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Persistent Abnormal Immunocytes Induced Systemic Bone Loss in Locally Irradiated Rats

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Abstract

Chronic and systemic bone complications frequently occur in patients who undergo radiotherapy; however, the pathological mechanisms underlying these complications remain unclear. This study aimed to observe persistent and systemic changes in locally irradiated rats and to determine the systemic pathological changes that persistently affect bone metabolism. We examined the inflammatory and oxidative stress responses that occurred after local irradiation using enzyme immunoassays and biochemical analyses. Lymphocytes obtained from the blood, spleen, thymus, and bone marrow were evaluated using flow cytometry. The proliferation and apoptosis characteristics of co-cultured bone marrow-derived mesenchymal stem cells (BMSCs) were detected by MTT assay and PI/Annexin V-FITC staining, respectively, and the differentiation of BMSCs was measured according to alkaline phosphatase (ALP) staining, alizarin red staining, and Oil Red O staining and by evaluating the mRNA expression of ALP, osteocalcin (OCN), osteopontin (OPN), collagen I, Runx2, and PPARγ. Our results revealed that no significant or continuous differences were present in the inflammatory response or the oxidative stress response throughout the body after local irradiation. B lymphocyte levels increased continuously in the blood, spleen, and bone marrow after local irradiation. T lymphocyte levels were decreased at 2 weeks after local irradiation, and CD8+T lymphocyte levels were increased in the blood, thymus, and bone marrow at 12 weeks after local irradiation. The ratio of CD4+/CD8+T lymphocytes began to decrease during the early phase after local irradiation and became significantly decreased at 12 weeks after local irradiation. Normal BMSCs co-cultured with lymphocytes derived from irradiated rats exhibited decreased proliferation and increased apoptosis, and the ALP staining intensity, alizarin red staining intensity, and mRNA expression of related genes were all also decreased. Oil Red O staining intensity and mRNA expression of PPARγ were both increased. Lymphocyte levels contribute to chronic and systemic bone complications after radiotherapy by inhibiting the proliferation and osteoblastogenesis of BMSCs.

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Abbreviations

TNF-α:

Tumor necrosis factor-α

IL-6:

Interleukin-6

IFN-γ:

Interferon-γ

IL-1α:

Interleukin-1α

IL-1β:

Interleukin-1β

MCP-1:

Monocyte chemoattractant protein 1

MIP:

Macrophage inflammatory protein

SOD:

Superoxide dismutase

CAT:

Catalase

MDA:

Malondialdehyde

GSH-Px:

Glutathione peroxidase

T-AOC:

Total antioxidant capacity

BMSCs:

Bone marrow-derived mesenchymal stem cells

RANKL:

Receptor activator of nuclear factor kappa B ligand

BV/TV:

Ratio of the segmented bone volume to the total volume

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Acknowledgements

We acknowledge financial support from the Shanghai Municipal Health Commission (No.201740028) and National Natural Science Foundation of China (No.81102071).

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Authors and Affiliations

  1. Department of Geriatrics and Gerontology, Huadong Hospital, Fudan University, 221 West Yan’an Road, Shanghai, 200040, China

    Wei Hong

  2. Shanghai Key Laboratory of Clinical Geriatric Medicine, Huadong Hospital, Fudan University, 221 West Yan’an Road, Shanghai, 200040, China

    Wei Hong

  3. Department of Breast Surgery, Shanghai Cancer Hospital, Fudan University, 270 Dongan Road, Shanghai, 200032, China

    Lichen Tang

  4. Department of General Surgery, Huadong Hospital, Fudan University, 221 West Yan’an Road, Shanghai, 200040, China

    Rui Ge, Weiping Li & Lixia Hong

  5. Department of Thoracic Surgery, Huadong Hospital, Fudan University, 221 West Yan’an Road, Shanghai, 200040, China

    Xiaoyong Shen

  6. Department of Radiation Biology, Institute of Radiation Medicine, Fudan University, 2094 Xietu Road, Shanghai, 200032, China

    Xiaoya Xu

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  1. Wei Hong
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  7. Xiaoya Xu
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Contributions

XX and WH designed the study and did the cell culture and answered all the questions of the reviewers; LT and XX performed the ELISA and flow cytometry experiments; WL, RG and LH contributed to the animal experiments; XX and XS analyzed the data; and WH wrote the manuscript. All authors have read and approved the final manuscript to be published.

Corresponding authors

Correspondence to Wei Hong or Xiaoya Xu.

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Conflict of interest

Wei Hong, Lichen Tang, Rui Ge, Weiping Li, Xiaoyong Shen, Lixia Hong and Xiaoya Xu declare that they have no conflict of interest.

Ethical Approval

Ethical approval was provided by the Committee on Animal Resources of Fudan University.

Informed Consent

For this type of study, no informed consent is required.

Human and Animal Rights

All procedures performed in studies involving animals were in accordance with the ethical standards of the Institutional Animal Ethics Committee of Fudan University at which the studies were conducted (NO.2012-03-FYS-GJJ-01).

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Hong, W., Tang, L., Ge, R. et al. Persistent Abnormal Immunocytes Induced Systemic Bone Loss in Locally Irradiated Rats. Calcif Tissue Int 109, 706–718 (2021). https://doi.org/10.1007/s00223-021-00883-8

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  • DOI: https://doi.org/10.1007/s00223-021-00883-8

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