The American Journal of Sports

Medicine
http://ajs.sagepub.com/

Revascularization Process of the BonePatellar TendonBone Autograft Evaluated by

Contrast-Enhanced Magnetic Resonance Imaging 6 and 12 Months After Anterior Cruciate Ligament
Reconstruction
Aikaterini Ntoulia, Frederica Papadopoulou, Stavros Ristanis, Maria Argyropoulou and Anastasios D. Georgoulis
Am J Sports Med 2011 39: 1478 originally published online March 10, 2011
DOI: 10.1177/0363546511398039
The online version of this article can be found at:
http://ajs.sagepub.com/content/39/7/1478

Published by:
http://www.sagepublications.com

On behalf of:
American Orthopaedic Society for Sports Medicine

Additional services and information for The American Journal of Sports Medicine can be found at:
Email Alerts: http://ajs.sagepub.com/cgi/alerts
Subscriptions: http://ajs.sagepub.com/subscriptions
Reprints: http://www.sagepub.com/journalsReprints.nav
Permissions: http://www.sagepub.com/journalsPermissions.nav

>> Version of Record - Jul 5, 2011

OnlineFirst Version of Record - Mar 10, 2011
What is This?

Downloaded from ajs.sagepub.com at TEXAS SOUTHERN UNIVERSITY on October 28, 2014

Revascularization Process of the

BonePatellar TendonBone Autograft
Evaluated by Contrast-Enhanced Magnetic
Resonance Imaging 6 and 12 Months After
Anterior Cruciate Ligament Reconstruction
Aikaterini Ntoulia,* MD, Frederica Papadopoulou,* MD, Stavros Ristanis,y MD,
Maria Argyropoulou,* MD, and Anastasios D. Georgoulis,yz MD
Investigation performed at the Department of Radiology, University Hospital of Ioannina, and the
Orthopaedic Sports Medicine Center, Department of Orthopaedic Surgery, University of
Ioannina, Ioannina, Greece
Background: Contrast-enhanced magnetic resonance imaging (MRI) studies conducted on animal models have shown that the
observed signal intensity changes are related to the degree of graft vascularity and its biomechanical properties.
Purpose: To evaluate by contrast-enhanced MRI the revascularization process at 3 distinct sites discerned in relation to the surrounding microenvironment along the course of bonepatellar tendonbone (BPTB) autograft in uncomplicated human anterior
cruciate ligament (ACL)reconstructed knees.
Study Design: Case series; Level of evidence, 4.
Methods: Thirty-two male patients were assessed with a 3-dimensional fast field echo/short tau inversion recovery (FFE/STIR)
MRI sequence at the third postoperative day and at time intervals of 6 and 12 months after surgery. Signal-to-noise quotient
(SNQ) was calculated for 3 specific graft sites (intra-articular site, intraosseous tibial tunnel site, and intraosseous juxta screw
site) before and after gadolinium administration. Comparisons of the enhancement index (EI: SNQafter/SNQbefore gadolinium)
were performed independently for each graft site and time interval.
Results: Three days postoperatively, insufficient vascularization was noticed at the 3 sites. Six and 12 months after surgery, the
enhancement index was significantly increased in all 3 sites (P \ .001). The intra-articular site, 6 months postoperatively, achieved
satisfactory contrast medium uptake (enhancement index .1), with significantly higher enhancement index values compared with
the other 2 sites (P \ .001). Twelve months after surgery, only the intraosseously enclosed sites displayed an increase of the
enhancement index, although nonsignificant (P = .09 and P = .07, respectively).
Conclusion: Revascularization of the graft occurs gradually along its length, with the intra-articular site being the first and the
faster part to complete this phase, while both the intraosseous sites are still in progress throughout the first postoperative
year. Revascularization is an important link at the intrinsic healing chain of the ACL graft. The surrounding microenvironment
does seem to play a significant role in this process, and the differences in its composition along the graft course are reflected
at the revascularization progress of the corresponding sites.
Keywords: anterior cruciate ligament reconstruction; graft revascularization; contrast-enhanced MRI; BPTB; enhancement index

After anterior cruciate ligament (ACL) arthroscopic reconstruction, the tendon autograft used to replace the injured
ligament undergoes a biological healing process consisting
of 4 successive phases: initial necrosis, revascularization,
cellular repopulation, and remodeling.4,7,11,18,19,50 Revascularization contributes significantly to this process by
providing the nutrient elements to sustain graft viability
and by facilitating the subsequent remodeling phases to
be conducted.4,18,41,50 The uneventful transition of these
histomorphological alterations will eventually lead to graft
ligamentization.4,19,30,31,41,46 As a neoligament, the tendon

z
Address correspondence to Anastasios D. Georgoulis, MD, Orthopaedic Sports Medicine Center, PO Box 1042, Ioannina 45110, Greece
(e-mail: georgoulis@osmci.gr).
*Department of Radiology, University Hospital of Ioannina, Ioannina,
Greece.
y
Department of Orthopaedic Surgery, Orthopaedic Sports Medicine
Center, University of Ioannina, Ioannina, Greece.
The authors declared that they have no conflicts of interest in the
authorship and publication of this contribution.

