Can an injured nerve be used as a donor nerve for distal nerve transfer?-An experimental study in rats

Abstract

Background:
Distal nerve transfer has proven efficacy. The purpose of this study was to investigate if an injured nerve can be used as a donor nerve for transfer, and to determine the threshold of injury.

Materials and methods:
Rat's left ulnar-nerves in the axilla with different degrees of injury were selected as the donor nerves for transfer, and the musculocutaneous-nerves the target nerves for being re-innervated. Six rats each served as positive and negative controls: Group A, intact ulnar-nerve transfer; and Group E, the ulnar-nerve was cut but no transfer. Ten rats each were assigned to Group B to Group D with 25%, 50%, and 75% transected ulnar-nerve, respectively and all were transferred to the musculocutaneous-nerve. After a 12-week recovery period, outcomes were evaluated.

Results:
Biceps muscle weight measurements showed all experimental groups-D 0.28 ± 0.02 g/72%, C 0.28 ± 0.03 g/73%, B 0.29 ± 0.04 g/74%, and A 0.29 ± 0.04 g/80%-were lighter than group H 0.36 ± 0.04 g, which were all statistically significant (P < 0.001). Muscle tetanus contraction force measurements were the lowest in group D35 ± 8.6 g/69%. Groups C and B measured 41 ± 8.5 g/75% and 40 ± 2.2 g/77% and group A 41 ± 9.4 g/95%, respectively. Group H showed muscle contraction force of 52 ± 7.2 g, which was statistically significant when compared to experimental groups (P < 0.05-0.001). EMG measurements of the biceps muscles showed: group D was 3.6 ± 0.7 mV/69%, group C was 3.6 ± 0.6 mV/75%, and group B was 4.2 mV ± 0.7/81%. Group H was5.1 ± 0.7 mV and statistically significant different when compared with experimental groups (P < 0.05-0.001).Axon counts of healthy ulnar-nerve (Group H) were 1849 ± 362. Axon counts of the injured ulnar-nerve were in group B 1447 ± 579/78%, group C 1051 ± 367/57% and group D 567 ± 230/31%.

Conclusion:
The donor nerve should be healthy in order to provide optimal result. A big nerve (e.g., ulnar nerve) but injured with at least 75% axon spared is still potentially effective for transfer. In contrast, a small nerve (e.g., intercostal nerve) once injured with 75%axon spared would be considered a suboptimal donor nerve.

Full text

RESEARCH ARTICLE
Can an injured nerve be used as a donor nerve for distal nerve
transfer?—An experimental study in rats
Chieh-Han John Tzou, MD, PhD
1,2
|
Chuieng-Yi Johnny Lu, MD
1
|
Tommy Naj-Jen Chang, MD
1
|
David Chwei-Chin Chuang, MD
1
1
Division of Reconstructive Microsurgery,
Department of Plastic Surgery, Chang Gung
Memorial Hospital, Taoyuan, Taiwan
2
Division of Plastic and Reconstructive
Surgery, Department of Surgery, Medical
University of Vienna, Vienna, Austria
Correspondence
David Chwei-Chin Chuang, MD, Professor,
Division of Reconstructive Microsurgery,
Department of Plastic Surgery, Chang Gung
Memorial Hospital, Chang Gung Medical
College, Chang Gung University, 5
Fu-HsingStreet, Kuei-Shan, Taoyuan 333,
Taiwan.
Email: dccchuang@gmail.com
Funding information
National Science Council Taiwan, Grant
Number: NSC 101-2314-B-182A-033-MY3
Background: Distal nerve transfer has proven efficacy. The purpose of this study was to investi-
gate if an injured nerve can be used as a donor nerve for transfer, and to determine the threshold
of injury.
Materials and Methods: Rat’s left ulnar-nerves in the axilla with different degrees of injury were
selected as the donor nerves for transfer, and the musculocutaneous-nerves the target nerves for
being re-innervated. Six rats each served as positive and negative controls: Group A, intact ulnar-
nerve transfer; and Group E, the ulnar-nerve was cut but no transfer. Ten rats each were assigned
to Group B to Group D with 25%, 50%, and 75% transected ulnar-nerve, respectively and all were
transferred to the musculocutaneous-nerve. After a 12-week recovery period, outcomes were
evaluated.
Results: Biceps muscle weight measurements showed all experimental groups—D0.2860.02 g/
72%, C 0.28 60.03 g/73%, B 0.29 60.04 g/74%, and A 0.29 60.04 g/80%—were lighter than
group H 0.36 60.04 g, which were all statistically significant (P<0.001). Muscle tetanus contrac-
tion force measurements were the lowest in group D35 68.6 g/69%. Groups C and B measured
41 68.5 g/75% and 40 62.2 g/77% and group A 41 69.4 g/95%, respectively. Group H showed
muscle contraction force of 52 67.2 g, which was statistically significant when compared to
experimental groups (P<0.05–0.001). EMG measurements of the biceps muscles showed: group
D was 3.6 60.7 mV/69%, group C was 3.6 60.6 mV/75%, and group B was 4.2 mV60.7/81%.
