The Study On The Potential Roles Of NRIP1 In The Pathologic Mechanism Of Psoriasis

BackgroundPsoriasis is a chronic inflammatory skin disorder affecting approximately 1-3% of the general population. It is characterized by keratinocytes hyper-proliferation and inflammatory cellular infiltrate in both dermis and epidermis. NF-κB is identified as a key mediator in the pathogenesis of psoriasis because of its involvement in inflammatory pathways, in proliferation and apoptosis of various cell types. Nuclear receptor interacting protein 1(NRIP1), also known as Receptor Interacting Protein of 140 kDa (RIP140), functions as a coactivator for NF-κB through direct interaction with Rel A and the transcriptional coactivator CREB-binding protein (CBP) and upregulates the downstream inflammatory gene expression such as TNFα and interleukin-6 in macrophages. NF-κB and the downstream inflammatory factors are involved in the pathogenesis of psoriasis, but the relationship between NRIP1 and psoriasis is still unclear.ObjectiveThe aim of this study is to detect the expression of NRIP1 of skin and PBMCs of psoriasis patients; investigate the role of NRIP1 on cell viability, apoptosis and NF-κB signaling pathway of HaCaT cells and CD4+T cells; and detect the effect of knocking out Nripl on IMQ-induced psoriasis-like models.Methods1. Immunohistochemistry and real-time PCR was used for detecting NRIP1 and NF-κB expression of skin biopsies and peripheral blood mononuclear cells (PBMCs) from psoriasis patients and healthy volunteers.2. Suppress the NRIP1 of HaCat cells by shRNA transfection, then apply western-blot, MTT assay, Annexin V/PI staining and real-time PCR to analyze the role of NRIP1 on cell viability, apoptosis and the expression of NF-κB.3. Expose sh-NRIP1 transfected HaCat cells to different dose of UVB, and then test the cell proliferation and apoptosis by MTT assay and Annexin V/PI staining.4. Suppress the NR1P1 of CD4+ T cells from psoriasis patients by siRNA transfection, then apply real-time PCR and ELISA to test the expression of NF-κB and secretion of IL-17.5. Apply IMQ-induced psoriasis model in Nripl knockout mice, evaluate the lesion by PASI score and immunohistochemistry, and test the expression of Nripl and NF-κB by immunofluorescence.Results1. Immunohistochemistry and real-time PCR analysis showed that NRJP1 elevates in psoriatic lesions, compared to psoriatic non-lesions and normal skin samples (PP vs NN, P=0.0218; PP vs PN, P=0.0091).2. Real-time PCR analysis showed that NF-κB elevated in psoriatic lesions, compared to psoriatic non-lesions and normal skins (PP vs NN, P= 0.0179; PP vs PN, P= 0.0006; PN vs NN, P= 0.4023).3. Real-time PCR analysis showed that NRIP1 elevates in PBMCs of patients with psoriasis compared to the healthy control (P= 0.0165)4. Real-time PCR analysis showed that NF-κB elevates in PBMCs of patients with psoriasis compared to the healthy control (P= 0.0385).5. Sh-NRIP1 transfection could effectively suppress the expression of NRIP1 of HaCat cells.6. Suppression of NRIP1 in HaCat cells could significantly suppress cells’viability and induce apoptosis (P=0.0005), and reduce the expression of NF-κB.7. UVB exposure could promote the induction of NRIP1 suppression on HaCat cells apoptosis.8. Si-NRIP1 transfection could effectively suppress the expression of NRIP1 of CD4+ T cells from the patients with psoriasis, and could reduce the expression of NF-κB (P=0.0011) and secretion of IL-17 (P=0.0129).9. Topical application of IMQ could induce psoriasis-like lesion on both C57BL/6 and Nrip1 KO mice.10. Nripl knockout could delay the IMQ-induced inflammation of skin.11. Nrip1 knockout could reduce the NF-κB levels of psoriasis-like lesion.ConclusionOur data suggests that NRIP1 and NF-κB over express both in skin and PBMCs of psoriasis patients, and NRIP1 may be involved in abnormal proliferation of keratinocytes and immune reaction by functioning as a coactivator for NF-κB in the pathogenesis of psoriasis.