The American Journal of Sports Medicine, Vol. 39, No. 7

DOI: 10.1177/0363546511398039
2011 The Author(s)

1478
Downloaded from ajs.sagepub.com at TEXAS SOUTHERN UNIVERSITY on October 28, 2014

Vol. 39, No. 7, 2011

Revascularization of BPTB Autograft Evaluated by Contrast-Enhanced MRI

autograft will achieve its incorporation to the new environment and therefore its functional adaptation to its new
role.4,9,18,31,56 The earlier this process is completed, the
sooner the ACL substitute graft will meet its new loading
demands, allowing the patient to resume earlier his or
her sport activities.4,19,25
In this healing process, the microenvironment surrounding the graft plays an important role because it provides the necessary elements for its healing.4,46
According to the composition of this environment, 3
distinct sites can be discerned along the graft course:
(1) the intra-articular site, (2) the site of the graft that
is located inside the osseous tunnels, and (3) the site of
the graft that is adjacent to the fixation material inside
the osseous tunnels. The first graft site interacts circumferentially with the surrounding synovial environment.
The site that is entirely enclosed in the osseous tunnels
interacts extensively with the cancellous bone, while the
graft site that is located between the fixation material
and the osseous tunnel wall interacts partially with
both of them. Given the prominent differences in the composition of these surrounding spaces, questions can be
raised concerning whether each of the aforementioned
graft sites could be differently affected in terms of
revascularization due to different nutrient elements
supply.
Several histological studies have been conducted in
dissimilar in vivo models documenting the revascularization process of the graft with variable results. On the
scope of imaging, MRI with the use of gadoliniumdiethylenetriamine pentacetic acid (Gd-DTPA) contrast
agent has been used as a valuable tool to provide noninvasively important information regarding the vascular status of the graft tissue. It is well established that
enhancement of a tissue after contrast agent administration results in increase of its signal intensity.8,27,36,37,51,52
Weiler et al,52 by means of contrast-enhanced MRI in
a sheep model, demonstrated that the signal intensity
changes observed in the graft tissue reflect its vascularity
and biomechanical properties. Additionally, evaluation of
quantitative imaging parameters, such as signal-to-noise
quotient (SNQ), was correlated with the degree of vascularity as it was histologically confirmed. However, in
this animal-based study, only the intra-articular part of
the graft was examined.
To our knowledge, no previous studies have been conducted on human patients to compare noninvasively the
degree of vascularity in the aforementioned distinct graft
sites that can be discerned along its course in relation to
the surrounding microenvironment. Therefore, the aim of
the present study was to quantitatively evaluate, by means
of contrast-enhanced MRI, the signal intensity changes of
the graft over time that correspond to its revascularization.
We hypothesized that the differences in the surrounding
microenvironment will be reflected in the vascularization
process.

References 4, 7, 11, 18, 19, 30, 31, 41, 46, 50, 56.

1479

MATERIALS AND METHODS

Patients
From June 2007 until September 2009, 215 patients underwent arthroscopically assisted ACL reconstruction by the
same orthopaedic surgeon (A.D.G.). In total, 144 bone
patellar tendonbone (BPTB) autografts were used in 137
male and 7 female patients, while 71 quadrupled hamstring tendon (semitendinosus/gracilis) autografts were
used in 56 male and 15 female patients.
Given the objective of the study, examination of a single,
although multiparametric, variable, as the vasculature is,
requires the establishment of conscientious and explicit
patient selection criteria to maintain the homogeneity of
the population sample and ensure the validity of the
results. Therefore, for the present study, we recruited
male patients who had undergone ACL reconstruction
with a BPTB autograft. Women were excluded because
hormonal differences have been related with modified vasculature responsiveness during revascularization.10,26,55
Patients with multiligament injuries, revision operation
of the ACL ligament, as well as serious chondral lesions
(Outerbridge classification III or IV), were also excluded
from this study because the severity of the concomitant
injuries may negatively affect the overall final outcome.
Moreover, smokers15,32 and overweight22,48 patients were
also excluded, given that their associated morbidities are
related to endothelial dysfunction that impairs vessels
microcirculation. Furthermore, patients with serious allergic reaction to pharmaceutical (previous administration of
contrast agent or other medication), environmental (severe
reactions to insects venom), or dietary allergens and
required medical treatment were also excluded from this
serial study for safety reasons.2,16 Finally, because of socioeconomic reasons, patients living more than 1 hour away
from the hospital where the present study was conducted
were also excluded.
According to the above criteria, 32 patients were finally
included in the study. The mean age of the selected
patients was 25.6 years (range, 22-32 years), and mean
body mass index (BMI) was 23.7. Informed consent was
obtained from all patients enrolled in this study, in accordance with the institutional review board policies of our
University Hospital.

Surgical Reconstruction With a BPTB Graft

The BPTB graft was taken from the medial third of the
patellar tendon.20,34 We did not harvest more than a third
of the total tendon width. The diameter of the harvested
graft was 9 to 10 mm. First, the tibial tunnel was drilled.
The hole in the tibial plateau was placed approximately
5 mm anterior and medial to the anatomic center of the
natural ACL attachment, so that the posterolateral part
of the tunnel circumference was located at the ACL attachment anatomic center.12 The drilling of the femoral tunnel
was performed arthroscopically through the anteromedial
approach, with the knee joint in 120 of flexion and at

Downloaded from ajs.sagepub.com at TEXAS SOUTHERN UNIVERSITY on October 28, 2014

1480 Ntoulia et al

The American Journal of Sports Medicine

Figure 1. Tibial (A) and femoral (B) tunnel placement in a left knee.The tibial tunnel was drilled in the center of the ACL footprint.
The femoral tunnel was drilled at the 10-oclock position of the left knee (or the 2-oclock position corresponding to the right knee)
according to the classification system of the scientific committee of the European Society of Sports Traumatology, Knee Surgery
and Arthroscopy.
about the 10-oclock position (for a right knee) or at the 2oclock position (for a left knee).
The placement of the graft in the femoral tunnel was
performed with the cortical side of the bone plug close to
the over-the-top position. In the tibia, we twisted the graft
90, so as to recreate the anatomic spiral of ACL fibers.20,34
Fixation of the graft was performed in all patients with the
same bioabsorbable interference screws, consisting of polylactic acid (PLA) and hydroxylapatite (HA) (BioRCI-HA,
Smith & Nephew Endoscopy, Andover, Massachusetts),
on both femur and tibia (size, 7 3 20 mm and 9 3 20 mm,
respectively). Interference screws were inserted at the cancellous side of the bone plugs. After the fixation to the
femur, maximal tension was performed manually by pulling
the graft from its tibial edge and pushing the tibia posteriorly. Holding the knee in 30 of flexion23 and holding the
graft tensioned as we described, we proceeded to the fixation
on the tibia with the second interference screw (Figure 1).
Finally, the graft was inspected both in full flexion and
full extension so as to exclude graft impingement at the
intercondylar roof, the lateral condyle, and the posterior
cruciate ligament. A notchplasty was not performed in any
of our cases.

