Home About us Editorial board Search Ahead of print Current issue Archives Submit article Instructions Subscribe Contacts Login 


 
 Table of Contents  
ORIGINAL ARTICLE
Year : 2016  |  Volume : 29  |  Issue : 4  |  Page : 1130-1135

Serum undercarboxylated osteocalcin and bone mineral density in premenopausal patients with rheumatoid arthritis


1 Department of Rheumatology, Physical Medicine and Rehabilitation, Faculty of Medicine, Zagazig University, Zagazig, Egypt
2 Department of Physical Medicine and Rehabilitation, Faculty of Medicine, Menoufia University, Menoufia, Egypt
3 Department of Clinical Pathology, Faculty of Medicine, Menoufia University, Menoufia, Egypt

Date of Submission24-Apr-2015
Date of Acceptance08-Jun-2015
Date of Web Publication21-Mar-2017

Correspondence Address:
Eman A Galbat
Department of Physical Medicine and Rehabilitation, Menoufia University, Al Shohdaa, Menoufia, 32958
Egypt
Login to access the Email id

Source of Support: None, Conflict of Interest: None


DOI: 10.4103/1110-2098.202498

Rights and Permissions
  Abstract 

Objectives
The aim of this study was to measure serum undercarboxylated osteocalcin (ucOC) and bone mineral density (BMD) in premenopausal patients with rheumatoid arthritis (RA) and their correlation with each other and with disease activity.
Background
Osteocalcin is a molecule synthesized as a prepro-osteocalcin. After proteolytic cleavage and vitamin K-dependent carboxylation processes, it is released and circulates in the blood as carboxylated osteocalcin or ucOC.
Participants and methods
In total, 60 premenopausal patients with RA and 30 healthy premenopausal controls with matched age were included in this study. All patients and controls were subjected to demographic data, clinical examination, and laboratory investigations including serum level of ucOC, disease activity assessment by DAS-28 score, and BMD measurement in the lumbar spine L2–L4, hip, and distal radius by dual-energy X-ray absorptiometry equipment.
Results; The level of ucOC was significantly higher in patients of RA than controls (P < 0.001). BMD in patients was found to be significantly lower than in controls in spine, femoral neck, and distal radius areas. The most frequent osteoporotic site according to Z-score was the spine (16.7%) followed by the femoral neck (8.3%) and then the distal radius (6.7%). Moreover, the commonest osteopenic site according to Z-score of -1 or less was the spine (31.7%) followed by the femoral neck (21.7%) and then (16.7%) the distal radius. This work showed that ucOC level was found to be high in premenopausal patients with RA with higher DAS values than those with lower DAS value (P < 0.001). In this work, BMD measured by dual-energy X-ray absorptiometry scan was found to be lower with higher DAS values, and vice versa.
Conclusion
Serum level of ucOC (which is a mirror of vitamin K deficiency) was found to be higher in premenopausal patients with RA than controls and correlated positively with disease activity and inversely with BMD measurement.

Keywords: bone mineral density, disease activity, rheumatoid arthritis, undercarboxylated osteocalcin


How to cite this article:
Al Hewala AI, Soliman SG, Esaily HA, Abo Ela DH, Galbat EA. Serum undercarboxylated osteocalcin and bone mineral density in premenopausal patients with rheumatoid arthritis. Menoufia Med J 2016;29:1130-5

How to cite this URL:
Al Hewala AI, Soliman SG, Esaily HA, Abo Ela DH, Galbat EA. Serum undercarboxylated osteocalcin and bone mineral density in premenopausal patients with rheumatoid arthritis. Menoufia Med J [serial online] 2016 [cited 2020 Jun 1];29:1130-5. Available from: http://www.mmj.eg.net/text.asp?2016/29/4/1130/202498


  Introduction Top


Rheumatoid arthritis (RA) is a chronic inflammatory autoimmune disease that causes chronic inflammation of the synovium with subsequent destruction and deformity of the joints [1].

