Luminal-Surface Claudin18.2 Expression Distinguishes Distal-Type Bronchiolar Adenoma from Mimicking Lesions: An Immunohistochemical Study

Luminal-Surface Claudin18.2 Expression Distinguishes Distal-Type Bronchiolar Adenoma from Mimicking Lesions: An Immunohistochemical Study | Research Square window.SnipcartSettings = { analytics: { enabled: false } }; (function() { var accessVector = localStorage.getItem('access_vector') || ''; window.dataLayer = window.dataLayer || []; if (accessVector) { window.dataLayer.push({ user: { profile: { profileInfo: { snid: accessVector } } } }); } })(); (function(w,d,s,l,i){w[l]=w[l]||[];w[l].push({'gtm.start':new Date().getTime(),event:'gtm.js'});var f=d.getElementsByTagName(s)[0],j=d.createElement(s),dl=l!='dataLayer'?'&l='+l:'';j.async=true;j.src='https://www.googletagmanager.com/gtm.js?id='+i+dl;f.parentNode.insertBefore(j,f);})(window,document,'script','dataLayer','GTM-K279D39R'); Browse Preprints In Review Journals COVID-19 Preprints AJE Video Bytes Research Tools Research Promotion AJE Professional Editing AJE Rubriq About Preprint Platform In Review Editorial Policies Our Team Advisory Board Help Center Sign In Submit a Preprint Cite Share Download PDF Research Article Luminal-Surface Claudin18.2 Expression Distinguishes Distal-Type Bronchiolar Adenoma from Mimicking Lesions: An Immunohistochemical Study Lin Xiangling, Xie Chuyu, Zhang Tingzhen, Liao Dingzhun, Li Xiangyun, and 1 more This is a preprint; it has not been peer reviewed by a journal. https://doi.org/ 10.21203/rs.3.rs-9167542/v1 This work is licensed under a CC BY 4.0 License Status: Posted Version 1 posted You are reading this latest preprint version Abstract Background Distal-type bronchiolar adenoma (BA) is a benign lung neoplasm characterized by a bilayered structure. However, atypical variants lacking continuous basal cell layers pose diagnostic challenges, particularly in distinguishing them from adenocarcinoma in situ (AIS), minimally invasive adenocarcinoma (MIA), and invasive lung adenocarcinoma (ILA) with acinar and papillary growth patterns. Aim To investigate the immunohistochemical (IHC) expression of claudin18.2 in distal-type BA and assess its diagnostic utility in differentiating BA from histologic mimics. Methods Distal-type BA, atypical distal-type BA, AIS, MIA, and ILAs with acinar and papillary growth patterns were collected and subjected to IHC staining for claudin18.2. Results Claudin18.2 was expressed in all distal-type BAs (100%, n = 6), including 3 atypical variants, with characteristic weak-to-moderate luminal-surface positivity. In contrast, AIS and MIA cases showed lateral and basal membrane staining (100%, n = 6), whereas the ILA cases were negative (0%, n = 3). Conclusion Claudin18.2 may serve as a valuable IHC marker for diagnosing distal-type BAs and distinguishing atypical variants from their histologic mimics. Distal-type Bronchiolar Adenoma Atypical distal-type Bronchiolar Adenoma Adenocarcinoma Claudin18.2 Immunohistochemical staining Figures Figure 1 Figure 2 Figure 3 Introduction Bronchiolar adenoma (BA) is a benign neoplasm of bronchiolar epithelial origin that was first described by Ishikawa et al. in 2002 as ciliated muconodular papillary tumor (CMPT). This rare peripheral lung nodule exhibits predominantly papillary architecture, comprising ciliated columnar cells, mucinous cells, and basal cells ( 1 ). In 2018, Chang et al. redefined BA and proposed two morphologic subtypes—proximal and distal—based on their resemblance to the corresponding segments of the bronchiolar-alveolar tree ( 2 ). The 5th edition of the WHO Classification of Thoracic Tumors (2021) recognized BA as a distinct entity, establishing the bilayered structure-comprising luminal cells and a continuous basal cell layer-as its essential diagnostic criterion( 3 ). Proximal-type BA is characterized by ciliated columnar cells, mucinous cells, and a continuous basal cell layer, whereas distal-type BAs typically show inconspicuous bilayered structures with loss of these cell types, rendering them difficult to distinguish from adenocarcinomas( 4 , 5 ). In such ambiguous cases, P40 immunohistochemical (IHC) staining serves as a critical adjunct to confirm a continuous basal cell layer( 6 ). In recent years, an increasing number of atypical distal-type BAs have been described ( 7 ). These lesions are characterized by either partial loss of the continuous basal cell layer (focally preserved) or its complete absence, as demonstrated by P40 IHC staining. This morphologic heterogeneity has sparked ongoing debate regarding the defining diagnostic criteria for BA. More critically, the recognition of atypical distal-type BA obscures the boundary between conventional BA and adenocarcinoma, thereby complicating clinicopathological differential diagnosis( 8 ). Therefore, reliable ancillary markers are urgently needed to differentiate BA from its adenocarcinoma mimics in clinical practice. In clinical pathology practice, distinguishing BA from adenocarcinoma in situ (AIS), minimally invasive adenocarcinoma (MIA), and invasive lung adenocarcinoma (ILA) with acinar and papillary growth patterns remains a significant diagnostic challenge ( 9 , 10 ). Morphologically, BA epithelial cells are compactly arranged with a smooth, even luminal surface. In contrast, AIS and MIA exhibit discontinuous luminal lining with characteristic hobnail cells—apically displaced nuclei that protrude into the alveolar space ( 11 , 12 ). This architectural contrast reflects fundamental differences in tight junction organization between these entities. Claudin18.2, a member of the tight junction protein family, plays a crucial role in tight junction formation( 13 ). Although claudin18.2 is physiologically expressed in gastric mucosa, it becomes pathologically exposed in gastric adenocarcinoma due to tight junction disruption ( 14 ). While early studies demonstrated absent claudin18.2 expression in normal lung tissue, recent investigations have revealed its weak expression in lepidic-predominant lung adenocarcinoma, with distinctive localization to the lateral and basal membranes ( 15 , 16 ). Notably, its expression in BA remains entirely unexplored. In this study, we investigated claudin18.2 IHC expression and subcellular localization in distal-type BA and atypical distal-type BA, comparing these findings with those in AIS, MIA, and ILAs with acinar and papillary growth patterns. Our results demonstrate that characteristic luminal-surface claudin-18.2 expression is a valuable diagnostic marker for distal-type BAs, particularly in distinguishing atypical variants from their histologic mimics. Materials and methods This retrospective study included 15 cases diagnosed at our hospital from September 2018 to December 2024: 3 distal-type BAs, 3 atypical distal-type BAs, 3 AISs, 3 MIAs, and 3 ILAs with acinar/papillary growth patterns. AIS, MIA, and ILA with acinar and papillary growth patterns were diagnosed according to the 2021 WHO Classification of Thoracic Tumours criteria. The study protocol was approved by the Institutional Review Board of the Seventh Affiliated Hospital of Sun Yat-sen University. H&E and IHC staining were performed on formalin-fixed paraffin-embedded (FFPE) tissue sections. IHC staining for claudin18.2 (clone number 43-14A; Zhongshan Golden Bridge Biotechnology, China) and P40 (clone number 513M2A7; Baidao, China) was performed on these samples. The Envision two-step method was used for IHC staining, with antigen retrieval conducted at high