CC BY-NC-ND
0The Egyptian Journal of Radiology and Nuclear Medicine (2014) xxx, xxx-xxx
Egyptian Society of Radiology and Nuclear Medicine The Egyptian Journal of Radiology and Nuclear Medicine
www.elsevier.com/locate/ejrnm www.sciencedirect.com
ORIGINAL ARTICLE
MR cartilage imaging in assessment of the regenerative power of autologous peripheral blood stem cell injection in knee osteoarthritis
Khaled A. Ahmad a,% Yosra A. Ibrahim a, Nayera Z. Saber b, Bassem A. Darwish
a Radiology Department, Ain Shams University, Egypt
b Physical Medicine, Rheumatology, Rehabilitation Department, Ain Shams University, Egypt c Orthopedic Department, Ain Shams University, Egypt
Received 9 March 2014; accepted 6 May 2014 Available online xxxx
KEYWORDS
PBSC intra-articular injection;
Cartilage imaging;
Osteoarthritis;
Abstract Background: Osteoarthritis (OA) describes an age-related, heterogeneous group of disorders characterized pathologically by focal areas of loss of articular cartilage in synovial joints, associated with varying degrees of osteophyte formation, subchondral bone change, and synovitis. Currently, cartilage repair remains a major challenge for physicians, being avascular with limited regenerative capacity. Stem cell therapy opened new horizons for hyaline cartilage repair. Peripheral blood stem cells (PBSC) due to their multi-lineage potential, immunosuppressive activities, and limited immunogenicity, were tried as an intra articular injection.
Aim of study: To find out the regenerative effect of repeated intra articular injections of autologous PBSC in knee joints of OA patients using MR cartilage imaging.
Methods: 10 patients (3 males and 7 females) diagnosed with bilateral knee joints OA were included in this study, they underwent history taking, clinical examination and MR cartilage imaging using the semi-quantitative whole joint assessment score of knee for OA (MOAKS). Three intra articular injections of 8 ml of autologous PBSC in each knee were administered. Clinical and MRI assessments were repeated after 1 year.
Results: A significant reduction was seen in all parameters post injection. MR images analysis showed increased cartilage thickness in 65 knee joint compartments out of 160 affected compartments.
Corresponding author. Address: Radiology Department, Faculty of Medicine, Ain Shams University, Cairo, Egypt. Tel.: +20 1285185185; fax: +20 1005089011.
E-mail addresses: Drkhaled_mohamed@med.asu.edu.eg (K.A. Ahmad), Yosra_zaher@yahoo.com (Y.A. Ibrahim).
Peer review under responsibility of Egyptian Society of Radiology and Nuclear Medicine.
0378-603X © 2014 Production and hosting by Elsevier B.V. on behalf of Egyptian Society of Radiology and Nuclear Medicine. http://dx.doi.org/10.1016/j.ejrnm.2014.05.012
Conclusion: Limited good level of evidence showed that repeated intra-articular injections of autologous PBSC resulted in an improvement of the quality of articular cartilage repair and physical function as observed by MRI and clinical assessment.
© 2014 Production and hosting by Elsevier B.V. on behalf of Egyptian Society of Radiology and Nuclear
Medicine.
1. Introduction
Osteoarthritis is a degenerative condition of the cartilage and other tissues such as the synovium in which immunological and inflammatory reactions occur contribute to the development of joint pathology (1) and clinically, patients with osteoarthritis (OA) express variable synovitis. Thickening of the lining layer containing predominantly macrophages produce elevated levels of pro-inflammatory factors and damage-associated molecular pattern molecules (2).
The release of cartilage matrix fragments from damaged cartilage may give a prolonged stimulation of synovial macrophages, thereby forming a positive feedback loop that drives deleterious synovitis. This sheds new light on the potential mechanism of action of mesenchymal stem cell (MSC) therapy in osteoarthritis (1).
