Next Generation Sequencing identifies lymph node lymphatic endothelial cell genes regulated by RANKL

Purpose: The objective was to identify genes regulted by RANK signalling in lymph node lymphatic endothelial cells Method: Lymphatic endothelial cells were FACS sorted from peripheral lymph nodes of mice deficient for stromal RANKL (Ccl19 Cre Rankl flox/flox) versus littermate controls (Rankl flox/flox) and processed for bulk RNA sequencing Results: Upregulated and downregulated genes were identified Lymphatic cell mRNA profiles of 8 week old littermate (Rankl flox/flox) and stromal RANKL-conditional knock out (Ccl19 Cre Rankl flox/flox) mice were generated by deep sequencing. The sequence reads that passed quality filters and key genes were validated by qRT-PCR using TaqMan and SYBR Green assays on lymphatic cells isolated from Prox-1 Cre Rank flox/flox mice.研究目的：本研究旨在鉴定淋巴结淋巴内皮细胞中受RANK信号通路调控的基因。 实验方法：从基质细胞RANKL缺陷型小鼠（Ccl19 Cre Rankl flox/flox）及其同窝对照小鼠（Rankl flox/flox）的外周淋巴结中通过FACS分选获取淋巴内皮细胞，随后进行批量RNA测序（bulk RNA sequencing）。 实验结果：本研究鉴定得到了差异上调与下调的基因；通过深度测序获取了8周龄同窝对照小鼠（Rankl flox/flox）及基质细胞RANKL条件性敲除小鼠（Ccl19 Cre Rankl flox/flox）的淋巴细胞mRNA表达谱。对通过质量过滤的测序读段与关键基因，采用TaqMan和SYBR Green分析法，对从Prox-1 Cre Rank flox/flox小鼠中分离的淋巴细胞开展qRT-PCR验证。