temperature. Whole-slide scanning was performed using the KF-PRO-005 scanner (KFBIO, Ningbo, China) at 40× magnification. Representative images were digitally extracted for publication. Results First, we examined claudin18.2 expression in terminal bronchioles lined by compactly arranged epithelial cells and in pulmonary alveolar epithelium characterized by loosely arranged cells with prominent intercellular gaps. The results revealed absent claudin18.2 expression in bronchiolar epithelium. In contrast, alveolar epithelium demonstrated weak expression localized to the lateral and basal membranes (Supplementary Fig. 1A–1B). We further examined claudin18.2 expression in distal-type BA, atypical distal-type BA, AIS, MIA, and ILAs with acinar and papillary growth patterns. Table 1 summarizes the clinicopathological features and claudin18.2 expression patterns across these lesions. The pathological features assessed included architectural growth patterns, bilayered structure integrity, cellular arrangement and luminal surface characteristics, cytologic atypia, and P40 immunoreactivity. Based on these features, lesions were classified into three categories: (I) distal-type BA, (II) atypical distal-type BA, and (III) adenocarcinomas (AIS, MIA, and ILAs with acinar and papillary growth patterns). Table 1 Clinicopathological features of BAs and its mimickers. Case Size (mm) Borders Patterns Bilayered structure Cells arrangement Cellular Atypia P40 expression Diagnosis Claudin18.2 expression 1 2.5 Well-defined Glandular/ Papillary Yes Compactly Mild Continuous BA Luminal surface 2 3.0 Well-defined Glandular/ Papillary No Compactly Mild Interspersed BA 3 4.8 Poorly defined Papillary/ Glandular No Compactly Mild Interspersed BA 4 4.3 Poorly defined Glandular Yes (focally) Compactly Mild Continuous (focally) Atypical BA 5 5.0 Well-defined Glandular No Compactly Mild Scattered Atypical BA 6 5.0 Well-defined Papillary/ Glandular No Compactly Mild Few Atypical BA 7 7.5 Poorly defined Lepidic No Loosely Hobnail cells Scattered AIS Lateral and basal membranes 8 8 Poorly defined Glandular/Lepidic No Loosely Hobnail cells No MIA 9 18 Poorly defined Papillary/ Glandular No Loosely Marked Scattered ILA* Negative BA: Bronchiolar adenoma; AIS: Lung adenocarcinoma in situ; MIA: minimally invasive adenocarcinoma; ILA: invasive lung adenocarcinoma; * invasive lung adenocarcinoma (ILA) with acinar and papillary growth patterns. Supplementary Fig. 1. Claudin18.2 expression in bronchiolar and pulmonary alveolar epithelium. (A) Terminal bronchiole lined by compactly arranged epithelial cells with smooth luminal surface, transitioning to alveolar epithelium characterized by loosely arranged cells with prominent intercellular gaps (H&E). (B) Claudin18.2 is negative in bronchiolar epithelium but weakly positive along lateral and basal membranes of alveolar epithelial cells. Cases 1 3 (Fig. 1 ) represent distal-type BAs (lesion diameters: 2.5–4.8 mm). Case 1 (Fig. 1 A–D) showed a well-circumscribed lesion with glandular/papillary architecture (Fig. 1 A). High-power examination revealed ciliated cells in compact arrangement with a distinct bilayered structure (Fig. 1 B). P40 IHC confirmed a continuous basal cell layer (Fig. 1 C), supporting the diagnosis. Case 2 (Fig. 1 E–H) also demonstrated well-defined borders. Tumor cells were arranged in dilated glandular/papillary structures separated by eosinophilic fibromyxoid stroma (Fig. 1 E), with compact architecture and a smooth luminal surface. Most cells exhibited abundant apical "snouting" secretion and basally located nuclei (Fig. 1 F). Although a distinct bilayered structure was not evident, P40 staining revealed interspersed basal cells (Fig. 1 G). Case 3 (Fig. 1 I–L) showed poorly defined borders, with tumor cells in papillary or glandular patterns (Fig. 1 I). Cells were compactly arranged with abundant apical secretion (Fig. 1 J). P40 staining revealed interspersed basal cells (Fig. 1 K). Thus, despite absent bilayered structures, Cases 2 and 3 were diagnosed as distal-type BA based on scattered P40-positive basal cells and characteristic cytomorphology. All three cases demonstrated weak-to-moderate claudin18.2 positivity localized to the luminal surface (Fig. 1 D, H, L). Figure 2 (Cases 4–6) illustrates atypical distal-type BAs (lesion diameters: 2.5–5.0 mm). Case 4 (Fig. 2 A–D) showed predominantly irregularly dilated glandular structures with variable calibers (Fig. 2 A). Ciliated cells and a bilayered structure were restricted to focal areas. Cells showed compact arrangement with apical "snouting" secretion and basally located nuclei (Fig. 2 B). P40 staining confirmed a focally continuous basal cell layer (Fig. 2 C), supporting the diagnosis of atypical BA (partially monolayered BA). Case 5 (Fig. 2 E–H) demonstrated well-defined borders, with tumor cells arranged in dilated glandular structures separated by eosinophilic fibromyxoid stroma (Fig. 2 E). No bilayered structure was identified; however, compactly arranged cells with abundant apical secretion and a smooth luminal surface were evident (Fig. 2 F). P40 staining revealed scattered positive cells (Fig. 2 G). Case 6 (Fig. 2 I–L) showed well-circumscribed borders, with tumor cells arranged in papillary or irregular glandular structures (Fig. 2 I). Compact arrangement with abundant apical secretion was evident (Fig. 2 J). P40 staining revealed few positive cells (Fig. 2 K). Based on morphology and P40 expression, Cases 5 and 6 were diagnosed as atypical BA (monolayered BA-like lesions). All three atypical BAs demonstrated claudin18.2 expression patterns identical to distal-type BAs—weak-to-moderate luminal surface positivity (Fig. 2 D, H, L). The third group comprises histologic mimics of BA, including AIS, MIA, and ILAs with acinar and papillary growth patterns. Case 7 (AIS) showed poorly defined borders, with tumor cells arranged in a single layer demonstrating predominantly lepidic growth (Fig. 3 A). Loosely arranged cells with "hobnail" appearance were evident (Fig. 3 B). P40 staining revealed scattered positive cells (Fig. 3 C). Claudin18.2 showed lateral and basal membrane positivity (Fig. 3 D). Case 8 (MIA) showed dilated glandular structures centrally and lepidic growth peripherally (Fig. 3 E). Most cells were loosely arranged, with prominent intercellular gaps, "hobnail" appearance, and uneven luminal surface (Fig. 3 F). P40 staining was negative (Fig. 3 G). Claudin18.2 positivity was localized to the lateral and basal membranes (Fig. 3 H). Case 9 (ILAs, acinar/papillary type) showed a lesion diameter exceeding 15 mm. Tumor cells were predominantly arranged in irregular glands and papillary structures (Fig. 3 I). Most cells showed marked atypia, with a nuclear-to-cytoplasmic ratio > 0.5. Cellular stratification and intercellular gaps were observed, with uneven luminal surface (Fig. 3 J). P40 staining revealed scattered positive cells in focal areas (Fig. 3 K). Claudin18.2 staining was negative (Fig. 3 L). Discussion In recent years, atypical distal-type BAs have been reported, including partial monolayered BA and monolayered BA-like lesions ( 8 , 10 ). These variants have challenged the initial diagnostic criterion—the bilayered structure with a continuous basal cell layer—of this entity. Therefore, additional ancillary markers are required to distinguish BA from its histologic mimics in routine diagnostic practice. In this study, we describe the morphologic features of six distal-type BAs, including three atypical variants. All six distal-type BAs