Although it is generally accepted that the primary effect of stem cell treatment occurs through tissue-specific differentiation, new data suggest that the therapeutic potential of these cells might also be related to their paracrine effect (3,4).
Some orthopedists try to treat OA patients with biologic reconstruction as soon as possible. Numerous procedures are available. These procedures include micro fracture or micro drilling surgery, autologous chondrocyte implantation (ACI), mosaicplasty, and matrix-guided autologous chondrocyte implantation, among other approaches (5).
However, the drawbacks of ACI include limited cell sources, difficulty in phenotype retention, and donor-site morbidity, all of which challenge autologous cell transfer procedures. Thus, new strategies rely upon cell therapies that explore the use of stem cells rather than primary chondrocytes for cartilage regeneration (6).
Therefore this study of repeated intra articular injections of PBSC in knees of OA patients without prior surgery will declare whether this non invasive application will also help to prevent joint destruction within the human OA joint.
Magnetic resonance imaging (MRI) provides high-spatial-resolution, multiplanar imaging and excellent tissue contrast. This enables a three-dimensional assessment of all components of the joint simultaneously, allowing direct visualization of articular cartilage. With the advances in techniques and development of dedicated sequences, MRI has become the imaging modality of choice in both clinical and research settings of musculoskeletal diseases, in particular osteoarthritis (OA), a chronic joint disease characterized by destruction and progressive loss of articular cartilage and clinical symptoms including pain, stiffness and impaired function (7).
2. Aim of the study
To find out the regenerative effect of repeated intra articular injections of autologous PBSC in knees of osteoarthritis patients using MR cartilage imaging.
3. Materials and methods
10 patients (3 males and 7 females; age range 38-64 years with bilateral knee joints OA were included in this study which was conducted during a period of one year, in a University Hospitals. The research carried out here on human subjects was in compliance with the Helsinki Declaration, and informed consent was obtained from all study subjects.
Inclusion criteria: include osteoarthritis diagnosed by X-ray and MRI and end stage osteoarthritis candidate for total knee replacement.
Exclusion criteria: include pregnancy or lactating, positive tests for HIV, HCV, and HBV, any bleeding disorders or blood diseases, active neurologic disorder, end organ damage, and uncontrolled endocrine disorders.
All patients underwent history taking and thorough clinical examination with emphasis on:
1- WOMAC index (8) questionnaire to evaluate the condition of patients, including pain (0-20), stiffness (0-8), and physical functioning of joints (0-68). 0 = None, 1 = Slight, 2 = Moderate, 3 = Very, 4 = Extremely (11). WOMAC questionnaire is used before and 12 months after treatment.
2- The 6-min walk distance (6MWD) is a test where the subject walks for 6 min on level ground, and the distance covered in 6 min is measured (9).
3- Plain X-ray A-P and lateral views before treatment to estimate joint space loss and any defect in bone or osteo-phytes using Kellgren-Lawrence grading scale (10).
4- MRI with cartilage imaging technique, to measure the thickness of the cartilage and number of affected compartments, presence of osteophytes, effusion, meniscal extrusion before and 12 months after treatment Fig. 1.
3.1. MR acquisition
MR images of both knees were obtained for all patients with a 1.5-T (Achiva; Philips Medical Systems, Best, the Netherlands) MR system with an extremity coil.
