Our finding suggest that levels of serum homocysteine are higher in SLE patients than in age-matched healthy female. Elevated levels of homocysteine in patients with SLE also reported in several researches [12
]. Homocysteine are regulated by a number of coenzymes and cofactors that required for homocysteine metabolism, such as folic acid, vitamin B6, and vitamin B12. Increased homocysteine levels in SLE patients are caused by a chronic inflammation and abnormal immune response leading to decrease of several vitamins. In SLE patients, the inflammatory process triggered by the presence of autoantibodies that induce further inflammatory reaction resulted in tissue damage [4
]. Activation of immune cells, particularly macrophages, will produce Reactive Oxygen Species (ROS) which cause the oxidation of folic acid, vitamin B6, and B12 [15
]. In addition, the inflammatory process also increase the proliferation of immune cells that would lead to an increased turnover of folic acid, vitamin B6, and B12 [16
]. Both are thought to be the cause of decreased levels of folic acid, vitamin B6, and B12 in inflammatory conditions. In this study, there are no significantly different of folic acid, vitamin B6, vitamin B12 level between SLE patients and control group. But the need of these vitamins is increase in SLE patients because increase of turnover and homocysteine metabolism [17
Several studies showed that high levels of homocystein were associated with decreased bone mass density and early onset of osteoporosis [7
]. Other studies mentioned that high levels of homocystiene and low levels of folic acid were associated with decreased bone mass density, whereas levels of vitamin B6 and vitamin B12 had no significant effect [8
]. Elshorbagy et al. reported that decrease of vitamin B12 and folate levels contribute to increased osteoclast activity which was characterized by high levels of biochemical markers of bone resorption, whereas there was no effect of vitamin B6 [18
]. In our study, a positive and significant correlation was observed between serum homocysteine and bCTx, whereas a negative and significant relationship was found between serum homocysteine and osteocalcin. It means that homocysteine contributes to the increase of bone resorption and decrease of bone formation process.
Recent studies tried to understand the mechanisms regarding the role of homocyteine, folic acid, vitamin B6, and vitamin B12 on bone metabolism. Homocysteine auto-oxidation results in increased production of intracellular ROS and stimulates p38 MAPK activation which influence the differentiation of osteoclast precursor cells [16
]. Homocysteine also induces activation of RANK, a receptor for RANKL, which is a key element in the process of osteoclast differentiation [19
]. There was positive and significant correlation between homocysteine level and MDA in this study. And this MDA level has positive and significant correlation with RANKL. Our result implicated that homocysteine cause an increase production of intracellular ROS that influences osteoclast differentiation through ROS-RANKL pathway.
In this study we also found a negative correlation between homocysteine and osteocalcin. It indicates that the high level of homocysteine cause decrease of osteoblast activity. Kim et al. reported that homocysteine induces apoptosis of human bone marrow stromal cells via caspase-dependent pathway [20
]. The intrinsic apoptotic signals derived from DNA damage and ROS production also induced by accumulation of homocysteine [21
]. In a study conducted by Park et al., reported that homocysteine induces osteoblast cell apoptosis through endoplasmic reticulum stress [22
]. Other studies shown that homocysteine weaken collagen crosslink's and in large amounts it can interfere bone remodeling process [6
The levels of osteocalcin were slightly more in SLE patients. Serum OC is considered a specific marker of osteoblast function, as its levels have been shown to correlate with bone formation rates. However, since it is also released from bone matrix during bone resorption, it reflects the overall turnover of bone and is considered as a bone turnover marker. In a higher activity of bone resorption such as in SLE patient, our body responds by increasing osteoclast activity to balance bone remodeling process [24
]. Some studies report that there were increasing serum osteocalcin levels in postmenopausal women with osteoporosis [25
]. So, this theory is explaining the slightly increasing of OC level in our study. Besides that, OC has a high affinity for calcium and has a compact a helical conformation. The carboxyglutamic acid (Gla) residues of OC are capable of binding to bone matrix hydroxyapatite, thus leading to bone mineralization. Osteoporotic patient may have a decreased rate of bone mineralization due to the reduction in hydroxyapatite crystal formation. In this condition, free OC may be present in the circulation, thus explaining the increased serum OC concentration in osteoporotic patient [28
Elevated level of homocysteine in SLE patients also produce high levels of inflammatory mediators which induces thrombosis of small blood vessels, and a low level of other angiogenic factors. It causes cellular necrosis of a number of osteocytes, followed by localized to bone demineralization. Ongoing inflammation in these localized areas will diminish vascular supply on the osteoblast precursors that available to replace bone demineralization. The longer it will experience a loss of bone fracture in the weakened bone [29
]. That is why osteoporosis occurs earlier in patients with SLE.
Osteoporosis in SLE patient also influenced by several kinds of drugs, such as glucocorticoids and immunosuppressants. All the subjects who contributed in this study are consuming glucocorticoids with average dose 26.87 mg/day. Approximately half of the patients who contribute to this research using immunosuppressant drugs, such as methotrexate, azathioprin, chloroquine, cyclophosphamide and mycophenolat mofetil. Among those immunosuppressants, methotrexate has the influence in decreasing bone mineral density. Study of Raghu Nadhanan et al. found that low dose of methotrexate can increase the risk of osteopenia [30
]. But, other immunosuppressants such as Azathioprin, Chloroquine, Cyclophosphamide and Mycophenolat mofetil have no interference in bone mass density [31
To diagnose osteoporosis we can use several methods, such as biochemical markers of bone turnover including bone formation and bone resorption. Of all the biochemical markers of bone turnover, the ones most commonly used in clinical practice are bone-specific alkaline phosphatase (BSAP), osteocalcin (OC), N-telopeptide of collagen cross-links (NTx), and C-telopeptide of collagen cross-links (CTx) [34
]. Plain radiographic findings can suggest the presence of osteopenia, bone loss, or if a fracture is suspected, but they cannot be used to diagnose osteoporosis. Dual-energy X-ray Absorptiometry (DXA) is currently the criterion standard for the evaluation of bone mineral density (BMD). Compared to imaging techniques, assays for biochemical markers of bone turnover are safe, cheap, easily performed, and can detect early changes in bone metabolism [35
]. In this study, we use bCTX and RANKL as bone resorption marker and Osteocalcin as bone formation marker. But, for further study we suggest to do BMD measurement to confirm the correlation.