Abstract
Elastic staining may aid in the diagnosis of pulmonary non-mucinous adenocarcinoma and its in situ lesions. This study evaluated elastic staining patterns and their diagnostic utility in a multi-cohort series. Elastic staining was performed in 120 lesions, including a principal cohort (n = 60), an internal validation cohort (n = 30), an external validation cohort (n = 30), and an interobserver agreement cohort (n = 40). Staining patterns were analyzed and their impact on diagnostic accuracy was assessed. Four elastic staining patterns were identified: a point-and-linear pattern, an intensified point-and-linear pattern, a disorderly thickened and recoiled pattern with continuous and discontinuous variants, and a decreased or disappeared pattern. All adenocarcinoma in situ lesions and the in situ components of minimally invasive adenocarcinoma and invasive adenocarcinoma demonstrated the continuous disorderly thickened and recoiled pattern. The decreased or disappeared pattern was observed in the majority of invasive adenocarcinomas and in all invasive components of minimally invasive adenocarcinoma, whereas a subset of invasive adenocarcinomas exhibited the discontinuous variant associated with extensive carcinomatous collapse. Non-neoplastic lesions displayed a point-and-linear pattern, with an intensified point-and-linear pattern observed in regions of iatrogenic or mechanical collapse. No significant differences were identified between validation cohorts (P > 0.05). Diagnostic accuracy and histologic identification improved with the use of elastic staining among both trainee and general pathologists, and interobserver agreement was high (Kappa = 0.945). Overall, elastic staining represents a useful adjunct for improving diagnostic accuracy and histologic classification of pulmonary non-mucinous adenocarcinoma and its in situ lesions.
Introduction
Lung cancer is one of the most common malignant tumors worldwide.1,2 Primary lung cancers are primarily classified into small cell lung cancer (SCLC) and non-small cell lung cancer (NSCLC). 3 In recent decades, adenocarcinoma has become the most prevalent type. 4 Various etiological factors and genetic alterations contribute to its pathogenesis, from precursor lesions to fully developed malignancies. 5 Accurate diagnosis still depends on pathological evaluation. However, the diverse and complex growth patterns of these tumors often lead to diagnostic challenges, particularly in distinguishing in situ from invasive components in certain contexts.
Elastin fibers, composed of elastin bundles, are a crucial component of the extracellular matrix (ECM), providing mesenchymal tissues with resilience, long-range deformability, and passive recoil.6,7 These fibers are abundant in organs and tissues such as the lungs, arteries, skin, tendons, and ligaments. 8 Elastin is essential for normal lung development and function. 9 Pathologically, elastic staining is used to assess pleural or alveolar invasion by malignancies.10,11 However, its application in adenocarcinomas and their precursor lesions is complicated by marked morphologic variability and therefore remains uncommon in routine diagnostic practice. While a few studies, including that by Thunnissen et al, have explored elastic staining in pulmonary pathology, detailed characterization of elastic staining patterns across adenocarcinoma subtypes and their in situ lesions remains limited.10,12,13 This study aims to provide a systematic, pattern-based analysis of elastic staining features in pulmonary non-mucinous adenocarcinoma and associated in situ lesions, and to evaluate their diagnostic utility across different histologic contexts and cohorts.
Materials and Methods
Study Design and Patient Selection
This retrospective study was designed as a multi-cohort analysis to evaluate elastic staining patterns in pulmonary non-mucinous adenocarcinoma and associated precursor lesions. All patients were diagnosed according to current WHO classification criteria and met the same predefined inclusion and exclusion criteria, including the availability of evaluable elastic-stained sections. 14 Based on study purpose, patients were allocated into a principal cohort, internal validation cohort, external validation cohort, and an interobserver concordance cohort, as detailed below.
