Numerous studies have demonstrated that regulatory T (Treg) cells play an important role in the tumour microenvironment (TME). The aim of this study was to investigate whether VEGFR2 affects the expression of miR-3200-3p in exosomes secreted by tumour cells, thereby influencing Treg senescence in the TME. The results showed that VEGFR2 expression level was the highest in Calu-1 cells, and after transfection with si-VEGFR2, the exosomes secreted from Calu-1 cells were extracted and characterised with no significant difference from the exosomes of the untransfected group, but the expression of miR-3200-3p in the exosomes of the transfected si-VEGFR2 group was elevated. The Cell Counting Kit-8 (CCK-8) and flow cytometry (FCM) results suggested that exosomes highly expressing miR-3200-3p could inhibit Treg cell viability and promote apoptosis levels when treated with Treg cells. Detection of the senescence-associated proteins p16 INK4A and MMP3 by western blot (WB) revealed that exosomes highly expressing miR-3200-3p were able to elevate their protein expression levels. Tumour xenograft experiments demonstrated that exosomes with high miR-3200-3p expression promoted Treg cell senescence and inhibited subcutaneous tumour growth in nude mice. Dual-luciferase reporter assays and RNA pull-down assays showed that miR-3200-3p could be linked with DDB1. Overexpression of DDB1 reverses changes in DCAF1/GSTP1/ROS protein expression caused by exosomes with high miR-3200-3p expression. In conclusion, inhibition of VEGFR2 expression in tumour cells promotes the expression of miR-3200-3p in exosomes secreted by tumour cells. miR-3200-3p enters the TME through exosomes and acts on DDB1 in Treg cells to promote senescence of Treg cells to inhibit tumour progression.

大量研究表明，调节性T（Treg）细胞在肿瘤微环境（TME）中发挥着重要作用。本研究的目的是探讨VEGFR2是否影响肿瘤细胞分泌的外泌体中miR-3200-3p的表达，从而影响TME中的Treg衰老。结果显示，Calu-1细胞中VEGFR2表达量最高，转染si-VEGFR2后，提取Calu-1细胞分泌的外泌体并进行表征，与未转染组的外泌体没有显着差异，但转染si-VEGFR2组外泌体中miR-3200-3p表达升高。细胞计数试剂盒-8 (CCK-8) 和流式细胞术 (FCM) 结果表明，高表达 miR-3200-3p 的外泌体在用 Treg 细胞处理时可以抑制 Treg 细胞活力并促进凋亡水平。通过蛋白质印迹（WB）检测衰老相关蛋白 p16 INK4A 和 MMP3 表明，高表达 miR-3200-3p 的外泌体能够提高其蛋白表达水平。肿瘤异种移植实验表明，高miR-3200-3p表达的外泌体促进Treg细胞衰老并抑制裸​​鼠皮下肿瘤生长。双荧光素酶报告基因检测和 RNA Pull-down 检测表明 miR-3200-3p 可以与 DDB1 连接。 DDB1 的过表达可逆转由 miR-3200-3p 高表达的外泌体引起的 DCAF1/GSTP1/ROS 蛋白表达的变化。总之，抑制肿瘤细胞中的 VEGFR2 表达可促进肿瘤细胞分泌的外泌体中 miR-3200-3p 的表达。 miR-3200-3p通过外泌体进入TME，作用于Treg细胞中的DDB1，促进Treg细胞衰老，从而抑制肿瘤进展。