Peter J. Evans

Walking track analysis : a long-term assessment of peripheral nerve recovery

Functional recovery following sciatic, tibial, and peroneal nerve injury was assessed over a 1-year period using walking track analysis in the rat. Internal neurolysis did not affect nerve function. Crush injury induced a temporary, but complete, loss of function that recovered to control levels by 4 weeks. Nerve transaction resulted in complete loss of function without any evidence of recovery. After nerve repair, functional recovery occurred, reaching near-optimal recovery by 12 weeks. The degree of functional recovery varied with the specific nerve involved. The sciatic nerve recovered 41 percent of function, whereas the tibial nerve recovered 54 percent of function. The peroneal nerve exhibited the highest degree of recovery, achieving functional levels similar to control values. Assessment of neural regeneration using walking track analysis appears to be a valuable addition to the traditional methods of histology and electrophysiology.

COLD PRESERVED NERVE ALLOGRAFTS: CHANGES IN BASEMENT MEMBRANE, VIABILITY, IMMUNOGENICITY, AND REGENERATION

Rat sciatic nerve graft segments were harvested and pretreated by either placement in the University of Wisconsin Cold Storage Solution at 5°C and storage from 1 to 26 weeks, or repeatedly freezing (−40°C) and thawing (20°C). Following pretreatment, grafts were transplanted as either syngeneic or allogeneic nerve grafts. Storage and freeze–thawing did not affect the Schwann cell basal lamina or laminin distribution of the peripheral nerve. Graft cell viability decreased with increasing time of storage, with some viable cells detectable even after 3 weeks of storage. Freeze–thawed grafts were not viable. Increasing time of storage led to decreasing immune response and graft rejection, but improved regeneration. Freeze–thawed and 26‐week stored allografts were nonimmunogenic and rejection was not seen, but regeneration was delayed compared to autografts. Graft storage may become a useful adjunct to clinical nerve allografting to permit elective scheduling of surgery, provide greater time for preoperative tissue testing, and possibly blunt the immune response.

Selective reinnervation: a comparison of recovery following microsuture and conduit nerve repair

Selective reinnervation was studied by comparing the regeneration across a conventional neurorraphy versus a conduit nerve repair. Lewis rats underwent right sciatic nerve transection followed by one of four different nerve repairs (n = 8/group). In groups I and II a conventional neurorraphy was performed and in groups III and IV the proximal and distal stumps were coapted by use of a silicone conduit with an interstump gap of 5 mm. The proximal and distal stumps in groups I and III were aligned anatomically correct and the proximal stump was rotated 180 degrees in groups II and IV (i.e. proximal peroneal nerve opposite the distal tibial nerve and the proximal tibial nerve opposite the distal peroneal nerve). By 14 weeks, there was an equivalent, but incomplete return in sciatic function index (SFI) in groups I, III, and IV as measured by walking track analysis. However, the SFI became unmeasurable by 6 weeks in all group II animals. At 14 weeks, the percent innervation of the tibialis anterior and medial gastronemius muscles by the peroneal and tibial nerves respectively was estimated by selective compound muscle action potential amplitude recordings. When fascicular alignment was reversed, there was greater tibial (P = 0.02) and lesser peroneal (P = 0.005) innervation of the gastrocnemius muscle in the conduit (group IV) versus the neurorraphy (group II) group. This suggests that the gastrocnemius muscle may be selectively reinnervated by the tibial nerve. However, there was no evidence of selective reinnervation of the tibialis anterior muscle. Despite these differences, the functional recovery in both conduit repair groups (III and IV) was equivalent to a correctly aligned microsuture repair (group I) and superior to that in the incorrectly aligned microsuture repair (group II).