Principal Cohort
This study was approved by the Institutional Review Board of The Second Peoples Hospital of Yibin, Department of Pathology (YBEY-P26002). The requirement for informed consent was waived due to the retrospective and minimal-risk nature of the study. Personal information from each sample was anonymized and handled in accordance with the principles of the Declaration of Helsinki. We retrospectively collected formalin-fixed, paraffin-embedded (FFPE) tissue specimens from 55 lesions with pulmonary adenocarcinoma and its precursor lesions, collected over a 3-year period, as the principal cohort. All invasive adenocarcinomas (IAs) included in the study were non-mucinous, given the rarity of other subtypes. An additional 5 lesions with non-neoplastic lesions (NNLs) with varying degrees of iatrogenic or mechanical collapse (IC/MC) were also included.
Pathologic Evaluation, Immunohistochemistry (IHC), and Elastic Staining
All representative slides were re-stained with hematoxylin–eosin (H&E) and independently reviewed by two senior pathologists (J.Z. and Q.S.). Evaluating pathologists were blinded to the original diagnoses and relevant clinical information during independent assessment. Pathological diagnoses included atypical adenomatous hyperplasia (AAH), adenocarcinoma in situ (AIS), minimally invasive adenocarcinoma (MIA), and IA. Morphological patterns of IA were documented, encompassing lepidic, acinar, papillary, and solid types.15,16 Two collapse patterns, cancer-related collapse (CRC) and iatrogenic/mechanical collapse, were also delineated within this cohort.10,17,18 Among the 47 resection specimens, a subset of tumors had undergone preoperative biopsy. Biopsy-related tissue alterations, including focal collapse or distortion of alveolar structures, were carefully recognized based on histologic context. These changes were distinguished from tumor-related elastic fiber alterations and from iatrogenic or mechanical collapse related to specimen handling, and were not used for the assessment of invasion-related elastic staining patterns. Lesions with background lung diseases, including emphysema or smoking-related interstitial fibrosis, were reviewed with careful correlation to hematoxylin–eosin morphology. Elastic fiber alterations attributable to background lung remodeling were distinguished from invasion-related changes during diagnostic assessment. Lesions in which background lung disease precluded reliable interpretation were not encountered in the present cohort. Sections of 4-μm thickness were obtained from 10% formalin-fixed, paraffin-embedded tissue blocks, followed by immunohistochemical staining using commercially available antibodies when necessary: keratin 7 (EP16, 1:200; ZSGB-BIO, Beijing, China), TTF-1 (also known as NKX2-1; 8G7G3/1, prediluted, Dako, Carpinteria, CA, USA), and Napsin A (polyclonal; prediluted, Ventana, Tucson, AZ, USA). An automated immunohistochemistry system (Ventana, Roche, Tucson, AZ, USA) was employed following the manufacturer's protocol. Elastic staining was performed on each selected slide using an Elastic Stain Kit (Verhoeff-Van Gieson, Baso, Zhuhai, China) as per the manufacturer's instructions, where elastic fibers appear blue to black, and collagen exhibits a red coloration. The results of immunohistochemistry and elastic staining were interpreted by Dr. J.Z. and Prof. X.Z..
Internal and Independent External Validation Cohort
An additional 30 consecutively collected resection lesions from the same institution, obtained during a subsequent period, were included as the internal validation cohort. The external validation cohort consisted of 30 consecutively collected resection lesions obtained from an independent institution and was approved by the Institutional Review Board of the Department of Pathology, Jinling Hospital, School of Medicine, Nanjing University. The same predefined inclusion and exclusion criteria as those applied to the principal cohort were uniformly used for both the internal and external validation cohorts. Immunohistochemical staining for keratin 7 and TTF-1 was performed in selected lesions when indicated, and elastic staining was applied to all lesions. The staining reagents, protocols, and evaluation criteria were identical to those used in the principal cohort. Diagnostic re-evaluation by a panel of pathologists (Prof. X.Z., Dr. Q.S., and X.W.), incorporating hematoxylin–eosin morphology, immunohistochemistry, and elastic staining, served as the reference diagnosis. Dr J.Z. independently reviewed all slides under identical conditions for comparison.
