Effect Of Interleukin-21on The Expression Of Keratin17in Keratinocytes And Its Molecular Mechanism

Psoriasis is a common, inflammatory skin disorder with high-relapse rate,and its pathogenesis is still not clear. At present, it is viewed as an auto-immunedisorder mainly mediated by Th cells, in which many inflammatory cells andcytokines are involved in.The keratin spectrum of keratinocyte (KC) in psoriatic lesions is changed,including significant elevation of keratin17(K17). K17,“psoriasis-proliferationrelated keratin”, is no or lowly expressed in normal skin but is highly expressedin psoriatic plaques, with its expression level correlating with the severity of thedisease to a certain extent. Preliminary studies of our group found that theremight be a role loop of “auto-reactive T cells/cytokines/K17” in psoriasis. K17can activate auto-reactive T cells to secrete some psoriasis-related cytokines suchas IFN-γ, IL-17et al, and these cytokines can up-regulate the expression of K17in KC, further activating auto-reactive T cells. In this reason, a malignant looptakes part in the occurrence and development of psoriasis.Interleukin-21(IL-21) comes mainly from CD4+T cells and has extensivebiological functions in many inflammatory and auto-immune diseases. IL-21 mRNA and protein levels in psoriatic plaques are significantly higher thanhealthy controls and it can stimulate the KCs of mice and normal skin in psoriasispatients to proliferate accompanied by the infiltration of inflammatory cells,consistent to the pathological changes in psoriatic plaques. Therefore it indicatesthat IL-21may take part in the occurrence and development of psoriasis. In thatway, whether IL-21are relevant with disease severity of psoriasis and IL-21haseffect on the expression of K17or not are included in the experiments.Objectives: To detect serum IL-21levels in patients with psoriasis vulgarisand investigate the correlation between serum IL-21levels and Psoriasis Area andSeverity Index(PASI) scores. Explore effect of IL-21on the expression of K17inKC and its molecular mechanism.Methods: Serum IL-21levels were measured by Enzyme-LinkedImmunosorbent Assay (ELISA). Statistical methods were used to analyse thedifferences between psoriasis patients and healthy controls and the correlationbetween IL-21and PASI scores in psoriasis patients.We used IL-21(0ng/mL,12.5ng/mL,25ng/mL,50ng/mL and100ng/mL)and IFN-γ (250U/ml) to stimulate KCs for24h and48h, and extracted RNA andprotein. Then we choosed Real-time PCR, Western blot and immunofluorescencestaining to detect K17mRNA and protein levels and analyzed the relationshipbetween them. In the meanwhile, choose a suitable concentration of IL-21tocontinue subsequent experiments in molecular pathway.In the next experiment, We used IL-21to stimulate KCs for0min,10min,20min,30min respectively, and choosed Real-time PCR and immunofluorescencestaining to detect the tyrosine phosphorylation of signal pathway molecules inKC. In order to validate whether activated signal pathways were related to thehigh-expression of K17in KC, we used molecular antagonists to inhibit KCs for 2hours, then stimulated KCs and choosed Western blot, Real-time PCR andimmunofluorescence staining to detect whether molecular antagonists couldinhibit the expression of K17in KC.Results: The levels of serum IL-21in psoriasis vulgaris were higher thanhealthy controls (P＜0.01), and positively correlated with PASI scores (r=0.471,P＜0.01). After stimulation by IL-21of12.5ng/mL,25ng/mL,50ng/mL and100ng/mL for24h and48h, all groups had high-expression of K17. Compared toblank group, the expressions of K17mRNA and protein in12.5ng/mL and25ng/mL groups were elevated with no statistical differences, but50ng/mL and100ng/mL groups had statistical significance (P＜0.05), with a dose-dependentrelationship. Then we used50ng/mL IL-21to stimulate KCs for10min,20min,30min, and found that Signal transducer and activator of transcription3(STAT3)and extracellular signal-regulated kinases1and2(ERK1/2) were activated. Atlast, we used molecular antagonists Piceatannol (STAT3) and PD-98059(ERK1/2) to pretreat KCs fou2hours, then used50ng/mL IL-21to stimulate itand found that mRNA and protein levels of K17were inhibited.Conclusions: This study demonstrates that the levels of serum IL-21inpatients with psoriasis vulgaris are higher than healthy controls, and positivelycorrelates with PASI scores. It indicates that IL-21has a close relationship withpsoriasis. IL-21can up-regulate the expression of K17in KC in a dose-dependentmode, and its specific regulation mechanism is by the activation of STAT3andERK1/2. Our experimental results prove that IL-21takes part in the pathogenesisof psoriasis on the level of keratin. It enriches and supplements the loophypothesis, and provides a new strategy toward pathogenesis and remedy inpsoriasis.