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Aggravated effects of human parvovirus B19 NS1 protein on bleomycin‑induced pulmonary fibrosis

  • Authors:
    • Tsai-Ching Hsu
    • Chih-Chen Tzang
    • Chia-Wei Kuo
    • Zhi-Han Wen
    • Der-Yuan Chen
    • Bor-Show Tzang
  • View Affiliations / Copyright

    Affiliations: Institute of Medicine, Chung Shan Medical University, Taichung 402, Taiwan, R.O.C., School of Medicine, College of Medicine, National Taiwan University, Taipei City 100, Taiwan, R.O.C.
    Copyright: © Hsu et al. This is an open access article distributed under the terms of Creative Commons Attribution License.
  • Article Number: 69
    |
    Published online on: December 16, 2025
       https://doi.org/10.3892/mmr.2025.13779
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Abstract

Interstitial lung diseases (ILDs) include various lung parenchymal disorders characterized by inflammation and fibrosis of the lung tissue, leading to progressive dyspnea and respiratory failure. Clinical evidence has suggested an association between human parvovirus B19 (B19V) infection and the progression of ILD and pulmonary fibrosis, but the mechanisms involved remain unclear. The present study screened 86 patients with connective tissue disease (CTD) and reported that B19V infection was significantly more prevalent among those with ILD than among those without (P<0.001). To investigate the potential underlying mechanisms, a bleomycin (BLM)‑treated mouse model was employed to assess the effect of B19V nonstructural protein 1 (NS1) on pulmonary fibrosis. Mice treated with BLM or BLM + NS1 exhibited markedly higher fibrosis scores, hydroxyproline content, and higher levels of transforming growth factor‑β and collagen I. Treatment with nintedanib attenuated fibrosis in both groups; however, lung fibrosis remained more pronounced in the BLM + NS1 group than in the BLM group. Furthermore, the levels of neutrophil‑associated markers, including citrullinated histone H3 and myeloperoxidase, as well as inflammasome‑related factors, such as IL‑18 and IL‑17A, were markedly elevated in lung tissues from both groups, with the highest levels observed in the BLM + NS1 group. These findings suggested that B19‑NS1 may exacerbate fibrosis in patients with ILD by increasing neutrophil‑driven responses and inflammasome activation, highlighting a need for nintedanib therapies to more effectively address B19V‑associated pulmonary fibrosis.
View Figures

Figure 1

Effects of nintedanib and B19V NS1 on
bleomycin-induced pulmonary fibrosis in mice. (A) Schematic diagram
of the experimental animal model. (B) Daily body weight
measurements across the PBS, BLM, BLM + nintedanib, BLM + NS1, and
BLM + NS1 + nintedanib groups (BLM vs. PBS, P=0.008; BLM +
nintedanib vs. PBS, P=0.002; BLM + NS1 vs. PBS, P<0.001; BLM +
nintedanib + NS1 vs. PBS, P<0.001; BLM + NS1 vs. BLM, P=0.031;
BLM + nintedanib + NS1 vs. BLM, P=0.048; BLM + nintedanib + NS1 vs.
BLM + nintedanib, P<0.001). (C) Lung weight of mice on day 19
(BLM vs. PBS, P=0.025; BLM + NS1 vs. PBS, P=0.037; BLM + nintedanib
vs BLM, P=0.035; BLM + NS1 + nintedanib vs BLM + NS1, P=0.044).
*P<0.05 vs. the PBS (Control) group, #P<0.05 vs.
the BLM group, and &P<0.05 vs. the BLM + NS1
group. B19V, human parvovirus B19; NS1, nonstructural protein 1;
BLM, bleomycin.

