Dataset Information

Intracellular trafficking and endocytosis of CXCR4 in fetal mesenchymal stem/stromal cells.

ABSTRACT:

Background

Fetal mesenchymal stem/stromal cells (MSC) represent a developmentally-advantageous cell type with translational potential.To enhance adult MSC migration, studies have focussed on the role of the chemokine receptor CXCR4 and its ligand SDF-1 (CXCL12), but more recent work implicates an intricate system of CXCR4 receptor dimerization, intracellular localization, multiple ligands, splice variants and nuclear accumulation. We investigated the intracellular localization of CXCR4 in fetal bone marrow-derived MSC and role of intracellular trafficking in CXCR4 surface expression and function.

Results

We found that up to 4% of human fetal MSC have detectable surface-localized CXCR4. In the majority of cells, CXCR4 is located not at the cell surface, as would be required for 'sensing' migratory cues, but intracellularly. CXCR4 was identified in early endosomes, recycling endosomes, and lysosomes, indicating only a small percentage of CXCR4 travelling to the plasma membrane. Notably CXCR4 was also found in and around the nucleus, as detected with an anti-CXCR4 antibody directed specifically against CXCR4 isoform 2 differing only in N-terminal sequence. After demonstrating that endocytosis of CXCR4 is largely independent of endogenously-produced SDF-1, we next applied the cytoskeletal inhibitors blebbistatin and dynasore to inhibit endocytotic recycling. These increased the number of cells expressing surface CXCR4 by 10 and 5 fold respectively, and enhanced the number of cells migrating to SDF1 in vitro (up to 2.6 fold). These molecules had a transient effect on cell morphology and adhesion, which abated after the removal of the inhibitors, and did not alter functional stem cell properties.

Conclusions

We conclude that constitutive endocytosis is implicated in the regulation of CXCR4 membrane expression, and suggest a novel pharmacological strategy to enhance migration of systemically-transplanted cells.

SUBMITTER: Pelekanos RA

PROVIDER: S-EPMC4065074 | biostudies-literature | 2014 May

REPOSITORIES: biostudies-literature

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Publications

Intracellular trafficking and endocytosis of CXCR4 in fetal mesenchymal stem/stromal cells.

Pelekanos Rebecca A RA Ting Michael J MJ Sardesai Varda S VS Ryan Jennifer M JM Lim Yaw-Chyn YC Chan Jerry K Y JK Fisk Nicholas M NM

BMC cell biology 20140516

<h4>Background</h4>Fetal mesenchymal stem/stromal cells (MSC) represent a developmentally-advantageous cell type with translational potential.To enhance adult MSC migration, studies have focussed on the role of the chemokine receptor CXCR4 and its ligand SDF-1 (CXCL12), but more recent work implicates an intricate system of CXCR4 receptor dimerization, intracellular localization, multiple ligands, splice variants and nuclear accumulation. We investigated the intracellular localization of CXCR4 i ...[more]

PMID: 24885150

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Project description:BackgroundOur previous studies indicated that MSC(CXCR4) improved cardiac function after myocardial infarction (MI). This study was aimed to investigate the specific role of MSC(CXCR4) in neovascularization of infarcted myocardium using a suicide gene approach.MethodsMSCs were transduced with either lentivirus-null vector/GFP (MSC(Null) as control) or vector encoding for overexpressing CXCR4/GFP. The MSC derived-endothelial cell (EC) differentiation was assessed by a tube formation assay, Dil-ac-LDL uptake, EC marker expression, and VE-cadherin promoter activity assay. Gene expression was analyzed by quantitative RT-PCR or Western blot. The suicide gene approach was under the control of VE-cadherin promoter. In vivo studies: Cell patches containing MSC(Null) or MSC(CXCR4) were transduced with suicide gene and implanted into the myocardium of MI rat. Rats received either ganciclovir (GCV) or vehicle after cell implantation. After one month, the cardiac functional changes and neovascularization were assessed by echocardiography, histological analysis, and micro-CT imaging.ResultsThe expression of VEGF-A and HIF-1? was significantly higher in MSC(CXCR4) as compared to MSC(Null) under hypoxia. Additionally, MSC(CXCR4) enhanced new vessel formation and EC differentiation, as well as STAT3 phosphorylation under hypoxia. STAT3 participated in the transcription of VE-cadherin in MSC(CXCR4) under hypoxia, which was inhibited by WP1066 (a STAT3 inhibitor). In addition, GCV specifically induced death of ECs with suicide gene activation. In vivo studies: MSC(CXCR4) implantation promoted cardiac functional restoration, reduced infarct size, improved cardiac remodeling, and enhanced neovascularization in ischemic heart tissue. New vessels derived from MSC(CXCR4) were observed at the injured heart margins and communicated with native coronary arteries. However, the derived vessel networks were reduced by GCV, reversing improvement of cardiac function.ConclusionThe transplanted MSC(CXCR4) enhanced neovascularization after MI by boosting release of angiogenic factors and increasing the potential of endothelial differentiation.

Dataset Information

Intracellular trafficking and endocytosis of CXCR4 in fetal mesenchymal stem/stromal cells.

Background

Results

Conclusions

Publications

Intracellular trafficking and endocytosis of CXCR4 in fetal mesenchymal stem/stromal cells.

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