3.2. MR imaging protocol included
Sagittal dual echo TSE sequence TR/TE 3000/50 and 80 ms, thickness, 3.5 mm; gap, 0.35 mm; matrix 256 x 200; FOV 170 x 170 x 84 mm. Sagittal and coronal intermediate weighted SE sequence with fat suppression "SPAIR" TR/TE 3000/50 ms; thickness, 3.5 mm; gap, 0.35 mm; matrix, 256 x 200; FOV 170 x 170 x 77 mm. Coronal T1W_TSE (TR/ TE 500/17 ms; section thickness, 4 mm; gap, 0.4 mm; sections, 20; matrix, 292 x 165; FOV 180 x 153 x 88 mm. Axial mFFE sequence "multislice fast field echo'' (TR/TE/delta TE, 940/
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WL: 1174 WW: 2183 T: 3.5mm L: -96.4mm
TR: 3000.0 TE: 6/3/2013 1:02:01
Fig. 1 (A) Sagittal fat suppressed intermediate weighted image (TE = 50 ms) along the lateral aspect of the knee joint. Normal cartilage thickness and signal is demonstrated along the lateral femoral (anterior, central and posterior) sub regions. The normal cartilage exhibits intermediate signal against the dark signal subchondral bone. (B) Sagittal fat suppressed intermediate weighted image of the same patient along the medial aspect of the knee joint. Full thickness cartilage loss is demonstrated at the medial femoral (anterior and central) sub regions. The posterior sub region shows lost articular cartilage along more than 50% of the sub region with a small area of intact residual cartilage superiorly.
WL: 968 WW: T: 3.0mm L: 6.4m
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TR: 20/ TE: 7.6 WL: 1103 WW: 1B17 [D] 6/27/2013/l(/l9:35 AM | T: 3.0mm L: 6.Vim
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WL: 863 WW: 1694 T: 3.0mm L: 52.5mm
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Fig. 2 (A) 3D WATS C axial image at the level of the patella. There is intact articular cartilage along the medial and lateral patellar facets. The cartilage demonstrates a hyper intense signal, while the joint fluid appears hypo intense. (B) 3D WATS F axial image was seen at the same level as A in the same patient. The cartilage demonstrates an intermediate signal, while the joint fluid appears hyper intense. (C) 3D WATS C axial image was seen in a different patient. There is partial thickness cartilage loss along the medial patellar sub region, while there is intact cartilage along the lateral patellar sub region.
9.2/8.2 ms; thickness, 4 mm; gap 0.4 mm; matrix, 232 x 187, FOV 160 x 160 x 88 mm. 3D_WATS F axial (TR/TE 20/ 7.4 ms, over contiguous slices; sections, 60; matrix, 204 x 203; FOV 140 x 140 x 90 mm. 3D_WATS C axial (TR/TE 20/ 7.6 ms, over contiguous slices; sections, 60; matrix, 252 x 252; FOV 150 x 150 x 90 mm Fig. 2.
3.3. MR Interpretation
MR examinations were selected for semi quantitative assessment. Two radiologists (Y.I. and K.A.), who were blinded to radiographic OA grade and clinical data, evaluated the MR images by using MRI Osteoarthritis Knee Score (MOAKS) (11).
Baseline and follow-up MR images were read in pairs, with the chronological order known to the readers. The following joint structures were assessed in this study: cartilage morphology and signal intensity, subchondral bone marrow lesions, meniscal status, effusion, and anterior cruciate ligament status. Cartilage signal intensity and morphology were scored according to the MOAKS system from 0 to 3 in five sub regions each in the medial and lateral tibifemoral (TF) compartments, for a total of 10 TF sub regions and two sub regions each in medial and lateral patellofemoral compartments for a total of 4 sub regions. The number of affected joint compartments was determined in each examined knee. On follow up scans, the number of responding compartments that showed increased cartilage thickness after treatment was also determined. The maximum scores of cartilage loss and full thickness cartilage loss were determined for each knee which is the maximum score obtained in any sub region. The percentage of change of these maximum scores was calculated in the pre and post-treatment MRI scans. Osteophyte formation was scored from 0 to 3 according to the osteophyte size. Bone marrow lesions were scored from 0 to 3 on the basis of the extent of regional involvement. Meniscal status was graded from 0 to 3 in the anterior horn, the body segment, and the posterior horn of the medial and lateral menisci. The anterior cruciate ligament was scored either as intact or torn. Joint effusion was graded from 0 to 3 in terms of the estimated maximum distention of the synovial cavity (11).