Figure 2

Proportions of immune cells in BALF
of mice. (A) Percentages of macrophage, neutrophil, and lymphocyte
in BALF of mice from different groups (Macrophages: BLM vs. PBS,
P<0.001; BLM + nintedanib vs. PBS, P<0.001; BLM + NS1 vs.
PBS, P=0.02; BLM + nintedanib + NS1 vs. PBS, P=0.005; BLM + NS1 vs.
BLM, P<0.001; Neutrophils: BLM vs. PBS, P<0.001; BLM +
nintedanib vs. PBS, P<0.001; BLM + NS1 vs. PBS, P<0.001; BLM
+ nintedanib + NS1 vs. PBS, P<0.001; BLM + NS1 vs. BLM, P=0.01).
The concentrations of (B) TNF-α (BLM vs. PBS, P=0.014; BLM + NS1
vs. PBS, P=0.003; BLM + nintedanib vs. BLM, P=0.039; BLM +
nintedanib + NS1 vs. BLM + NS1 P=0.009), (C) KL-6 (BLM vs. PBS,
P<0.001; BLM + NS1 vs. PBS, P<0.001; BLM + nintedanib vs.
BLM, P<0.001; BLM + nintedanib + NS1 vs. BLM + NS1 P<0.001)
and (D) IL-6 (BLM vs. PBS, P= 0.039; BLM + NS1 vs. PBS, P=0.009;
BLM + nintedanib vs. BLM, P=0.013; BLM + nintedanib + NS1 vs. BLM +
NS1, P=0.014) in BALF of mice from different groups. *P<0.05 vs.
the PBS (Control) group, #P<0.05 vs. the BLM group,
and &P<0.05 vs. the BLM + NS1 group. BALF,
bronchoalveolar lavage fluid; BLM, bleomycin; NS1, nonstructural
protein 1; IL, interleukin.

Figure 3

Histological staining of the lung
tissues of mice. (A) Representative images of H&E and Masson's
trichrome-stained mouse lung sections. Scale bar, 100 µm.
Quantified result of (B) fibrosis score (BLM vs. PBS, P<0.05;
BLM + NS1 vs. PBS, P<0.001) and (C) hydroxyproline content (BLM
vs. PBS, P<0.001; BLM + NS1 vs. PBS, P<0.001; BLM +
nintedanib vs. BLM, P<0.001; BLM + NS1 vs. BLM, P= P=0.016; BLM
+ nintedanib + NS1 vs. BLM + NS1, P=0.036; BLM + NS1 + nintedanib
vs. BLM + nintedanib, P=0.006). *P<0.05 vs. the PBS (Control)
group, #P<0.05 vs. the BLM group,
&P<0.05 vs. the BLM + NS1 group, and
§P<0.05 vs. the BLM + nintedanib group. H&E,
hematoxylin and eosin; BLM, bleomycin; NS1, nonstructural protein
1.

Figure 4

IHC staining of lung tissue of mice.
(A) Representative images of lung sections of mice stained with
TGF-β and collagen I. Scale bar, 100 µm. Quantified results of (B)
TGF-β (BLM vs. PBS, P<0.001; BLM + NS1 vs. PBS, P<0.001; BLM
+ nintedanib vs. BLM, P=0.001; BLM + NS1 vs. BLM, P=0.015; BLM +
nintedanib + NS1 vs. BLM + NS1, P<0.001) and (C) collagen I
positive cells (BLM vs. PBS, P=0.019; BLM + NS1 vs. PBS,
P<0.001; BLM + nintedanib vs. BLM, P=0.024; BLM + NS1 vs. BLM,
P<0.001; BLM + nintedanib + NS1 vs. BLM + NS1, P<0.001) in
lung sections of mice. *P<0.05 vs. the PBS (Control) group,
#P<0.05 vs. the BLM group, and
&P<0.05 vs. the BLM + NS1 group. IHC,
immunohistochemistry; TGF transforming growth factor; BLM,
bleomycin; NS1, nonstructural protein 1.