Clinical Evaluation
All ACL-reconstructed patients followed the same carefully
controlled rehabilitation protocol. They were allowed to
return to sports-related activities at 6 months after surgery, provided that they had regained full functional

strength and stability. Strength at that time was determined with the BIODEX (System-3, Biodex Corp, Shirley,
New York) isokinetic dynamometer, revealing acceptable
symmetry in quadriceps and hamstring strength.45 Clinical examination was performed by the same experienced
orthopaedic surgeon at the same day that the MRI examination was performed and included several clinical tests
for objective and subjective evaluation of knee joint stability. Specifically, the subjective examination included the
Lachman, anterior drawer, and pivot-shift tests, as well
as International Knee Documentation Committee (IKDC),
Tegner, and Lysholm activity scores. In addition, anterior
tibial translation was evaluated using the KT-1000 knee
arthrometer (MEDmetric Corp, San Diego, California).49
The measurements were performed with the application
at the tibia of 134-N posterior-anterior external force, as
well as maximum manual posterior-anterior external force
until heel clearance.

MRI Evaluation
Each patient underwent static MRI of the knee joint at 3
time intervals: third postoperative day and 6 and 12
months after surgery. The examination at the third postoperative day was performed after the removal of the drainage tubes. Clinical examination and routine laboratory
profile of the patients, including total white blood cell
(WBC) count, C-reactive protein, erythrocyte sedimentation rate (ESR), were performed, and results were normal
with no evidence of surgery-related infection. All the

Downloaded from ajs.sagepub.com at TEXAS SOUTHERN UNIVERSITY on October 28, 2014

Vol. 39, No. 7, 2011

Revascularization of BPTB Autograft Evaluated by Contrast-Enhanced MRI

1481

TABLE 1
MR Imaging Protocol Series of Different Sequences Performed in Orthogonal Planes to Obtain an Overview of
All Knee Joint Structures and to Evaluate Graft Integritya
Parameters Sequence
PDW/TSE/SPIR
PDW/TSE
STIR/long TE
PDW/TSE
T2W/TSE/SPIR
T1W/TSE
3D/FFE/SPIR

Plane

TR (ms)

TE

FA

NSA

Thickness/Gap (mm)

FOV/RFOV

Axial
Coronal
Coronal
Sagittal
Sagittal
Oblique axial
Oblique sagittal

3455
3092
1798
2711
2600
500
32

15
16
55
16
70
17
5.1

90
90
90
90
90
90
25

2
2
3
2
2
3
2

4.00/0.4
3.3/0.33
3.3/0.33
3.3/0.33
3.3/0.33
3.0/0.3
2.0

180/80
180/90
180/90
180/90
180/90
180/90
180/80

PDW, proton density weighted; TSE, turbo spin echo; SPIR, spectral presaturation by inversion recovery; STIR, short tau inversion
recovery; TE, echo time; T2W, T2 weighted; T1W, T1 weighted; 3D/FFE, 3-dimensional fast field echo; TR, repetition time; FA, flip angle;
NSA, number of sample averages; FOV, field of view; RFOV, rectangular field of view.

examinations were performed with a 1.5-T imager (Gyroscan ACS NT, Philips, Medical System Best, Best, the Netherlands) and a circular receiver-transmission extremity coil.
The knee was positioned in full extension, was comfortable
to the patient, and was immobilized with foam padding. An
infusion line was placed in the patients antecubital vein
before the examination, so that the patient could remain
totally immobile during the examination. The unenhanced
examination protocol included sequences in all 3 planes to
obtain an overview of the graft and joint structures. The
implementation of a 3-dimensional spoiled gradient echo
sequence with fat suppression (3-D FFE/STIR) in an oblique
sagittal plane along the long axis of the graft, before and 1
minute after gadolinium (Gd-DTPA) contrast administration (0.1 mmol/kg body weight), was used for the analysis
of signal intensity, which was performed with the computer
software ImageJ (version 1.43, National Institutes of
Health, Bethesda, Maryland). The detailed parameters of
the MRI protocol are outlined in Table 1.

Data Analysis
Two radiologists with experience in musculoskeletal MRI
evaluated blindly all MRI examinations, with disagreements resolved by consensus. The image that showed the
full extent of the graft from its femoral tunnel exit through
the joint space up to the tibial tunnel exit was selected for
the analysis. Three sites of the graft were distinguished
according to the surrounding microenvironment, and
they were evaluated independently: (1) the intra-articular
part of the graft located centrally within the joint (intraarticular site), (2) the intraosseous part of the graft
surrounded by the osseous tibial tunnel wall (intraosseous
tibial tunnel site), and finally (3) the part of the graft
located at the interface between the osseous tibial tunnel
wall and the biodegradable interference screw (intraosseous juxta screw site) (Figure 2).
The signal intensity (SI) was calculated at the 3 aforementioned graft sites using an 8-mm2 elliptical region of interest
(ROI). The SNQ was then calculated at each of the 3 sites by
dividing the SI of the graft by the noise of the image background on the images obtained before contrast agent

administration29,43,52: SNQbefore = SIspecific site of the graft O

noise background (Figure 3). Similarly, on the images
obtained after the administration of gadolinium, ROIs were
placed at identical positions by the computer software, and
the measurements were repeated exactly the same way:
SNQafter =SIspecific site of the graft O noise background (Figure
4). Noise was defined as the standard deviation of the signal
in a 250-mm2 ROI placed outside the patient within a 1-cm
distance from the tibial tubercle, at the lower part of the
image29,42 (Figures 3 and 4).
Each measurement was repeated 3 times for the intraobserver variation analysis as well as the calculation of
the mean values. By dividing the SNQ mean values after
and before contrast administration, a new ratio was
derived to provide direct information about revascularization. This ratio was named the enhancement index (EI):
EI = SNQafter O SNQbefore. Calculation of the enhancement
index was made independently for each graft site and time
interval. Enhancement index values above 1 were considered as satisfactory revascularization of any graft site,
while values equal or less than 1 indicated insufficient
revascularization (Table 2).
Comparison of the enhancement index values was performed using analysis of variance (ANOVA), and P values
\.05 were considered significant. Tukey post hoc tests
were used for comparison between mean values. Power
analysis was conducted for every comparison we made.
Effect sizes .0.8 were considered large according to
Cohen.13 Finally, k correlation coefficient with 95% confidence interval (CI) was used to measure the overall intraobserver and interobserver agreement regarding the ROI
placement. The agreement was classified as poor (k \ 0.4),
moderate (k = 0.4-0.8), or excellent (k  0.8). The statistical
analysis was performed with the Statistica v.8 program
(StatSoft, Tulsa, Oklahoma).