3.4. Statistical methodology
Analysis of data was done using SPSS v12 to describe quantitative and qualitative variables. Wilcoxon signed test was used in non-parametric data and paired t-test in parametric data. Spearman correlation test was used to rank variables versus each other's positively or inversely.
4. Results
This study included 7 females and 3 males with moderate to severe OA diagnosed by plain X-ray as asymmetrical loss of joint space, osteophytes, and subchondral bone sclerosis. Their mean age was 51 ± 13 years. range (38-64 years) and mean BMI 32 ± 1.2.
Clinical knee examination revealed painful limited flexion 110° in 8 patients while unlimited in 2. Genu varum deformities less than 10°, MCL laxity and patellofemoral friction were evident in all patients. 15% had moderate effusion with
synovial hypertrophy, while 50% had positive Mc Murray test and further subjective evaluation of osteoarthritis by WOMAC index was done 1 week prior to treatment by one physiatrist and the results illustrated in (Table 1).
All patients performed the 6MWD test by the same physiat-rist 1 week prior to treatment and mean distance measured was 306 ± 62 m range (200-400).
Plain X-ray of 20 knees revealed that all patients had radiological features of OA, severity grades are listed as 10% of knees was minimal grade, 50% was moderate and 40% was severe.
MRI findings before PBSC injection, mean number of affected compartments was 8 ± 2, while osteophyte scoring was maximum 3 in all patients, effusion was mild in 80% of knees, moderate in 10% and severe in 5% of knees while 5% had no effusion. Ligaments/tendons assessment showed MCL sprain was evident in 18 knees while 35% knees revealed ACL tear. 2 patients had edematous subchondral lesions. Other MRI findings, meniscal affection was evident in 95% of knees. 55% of knees had grade 3 extrusion, 15% of knees showed grade 2, while 30% showed grade 1 meniscal extrusion Fig. 3.
The values of number of affected compartments in knees prior to PBSC injection and number of responding compartments post injection and comparison between them were illustrated in (Table 2).
Upon studying MRI findings with clinical tests, there were significant positive correlation between physical functional
Table 1 WOMAC index results before treatment.
Variable Mean ± SD
Stiffness 6.2 ± 0.9
Pain 15.4 ± 2.4
Physical function limitations 51 + 7
Total WOMAC 72.7 ± 9
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TR: 3000.0 TE: 50.0 6/3/2013 1:07:36 PM
Fig. 3 Coronal fat suppressed intermediate weighted image (TE = 50 ms) of the knee joint. There is subchondral marrow edema along the femoral and tibial aspects, most appreciated at the subspinous tibial region, extruded medial meniscus and loss of articular cartilage at the medial compartment on both opposing articular surfaces.
Table 2 Comparison between number of affected and number of responding compartments.
Variables Mean ± Z P
Number of affected compartments 8 ± 2 -3.919 60.01
Number of responding compartments 3.5 ± 1.8
limitation and number of affected compartments before injection (Table 3). On the other hand, 6MWD test was not significantly correlated with meniscal extrusion score.
Twelve months Post PBSC intra articular injection from day 1 in both knees, no signs of infection or post operative complications were reported except swelling, warmth in knee, difficulty in moving knee, and pain at injection site within the first 2 weeks. Most of the patients reported gradual improvement; regarding pain, stiffness, walking on a flat surface and light domestic duties but not in ascending or descending stairs; starting after the 4th month and persisting till the end of the study period; except one patient who showed no improvement at the end of the study period and underwent total knee replacement in his RT knee. Flexion ROM was improved to 140° in the 8 patients, and effusion disappeared in all patients. Post injection values of WOMAC score and 6MWD are expressed and compared to pre injection values in (Table 4).
Table 4 showed that, stiffness, pain, functional limitation and WOMAC score declined, while 6MWD increased post treatment with statistically significant difference in comparison to pretreatment results using the paired test. WOMAC index sections revealed a significant reduction in all parameters with maximum improvement in pain score as the percentage of change was 29%. 6MWD was increased by more than 54 m which reflect significant improvement in physical function. Moreover, post injection pain score was significantly correlated inversely with number of responding compartments (r = -0.63, P 6 0.05).