Figure 5

Expressions of neutrophil-related and
inflammasome markers. (A) Representative immunohistochemistry
images of the lung section of mice stained with (A) Cit-H3, MPO,
IL-18 and IL-17. Scale bar, 100 µm. Quantification of (B) Cit-H3
(BLM vs. PBS, P<0.001; BLM + NS1 vs. PBS, P<0.001; BLM +
nintedanib vs. BLM, P=0.013; BLM + NS1 vs. BLM, P<0.001; BLM +
nintedanib + NS1 vs. BLM + NS1, P<0.001), (C) MPO (BLM vs. PBS,
P=0.001; BLM + NS1 vs. PBS, P<0.001; BLM + nintedanib vs. BLM,
P<0.001; BLM + NS1 vs. BLM, P= P<0.001; BLM + nintedanib +
NS1 vs. BLM + NS1, P<0.001; BLM + nintedanib + NS1 vs. BLM +
nintedanib, P=0.015), (D) IL-18 (BLM vs. PBS, P<0.001; BLM + NS1
vs. PBS, P<0.001; BLM + nintedanib vs. BLM, P=0.043; BLM + NS1
vs. BLM, P<0.001; BLM + nintedanib + NS1 vs. BLM + NS1,
P<0.001), and (E) IL-17 (BLM vs. PBS, P<0.001; BLM + NS1 vs.
PBS, P<0.001; BLM + nintedanib vs. BLM, P<0.03; BLM + NS1 vs.
BLM, P<0.001; BLM + nintedanib + NS1 vs. BLM + NS1, P<0.001)
positive cells in lung sections of mice. *P<0.05 vs. the PBS
(Control) group, #P<0.05 vs. the BLM group,
&P<0.05 vs. the BLM + NS1 group, and
§P<0.05 vs. the BLM + nintedanib group. Cit-H3,
citrullinated histone H3; MPO, myeloperoxidase; IL, interleukin;
BLM, bleomycin; NS1, nonstructural protein 1.
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Copy and paste a formatted citation
Spandidos Publications style
Hsu T, Tzang C, Kuo C, Wen Z, Chen D and Tzang B: Aggravated effects of human parvovirus B19 NS1 protein on bleomycin‑induced pulmonary fibrosis. Mol Med Rep 33: 69, 2026.
APA
Hsu, T., Tzang, C., Kuo, C., Wen, Z., Chen, D., & Tzang, B. (2026). Aggravated effects of human parvovirus B19 NS1 protein on bleomycin‑induced pulmonary fibrosis. Molecular Medicine Reports, 33, 69. https://doi.org/10.3892/mmr.2025.13779
MLA
Hsu, T., Tzang, C., Kuo, C., Wen, Z., Chen, D., Tzang, B."Aggravated effects of human parvovirus B19 NS1 protein on bleomycin‑induced pulmonary fibrosis". Molecular Medicine Reports 33.2 (2026): 69.
Chicago
Hsu, T., Tzang, C., Kuo, C., Wen, Z., Chen, D., Tzang, B."Aggravated effects of human parvovirus B19 NS1 protein on bleomycin‑induced pulmonary fibrosis". Molecular Medicine Reports 33, no. 2 (2026): 69. https://doi.org/10.3892/mmr.2025.13779
Copy and paste a formatted citation
x
Spandidos Publications style
Hsu T, Tzang C, Kuo C, Wen Z, Chen D and Tzang B: Aggravated effects of human parvovirus B19 NS1 protein on bleomycin‑induced pulmonary fibrosis. Mol Med Rep 33: 69, 2026.
APA
Hsu, T., Tzang, C., Kuo, C., Wen, Z., Chen, D., & Tzang, B. (2026). Aggravated effects of human parvovirus B19 NS1 protein on bleomycin‑induced pulmonary fibrosis. Molecular Medicine Reports, 33, 69. https://doi.org/10.3892/mmr.2025.13779
MLA
Hsu, T., Tzang, C., Kuo, C., Wen, Z., Chen, D., Tzang, B."Aggravated effects of human parvovirus B19 NS1 protein on bleomycin‑induced pulmonary fibrosis". Molecular Medicine Reports 33.2 (2026): 69.
Chicago
Hsu, T., Tzang, C., Kuo, C., Wen, Z., Chen, D., Tzang, B."Aggravated effects of human parvovirus B19 NS1 protein on bleomycin‑induced pulmonary fibrosis". Molecular Medicine Reports 33, no. 2 (2026): 69. https://doi.org/10.3892/mmr.2025.13779
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