YAP1/Piezo1 involve in the dynamic changes of lymphatic vessels in UVR-induced photoaging progress to squamous cell carcinoma
Authors: Yang Yuling , Zhou Chu , Chen Qi , Shen Shuzhan , Li Jiandan , Wang Xiuli , Wang Peiru 
Published: April 2023

Abstract
Background UV-induced cutaneous squamous cell carcinoma (cSCC) is the most common skin cancer. The constant alterations of the lymphatic-centered immune microenvironment are essential in transforming from photoaging to cSCC. Studying the mechanism will be beneficial for finding new targets for the early prediction of cSCC.Aims To investigate the dynamic changes and mechanism of the lymphatic-centered immune microenvironment in transforming from photoaging to cSCC induced by ultraviolet irradiation (UVR).Methods TIMER2.0 was used to analyze whether YAP1/VEGFC signaling pathway is involved in lymphangiogenesis in head and neck squamous cell carcinoma (HNSCC). Meanwhile, lymphatic-centered immune microenvironments alterations and the related cumulative survival time were also analyzed. With accumulated UVR at 8, 16–18, and 20–24 weeks, skin photoaging developed and gradually progressed into actinic keratosis and cSCC on SKH-1 hairless mice. The skin lymphatic-centered immune microenvironment was evaluated at the 0th, 8th, 12th, 16-18th, and 20-24th week of UVR. Skin phenotype was assessed using optical coherence tomography (OCT) and skin image. The structure of lymphatic vessels (LVs), blood vessels, and different types of T cells was evaluated by immunohistochemistry staining. The expression of Piezo1, whose deletion in adult lymphatics led to substantial valve degeneration, VE-cadherin, which maintained the drainage function of LVs, and YAP1 were evaluated by immunohistochemistry staining as well. Besides, the drainage function of LVs was assessed by Evans Blue assay in vivo. H&E and Masson’s trichrome staining evaluated the skin dermis and collagen structure.Results The lymphatic function and immune cell infiltration have adapted under continuous UVR. TIMER2.0 analysis indicated that YAP1 and VEGFC genes high expressed in HNSCC. LV density increased in human cSCC. More LVs in HNSCC were beneficial to prolong the survival time. VEGFC and YAP1 gene overexpression was correlated to CD8+T cell infiltration. More CD8A+ T cells and CD8B+ T cell infiltration in HNSCC also extended survival time. However, YAP1 gene overexpression and more CD4+T cell infiltration at the same time might be against survival time. In animal studies, UVR-induced eight weeks (photoaging skin) and 16–18 weeks (actinic keratosis) were two turning points in the lymphatic-centered immune microenvironment. LVs in UV-8w and UV-12w groups were the lowest in density and lower than that in normal mice. When normal skin developed into AK lesions (UV-16-18w), LV slightly exceeded healthy skin and proliferated sharply in cSCC (UV-20-24w). YAP1 expression was consistent with LV but rose after the photoaging stage. The drainage of cSCC induced by UVR was better than that of photoaging skin and worse than that of normal skin. The dynamic alterations of LVs number, Piezo1 expression, and collagen content were reasons for it. Piezo1 expression represented the highest point in the UV-8w group, then gradually descended to the platform. The dermis slowly diminished after chronic UVR, especially the collagen. The total T cells maintained steady, but the number of CD4+T cells increased, and CD8+T cells decreased after eight weeks of UVR. The whole T cells and CD4+T cells increased sharply in UV-16-18w and UV-20-24w groups.Conclusion The lymphatic-centered immune microenvironment has adapted under continuous UVR via regulating YAP1/VEGFC and Piezo1. UVR-induced eight weeks (photoaging) and 16–18 weeks (precancerous), two turning points. YAP1, Piezo1, LVs, and immune cells constantly changed with the skin state induced by UVR. According to changes in photoaging, UV-12w, and precancerous, identify the process of cSCC in advance and intervene timely.