demonstrated compactly arranged cells lacking discernible intercellular gaps—a key morphologic feature distinguishing this entity from AIS and MIA. Furthermore, we observed that claudin18.2 is expressed in distal-type BAs with characteristic luminal surface positivity. This luminal localization contrasts with the absence of claudin18.2 in bronchiolar epithelium and its lateral–basal distribution in alveolar epithelium. Claudin18.2 is essential for maintaining tight junctions between epithelial cells ( 17 ). Previous studies have shown that carcinogenesis disrupts tight junctions in epithelial cells, resulting in claudin18.2 exposure ( 16 , 17 ). As distal-type BAs are believed to arise from terminal bronchiolar epithelium, this luminal localization may reflect distinct tight junction architecture in BAs, possibly related to their terminal bronchiolar origin and compact cellular arrangement. In AIS, MIA, and ILA with acinar/papillary growth patterns, IHC staining revealed claudin18.2 expression patterns distinct from those in BAs. Claudin18.2 was expressed in AIS and MIA with lateral and basal membrane positivity, resembling that of alveolar epithelium; however, it was absent in ILA with acinar and papillary patterns. We hypothesize that AIS and MIA share tight junction characteristics with alveolar epithelium, as these lesions exhibit loosely arranged cells and uneven luminal surfaces. Conversely, in ILA, tight junctions may be disrupted or undergo significant alterations, leading to complete loss of claudin18.2 expression. Our findings align with a recent report demonstrating absent claudin18.2 expression in non-mucinous ILA with solid, papillary, acinar, or micropapillary patterns, in contrast to weak expression in lepidic-predominant adenocarcinoma ( 15 ). Collectively, these findings establish a diagnostic algorithm based on claudin18.2 localization: luminal-surface expression defines distal-type BA (including atypical variants); lateral and basal membrane positivity indicates AIS or MIA; and absent expression supports ILA. In routine diagnostic practice, we propose a tiered IHC staining strategy: initial assessment using P40 to evaluate basal cell layer integrity; when P40 shows indeterminate results (discontinuous or rare basal cells), claudin18.2 staining is added—luminal-surface positivity supports BA diagnosis, whereas lateral–basal membrane positivity or negative staining suggests non-BA lesions (AIS/MIA/ILA). This study has several limitations. First, the sample size is limited, with claudin18.2 expression examined in only six distal-type BAs (including three atypical variants). Moreover, proximal-type BAs were not included in this analysis. Consequently, the characteristic luminal surface expression pattern requires validation in a larger cohort, particularly one encompassing the full morphologic spectrum of BA subtypes. Second, integrated molecular profiling was not performed. Whether the observed claudin18.2 expression pattern correlates with specific genetic alterations (e.g., BRAF V600E or EGFR mutations) remains to be determined in future studies combining morphological, immunophenotypic, and molecular analyses. Conclusions This study identifies characteristic luminal-surface claudin18.2 expression as a diagnostic marker for distal-type BAs, including atypical variants with incomplete or absent basal cell layers. When integrated with P40 immunostaining, claudin18.2 enables a tiered diagnostic approach to distinguish BA from AIS, MIA, and ILA with acinar and papillary growth patterns. Declarations 1. Ethics approval and consent to participate The study was conducted in accordance with the Declaration of Helsinki and was approved by the Institutional Review Board (IRB) of the Seventh Affiliated Hospital of Sun Yat-sen University. The requirement for informed consent was waived due to the retrospective nature by the IRB of the Seventh Affiliated Hospital (application ID: KY-2025-399-01). 2. Consent for publication Written informed consent for the publication of potentially identifiable information (age, sex, and pathological images, but excluding photographs with facial features, ID numbers, and hospital admission numbers) was obtained from all patients. 3. Availability of data and materials Aggregated data are available from the corresponding author upon reasonable request. 4. Competing interests The authors declare no competing interests. 5. Funding No funding was involved in this study. 6. Authors' contributions QCF, LXY and XCY: Literature search, study design and concept drafting, references and writing of the manuscript. LXL and ZTZ: Literature search, study design, writing of manuscript and interpretation of results. LDZ: IHC staining and review of the final manuscript. 7. Acknowledgment None References Kamata T, Yoshida A, Kosuge T, Watanabe S, Asamura H, Tsuta K. Ciliated muconodular papillary tumors of the lung: a clinicopathologic analysis of 10 cases. Am J Surg Pathol. 2015;39(6):753–60. Chang JC, Montecalvo J, Borsu L, Lu S, Larsen BT, Wallace WD, et al. Bronchiolar Adenoma: Expansion of the Concept of Ciliated Muconodular Papillary Tumors With Proposal for Revised Terminology Based on Morphologic, Immunophenotypic, and Genomic Analysis of 25 Cases. Am J Surg Pathol. 2018;42(8):1010–26. Nicholson AG, Tsao MS, Beasley MB, Borczuk AC, Brambilla E, Cooper WA, et al. The 2021 WHO Classification of Lung Tumors: Impact of Advances Since 2015. J Thorac oncology: official publication Int Association Study Lung Cancer. 2022;17(3):362–87. Zhu M, Yang Q, Zhan S, Liu W, Liu W, Guo L, et al. Clinicopathological analysis of bronchiolar adenoma combined with lung adenocarcinoma: Report of eight cases and literature review. Histol Histopathol. 2024;39(6):783–94. Shirsat H, Zhou F, Chang JC, Rekhtman N, Saqi A, Argyropoulos K, et al. Bronchiolar Adenoma/Pulmonary Ciliated Muconodular Papillary Tumor. Am J Clin Pathol. 2021;155(6):832–44. Liu S, Cai X, Pan J, Liu S, Lin J, Yue X. Bronchiole adenoma/pulmonary ciliated mucinous nodular papillary tumor: Case series and literature review. Medicine. 2023;102(50):e36559. Shao J, Yin JC, Bao H, Zhao R, Han Y, Zhu L, et al. Morphological, immunohistochemical, and genetic analyses of bronchiolar adenoma and its putative variants. J Pathol Clin Res. 2021;7(3):287–300. Bo J, Chen X, Zhang T, Zhu X, Zhang L, Liu Y, et al. Clinicopathological features and genomic analysis of bronchiolar adenoma. Histol Histopathol. 2023;38(12):1465–74. Ding B, Shang Z, Xiang Z, Han Y. Clinicopathologic Features and Frozen Diagnostic Pitfalls of Bronchiolar Adenoma/Ciliated Muconodular Papillary Tumors (BA/CMPTs). Am J Surg Pathol. 2023;47(4):431–9. Lin DL, Ding L, Shao SH, Xin FJ, Zhang LX, Li GQ, et al. Bronchiolar adenoma-like tumour with monolayered component: Represent malignant transformation of bronchiolar adenoma? A series of five cases. Pathol Res Pract. 2022;238:154079. Travis WD, Brambilla E, Noguchi M, Nicholson AG, Geisinger KR, Yatabe Y, et al. International association for the study of lung cancer/american thoracic society/european respiratory society international multidisciplinary classification of lung adenocarcinoma. J Thorac oncology: official publication Int Association Study Lung Cancer. 2011;6(2):244–85. Takanashi Y, Tajima S, Tsukui M, Shinmura K, Hayakawa T, Takahashi T, et al. Non-Mucinous Lepidic Predominant Adenocarcinoma Presenting with Extensive Aerogenous Spread. Rare tumors. 