Interobserver Agreement
The interobserver reproducibility in the interpretation of elastic staining was assessed by two pathologists: a general pathologist (W.Z.) who routinely handles all types of histologic specimens without a specific focus on thoracic pathology, and a second-year surgical pathology trainee (A.W.). Each pathologist underwent a brief training session on the interpretation of elastic staining in pulmonary adenocarcinomas and in situ lesions by reviewing a teaching set of 10 representative lesions. They then independently reviewed a test set of 40 randomly selected lesions (biopsy, n = 5; resection, n = 35), confirmed by thoracic pathologists (Prof. X.Z.), which included H&E, IHC, and elastic staining slides.
Data Analysis
Diagnostic accuracy and histological identification were calculated using standard formulas. Diagnostic accuracy was defined as concordance between the evaluating pathologist's diagnosis and the reference diagnosis established by consensus review. Differences between any two observation groups were evaluated using the Chi-squared test. Interobserver agreement was statistically assessed using Fleiss’ or Cohen's generalized kappa. All statistical analyses were performed using IBM SPSS Statistics, version 26.0 (IBM Corp., Armonk, NY, USA).
Results
Morphologic Features of the Principal Cohort
The reviewed principal cohort included 13 core-needle biopsies and 47 surgical specimens, comprising 5 NNLs and 55 neoplastic lesions (Table 1). The initial diagnoses of AAH, AIS, MIA, and IA were 2, 3, 12, and 38 patients, respectively. However, the final diagnostic proportions changed to AAH (3/55), AIS (5/55), MIA (10/55), and IA (37/55). Some lesions initially classified as minimally invasive or IA were reclassified as AIS based on distinct elastic staining patterns, particularly the collapse of the lepidic pattern with interstitial fibrosis. In contrast, some lesions with IA were reclassified from MIA due to the more apparent papillary features. Figure 1 summarizes the elastic staining patterns observed across different entities. The detailed features of elastic fiber staining patterns across different lesions and histologic growth patterns are summarized in Supplementary Table S1. These patterns represent recurring alterations in elastic fibers observed across the spectrum of non-neoplastic lung tissue, precursor lesions, and IA in this cohort. In general, several distinct staining patterns were identified (Figure 1): (a) a point-and-linear pattern (PL), a normal staining pattern characterized by discontinuous linear figures and pointed condensations (supporting rings) representing alveolar walls and openings (Figure 1A); (b) an intensified point-and-linear pattern (IPL), showing increased intensity and extent of point-and-linear staining (Figure 1B); (c) a disorderly thickened and recoiled pattern, characterized by non-parallel and apparently recoiled elastic fibers, with continuous (CDTR; Figure 1C) and discontinuous (DDTR; Figure 1D) variants; and (d) a decreased or disappeared pattern (D/D), characterized by a marked reduction in elastic staining resulting from effacement of the alveolar wall–associated elastic fiber framework (Figure 1E, F). These patterns identified in the principal cohort were subsequently applied, without modification, to the internal and external validation cohorts for independent assessment. All lesions with AAH (100%, 3/3) exhibited relatively increased type II pneumocytes lining the alveolar septa, highlighted by keratin 7 and TTF-1. The elastin staining in these lesions varied between point and linear and disorderly thickened and recoiled patterns, with a majority resembling a point and linear-like pattern. Specifically, two lesions resembled normal alveolar tissue, displaying a point and linear-like pattern without a visually appreciable increase in staining intensity (Figure 2A, B), while one lesion showed a focally discontinuous pattern with disorderly thickened and recoiled elastic fibers. All lesions with AIS (100%, 