Sirt2Suppresses Inflammatory Responses In Collagen-induced Arthritis

Arthritisis a common autoimmune disease that is associated with progressive disability, systemic complications and early death. However, the underling mechanisms of arthritis are still unclear. Usually we think their own immune response, metabolic disorders, inflammation, trauma, infection, and degenerative diseases such as arthritis is the main factors of complex etiology. Arthritis sort is more, the treatment methods are also different.Treatments including various types of medications, assistive devices, surgery and physical therapy. Adherence to non-steroidalanti-inflammatorydrugs and slow acting anti-rheumatic drugs therapy is beneficial in disease controlling, but it can’t change the disease progression or cure the disease. The surgical treatment methods such as joint replacement, orthopaedic, fusion can also reduce pain while control the disease worsens. In addition, the appropriate physical therapy for arthritis patients recovery also has important significance.Sirtuins are a NAD+-dependent class III deacetylase family, and regulate cellular stress, inflammation, genomic stability, carcinogenesis, and energy metabolism. Among the sirtuin family members, Sirtl and Sirt6are critically involved in the development of arthritis. It remains unknown whether other sirtuin family members participate in arthritis. Here in this study, we demonstrate that Sirt2inhibits collagen-induced arthritis (CIA) using in vivo and in vitro evidence.1Sirt2-KO affect the development of collagen induced arthritis(CIA) in rats.Objective:To probe whether Sirt2-KO affect the development of collagen induced arthritis(CIA) in rats.Methods:DBA/1mice were immunized intradermally with type II collagen(CII)as CIA animal model. Joint tissues were prepared from normal or CIA mice and subjected to western blot and RT-QPCR analysis. The statistical result of Sirt2vs. Gapdh radio was showed RT-QPCR showing the relative mRNA levels of Sirt2in normal and CIA mice.WT and Sirt2-KO mice were immunized intradermally with CII, the cumulative incidence of arthritis should be recorded. Furthermore, arthritis scores and hind paw thickness were evaluated every five days from day20to day45. Results:Sirt2protein and mRNA levels were markedly reduced compared to control mice when immunized with collagen, suggesting that Sirt2may play certain roles in arthritis. when Sirt2was knocked out, mice suffered arthritis as early on day23(two days after secondary immunization), while the WT mice occurred arthritis from day25(Fig.1C). Even though all WT and Sirt2-KO mice developed arthritis, Sirt2-KO group reached a100%cumulative incidence on day34whereas WT group reached a100%cumulative incidence on day37. Those findings indicated that Sirt2-deficiency facilitated the development of collagen-induced arthritis. We found that Sirt2-KO mice developed more severe swelling, erythema and joint rigidity in the hind paws. However, WT arthritic mice showed markedly less bone destruction in the hind paws.Conclusions:Those data indicated that Sirt2considerably participated in arthritis and Sirt2-KO significantly promoted the development of pathologic arthritic disease and increased disease severity.2Sirt2rescue reduces severity of CIA in Sirt2-KO miceObjective:In order to study sirt2rescue whether reduces severity of CIA in Sirt2-KO mice.Method:Sirt2-deficient DBA/1mice were immunized intradermally with CII on day0and given a booster by intradermally injection with CII on day21. The day before secondary immunization, Sirt2-KO mice received a single intraarticular delivery of10ul of adenovirus (3×109pfu) or PBS to each ankle joint. Finally, we performed ELISA assay to detect the cytokine levels in serum.Results:Our rescue results showed that Sirt2re-expression significantly reduced the mean arthritis scores and hind paw thickness in Sirt2-KO mice. In addition, our histopathologic analysis using hematoxylin and eosin staining of ankle joints also revealed a marked reduction of cell infiltration, synovial hyperplasia and bone erosion in mice injected with Ad-Sirt2. Further assessment by micro-CT showed that Sirt2rescue abated bone destruction.Finally, we performed ELISA assay to detect the cytokine levels in serum, and we found that Sirt2rescue significantly reduced the levels of pro-inflammatory cytokines. Conclusions:All of these results demonstrated that Sirt2rescue markedly repressed the development and severity of arthritis in Sirt2-KO mice, and it maybe at the way that reduced the levels of pro-inflammatory cytokines.3Sirt2-deficiency have a impact on pro-inflammatory cytokine levelsObjective:In order to study whether sirt2-deficiency increases have a impact on pro-inflammatory cytokine levels.Method:We measured pro-inflammatory cytokine levels in the serum by ELISA. And we analyzed the mRNA levels of pro-inflammatory cytokines (especially those controlled by NF-κB) in joint tissues by q-PCR.Results:Compared to WT arthritic mice, significantly increase in IL-1β, IL-6, TNF-α, IL-17, IL-33and MCP-1was observed in the serum of Sirt2-KO arthritic mice Pro-inflammatory cytokines such as IL-1β, TNF-α and IL-6play pivotal roles in the pathogenesis of arthritis, and they affect each other. Since arthritis is a systemic inflammatory disease, we wanted to know the inflammatory status of local tissues. Consistent with the serum ELISA results, the relative mRNA levels of IL-1β, TNF-α and MCP-1were markedly increased in Sirt2-KO arthritic mice. In the joint tissues, synovial fibroblasts are key players in joint damage through secreting matrix metalloproteinase (MMPs) and cathepsins.Conclusions:We found that the levels of MMP-9and MMP-13were significantly up-regulated in Sirt2-KO arthritic mice.4Sirt2deacetylates p65and regulates TNF-a-induced expression of inflammatory genesObjective:To verify whether Sirt2genes regulates NF-kB expression of inflammatory.Method:we analyzed protein levels of acetylated p65and total p65of the joint tissues of normal and CIA mice, we further investigated whether Sirt2overexpression could inhibit NF-κB-dependent gene expression in Sirt2-KO mouse FLS by q-PCR.Results:Those findings implicated that Sirt2regulated NF-κB at basal and CIA conditions by regulating acetylation of p65.And our data demonstrated that Sirt2regulated NF-κB by deacetylating p65at lysine310in joint tissues and mouse FLS.Conclusions:Those results demonstrated that Sirt2inhibited TNF-α-induced NF-KB-dependent gene expression.