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Mitochondrial fission protein 1 up-regulation ameliorates senescence-related endothelial dysfunction of human endothelial progenitor cells

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Abstract

Background

We investigated the contribution of mitochondrial dysfunction to the senescence of human endothelial progenitor cells (EPCs) expanded in vitro and the underlying molecular mechanism.

Methods and results

Serial passage increased cell doubling time and those cells reaching the doubling time for more than 100% were defined as senescent EPCs, of which the activity of therapeutic angiogenesis was attenuated in mouse ischemic hindlimbs. The senescent cells, in medium free of glucose and bicarbonate, showed impaired activity in migration and tube formation. Flow cytometry indicated increased content of reactive oxygen species, mitochondria, and calcium, while bioenergetic analysis showed increased oxygen consumption and reduced ATP content. Examination of mitochondrial network showed that senescence increased the length of the network and ultrastructure analysis exhibited elongated mitochondria. Immunoblotting of the senescent EPCs demonstrated decreased expression level of fission protein1 (Fis1). In rat EPCs, the Fis1 level was decreased in the animals aged 24 months or older, compared to those of 3 months. Silencing of Fis1 in the young EPCs using Fis1-specific siRNA leads to appearance of phenotype resembling those of senescent cells, including elevated oxidative stress, disturbed mitochondrial network, reduced mitochondria membrane potential, decreasing ATP content, lower proliferation activity, and loss of therapeutic potential in ischemic hindlimbs. Fis1 over-expression in senescent EPCs reduced the oxidative stress, increased the proliferation, and restored the cobble stone-like morphology, senescence, bioenergetics, angiogenic potential, and therapeutic activity.

Conclusion

In human EPCs, down-regulation of Fis1 is involved in mitochondrial dysfunction and contributes to the impaired activity of EPCs during the senescence process. Enhanced expression of Fis1 in senescent EPCs restores the youthful phenotype.

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Abbreviations

DCFDA:

2′,7′-dichlorofluorescin diacetate

DHE:

Dihydroethidine

Drp1:

Dynamin-related protein 1

EPCs:

Endothelial progenitor cells

Fis1:

Fission protein 1

MFN:

Mitofusin

mtDNA:

Mitochondrial deoxyribonucleic acid

MTR:

MitoTracker Red CMXRos

NAO:

Nonyl acridine orange

OPA1:

Optic atrophy 1

OCR:

Oxygen consumption rate

PBMCs:

Peripheral blood mononuclear cells

ROS:

Reactive oxygen species

VDAC:

Voltage-dependent anion channel

SA-β-Gal:

Senescence-associated β-galactosidase

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Acknowledgements

The authors acknowledge the supported provided from the Ministry of Science and Technology, Taiwan (Grant Nos. MOST 103-2320-B-715-004, MOST 104-2320-B-715-008-MY2, MOST 105-2632-B-715-001, MOST 106-2632-B-715-001, and MOST 107-2632-B-715-001), MacKay Memorial Hospital (Grant Nos. MMH-E-102003, MMH-MM-10703, and MMH-MM-10810), and Mackay Medical College (Grant Nos. 1041B16 and 1061B21), Taiwan.

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  1. Department of Medicine, Mackay Medical College, No. 46, Sec. 3, Zhongzheng Rd, Sanzhi District, New Taipei City, 252, Taiwan

    Hsueh-Hsiao Wang, Yih-Jer Wu, Ya-Ming Tseng, Cheng-Huang Su & Hung-I Yeh

  2. Departments of Internal Medicine and Medical Research, MacKay Memorial Hospital, Taipei, Taiwan

    Yih-Jer Wu, Cheng-Huang Su, Chin-Ling Hsieh & Hung-I Yeh

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Wang, HH., Wu, YJ., Tseng, YM. et al. Mitochondrial fission protein 1 up-regulation ameliorates senescence-related endothelial dysfunction of human endothelial progenitor cells. Angiogenesis 22, 569–582 (2019). https://doi.org/10.1007/s10456-019-09680-2

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