5/5, Figure 2C-E), as well as the in situ components of both MIA (100%, 10/10) and IA (so-called “lepidic-predominant adenocarcinoma,” 100%, 4/4), demonstrated a continuously disorderly thickened and recoiled pattern, delineating the alveolar contour through elastic staining. Interestingly, the features observed at the periphery of a few AIS lesions were similar to those in most AAH, characterized by a point and linear pattern (Figure 2F). There were two lesions exhibiting the collapse with interstitial fibrosis pattern in the AIS group (40%, 2/5) and two in the MIA group (20%, 2/10). For IA (73%, 27/37) and the invasive components of MIA (100%, 10/10), the normal lung structure was effaced, resulting in a decreased or disappeared pattern. The acinar type (Figure 2G, H) displayed varying extents of this pattern, while the papillary (Figure 2I∼K) and solid types (Figure 2M, N) exhibited more severe architectural destruction, with few areas retaining a normal point-and-linear elastic staining pattern. Notably, there was a complete lack of elastin in the micropapillary cores (Figure 2O, P). Among the IAs, 27.0% (10/37) involved bronchioles and adjacent vasculature, resulting in extensive carcinomatous collapse, where voluminous elastin fibers were compressed and aggregated, producing numerous aggregates of discontinuously disorderly thickened and recoiled elastic fibers (Figure 2I, L). Five lesions with NNLs (100%, 5/5) typically exhibited a point and linear pattern (Figure 2Q, R), with intensified point and linear pattern observed in their iatrogenic/mechanical collapse regions (100%, 5/5; Figure 2S, T). All normal lung tissue components in this cohort demonstrated point-and-linear staining features.
Schematic illustration of four elastic staining patterns: point and linear (PL) type, the normal staining pattern (A); intensified PL type (IPL), usually appreciated in regions with iatrogenic or mechanical collapse (B); disorderly thickened and recoiled (DTR) pattern, including continuous (CDTR, [C]) and discontinuous (DDTR, [D]) variants; the former is characteristic of adenocarcinoma in situ (AIS), and the latter usually results from cancer-related collapse; decreased or disappeared pattern (D/D), usually due to effacement of the normal alveolar wall–associated elastic fiber framework by invasive adenocarcinomas, regardless of growth pattern (E, acinar or solid; F, papillary). The lines represent elastin, and the small bumps represent pneumocytes. IA, invasive adenocarcinoma; B, bronchiole; V, vasculature.
Elastic staining features in different pulmonary lesions. Atypical adenomatous hyperplasia (A, H&E, 20×) shows a point-and-linear elastic staining pattern (B, ES, 10×). Adenocarcinoma in situ (C, H&E, 10×) demonstrates a continuous disorderly thickened and recoiled pattern (D, 10×; E, ES, 40×), with minimal focal point-and-linear-like staining at the extreme periphery in some lesions (F, ES, 10×; highlighted by the blue arrowheads). In invasive adenocarcinoma (G, H&E, 10×), pulmonary architecture is effaced, resulting in decreased or disappeared elastic staining (H, ES, 10×). Papillary growth shows fibrovascular cores with marked reduction of alveolar wall–associated elastic fibers (I, H&E, 4×; J, H&E, 10×; K, ES, 20×). Prominent carcinomatous collapse (I, the upper right regions highlighted by a blue arrowhead) exhibits a discontinuous disorderly thickened and recoiled pattern (L, ES, 40×; blue arrowhead). Solid (M, H&E, 10×; N, ES, 10×) and micropapillary growth (O, H&E, 20×; P, ES, 20×) patterns typically show prominent decreased or disappeared pattern elastic staining. Normal alveoli (Q, H&E, 10×) display a point and linear pattern (R, ES, 10×; highlighted by the red arrows [points] and blue arrowheads [lines]), which becomes more intensified in areas of mechanical or iatrogenic collapse (S, H&E 10×; T, ES, 10×). H&E, hematoxylin and eosin; ES, elastic staining.
Diagnoses and Pathologic Patterns in Principal Cohort and Validation Sets With the Help of Elastic Staining.