Post injection analysis of MR images of cartilage thickness were scored according to the MOAKS system from 0 to 3 in 14 sub regions. Improvement was expressed as a percentage of change of the maximum scores and was calculated in the pre and post treatment MRI scans. The mean percentage of change of maximum score of cartilage loss was 33 ± 0.0 and
the mean percentage of change of the maximum score of full thickness cartilage loss was 44 + 27 Fig. 1.
Moreover, total WOMAC index was significantly inversely correlated with percentage of change of maximum score of full thickness cartilage loss; denoting cartilage repair (r = -0.073, P 6 0.05).
5. Discussion
Osteoarthritis (OA) is a progressive disorder of the joints caused by gradual loss of articular cartilage, which naturally possesses a limited regenerative capacity (12).
The goals of successful cartilage repair include reducing pain, improving symptoms and long term function; preventing early osteoarthritis and subsequent total knee replacements; and rebuilding hyaline cartilage instead of fibrous cartilage
Conventional treatment is aimed at reducing pain, maintaining mobility and minimizing disability (1). However, till date, no technique has reliably regenerated the biological composition and biomechanical properties of native cartilage, leaving unresolved pain and loss of joint function for millions of patients with defective cartilage from aging, injury, or disease. Emerging evidence indicates that direct intra-articular injection of stem cells may boost the normally limited repair and limit the destructive process (14).
In our study, the authors proposed autologous peripheral blood stem cells (PBSC) as opposed to cultured MSC or marrow aspirate due to the ease of harvest and the increased potential of this cell line. This in in congruency with a recent study by Saw et al., who injected intra articular autologous PBSC in combination with hyaluronic acid (HA) a cartilage regeneration protocol (15).
In our study all patients with osteoarthritis had positive X-ray findings suggestive from mild to severe OA. Subjective assessment of the patients was done by the WOMAC index as a reliable index of disability for OA and high scores which indicate worse pain, stiffness, and functional limitations (16). The mean of pain, stiffness and physical function scores was 6.2 ± 0.9, 15.4 ± 2.4 and 51 ± 7, respectively. 6MWD was used as a predictor of functional outcomes in rehabilitation purposes. It was originally designed as a "useful measure of exercise capacity'' by researchers working with patients demonstrating chronic heart failure (9). In this study it was used before injection to reveal functional disability as mean of 6MWD was 306 ± 62 which was lower than normal values (9).
We studied the relation between clinical parameters and MRI findings, there was significant positive correlation between physical functional limitation and the number of affected compartments before injection Table 2. This could be explained by many activities of the WOMAC index; as
Table 3 Correlation between functional limitations versus number of affected compartments.
Variables Physical functional
Number of affected compartments r P
0.59 60.05 NS
Table 4 Comparison between clinical parameters pre and post injection.
Variables Pre (mean + SD) Post (mean + SD) % of change t P
Stiffness 6.2 + 0.9 5 + 1.5 17 4.8 <0.001 HS
Pain 15.4 + 2.4 11 + 2.8 29 8.4 <0.001 HS
Function limitation 51+7 43 + 6.5 24 9.3 <0.001 HS
WOMAC (total) 72.7 + 9 59 + 10 19.1 13 <0.001 HS
6 m WD 306 + 62 387 + 87 26 6 <0.001 HS
going upstairs and downstairs, getting in/out of bath, heavy domestic duties; which produce maximum compression of all compartments.
Most of the patients reported gradual improvement in pain, stiffness and physical function and this was clear when compared with pre injection values in Table 4. The amount of improvement was 29% and physical function improved by 24%. Moreover, comparing pretreatment values of 6MWD with post treatment ones, revealed significantly increased distance i.e. improved functional performance and post treatment WOMAC index inversely correlated with percentage of change of cartilage thickness i.e. cartilage repair (r = —0.073, P 6 0.05).
