Keywords
Shoulder anomalies following brachial plexus birth palsy - Shoulder internal rotation
contracture - Glenohumeral dysplasia - Erb’s palsy
Introduction
Shoulder abnormalities are the most common long-term complication and the major cause
of morbidity in upper trunk brachial plexus birth palsy (BPBP) [[1]]. Shoulder functional impairment is due to a progressive development of muscle abnormalities,
with shoulder internal rotation contracture leading to major joint deformities (i.e.
glenohumeral dysplasia) [[2],[3],[4]]. Shoulder joint deformity has been extensively studied while the pathogenesis of
muscular changes remains barely understood [[5],[6],[7]]. An animal model of shoulder disorders after BPBP would permit the assessment of
these deformities, help define the muscle changes and would also provide a base for
treatment research. Previous murine models only focused on joint changes after upper
brachial plexus trunk neurectomy and came up with equivocal conclusions [[8],[9],[10]]. The purpose of this study is to use a similar rat model, combined with MRI evaluation,
to better delineate muscle and joint shoulder changes after BPBP. We would also discuss
its usefulness as a preclinical model.
Methods
This study was carried out following the National Institutes of Health Guidelines
for the use of laboratory animals and with the approval of the local Ethics Committee
for experimental animal use. Forty three rat newborns from four pregnant Sprague-Dowley
OFA rats were used in this study. Four rat pups died during or after surgery, and
were removed from the study.
Rat newborn surgery
5-day-old rat pups underwent right brachial plexus surgery under general anaesthesia
with isofluorane. A surgical microscope was used for dissection. A transverse incision
below the clavicle with splitting of pectoralis major and pectoralis minor muscles
was done to expose the brachial plexus. By locating the suprascapular nerve, we reached
the upper trunk of the brachial plexus. A segmental excision of the upper trunk using
microscissors was then performed to simulate upper trunk neurtometic injury.
Clinical evaluation
Glenohumeral passive external rotation range of motion was evaluated immediately after
sacrifice and right before MRI evaluation. The neutral position of the shoulder was
defined as the shoulder in 0° abduction and the elbow in 90° of flexion with the front
limbs up ventrally, perpendicular to the examination table. After scapular stabilization
with the thumb, external glenohumeral rotation was measured using a goniometer. The
left side was also measured and used as a control.
MRI evaluation
All animals were sacrificed using sodium pentobarbital injected intraperitoneally
after sedation. The animal was then placed into a 7.2T MRI Biospect system (Bruker,
Germany) in supine position with the front limbs onto its abdomen. Both shoulders
were independently evaluated using RARE 1mm axial TR 4000 T 30 sequence in axial and
sagital oblique planes parallel to the scapula. Each shoulder was analyzed independently.
MRI was obtained at different times after surgery: 10 rats after 3 weeks, 10 rats
after 4 weeks, 7 at 8 weeks, 6 at 15 weeks and 6 at 20 weeks. MRI analysis included
mesurement of the glenoid version, PHHA (percentage of the humeral head anterior to
the middle of the glenoid fossa), humeral head section area and infraspinatus and
subscapular muscle thickness. As a measurement tool, we used Osirix (Apple). All mesurements
were done by the same examiner.
Statistical analysis
In order to improve statistical power, rats were grouped into two groups. The early
group (n=20) included the rats sacrificed and imaged less than 4 weeks after surgery
while the late group (n=19) was formed with the remaning rats (all sacrificed and
studies at least 8 weeks after surgery). We relied on SPSS 15.0 to compare data. We
used Paired T-test to compare the involved and uninvolved shoulders. Non parametric
tests (Wilcoxon test and Mann–Whitney) were employed to compare between groups when
necesary.
Results
The results are reproduced in [Table 1]. All animals showed right shoulder and elbow flexion paralysis after surgery, with
an adduction and internal rotation posturing of the shoulder and lack of elbow flexion
([Figure 1]). There was a statistically significant decrease of passive external rotation in
the involved shoulder compared to uninvolved shoulder (23.85°±27.5 vs 79.62°±3.8 respectively)
(P<0,01) ([Figure 2]). When comparing early vs late group, no significant differences were found in mean
passive shoulder external rotation (p=0.08).
