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Abstract Number: 2087

Regulatory Effect of the Combination of Methotrexate and 1,25-Dihydroxyvitamin D3 On the Balance of Treg and Th17 in Collagen-Induced Arthritis

Jing Luo, Department of Rheumatoloy, the Second Hospital of Shanxi Medical University, Taiyuan, China

Meeting: 2012 ACR/ARHP Annual Meeting

Keywords: Type II collagen

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Session Information

Title: Rheumatoid Arthritis: Animal Models

Session Type: Abstract Submissions (ACR)

Background/Purpose: 1,25-dihydroxyvitamin D3 (1,25-(OH)2D3) is the physiologically active metabolite of vitamin D, and it may modulate inflammatory response, cell maturation and cell differentiation. More recently, animal and human studies have suggested that vitamin D is a potential modifier of autoimmune diseases such as rheumatoid arthritis (RA), but the mechanism is not yet clear.In this study, we aimed to determine the effect of methotrexate (MTX) and 1,25-(OH)2D3, used alone or in combination, in the balance of CD4(+) CD25(+) Tregs and Th17, in a rat model of collagen induced arthritis (CIA).

 Methods: Arthritis was induced in 50 female Sprague-Dawley rats. After the clinical onset of CIA, rats were assigned to treatment with MTX (1 mg/kg/week), 1,25-(OH)2D3 (5 mg/kg twice weekly), both treatments at the same regimens, or vehicle. Arthritis score and paw thickness were recorded twice weekly.At termination of the experiment (day 56), bone mineral density (BMD) was analyzed by densitometry, and hind paws were removed for radiographic, histological and immunohistochemical analysis. mRNA expression of transcription factor and proinflammatory mediators was determined by reverse transcriptase–polymerase chain reaction (RT-PCR), and purified T cell proliferation was assessed using [3H]-thymidine incorporated assay, and T-cell phenotypes and activation were assessed by fluorescence-activated cell sorting analysis and T helper (Th) 17/Th1/Th2/Tregs type cytokine production from supernatants from spleen, lymph node and paw cultures was examined by ELISA.

Results: The combination of 1,25-(OH)2D3 with MTX was more effective than MTX alone for reducing the incidence and severity of CIA, and inhibited the production of interleukin (IL)-1β, tumor necrosis factor (TNF)-α and IL-6, IL-2, interferon (IFN)-γ, and matrix metalloproteinases (MMP)-3 and up-regulated anti-inflammatory cytokines IL-4, IL-10. Cytokine analysis indicated that the combination of 1,25-(OH)2D3 with MTX not only modulated the T-helper cell balance from Th1 to Th2 effector function but also associated with the upregulation of CD4(+) CD25(+) Tregs, downregulation of Th17 differentiation. Correspondingly, the mRNA expression of IL-17A and RORrt (a specific transcription factor for Th17) was also reduced.

Conclusion: A combination of MTX and 1,25-(OH)2D3 had beneficial effects on CIA by regulating the balance of CD4(+) CD25(+) Tregs and Th17. These two different mechanisms of action provide support for the use of a combination of these two drugs to improve the prevention of structural joint damage in RA.


Disclosure:

J. Luo,
None;

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