Introduction:
Cartilage oligomeric matrix protein (COMP) is a tissue specific non-collagenous, five-armed 435 KD matrix protein primarily found in cartilage required for normal development and function of cartilage.[1] It interacts with collagen and plays a role in regulating fibril assembly to stabilize the mature collagen network.[2,3] It is a soluble biomarker for cartilage degradation useful in knee Osteoarthritis (KOA) research; for differentiating between healthy individuals who clinically have KOA and those who do not have it.[4] COMP can also be used to determine the burden and prognosis of KOA.[5,6]
KOA develops over decades characterized by a prolonged asymptomatic molecular phase, followed by a preradiographic phase, and a later radiographic phase during which structural joint changes become evident. Biomarkers such as COMP have the potential to provide an early warning of the initiation of matrix breakdown that could prompt early treatment to prevent the cartilage and bone destruction that leads to disability.[7] Some researchers have demonstrated that changes in levels of serum COMP reflect metabolic processes and changes occurring in the cartilage matrix of diseased joints[8,9] and that the level of serum COMP is elevated and correlate with cartilage degradation in diseases such as KOA.[10,11] It has also been shown to be consistently elevated in the serum of patients with radiographically diagnosed KOA and the serum levels were elevated according to KOA severity when compared to healthy controls.[12]
COMP has been identified in the literature to diagnose pre-radiographic KOA.[13,14] and elevated levels of serum COMP are detected in the serum long before radiological features of osteoarthritis become obvious making early detection of KOA possible.[15] The feasibility of serum COMP as a prognostic indicator of future joint damage and as a marker of ongoing joint damage in osteoarthritis, has also been suggested by some researchers.[16,17] The fact that the current diagnostic tools for KOA are mainly clinical and radiological, and that early osteoarthritic changes such as articular cartilage abnormalities are silent, diagnosis of KOA is often delayed until it has reached an irremediable and disabling stage. Hence the need to establish other tools capable of diagnosing this condition at the early stage[18] when early implementation of effective intervention strategies may lead to a better prognosis.
Establishing reference intervals for COMP in healthy individuals who do not have clinical symptoms and radiographic evidence of KOA; Kellgren and Lawrence (K/L) grade 0 [19] provides reference values for comparison of clinical laboratory test results of symptomatic KOA patients without radiological features. The fact that levels of serum COMP is sensitive to differences between healthy individuals and those with KOA,[6,20] makes it a potential tool for making early diagnosis of KOA in symptomatic patients whose laboratory results are above the corresponding upper reference limit even when there are no radiological features. To the knowledge of the researchers, serum COMP has not been introduced into routine clinical practice and there are presently no established reference intervals for COMP in Nigeria. The aim of this study was to determine the reference intervals of serum COMP in Nigerian adults of Igbo ethnic group at Nnewi, a town in south eastern Nigeria.
Methods
Sample Population
Eighty healthy volunteers with no clinical features or radiographic evidence of KOA Kellgren/Lawrence (K/L) grade 0 were recruited into the study. Nnewi is the second largest city in southeastern Nigeria inhabited by the Igbo ethnic group and all those who participated in the study were Igbos. Approval was obtained from the Hospital Ethical Committee and written informed consent was given by all the participants. After due documentation of the clinical history and physical examination findings of the participants, anthropometric measurements of weight, height and body mass index (BMI) were recorded in kilograms (KG), meters (M) and kilograms/meters2 (kg/m2) respectively. Age was recorded in years. Bilateral anterior-posterior and lateral weight-bearing plain radiographs of the knees of the participants were obtained after collection of blood samples. The radiographs were reported by a single radiologist using the K/L atlas for overall radiographic grading: K/L grade 0 was defined as showing no radiographic features of OA.[18]
Selection Criteria
Only volunteers who had no clinical or radiographic features of OA or rheumatoid arthritis (RA) in the knee, hand joints, hips, shoulders or spine were included in the study. Volunteers who had liver, renal and lung pathologies, systemic lupus erythematosus, systemic sclerosis and scleroderma dermal fibroblasts were excluded from the study. Patients with inflammatory joint disorders, previous anterior cruciate ligament injury, or known injury to the menisci were also excluded.
