Multiplexed Molecular Imaging Strategy Integrated with RNA Sequencing in the Assessment of the Therapeutic Effect of Wharton's Jelly Mesenchymal Stem Cell-Derived Extracellular Vesicles for Osteoporosis

Cheng-Hsiu Lu; Yi-An Chen; Chien-Chih Ke; Sain-Jhih Chiu; Fong-Shya Jeng; Chao-Cheng Chen; Ya-Ju Hsieh; Bang-Hung Yang; Chi-Wei Chang; Feng-Sheng Wang; Ren-Shyan Liu

doi:10.2147/IJN.S335757

Multiplexed Molecular Imaging Strategy Integrated with RNA Sequencing in the Assessment of the Therapeutic Effect of Wharton's Jelly Mesenchymal Stem Cell-Derived Extracellular Vesicles for Osteoporosis

Int J Nanomedicine. 2021 Nov 30:16:7813-7830. doi: 10.2147/IJN.S335757. eCollection 2021.

Authors

Cheng-Hsiu Lu^{1

2

3}, Yi-An Chen^{4

5}, Chien-Chih Ke^{6

7

8}, Sain-Jhih Chiu⁵, Fong-Shya Jeng⁵, Chao-Cheng Chen⁴, Ya-Ju Hsieh^{6

7

8}, Bang-Hung Yang^{4

9}, Chi-Wei Chang⁹, Feng-Sheng Wang^{2

3

10

11}, Ren-Shyan Liu^{1

4

9

12}

Affiliations

¹ Industrial Ph.D. Program of Biomedical Science and Engineering, National Yang Ming Chiao Tung University, Taipei, Taiwan.
² Core Facility for Phenomics and Diagnostics, Kaohsiung Chang Gung Memorial Hospital, Kaohsiung, Taiwan.
³ Department of Medical Research, Kaohsiung Chang Gung Memorial Hospital, Kaohsiung, Taiwan.
⁴ Department of Biomedical Imaging and Radiological Sciences, National Yang Ming Chiao Tung University, Taipei, Taiwan.
⁵ Molecular and Genetic Imaging Core/Taiwan Mouse Clinic, National Comprehensive Mouse Phenotyping and Drug Testing Center, Taipei, Taiwan.
⁶ Department of Medical Imaging and Radiological Sciences, Kaohsiung Medical University, Kaohsiung, Taiwan.
⁷ Drug Development and Value Creation Research Center, Kaohsiung Medical University, Kaohsiung, Taiwan.
⁸ Department of Medical Research, Kaohsiung Medical University Hospital, Kaohsiung, Taiwan.
⁹ PET Center, Department of Nuclear Medicine, Taipei Veterans General Hospital, Taipei, Taiwan.
¹⁰ Center for Mitochondrial Research and Medicine, Kaohsiung Chang Gung Memorial Hospital, Kaohsiung, Taiwan.
¹¹ Graduate Institute of Clinical Medical Science, Chang Gung University College of Medicine, Kaohsiung, Taiwan.
¹² Department of Nuclear Medicine, Cheng Hsin Hospital, Taipei, Taiwan.

Abstract

Introduction: Osteoporosis is a result of an imbalance in bone remodeling. Mesenchymal stem cell-derived extracellular vesicles (MSC-EVs) have been considered as a potentially promising treatment for osteoporosis. However, the therapeutic effect, genetic alterations, and in vivo behavior of exogenous EVs for osteoporosis in mice models remain poorly understood.

Methods: A multiplexed molecular imaging strategy was constructed by micro-positron emission tomography (µPET)/computed tomography (CT), µCT, and optical imaging modality which reflected the osteoblastic activity, microstructure, and in vivo behavior of EVs, respectively. RNA sequencing was used to analyze the cargo of EVs, and the bone tissues of ovariectomized (OVX) mice post EV treatment.

Results: The result of [¹⁸F]NaF µPET showed an increase in osteoblastic activity in the distal femur of EV-treated mice, and the bone structural parameters derived from µCT were also improved. In terms of in vivo behavior of exogenous EVs, fluorescent dye-labeled EVs could target the distal femur of mice, whereas the uptakes of bone tissues were not significantly different between OVX mice and healthy mice. RNA sequencing demonstrated upregulation of ECM-related genes, which might associate with the PI3K/AKT signaling pathway, in line with the results of microRNA analysis showing that mir-21, mir-29, mir-221, and let-7a were enriched in Wharton's jelly-MSC-EVs and correlated to the BMP and PI3K/AKT signaling pathways.

Conclusion: The therapeutic effect of exogenous WJ-MSC-EVs in the treatment of osteoporosis was successfully assessed by a multiplexed molecular imaging strategy. The RNA sequencing demonstrated the possible molecular targets in the regulation of bone remodeling. The results highlight the novelty of diagnostic and therapeutic strategies of EV-based treatment for osteoporosis.

Keywords: RNA sequencing; Wharton’s jelly mesenchymal stem cells; [18F]NaF; extracellular vesicles; osteoporosis.

MeSH terms

Animals
Extracellular Vesicles*
Mesenchymal Stem Cells*
Mice
MicroRNAs*
Molecular Imaging
Osteoporosis* / diagnostic imaging
Osteoporosis* / therapy
Phosphatidylinositol 3-Kinases
Sequence Analysis, RNA
Wharton Jelly*

Substances

MIRN29 microRNA, mouse
MicroRNAs

Grants and funding

This work was financially supported by Ministry of Science and Technology, Taiwan (MOST 110-2314-B-350-002) and Molecular and Genetic Imaging Center, National Yang Ming Chiao Tung University and Taiwan Animal Consortium (MOST 110‐2740‐B‐001‐002), and Kaohsiung Medical University Research Foundation (KMU-DK110005 and KMU-M11010).