2016;8(4):6580. Tsukita S, Furuse M. Pores in the wall: claudins constitute tight junction strands containing aqueous pores. J Cell Biol. 2000;149(1):13–6. Zhang J, Dong R, Shen L. Evaluation and reflection on claudin 18.2 targeting therapy in advanced gastric cancer. Chin J cancer Res = Chung-kuo yen cheng yen chiu. 2020;32(2):263–70. Yan P, Dong Y, Zhang F, Zhen T, Liang J, Shi H, et al. Claudin18.2 expression and its clinicopathological feature in adenocarcinoma from various parts. J Clin Pathol. 2025;78(12):815–21. Sahin U, Koslowski M, Dhaene K, Usener D, Brandenburg G, Seitz G, et al. Claudin-18 splice variant 2 is a pan-cancer target suitable for therapeutic antibody development. Clin cancer research: official J Am Association Cancer Res. 2008;14(23):7624–34. Pak S, Simon AG, Lyu SI, Tolkach Y, Alakus H, Zander T, et al. The expression of the tight junction protein and therapeutical target Claudin 18.2 is heterogeneously distributed within esophageal and gastric adenocarcinoma. Sci Rep. 2025;15(1):28958. Additional Declarations No competing interests reported. Supplementary Files floatimage4.jpeg Supplementary Figure 1. Claudin18.2 expression in bronchiolar and pulmonary alveolar epithelium. (A) Terminal bronchiole lined by compactly arranged epithelial cells with smooth luminal surface, transitioning to alveolar epithelium characterized by loosely arranged cells with prominent intercellular gaps (H&E). (B) Claudin18.2 is negative in bronchiolar epithelium but weakly positive along lateral and basal membranes of alveolar epithelial cells. Cite Share Download PDF Status: Posted Version 1 posted You are reading this latest preprint version Research Square lets you share your work early, gain feedback from the community, and start making changes to your manuscript prior to peer review in a journal. As a division of Research Square Company, we’re committed to making research communication faster, fairer, and more useful. We do this by developing innovative software and high quality services for the global research community. Our growing team is made up of researchers and industry professionals working together to solve the most critical problems facing scientific publishing. Also discoverable on Platform About Our Team In Review Editorial Policies Advisory Board Help Center Resources Author Services Accessibility API Access RSS feed Manage Cookie Preferences © Research Square 2026 | ISSN 2693-5015 (online) Privacy Policy Terms of Service Do Not Sell My Personal Information {"props":{"pageProps":{"initialData":{"identity":"rs-9167542","acceptedTermsAndConditions":true,"allowDirectSubmit":true,"archivedVersions":[],"articleType":"Research Article","associatedPublications":[],"authors":[{"id":627360023,"identity":"09120082-71d8-4d69-8c0f-7e3254966d17","order_by":0,"name":"Lin Xiangling","email":"","orcid":"","institution":"Seventh Affiliated Hospital of Sun Yat-sen University","correspondingAuthor":false,"prefix":"","firstName":"Lin","middleName":"","lastName":"Xiangling","suffix":""},{"id":627360024,"identity":"cc359b0b-d854-4c34-8c95-0784a5f31e11","order_by":1,"name":"Xie Chuyu","email":"","orcid":"","institution":"Seventh Affiliated Hospital of Sun Yat-sen University","correspondingAuthor":false,"prefix":"","firstName":"Xie","middleName":"","lastName":"Chuyu","suffix":""},{"id":627360025,"identity":"68bbfb68-1315-4c52-b5a7-4e1aa22d1680","order_by":2,"name":"Zhang Tingzhen","email":"","orcid":"","institution":"Seventh Affiliated Hospital of Sun Yat-sen University","correspondingAuthor":false,"prefix":"","firstName":"Zhang","middleName":"","lastName":"Tingzhen","suffix":""},{"id":627360026,"identity":"515c1087-986e-480a-be1e-a83b33476a1c","order_by":3,"name":"Liao Dingzhun","email":"","orcid":"","institution":"Seventh Affiliated Hospital of Sun Yat-sen University","correspondingAuthor":false,"prefix":"","firstName":"Liao","middleName":"","lastName":"Dingzhun","suffix":""},{"id":627360027,"identity":"a63276c7-de45-4959-adb5-0c0472e82335","order_by":4,"name":"Li Xiangyun","email":"","orcid":"","institution":"Seventh Affiliated Hospital of Sun Yat-sen University","correspondingAuthor":false,"prefix":"","firstName":"Li","middleName":"","lastName":"Xiangyun","suffix":""},{"id":627360028,"identity":"a2bf0e9d-c9f1-48b4-8a8b-946ecad06eb3","order_by":5,"name":"Qin Changfei","email":"data:image/png;base64,iVBORw0KGgoAAAANSUhEUgAAAZAAAAAyAQMAAABI0h/eAAAABlBMVEX///8AAABVwtN+AAAACXBIWXMAAA7EAAAOxAGVKw4bAAABAUlEQVRIiWNgGAWjYDACZhCyqZFjYGBsPAATlCCsJe2YMVBLA5FawLrSmBMbgAzitPAd5z34uSCBLX1t+2GgLX8O2xscYD54m4fBLg+XFsnDfMnSMxJkcredSWw4wNh2OHHDAbZkax6G5GJcWgwO85gx8/5gy912AKSl4XCCwQEeM2kehgNgp+LUwpPAnG52/iHMYfzfiNKSYHYDaAsD22HGDQd42PBqkTzMYyzNk3DMcNsNoC2JbemJMw+zGVvOMUjGqYXv/BnDzzwJNfJm59MfPvjwx9qe73jzwxtvKuxwakHEBQgkMDSDIxfoYFzq0bUwMNThUToKRsEoGAUjFQAA23VaGGHAoQoAAAAASUVORK5CYII=","orcid":"","institution":"Seventh Affiliated Hospital of Sun Yat-sen University","correspondingAuthor":true,"prefix":"","firstName":"Qin","middleName":"","lastName":"Changfei","suffix":""}],"badges":[],"createdAt":"2026-03-19 09:23:18","currentVersionCode":1,"declarations":"","doi":"10.21203/rs.3.rs-9167542/v1","doiUrl":"https://doi.org/10.21203/rs.3.rs-9167542/v1","draftVersion":[],"editorialEvents":[],"editorialNote":"","failedWorkflow":false,"files":[{"id":107651750,"identity":"55f4be4d-5098-4a20-9364-0205ff977807","added_by":"auto","created_at":"2026-04-23 15:10:33","extension":"jpeg","order_by":1,"title":"Figure 1","display":"","copyAsset":false,"role":"figure","size":484009,"visible":true,"origin":"","legend":"\u003cp\u003eClaudin18.2 expression in distal-type BA. (A-D) Case 1: H\u0026amp;E staining showing a well-circumscribed lesion with glandular or papillary architecture (A), bilayered structure with ciliated cells arranged in a compact pattern (B). P40 immunostaining highlights a continuous basal cell layer (C). Claudin18.2 shows moderate luminal surface positivity (D). (E-H) Case 2: Dilated glandular/papillary structures separated by eosinophilic fibromyxoid stroma (E). Compactly arranged tumor cells exhibit apical snouting secretion and basally-located nuclei (F). P40 shows interspersedbasal cells (G). Claudin18.2 shows weak positivity on the luminal surface \u0026nbsp;(H). (I-L) Case 3: Papillary/glandular growth pattern (I), Compactly arranged cells with abundant apical secretion (J). P40 demonstrates interspersed basal cells (K). Claudin18.2 shows luminal surface weak positivity (L).\u003c/p\u003e","description":"","filename":"floatimage1.jpeg","url":"https://assets-eu.researchsquare.com/files/rs-9167542/v1/2ddae307a9d715467d20b4d4.jpeg"},{"id":107651721,"identity":"f782f775-8c87-4b8c-980a-7b2d61a4e36b","added_by":"auto","created_at":"2026-04-23 15:10:28","extension":"jpeg","order_by":2,"title":"Figure 2","display":"","copyAsset":false,"role":"figure","size":1238821,"visible":true,"origin":"","legend":"\u003cp\u003eClaudin18.2 expression in atypical BAs. (A-D) Case 4 (partially monolayered BA): Irregularly dilated glands (A), focally identifiable bilayered structure (upper panel), compactly arranged cells and a smooth luminal surface (B). P40 demonstrates focally continuous basal cells (C). Claudin18.2 exhibits moderate luminal surface positivity (D). (E-H) Case 5 (monolayered BA-like): Dilated glands with eosinophilic fibromyxoid stroma (E), smooth luminal surface, compactly arranged cells with abundant apical secretion (F). P40 demonstrates scattered positive cells (G). Claudin18.2 shows moderate positivity on the luminal surface (H). (I-L) Case 6 (monolayered BA-like): Papillary/irregular glandular structures (I), compactly arranged cells with abundant apical secretion (J). P40 demonstrates few positive cells (K). Claudin18.2 shows weak luminal surface positivity (L).