COMP Analyses
Five milliliters (5ml) of venous blood sample was collected from the vena mediana cubitii of each participant after resting for half an hour in a seated position to avoid the effect of physical activity on serum COMP levels [21] between 12 noon and 4pm to avoid diurnal variations [22] by a board certified laboratory scientist. After clotting for 60 minutes at room temperature, the sera were separated by centrifugation and stored at -20°C until all the samples were obtained. The samples were then stored at -80°C until analysis. Serum COMP levels were analysed with the COMP ELISA (AnaMar Medical AB, Lund, Sweden). The COMP ELISA is a quantitative solid-phase, two-site enzyme-linked Immunosorbent assay for the determination of COMP in human serum. It is based on the direct sandwich technique in which two monoclonal antibodies are directed against separate antigenic determinants on the COMP molecule.
Statistical Analysis
The statistical package for social sciences (SPSS) software version 20.0 was used for data entry and analysis. Descriptive statistics of mean and standard deviation were calculated for all measurements taken and independent t-test was used for the between group comparisons for mean scores of all parameters. Pearson’s coefficient was used to calculate correlations and Alpha level for all statistics employed was set at p < 0.05. Data was log transformed. Reference intervals for serum COMP for the various groups were obtained after back transformation of the calculated values obtained using the formula: reference interval = m - 2s to m + 2s for a variable that follows a normal distribution where m was the mean and s was the standard deviation.
Results
The physical characteristics of the eighty participants who were recruited into the study are shown in Table 1. There were forty males and forty females; partitioned into two age groups: group 1 comprising those 29 years and younger and group 2 comprising those 30 years and older. Comparison of the physical characteristics of the two groups (Table 2) shows that the difference in height between the two groups was not statistically significant p=.550 while the difference in age, weight, BMI and serum COMP was statistically significant: p=.001, p=.006, p=.0001 and p=.001 respectively.
| Table 1: Physical characteristics of the participants |
|
N |
Minimum |
Maximum |
Mean |
Standard Deviation |
Age (years) |
80 |
19 |
56 |
31.81 |
10.10 |
Height (m) |
80 |
1.62 |
1.95 |
1.77 |
.07 |
Weight (Kg) |
80 |
62.00 |
90.00 |
75.09 |
4.93 |
BMI (Kg/m) |
80 |
21.68 |
26.67 |
23.87 |
1.05 |
COMP (ng/dl) |
80 |
170.30 |
512.20 |
267.06 |
95.05 |
| Table 2: Comparison of the physical characteristics of both groups (group1: participants 29 years and below, group 2: participants 30 years and above) |
|
Groups |
N |
Mean |
Standard Deviation |
t-value |
p-value |
Age(years) |
Group 1 |
40 |
22.35 |
1.97 |
25.040 |
.000 |
Group 2 |
40 |
41.28 |
4.36 |
|
|
Height (m) |
Group 1 |
40 |
1.77 |
.07 |
.600 |
.550 |
Group 2 |
40 |
1.78 |
.06 |
|
|
Weight (Kg) |
Group 1 |
40 |
73.58 |
5.69 |
2.870 |
.006 |
Group 2 |
40 |
76.60 |
3.48 |
|
|
BMI (Kg/m2) |
Group 1 |
40 |
23.49 |
.80 |
3.445 |
.001 |
Group 2 |
40 |
24.25 |
1.15 |
|
|
COMP (ng/dl) |
Group 1 |
40 |
193.58 |
9.89 |
10.938 |
.000 |
Group 2 |
40 |
340.55 |
84.41 |
|
|
There were twenty males and twenty females in each group. In group 1 (Table 3), there was a difference in the mean (SD) age and BMI between the males and females that was not statistically significant p=.875 and p=.978 respectively while the difference in the mean (SD) height, weight, and serum COMP was statistically significant p=.001, p=.001 and p=.002 respectively.