AAH, atypical adenomatous hyperplasia; AIS, adenocarcinoma in situ; IA, invasive adenocarcinoma; CLPIF, collapse of the lepidic pattern with interstitial fibrosis; CRC, cancer-related collapse (CRC); IC/MC, iatrogenic/mechanical collapse; MIA, minimally invasive adenocarcinoma; MP, micropapillary; NNL, non-neoplastic lesion; PA, papillary; SO, solid.
Accuracy values reflect complete concordance within the limited sample size of the validation cohorts.
Internal and External Test Validation
The results are summarized in Table 1. With the use of elastic staining and immunohistochemistry as indicated, all diagnoses were accurately established in both internal (n = 30) and external (n = 30) validation cohorts, with no significant difference in diagnostic accuracy (100%, P = 1.000). All lepidic, solid, micropapillary, carcinomatous collapse, and collapse with interstitial fibrosis patterns were correctly identified, achieving an accuracy of 100% in the test validations (Figures 3A–E). However, the papillary pattern demonstrated comparable performance between the internal and external cohorts, with an accuracy of 98% in both. The acinar pattern presented greater challenges, with accuracies of 81% and 98% in the internal and external validation cohorts, respectively, indicating some degree of mutual misinterpretation between the groups. For the identification of interstitial collapse/mechanical collapse in NNLs and adjacent neoplastic lesions, the accuracy was 98% in both the internal and external validation cohorts. Although a mild discordance in identification was observed, this was deemed acceptable given the variability in recognition thresholds among pathologists. Nonetheless, no significant identification differences were detected between the two test groups (P = 1.000).
Representative examples illustrating how elastic staining improves diagnostic accuracy and histologic pattern recognition. (A–E) A non-mucinous adenocarcinoma (A, H&E, 10×) showed expression of keratin 7 (B, IHC, 10×), TTF-1 (C, IHC, 10×), and Napsin A (D, IHC, 10×), despite abundant intracytoplasmic mucin; near-complete loss of elastin supported an invasive component (E, ES, 10×). (F–H) Collapse of the lepidic pattern with interstitial fibrosis may mimic invasion on H&E (F, 10×) and was highlighted by keratin 7 immunohistochemistry (G, IHC, 10×); elastic staining clarifies the underlying pattern (H, ES, 10×). (I–J) Collapsed alveolar walls may resemble papillary structures on H&E (I, 10×); preservation of a point-and-linear elastic staining pattern helps avoid overdiagnosis (J, ES, 10×). (K–L) In a minimally invasive adenocarcinoma, focal loss of elastin indicates the presence of an invasive component (K, H&E, 10×; L, ES, 10×).
Interobserver Reproducibility
The diagnostic results and pathologic patterns identified by the trainee and the general pathologist, with or without the assistance of elastic staining, are summarized in Table 2. When reviewing only hematoxylin and eosin and immunohistochemistry, both pathologists correctly diagnosed most patients, with no significant difference in overall accuracy (P = 0.956), although accuracy remained relatively lower. The trainee and senior pathologist misinterpreted the collapse with interstitial fibrosis pattern (3/3 for the trainee and 2/3 for the general pathologist) as an acinar pattern, leading to reclassifications of some patients from AIS to MIA (trainee, n = 2) or IA (general pathologist, n = 1; Figure 3G, H). Within the IA group, some papillary structures were incorrectly identified as lepidic (trainee, 6/11) or acinar (general pathologist, 1/11) patterns. A