Table 1
Shoulder rotation and MRI evaluation
|
Shoulder Passive external rotation (degrees)
|
Glenoid angle (degrees)
|
PHHA (%)
|
Humeral head sectional area (cm)
|
Subescapular thickness (cm)
|
Infraespinatus thickness (cm)
|
|
Involved shoulder m(SD)
|
23.85
|
91.41
|
50.85
|
0.15
|
0.12
|
0.13(cm)
|
|
(27.5)
|
(6.0)
|
(6.7)
|
(0.05)
|
(0.03)
|
(0.03)
|
|
Healthy shoulder
|
79.62
|
87.33
|
48.69
|
0.19
|
0.25
|
0.26
|
|
(3.8)
|
(4.4)
|
(6.17)
|
(0.06)
|
(0.08)
|
(0.08)
|
|
p
|
0.000
|
0.000
|
0.058
|
0.000
|
0.000
|
0.000
|
Figure 1 10-days-old rat (five days after surgery) showing shoulder paralysis and
lack of elbow flexion after surgery.
Figure 2 Axial vision of a rat showing a decrease of right passive shoulder external
rotation 8 weeks after surgery.
Mean glenoid version in the involved shoulder was 91.41°±6.0 and 87.33°±4.4 in the
uninvolved. There was an statistically significant increase in the glenoid version
in the involved shoulder, with an average increase of 4,08° (IC95% 2,31;5,83) (P<0,01).
However, separate comparison between groups, showed a significative increase of glenoid
anteversion in the early group, reaching 6,6° (IC95% 4.5° ; 8.6°) (P<0,01), but not
in late group, 1,4° (IC95% -1.1°;3.9°) (P=0,4). Significant differences were found
when comparing the increase of glenoid anteversion of both groups (P<0,01) ([Figure 3]). No statistically significant differences were found in PHHA between involved and
healthy shoulder ( 50,85 ± 6.7% vs 48,69 ± 6.1% ) (P=0,1) ([Figure 3]).
Figure 3 Shoulder MRI 8 weeks after surgery showing symmetrical measures of glenohumeral
joint alignment: glenoid version and PHHA. This rat model does not reproduce the joint luxation found in children.
Involved humeral head sectional area was significantly smaller than the healthy one
with a mean size decrease of 19,9% (IC95% 16.9;23.3) (p<0.01). When comparing early
and late groups, no significant differences were found in the decrease of humeral
sectional area (p=0.73) ([Figure 4]). Involved infraspinatus muscle thickness was significatively smaller than the healthy
side with a relative 46.3% (IC95 40.6;52) mean thickness decrease (P<0,01) ([Figure 5]). When comparing both groups, significantly more muscle thickness loss was found
in the late group (56.9% (IC95% 50.6;63.3) vs. early group 36.21% (IC95 29.17;43.25)
( P<0,01)). Involved subscapular muscle thickness was significatively smaller than
healthy one with a relative 52.5% (IC95 49.4;55.8) mean thickness decrease (P<0,01)
([Figure 5]). When comparing early and late groups, no significant differences were found in
muscle thickness loss (p=0.02). When comparing loss of thickness between subscapular
and infraspinatus muscle no statistical significant differences were found (P=0,02).
However when comparing between early and late groups a statistically significative
diference was found in the early group with a more severe atrophy in subscapular muscle
(49.7% IC95% 44.6;54.7) than infraspinatus muscle (36.21% IC95% 29.17;43.25) p<0.01.
Those differences were not found in the late group (p=0.55). Shoulder MRI of older
rats showed severe deformity of the humeral head, the glenohumeral joint and the glenoid,
but with no shoulder dislocation being found whatsoever ([Figure 6]).
Figure 4 Shoulder MRI 8 weeks after surgery showing a decrease of humeral head sectional
area that reproduces the joint hypoplasia found in children.
