuring the cross-sectional area and calculating the total number of myelinated axons. The number of regenerated myelinated axons was set in relation to the whole cross-section caliber of the nerve to determine the nerve fiber density (myelinated axons/mm 2 ).…”

“…16,17 It has the advantage of being unaffected by automutilation, toe contracture, or animal behavior, but it is often only done as a final evaluation step prior to euthanizing the animal and harvesting the nerve tissue for further analyses. 11,[18][19][20][21] There have been only a few studies describing repeated electrodiagnostic measurements during long-term investigations of peripheral nerve regeneration. [22][23][24] The advantage of minimally invasive electrodiagnostic methods that allow periodic evaluation of reinnervation at predefined time intervals without euthanizing the animals or interfering with the progress of regeneration 16 seems not to be widely recognized or routinely used to assess the progress of peripheral nerve regeneration.…”