RESULTS
Clinical Evaluation
All ACL-reconstructed patients were satisfied with the outcome of the surgery and resumed their preinjury level of

Downloaded from ajs.sagepub.com at TEXAS SOUTHERN UNIVERSITY on October 28, 2014

1482 Ntoulia et al

The American Journal of Sports Medicine

Figure 2. Graft sites. Three sites can be discerned along the

graft course according to the composition of the surrounding
microenvironment: the intra-articular site, the intraosseous
tibial tunnel site, and the intraosseous juxta screw site.
sports participation. Negative Lachman, anterior drawer,
and pivot-shift tests indicated that the knee joint stability
was regained clinically for all the patients. For the BPTB
graftreconstructed patients at 6 months postoperative
time interval, the median Lysholm score was 93 (range,
83-100) and the Tegner score was 6 (range, 4-8) at the
time of examination, while for patients at 12 months after
surgery, the median Lysholm score was 98 (range, 93-100)
and the Tegner score was 7 (range, 6-8). The KT-1000
arthrometer results revealed that the mean difference
between the anterior tibial translation of the reconstructed
and intact sides at 6 months was 1.5 mm (range, 0-3 mm)
for the 134-N test and 1.9 mm (range, 1-3 mm) for the maximum manual test, respectively. The KT-1000 arthrometer
results at 12 months were 1.2 mm (range, 0-2 mm) for the
134-N test and 1.5 mm (range, 1-3 mm) for the maximum
manual test.

MRI Evaluation
Calculation of the enhancement index at the 3 graft sites
on the third postoperative day MRI examination showed
mean values of 0.95 6 0.03, 0.93 6 0.02, and 0.90 6 0.02
for the intra-articular, intraosseous tibial tunnel, and
intraosseous juxta screw sites, respectively. According to
the biomedical meaning of the enhancement index, no significant vascularization can be assumed at this time. However, 6 months after surgery, the enhancement index mean
values were 1.52 6 0.14, 1.36 6 0.16, and 1.29 6 0.15,
respectively. All these values were found to be significantly
increased above the threshold value of 1, demonstrating

Figure 3. Before contrast agent administration. A sagittal

section through the reconstructed knee obtained before
intravenous administration of gadolinium. Three regions of
interest (ROIs), seen as numbered circles, were assigned at
the midportion of every graft site (ROI 1, 2, and 3). Calculation of the SNQ values was performed independently for
each graft site and every time interval. The noise of the image
background was measured with the assignment of a ROI (4)
at the lower part of the image and within a distance of 1 cm
from the tibial tubercle.
satisfactory revascularization. Similarly, 12 months after
surgery, the enhancement index mean values were 1.53 6
0.15, 1.45 6 0.13, and 1.38 6 0.15, respectively, indicating
also satisfactory revascularization. Post hoc analysis with
Tukey tests revealed significant differences in the enhancement index values of the 3 graft sites at the 3 time intervals
on respective MRI examinations.
Intra-articular Site. Three days after surgery, the intraarticular site of the graft demonstrated insufficient
enhancement index (0.95 6 0.03). However, 6 months after
surgery, this site of the graft achieved satisfactory contrast
medium uptake, displaying at the same time significantly
higher enhancement index values compared with the other
2 sites (P \ .001). Nevertheless, by 12 months, no further
significant increase of the enhancement index was noted
compared to the 6 months (P = .1).
Intraosseous Tibial Tunnel Site. Similarly, 3 days after
surgery, the graft site that was enclosed inside the tibial
tunnel displayed enhancement index values inferior to
the threshold value (0.93 6 0.02). On the other hand, 6
and 12 months after surgery, the enhancement index
was clearly increased as judged by the effect size,
although this change was not statistically significant (P
= .09). This indicates that revascularization was still in
progress.

Downloaded from ajs.sagepub.com at TEXAS SOUTHERN UNIVERSITY on October 28, 2014

Vol. 39, No. 7, 2011

Revascularization of BPTB Autograft Evaluated by Contrast-Enhanced MRI

1483

TABLE 2
Enhancement Index Numerical and Biomedical Definitiona
Enhancement Index EI

SNQafter
SNQbefore

SNQafter . SNQbefore, EI . 1
SNQafter  SNQbefore, EI  1

Satisfactory revascularization
Insufficient revascularization

a
Enhancement index ratio provides directly interpretative
information regarding the presence, or not, of revascularization.
Uptake of the contrast medium will result in the SNQ values
obtained after its administration being greater than before, and
therefore, their resultant ratio will be greater or at least equal
to the numeric value of 1. This signifies satisfactory revascularization for the specific graft site. On the contrary, enhancement index
values lower than 1 mean insufficient revascularization. Therefore, 1 is the threshold value for demonstrating revascularization.