Strain differences in autotomy in rats undergoing sciatic nerve transection or repair

&NA; The degree of self‐mutilation (autotomy) following sciatic nerve injury was assessed in 6 rat strains. Experimental groups included sciatic nerve transection with and without repair and crush lesions. The degree of autotomy was measured using a standard grading system. There were statistically significant differences between the strains in the transected group in terms of both degree of autotomy and the time of onset of its appearance. The repair group showed a shorter mean time of onset, lower maximal scores, and lower percentage of affected animals. Autotomy was absent in the nerve crush group and in all Lewis rats. In experimental studies when the status of the foot is critical (e.g., to evaluate functional walking patterns) Lewis rats appear to be the most appropriate strain to utilize. These results suggest that there are definite strain differences in the degree of autotomy following nerve transection or repair.

Walking Track Analysis: Utilization of Individual Footprint Parameters

Functional assessment of rat sciatic, tibial, and peroneal nerve injuries was performed using walking track analysis. Individual walking print length (PL), toe spread (TS), and intermediate toe spread (ITS) values were measured up to 24 weeks after specific nerve transection, with or without repair. Sciatic and tibial nerve manipulation initially affected all footprint measurements, consistent with loss of intrinsic and extrinsic motor function. After sciatic repair, TS demonstrated partial recovery without any substantial recovery in PL or ITS, compared with sciatic transection values. By contrast, after tibial repair, PL values recovered dramatically, between 16 and 24 weeks, to levels not significantly different from control subjects. This was not observed after tibial transection without repair. TS recovered partially, whereas ITS recovered to control levels by 20 weeks after tibial repair. Peroneal transection resulted in multiple contractures, rendering this group unmeasurable at 4 weeks. After peroneal repair, only the PL reflected significant loss of function at 2 weeks, recovering to control values by 8 weeks. Manual TS measurements in nonwalking rats did not reflect functional nerve regeneration. Thus, individual PL measurements alone can be used to characterize functional recovery after tibial and peroneal nerve injury, whereas TS reflected recovery after sciatic nerve injury.

Regeneration across cold preserved peripheral nerve allografts

The feasibility of peripheral nerve allograft pretreatment utilizing cold storage (5°C in the University of Wisconsin Cold Storage Solution) or freeze‐thawing to prevent rejection was investigated. Regeneration across cold‐stored (3 or 5 weeks) or freeze‐thawed (FT), 3.0‐cm sciatic nerve allografts were compared to fresh auto‐ and allografts in an inbred rat model. At 16‐week post‐engraftment, only FT allografts appeared similar to autografts on gross inspection; FT grafts were neither shrunken nor adherent to the surrounding tissue as seen in the other allograft groups. Qualitatively, the pattern of regeneration in the graft segments of the fresh allograft and to a lesser extent of pretreated allografts was inferior to that of autografts as evidenced by a disruption in the perineurium, more extrafascicular axons, smaller and fewer myelinated axons, increased intrafascicular collagen deposition, and the persistence of perineurial cell compartmentation and perivascular infiltrates. Distal to these grafts, the regeneration became more homogenous between groups, although areas of ongoing Wallerian degeneration, new regeneration as well as compartmentation, were more prevalent in fresh and pretreated allografts. Although the number of myelinated fibres was equivalent to autografts, the fibre diameters, the number of large diameter fibres, and the G‐ratio were significantly decreased in the allograft groups, which, in part, accounted for the significant decrease in conduction velocity in the 3‐week stored and fresh allograft, and the slight decrease in the 5‐week stored and FT allograft groups. There was a small return in the Sciatic Function Index towards normal, but no consistent differences between groups were found. Prolonged cold storage and freeze‐thawing of nerve allografts resulted in regeneration that was better than fresh allografts, but inferior to autografts. With the concomitant use of host immunosuppression or other immunotherapies, these storage techniques can provide a means of transporting nerve allografts between medical centres and for converting urgent into elective procedures.