| Table 3: Comparison of the physical characteristics of the males and females in group 1 (participants 29 years and younger) |
|
Gender |
N |
Mean |
Standard Deviation |
T-value |
P-value |
Age (years) |
M |
20 |
22.30 |
1.81 |
-.159 |
.875 |
F |
20 |
22.40 |
2.16 |
|
|
Height (m) |
M |
20 |
1.81 |
.06 |
4.793 |
.000 |
F |
20 |
1.73 |
.06 |
|
|
Weight (Kg) |
M |
20 |
77.25 |
4.68 |
5.334 |
.000 |
F |
20 |
69.90 |
4.01 |
|
|
BMI (Kg/m2) |
M |
20 |
23.49 |
.93 |
.028 |
.978 |
F |
20 |
23.49 |
.66 |
|
|
COMP (ng/dl) |
M |
20 |
198.21 |
8.10 |
3.326 |
.002 |
F |
20 |
188.94 |
9.47 |
|
|
| M-male; F-female |
The difference in the mean (SD) age between the males and females in group 2 was not statistically significant p=.915 while the difference in the mean (SD) height, weight, BMI and serum COMP was statistically significant; p=.001, p=.001, p=.003 and p=.006 respectively (Table 4).
Table 4: Comparison of the physical characteristics of the males and females in group 2 (participants 30years and older) |
|
Gender |
N |
Mean |
Standard Deviation |
T-value |
P-value |
Age(years) |
M |
20 |
41.35 |
4.61 |
.108 |
.915 |
F |
20 |
41.20 |
4.20 |
|
|
Height(m) |
M |
20 |
1.82 |
.06 |
4.773 |
.000 |
F |
20 |
1.74 |
.04 |
|
|
Weight(Kg) |
M |
20 |
78.30 |
3.31 |
3.51 |
.001 |
F |
20 |
74.90 |
2.79 |
|
|
BMI (Kg/m2) |
M |
20 |
23.73 |
1.03 |
-3.198 |
.003 |
F |
20 |
24.77 |
1.04 |
|
|
COMP (ng/dl) |
M |
20 |
376.25 |
86.61 |
2.922 |
.006 |
F |
20 |
304.85 |
66.64 |
|
|
| M-male; F-female |
In group 1, serum COMP showed moderate positive correlation r=.546 with age which was statistically significant p=.001 but correlation with height, weight and BMI was not statistically significant (Table 5). In group 2, there was good positive correlation with age r=.624 which was statistically significant p=.001. Correlation with height, weight and BMI was not statistically significant (Table 5).
| Table 5: Pearson’s Correlation test for COMP with physical characteristics of participants in the two groups |
Group 1 (29 years and younger) |
|
Age |
Height |
Weight |
BMI |
COMP (ng/dl) Pearson’s Correlation |
.546 |
.147 |
.272 |
.262 |
P-value |
.000 |
.367 |
.090 |
.102 |
Group 2 (30 years and older) |
|
Age |
Height |
Weight |
BMI |
COMP (ng/dl) Pearson’s Correlation |
.624 |
.196 |
.051 |
-.241 |
P-value |
.000 |
.227 |
.753 |
.133 |
The reference interval for the males in group 1 was 182.87 - 214.52ng/dl while for the females it was 170.67 - 208.66ng/dl. For the males in group 2 it was 174.54 – 608.22ng/dl and 182.97 – 482.19ng/dl for the females (Table 6). The upper reference limits were higher for the males in both groups.
