مقالات پذیرفته شده در نهمین کنگره بین المللی زیست پزشکی
Studying the gene expression differentiation between mesenchymal stem cells and osteogenics
Studying the gene expression differentiation between mesenchymal stem cells and osteogenics
Amir Arsalan Jourabloo,1Shirin Farivar,2,*
1. M.Sc candidate in Biotechnology; Department of Microbiology and Biotechnology, Faculty of Life Sciences and Biotechnology, Shahid Beheshti University, Tehran, IRAN. (a.jourabloo@sbu.ac.ir ) 2. Associate Professor; Department of Cell and Molecular Biology, Faculty of Life Sciences and Biotechnology, Shahid Beheshti University, Tehran, IRAN. (s_farivar@sbu.ac.ir )
Introduction: The investigation of disparities between gene expression profiles in mesenchymal stem cells (MSCs) and osteogenic stem cells is a significant topic of interest in the field of tissue engineering and regenerative medicine. MSCs are an interesting class of multipotent cells known for their ability to differentiate into numerous cell types, such as osteoblasts, which are important for the formation of new bone and healing bone. As we assist in organizing and identifying additional mechanisms to improve clinical concepts and approaches in bone disease and injury, it is essential to deepen
an improved understanding of the molecular mechanisms regulating the processes of these cells differentiating, through gene expression. Most recently, highlighted the role of key transcription factors and for signaling pathways contributing to the importance of the extracellular matrix, which has an applied role in MSCs to osteogenic cell lines. This exciting area not only enhances our qualitative understanding of biological processes, but also lends itself to new areas of exploration, like improving bone regeneration. And here, possible engagement with biomaterials for clinical practices in orthopedics.
Methods: In order to assess RNAseq data and identify differences in gene expression, we utilized the Galaxy server, a powerful and easy to use platform for bioinformatics data analysis. We obtained raw sequencing data from the NCBI Sequence Read Archive (SRA) containing paired-end reads in accessions SRR23920425, SRR23920427, SRR12782290, and SRR12782291, and used FastQC for initial quality control checks. FastQC provides quality statistics including things like sequence length distribution, GC content, and per-base sequence quality. Because FastQC generates a large amount of quality report statistics, we also used MultiQC to generate a report that summarized all of the individual FastQC metrics into a single overview of all samples. Once quality coverage was confirmed, we then used HISAT2, a splice-aware aligner, to align high-quality reads to a reference genome (using the reference genome from Ensembl v.93). HISAT2 is noted for its high accuracy aligning RNAseq data. We used FeatureCounts to quantify gene expression levels to produce raw count matrices of transcription profiles for each sample. The paired-end sequencing approach improved the accuracy of alignment and transcriptome mapping, thus, assuring robustness of the analysis pipeline.
Results: The analysis of RNA sequencing data revealed different patterns of gene expression associated with the differentiation of mesenchymal stem cells (MSCs) to osteogenic lineages. Significant upregulation of osteogenic-related genes, such as VCAM1, members of the collagen (COL) family, and ALPL, demonstrated their essential role in regulating osteogenic differentiation. Likewise, genes such as FGF18, members of the HOX family, and SOX transcription factors exhibited downregulation, suggesting a reduced involvement in MSC differentiation to osteogenic cells. The differential gene expression was demonstrated utilizing a heatmap configuration to clearly illustrate the transcriptional profiles for the relative samples. The heatmap also depicted clusters of genes exhibiting similar patterns of expression, which conveyed added descriptions for underlying molecular mechanisms associated with differentiation of MSCs to osteogenic cells.
Conclusion: The research successfully illustrates unique gene expression profiles of mesenchymal stem cells (MSCs) to their osteogenic lineage, highlighting specific molecular networks involved in this process of differentiation. A prominent feature of this study was the identification of key osteogenic markers, such as VCAM1, members of the collagen (COL) family, and ALPL, robustly upregulated during differentiation which are essential to support the process of osteogenesis and cellular maturation. Conversely, some gene candidates such as FGF18, members of the HOX family, or SOX transcription factors were downregulated and indicate either a reduced role or function in this context. These changes in gene regulation from MSCs to the osteogenic lineage were visualized using heatmaps to reflecting the dynamic transcriptomic shifts in regulating osteogenic differentiation; the altered O-CL, D-CL, or other previously identified differentiation networks. The findings from this study contribute to the identification of novel therapeutic targets of regenerative medicine, and tissue engineering. Collectively, this work advances our understanding of the transcriptomic landscape occurring when MSCs differentiate toward an osteogenic lineage for potential clinical applications.