Osteoporosis is a multifactorial skeletal disease characterized by a reduction in bone mass and deterioration of the microarchitectural structure of bone tissue, with resulting increase in bone fragility and fracture risk [2].

Osteoporosis is more common in patients with RA than in the general population. The prevalence of concurrent osteoporosis is 50%. Osteoporosis can cause pain and loss of height, and it increases the risk of fractures after falling [3].

The chronic synovial inflammation in RA can promote osteoclastogenesis, leading directly to both focal and generalized bone loss and increased risk of fractures [4].

Vitamin K is a cofactor of γ-carboxylase, which converts three glutamic acid (Glu) residues in osteocalcin (OC) to γ-carboxyglutamic acid (Gla) and is thus essential for γ-carboxylation of OC. Without this modification, OC becomes undercarboxylated osteocalcin (ucOC), which lacks structural integrity and the ability to bind to the mineral hydroxyapatite. Vitamin K deficiency impairs γ-carboxylation of OC, resulting in high serum concentrations of ucOC [5].


  Participants and Methods Top


The study was done by Clinical Pathology Department in collaboration with Physical Medicine and Rehabilitation Department.

Participants

Participants were classified into two groups: group I included

60 premenopausal patients diagnosed as having RA according ACR/EULAR 2010 criteria [6], with age greater than 16 years and disease duration of more than 2 years, and group II included

30 age-matched healthy premenopausal female patients as control who were free of earlier fractures and chronic diseases and were not taking medications that influence bone metabolism (e.g., anticonvulsants and thyroxine).

Exclusion criteria

Postmenopausal female patients with RA, male patients with RA, those experiencing serious cardiovascular and/or pulmonary disease, those having an abnormal thyroid function or a serious infection, those with hyperparathyroidism, those with hepatic or renal dysfunction, those taking any drugs or hormones that affect bone metabolism (e.g., sex steroids) were excluded.

Methods

All patients and controls were subjected to the following:

  1. Demographic data recording
  2. Full history taking and thorough clinical examination
  3. Disease activity assessment.


  4. Disease Activity Score (DAS-28) with three variables [erythrocyte sedimentation rate (ESR), the number of swelled joints, and number of tender joints] was used [7].



    The level of disease activity was interpreted as follows:

    1. DAS-28 ≤ 3.2 = low;
    2. 3.2 <DAS-28 ≤ 5.1 = moderate;
    3. DAS-28 > 5.1 = high
    4. DAS < 2.6 = disease being in remission according to the American Rheumatism Association (ARA) criteria [8].


  5. Laboratory investigations


  6. ESR [9] was obtained by Westergren method, rheumatoid factor and C-reactive protein levels by latex agglutination [10],[11], and serum levels of ucOC and anti-CCP level were measured by using Cobas Elecsys 2010 (Roche Diagnostics GmbH, Mannheim, Germany).

    Serum levels of undercarboxylated osteocalcin

    Serum levels of ucOC were obtained using the sandwich principle test (enzyme-linked immunosorbent assay). The total duration of the assay was 18 min.

    First incubation

    A 20 µl sample, a biotinylated monoclonal N-MID OC-specific antibody, and a monoclonal N-MID OC-specific antibody labeled with a ruthenium complex [Tris (2,2’-bipyridyl) ruthenium (II)-complex (Ru (bpy)] react to form a sandwich complex.

    Second incubation

    After addition of streptavidin-coated microparticles, the complex becomes bound to the solid phase through interaction of biotin and streptavidin. The reaction mixture is aspirated into the measuring cell where the microparticles are magnetically captured onto the surface of the electrode. Unbound substances are then removed with ProCell/ProCell M. Application of a voltage to the electrode then induces chemiluminescent emission which is measured by a photomultiplier.

    Results are determined using a calibration curve, which is an instrument specifically generated by two-point calibration and a master curve provided through the reagent barcode. Normal level is less than 4.5 ng/ml.

