Transcriptome analysis of human dental pulp cells cultured on a novel cell-adhesive fragment by RNA sequencing.

AIM

The aim of the present work was to find an efficient method for safe and reliable expansion of human dental pulp cells (hDPCs) in vitro. Here, we examined the effect of a novel recombinant E8 fragment of Laminin-511 (iMatrix-511) in hDPCs regarding viability and cell spreading. Further, we investigated the underlying mechanisms governing its effects in hDPCs using RNA sequencing (RNA-seq).

METHODOLOGY

hDPCs were obtained from caries-free maxilla third molars (n = 3). CCK-8 assay was conducted to measure the viability of cells cultured on iMatrix-511 and two other ECM proteins. Cell morphology was observed by phase contrast microscope. RNA-seq of hDPCs cultured on iMatrix-511 or noncoated control was performed on Illumina Novaseq 6000 platform.

RESULTS

iMatrix-511 (0.5 μg/cm) enhanced the viability of hDPCs to an extent better than COL-1 and gelatin. Short term culture of hDPCs on iMatrix-511 resulted in 233 differentially expressed genes (DEGs). The top 12 most upregulated genes were XIAP, AL354740, MRFAP1, AC012321, KCND3, TMEM120B, AC009812, GET1-SH3BGR, CNTN3, AC090409, GEN1 and PIK3IP1, whereas the top 12 most downregulated genes were SFN, KRT17, RAB4B-EGLN2, CSTA, KCTD11, ATP6V1G2-DDX39B, AC010323, SBSN, LYPD3, FOSB, AC022400 and CHI3L1. qPCR validation confirmed the significant upregulation of GEN1, KCND3, PIK3IP1 and MRFAP1. Gene Ontology (GO) and Kyoto Encyclopedia of Genes and Genomes (KEGG) analyses were performed, with genes enriched in various extracellular matrix interaction, estrogen and fat metabolism-related functions and pathways.

CONCLUSIONS

iMatrix-511 facilitated spreading and proliferation of hDPCs. It enhances expression of anti-apoptotic genes, while inhibits expression of epidermis development-related genes.