Materials
Activity assay kits of GSH-Px (D-89075), SOD (K335-100) and Catalase (D-89075) from Biocore Diagnosik Ulm GmbH Co (Germany), BioVision Co (USA) and Biocore Diagnosik Ulm GmbH Co (Germany), respectively. Quantity assay kits of ox.LDL (10-1143-01), IL1β (850.006.048) and IL6 (860.020.048, 860.020.096, 860.020.192) from Mercodia Co (Sweden), DIACLONE Co (France), DIACLONE Co (France) and DIACLONE Co (France), respectively. Quantity assay kits of MDA (10009055) and TNF-α (DTA00C) from Cayman Co (USA) and R&D SYSTEMS Co (USA), respectively.
Methods
Samples
10 ml of venous blood sample of each patient was drawn in the hospital lab. Then, each sample was mixed with anticoagulant ethylenediaminetetraacetic acid. The blood sample was centrifuged at 2500 × g for 10 min, and the serum was then separated and aliquoted into tubes. Samples were stored at −70°C until assayed.
Measuring markers
MDA
The serum level of MDA was assessed by Colorimetric method and Cayman kit, USA). Microplate reader instrument (Sunrise model) (Teacan Co, Austria) was applied to determine mentioned parameter.
ox.LDL
Quantity level of ox.LDL in rat’s serum was determined by ELISA method and Mercodia kit (Sweden). Mindray ELISA reader apparatus (MR-96A model, Germany) was used to measure related amounts.
CAT
Catalase activity of patient’s serum was assessed by activity assay kit of Biocore Diagnostik Ulm GmbH Co (Germany) and colorimetric technique. Microplate reader instrument (Sunrise model) (Teacan Co, Austria) was applied to determine mentioned parameter.
GSH-Px
Glutathione peroxidase activity of patient’s serum was determined by activity assay kit of Biocore Diagnostik Ulm GmbH Co (Germany) and colorimetric technique. Microplate reader instrument (Sunrise model) (Teacan Co, Austria) was applied to determine mentioned parameter.
SOD
Superoxide Dismutase activity of patient’s serum was determined by activity assay kit of BioVision Co (USA) and colorimetric technique. Microplate reader instrument (Sunrise model) (Teacan Co, Austria) was applied to determine mentioned parameter.
Interleukins
The serum levels of IL1β and IL6 were assessed by ELISA method and immunoenzymometric assay (DIACLONE kit, France). All measurements were done using a Mindray ELISA reader instrument (MR-96A model, Germany).
TNF-α
The serum levels of TNF-α were assessed by ELISA method and immunoenzymometric assay (R&D SYSTEMS kit, USA). All measurements were done using a Mindray ELISA reader instrument (MR-96A model, Germany).
Study population
43 RA patients who referred to the rheumatology clinic of Imam Khomeini Hospital complex in 2017 were chosen as a case group. All of RA patients were diagnosed based on American College of Rheumatology criteria [10]. 43 healthy people who were matched with the case group were selected as a control group. The exclusion criteria were smoking (smoking longer than past 5 years), alcohol intake (drinking longer than past 12 months), using narcotics (any type of narcotic drugs at any frequency anytime), hypertension, diabetes mellitus, hypothyroidism, hyperthyroidism, and any other form of inflammatory arthritis except RA, receiving alternative and complementary treatments such as Ayurveda, homeopathy, and siddha.
This study was approved by ethical committee of Tehran University of Medical sciences and all participants completed the informed consent form.
Data collection
An expert rheumatologist discussed the study to the patients and questionnaires including demographic factors, disease activity Index (DAS28) and Global Health assessment were completed. Both cases and control groups were referred to the central endocrinology lab located in Imam Khomeini Hospital to evaluate the levels of oxidants, antioxidants, and other inflammatory factors. We used DAS28 to evaluate RA activity. Patients with DAS28<2.6 were considered in remission and above it were grouped as active RA [4].
Data analysis
The data was analyzed by SPSS, version 15.0. Values are expressed as the mean and standard deviation. The differences were assessed