| Table 6: Reference intervals for the different gender age groups |
|
Gender |
Reference Interval(ng/dl) |
Group 1 (29 years and younger) |
M |
182.87 – 214.52 |
F |
170.67 – 208.66 |
Group 2 (30 years and older) |
M |
174.54 – 608.22 |
F |
182.97 – 482.19 |
M-male; F-female |
Discussion
The International Federation of Clinical Chemistry and Laboratory Medicine (IFCC) and Clinical and Laboratory Standards Institute (CLSI) have defined reference interval as the interval between two reference limits; an upper and lower reference limit, which are estimated to include a specified percentage (usually 95%) of the values for a population from which the reference subjects are drawn.[23] Reference intervals are necessary to enable physicians interpret clinical laboratory results when making specific clinical decisions. COMP is one of the biomarkers of KOA that has been studied to potentially aid in early diagnosis and assessment of minor changes in bone and cartilage that are predictive of further development of KOA.[14, 24] Establishing reference intervals for serum COMP in healthy Nigerians potentially provides a tool with which laboratory results can be compared to diagnose early KOA in the absence of radiological features.
In this study there was a significant gender difference in levels of serum COMP between the males and females in the two age groups. The serum COMP was higher in the males than in the females in both groups and the difference was statistically significant; p=.002 in group 1and p=.006 in group 2. Though the difference in the height and weight between the males and females in group 1 and the height, weight and BMI between the males and females in group 2 was statistically significant, the association between COMP and these physical attributes was not statistically significant. We can infer from the correlation results that the significant gender difference in serum levels of COMP may not be due to the differences in physical characteristics. The results of this study concur with some other studies that have reported higher serum COMP in males. Jordan in his study reported higher serum COMP level in Caucasian men than Caucasian women which was statistically significant.[25] Verma in his study also found gender difference with 52% higher serum COMP in males as compared to that of females.[26] On the contrary, some studies have also reported no gender difference. Clark in his study reported no gender bias with respect to serum COMP levels in the males and females [6] and Jordan also found no significant difference in serum COMP values in African American men and women.[25] More physical activity by the males in the local population, because they are the main bread winners may translate into more load on male knees with increased cartilage degradation and consequent higher serum COMP levels. There may also be some contributory hormonal and racial factors which we are not able to explain from this study.
There was a good positive correlation between serum COMP and age in both groups which was statistically significant r=.546 (p=.001) in group 1 and r=.624(p=.001) in group 2. Good positive correlation has also been reported in other studies. Verma and Jordan reported in their studies that serum COMP values steadily increased with age.[25,26] Age is a risk factor for KOA. With increasing age, there is increase in the cartilage degradation of the joints with consequent increase in serum COMP levels. Age has been negatively associated with knee cartilage thickness and positively associated with knee cartilage defect.[27] knee cartilage defects are positively associated with KOA and quite common in older subjects, even in those without radiological evidence of KOA.[28]
There is also a significant difference in the values of the reference intervals between the different gender age groups reflecting the gender bias and positive correlation between serum COMP and age. Establishing reference intervals for the different gender age groups makes the values more clinically precise and useful for interpreting laboratory serum COMP results. Levels of serum COMP above the upper reference limits for a particular gender and age group may indicate early KOA in the absence of radiological features. Pre-radiographic diagnosis may have many advantages including early commencement of treatment with better prospects of response to treatment and prognosis. Also early commencement of preventive measures in those whose serum COMP is above the upper reference limit may prevent development of KOA. Radiographic diagnosis of KOA signifies advanced disease which is not curable. Those with serum COMP at or above the upper limit of the reference interval may be at risk of developing and having rapidly progressing KOA as reported by Verma PI and Dalal K.[26]
Presently, to the knowledge of the researchers, the diagnostic ranges of reference intervals for serum COMP have not been defined for the Nigerian population and there are no reference intervals for the Nigerian population in the literature. Further research is necessary to validate the results of this study so that formal reference intervals can be established specifically for the Nigerian population as there may be a racial influence as suggested by the results from Jordan’s study.[23] Establishing reference intervals for the Nigerian population may provide the orthopaedic surgeons, rheumatologists, physiotherapists and others involved in the management of patients with KOA a promising tool to enable them diagnose KOA serologically early in the pre radiographic stage at a lower cost than the more expensive modalities of investigation like MRI and ultrasound.[29,30] Serum COMP above the upper limit of the reference intervals when other clinical and laboratory findings are suggestive of KOA in the absence of radiological features may be an indication to commence treatment.
