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Stain-Free Quantitative Two-Photon Fiber Histology for Streamlined Pathological Diagnosis in Interstitial Lung Diseases
40 Pages Posted: 14 May 2024
More...Abstract
Interstitial lung disease (ILD), characterized by inflammation and fibrosis, often exhibits low diagnostic accuracy and consistency. Traditional H&E staining provides limited visualization of fibrosis. In contrast, two-photon microscopy enables simultaneous, label-free imaging of collagen and elastic fibers, enhancing the depiction of fibrosis. In this paper, we introduce a pioneering, stain-free quantitative Two-Photon Fiber Histology (TPFH) technique into the traditional diagnostic process based on H&E staining. This innovative method offers precise, co-localized visualization of collagen, elastin fibers, and cells, significantly improving the identification of fibrotic ILD (f-ILD) subtypes. Our study demonstrated that fibrotic characteristics such as type, distribution, and degree of fibrotic proliferation can effectively distinguish between subtypes. We rigorously tested this approach in a blinded trial involving seven pathologists, who assessed 14 patients with normal lung areas and 31 with ILD (11 idiopathic pulmonary fibrosis/usual interstitial pneumonia (IPF/UIP), 14 nonspecific interstitial pneumonia (NSIP), and 6 pleuroparenchymal fibroelastosis (PPFE)). TPFH significantly improved diagnostic consistency (kappa values 0.56 to 0.72) and accuracy (74% to 82%). Our customized algorithm extracted quantitative fiber indicators that effectively aided f-ILD subtype differentiation. Indices like CEFAR, ASW, and collagen fiber heterogeneity differentiated normal and UIP from cellular and fibrotic NSIP, with area under the receiver operating characteristic curve (AUC) scores of 0.98, 0.92, and 0.93, respectively. The EAR index was particularly effective in distinguishing PPFE, with an AUC of 0.93. TPFH significantly enhances diagnostic accuracy and consistency, offering valuable support for differential diagnosis by adding just a few minutes of two-photon imaging to the standard diagnostic process.
Funding: The research was supported by the National High-Level Hospital Clinical Research Funding of China No. 2022-NHLHCRF-LX-01-0206.
Declaration of Interest: The authors declare that they have no conflict of interest.
Ethical Approval: Informed consent was obtained from each participating patient. Ethical approval and experimental procedures were granted by the Clinical Research Ethics Committee of the China-Japan Friendship Hospital (2023-key-023).
Keywords: Stain-free two-photon fiber histology, quantitative analysis, fibrotic interstitial lung diseases, diagnosis and differential diagnosis
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