    Sampling

    Under complete aseptic conditions, 5 ml of venous blood was collected. Then, 4 ml of whole blood was added to a sterile plain tube and allowed to clot at 37°C. Serum was separated by centrifugation and used for measuring levels of ucOC, anti-CCP, rheumatoid factor, and C-reactive protein. Moreover, 0.8 ml of whole blood was added to 0.2-ml sodium citrate tube to measure ESR.

  7. Radiographic assessment


Bone mineral density (BMD) was measured in the lumbar spine L2–L4, hip, and distal radius by dual-energy X-ray absorptiometry (DEXA) equipment [12].

BMD was expressed in SD from the mean of healthy age-matched and sex-matched people (the Z-score) and as the number of SD from the mean of healthy, young sex-matched people (the T-score). By the WHO classification and the 2005 International Society for Clinical Densitometry (ISCD) official positions, a T- score of -2.5 or lower in postmenopausal women was classified as osteoporosis, a T-score between >-2.5 and <-1 as osteopenia, and a T-score ≥-1 as normal, whereas osteoporosis according to Z-score of -2.0 or lower in females before menopause was defined as 'below the expected range for age' [13].

Statistical analysis

The data collected were tabulated and analyzed by SPSS (Statistical Package for the Social Science software) statistical package version 20 on IBM compatible Computer (IBM Corp. Released 2013. IBM SPSS Statistics for Windows, Version 22.0. Armonk, NY: IBM Corp., USA). A total of two types of statistics were done: descriptive statistics included percentage (%), range, mean (X), and SD, and analytical statistics included Student's t- test, Mann–Whitney test, and Fisher's exact test. P value greater than 0.05 was considered a nonsignificant difference, P value less than 0.05 as a significant difference, and P value less than 0.001 as a highly significant difference.


  Results Top


In this work, the level of ucOC was significantly higher in patients with RA than in controls (P < 0.001, [Table 1]) and higher in patients with osteoporosis than those with osteopenia and those with normal BMD ([Table 2]). BMD in patients was found to be significantly lower than controls in all sites (spine, femoral neck, and distal radius areas). The most frequent osteoporotic site according to Z-score was the spine (16.7%) followed by the femoral neck (8.3%) and then the distal radius (6.7%). According to T-score, the commonest osteoporotic site was the spine (13.3%), whereas the commonest osteopenic site according to Z-score of -1 or less was the spine (31.7%) followed by the femoral neck (21.7%) and then the distal radius (16.7%) ([Table 3]). Level of ucOC was found to be positively correlated with DAS score (P < 0.001, [Table 4]). BMD measured by DEXA scan was found to be lower with higher DAS values and vice versa, as shown in [Table 5].
Table 1 A comparison between the studied patient group and controls regarding undercarboxylated osteocalcin (ng/ml)

Click here to view
Table 2 A comparison between female patients with osteoporosis, female patients with osteopenia, and normal bone mineral density patients regarding undercarboxylated osteocalcin (ng/ml)

Click here to view
Table 3 A comparison between the studied patient group and control regarding dual-energy X-ray absorptiometry parameters(n (%))

Click here to view
Table 4 Relationship between disease activity and undercarboxylated osteocalcin level (ng/ml) among studied group of patients (n=60)

Click here to view
Table 5 Relationship between disease activity and dual-energy X-ray absorptiometry parameters among studied group of patients (n=60)

Click here to view



  Discussion Top


RA is a chronic inflammatory autoimmune disease that causes chronic inflammation of the synovium with subsequent destruction and deformity of the joints. Osteoporosis is more common in patients with RA than in the general population. Vitamin K is a cofactor of γ-carboxylase, which converts three glutamic acid (Glu) residues in OC to γ-carboxyglutamic acid (Gla) and is thus essential for γ-carboxylation of OC. Without this modification, OC becomes ucOC, which lacks structural integrity and the ability to bind to the mineral hydroxyapatite, and secreted OC can no longer be carboxylated. Vitamin K deficiency impairs γ-carboxylation of OC, resulting in high serum concentrations of ucOC.