\u003c/p\u003e","description":"","filename":"floatimage2.jpeg","url":"https://assets-eu.researchsquare.com/files/rs-9167542/v1/1f3c839873d6113dfc116194.jpeg"},{"id":107651720,"identity":"2fd1a319-b2d8-4e5c-8b23-2c032a5ce286","added_by":"auto","created_at":"2026-04-23 15:10:27","extension":"jpeg","order_by":3,"title":"Figure 3","display":"","copyAsset":false,"role":"figure","size":464239,"visible":true,"origin":"","legend":"\u003cp\u003eClaudin18.2 expression in AIS, MIA, and lung adenocarcinoma with acinar and papillary growth patterns. (A-D) AIS: Poorly defined borders with lepidic growth (A), hobnail cells with discontinuous luminal surface (B). P40 demonstrates scattered positive cells (C). Claudin18.2 shows moderate positivity localized to the lateral and basal membranes (D). (E-H) MIA: Central dilated glands and peripheral lepidic pattern (E), hobnail appearance with uneven luminal surface (F). P40 demonstrates negative staining (G). Claudin18.2 shows lateral and basal membrane moderate positivity (H). (I-L) Invasive adenocarcinoma (acinar/papillary): Irregular glands/papillary structures, cellular stratification (upper panel), nuclear-to-cytoplasmic ratio \u0026gt;0.5, uneven luminal surface (I, J). P40 demonstrates focally scattered cells (K). Claudin18.2 is negative (L).\u003c/p\u003e","description":"","filename":"floatimage3.jpeg","url":"https://assets-eu.researchsquare.com/files/rs-9167542/v1/f020296f37a88b0b2004e65d.jpeg"},{"id":107869799,"identity":"d4e667c1-8c24-4022-a573-02dbb698f0d0","added_by":"auto","created_at":"2026-04-27 07:38:10","extension":"pdf","order_by":0,"title":"","display":"","copyAsset":false,"role":"manuscript-pdf","size":2410289,"visible":true,"origin":"","legend":"","description":"","filename":"manuscript.pdf","url":"https://assets-eu.researchsquare.com/files/rs-9167542/v1/12bedc12-9d28-40fb-88bf-6f19352c4a23.pdf"},{"id":107651822,"identity":"a9aabe26-d768-4fdf-b4a4-005782023c6e","added_by":"auto","created_at":"2026-04-23 15:10:48","extension":"jpeg","order_by":1,"title":"","display":"","copyAsset":false,"role":"supplement","size":102723,"visible":true,"origin":"","legend":"\u003cp\u003eSupplementary Figure 1. Claudin18.2 expression in bronchiolar and pulmonary alveolar epithelium. (A) Terminal bronchiole lined by compactly arranged epithelial cells with smooth luminal surface, transitioning to alveolar epithelium characterized by loosely arranged cells with prominent intercellular gaps (H\u0026amp;E). (B) Claudin18.2 is negative in bronchiolar epithelium but weakly positive along lateral and basal membranes of alveolar epithelial cells.\u003c/p\u003e","description":"","filename":"floatimage4.jpeg","url":"https://assets-eu.researchsquare.com/files/rs-9167542/v1/89d5e0ecf67b6cbc5cf54bbe.jpeg"}],"financialInterests":"No competing interests reported.","formattedTitle":"Luminal-Surface Claudin18.2 Expression Distinguishes Distal-Type Bronchiolar Adenoma from Mimicking Lesions: An Immunohistochemical Study","fulltext":[{"header":"Introduction","content":"\u003cp\u003eBronchiolar adenoma (BA) is a benign neoplasm of bronchiolar epithelial origin that was first described by Ishikawa et al. in 2002 as ciliated muconodular papillary tumor (CMPT). This rare peripheral lung nodule exhibits predominantly papillary architecture, comprising ciliated columnar cells, mucinous cells, and basal cells (\u003cspan citationid=\"CR1\" class=\"CitationRef\"\u003e1\u003c/span\u003e). In 2018, Chang et al. redefined BA and proposed two morphologic subtypes\u0026mdash;proximal and distal\u0026mdash;based on their resemblance to the corresponding segments of the bronchiolar-alveolar tree (\u003cspan citationid=\"CR2\" class=\"CitationRef\"\u003e2\u003c/span\u003e). The 5th edition of the WHO Classification of Thoracic Tumors (2021) recognized BA as a distinct entity, establishing the bilayered structure-comprising luminal cells and a continuous basal cell layer-as its essential diagnostic criterion(\u003cspan citationid=\"CR3\" class=\"CitationRef\"\u003e3\u003c/span\u003e). Proximal-type BA is characterized by ciliated columnar cells, mucinous cells, and a continuous basal cell layer, whereas distal-type BAs typically show inconspicuous bilayered structures with loss of these cell types, rendering them difficult to distinguish from adenocarcinomas(\u003cspan citationid=\"CR4\" class=\"CitationRef\"\u003e4\u003c/span\u003e, \u003cspan citationid=\"CR5\" class=\"CitationRef\"\u003e5\u003c/span\u003e). In such ambiguous cases, P40 immunohistochemical (IHC) staining serves as a critical adjunct to confirm a continuous basal cell layer(\u003cspan citationid=\"CR6\" class=\"CitationRef\"\u003e6\u003c/span\u003e).\u003c/p\u003e \u003cp\u003eIn recent years, an increasing number of atypical distal-type BAs have been described (\u003cspan citationid=\"CR7\" class=\"CitationRef\"\u003e7\u003c/span\u003e). These lesions are characterized by either partial loss of the continuous basal cell layer (focally preserved) or its complete absence, as demonstrated by P40 IHC staining. This morphologic heterogeneity has sparked ongoing debate regarding the defining diagnostic criteria for BA. More critically, the recognition of atypical distal-type BA obscures the boundary between conventional BA and adenocarcinoma, thereby complicating clinicopathological differential diagnosis(\u003cspan citationid=\"CR8\" class=\"CitationRef\"\u003e8\u003c/span\u003e). Therefore, reliable ancillary markers are urgently needed to differentiate BA from its adenocarcinoma mimics in clinical practice.\u003c/p\u003e \u003cp\u003eIn clinical pathology practice, distinguishing BA from adenocarcinoma in situ (AIS), minimally invasive adenocarcinoma (MIA), and invasive lung adenocarcinoma (ILA) with acinar and papillary growth patterns remains a significant diagnostic challenge (\u003cspan citationid=\"CR9\" class=\"CitationRef\"\u003e9\u003c/span\u003e, \u003cspan citationid=\"CR10\" class=\"CitationRef\"\u003e10\u003c/span\u003e). Morphologically, BA epithelial cells are compactly arranged with a smooth, even luminal surface. In contrast, AIS and MIA exhibit discontinuous luminal lining with characteristic hobnail cells\u0026mdash;apically displaced nuclei that protrude into the alveolar space (\u003cspan citationid=\"CR11\" class=\"CitationRef\"\u003e11\u003c/span\u003e, \u003cspan citationid=\"CR12\" class=\"CitationRef\"\u003e12\u003c/span\u003e). This architectural contrast reflects fundamental differences in tight junction organization between these entities.\u003c/p\u003e \u003cp\u003eClaudin18.2, a member of the tight junction protein family, plays a crucial role in tight junction formation(\u003cspan citationid=\"CR13\" class=\"CitationRef\"\u003e13\u003c/span\u003e). Although claudin18.2 is physiologically expressed in gastric mucosa, it becomes pathologically exposed in gastric adenocarcinoma due to tight junction disruption (\u003cspan citationid=\"CR14\" class=\"CitationRef\"\u003e14\u003c/span\u003e). While early studies demonstrated absent claudin18.2 expression in normal lung tissue, recent investigations have revealed its weak expression in lepidic-predominant lung adenocarcinoma, with distinctive localization to the lateral and basal membranes (\u003cspan citationid=\"CR15\" class=\"CitationRef\"\u003e15\u003c/span\u003e, \u003cspan citationid=\"CR16\" class=\"CitationRef\"\u003e16\u003c/span\u003e). Notably, its expression in BA remains entirely unexplored. In this study, we investigated claudin18.2 IHC expression and subcellular localization in distal-type BA and atypical distal-type BA, comparing these findings with those in AIS, MIA, and ILAs with acinar and papillary growth patterns. Our results demonstrate that characteristic luminal-surface claudin-18.2 expression is a valuable diagnostic marker for distal-type BAs, particularly in distinguishing atypical variants from their histologic mimics.