Group H was5.1 60.7 mV and statistically significant different when compared with experimental
groups (P<0.05–0.001).Axon counts of healthy ulnar-nerve (Group H) were 18496362. Axon
counts of the injured ulnar-nerve were in group B 14476579/78%, group C 1051 6367/57%
and group D 5676230/31%.
Conclusion: The donor nerve should be healthy in order to provide optimal result. A big nerve (e.
g., ulnar nerve) but injured with at least 75% axon spared is still potentially effective for transfer.
In contrast, a small nerve (e.g., intercostal nerve) once injured with 75%axon spared would be con-
sidered a suboptimal donor nerve.
1
|
INTRODUCTION
In brachial plexus injury (BPI), nerve transfer (or neurotization) is a
nerve reconstruction technique, and very often being used to reinner-
vate the distal part of an avulsed, irreparable, but important nerve with
low sequel (Chuang, 2005; Narakas, 1978). The majority of nerve trans-
fers are for motor neurotization (Alnot, 1978; Bertelli & Ghizoni, 2004;
Brown, Shah, & Mackinnon, 2009; Chuang, 2005, 2006, 2008; Chuang,
Lee, Hashem, & Wei, 1995; Mackinnon, Novak, Myckatyn, & Tung,
2005; Narakas, 1978; Oberlin, Beal, Leechavengvongs, Salon, Dauge, &
Sarcy, 1994). Functioning free muscle transplantation in BPI is also
another example of the use of nerve transfer (Chuang, 2008).
Nerve transfer in BPI can be broadly classified into proximal and
distal nerve transfer (Chuang, 2005). Proximal nerve transfer (Alnot,
1978; Chuang, 2006; Chuang, Lee, Hashem, & Wei, 1995) is a tradi-
tional technique, involving brachial plexus exploration which permits
Microsurgery 2017; 1–8;
DOI: 10.1002/micr.30153
wileyonlinelibrary.com/journal/micr V
C2017 Wiley Periodicals, Inc.
|
1
Received: 19 June 2015
|
Revised: 1 December 2016
|
Accepted: 16 December 2016
DOI 10.1002/micr.30153

confirmation of the diagnosis of root avulsion, and the nerve coaptation
is inside the brachial plexus zones (supra-or infraclavicular). Distal nerve
transfer (Bertelli & Ghizoni, 2004; Brown, Shah, & Mackinnon, 2009;
Mackinnon, Novak, Myckatyn, & Tung, 2005; Oberlin, Beal, Leecha-
vengvongs, Salon, Dauge, & Sarcy, 1994; Tzou, Chuang, Chang, & Lu,
2014) is a newer strategy in which the nerve coaptation is performed
distal to the brachial plexus. Using parts of fascicles of ulnar/median
nerve transfer to biceps/brachialis branch of the musculocutaneous
nerve in C5-C6 root avulsion injuries is a classic example of distal nerve
transfer (Bertelli & Ghizoni, 2004; Brown, Shah, & Mackinnon, 2009;
Chuang, 2006; Mackinnon, Novak, Myckatyn, & Tung, 2005; Oberlin,
Beal, Leechavengvongs, Salon, Dauge, & Sarcy, 1994). Distal nerve
transfers have benefits of direct nerve repair without nerve graft, no
scar disturbance, short operation and rehabilitation times. Distal nerve
transfers have become increasingly popular currently.
Although most patients achieved good elbow flexion by ulnar
nerve fascicular transfer to the musculocutaneous nerve, some cases
still required long rehabilitation or additional procedure to enhance
the result (Bertelli & Ghizoni, 2004; Mackinnon, Novak, Myckatyn, &
Tung, 2005; Oberlin, Beal, Leechavengvongs, Salon, Dauge, & Sarcy,
1994). One reason for this variability is the fact that in traction avul-
sion injury of the BPI, multiple roots are often involved beyond the
expectation. For example, a patient has a functional hand with grossly
good grip strength. He might be a case of C5-6 two root avulsion, or
C5-7 three root avulsion with intact C8-T1, or C5-8 four root avulsion
with intact T1. In such circumstance the health of ulnar nerve (fibers
from C8-T1), or median (C6-T1) is actually questionable. Although few
studies have shown that an injured nerve can be used as a donor
(Chuang & Hernon, 2012; Totosy de Zepetnek, Zung, Erdebil, &
Gordon, 1992; Tzou, Chuang, Chang, & Lu, 2014), it is often difficult
for a surgeon to judge whether a macroscopically normal-looking
nerve is suitable for transfer. The goal of this study was through
experimental rats to investigate if an injured nerve can be used as a
donor nerve for transfer, and to determine the threshold of injury
beyond which the donor nerve is no longer effective.