This study revealed that the level of ucOC was significantly higher in patients with RA than in controls. This is in accordance with Sakai et al. [14] who showed that serum levels of ucOC were increased in higher state of bone turnover as indicated by biochemical markers, and vice versa.

No other searchers discussed ucOC in adult patients with RA, although van Summeren et al. [15] found that children with JIA (Juvenile Idiopathic Arthritis) have a high ratio of ucOC/carboxylated osteocalcin, indicating low vitamin K status, and associated with low bone ultrasound parameters, whereas children with a high vitamin K status had markedly higher bone properties.

Moreover, Iwamoto et al. [16] found that serum ucOC is an index of vitamin K nutritional status in treatment-naïve postmenopausal women with osteoporosis and concluded that high level of ucOC is a risk factor for osteoporotic fracture, which is decreased by vitamin K intake.

In addition, in this work, ucOC level was found to be positively correlated with disease activity in premenopausal patients with RA.

To our knowledge, no other studies discuss the relationship between ucOC in premenopausal patients with RA and DAS score.

In this work, BMD in patients was found to be significantly lower than in controls in the spine, the femoral neck, and the distal radius areas. The most frequent osteoporotic site according to Z-score was the spine (16.7%) followed by the femoral neck (8.3%) and then the distal radius (6.7%), and the commonest osteopenic site according to Z-score of -1 or less was spine (31.7%) followed by the femoral neck (21.7%) and then the distal radius (16.7%).

This is in accordance with Lee et al. [17], who found that the prevalence of osteoporosis in patients with RA was 1.9 times higher than in healthy subjects.

Moreover, Lodder and colleagues found that the frequency of osteoporosis (T-score≤−2.5 SD) is 12.6 and 6.5% in the spine and the femoral neck, respectively. Reduced bone mass or osteopenia occurred in the spine and the femoral neck at 20.7 and 18.9%, respectively [18].

Laan and colleagues examined BMD in 97 patients with recent-onset RA and a mean disease duration of 30 months. Low bone mass (Z-score≤-1.0) was found in 32.0% of the patients in the lumbar spine and in 24.2% in the hip. They confirm that in comparison with healthy age-matched and sex-matched controls, low bone mass occurs more frequently in patients with RA [19].

Another research by Haugeberg and colleagues studied a representative sample of 394 female patients with RA with a mean disease duration of 13 years, using both T- scores and Z- scores. The prevalence of osteoporosis (T-score≤-2.5) was 16.8% in the spine, 14.7% in the femoral neck, and 14.7% in the total hip. The prevalence of reduced bone mass (Z-score≤-1.0) was greater than expected (15.9%) in the femoral neck (27.6%), the total hip (31.6%), and the spine (19.6%) [20].

However, Hamalainen et al. [21] concluded that according to bone mineral concentration, premenopausal women with RA both with and without prednisolone treatment and controls lost bone statistically similarly.

Hui et al. [22] had followed up 130 healthy premenopausal white women for 1–9 years and found a significant decrease in femoral neck BMD over time.

In our work, BMD measured by DEXA scan was found to be lower with higher DAS values, and vice versa.

This is in accordance with studies by Lodder et al. [18], Shenstone et al. [23], and Gough et al. [24]; their studies concluded that patients with active disease lost more bone at the spine and hip than patients with inactive disease.

However, Hamalainen et al. [21] conducted their study on premenopausal women and found that women in the RA with PRED group had lower BMD values than those in the RA without PRED group at the start of follow-up, in both lumbar spine and in femoral neck; however, finally in the premenopausal women, bone status is stable or bone loss minimal in the lumbar spine, whereas some bone loss takes place in the femoral neck.