\u003c/p\u003e"},{"header":"Materials and methods","content":"\u003cp\u003eThis retrospective study included 15 cases diagnosed at our hospital from September 2018 to December 2024: 3 distal-type BAs, 3 atypical distal-type BAs, 3 AISs, 3 MIAs, and 3 ILAs with acinar/papillary growth patterns. AIS, MIA, and ILA with acinar and papillary growth patterns were diagnosed according to the 2021 WHO Classification of Thoracic Tumours criteria. The study protocol was approved by the Institutional Review Board of the Seventh Affiliated Hospital of Sun Yat-sen University.\u003c/p\u003e \u003cp\u003eH\u0026amp;E and IHC staining were performed on formalin-fixed paraffin-embedded (FFPE) tissue sections. IHC staining for claudin18.2 (clone number 43-14A; Zhongshan Golden Bridge Biotechnology, China) and P40 (clone number 513M2A7; Baidao, China) was performed on these samples. The Envision two-step method was used for IHC staining, with antigen retrieval conducted at high temperature.\u003c/p\u003e \u003cp\u003eWhole-slide scanning was performed using the KF-PRO-005 scanner (KFBIO, Ningbo, China) at 40\u0026times; magnification. Representative images were digitally extracted for publication.\u003c/p\u003e"},{"header":"Results","content":"\u003cp\u003eFirst, we examined claudin18.2 expression in terminal bronchioles lined by compactly arranged epithelial cells and in pulmonary alveolar epithelium characterized by loosely arranged cells with prominent intercellular gaps. The results revealed absent claudin18.2 expression in bronchiolar epithelium. In contrast, alveolar epithelium demonstrated weak expression localized to the lateral and basal membranes (Supplementary Fig.\u0026nbsp;1A\u0026ndash;1B).\u003c/p\u003e \u003cp\u003eWe further examined claudin18.2 expression in distal-type BA, atypical distal-type BA, AIS, MIA, and ILAs with acinar and papillary growth patterns. Table\u0026nbsp;\u003cspan refid=\"Tab1\" class=\"InternalRef\"\u003e1\u003c/span\u003e summarizes the clinicopathological features and claudin18.2 expression patterns across these lesions. The pathological features assessed included architectural growth patterns, bilayered structure integrity, cellular arrangement and luminal surface characteristics, cytologic atypia, and P40 immunoreactivity. Based on these features, lesions were classified into three categories: (I) distal-type BA, (II) atypical distal-type BA, and (III) adenocarcinomas (AIS, MIA, and ILAs with acinar and papillary growth patterns).\u003c/p\u003e \u003cp\u003e \u003cdiv class=\"gridtable\"\u003e\u003ctable float=\"Yes\" id=\"Tab1\" border=\"1\"\u003e \u003ccaption language=\"En\"\u003e \u003cdiv class=\"CaptionNumber\"\u003eTable 1\u003c/div\u003e \u003cdiv class=\"CaptionContent\"\u003e \u003cp\u003eClinicopathological features of BAs and its mimickers.\u003c/p\u003e \u003c/div\u003e \u003c/caption\u003e \u003ccolgroup cols=\"10\"\u003e \u003cdiv align=\"left\" class=\"colspec\" colname=\"c1\" colnum=\"1\"\u003e\u003c/div\u003e \u003cdiv align=\"left\" class=\"colspec\" colname=\"c2\" colnum=\"2\"\u003e\u003c/div\u003e \u003cdiv align=\"left\" class=\"colspec\" colname=\"c3\" colnum=\"3\"\u003e\u003c/div\u003e \u003cdiv align=\"left\" class=\"colspec\" colname=\"c4\" colnum=\"4\"\u003e\u003c/div\u003e \u003cdiv align=\"left\" class=\"colspec\" colname=\"c5\" colnum=\"5\"\u003e\u003c/div\u003e \u003cdiv align=\"left\" class=\"colspec\" colname=\"c6\" colnum=\"6\"\u003e\u003c/div\u003e \u003cdiv align=\"left\" class=\"colspec\" colname=\"c7\" colnum=\"7\"\u003e\u003c/div\u003e \u003cdiv align=\"left\" class=\"colspec\" colname=\"c8\" colnum=\"8\"\u003e\u003c/div\u003e \u003cdiv align=\"left\" class=\"colspec\" colname=\"c9\" colnum=\"9\"\u003e\u003c/div\u003e \u003cdiv align=\"left\" class=\"colspec\" colname=\"c10\" colnum=\"10\"\u003e\u003c/div\u003e \u003cthead\u003e \u003ctr\u003e \u003cth align=\"left\" colname=\"c1\"\u003e \u003cp\u003eCase\u003c/p\u003e \u003c/th\u003e \u003cth align=\"left\" colname=\"c2\"\u003e \u003cp\u003eSize\u003c/p\u003e \u003cp\u003e(mm)\u003c/p\u003e \u003c/th\u003e \u003cth align=\"left\" colname=\"c3\"\u003e \u003cp\u003eBorders\u003c/p\u003e \u003c/th\u003e \u003cth align=\"left\" colname=\"c4\"\u003e \u003cp\u003ePatterns\u003c/p\u003e \u003c/th\u003e \u003cth align=\"left\" colname=\"c5\"\u003e \u003cp\u003eBilayered structure\u003c/p\u003e \u003c/th\u003e \u003cth align=\"left\" colname=\"c6\"\u003e \u003cp\u003eCells arrangement\u003c/p\u003e \u003c/th\u003e \u003cth align=\"left\" colname=\"c7\"\u003e \u003cp\u003eCellular Atypia\u003c/p\u003e \u003c/th\u003e \u003cth align=\"left\" colname=\"c8\"\u003e \u003cp\u003eP40 expression\u003c/p\u003e \u003c/th\u003e \u003cth align=\"left\" colname=\"c9\"\u003e \u003cp\u003eDiagnosis\u003c/p\u003e \u003c/th\u003e \u003cth align=\"left\" colname=\"c10\"\u003e \u003cp\u003eClaudin18.2 expression\u003c/p\u003e \u003c/th\u003e \u003c/tr\u003e \u003c/thead\u003e \u003ctbody\u003e \u003ctr\u003e \u003ctd align=\"left\" colname=\"c1\"\u003e \u003cp\u003e1\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c2\"\u003e \u003cp\u003e2.5\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c3\"\u003e \u003cp\u003eWell-defined\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c4\"\u003e \u003cp\u003eGlandular/\u003c/p\u003e \u003cp\u003ePapillary\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c5\"\u003e \u003cp\u003eYes\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c6\"\u003e \u003cp\u003eCompactly\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c7\"\u003e \u003cp\u003eMild\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c8\"\u003e \u003cp\u003eContinuous\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c9\"\u003e \u003cp\u003eBA\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c10\" morerows=\"5\" rowspan=\"6\"\u003e \u003cp\u003eLuminal surface\u003c/p\u003e \u003c/td\u003e \u003c/tr\u003e \u003ctr\u003e \u003ctd align=\"left\" colname=\"c1\"\u003e \u003cp\u003e2\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c2\"\u003e \u003cp\u003e3.0\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c3\"\u003e \u003cp\u003eWell-defined\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c4\"\u003e \u003cp\u003eGlandular/ Papillary\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c5\"\u003e \u003cp\u003eNo\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c6\"\u003e \u003cp\u003eCompactly\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c7\"\u003e \u003cp\u003eMild\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c8\"\u003e \u003cp\u003eInterspersed\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c9\"\u003e \u003cp\u003eBA\u003c/p\u003e \u003c/td\u003e \u003c/tr\u003e \u003ctr\u003e \u003ctd align=\"left\" colname=\"c1\"\u003e \u003cp\u003e3\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c2\"\u003e \u003cp\u003e4.8\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c3\"\u003e \u003cp\u003ePoorly defined\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c4\"\u003e \u003cp\u003ePapillary/ Glandular\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c5\"\u003e \u003cp\u003eNo\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c6\"\u003e \u003cp\u003eCompactly\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c7\"\u003e \u003cp\u003eMild\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c8\"\u003e \u003cp\u003eInterspersed\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c9\"\u003e \u003cp\u003eBA\u003c/p\u003e \u003c/td\u003e \u003c/tr\u003e \u003ctr\u003e \u003ctd align=\"left\" colname=\"c1\"\u003e \u003cp\u003e4\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c2\"\u003e \u003cp\u003e4.3\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c3\"\u003e \u003cp\u003ePoorly defined\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c4\"\u003e \u003cp\u003eGlandular\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c5\"\u003e \u003cp\u003eYes\u003c/p\u003e \u003cp\u003e(focally)\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c6\"\u003e \u003cp\u003eCompactly\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c7\"\u003e \u003cp\u003eMild\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c8\"\u003e \u003cp\u003eContinuous\u003c/p\u003e \u003cp\u003e(focally)\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c9\"\u003e \u003cp\u003eAtypical BA\u003c/p\u003e \u003c/td\u003e \u003c/tr\u003e \u003ctr\u003e \u003ctd align=\"left\" colname=\"c1\"\u003e \u003cp\u003e5\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c2\"\u003e \u003cp\u003e5.0\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c3\"\u003e \u003cp\u003eWell-defined\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c4\"\u003e \u003cp\u003eGlandular\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c5\"\u003e \u003cp\u003eNo\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c6\"\u003e \u003cp\u003eCompactly\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c7\"\u003e \u003cp\u003eMild\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c8\"\u003e \u003cp\u003eScattered\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c9\"\u003e \u003cp\u003eAtypical BA\u003c/p\u003e \u003c/td\u003e \u003c/tr\u003e \u003ctr\u003e \u003ctd align=\"left\" colname=\"c1\"\u003e \u003cp\u003e6\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c2\"\u003e \u003cp\u003e5.0\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c3\"\u003e \u003cp\u003eWell-defined\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c4\"\u003e \u003cp\u003ePapillary/ Glandular\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c5\"\u003e \u003cp\u003eNo\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c6\"\u003e \u003cp\u003eCompactly\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c7\"\u003e \u003cp\u003eMild\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c8\"\u003e \u003cp\u003eFew\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c9\"\u003e \u003cp\u003eAtypical BA\u003c/p\u003e \u003c/td\u003e \u003c/tr\u003e \u003ctr\u003e \u003ctd align=\"left\" colname=\"c1\"\u003e \u003cp\u003e7\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c2\"\u003e \u003cp\u003e7.5\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c3\"\u003e \u003cp\u003ePoorly defined\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c4\"\u003e \u003cp\u003eLepidic\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c5\"\u003e \u003cp\u003eNo\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c6\"\u003e \u003cp\u003eLoosely\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c7\"\u003e \u003cp\u003eHobnail cells\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c8\"\u003e \u003cp\u003eScattered\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c9\"\u003e \u003cp\u003eAIS\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c10\" morerows=\"1\" rowspan=\"2\"\u003e \u003cp\u003eLateral and basal membranes\u003c/p\u003e \u003c/td\u003e \u003c/tr\u003e \u003ctr\u003e \u003ctd align=\"left\" colname=\"c1\"\u003e \u003cp\u003e8\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c2\"\u003e \u003cp\u003e8\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c3\"\u003e \u003cp\u003ePoorly defined\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c4\"\u003e \u003cp\u003eGlandular/Lepidic\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c5\"\u003e \u003cp\u003eNo\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c6\"\u003e \u003cp\u003eLoosely\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c7\"\u003e \u003cp\u003eHobnail cells\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c8\"\u003e \u003cp\u003eNo\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c9\"\u003e \u003cp\u003eMIA\u003c/p\u003e \u003c/td\u003e \u003c/tr\u003e \u003ctr\u003e \u003ctd align=\"left\" colname=\"c1\"\u003e \u003cp\u003e9\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c2\"\u003e \u003cp\u003e18\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c3\"\u003e \u003cp\u003ePoorly defined\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c4\"\u003e \u003cp\u003ePapillary/ Glandular\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c5\"\u003e \u003cp\u003eNo\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c6\"\u003e \u003cp\u003eLoosely\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c7\"\u003e \u003cp\u003eMarked\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c8\"\u003e \u003cp\u003eScattered\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c9\"\u003e \u003cp\u003eILA*\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c10\"\u003e \u003cp\u003eNegative\u003c/p\u003e \u003c/td\u003e \u003c/tr\u003e \u003c/tbody\u003e \u003c/colgroup\u003e \u003ctfoot\u003e \u003ctr\u003e\u003ctd colspan=\"10\"\u003eBA: Bronchiolar adenoma; AIS: Lung adenocarcinoma in situ; MIA: minimally invasive adenocarcinoma; ILA: invasive lung adenocarcinoma; * invasive lung adenocarcinoma (ILA) with acinar and papillary growth patterns.\u003c/td\u003e\u003c/tr\u003e \u003ctr\u003e\u003ctd colspan=\"10\"\u003eSupplementary Fig.\u0026nbsp;1. Claudin18.2 expression in bronchiolar and pulmonary alveolar epithelium. (A) Terminal bronchiole lined by compactly arranged epithelial cells with smooth luminal surface, transitioning to alveolar epithelium characterized by loosely arranged cells with prominent intercellular gaps (H\u0026amp;E). (B) Claudin18.2 is negative in bronchiolar epithelium but weakly positive along lateral and basal membranes of alveolar epithelial cells.\u003c/td\u003e\u003c/tr\u003e \u003c/tfoot\u003e \u003c/table\u003e\u003c/div\u003e \u003c/p\u003e \u003cp\u003e \u003cstrong\u003eCases 1\u003c/strong\u003e \u003cp\u003e3 (Fig.\u0026nbsp;\u003cspan refid=\"Fig1\" class=\"InternalRef\"\u003e1\u003c/span\u003e) represent distal-type BAs (lesion diameters: 2.5\u0026ndash;4.8 mm). Case \u003cspan refid=\"FPar1\" class=\"InternalRef\"\u003e1\u003c/span\u003e (Fig.\u0026nbsp;\u003cspan refid=\"Fig1\" class=\"InternalRef\"\u003e1\u003c/span\u003eA\u0026ndash;D) showed a well-circumscribed lesion with glandular/papillary architecture (Fig.\u0026nbsp;\u003cspan refid=\"Fig1\" class=\"InternalRef\"\u003e1\u003c/span\u003eA). High-power examination revealed ciliated cells in compact arrangement with a distinct bilayered structure (Fig.\u0026nbsp;\u003cspan refid=\"Fig1\" class=\"InternalRef\"\u003e1\u003c/span\u003eB). P40 IHC confirmed a continuous basal cell layer (Fig.\u0026nbsp;\u003cspan refid=\"Fig1\" class=\"InternalRef\"\u003e1\u003c/span\u003eC), supporting the diagnosis. Case 2 (Fig.