Reparo de DNA em células-tronco mesenquimais humanas submetidas a diferentes condições de cultivo

Abstract

Human mesenchymal stem cells (HSC) are multipotent adult cells that have the ability to differentiate into many different lineages. Previous works by our group have shown that HMSC present a high frequency of nuclear alterations such as bridges and nucleoplasmic buds, both markers of tumor cells and chromosomal instability when cells enter senescence or when they are exposed to biomaterials. Thus, the aim of this work was to perform a differential expression analysis of DNA repair genes using a PCR-array matrix. For this, the MSC were cultivated under different culture conditions, such as adipogenic differentiation (ADC), with hydroxyapatite microparticles (CTMHAp) and senescence (CSs). In total, 84 genes involved in the main DNA repair mechanisms were analyzed. In the ADCs, there was a decrease in the expression of the XPC, APEX1 and XRCC4 genes. CTMHAp activated several repair pathways, especially the DSBr and MMR pathways. CSs also had differentially expressed genes (BRCA1, XRCC4, APEX1), which in turn were linked to the DSBr and BER pathways. PPI networks were built to investigate possible interactions between proteins that act in DNA repair and their behavior in each culture condition. At ADC, the network consisted of 8 nodes and 17 connections, with EXO1 and APEX1 as the bottleneck nodes, related to the MMR and BER routes. In CTMHAp, the network had 26 nodes and 220 connections, where EXO1, ERCC5 and RAD52 were identified as bottlenecks, related to MMR, NER and DSBR pathways. In the CSs, 55 nodes and 878 connections were identified, with XRCC1 as the bottleneck, being related to the SSBr and BER routes. We identified the transcription factors involved in each of the constructed networks, and those of the E2F family, involved with cell cycle progression, were common to the three conditions. Together, the data from this study showed that, when subjected to different conditions of cell culture/stress, MSCs respond effectively and quickly, being able to prevent damage to the DNA and maintain the integrity of the genome.