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Background <p>Endothelial colony-forming cells (ECFCs) are progenitors of endothelial cells and have shown angiogenic effects in pre-clinical studies in ischaemic tissues. ECFCs may represent a potential therapy for patients who require vascular repair, including those with diabetes mellitus (DM). However, some, but not all, studies have reported a decrease in the number and impaired function of peripheral blood-derived ECFCs (PB-ECFCs) from patients with diabetes. This study aimed to compare the gene expression profiles and pathways between control and diabetic PB-ECFCs in order to understand mechanisms of diabetes-induced dysfunction.</p> Methods <p>Bulk RNA sequencing (RNA-Seq) was performed to compare the gene expression patterns of PB-ECFCs from control and diabetes groups. Subsequently, gene enrichment analysis (GSEA) was used to identify enriched pathways in PB-ECFCs from the diabetes group. To mimic the inflammatory environment of DM, PB-ECFCs were treated with TNF-α. Changes in gene and microRNA (miRNA) expression following TNF-α exposure were analysed via qRT-PCR. The proliferation, cell adhesion and tube formation ability of PB-ECFCs were assessed using Prestoblue, THP-1 cell adhesion, and in vitro Matrigel assays, respectively.</p> Results <p>We identified 66 differentially expressed genes (28 upregulated and 38 downregulated) between control and diabetic PB-ECFCs. GSEA showed that the TNF-α pathway was markedly enriched in PB-ECFCs from the diabetes group. We then exposed ECFCs to TNF-α and demonstrated activation of inflammatory pathways, which resulted in inhibition of proliferation and potential impairment of tube formation (reducing the number of nodes and total length) in control PB-ECFCs, and enhanced cell adhesion to THP-1. Additionally, we found that three miRNAs (miR-146a-5p, miR-199a-3p, and miR-199a-5p) were upregulated in control PB-ECFCs upon exposure to TNF-α.</p> Conclusion <p>In summary, this study suggested that the TNF-α pathway may play an important role in the pathophysiology of diabetic PB-ECFCs. The upregulation of miR-146a-5p and miR-199a-3p was likely a compensatory response to the inflammation induced by TNF-α. These findings may provide a rationale for the intervention of intracellular miRNAs as possible targets for improving PB-ECFC function in the inflammatory environment.</p>

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TNF-α Pathway Activation and Altered microRNA Expression in Peripheral Blood-Derived Endothelial Colony-Forming Cells: Insights into Diabetes-Induced Dysfunction

  • 期刊论文

  • 发表日期：

    2025年7月9日

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  • Yaqiong Liu,
  • Caomhán J. Lyons,
  • Alan Keane,
  • Stuart McKeown,
  • Edoardo Pedrini,
  • Michael Creane,
  • Nadeem Soomro,
  • Alicja Straszewicz,
  • Tomás Griffin,
  • Alan Stitt,
  • Timothy O’Brien

摘要

Background

Endothelial colony-forming cells (ECFCs) are progenitors of endothelial cells and have shown angiogenic effects in pre-clinical studies in ischaemic tissues. ECFCs may represent a potential therapy for patients who require vascular repair, including those with diabetes mellitus (DM). However, some, but not all, studies have reported a decrease in the number and impaired function of peripheral blood-derived ECFCs (PB-ECFCs) from patients with diabetes. This study aimed to compare the gene expression profiles and pathways between control and diabetic PB-ECFCs in order to understand mechanisms of diabetes-induced dysfunction.

Methods

Bulk RNA sequencing (RNA-Seq) was performed to compare the gene expression patterns of PB-ECFCs from control and diabetes groups. Subsequently, gene enrichment analysis (GSEA) was used to identify enriched pathways in PB-ECFCs from the diabetes group. To mimic the inflammatory environment of DM, PB-ECFCs were treated with TNF-α. Changes in gene and microRNA (miRNA) expression following TNF-α exposure were analysed via qRT-PCR. The proliferation, cell adhesion and tube formation ability of PB-ECFCs were assessed using Prestoblue, THP-1 cell adhesion, and in vitro Matrigel assays, respectively.

Results

We identified 66 differentially expressed genes (28 upregulated and 38 downregulated) between control and diabetic PB-ECFCs. GSEA showed that the TNF-α pathway was markedly enriched in PB-ECFCs from the diabetes group. We then exposed ECFCs to TNF-α and demonstrated activation of inflammatory pathways, which resulted in inhibition of proliferation and potential impairment of tube formation (reducing the number of nodes and total length) in control PB-ECFCs, and enhanced cell adhesion to THP-1. Additionally, we found that three miRNAs (miR-146a-5p, miR-199a-3p, and miR-199a-5p) were upregulated in control PB-ECFCs upon exposure to TNF-α.

Conclusion

In summary, this study suggested that the TNF-α pathway may play an important role in the pathophysiology of diabetic PB-ECFCs. The upregulation of miR-146a-5p and miR-199a-3p was likely a compensatory response to the inflammation induced by TNF-α. These findings may provide a rationale for the intervention of intracellular miRNAs as possible targets for improving PB-ECFC function in the inflammatory environment.