2
|
MATERIALS AND METHODS
In forty-two female Sprague–Dawley rats, each weighing 250 g, the
left upper limb was used as the experimental site. The ulnar nerve in
the axilla was chosen as the motor donor nerve for transfer, and its
neighboring musculocutaneous nerve as the target nerve to be neuro-
tized. They were randomly separated into five groups based on health
of the ulnar nerves and procedure:
Group A (n56) was the positive control group, in which the ulnar
nerve was intact. In group B (n510), one quarter (25%) of the ulnar
nerve, in group C (n510), one half (50%) of the ulnar nerve and in
group D (n510), three quarter (75%) of the ulnar nerve was trans-
ected. Group E (n56) was the negative control group, in which the
whole ulnar nerve was transected, but not transferred.
The musculocutaneous nerve was divided at 1.0 cm from the
medial biceps muscle edge. The proximal stump was embedded in the
nearby pectoralis muscle to avoid collateral sprouting. In Group A-D,
the donor ulnar nerves were transected and coapted to the musculocu-
taneous nerve (Figure 1A–D). In all experimental rats, the musculocuta-
neous nerve distal to the biceps branch nerve was uniformly cut and
transferred back into the biceps muscle to avoid loss of regenerated
axons (Rodriguez, Chuang, Chen, Chen, Lyu, & Ko, 2011).
2.1
|
Surgical procedures
All surgical procedures were performed by the first author using sterile
conditions and under general anesthesia by inhalation of isofluorane
(Halocarbon Laboratories, River Edge, NJ). The animals were placed in
the supine position, shaved, and prepared with antiseptic solution.
Under an operating microscope, a transverse ventral incision was made
from the clavicle down to the medial arm of the left upper forelimb.
The pectoralis muscles were elevated. The ulnar and nearby musculo-
cutaneous nerves were identified using a nerve stimulator (Vari-Stim
hand-held nerve locator/stimulator, Medtronic Xomed, Minneapolis,
MN).
At 1.5 cm distal to the median nerve origin (merging point of lat-
eral and medial cords), the ulnar nerve was penetrated by a 9–0nylon
needle with a variously cut surface (25, 50, and 75%), and ligated. The
ligated stitch was used for traction. Through ventero-laterally intraneu-
ral dissection, the ulnar nerve was split proximally to the point level of
median nerve bifurcation. The stump with the traction stitch was
implanted into the chest muscle to avoid collateral sprouting. The split
ulnar nerve at the point of the beginning dissection was divided and
preparing for transfer (Figure 2). The nearby musculocutaneous nerve
was cut at 1.0 cm from the medial biceps muscle edge. The transect
ulnar nerve was coapted to the musculocutaneous nerve in a tension-
free end-to-end fashion, using two 11–0 nylon sutures.
Closure of wounds was carried out with nylon 4–0sutures.Post-
operatively, animals were re-warmed and returned to their cage and
subsequently maintained on standard rat chow and water ad libitum
with a 12-h light-dark cycle. All groups were blinded for postoperative
assessments.
Regeneration for 12 weeks was allowed. Functional outcomes
were evaluated with behavioral assessment (grooming test), compound
muscle action potentials (CMAPs), biceps muscle weight, biceps tetanus
contraction force, and histomorphometry of the ulnar and musculocu-
taneous nerves with axon counts. All assessments were run bilaterally
with the nonoperative side serving as each animal’sowncontrol.
2.2
|
Behavioral analysis—grooming test
Functional outcomes of the biceps muscle were evaluated using the
grooming test developed by Bertelli & Mira (1993) and Inciong,
Marrocco, & Terzis (2000) which assesses shoulder abduction and
elbow flexion. Water (1–3 ml) was applied over the animal’s snout to
provoke a reproducible grooming response: attempting to remove
drops of water from their heads, animals raised and elevated their
forelimbs behind the ears, and then brought them down to the snout
and licked. Digital video recordings were assessed in slow motion to
2
|
TZOU ET AL.

categorize the forelimb function on a five-point scale: 5 points if the
paw reached behind the ear, 3 points if the paw passed the snout but
did not reach the eye, and 1 point if the paw moved but did not reach
the snout. Multiple assessments were performed and the best score
was recorded.
2.3
|
Biceps compound muscle action potentials
At 12 weeks, after the grooming test, the rats’operative and nonope-
rative biceps muscle and musculocutaneous nerve were exposed and
examined. Under inhalation anesthesia, a recording electrode was
placed in the biceps muscle and a subcutaneous ground electrode posi-
tioned adjacently in the electromyogram setup. Two small hook-
shaped stimulating electrodes (2-mm apart) held the musculocutaneous
gently. Stimulation was delivered for each trial by an electrical stimula-
tor (Biopac System, BSL Software Installation Package, Windows,
Goleta, CA), fixed at 1 msec at a constant current between 10 mAupto
10 mA. A single shock was delivered for each trial. The musculocutane-
ous nerve CMAPs, were recorded. The similar procedure was per-
formed on the non-operativeside for control data.