Figure 4. After contrast agent administration. The same sagittal section is depicted after intravenous administration of
gadolinium. Uptake of the contrast medium can be seen as
increased signal intensity within the vessels of the popliteal
region. Identical ROIs were assigned at the exact same positions of the 3 graft sites by the software program, and calculations of the SNQ values were similarly repeated.
Intraosseous Juxta Screw Site. Finally, at the third
postoperative day, the intraosseous juxta screw site
exhibited enhancement index values lower than 1 as the
other 2 sites did. Six and 12 months after surgery, increase
of the enhancement index values was noted, although nonsignificant as well (P = .07).
Moreover, comparison of the enhancement index values
between the 2 intraosseous graft sites showed that the site
that was entirely surrounded by the cancellous bone achieved
higher values than the juxta screw site during the whole
observation period. This finding was supported by the large
effect sizes, although significant differences were also not
detected. The above findings are summarized in Figure 5.
Power analysis revealed that all effect sizes of our comparisons were .0.8. This enabled us to confidently regard
as sufficient the population sample for detecting correctly
the true experimental outcome (lack or presence of differences) between graft sites at every time interval. The
intraobserver and interobserver agreement, as assessed
by the k coefficient, was excellent in every case (0.92: CI,
0.9-0.94 and 0.9: CI, 0.89-0.91, respectively).

DISCUSSION
The successful outcome of ACL reconstruction requires the
incorporation of the tendon autograft into its new environment.4,30,56 This process will take the intrinsic metamorphosis

of a tendinous tissue into a neoligament with the uneventful

transition through 4 distinct phases: avascular necrosis, revascularization, cellular repopulation, and finally remodeling and
reorganization of the collagen fibers.|| The phase at which the
graft tissue recovers its vascular supply plays the key role in
this process by acting as a prerequisite for the other phases
to be conducted and ensuring the long-term viability of the
graft. Therefore, it is important to elucidate how this phase
proceeds over time along the graft course.
The present study is an MRI-based quantitative analysis of the enhancement that the BPTB autograft achieves
after contrast medium administration corresponding to
its vascularity at the 3 specific sites examined along its
length. These 3 sites are namely the intra-articular site
located centrally within the joint, the intraosseous site
enclosed in the osseous tibial tunnel, and the site located
at the interface between the osseous tibial tunnel wall
and the fixation material. Our hypothesis was that because
the microenvironment of the graft is different at these 3
sites, the graft tissue would also be differently affected
during its revascularization process.
Our results show that, initially, the absence of functioning vasculature due to necrosis after graft harvesting
results in absence of contrast agent uptake and therefore
insufficient enhancement of the graft tissue with values
of the corresponding index below 1 in all cases. This imaging finding is in agreement with the results from previous
histological studies that reported extensive ischemic necrosis taking place throughout the graft substance initially
after its harvesting.28,47
Six and 12 months after surgery, however, satisfactory
enhancement was noticed in all graft sites, indicating adequate perfusion of the harvested tissue. Interestingly, the
intra-articular site was found to be the first graft site
that reached maximum enhancement index values at 6
months without any significant alteration noted at 12
months, indicating the completion of the revascularization
process for this site by this time period. On the other hand,
the site of the graft that was entirely enclosed by the
||

References 4, 7, 11, 18, 19, 28, 30, 31, 41, 46, 50.

Downloaded from ajs.sagepub.com at TEXAS SOUTHERN UNIVERSITY on October 28, 2014

1484 Ntoulia et al

The American Journal of Sports Medicine

Figure 5. Changes over time of the enhancement index

mean values at the 3 examined graft sites. The intra-articular
site is the first to complete this phase, while for both the other
2 intraosseously enclosed sites, the revascularization process is still in progress throughout the first postoperative
year.

osseous tibial tunnel wall, as well as the site that partially

interacted with the interference screw, showed a slower
progression of revascularization process. Both these sites
at 6 months, despite the fact that they exhibited satisfactory enhancement, had indices that were significantly inferior compared with the intra-articular site (P \ .001,
respectively). By 12 months, the increase of the enhancement index was clearly noted in both these sites, suggesting that graft revascularization was still in progress
during all this period. However, mean enhancement index
values were inferior to those of the intra-articular site,
although no significant differences were observed at that
time. Nonsignificant differences as well were found when
mean enhancement index values of the intraosseous tibial
tunnel and intraosseous juxta screw sites between 6 and 12
months time interval were compared (P = .07 and .09,
respectively), although the increasing trend of their mean
values during this time period is supported by large effect
sizes (.0.8), as a power analysis revealed. The abovementioned differences in the enhancement index values
reflect the differences in the revascularization progress
at the different graft sites, supporting our initial hypothesis that the microenvironment conditions existing around
the graft would affect its revascularization process.
The rapid completion of revascularization at the intraarticular site could be explained by the fact that its free
surface facilitates the provision of the nutrient elements
directly from synovial fluid in the initial postoperative
period.3,4,14,17,21,37,54 Several studies have established the
role of diffusion as an important instantaneous pathway
between interstitial synovial fluid and synovial lining neovessels for the delivery of the nutrients.14,54 Later, microvessels and neovessels originating from the infrapatellar
fat pad and the posterior synovial tissues embrace circumferentially its surface to form a rich vascular envelop that