lesion with collapsed alveolar walls that resembled papillary structures was misdiagnosed as IA by the trainee, but the characteristic point and linear pattern revealed by elastic staining corrected the diagnosis (Figure 3I, J). The overall diagnostic accuracy for the trainee and general pathologist was 75% and 95%, respectively, while the overall accuracy for histologic identification was 68.1% for the trainee and 93.1% for the general pathologist. With the aid of elastic staining, the senior pathologist achieved a diagnostic accuracy of 100%, and the trainee's accuracy significantly improved, showing no significant difference between the two (P = 0.995). The trainee effectively removed six misclassifications of lepidic patterns (100%, 6/11) that were actually papillary types; however, two lesions were still incorrectly identified as acinar patterns. A specific challenge arose in identifying invasive components approaching the 5 mm threshold for the diagnosis of IA, particularly concerning lepidic-like areas without a continuous elastin staining pattern around AIS. This discrepancy accounted for two misclassifications. In our assessment, a prominent loss of elastic staining is more indicative of an invasive component rather than peripheral features of AIS, as any staining loss in AIS typically appears minimal (Figure 3K, L). Ultimately, the overall diagnostic accuracies reached 95% for the trainee and 100% for the general pathologist, with histologic identification accuracies of 84.7% and 98.6%, respectively. As depicted in Table 3, the agreement between the two pathologists regarding diagnoses and histologic patterns showed substantial agreement (Kappa = 0.765, P = 0.000) and almost perfect agreement (Kappa = 0.849, P = 0.000), respectively. Agreement was slightly lower when compared to the confirmed diagnoses by thoracic pathologists (trainee, Kappa = 0.650, P = 0.000; general pathologist, Kappa = 0.800, P = 0.000). Under the guidance of elastic staining, all diagnostic and pattern identification agreements reached almost perfect levels between the two pathologists (Table 3), particularly between the general and thoracic pathologists (Kappa = 1.000, P = 0.000). The overall agreement for diagnoses with and without elastic staining assistance was 77.5% (Kappa = 0.76, P = 0.000) and 95% (Kappa = 0.945, P = 0.000), respectively. Furthermore, the overall interobserver agreement for histologic patterns was 79.2% (Kappa = 0.857, P = 0.000) and 95.8% (Kappa = 0.977, P = 0.000), respectively.
Diagnoses and Pathologic Patterns in Test Cohort by Trainee and General Pathologist With or Without the Help of Elastic Staining.
AAH, atypical adenomatous hyperplasia; AIS, adenocarcinoma in situ; IA, invasive adenocarcinoma; CLPIF, collapse of the lepidic pattern with interstitial fibrosis; CRC, cancer-related collapse (CRC); IC/MC, iatrogenic/mechanic collapse; MIA, minimally invasive adenocarcinoma; MP, Micropapillary; NNL, non-neoplastic lesion; PA, papillary; SO, solid; WOES, without elastic staining; WES, with elastic staining.
Interobserver Agreement Among Thoracic Pathologists, General Pathologist and Trainee in Diagnosis and Pattern Identification.
Overall agreement of the diagnoses without and with the help of elastic staining were 77.5% (Kappa** = 0.76, P = 0.000), 95% (Kappa** = 0.945, P = 0.000), respectively, and overall interobserver agreement of histologic patterns were 79.2% (Kappa** = 0.857, P = 0.000), 95.8% (Kappa** = 0.977, P = 0.000), respectively.
* Cohen's Kappa; ** Fleiss’ Kappa; AG, agreement; WOES, without elastic staining; WES, with elastic staining.