Figure 5 Shoulder MRI 8 weeks after surgery showing a marked atrophy of both infraspinatus
and subscapular muscle that reproduces muscle changes found in children.
Figure 6 Shoulder MRI 16 weeks after surgery showing a loss of relationship between
the glenoid -which presents a normal version- and a marked deformed humeral head. An advanced right glenohumeral joint hypoplasia is present. The distinctive shoulder
dysplasia found in humans is absent.
Discussion
The present study compares and contrasts our experimental findings to previous publications
on shoulder structural changes after BPBP in animals and humans. Shoulder abnormalities
are the most common long-term complication and the major cause of morbidity in BPBP.
Nearly 30% of children with incomplete neuronal recovery go on into developing glenohumeral
dysplasia [[1],[3],[4],[11],[12],[13]]. An adequate preclinical model would be beneficial for the development of studies
regarding pathogenesis, prevention or even treatment of this disabling disorder. The
anatomy of the brachial plexus and its nerve distribution in rat is similar to that
of humans and injury of upper brachial plexus roots leads to a clinical pattern similar
to Erb´s palsy [[14],[15]]. Although the murine model has been advocated as an adequate preclinical model
of shoulder dysplasia following BPBP, our results lead us to disagree with this statement
[[8],[9],[10]]. Shoulder anomalies following BPBP are characterized by muscle atrophy and contracture,
as well as a progressive joint subluxation and joint hypoplasia [[6]]. Lack of shoulder external rotation and consequent joint deformities are progressive
and appear early on. Initial changes can occur as early as the third month of life,
while advanced deformities appear starting the second year [[2],[3]]. Our model could show an early development of shoulder internal rotation contracture
and joint changes. Natural history of shoulder joint and muscle changes after BPBP
are been best followed with imaging studies [[6],[12],[13]]. MRI is the most accurate image tool to assess the pediatric glenohumeral joint
due to its high percentage of cartilage. MRI would further allow a better standardization
of radiologic measurements and muscle assessment [[16],[17]]. This explains the choice of this imaging modality in our animal model. In fact,
MRI was highly reliable in characterizing the normal anatomy and pathological shoulders
changes in our rat model.
According to reported in MRI findings in children with BPBP, muscle atrophy is a common
denominator in all rotator cuff muscles, but most prominently affects the subscapularis
muscle [[3],[6],[7]]. The pathogenesis of the muscle changes that lead to the shoulder internal rotation
contracture are barely understood with a few reports focusing on this topic [[5],[6],[7],[18]]. The atrophy and contracture of the subscapularis muscle is thought to be the main
contributor to shoulder contracture [[19]]. A recent experimental report concludes that these subscapularis changes are caused
by muscle denervation-reinnervation. However recent clinical and radiological data
supports that muscle imbalance around the shoulder might also play a role [[19]]. In our study, we chose to properly assess eventful muscle changes through the
use of the MRI [[8],[9],[10]]. All our animals, similarly to the previously reported murine models, uniformly
developed an early and severe shoulder internal rotation contracture [[8],[9],[10]]. Moreover, our model showed a severe atrophy of both subscapular and infraspinatus
muscle. Infraspinatus muscle wasting seemed to appear more progressively than in the
subscapular muscle, in which severe atrophy appeared very early. It must be emphasized
that this phenomenon is similar to that encountered in children with shoulder dysplasia.
In these, incomplete reinnervation leads to external rotator muscles atrophy and to
an abnormal growth of the subscapularis muscle, further evolving into atrophy and
contracture [[20]]. Further experimental studies including mechanical and histological assessment
of periarticular muscles are recommended to define the exact pathogenesis of internal
rotation contracture [[5]]. Though our model reproduced the muscular changes and the shoulder contracture,
it failed to reproduce the typical glenoid deformity. Thus, the internal rotation
contracture might not be the only causative factor for shoulder subluxation in BPBP.
A possible explanation for these findings might be the quadripedal gait of the rat,
with the shoulder being in a different mechanical disposition.