contributes with numerous intraligamentous branches to

perfusion of this graft site.2,3,5,19,41,44,47 On the other
hand, the slower revascularization progress of both intraosseous graft sites could be explained by the fact that
nutrient elements are provided mainly by endosteal vessel
branches developing from the adjacent cancellous bone.6,28
Specifically, the graft site entirely located in the osseous
tunnel and which interacted perimetrically with the cancellous bone demonstrated higher enhancement index values during the whole examination period, compared with
the juxta screw site, which interacted partially with the
cancellous bone. And despite the fact that we did not notice
statistical significant differences in this case, these findings seem to have remarkable clinical significance.
Contrary to what was previously suggested based on the
results from animal studies,11 that the femoral and tibial
tunnels contribute predominantly to early vasculature formation at the corresponding graft sites, our findings show
that the intra-articular site was the first graft site to complete this crucial phase. These differences could be attributed
to substantial differences in the precision of the surgical technique used and the overall success of the reconstructed surgery at the knees of experimental animal models that may
affect the final healing outcome.7,11,50 Results from previous
histological studies conducted on human ACL specimens,
however, are in accordance with our findings. Falconiero
et al19 studied the maturation characteristics of BPTB autografts at the midportion and noted maximum vascular
response of the graft tissue 6 months after surgery, while
at 12 months, the degree of graft vascularity closely resembled that of normal ACL. Abe et al,1 analyzing biopsy specimens obtained at the time of second-look arthroscopy in
human ACL-reconstructed knees, reported that revascularization of the graft occurs during the early postoperative
period with no differences detected between 24 and 52 weeks
after surgery. Similarly, Rougraff and Shelbourne41 obtained
arthroscopic biopsy samples from a limited area of the intraarticular portion of BPTB autografts in uninjured ACLreconstructed knees and revealed that although this process
was not entirely equivalent with what was observed in animal models, revascularization of the graft does take place,
ensuring the grafts long-term survival.
Among noninvasive procedures, MRI has been widely
used for the postoperative evaluation of ACL graft integrity.33,39,40,53 Several MRI studies were performed to
describe the longitudinal imaging appearance of the ACL
graft substitute. Many of these studies were performed
without the administration of contrast agent, and therefore, the serial changes noted on the graft were characterized on a visual scoring basis, reflecting indirectly its
overall healing course.24,35,38 However, without the objective measurements of contrast medium uptake, the temporal changes of signal intensity could only potentially be
attributed to vascular supply.
The use of a paramagnetic contrast agent such as GdDTPA can improve the diagnostic sensitivity and specificity by direct visualization of tissue vascularity.27 It is
known that enhancement of a tissue after contrast agent
administration results in an increase of signal intensity.8,27,36,51,52 Therefore, contrast-enhanced MRI can

Downloaded from ajs.sagepub.com at TEXAS SOUTHERN UNIVERSITY on October 28, 2014

Vol. 39, No. 7, 2011

Revascularization of BPTB Autograft Evaluated by Contrast-Enhanced MRI

noninvasively monitor the serial signal intensity changes

noted during the revascularization process of the ACL
graft. On the basis of these changes before and after the
administration of contrast agent, we calculated the
enhancement index, which reflects this time-dependent
biological phenomenon and most importantly gives directly
quantitative information about the revascularization
degree. Muramatsu et al36 compared the revascularization
progress in autografts and allografts in humans, establishing a fundamental methodological basis for quantitative
analysis by means of contrast-enhanced MRI. Weiler
et al,52 similarly in a sheep model, demonstrated that the
signal intensity changes observed in the graft tissue reflect
its vascularity. However, in this animal-based study, only
the intra-articular part of the graft was examined. In our
study, we assessed graft revascularization in 3 specific
sites identified along its length. To our knowledge, this is
the first study to evaluate the outcome of the revascularization process in association with the microenvironment
surrounding each site of the graft. More interestingly,
with the use of the enhancement index as a quantitative
parameter, this can be performed in a comprehensive
and reproducible way, which provides directly comparable
results for every patient at each time interval.
Limitations of our study might be considered in that we
have not included in our study the corresponding sites of
the graft that are located inside the femoral tunnel. However, because the surgical technique ensures the same
intraosseous environment for both tibial and femoral tunnels, we can presume that the microenvironment conditions are identical, and therefore, the revascularization of
the graft in the tibial tunnel itself can be provided as
a model for the interaction between cancellous bone and
fixation material with the graft. Imaging of the intrafemoral graft portion and calculation of its signal intensity
changes require a different MRI scan design in an oblique
coronal plane. Future studies in this field would provide
valuable information regarding the revascularization rate
of this graft portion as compared with the intra-articular
and intratibial parts.
Another limitation of this study is the lack of correlation
between our findings and histological or biomechanical
analysis of graft material properties. However, the design
of our study included uncomplicated reconstructed knees
of human patients, and thus, any kind of interventional
procedure was inappropriate.
As far as the future prospective of the present study is
concerned, new hypotheses in the field of several in vivo
clinical applications can be developed and tested, benefiting from the level of quantitative information that can be
acquired noninvasively by contrast-enhanced MRI. Administration of various blood products, such as platelet-rich
plasma and bone morphogenic proteins and examination
of their acceleration effect on the revascularization rate
as well as the effect that different fixation materials (metal
screws or biodegradable screws) and surgical techniques
might have on this process, can be accurately demonstrated with this imaging modality.
To conclude, in an uncomplicated knee joint having
undergone ACL reconstruction, the graft gradually achieves

1485

revascularization, which is clearly and objectively demonstrated by contrast-enhanced MRI. This process occurs in
association with the surrounding microenvironment, with
the intra-articular site of the graft being the first fully
revascularized site even 6 months after surgery, while the
intraosseous sites were still in progress throughout the first
postoperative year. Enhancement index is an objective
imaging parameter to directly assess this phenomenon
with contrast-enhanced MRI, which can be used as a valuable noninvasive tool to evaluate the healing process of
the ACL graft.