FIGURE 2 The detail of ulnar nerve transect and preparation for transfer (see Text)
FIGURE 1 (A) Schematic drawing of the Group A (positive control). UNp, ulnar nerve proximal stump;,UNd, ulnar nerve distal stump; MCn,
musculocutaneous nerve; Biceps, Biceps muscle. (B) Schematic drawing of the experimental Group B (25% transected). UNp, ulnar nerve
proximal stump; UNd, ulnar nerve distal stump; MCn, musculocutaneous nerve; Biceps, Biceps muscle. (C) Schematic drawing of the
experimental Group C (50% transected). UNp, ulnar nerve proximal stump; UNd, ulnar nerve distal stump; MCn,musculocutaneous nerve;
Biceps, Biceps muscle. (D) Schematic drawing of the experimental Group D (75% transected). UNp, ulnar nerve proximal stump; UNd, ulnar
nerve distal stump; MCn, musculocutaneous nerve; Biceps, Biceps muscle
TZOU ET AL.
|
3

2.4
|
Biceps muscle tetanus contraction
force measurement
After the electrophysiological study, the biceps muscle contraction
force was assessed with a force displacement transducer and compu-
terized recording software (FT03 Force Displacement Transducers,
Grass Instruments, Quincy, MA). Force measurement followed a modi-
fied protocol for rats based on Terzis, Sweet, Dykes, & Williams (1978)
and Shibata, Breidenbach, Ogden, & Firrell (1991)procedure. The rest-
ing muscle length of the biceps was determined, and its insertion was
detached from the cubital region and then sutured to the force trans-
ducer with the muscle resting length. The shoulder, elbow and wrist
joints were immobilized with fixation pins to avoid motion artifacts dur-
ing muscle-contraction measurements. Stimulating current was applied
with a bipolar platinum electrode to the musculocutaneous nerve. Per
definition, the threshold stimulus was a stimulus to produce an observ-
able muscle twitch (activating the biceps muscle) at different thresholds
(1–10 times the threshold), voltages (range: 0.6 and 1.2 V) and frequen-
cies (range: 1.0–60 Hz). With various stimuli, the maximal tetanic
strength was determined at 1 V and 60 Hz, and the mean maximal iso-
metric tetanic muscle contraction forces of the repeated muscle con-
traction (five times with a pulse duration of 1.0 msec) were recorded as
grams/weight. All data were analyzed and documented with MacLab
Systems (AD Instruments, Colorado Springs, CO). The optimal rest and
the maximal tetanic tension were assessed under isometric conditions.
2.5
|
Biceps muscle weights
After all in vivo experiments were finished the animals were euthanized
with an overdose of pentobarbital. Biceps muscles were harvested and
weighed immediately.
2.6
|
Axon counts
Nerve specimens (3–5 mm in length in each) were obtained: injured
ulnar nerve before the split and before the coaptation site, musculocu-
taneous nerves before its entry into the biceps muscle (Figure 3), and
the healthy ulnar nerve on the nonoperative limb. Samples were fixed
in 2.5% glutaraldehyde and postfixed in 2% osmium tetroxide. Each
nerve was embedded in 100% Epon. One-micrometer–thick transverse
sections were made from the nerve to obtain successive sections in 1-
mm intervals, which were stained with 2% toluidine blue and photo-
graphed under a light microscope with 4003magnification. Myelinated
axons were counted with 400x magnification under an Olympus BX53
microscope (Olympus Corp., Japan) using Image-Pro Plus software
(Media Cybernetics, Rockville, MA).
2.7
|
Statistical analysis
Data were expressed as mean 6standard deviation in each group. All
results were compared with values of the nonoperative right limb. To
avoid animals’weight-dependent discrepancies among animals and to
provide an internal control, results were expressed in mean absolute
values 6standard deviation and as ratios (operative: nonoperative limbs
of the same animal) (Table 1).
The Kruskal–Wallis test was performed to compare performances
of each group regarding grooming, muscle–force contraction, muscle
weight and electromyography. Dunn’s test was used as a post-hoc test
when groups were significantly different (P<0.05) by the Kruskal–
Wallis test. A Mann–Whitney test was used to compare the axon
counts between the groups. Pvalues <0.05 was considered statis-
tically significant. Statistical analysis was performed with IBM SPSS
Statistics software, version 20.0 (IBM, Armonk, NY, 2011) by the Bio-
statistical Center for Clinical Research, Chang Gung Memorial Hospital,
Linkou, Taiwan.
3
|
RESULTS
None of the experimental rats (Group A–D) reached normal status (the
value of the contralateral, nonoperative side) for any of the four param-
eters: grooming test, CMAPs, tetanic muscle contraction force, and
biceps muscle weight. Regardless of the amount of injury to the donor
nerve, all showed significant differences (with Kruskal–Wallis statistical
analysis) but with various pvalues: when compared with the non-
operative side (Group H), Group D was the worst, all four tests showed
P<0.001; Group B, two tests (muscle weight and tetanic contracture
force) showed P<0.001, but two test (CMAP and grooming test)
P<0.05; Group A only one (muscle weight) showed P<0.001. (Table
1, Column I).
3.1
|
Behavioral analysis (grooming test)
The grooming test on the healthy (right) side showed the paw could
pass at least the snout and scored on average 3.3360.50 in all groups.
