SFTPB in serum extracellular vesicles as a biomarker of progressive pulmonary fibrosis
Takatoshi Enomoto, et al.
Takatoshi Enomoto, et al.
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Research Article Pulmonology

SFTPB in serum extracellular vesicles as a biomarker of progressive pulmonary fibrosis

Abstract

Progressive pulmonary fibrosis (PPF), defined as the worsening of various interstitial lung diseases (ILDs), currently lacks useful biomarkers. To identify novel biomarkers for early detection of patients at risk of PPF, we performed a proteomic analysis of serum extracellular vesicles (EVs). Notably, the identified candidate biomarkers were enriched for lung-derived proteins participating in fibrosis-related pathways. Among them, pulmonary surfactant-associated protein B (SFTPB) in serum EVs could predict ILD progression better than the known biomarkers, serum KL-6 and SP-D, and it was identified as an independent prognostic factor from ILD-gender-age-physiology index. Subsequently, the utility of SFTPB for predicting ILD progression was evaluated further in 2 cohorts using serum EVs and serum, respectively, suggesting that SFTPB in serum EVs but not in serum was helpful. Among SFTPB forms, pro-SFTPB levels were increased in both serum EVs and lungs of patients with PPF compared with those of the control. Consistently, in a mouse model, the levels of pro-SFTPB, primarily originating from alveolar epithelial type 2 cells, were increased similarly in serum EVs and lungs, reflecting pro-fibrotic changes in the lungs, as supported by single-cell RNA sequencing. SFTPB, especially its pro-form, in serum EVs could serve as a biomarker for predicting ILD progression.

Authors

Takatoshi Enomoto, Yuya Shirai, Yoshito Takeda, Ryuya Edahiro, Shigeyuki Shichino, Mana Nakayama, Miho Takahashi-Itoh, Yoshimi Noda, Yuichi Adachi, Takahiro Kawasaki, Taro Koba, Yu Futami, Moto Yaga, Yuki Hosono, Hanako Yoshimura, Saori Amiya, Reina Hara, Makoto Yamamoto, Daisuke Nakatsubo, Yasuhiko Suga, Maiko Naito, Kentaro Masuhiro, Haruhiko Hirata, Kota Iwahori, Izumi Nagatomo, Kotaro Miyake, Shohei Koyama, Kiyoharu Fukushima, Takayuki Shiroyama, Yujiro Naito, Shinji Futami, Yayoi Natsume-Kitatani, Satoshi Nojima, Masahiro Yanagawa, Yasushi Shintani, Mari Nogami-Itoh, Kenji Mizuguchi, Jun Adachi, Takeshi Tomonaga, Yoshikazu Inoue, Atsushi Kumanogoh

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Figure 4

Reproducibility of associations between SFTPB in serum EVs but not in serum and ILD progression.

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Reproducibility of associations between SFTPB in serum EVs but not in se...
(A and E) Schematic representation of the project designs in the validation cohort (A) and in the combined-sample cohort (E). In the validation cohort, serum EV levels of SFTPB were analyzed by data-independent acquisition. In the combined-sample cohort, serum levels of SFTPB were measured by ELISA. (B) Serum EV levels of SFTPB in the validation cohort. Numbers of samples: HC (n = 23), non-PPF (n = 20), PPF (n = 14). The data were subjected to ANOVA, and Holm’s method was applied to adjust for the ANOVA P values. (F) Serum levels of SFTPB in the combined-sample cohort. Numbers of samples: HC (n = 49), BA (n = 20), COPD (n = 20), LC (n = 22), NTM (n = 15), non-PPF (n = 100), and PPF (n = 60). The expression levels were compared by ANOVA, and Dunnett’s method was applied to adjust for the ANOVA P values. Subsequently, differences between PPF and non-PPF were compared by ANOVA, and Holm’s method was applied to adjust for the ANOVA P values. (B and F) *P < 0.05, **P < 0.01, and ***P < 0.001 for significant differences from healthy control. N.S., no significant difference between non-PPF and PPF. (C and G) ROC curves for evaluating SFTPB in serum EVs as predicting composite outcome (relative decline in %FVC ≥ 10%, acute exacerbation, or death) within a year in 23 evaluable ILD cases in the validation cohort (C) and ROC curves for evaluating SFTPB in serum as predicting composite outcome in 78 evaluable ILD cases in the combined-sample cohort (G). (D and H) Kaplan-Meier curves estimating the probability of overall survival (OS) stratified by the serum EV levels of SFTPB in the validation cohort (D) and the serum levels of SFTPB in the combined-sample cohort (H). OS was defined as the period from the date of blood collection to the date of death from any cause. BA, bronchial asthma; COPD, chronic obstructive pulmonary disease; LC, lung cancer; NTM, nontuberculous mycobacterial lung disease.