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Laser capture microdissection for detecting the expression of epithelial–mesenchymal transition‐related genes in epithelial and spindle cells of paraffin‐embedded formalin‐fixed biphasic synovial sarcoma
Author(s) -
Wang Ning,
Liu ZiHan,
Zou Hong,
Pang Lijuan,
Gu Wenyi,
Hu JianMing,
Li Dongmei,
Zhao Jin,
Zhang Jun,
Liu ChunXia,
Zhang WenJie,
Qi Yan,
Li Feng
Publication year - 2018
Publication title -
clinical and experimental pharmacology and physiology
Language(s) - English
Resource type - Journals
SCImago Journal Rank - 0.752
H-Index - 103
eISSN - 1440-1681
pISSN - 0305-1870
DOI - 10.1111/1440-1681.12936
Subject(s) - laser capture microdissection , mesenchymal stem cell , biology , microbiology and biotechnology , pathology , epithelial–mesenchymal transition , microdissection , synovial sarcoma , cell , population , chemistry , gene expression , metastasis , sarcoma , gene , cancer , medicine , biochemistry , genetics , environmental health
Summary Synovial sarcoma ( SS ) is a mesenchymal malignant neoplasm showing characteristics of epithelial–mesenchymal biphasic differentiation. SS is of uncertain cellular origin; however, studies have suggested that SS originates from a somatic stem cell population. In this study, we aim to determine whether differential morphological features of the epithelial–mesenchymal transition ( EMT ) contributed to the tumourigenesis of SS invasion and metastasis. Twelve paraffin‐embedded formalin‐fixed tissue ( FFPE ) SS tissue specimens were obtained, and laser capture microdissection ( LCM ) with the Arcturus XT system and small chip method ( SCM ) were used to isolate and purify spindle and epithelial cells from SS specimens. The TRI zol method was used to extract RNA , and the mRNA levels of EMT ‐related genes in epithelial and spindle cells of SS specimens were measured using real‐time fluorescent quantitative reverse transcription polymerase chain reaction ( qRT ‐ PCR ). The results show that collection of about 2 × 10 4 cells from FFPE samples using LCM was sufficient for qRT ‐ PCR , with an efficiency of 75%. Compared with LCM , 72.2% (13 of 18) RNA samples were successfully extracted using SCM to isolate cells from FFPE SS tissues. In the 16 samples (11 spindle cell samples and 5 epithelial cell samples), Snail mRNA was significantly upregulated in spindle cell areas compared with that in epithelial cell areas ( P = .001). Expression levels of the epithelial marker E‐cadherin and the mesenchymal marker N‐cadherin were not significantly different between epithelial and spindle cell areas. In spindle cells of recurrent SS samples, the mRNA levels of E‐cadherin, N‐cadherin, Snail , and Slug were higher in primary SS samples than in recurrent samples. Taken together, our results indicated that in SS samples, Snail mRNA was upregulated in spindle cell areas compared with that in epithelial cell areas and that the expression of EMT ‐related genes was increased in primary SS . LCM could be used to isolate and purify RNA from FFPE samples.