✍🏼 Yunmei Dong, Kan Zeng, Ruixue Ai, Chengli Zhang, Fei Mao, Hongxia Dan, Xin Zeng, Ning Ji, Jing Li, Xin Jin, Qianming Chen, Yu Zhou, Taiwen L

🏠 State Key Laboratory of Oral Diseases, National Clinical Research Center for Oral Diseases, Chinese Academy of Medical Sciences Research Unit of Oral Carcinogenesis and Management, West China Hospital of Stomatology, Sichuan University, Chengdu, China

📑 MedComm (2024)

Abstract
Oral squamous cell carcinoma (OSCC) stands as a predominant and perilous malignant neoplasm globally, with the majority of cases originating from oral potential malignant disorders (OPMDs). Despite this, effective strategies to impede the progression of OPMDs to OSCC remain elusive. In this study, we established mouse models of oral carcinogenesis via 4-nitroquinoline 1-oxide induction, mirroring the sequential transformation from normal oral mucosa to OPMDs, culminating in OSCC development. By intervening during the OPMDs stage, we observed that combining PD1 blockade with photodynamic therapy (PDT) significantly mitigated oral carcinogenesis progression. Single-cell transcriptomic sequencing unveiled microenvironmental dysregulation occurring predominantly from OPMDs to OSCC stages, fostering a tumor-promoting milieu characterized by increased Treg proportion, heightened S100A8 expression, and decreased Fib_Igfbp5 (a specific fibroblast subtype) proportion, among others. Notably, intervening with PD1 blockade and PDT during the OPMDs stage hindered the formation of the tumor-promoting microenvironment, resulting in decreased Treg proportion, reduced S100A8 expression, and increased Fib_Igfbp5 proportion. Moreover, combination therapy elicited a more robust treatment-associated immune response compared with monotherapy. In essence, our findings present a novel strategy for curtailing the progression of oral carcinogenesis.

How the WOLF was used in this study
The researchers utilized the WOLF cell sorter as part of their preparation of single‑cell suspensions for downstream transcriptomic analysis. After enzymatically digesting mouse tongue tissues from different experimental groups, the investigators stained cells with antibodies against CD45 and viability dye 7AAD to distinguish immune from non‑immune and live from dead cells. The WOLF sorter was then used to physically separate and collect viable CD45⁺7AAD⁻ immune cells and CD45⁻7AAD⁻ non‑immune cells, which were subsequently processed for single‑cell RNA sequencing to build a comprehensive atlas of microenvironmental cell types and states during carcinogenesis and after PD‑1 blockade ± photodynamic therapy. This sorting step was critical to enrich for live, defined cell populations prior to high‑resolution single‑cell transcriptomic profiling.