REFERENCES
1. Abe S, Kurosaka M, Iguchi T, Yoshiya S, Hirohata K. Light and electron microscopic study of remodeling and maturation process in
autogenous graft for anterior cruciate ligament reconstruction.
Arthroscopy. 1993;9(4):394-405.
2. Abujudeh HH, Kosaraju VK, Kaewlai R. Acute adverse reactions to
gadopentetate dimeglumine and gadobenate dimeglumine: experience with 32,659 injections. AJR Am J Roentgenol. 2010;194(2):
430-434.
3. Amiel D, Akeson WH, Renzoni S, Harwood F, Abel M. Nutrition of cruciate ligament reconstruction by diffusion: collagen synthesis studied
in rabbits. Acta Orthop Scand. 1986;57(3):201-203.
4. Amiel D, Kleiner JB, Roux RD, Harwood FL, Akeson WH. The phenomenon of ligamentization: anterior cruciate ligament reconstruction with autogenous patellar tendon. J Orthop Res. 1986;4(2):
162-172.
5. Arai Y, Hara K, Takahashi T, et al. Evaluation of the vascular status of
autogenous hamstring tendon grafts after anterior cruciate ligament
reconstruction in humans using magnetic resonance angiography.
Knee Surg Sports Traumatol Arthrosc. 2008;16(4):342-347.
6. Arnoczky SP, Rubin RM, Marshall JL. Microvasculature of the cruciate ligaments and its response to injury: an experimental study in
dogs. J Bone Joint Surg Am. 1979;61(8):1221-1229.
7. Arnoczky SP, Tarvin GB, Marshall JL. Anterior cruciate ligament
replacement using patellar tendon: an evaluation of graft revascularization in the dog. J Bone Joint Surg Am. 1982;64:217-224.
8. Beltran J, Chandnani V, McGhee RA Jr, Kursunoglu-Brahme S.
Gadopentetate dimeglumine-enhanced MR imaging of the musculoskeletal system. AJR Am J Roentgenol. 1991;156(3):457-466.
9. Bicer EK, Lustig S, Servien E, Selmi TA, Neyret P. Current knowledge
in the anatomy of the human anterior cruciate ligament. Knee Surg
Sports Traumatol Arthrosc. 2010;18(8):1075-1084.
10. Bulut D, Albrecht N, Imohl M, et al. Hormonal status modulates circulating endothelial progenitor cells. Clin Res Cardiol. 2007;96(5):258-263.
11. Clancy WG Jr, Narechania RG, Rosenberg TD, Gmeiner JG,
Wisnefske DD, Lange TA. Anterior and posterior cruciate ligament
reconstruction in rhesus monkeys: a histological, microangiographic,
and biomechanical analysis. J Bone Joint Surg Am. 1981;63:12701284.
12. Clancy WG Jr, Nelson DA, Reider B, Narechania RG. Anterior cruciate ligament reconstruction using one-third of the patellar ligament,
augmented by extra-articular tendon transfers. J Bone Joint Surg
Am. 1982;64(3):352-359.
13. Cohen J. Statistical Power Analysis for the Behavioral Sciences. Mahwah, New Jersey: Lawrence Erlbaum Associates; 1988.
14. Dahlin LB, Hanff G, Myrhage R. Healing of ligaments in synovial fluid:
an experimental study in rabbits. Scand J Plast Reconstr Surg Hand
Surg. 1991;25(2):97-102.
15. Di Stefano R, Barsotti MC, Felice F, et al. Smoking and endothelial
progenitor cells: a revision of literature. Curr Pharm Des.
2010;16(23):2559-2566.
16. Dillman JR, Ellis JH, Cohan RH, Strouse PJ, Jan SC. Frequency and
severity of acute allergic-like reactions to gadolinium-containing i.v.

Downloaded from ajs.sagepub.com at TEXAS SOUTHERN UNIVERSITY on October 28, 2014

1486 Ntoulia et al

17.

18.

19.

20.

21.

22.

23.

24.

25.

26.

27.
28.

29.

30.

31.

32.

33.

34.

35.

The American Journal of Sports Medicine

contrast media in children and adults. AJR Am J Roentgenol.

2007;189(6):1533-1538.
Dunlap J, McCarthy JA, Joyce ME, Ogata K, Shively RA. Quantification of the perfusion of the anterior cruciate ligament and the effects
of stress and injury to supporting structures. Am J Sports Med.
1989;17(6):808-810.
Espregueira-Mendes J, Lopes JM, Castro C, Oliveira J. Time of
remodelling of the patella tendon graft in anterior cruciate ligament
surgery: an histological and immunohistochemical study in a rabbit
model. Knee. 1998;5:9-19.
Falconiero RP, DiStefano VJ, Cook TM. Revascularization and ligamentization of autogenous anterior cruciate ligament grafts in
humans. Arthroscopy. 1998;14(2):197-205.
Georgoulis AD, Papageorgiou CD, Makris CA, Moebius UG, Soucacos PN. Anterior cruciate ligament reconstruction with the press-fit
technique: 2-5 years followed-up of 42 patients. Acta Orthop Scand
Suppl. 1997;275:42-45.
Ginsburg JH, Whiteside LA, Piper TL. Nutrient pathways in transferred patellar tendon used for anterior cruciate ligament reconstruction. Am J Sports Med. 1980;8(1):15-18.
Grassi G, Seravalle G, Brambilla G, et al. Impact of the metabolic
syndrome on subcutaneous microcirculation in obese patients. J
Hypertens. 2010;28(8):1708-1714.
Hoher J, Kanamori A, Zeminski J, Fu FH, Woo SL. The position of the
tibia during graft fixation affects knee kinematics and graft forces for
anterior cruciate ligament reconstruction. Am J Sports Med.
2001;29:771-776.
Howell SM, Clark JA, Farley TE. Serial magnetic resonance study
assessing the effects of impingement on the MR image of the patellar
tendon graft. Arthroscopy. 1992;8:350-358.
Irrgang J, Safran M, Fu F. The knee: ligamentous and meniscal injuries. In: Zachazewski J, Magee D, Quillen W, eds. Athletic Injuries and
Rehabilitation. Philadelphia: W.B. Saunders; 1996:623-692.
Iwakura A, Luedemann C, Shastry S, et al. Estrogen-mediated, endothelial nitric oxide synthase-dependent mobilization of bone marrowderived endothelial progenitor cells contributes to reendothelialization after arterial injury. Circulation. 2003;108(25):3115-3121.
Kirsch JE. Basic principles of magnetic resonance contrast agents.
Top Magn Reson Imaging. 1991;3(2):1-18.
Kleiner JB, Amiel D, Roux RD, Akeson WH. Origin of replacement
cells for the anterior cruciate ligament autograft. J Orthop Res.
1986;4(4):466-474.
Leung DA, Pelkonen P, Hany TF, Zimmermann G, Pfammatter T,
Debatin JF. Value of image subtraction in 3D gadolinium-enhanced
MR angiography of the renal arteries. J Magn Reson Imaging.
1998;8(3):598-602.
Marumo K, Saito M, Yamagishi T, Fujii K. The ligamentization process in human anterior cruciate ligament reconstruction with autogenous patellar and hamstring tendons: a biochemical study. Am J
Sports Med. 2005;33(8):1166-1173.
Menetrey J, Duthon VB, Laumonier T, Fritschy D. Biological failure
of the anterior cruciate ligament graft. Knee Surg Sports Traumatol
Arthrosc. 2008;16(3):224-231.
Michaud SE, Dussault S, Haddad P, Groleau J, Rivard A. Circulating
endothelial progenitor cells from healthy smokers exhibit impaired
functional activities. Atherosclerosis. 2006;187(2):423-432.
Min BH, Chung WY, Cho JH. Magnetic resonance imaging of reconstructed anterior cruciate ligament. Clin Orthop Relat Res.
2001;393:237-243.
Moebius U, Georgoulis A, Papageorgiou C, Papadonikolakis A,
Rossis J, Soucacos P. Alterations of the extensor apparatus after
anterior cruciate ligament reconstruction using the medial third of
the patellar tendon. Arthroscopy. 2001;17(9):953-959.
Murakami Y, Sumen Y, Ochi M, Fujimoto E, Deie M, Ikuta Y. Appearance of anterior cruciate ligament autografts in their tibial bone tunnels on oblique axial MRI. Magn Reson Imaging. 1999;17(5):679-687.