  Conclusion Top


From the present study, we concluded that patients with RA are more susceptible to develop generalized osteoporosis than other population even if they are in premenopausal phase. Serum level of ucOC is an indicator of BMD in premenopausal patients with RA. Patients with high serum level of ucOC are more susceptible to osteoporosis than other patients are. Patients with active RA disease are associated with high level of ucOC and lower BMD values. The commonest sites of osteoporosis in premenopausal patients with RA were in lumbar spine (16.7%) followed by hip (8.3%) and the distal radius (6.7%) according to Z-score.

Financial support and sponsorship

Nil.

Conflicts of interest

There are no conflicts of interest.

 
  References Top

1.
Chung SJ, Kwon YJ, Park MC, Park YB, Lee SK. The correlation between increased serum concentrations of interleukin-6 family cytokines and disease activity in rheumatoid arthritis patients. Yonsei Med J 2011; 52:113–120.  Back to cited text no. 1
    
2.
Role of family medicine in the early detection and management of osteoporosis. Mohammed Omaima Abo Elfatth, Shaheen Hala Mohammed El Moselhey, Kaoud Yasmin El Gamil 2014: 27:833-839.  Back to cited text no. 2
    
3.
Wijbrandts CA, Klaasen R, Dijkgraaf MG, Gerlag DM, van Eck-Smit BL, Tak PP. Bone mineral density in rheumatoid arthritis patients 1 year after adalimumab therapy: arrest of bone loss. Ann Rheum Dis 2009; 68:373–376.  Back to cited text no. 3
    
4.
Aizer J, Reed G, Onofrei A, Harisson MJ. Predictors of bone density testing in patients with rheumatoid arthritis. Rheumatol Int 2009; 29:897–905.  Back to cited text no. 4
    
5.
Koshihara Y, Hoshi K. Vitamin K2 enhances osteocalcin accumulation in the extracellular matrix of human osteoblasts in vitro. J Bone Miner Res 1997; 12:431–438.  Back to cited text no. 5
    
6.
Funovits J, Aletaha D, Bykerk V, Combe B, Dougados M, EmeryP, et al. American College of Rheumatology/European League Against Rheumatism classification criteria for rheumatoid arthritis: methodological report phase I. Ann Rheum Dis 2010; 69:1589–1595.  Back to cited text no. 6
    
7.
Prevoo ML, van 't Hof MA, Kuper HH, van Leeuwen MA, van de Putte LB, van Riel PL. Modified disease activity scores that include twenty-eight-joint counts. Development and validation in a prospective longitudinal study of patients with rheumatoid arthritis. Arthritis Rheum 1995; 38:44–48.  Back to cited text no. 7
    
8.
Fransen J, Riel PL. The disease activity score and EULAR response criteria. Clin Exp Rheum 2005; 23:93–109.  Back to cited text no. 8
    
9.
Westergren A. Diagnostic tests: the erythrocyte sedimentation rate range and limitations of the technique. Triangle 1957; 3:20–25.  Back to cited text no. 9
    
10.
Nikolaisen C, Rekvig OP, Nossent HC. Rheumatoid factor by laser nephelometry and Waaler-Rose assay: prognostic value in patients with recent-onset rheumatoid arthritis. Scand J Rheumatol 2005; 34:269–276.  Back to cited text no. 10
    
11.
Dominici R, Luraschi P, Franzini C. Measurement of C-reactive protein: two high sensitivity methods compared. J Clin Lab Anal 2004; 18:280–284.  Back to cited text no. 11
    
12.
Dacie GV, Lwis SM Practical hematology, 6th ed. London: Churchill Livingstone; 1985. 33–35.  Back to cited text no. 12
    
13.
Leslie WD, Adler RA, El-Hajj Fuleihan G, Hodsman AB, Kendler DL, McClung M, et al. Application of the 1994 WHO classification to populations other than postmenopausal Caucasian women: the 2005 ISCD Official Positions. J Clin Densitom 2006; 9:22–30.  Back to cited text no. 13
    