The experimental (left) sides showed scores of 2.6360.74 (80% of
healthy side) in group A; 2.56 60.53 (77% of healthy side) in group B;
and 2.57 60.53 (64% of healthy side) in groups C and D. Statistically
significant differences (from P<0.001 to P<0.05) were observed
between the experimental and healthy limbs (Table 1). Regardless of
the amount of injury to the donor nerve, none of the animals reached
FIGURE 3 Schematic drawing to illustrate the nerve biopsy sites
for istomorphometry
4
|
TZOU ET AL.

TABLE 1 Results in absolute values (gram or mV) and in ratio (% of healthy side)
Groups Biceps muscle weight Tetanic muscle contraction force Compound muscle action potentials Grooming test
Gram (ratio, %
of healthy side)
Gram
H: healthy
Gram (ratio, %
of healthy side)
Gram
H: healthy
mV (ratio, %
of healthy side)
mV
H: healthy
Score (ratio, %
of healthy side)
Score
H: healthy
Experiment side side Experiment side side Experiment side side Experiment side side
A: positive control 0.29 60.04
(80% 66)
0.36 60.04 40.95 69.37
(95% 613)
44.26 68.93 4.53 60.64
(95% 616)
4.81 60.65 2.63 60.74
(80% 626)
3.38 60.52
B: ulnar nerve—25% Injury 0.29 60.04
(74% 611)
0.40 60.04 40.18 62.19
(77% 67)
52.41 65.40 4.24 60.72
(81% 612)
5.24 60.8 2.56 60.53
(77% 614)
3.33 60.50
C: ulnar nerve—50% Injury 0.28 60.03
(73% 67)
0.39 60.03 40.92 68.51
(75% 69)
54.54 68.94 3.59 60.58
(75% 613)
4.85 60.7 2.57 60.53
(64% 613)
4.00 60.64
D: ulnar nerve—75% Injury 0.28 60.02
(72% 66)
0.38 60.02 34.70 68.64
( 69% 615)
55.44 65.4 3.62 60.67
(69% 614)
5.27 60.61 2.57 60.53
(64% 613)
4.00 60.64
(I) Pvalues in absolute
values, compared to
healthy side (KW test)
Group A vs. Group H
(P<0.001), Group B vs.
Group H (P<0.001),
Group C vs. Group H
(P<0.001), Group D vs.
Group H(P<0.001)
Group B vs. Group H
(P<0.001), Group C vs.
Group H (P<0.05), Group
D vs. Group H (P<0.001)
Group B vs. Group H
(P<0.05), Group C
vs. Group H
(P<0.001), Group
D vs. Group H
(P<0.001)
Group A vs. Group H
(P<0.05), Group B
vs. Group H
(P<0.05),
Group C vs. Group H
(P<0.001),
Group D vs. Group H
(P<0.001)
(II)
Pvalues in absolute values
intergroup comparison [g,
mV] (Sig. diffby post-hoc
Dunn test)
Group A vs. Group B,
Group A vs. Group C, ...No
statistic significance.
No statistic significance. No statistic
significance.
No statistic signifi-
cance.
(III) Pvalues in ratio [% of
healthy side] (Sig. diffby
post-hoc Dunn test)
Group A vs. Group C
(P50.0378),
Group A vs. Group B
(P50.0026), Group A vs.
Group C (P50.0011),
Group A vs. Group D
(P<0.0001), Group B vs.
Group D (P50.02),
Group A vs. Group B
(P50.04), Group A
vs. Group C
(P50.003), Group
A vs. Group D
(P50.0004),
Group B vs. Group
D(P50.05),
No statistic
significance.
Group H: nonoperative healthy side; Dunn test (pairwise comparison), Pvalues by Kruskal Wallis test was performed.
Ratio 5Experimental Side/Healthy Side 5% of healthy side.
TZOU ET AL.
|
5

normal status. Although the degree of injury was different between
groups C (50% damage) and D (75% damage), the results of both
groups were the worst (P<0.001) (Table 1).
3.2
|
Muscle weights
Biceps muscle weight was heaviest in the healthy control group, with
at least on average 0.36 60.04 g. In the experimental group, it showed
0.29 60.04 g (80% of healthy side) in group A; 0.2960.04 g (74%) in
group B; 0.28 60.03 g (73%) in group C; and 0.28 60.02 g (72%) in
group D. Biceps muscle weight following the transfers did not reach
the normal status in all experimental rats, with a significant difference
(P<0.001). Group D showed the lowest muscle weight (Table 1).
3.3
|
Muscle tetanus contraction force measurement
Biceps muscle tetanus contraction force decreased with the increased
injury of the ulnar nerve: group A 40.95 69.37 g (95% of healthy side),
group B 40.18 62.19 g (77%), group C 40.9268.51 g (75%), and
Group D 34.70 68.64 g (69%). All experimental groups showed statis-
tically significant differences when compared with the healthy side,
51.66 67.16 g (P<0.05), except Group A (P50.4). The Pvalues for
the ratio (%) showed statistically significant differences between groups
AandB(P50.0023), groups A and C (P50.001), groups A and D
(P<0.001), and groups B and D (P50.02) (Table 1). Group D was the
worst but similar to group C (P50.06) (Table 1).