Discussion
Although previous literature has addressed elastic fiber degeneration and described staining patterns in lung carcinoma, a comprehensive analysis of elastic staining characteristics, particularly for diagnosing pulmonary adenocarcinoma, is lacking.10,19 In this study, we summarize four distinct elastic staining patterns: a point-and-linear pattern, an IPL, a disorderly thickened and recoiled pattern (with continuous and discontinuous variants), and a decreased or disappeared pattern. Recognition of these staining types is critical for improving diagnostic accuracy and histologic pattern identification. In normal alveolar walls, elastic fibers are slender and sparse, often presenting as discontinuous point and linear patterns in conventional tissue sections of 3 to 5 μm. 10 Consistent with prior reports, continuous thickening of elastic fibers delineates the alveolar walls in AIS. 10 While the underlying reasons for these observations are not fully understood, they may relate to pathological fibroblast proliferation. Notably, prominent elastin deposition does not appear to be an initial event; some lesions with organizing pneumonia (not included in this study) support this notion. Other studies have documented elastic fiber expression at different developmental stages of malignant epithelial tumors and cancer-related desmoplastic reactions.20–23 In lesions characterized by prolonged fibrosis, such as pulmonary fibrosis and encapsulated tumors, there is typically an increase in elastic fibers.21,24 Thus, elastic staining aids in differentiating AIS from reactive mimickers, particularly those featuring proliferative type II pneumocytes in acute pneumonia settings. 25 In contrast to IPL, the peripheral regions of AIS exhibit a point-and-linear pattern staining pattern, indicating developing areas without marked elastin deposition. However, it remains unclear whether these regions represent early de novo development or secondary outgrowth from adjacent AIS.15,26 This ambiguity further complicates diagnostic interpretation, particularly when point and linear-type lepidic patterns intermingle with invasive components approaching the 5 mm threshold for IA diagnosis. Such was observed in two lesions within the agreement test cohort between the general/thoracic pathologist and trainee. Further investigation in larger, multi-institutional cohorts is warranted to clarify these diagnostic ambiguities. Although elastic staining serves as a robust auxiliary method for identifying AIS, MIA, and IA, focal, discontinuous, disorderly thickened and recoiled patterns may appear in some AIS cross-sections or as artifacts related to thin sectioning or suboptimal staining, thereby complicating the distinction between subtle MIA and IA. Consequently, a combination of histological and immunohistochemical features remains essential.
The presence of a papillary structure is a critical pathological parameter influencing lung adenocarcinoma grading; however, its interpretation poses challenges in pathological practice. 27 Loss of elastin is helpful in distinguishing papillary from lepidic configurations, the latter typically displaying continuously disorderly thickened/recoiled elastic staining. This distinction may suggest that papillary structures arise from the destruction of normal pulmonary alveolar wall structures or de novo development, a notion supported by other studies. 10 Young or non-specialist thoracic pathologists often struggle to differentiate papillary from acinar growth patterns, although this confusion does not alter the diagnosis of IA. Unfortunately, elastic staining does not resolve this issue due to their similar staining patterns. In practice, evaluation remains primarily reliant on hematoxylin and eosin morphology, with useful features identified from our experience, such as increased density of “alveolar wall"-like structures, incomplete structures with traffic-like or labyrinth-like growth patterns, and obvious branching structures. Nevertheless, reproducibility issues persist, particularly among inexperienced practitioners, as evidenced by challenges faced by both the trainee and general pathologist in our study.
Two types of lung collapse are recognized: carcinomatous collapse, also known as adenocarcinoma with a central “scar,” and interstitial collapse/micro-collapse.18,28,29 Japanese researchers suggest that the central “scar” may represent fibrosis and compressed elastic fibers, while Western studies attribute it mainly to the collapse of elastic fibers without fibrosis, suggesting that carcinomatous collapse reflects a “physiological” collapse characterized by the loss of alveolar structure. 10 In the present study, carcinomatous collapse was characterized by a distinctive discontinuously disorderly thickened/recoiled elastic staining pattern, which is morphologically compatible with collapse and condensation of elastic fibers. This pattern was also observed in proximity to bronchioles and adjacent vasculature, where tumor involvement was associated with prominent aggregation of elastic fibers. Compared with invasive carcinoma involving peripheral lung parenchyma, these regions showed relatively increased elastic fiber density, reflecting a different pattern of elastic fiber remodeling. Importantly, these interpretations are based on morphologic and elastic staining features, and underlying biological mechanisms were not directly examined in this study. In the absence of direct evidence for active elastic fiber proliferation within interstitial collapse/micro-collapse, the observed increase in elastic fiber density is most plausibly attributable to spatial compression or architectural rearrangement of pre-existing alveolar walls, although this remains speculative. In the present study, these biopsy-related changes were readily distinguishable from invasion-related elastic fiber patterns based on their limited extent and preserved point-and-linear elastic staining. Importantly, classic iatrogenic or mechanical collapse related to specimen handling also showed characteristic elastic staining features that were distinct from tumor-associated alterations. Therefore, prior biopsy procedures did not compromise the interpretation of elastic staining when appropriate morphologic correlation was applied. Variability in the identification of interstitial collapse/micro-collapse among the trainee, general pathologist, and thoracic pathologists may reflect differences in lesion extent, as we did not establish quantitative criteria in our study. Recognizing these morphologic patterns and their potential interpretations can help avoid misinterpretation of elastic staining results in routine practice. Differentiating interstitial collapse/mechanical collapse from IA remains another diagnostic challenge, often leading to erroneous diagnoses by both inexperienced and senior pathologists. Our study highlights how distinct elastin staining patterns facilitate this differentiation, underscoring its practical importance.