Emadedin et al. reported another case series involving six patients. They found that patients were partly satisfied, walking ability was slightly decreased 6 months post injection,
and this was coincident with an increase in cartilage thickness and extension of the repair tissue over the subchondral bone (12).
5.1. MR images
MRI allows comprehensive imaging of joint structures and structural changes over time in patients with knee OA. A number of semi quantitative scoring methods and quantitative technologies have been developed and validated. They have been shown to be reliable in the assessment of structural changes cross-sectional and longitudinal (17).
Meniscal degeneration was evident in 95% of examined knees, while 55% of knees had grade 3 extrusion, 15% of knees showed grade 2, and 30% showed grade1 meniscal extrusion. Meniscal tear was found in one knee. The strong
Fig. 4 Axial 3D WATS of a knee of a 64 years old female with osteoarthritis before (A) and after (B) intraarticular injection of autologous stem cells. (A) There is full thickness cartilage loss over more than 75% of the surface area of the medial patellar sub region (grade 3 full thickness cartilage loss). (B) A thin rim of hyper intense cartilage signal formed denoting partial response to therapy.
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Fig. 5 Sagittal intermediate density weighted image with fat suppression (TE = 50 ms) of a knee of a 65 years old female with osteoarthritis before (A) and after (B) intraarticular injection of autologous stem cells. (A) Shows hyper intense subchondral marrow edema at the medial central femoral sub region (grade 2) and the medial central tibial sub region (grade 3). (B) There is regression of the signal abnormality that is down staged to grade 1 on the femoral side and grade 2 of the tibial side.
association of baseline medial meniscal damage and malposition with consequent cartilage loss in the same compartment in subjects with OA is well established (18,19).
In our study, we examined the patient's articular cartilage with MRI and accordingly cartilage thickness appeared to be increased in 65 knee joint compartments out of 160 affected compartments, indicating that injected stem cells participate in repair of the damaged cartilage in OA knees Fig. 4. In the study done by Emadedin et al., (12) cartilage thickness appeared to be increased in three out of six patients following intra articular injection of autologous MSC. This was similar to our study regarding the way of articular cartilage assessment as both studies used the semi quantitative scoring methods.
This is in common with a recent study by Saw et al., who concluded that after arthroscopic subchondral drilling into grade 3 and 4 chondral lesions, postoperative intra-articular injections of autologous PBSC in combination with HA resulted in an improvement of the quality of articular cartilage repair over the same treatment without PBSC, as shown by histologic and MRI evaluation (20).
One recent study showed that MRI quantitative methodology may be more sensitive to change during a 2-year observation period than the semi quantitative scoring methods to assess cartilage loss in the context of clinical trials (21). This relative lack of sensitivity to change compared with quantitative methods is a known weakness of semi quantitative assessments (22).
In the same study of Emadedin et al., there was decrease in the size of edematous subchondral patches following the intra-articular injection in the MRI images. In our study, there was decrease in size in the edematous subchondral lesions in two patients, in three compartments in one, and two compartments in the other Fig. 5 and such effects can be attributed to anti-inflammatory influences of the injected stem cells. (23).
In our study MR images showed complete ACL tear in two knees of two different patients and partial tear in one knee in another one. An increased risk of subsequent cartilage loss for knees with established OA and baseline complete anterior cruciate ligament tears has been reported previously (24).
Repeated injection for 3 times were suggested in this study as a compensatory method for other adjuvants such as HA to overcome cells dilution and dispersion inside the joint cavity.
6. Conclusion
Limited good level of evidence showed that repeated intra-articular injections of autologous PBSC resulted in an improvement of the quality of articular cartilage repair and physical function as observed from the MRI and clinical assessment.
No funds, sponsorship or financial support to be disclosed. Conflict of interest
No conflict of interest.
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