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Abstract：
Retinoblastoma (RB) is the most common intraocular malignancy in children. It has been previously reported that p38 MAPK is related to the pathogenesis of RB. Here we aim at investigating how p38 MAPK affected RB progression through mediating USP22/SIRT1/SOST axis. In this study, Thirty-two cases of RB and normal retinal tissues were collected. The expression of p38 MAPK, phosphorylation of p38 MAPK (P-p38 MAPK), USP22, SIRT1 and SOST in clinical tissues and cells was measured using RT-qPCR, IHC assay or western blot analysis. Cell proliferation was detected by CCK-8. Apoptosis rate of cells was examined by flow cytometry. Cell migration was evaluated using scratch test. Cell invasion ability was examined by Transwell assay. Co-immunoprecipitation (CO-IP) was utilized to measure the deubiquitination of USP22 on SIRT1. In vivo, mice were respectively injected with plasmids and the tumor growth as well as the tumor weight were detected. Results showed that p38 MAPK, P-p38 MAPK and SOST were poorly expressed in RB tissues and cells whereas USP22 and SIRT1 were overly expressed. P-p38 MAPK inhibited the expression of USP22, and overexpression of USP22 eliminated the inhibitory roles of P-p38 MAPK on tumor growth, as well as cell proliferation, migration and invasion. USP22 stabilized and promoted the expression of SIRT1 through its deubiquitination function. Silencing the expression of SIRT1 contributed to boosted expression of SOST, thus suppressing the growth of tumor cells. Collectively, the phosphorylation of p38 MAPK regulates the SIRT1/SOST axis to protect against RB via silencing USP22. The findings present some cues for a further approach to RB.
摘要：
视网膜母细胞瘤（RB）是儿童最常见的眼内恶性肿瘤。已有文献报道p38MAPK与RB的发病机制有关。在这里，我们旨在研究p38MAPK如何通过介导USP22/SIRT1/SOST轴影响RB进展。在这项研究中,收集RB和正常视网膜组织32例。p38MAPK的表达,磷酸化p38MAPK（P-p38MAPK），使用RT-qPCR测量临床组织和细胞中的USP22，SIRT1和SOST，IHC测定或蛋白质印迹分析。CCK-8检测细胞增殖。流式细胞术检测细胞凋亡率。使用划痕测试评估细胞迁移。通过Transwell测定法检查细胞侵袭能力。免疫共沉淀(CO-IP)用于测量USP22在SIRT1上的去泛素化。在体内,小鼠分别注射质粒，检测肿瘤生长情况和肿瘤重量。结果显示p38MAPK,P-p38MAPK和SOST在RB组织和细胞中表达不佳，而USP22和SIRT1过度表达。P-p38MAPK抑制USP22的表达，USP22的过表达消除了P-p38MAPK对肿瘤生长的抑制作用，以及细胞增殖，移民和入侵。USP22通过其去泛素化功能稳定并促进SIRT1的表达。沉默SIRT1的表达有助于SOST的增强表达，从而抑制肿瘤细胞的生长。总的来说，p38MAPK的磷酸化调节SIRT1/SOST轴，通过沉默USP22保护RB。这些发现为进一步研究RB提供了一些线索。