14.
Sakai Yoshitada Hashiramoto, Akira Hashimoto, Teppei Kawasaki, Yoshiko Tanaka, Chihiro Shiozawa, Shunichi Kurosaka Masahiro. et al. The serum level of undercarboxylated osteocalcin (ucOC) in patients with rheumatoid arthritis: a high dose of predonisolone decrease the serum level of ucOC [abstract]. Arthritis Rheum 2010; 62 (Suppl 10):983.  Back to cited text no. 14
    
15.
MJH van Summeren, Vermeer C, Engelbert RHH, Schurgers LJ, Takken T, et al. Extremes in vitamin K status of bone are related to bone ultrasound properties in children with juvenile idiopathic arthritis. Clin Exp Rheumatol 2008; 26:484–491.  Back to cited text no. 15
    
16.
Iwamoto J, Takada T, Sato Y. Vitamin K nutritional status and undercarboxylated osteocalcin in postmenopausal osteoporotic women treated with bisphosphonates. Asia Pac J ClinNutr 2014; 23:256–262.  Back to cited text no. 16
    
17.
Lee SG, Park YE, Park SH, Kim TK, Choi HJ, Lee SJ, et al. Increased frequency of osteoporosis and BMD below the expected range for age among South Korean women with rheumatoid arthritis. Int J Rheum Dis 2012; 15:289–296.  Back to cited text no. 17
    
18.
Lodder MC, Haugeberg G, Lems WF, Uhlig T, Orstavik RE, Kostense P, et al. Radiological damage is associated with low BMD and vertebral deformities in rheumatoid arthritis. Accepted 18 January 2004 The Oslo-Truro-Amsterdam (OSTRA) collaborative study. Arthritis Rheum Arthritis Care Res 2003; 49:209–215.  Back to cited text no. 18
    
19.
Laan RF, Buijs WC, Verbeek AL, Draad MP, Corstens FH, van de Putte LB, et al. Bone mineral density in patients with recent onset rheumatoid arthritis: influence of disease activity and functional capacity. Ann Rheum Dis 1993; 52:21–26.  Back to cited text no. 19
    
20.
Haugeberg G, Uhlig T, Falch JA, Halse JI, Kvien TK. Bone mineral density and frequency of osteoporosis in female patients with rheumatoid arthritis: results from 394 patients in the Oslo County Rheumatoid Arthritis register. Arthritis Rheum 2000; 43:522–530.  Back to cited text no. 20
    
21.
Hamalainen H, Kaarela K, Kroger H, Kauppi M, Järvenpää S, Hakala M, et al. Changes in bone mineral density in premenopausal women with rheumatoid arthritis during a two-year follow-up. Joint Bone Spine 2007; 74:482–487.  Back to cited text no. 21
    
22.
Hui SL, Perkins AJ, Zhou L, Longcope C, Econs MJ. Bone loss at the femoral neck in premenopausal white women: effects of weight change and sex-hormone levels. J Clin Endocrinol Metab 2002; 87:1539–1543.  Back to cited text no. 22
    
23.
Shenstone BD, Mahmoud A, Woodward R, Elvins D, Palmer R, Ring EF, et al. Longitudinal bone mineral density changes in early rheumatoid arthritis. Br J Rheumatol 1994; 33:541–545.  Back to cited text no. 23
    
24.
Gough AK, Lilley J, Eyre S, Holder RL, Emery P. Generalised bone loss in patients with early rheumatoid arthritis. Lancet 1994; 344:23–27.  Back to cited text no. 24
    



 
 
    Tables

  [Table 1], [Table 2], [Table 3], [Table 4], [Table 5]



 

Top
 
 
  Search
 
Similar in PUBMED
   Search Pubmed for
   Search in Google Scholar for
 Related articles
Access Statistics
Email Alert *
Add to My List *
* Registration required (free)

 
  In this article
Abstract
Introduction
Participants and...
Results
Discussion
Conclusion
References
Article Tables

 Article Access Statistics
    Viewed600    
    Printed3    
    Emailed0    
    PDF Downloaded49    
    Comments [Add]    

Recommend this journal


[TAG2]
[TAG3]
[TAG4]