Conclusion
Serum COMP is a potential tool for making early diagnosis of KOA but multi-center corroborative studies using large sample sizes will be required before reference intervals can be established for the Nigerian population.
Funding
This research did not receive any specific grant from funding agencies in the public, commercial or not – for – profit sectors.
References
- Saxne T, Heinegard D. Cartilage oligomeric matrix protein: a novel marker of cartilage turnover detectable in synovial fluid and blood. Br J Rheumatol. 1992;31:583-591.
- Heinegård D, Lorenzo P, Saxne T. Matrix Glycoproteins and Proteoglycans in Cartilage. In Harris ED, Budd RC, Firestein GS, Genovese MC, Ruddy S.(Eds) Kelley's Textbook of Rheumatology. 2005, Philadelphia: Elsevier Saunders. pp 48-62.
- Muller G, Michel A, Altenburg E. COMP (cartilage oligomeric matrix protein) is synthesized in ligament, tendon, meniscus, and articular cartilage. Connect Tissue Res. 1998;39(4):233–244.
- Neidhart M, Hauser N, Paulsson M, DiCesare PE, Michel BA, Hauselmann HJ. Small fragments of cartilage oligomeric matrix protein in synovial fluid and serum as markers for cartilage degradation. Br J Rheumatol. 1997;36(11):1151–1160.
- Hunter DJ, Li J, LaValley M et al. Cartilage markers and their association with cartilage loss on magnetic resonance imaging in knee osteoarthritis: the Boston Osteoarthritis Knee Study. Arthritis Research and Therapy. 2007;9(5):R108
- Clark G, Jordan JM, Vilim V et al. Serum cartilage oligomeric matrix protein reflects osteoarthritis presence and severity: the Johnston County Osteoarthritis Project. Arthritis Rheum. 1999;42(11):2356–2364.
- Kraus VB1, Burnett B, Coindreau J et al. OARSI FDA Osteoarthritis Biomarkers Working Group. Application of biomarkers in the development of drugs intended for the treatment of osteoarthritis. Osteoarthritis Cartilage. 2011 May;19(5):515-42.
- Larsson E, Erlandsson Harris H, Larsson A, Mansson B, Saxne T, Klareskog L. Corticosteroid treatment of experimental arthritis retards cartilage destruction as determined by histology and serum COMP. Rheumatology (Oxford). 2004;43:428-434.
- Crnkic M, Mansson B, Larsson L, Geborek P, Heinegard D, Saxne T. Serum cartilage oligomeric matrix protein (COMP) decreases in rheumatoid arthritis patients treated with infliximab or etanercept. Arthritis Res Ther. 2003;5:R181-185.
- Garnero P. New biochemical markers of cartilage turnover in osteoarthritis: recent developments and remaining challenges. International Bone and Mineral Society Bonekey. 2007;4(1):7-18
- Vilim V, Vytasek R, Olejarova M et al. Serum cartilage oligomeric matrix protein reflects the presence of clinically diagnosed synovitis in patients with knee osteoarthritis. Osteoarthritis Cartilage. 2001;9(7):612–618.
- Hoch JM, Mattacola CG, Medina McKeon JM, Howard JS, Lattermann C. Serum cartilage oligomeric matrix protein (sCOMP) is elevated in patients with knee osteoarthritis: a systematic review and meta-analysis. Osteoarthritis Cartilage. 2011 Dec;19(12):1396-404.
- Fernandes FA, Pucinelli MLC, da Silva NP, Feldman D. Serum cartilage oligomeric matrix protein (COMP) levels in knee osteoarthritis in a Brazilian population: clinical and radiological correlation. Scand J Rheumatol. 2007;36(3):211–215.