36. Muramatsu K, Hachiya Y, Izawa H. Serial evaluation of human anterior cruciate ligament grafts by contrast-enhanced magnetic resonance imaging: comparison of allografts and autografts.
Arthroscopy. 2008;24(9):1038-1044.
37. Petersen W, Tillmann B. Structure and vascularization of the cruciate
ligaments of the human knee joint. Anat Embryol (Berl).
1999;200(3):325-334.
38. Rak KM, Gillogly SD, Schaefer RA, Yakes WF, Liljedahl RR. Anterior
cruciate ligament reconstruction: evaluation with MR imaging. Radiology. 1991;178(2):553-556.
39. Recht MP, Kramer J. MR imaging of the postoperative knee: a pictorial essay. Radiographics. 2002;22(4):765-774.
40. Recht MP, Parker RD, Irizarry JM. Second time around: evaluating
the postoperative anterior cruciate ligament. Magn Reson Imaging
Clin N Am. 2000;8(2):285-297.
41. Rougraff BT, Shelbourne KD. Early histologic appearance of human
patellar tendon autografts used for anterior cruciate ligament reconstruction. Knee Surg Sports Traumatol Arthrosc. 1999;7(1):9-14.
42. Ruehm SG, Nanz D, Baumann A, Schmid M, Debatin JF. 3D contrast-enhanced MR angiography of the run-off vessels: value of
image subtraction. J Magn Reson Imaging. 2001;13(3):402-411.
43. Runge VM, Wood ML, Kaufman D, Price AC. Gd DTPA: future applications with advanced imaging techniques. Radiographics. 1988;
8(1):161-179.
44. Saddik D, McNally EG, Richardson M. MRI of Hoffas fat pad. Skeletal Radiol. 2004;33(8):433-444.
45. Schatzmann L, Brunner P, Staubli HU. Effect of cyclic preconditioning on the tensile properties of human quadriceps tendons and patellar ligaments. Knee Surg Sports Traumatol Arthrosc. 1998;6:
S56-S61.
46. Scheffler SU, Unterhauser FN, Weiler A. Graft remodeling and ligamentization after cruciate ligament reconstruction. Knee Surg Sports
Traumatol Arthrosc. 2008;16(9):834-842.
47. Sckell A, Leunig M, Fraitzl CR, Ganz R, Ballmer FT. The connectivetissue envelope in revascularisation of patellar tendon grafts. J Bone
Joint Surg Br. 1999;81(5):915-920.
48. Shankar SS, Steinberg HO. Obesity and endothelial dysfunction.
Semin Vasc Med. 2005;5(1):56-64.
49. Steiner ME, Brown C, Zarins B, Brownstein B, Koval PS, Stone P.
Measurement of anterior-posterior displacement of the knee: a comparison of the results with instrumented devices and with clinical
examination. J Bone Joint Surg Am. 1990;72(9):1307-1315.
50. Unterhauser FN, Bail HJ, Hoher J, Haas NP, Weiler A. Endoligamentous revascularization of an anterior cruciate ligament graft. Clin
Orthop Relat Res. 2003;414:276-288.
51. Vogl TJ, Schmitt J, Lubrich J, et al. Reconstructed anterior cruciate
ligaments using patellar tendon ligament grafts: diagnostic value of
contrast-enhanced MRI in a 2-year follow-up regimen. Eur Radiol.
2001;11(8):1450-1456.
52. Weiler A, Peters G, Maurer J, Unterhauser FN, Sudkamp NP. Biomechanical properties and vascularity of an anterior cruciate ligament
graft can be predicted by contrast-enhanced magnetic resonance
imaging: a two-year study in sheep. Am J Sports Med. 2001;
29(6):751-761.
53. White LM, Kramer J, Recht MP. MR imaging evaluation of the postoperative knee: ligaments, menisci, and articular cartilage. Skeletal
Radiol. 2005;34:431-452.
54. Whiteside LA, Sweeney RE Jr. Nutrient pathways of the cruciate ligaments: an experimental study using the hydrogen wash-out technique. J Bone Joint Surg Am. 1980;62(7):1176-1180.
55. Yu WD, Liu SH, Hatch JD, Panossian V, Finerman GA. Effect of estrogen on cellular metabolism of the human anterior cruciate ligament.
Clin Orthop Relat Res. 1999;366:229-238.
56. Zaffagnini S, De Pasquale V, Marchesini Reggiani L, et al. Neoligamentization process of BTPB used for ACL graft: histological evaluation from 6 months to 10 years. Knee. 2007;14(2):87-93.

For reprints and permission queries, please visit SAGEs Web site at http://www.sagepub.com/journalsPermissions.nav

Downloaded from ajs.sagepub.com at TEXAS SOUTHERN UNIVERSITY on October 28, 2014