The glenoid version has been used to quantify the degree of glenoid deformity and
the PHHA to quantify the degree of posterior humeral subluxation in children with
shoulder dysplasia, as the typical human joint glenohumeral deformity combines a glenoid
version change with humeral head subluxation [[2],[5]]. The degree of change of both parameters is mutually proportional since the deformity
occurs progressively and consequently. As a complete neurectomy of the upper trunk
was performed (i.e. neurotmesis), a severe glenohumeral dysplasia combining a severe
change in the glenoid version (or the development of a pseudoglenoid) and a severe
humeral head subluxation, might be expected. However, our rats showed only a slight
increase of glenoid anteversion, a finding that was simply present in the early group.
Similarly, none of the previous murine models had reported severe glenoid version
changes after a surgical neurotmesis of the upper brachial plexus trunk in the neonatal
age [[8],[9],[10]]. Li’s first study speaks of a 10° increase of the glenoid anteversion while a contradictory
10° retroversion was obtained in his second study [[8],[9]]. Kim’s mouse model also shows an 8.3° glenoid retroversion [[10]]. The discrepancy of the results between these studies might be due to the measurement
method or to the fact that no intraobsever reliability test was conducted. In our
study, the differences between early and late group might be due to these same reasons.
We think that the shoulder dislocations reported in the mentioned papers do not correspond
to a truly glenohumeral dysplasia but rather to an advanced degree of joint hypoplasia
since they were associated to a very mild glenoid version change. The advanced joint
hypoplasia might be related to the joint denervation that follows an upper trunk neurotmesis.
Shoulder joint growth anomalies in children with incompletely recovered BPBP manifest
by a delay in ossification nucleus growth and a decrease of humeral head size [[2],[4],[21]]. Abnormal mechanical transarticular loads due to muscle weakness and denervation
of the physes and joint might play a role in joint underdevelopment. The marked joint
hypoplasia that occurred in the oldest rats of our study, as well as those of the
previously mentioned studies, might be explained by the severe joint denervation that
follows the complete upper trunk neurectomy (i.e. neurotmesis). Both groups, (<4weeks
and >8weeks), showed a significant decrease of the humeral head transverse sectional
area but no statistical differences were found between them. However, the shoulder
joint of the older rats (20 weeks) were very deformed. Severe deformity of the humeral
head and loss of the articular space were present. These findings are similar to those
reported in Li’s and Kim’s papers for the older animals (16 weeks old rats or 30 week
old mice, respectively) [[8],[9],[10]]. All rats included in our study developed a marked shoulder contracture and subscapular
changes in consonance with the changes occurring in children with BPBP. In addition,
our model reproduced the joint hypoplastic changes found in human. However, our model
failed to reproduce the typical human joint deformity with proportional glenoid retroversion
and humeral head subluxation [[1],[2],[3],[4]] as, in our opinion, did fail the previous experimental studies. A better understanding
of the pathogenesis of shoulder internal rotation contracture after BPBP with a deep
insight to the role of subscapular muscle changes are necessary to develop future
strategies in the prevention and treatment of this disorder.
Conclusions
The rat model might be an adequate preclinical model of shoulder contracture following
BPBP but not of glenohumeral dysplasia. Studies regarding pathogenesis, prevention
or treatment of shoulder contracture could be performed using this model.
Competing interests
This work was funded by Instituto de Salud Carlos III, grant FIS PI10/01357 cofinanced
by the European Regional Development Fund (ERDF), Fundacio ? Privada A. Bosch and
by Fundação Santa-Maria-Silva. The authors declare that they have no conflict of interest.
Authors’ contributions
FS, BCD, KSH and HS conceived of the study, and participated in its design and coordination
and draft the manuscript. MM, MME and FCG participated in the design of the study
and performed the statistical analysis. IB and MVV carried out the radiological analysis.
All authors read and approved the final manuscript.
Cite this article as: Soldado et al.: Muscular and glenohumeral changes in the shoulder after brachial plexus birth palsy:
an MRI study in a rat model. Journal of Brachial Plexus and Peripheral Nerve Injury 2012 7:9.