3.4
|
Musculocutaneous nerve motor action potentials
The mean peak amplitude of the compound muscle action potentials
following individual donor stimulation on the experimental side showed
low amplitudes in groups C and D, with an average of 3.59 60.58 mV
(75%) and 3.62 60.67 mV (69%), respectively, but high amplitude in
group B scored 4.24 60.72 mV (81%), and group A 4.5360.64 mV
(95% of the healthy side). These results were significant in groups C
and D (P<0.001). Group B (P<0.05) also showed a statistically signi-
ficant difference, compared with the healthy control side (5.0560.69
mV) (Table 1).
When using Dunn’s statistical analysis with absolute values for
intergroup comparisons (Table 1, Column II) it showed no significant
difference. However, with ratio (% of the healthy side) for internal
control (operative: non-operative, Table 1, Column III), comparison
between Group D/Group A showed most significant difference, then
Group C/Group A, and then Group B/Group A.
3.5
|
Axon counts
Axon counts of the musculocutaneous nerve distal to the ulnar-
musculocutaneous coaptation site, the regenerative axons in the exper-
imental rats all showed small but more sprouting axons. Group D
showed the lowest number of regenerated axons. Similar to the previ-
ous studies, intergroup comparison demonstrated (1) Group B was sim-
ilar to Group A, showing no significant difference with non-operative
side, Group H; and (2)Group C and Group D were similar, significantly
lower axon counts than others.
Axon counts of the healthy ulnar nerve (Group H, Table 2) were
1849 6362 on average (normal value). Axon counts of the injured
ulnar nerve proximal of the cut surface were 1447 6579 (78%) in
group B, 1051 6367 in group C (57%), and 5676230 in group D
(31% of the healthy ulnar nerve). Axon counts of the musculocutane-
ous nerve distally to the ulnar-musculocutaneous coaptation site in
group A were 2149 6366, group B 1753 6110, group C 1431 6565,
and group D 1366 6305. The regenerative axons in all groups showed
TABLE 2 Axon counts of healthy and injured ulnar nerve as innervated musculocutaneous nerve
Groups
Axon counts of
injured Uln
Percentage of
control Uln
Axon counts
of MCn
Percentage of
control Uln
A: positive control ––2149 6366 116%
B: ulnar nerve - 25% Injury/MCn 1447 6579 78% 1753 6110 85%
C: ulnar nerve - 50% Injury/MCn 1051 6367 57% 1431 6565 77%
D: ulnar nerve - 75% Injury/MCn 567 6230 31% 1366 6305 74%
H: Control Ulnar Nerve (healthy) 1849 6362 100% 100%
p values
Group H (healthy) vs. A, B, C, D
(experimental)
x Group A vs. Group B
(P50.044),
Group A vs. Group C
(P50.005),
Group A vs. Group D
(P50.002),
Group B vs. Group D
(P50.02),
Group B vs Group H
(P50.05),
Group C vs. Group H
(P50.005),
Group D vs Group H
(P<0.0001)
Group B vs. Group D
(P50.049),
Group C vs. Group H
(P50.04), Group D vs. Group
H(P50.02)
6
|
TZOU ET AL.

smaller in axon diameter, but higher axon counts. Axon counts in the
musculocutaneous nerve were all higher than the donor nerve due to
axon sprouting. Statistically significant differences of axon counts were
seen in groups C and D when compared with the healthy ulnar nerve
group H: C/H, P50.005 and D/H, P<0.0001. Group D had the low-
est number of regenerated axons. There is no significant difference
between Group C and D (P50.81).
Statistically significant correlation coefficients were observed only
between musculocutaneous nerve axon counts and muscle weight in
group A (corr 52861; P50.028), and between musculocutaneous
nerve axon counts and muscle action potentials in group B
(corr 50.882; P50.048).
4
|
DISCUSSION
In BPI, nerve transfer is indicated in root avulsion injury within the
golden period, 5 months after injury (Chuang, 2006). Nerve transfer is
growing in importance and popularity. Oberlin’s method (Oberlin, Beal,
Leechavengvongs, Salon, Dauge, & Sarcy, 1994)of using one or two
fascicles of the ulnar or median nerve to the biceps motor branch of
the musculocutaneous nerve to achieve elbow flexion is a good exam-
ple, but some require second procedure (such as functioning free mus-
cle transplantation) to achieve satisfactory elbow flexion (Cho, Paulos,
de Resende, Kiyohara, Sorrenti, Wei,... Mattar, 2014; Chuang, 2008;
Nath, Lyons, & Bietz, 2006). Mackinnon’s“double fascicular transfer”
(Mackinnon & Novak, 2005) using ulnar and median nerve fascicles to
the biceps and brachialis motor branches for elbow flexion is an
enhanced procedure for elbow flexion.
Clinically, we have seen patients with plexus avulsion injury, two
roots (C5-6), or three roots (C5-6–7) or even four roots (C5-6–7-8)
injury with intact T1 all could show good hand functions. Those
patients were reconstructed by “double fascicular transfer”technique
for elbow flexion. Most of them achieved good elbow flexion (M4 mus-
cle strength) within 2-years follow-up. However, some obtain fair
results, M3, requiring long rehabilitation (even longer than 4 years). The
main reason for such differences is due to uncertain degree of injury of
the donor nerves.
