Effect of Transforming Growth Factor-beta 1 on the Rat Renal Mesangial Cell Transcriptome

Rat renal mesangial cells (SciencCell, #R4200) were grown to sub-confluence on poly-L-lysine coated flasks in mesgnaial cell medium (SciencCell, #R4201) supplemented with 2�S, growth supplement (SciencCell, #R4252) and pen/strep. Cells were quiesced for 48h and then treated for 24h with human recombinant TGF-beta 1 (5 ng/mL; Sigma#T7039) in complete media (n=4) or fresh media without growth factor (n=4). Total RNA was extraced from cell pellets using standard protocols. Whole RNA stranded libraries were prepared using Illumina's Ribo-zero Gold protocol and sequenced on an Illumina NovaSeq platform (150bp PE). After demultiplexing and quality control, the data was then processed through RNA-seq expression analysis workflow, which included trimming, alignment, transcript assembly, feature quantification and, differential expression analysis with edgeR (verison 3.38.4).

Rat renal mesangial cells (SciencCell, #R4200) 于poly-L-lysine涂层瓶中，在肾小球细胞培养基（SciencCell, #R4201）中增殖，该培养基中添加了2μS生长补充剂（SciencCell, #R4252）及青霉素/链霉素。细胞经过48小时的静息处理，随后在含有完整培养基（n=4）或不含生长因子的新鲜培养基（n=4）中，用人类重组TGF-β1（5 ng/mL；Sigma#T7039）处理24小时。通过标准程序从细胞沉淀中提取总RNA。利用Illumina的Ribo-zero Gold方案制备全RNA链文库，并在Illumina NovaSeq平台上进行测序（150bp PE）。在去除混合样本和进行质量控制后，数据通过RNA-seq表达分析工作流程进行处理，包括剪切、对齐、转录组装、特征定量以及使用edgeR（版本3.38.4）进行差异表达分析。