- Cibere J, Zhang H, Garnero P et al. Association of biomarkers with pre-radiographically defined and radiographically defined knee osteoarthritis in a population-based study. Arthritis Rheum. 2009;60(5):1372–1380.
- Kane ED. Biomarkers aid early detection of joint disease, bone damage. DVM newsmagazine. 2007;24:46–49.
- Petersson IF, Boegard T, Svensson B, Heinegard D, Saxne T. Changes in cartilage and bone metabolism identified by serum markers in early osteoarthritis of the knee joint. Br J Rheumatol 1998;37:46-50.
- Sharif M, Kirwan JR, Elson CJ, Granell R, Clarke S. Suggestion of nonlinear or phasic progression of knee osteoarthritis based on measurements of serum cartilage oligomeric matrix protein levels over five years. Arthritis Rheum 2004;50:2479-2488.
- Garnero P, Rousseau JC, Demas PD. Molecular basis and clinical use of biochemical marker of bone, cartilage, and synovium in joint disease. Arthritis Rheum. 2000;43:953–68.
- Kellgren JH, Lawrence JS. Radiological assessment of osteoarthrosis. Ann Rheum Dis 1957;16:494–502.
- Otterness IG, Swindell AC, Zimmerer RO, Poole AR, Ionescu M, Weiner E. An analysis of 14 molecular markers for monitoring osteoarthritis: segregation of the markers into clusters and distinguishing osteoarthritis at baseline. Osteoarthritis Cartilage. 2000;8(3):180–185.
- Mündermann A, Dyrby CO, Andriacchi TP, King KB. Serum concentration of cartilage oligomeric matrix protein (COMP) is sensitive to physiological cyclic loading in healthy adults. Osteoarthritis and Cartilage. 2005;(1):34-8.
- Andersson MLE, Petersson IF, Karlsson KE, Jonsson EN, Mansson B. Diurnal variation in serum levels of cartilage oligomeric matrix protein in patients with knee osteoarthritis or rheumatoid arthritis. Annals of Rheumatic Diseases. 2006;65(11):1490-1494.
- CLSI. Defining, establishing, and verifying reference intervals in the clinical laboratory; Approved guideline, 3rd edition. CLSI Document EP28-A3c. Wayne, PA; Clinical and Laboratory Standards Institute; 2008.
- Bruyere O, Collette JH, Ethgen O et al. Biochemical markers of bone and cartilage remodelling in prediction of long term progression of knee osteoarthritis. J Rheumatol. 2003;30:1043–1050
- Jordan JM, Luta G, Stabler T et al. Ethnic and sex differences in serum levels of cartilage oligomeric matrix protein: the Johnston County Osteoarthritis Project. Arthritis Rheum. 2003 Mar;48(3):675-81.
- Verma P, Dalal K. Serum cartilage oligomeric matrix protein (COMP) in knee osteoarthritis: a novel diagnostic and prognostic biomarker. J Orthop Res. 2013 Jul;31(7):999-1006.
- Ding C, Cicuttini F, Scott F, Cooley H, Jones G. Association between age and knee structural change: a cross sectional MRI based study. Ann Rheum Dis. 2005 Apr; 64(4): 549–555.
- Joensen AM, Hahn T, Gelineck J, Overvad K, Ingemann-Hansen T. Articular cartilage lesions and anterior knee pain. Scand J Med Sci Sports 2001;11:115–9.
- Kamei G, Sumen Y, Sakaridani K. Evaluation of cartilage defect at medial femoral condyle in early osteoarthritis of the knee. Magn Reson Imaging. 2008;26:567–571
- Lee CL, Huang MH, Chai CY, Chen CH, Su JY, Tien YC. The validity of in vivo ultrasonographic grading of osteoarthritic femoral condylar cartilage: a comparison with in vitro ultrasonographic and histologic gradings. Osteoarthritis Cartilage. 2008;16:352–358.
|
