Spatio-temporal landscape of mouse epididymal cells and specific mitochondria-rich segments defined by large-scale single-cell RNA-seq

In total, 40,623 cells were detected and further clustered into 8 identified cell populations. Focused analyses revealed the subpopulations of principal cells, basal cells, clear/narrow cells, and halo/T cells. Notably, two subtypes of principal cells, the Prc7 and Prc8 subpopulations were enriched as stereocilia-like cells according to GO analysis. Further analysis demonstrated the spatially specific pattern of the DEGs in each cell cluster. Unexpectedly, the abundance of mitochondria and mitochondrial transcription (MT) was found to be higher in the corpus and cauda epididymis than in the caput epididymis by scRNA-seq, immunostaining, and qPCR validation. In addition, the spatio-temporal profile of the DEGs from the P42 and P56 epididymis, including transiting spermatozoa, was depicted. Five 42 days (P42) and ten 56 days (P56) old of wild-type C57Bl/6J mice were sacrified, then the epididymis were dissected and divided into three segments (caput, corpus, and cauda). Ten epididymis from five mice were cut into the different segment and immediately transferred into the GEXSCOPETM Tissue Preservation Solution (Singleron Biotechnologies) in an Eppendorf tube labeled as EP (caput), EO (corpus) and ED (cauda) on ice. For P56 epididymis, samples were two replicates. The procedure was repeated in a mice until all segments were collected in each tube. The pooled segments were further to the procedure of tissue dissociation and single cell preparation.