Expression Of Sirt1 In CD4~+T Cell And The Effect Of Resveratrol On Percentage Of CD4~+T Cell Subgroups And Their Cytokines And On Fuction Of CD4~+T Cell And PBMC Isolated From Peripheral Blood Of Patients With Systemic Lupus Erythematosus

BackgroundSystemic lupus erythematosus(SLE), clinically characterized by multisystem involvement and various antoantibodies positive in serum, is a chronic, immue-complex mediated diffuse connective tissue disease. Abnormal proliferation and hyperactivation of CD4+T cell is crucial to induce autoreactive B cell hyperplasia and autoantibodies production. According to the differentiation and functional features, CD4+T cell can be divided into Th1,Th2,Th17 and Treg cell subgroups. Each cell subgroup secretes characteristic cytokines, such as Th1,Th2 and Th17 cells secrete IFN-gamma,IL-4/ IL-10 and IL-17 respectively. While Treg cell is in charge of immunregulation. The imbalances among four subgroups and among the cytokines secreted by four subgroups play a central role in the pathogenesis of SLE.Silent information regulator 2(Sir2), which is remarkably conserved throughout the evolution,was first recognized as a gene that is required to maintain cell-mating type in Saccharomyces cerevisiae. The ortholog of the Sir2 is collectively called Sirtuin family. Seven homologs(Sirtl-7) in mammals have been identified.Among the seven members, Sirtl is the closest homolog to Sir2 and is best characterized so far. Sirtl, a member of classIII histone deacetylase (HDAC), deacetylates a variety of proteins including histones HI, H3, H4 and several other proteins requiring NAD+as cofactor. With the development of epigenetics, DNA hypomethylation status was found in CD4+Tcell from SLE patients. But the view on acetylation status in SLE patients is controversial. It was reported that hyperacetylation is helpful to induce autoantibodies production, while hypoacetylation of H3 and H4 was found in SLE patients and MRL/lpr mouse. So abnormal Sirtl expression may be involved in the pathology of SLE. Sirtl is abundant in the thymus, and Jedon reported that immune complex deposited in kidney and anti-nuclear antibody was found in serum.These evidence support the above-mentioned hypothesis.Resveratrol, originally described as an anti-fungal plant, is polyphenol stilbene found widely in plants, especially in grape skins, giant knotweed rhizome, and nuts. Resveratrol has many fuctions, such as anti-inflammatory,anti-oxidant effects. Recently, it is found that Resveratrol has immune regulatory effects, which has a lot of attractions.In addition, Resveratrol is a natural effective activator of Sirtl. So it is important to make researchs about the expression of Sirtl in CD4+T cell and the effect of Resveratrol on percentage of CD4+T cell subgroups and their cytokines and on fuction of CD4+T cell and PBMC isolated from peripheral blood of patients with systemic lupus erythematosus. Which may clue a new target therapy.Methods Part one, seventeen patients diagnosed of SLE with available clinical data and nine healthy controls(HC) were recuited. SirtlmRNA was measured and quantified by real-time polymerase chain reaction. Patients were classified into subgroups according to organ involvement and/or SLEDAI scores. Part two, CD4+T cell subgroups and their cytokines were measured in eigh patients and eight controls by FCS. And six active SLE patients and controls were given Resveratrol with different dosages, and the percentage of CD4+T cell subgroups with cytokines in culture supernatants were measured by FCS. Part three, PBMC and CD4+T cell were isolated from 4 patients and 5 controls, certain amount of PBMC and CD4+T cell were stained with CFSE under sterile conditions and cocultured with Resveratrol with different dosages in the presence of stimulator (anti-CD3/CD28) for 96 hours. The status of proliferation was detected by flow cytometer. In addition, CD25 as a T cell activation marker was also detected in 5 patients and 5 controls, but the co-culture time of Resveratrol with stimulator (anti-CD3/CD28) was 24 hours.Results Part one, There was no significant difference of SirtlmRNA expression in CD4+T cell between SLE patients and HC. Active SLE paients had significantly decreased SirtlmRNA expression than inactive and HC. SLE paients with butterfly erythema had increased SirtlmRNA expression than paients without butterfly erythema. No significant difference was found between patients with renal involvement and those without. SirtlmRNA expression correlated positively with C3 level. Part two, SLE patients had higher percentages of Th1,Th2,Th17 subgroups and higher levels of IFN-γ,IL-4,IL-10,IL-17 than that in HC. There is no significant difference of Treg cell. There is no obvious effect of Resveratrol on percentages of Thl,Th2,Treg cell and levels of IFN-γ,IL-4. IL-10 in culture supernatant can be decreased sharply with Resveratrol 40μM. Percentage of Th17 cell and level of IL-17 in culture supernatant can be decreased with Resveratrol 40μM. Part three, The proliferation of activated PBMC and CD4+T cell isolated from SLE patients and HC was inhibited by Resveratrol. And the inhibition capacity is dependent on the dosage. In addition, Resveratrol can decrease the percentage of activated CD4+T cell.Conclusion Our findings revealed that Sirtl may be involved in the pathogenesis of SLE. It may be a new therapy target. Resveratrol can be an attractive therapy through decreasing the percentage of Th17 cell,the level of IL-l0,IL-17 and inhibiting proliferation and activation of CD4+T cell.