Despite these findings, several limitations should be acknowledged. Although the interobserver concordance analysis involved participants from different institutions, the number of observers was limited, and all were part of the same research project with standardized training. This design may have contributed to relatively high agreement rates. Accordingly, the reported concordance should be interpreted as evidence from a small-scale interobserver study, and further validation in larger, multi-institutional cohorts with more heterogeneous observer backgrounds will be important to assess generalizability in real-world practice. Given the limited sample size of the interobserver concordance cohort, confidence intervals for agreement statistics were not reported, and the observed Kappa values should be interpreted as preliminary estimates rather than precise measures of reproducibility. Taken together, the reported accuracy and agreement metrics should not be regarded as definitive performance benchmarks, but rather as evidence of potential diagnostic utility under controlled study conditions. In addition, this study focused exclusively on non-mucinous adenocarcinomas; therefore, the applicability of elastic staining patterns to mucinous adenocarcinoma subtypes remains uncertain and warrants further investigation.
In conclusion, we comprehensively analyzed the patterns of elastic staining in one of the largest reported series of pulmonary adenocarcinoma and its in situ lesions to date. This study underscores the robust auxiliary value of elastic staining in diagnostic practice and highlights its potential role in improving histological reproducibility. Familiarity with these characteristic staining patterns not only enhances our understanding of the pathological effects of lung adenocarcinoma on pulmonary architecture but also facilitates accurate recognition of diverse histological structures, ultimately supporting correct diagnosis.
Supplemental Material
sj-docx-1-ijs-10.1177_10668969261439152 - Supplemental material for Elastic Staining Features in Pulmonary Non-Mucinous Adenocarcinoma and Associated In Situ Lesions: Diagnostic Implications
Supplemental material, sj-docx-1-ijs-10.1177_10668969261439152 for Elastic Staining Features in Pulmonary Non-Mucinous Adenocarcinoma and Associated In Situ Lesions: Diagnostic Implications by Jun Zhou, Anran Wang, Xue Wang, Wenjing Zhang, Qin Shen and Xiao-Jun Zhou in International Journal of Surgical Pathology
Footnotes
Ethics Approval
This study was approved by the Institutional Review Board of The Second People's Hospital of Yibin, Department of Pathology (YBEY-P26002).
Consent to Participate
The requirement for informed consent was waived due to the retrospective and minimal-risk nature of the study.
Consent for Publication
All authors consent to publication.
Data are available without commercial interests through Jun Zhou (e-mail: zhoujun2006305@126.com).
Author Contributions
Conception and design of the study, and approval of final version of manuscript were done by J.Z., X.Z.; acquisition and analysis of data were done by X.W., W.Z.; data analysis and/or interpretation were done by J.Z., A. W., X.Z., Q.S.; drafting the manuscript or figures was done by J.Z.
Funding
The authors received no financial support for the research, authorship, and/or publication of this article.
Declaration of Conflicting Interests
The authors declared no potential conflicts of interest with respect to the research, authorship, and/or publication of this article.
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References
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