Improvements and challenges of tissue preparation for spatial transcriptome analysis of skull base tumors
Yan Fang,
Zhouying Peng,
Yumin Wang,
Xiaotian Yuan,
Kelei Gao,
Ruohao Fan,
Ruijie Liu,
Yalan Liu,
Hua Zhang,
Zhihai Xie,
Weihong Jiang
Affiliations
Yan Fang
Department of Otolaryngology Head and Neck Surgery, Xiangya Hospital, Central South University, Changsha, Hunan, 410008, China; Otolaryngology Major Disease Research Key Laboratory of Hunan Province, Changsha, Hunan, 410008, China; National Clinical Research Center for Geriatric Disorders, Xiangya Hospital, Central South University, Changsha, Hunan, 410008, China; Anatomy Laboratory of Division of Nose and Cranial Base, Clinical Anatomy Center of Xiangya Hospital, Central South University, Changsha, Hunan, 410008, China
Zhouying Peng
Department of Otolaryngology Head and Neck Surgery, Xiangya Hospital, Central South University, Changsha, Hunan, 410008, China; Otolaryngology Major Disease Research Key Laboratory of Hunan Province, Changsha, Hunan, 410008, China; National Clinical Research Center for Geriatric Disorders, Xiangya Hospital, Central South University, Changsha, Hunan, 410008, China; Anatomy Laboratory of Division of Nose and Cranial Base, Clinical Anatomy Center of Xiangya Hospital, Central South University, Changsha, Hunan, 410008, China
Yumin Wang
Department of Otolaryngology Head and Neck Surgery, Xiangya Hospital, Central South University, Changsha, Hunan, 410008, China; Otolaryngology Major Disease Research Key Laboratory of Hunan Province, Changsha, Hunan, 410008, China; National Clinical Research Center for Geriatric Disorders, Xiangya Hospital, Central South University, Changsha, Hunan, 410008, China; Anatomy Laboratory of Division of Nose and Cranial Base, Clinical Anatomy Center of Xiangya Hospital, Central South University, Changsha, Hunan, 410008, China
Xiaotian Yuan
Department of Otolaryngology Head and Neck Surgery, Xiangya Hospital, Central South University, Changsha, Hunan, 410008, China; Otolaryngology Major Disease Research Key Laboratory of Hunan Province, Changsha, Hunan, 410008, China; National Clinical Research Center for Geriatric Disorders, Xiangya Hospital, Central South University, Changsha, Hunan, 410008, China
Kelei Gao
Department of Otolaryngology Head and Neck Surgery, Xiangya Hospital, Central South University, Changsha, Hunan, 410008, China; Otolaryngology Major Disease Research Key Laboratory of Hunan Province, Changsha, Hunan, 410008, China; Anatomy Laboratory of Division of Nose and Cranial Base, Clinical Anatomy Center of Xiangya Hospital, Central South University, Changsha, Hunan, 410008, China
Ruohao Fan
Department of Otolaryngology Head and Neck Surgery, Xiangya Hospital, Central South University, Changsha, Hunan, 410008, China; Otolaryngology Major Disease Research Key Laboratory of Hunan Province, Changsha, Hunan, 410008, China; National Clinical Research Center for Geriatric Disorders, Xiangya Hospital, Central South University, Changsha, Hunan, 410008, China; Anatomy Laboratory of Division of Nose and Cranial Base, Clinical Anatomy Center of Xiangya Hospital, Central South University, Changsha, Hunan, 410008, China
Ruijie Liu
National Clinical Research Center for Geriatric Disorders, Xiangya Hospital, Central South University, Changsha, Hunan, 410008, China; Department of Pathology, Xiangya Hospital, Central South University, Changsha, Hunan, 410008, China
Yalan Liu
Department of Otolaryngology Head and Neck Surgery, Xiangya Hospital, Central South University, Changsha, Hunan, 410008, China; Otolaryngology Major Disease Research Key Laboratory of Hunan Province, Changsha, Hunan, 410008, China; National Clinical Research Center for Geriatric Disorders, Xiangya Hospital, Central South University, Changsha, Hunan, 410008, China
Hua Zhang
Department of Otolaryngology Head and Neck Surgery, Xiangya Hospital, Central South University, Changsha, Hunan, 410008, China; Otolaryngology Major Disease Research Key Laboratory of Hunan Province, Changsha, Hunan, 410008, China; National Clinical Research Center for Geriatric Disorders, Xiangya Hospital, Central South University, Changsha, Hunan, 410008, China; Anatomy Laboratory of Division of Nose and Cranial Base, Clinical Anatomy Center of Xiangya Hospital, Central South University, Changsha, Hunan, 410008, China
Zhihai Xie
Department of Otolaryngology Head and Neck Surgery, Xiangya Hospital, Central South University, Changsha, Hunan, 410008, China; Otolaryngology Major Disease Research Key Laboratory of Hunan Province, Changsha, Hunan, 410008, China; National Clinical Research Center for Geriatric Disorders, Xiangya Hospital, Central South University, Changsha, Hunan, 410008, China; Anatomy Laboratory of Division of Nose and Cranial Base, Clinical Anatomy Center of Xiangya Hospital, Central South University, Changsha, Hunan, 410008, China
Weihong Jiang
Department of Otolaryngology Head and Neck Surgery, Xiangya Hospital, Central South University, Changsha, Hunan, 410008, China; Otolaryngology Major Disease Research Key Laboratory of Hunan Province, Changsha, Hunan, 410008, China; National Clinical Research Center for Geriatric Disorders, Xiangya Hospital, Central South University, Changsha, Hunan, 410008, China; Anatomy Laboratory of Division of Nose and Cranial Base, Clinical Anatomy Center of Xiangya Hospital, Central South University, Changsha, Hunan, 410008, China; Corresponding author. Department of Otolaryngology Head and Neck Surgery, Xiangya Hospital, Central South University, Changsha, Hunan, 410008, China.
Background: Spatial transcriptome (ST) provides molecular profiles of tumor cells at the spatial level, which brings new progress to the research of tumors and the tumor microenvironment. This study summarizes the experiences and lessons learned in the spatial section preparation of two different pathological types of nose and skull base tumors at our institution, with the aim of offering guidelines to researchers to avoid wasting precious samples and provide a basis for the application of ST in clinical practice. Methods: Frozen tissue blocks from patients with squamous cell carcinoma and adenocarcinoma of the nose and skull base diagnosed at our institution were prepared. The effects of different procedures and pathological tissue types on slide quality were explored and evaluated using RNA integrity number (RIN) and HE scores as criteria. The effects of different RIN values on ST sequencing data were explored. Results: A total of 43 samples were obtained from 26 patients, including 22 with squamous carcinomas and 21 with adenocarcinomas. Thirteen samples with satisfactory RNA quality control and good histological morphology were sequenced for ST. Sample isolation time 7 in ST sequencing results was found, indicating that RIN ≥6 can be used as a criterion for qualified RNA quality control. Therefore, fresh tissues washed as soon as possible with cold PBS and then dried using OCT for snap freezing are currently the best method for preparing spatial sections of nose and skull base tumor tissues of different pathological types. Conclusion: This study is the first to investigate the feasibility of applying ST to different pathological types of nose and skull base tumors and to demonstrate the widespread application of ST in tumors. Rational optimization of spatial slide preparation procedures and exploration of individualized pre-sequencing protocols are used as the first stage to ensure the quality of spatial sequencing and lay the foundation for subsequent spatial analysis.