An experimental study of rabbits (Tubiana, 1988) observes that an
average of 50% of normal muscle power is achieved when one-third
(33%) of ulnar nerve is used as a donor for end-to-side nerve transfer.
Gordon et al. (1993) demonstrate that 20% motor neurons of the
donor nerve are sufficient for muscle function because motor units can
enlarge up to about five times their original size. Studies in the litera-
ture show that 30–45% of axons of a donor nerve are sufficient for
reinnervation of muscle to gain good muscle function (Lutz, Chuang,
Chuang, Hsu, Ma, & Wei, 2000; Witoonchart, Leechavengvongs,
Uerpairojkit, Thuvasethakul, & Wongnopsuwan 2003) which we usually
interpret as M4 in muscle strength (Midha, 2004). In fact, interpretation
of the M4 muscle strength for an acceptable functional result is not all
the same with some range of differences. Schreiber, Byun, Khair, Rose-
nblatt, Lee, & Wolfe (2015), published an overview of axonal counts of
donor nerves and showed optimal axon counts for brachial plexus
nerve transfers to restore elbow flexion, including intercostal nerve,
pectoral nerve, spinal accessory nerve, branch of median, branch of
ulnar, and thoracodorsal nerve.
In our designed experimental rat study, all four test parameters
(grooming test, biceps muscle CMAPs, tetanus contraction force, and
muscle weight recovery) showed that no group reached normal status.
Regardless of the amount of injury to the ulnar nerve, all showed poor
results, significant difference with non-operative side. For intergroup
comparison, the results of the Group D (75% transected) were the
worst. Group C (50% transected) showed similarity with Group D with
poor results. But Group B (25% transected) showed mostly acceptable
results as Group A (intact ulnar nerve) with no significant differences.
There were two facts of weakness in our designed study. First, a
12-week regeneration time might be not enough for such muscle com-
parison. Second, the ulnar nerve we chosen as a motor donor in this
study, is actually a big and powerful nerve. When performing ulnar
nerve 50% transected (Group C, for exampling) and intraneural dissec-
tion for 1.5 cm in distance, its damage was actually higher than 50%
(Figure 2). Similarly, 25% transected ulnar nerve in the Group B, its
axons preserved for transfer was actually <75%. This is why we highly
expect that the Group B with 75% axons preserved will potentially
provide the similar effects as the Group A. Further studies for these
questions are warranted.
5
|
CONCLUSION
The results of using an injured motor nerve for transfer are directly
proportional to the size (axon counts) and damage of the donor nerve
sustained. The donor nerve should be healthy in order to provide opti-
mal result. If the donor nerve is a big nerve (like ulnar nerve) but
injured, at least 75% axon spared is still effective for transfer. In con-
trast, a small nerve (like intercostal nerve) once injured, although with
75% axon spared, would be considered a suboptimal donor nerve.
ACKNOWLEDGMENTS
The authors thank research assistants Ms. Pei-Ju Chen and Ms. Ruby
Yih-Ru Chang for their professional and continuous technical assis-
tance throughout this study; also Ms. Hsiao-Jung Tseng M.P.H. from
Biostatistical Center for Clinical Research, Chang Gung Memorial Hos-
pital, Taoyuan, Taiwan, for her statistical support and analysis of the
data, as Maria Prieto Barea for creating parts of the figures.
DISCLOSURES
The authors hereby declare that they have no conflict of interest in
any products used/tested in this study and have nothing to declare.
REFERENCES
Alnot JY (1987). Traumatic paralysis of the brachial plexus: Preoperative
problems and therapeutic indications. In J. K. Terzis (Eds.), Microre-
construction of nerve injuries (p 325). Philadelphia: WB Saunders.
Bertelli JA, Ghizoni MF. (2004). Reconstruction of C5 and C6 brachial
plexus avulsion injury by multiple nerve transfers: Spinal accessory to
TZOU ET AL.
|
7

suprascapular, ulnar fascicles to biceps branch, and triceps long or
lateral head branch to axillary nerve. Journal of Hand Surgery,29,
131–139.
Bertelli JA, Mira JC. (1993). Behavioral evaluating methods in the objec-
tive clinical assessment of motor function after experimental brachial
plexus reconstruction in the rat. Journal of Neuroscience Methods,46,
203–208.
Brown JM, Shah MN, Mackinnon SE. (2009). Distal nerve transfers: A
biology-based rationale. Neurosurgery Focus,26, E12.
Cho AB, Paulos RG, de Resende MR, Kiyohara LY, Sorrenti L, Wei TH,
BolligerNeto R, Mattar Junior R. (2014). Median nerve fascicle trans-
fer versus ulnar nerve fascicle transfer to the biceps motor branch in
C5-C6 and C5-C7 brachial plexus injuries: Nonrandomized prospec-
tive study of 23 consecutive patients. Microsurgery 34, 511–515.