The peripheral nerve allograft in the primate immunosuppressed with cyclosporin A. I: Histologic and electrophysiologic assessment

Nerve regeneration across peripheral nerve allografts and control autografts in primates immunosuppressed with Cyclosporin A was quantitatively evaluated by electrophysiologic and histologic methods. Twelve cynomolgus monkeys received 3-cm autografts and allografts in contralateral ulnar nerves. They were immunosuppressed with Cyclosporin A at 25 mg/kg per day or placebo vehicle. Morphometric analysis of nerve graft and distal nerve segments was assessed at 1 year after engraftment. Quantitative electrophysiologic studies were performed percutaneously at 6 and 12 months, and compound action potentials were measured directly across the nerve grafts at 1 year. Excellent regeneration was seen across autografts and allografts in Cyclosporin A-treated and placebo-treated recipients.

Phenotypic analysis of migrant, efferent lymphocytes after implantation of cold preserved, peripheral nerve allografts

Cold-preservation of peripheral nerve allografts in vitro (3 weeks, 5 degrees C) was performed to determine its effect on local lymphocyte migration patterns in vivo. Lymphocyte migration was assessed by continuously monitoring the cell output in the regional lymph for nearly 1 month. Cold-preservation delayed or prevented the typical biphasic increase in efferent lymphocyte output observed after fresh allograft implantation. It also decreased the output of activated lymphocytes (CD 5 and MHC class II positive) compared with that seen in the fresh allograft response. These changes suggest that the host immune response to preserved nerve allografts is altered over a prolonged period in vivo (3 weeks). Cold-preservation may be a useful method of reducing allograft immunogenicity, thereby limiting systemic immunosuppression requirements for the successful clinical utilization of peripheral nerve allografts.

Verification of a free vascularized nerve graft model in the rat with application to the peripheral nerve allograft.

We report a vascularized sciatic nerve graft in the rat, based upon the femoral popliteal superior muscular artery pedicle. The pedicle may be raised from the common femoral artery via the popliteal artery, with accompanying venous drainage to the femoral vein. We have characterized two vessels: the middle tibial artery to the posterior tibial nerve and the arteria comitas nervus peroneus to the peroneal nerve. Plastic monomer and Evans blue labeled albumin injections established the anatomic model. The graft was reliably perfused at the time of isolation and 24 hours post-transplantation. The model was used to evaluate regeneration across allogeneic (ACI to Lewis, n = 12) versus syngeneic (Lewis to Lewis, n = 6) nerve grafts. Electrophysiologic and histomorphometric assessments demonstrated that the vascularized immunosuppressed allograft was similar to the vascularized syngeneic graft. Both were suerior to the vascularized allograft without immunosuppression. (Plast. Reconstr. Surg. 92: 516, 1993.)

A Comparison of Volume Loading and Atrial Pacing Following Aortocoronary Bypass

Although cold potassium cardioplegia provides adequate myocardial protection, transient hemodynamic and metabolic instability occasionally occurs after uncomplicated coronary bypass surgery. Two methods to increase cardiac output were compared 2 to 6 hours postoperatively in 24 patients recovering from elective coronary bypass operation. Volume loading increased cardiac index (CI) from 2.1 +/- 0.5 to 2.7 +/- 0.6 L/min/m2 by increasing left atrial pressure (LAP) from 8.6 +/- 3.6 to 13.0 +/- 4.1 mm Hg. Atrial pacing at a rate of 112 +/- 8 beats per minute increased CI from 2.4 +/- 0.5 to 2.7 +/- 0.8 L/min/m2 without a change in LAP. Ejection fraction by nuclear angiography did not change, but the calculated left ventricular end-diastolic volume index (stroke index/ejection fraction) increased with volume loading and decreased with atrial pacing--a decrease in diastolic compliance. Myocardial oxygen extraction did not change, but myocardial lactate extraction increased with volume loading and decreased with atrial pacing. Coronary sinus blood flow was measured in 5 patients and increased with both methods studied. Volume loading demonstrated that myocardial performance was normal and myocardial metabolism increased commensurate with the increase in work. Atrial pacing increased CI but resulted in anaerobic metabolism and a decrease in diastolic compliance. Volume loading rather than atrial pacing will improve CI without producing ischemia in the early postoperative period.