Chuang DC. (2005). Nerve transfers in adult brachial plexus injuries: My
methods. Hand Clinics,21,71–82.
Chuang DC. (2006). Adult brachial plexus injuries. In S. J. Mathes and V.
R. Hentz (Eds.), Plastic surgery (Vol. 7, pp. 515–538). Philadelphia:
Saunders Elsevier.
Chuang DC. (2008). Nerve transfer with functioning free muscle trans-
plantation. Hand Clinics,24, 377–388.
Chuang DC, Hernon C. (2012). Minimum 4-year follow-up on contralat-
eral C7 nerve transfers for brachial plexus injuries. Journal of Hand
Surgery,37(A), 270–276.
Chuang DC, Lee GW, Hashem F, Wei FC. (1995). Restoration of
shoulder abduction by nerve transfer in avulsed brachial plexus
injury: Evaluation of 99 patients with various nerve transfers. Plastic
and Reconstruction Surgery,96, 122–128.
Gordon T, Yang JF, Ayer K, Stein RB, Tyreman N. (1993). Recovery
potential of muscle after partial denervation: A comparison between
rats and humans. Brain Research Bulletin,30, 477–482.
Inciong JG, Marrocco WC, Terzis JK. (2000). Efficacy of intervention
strategies in a brachial plexus global avulsion model in the rat. Plastic
Reconstruction Surgery,105, 2059–2071.
Lutz BS, Chuang DC, Chuang SS, Hsu JC, Ma SF, Wei FC. (2000). Nerve
transfer to the median nerve using parts of the ulnar and radial
nerves in the rabbit—Effects on motor recovery of the median nerve
and donor nerve morbidity. Journal of Hand Surgery,25, 329–335.
Mackinnon SE, Novak CB, Myckatyn TM, Tung TH. (2005). Results of
reinnervation of the biceps and brachialis muscles with a double fas-
cicular transfer for elbow flexion. Journal of Hand Surgery (American
Volume) 30, 978–985.
Midha R. (2004). Nerve transfers for severe brachial plexus injuries: A
review. Neurosurgery Focus,1516, E5.
Narakas A. (1978). Surgical treatment of traction injuries of the brachial
plexus. Clinical Orthopaedics Related Research,133,71–90.
Nath RK, Lyons AB, Bietz G. (2006). Physiological and clinical advantages
of median nerve fascicle transfer to the musculocutaneous nerve fol-
lowing brachial plexus root avulsion injury. Journal of Neurosurgery,
105, 830–834.
Oberlin C, Beal D, Leechavengvongs S, Salon A, Dauge MC, Sarcy JJ.
(1994). Nerve transfer to biceps muscle using a part of ulnar nerve
for C5-C6 avulsion of the brachial plexus: Anatomical study and
report of four cases. Journal of Hand Surgery (American Volume),19,
232–237.
Rodriguez A, Chuang DCC, Chen KT, Chen RF, Lyu RK, Ko YS. (2011).
Comparative study of single-, double- and triple-nerve transfer to a
common target: Experimental study of rat brachial plexus. Plastic and
Reconstruction Surgery,127, 1155–1162.
Schreiber JJ, Byun DJ, Khair MM, Rosenblatt L, Lee SK, Wolfe SW.
(2015). Optimal axon counts for brachial plexus nerve transfers to
restore elbow flexion. Plastic and Reconstruction Surgery,135,
135e–141.
Shibata M, Breidenbach WC, Ogden L, Firrell J. (1991). Comparison of
one- and two-stage nerve grafting of the rabbit median nerve. Jour-
nal of Hand Surgery,16(A), 262–268.
Terzis JK, Sweet RC, Dykes RW, Williams HB. (1978). Recovery of func-
tion in free muscle transplants using microneurovascular anastomo-
ses. Journal of Hand Surgery,3(A), 37–59.
Totosy de Zepetnek JE, Zung HV, Erdebil S, Gordon T. (1992). Innerva-
tion ratio is an important determinant of force in normal and reinner-
vated rat tibialis anterior muscles. Journal of Neurophysiology,67,
1385–1403.
Tubiana R. (1988). Clinical examination and functional assessment of the
upper limb after peripheral nerve lesion. In R. Tubiana (Ed.), The hand
(pp. 455). Philadelphia: WB Saunders.
Tzou CH, Chuang DC, Chang TN, Lu JC. (2014). The impact of different
degrees of injured C7 nerve transfer: An experimental rat study. Plas-
tic Reconstruction Surgery Global Open,2, e230.
Witoonchart K, Leechavengvongs S, Uerpairojkit C, Thuvasethakul P,
Wongnopsuwan V. (2003). Nerve transfer to deltoid muscle using
the nerve to the long head of the triceps, Part I: An anatomic feasi-
bility study. J Hand Surg Am,28, 628–632.
How to cite this article: Tzou-CH, Lu C-YJ, Chang TN-J, and
Chuang DC-C. Can an injured nerve be used as a donor nerve
for distal nerve transfer?—An experimental study in rats. Micro-
surgery. 2016;00:1–8. doi:10.1002/micr.30153.
8
|
TZOU ET AL.