\u0026nbsp;\u003cspan refid=\"Fig1\" class=\"InternalRef\"\u003e1\u003c/span\u003eE\u0026ndash;H) also demonstrated well-defined borders. Tumor cells were arranged in dilated glandular/papillary structures separated by eosinophilic fibromyxoid stroma (Fig.\u0026nbsp;\u003cspan refid=\"Fig1\" class=\"InternalRef\"\u003e1\u003c/span\u003eE), with compact architecture and a smooth luminal surface. Most cells exhibited abundant apical \"snouting\" secretion and basally located nuclei (Fig.\u0026nbsp;\u003cspan refid=\"Fig1\" class=\"InternalRef\"\u003e1\u003c/span\u003eF). Although a distinct bilayered structure was not evident, P40 staining revealed interspersed basal cells (Fig.\u0026nbsp;\u003cspan refid=\"Fig1\" class=\"InternalRef\"\u003e1\u003c/span\u003eG). Case 3 (Fig.\u0026nbsp;\u003cspan refid=\"Fig1\" class=\"InternalRef\"\u003e1\u003c/span\u003eI\u0026ndash;L) showed poorly defined borders, with tumor cells in papillary or glandular patterns (Fig.\u0026nbsp;\u003cspan refid=\"Fig1\" class=\"InternalRef\"\u003e1\u003c/span\u003eI). Cells were compactly arranged with abundant apical secretion (Fig.\u0026nbsp;\u003cspan refid=\"Fig1\" class=\"InternalRef\"\u003e1\u003c/span\u003eJ). P40 staining revealed interspersed basal cells (Fig.\u0026nbsp;\u003cspan refid=\"Fig1\" class=\"InternalRef\"\u003e1\u003c/span\u003eK). Thus, despite absent bilayered structures, Cases 2 and 3 were diagnosed as distal-type BA based on scattered P40-positive basal cells and characteristic cytomorphology. All three cases demonstrated weak-to-moderate claudin18.2 positivity localized to the luminal surface (Fig.\u0026nbsp;\u003cspan refid=\"Fig1\" class=\"InternalRef\"\u003e1\u003c/span\u003eD, H, L).\u003c/p\u003e \u003cp\u003e \u003c/p\u003e \u003c/p\u003e \u003cp\u003eFigure \u003cspan refid=\"Fig2\" class=\"InternalRef\"\u003e2\u003c/span\u003e (Cases 4\u0026ndash;6) illustrates atypical distal-type BAs (lesion diameters: 2.5\u0026ndash;5.0 mm). Case 4 (Fig.\u0026nbsp;\u003cspan refid=\"Fig2\" class=\"InternalRef\"\u003e2\u003c/span\u003eA\u0026ndash;D) showed predominantly irregularly dilated glandular structures with variable calibers (Fig.\u0026nbsp;\u003cspan refid=\"Fig2\" class=\"InternalRef\"\u003e2\u003c/span\u003eA). Ciliated cells and a bilayered structure were restricted to focal areas. Cells showed compact arrangement with apical \"snouting\" secretion and basally located nuclei (Fig.\u0026nbsp;\u003cspan refid=\"Fig2\" class=\"InternalRef\"\u003e2\u003c/span\u003eB). P40 staining confirmed a focally continuous basal cell layer (Fig.\u0026nbsp;\u003cspan refid=\"Fig2\" class=\"InternalRef\"\u003e2\u003c/span\u003eC), supporting the diagnosis of atypical BA (partially monolayered BA). Case 5 (Fig.\u0026nbsp;\u003cspan refid=\"Fig2\" class=\"InternalRef\"\u003e2\u003c/span\u003eE\u0026ndash;H) demonstrated well-defined borders, with tumor cells arranged in dilated glandular structures separated by eosinophilic fibromyxoid stroma (Fig.\u0026nbsp;\u003cspan refid=\"Fig2\" class=\"InternalRef\"\u003e2\u003c/span\u003eE). No bilayered structure was identified; however, compactly arranged cells with abundant apical secretion and a smooth luminal surface were evident (Fig.\u0026nbsp;\u003cspan refid=\"Fig2\" class=\"InternalRef\"\u003e2\u003c/span\u003eF). P40 staining revealed scattered positive cells (Fig.\u0026nbsp;\u003cspan refid=\"Fig2\" class=\"InternalRef\"\u003e2\u003c/span\u003eG). Case 6 (Fig.\u0026nbsp;\u003cspan refid=\"Fig2\" class=\"InternalRef\"\u003e2\u003c/span\u003eI\u0026ndash;L) showed well-circumscribed borders, with tumor cells arranged in papillary or irregular glandular structures (Fig.\u0026nbsp;\u003cspan refid=\"Fig2\" class=\"InternalRef\"\u003e2\u003c/span\u003eI). Compact arrangement with abundant apical secretion was evident (Fig.\u0026nbsp;\u003cspan refid=\"Fig2\" class=\"InternalRef\"\u003e2\u003c/span\u003eJ). P40 staining revealed few positive cells (Fig.\u0026nbsp;\u003cspan refid=\"Fig2\" class=\"InternalRef\"\u003e2\u003c/span\u003eK). Based on morphology and P40 expression, Cases 5 and 6 were diagnosed as atypical BA (monolayered BA-like lesions). All three atypical BAs demonstrated claudin18.2 expression patterns identical to distal-type BAs\u0026mdash;weak-to-moderate luminal surface positivity (Fig.\u0026nbsp;\u003cspan refid=\"Fig2\" class=\"InternalRef\"\u003e2\u003c/span\u003eD, H, L).\u003c/p\u003e \u003cp\u003e \u003c/p\u003e \u003cp\u003eThe third group comprises histologic mimics of BA, including AIS, MIA, and ILAs with acinar and papillary growth patterns. Case 7 (AIS) showed poorly defined borders, with tumor cells arranged in a single layer demonstrating predominantly lepidic growth (Fig.\u0026nbsp;\u003cspan refid=\"Fig3\" class=\"InternalRef\"\u003e3\u003c/span\u003eA). Loosely arranged cells with \"hobnail\" appearance were evident (Fig.\u0026nbsp;\u003cspan refid=\"Fig3\" class=\"InternalRef\"\u003e3\u003c/span\u003eB). P40 staining revealed scattered positive cells (Fig.\u0026nbsp;\u003cspan refid=\"Fig3\" class=\"InternalRef\"\u003e3\u003c/span\u003eC). Claudin18.2 showed lateral and basal membrane positivity (Fig.\u0026nbsp;\u003cspan refid=\"Fig3\" class=\"InternalRef\"\u003e3\u003c/span\u003eD). Case 8 (MIA) showed dilated glandular structures centrally and lepidic growth peripherally (Fig.\u0026nbsp;\u003cspan refid=\"Fig3\" class=\"InternalRef\"\u003e3\u003c/span\u003eE). Most cells were loosely arranged, with prominent intercellular gaps, \"hobnail\" appearance, and uneven luminal surface (Fig.\u0026nbsp;\u003cspan refid=\"Fig3\" class=\"InternalRef\"\u003e3\u003c/span\u003eF). P40 staining was negative (Fig.\u0026nbsp;\u003cspan refid=\"Fig3\" class=\"InternalRef\"\u003e3\u003c/span\u003eG). Claudin18.2 positivity was localized to the lateral and basal membranes (Fig.\u0026nbsp;\u003cspan refid=\"Fig3\" class=\"InternalRef\"\u003e3\u003c/span\u003eH). Case 9 (ILAs, acinar/papillary type) showed a lesion diameter exceeding 15 mm. Tumor cells were predominantly arranged in irregular glands and papillary structures (Fig.\u0026nbsp;\u003cspan refid=\"Fig3\" class=\"InternalRef\"\u003e3\u003c/span\u003eI). Most cells showed marked atypia, with a nuclear-to-cytoplasmic ratio\u0026thinsp;\u0026gt;\u0026thinsp;0.5. Cellular stratification and intercellular gaps were observed, with uneven luminal surface (Fig.\u0026nbsp;\u003cspan refid=\"Fig3\" class=\"InternalRef\"\u003e3\u003c/span\u003eJ). P40 staining revealed scattered positive cells in focal areas (Fig.\u0026nbsp;\u003cspan refid=\"Fig3\" class=\"InternalRef\"\u003e3\u003c/span\u003eK). Claudin18.2 staining was negative (Fig.\u0026nbsp;\u003cspan refid=\"Fig3\" class=\"InternalRef\"\u003e3\u003c/span\u003eL).\u003c/p\u003e \u003cp\u003e \u003c/p\u003e"},{"header":"Discussion","content":"\u003cp\u003eIn recent years, atypical distal-type BAs have been reported, including partial monolayered BA and monolayered BA-like lesions (\u003cspan citationid=\"CR8\" class=\"CitationRef\"\u003e8\u003c/span\u003e, \u003cspan citationid=\"CR10\" class=\"CitationRef\"\u003e10\u003c/span\u003e). These variants have challenged the initial diagnostic criterion\u0026mdash;the bilayered structure with a continuous basal cell layer\u0026mdash;of this entity. Therefore, additional ancillary markers are required to distinguish BA from its histologic mimics in routine diagnostic practice.\u003c/p\u003e \u003cp\u003eIn this study, we describe the morphologic features of six distal-type BAs, including three atypical variants. All six distal-type BAs demonstrated compactly arranged cells lacking discernible intercellular gaps\u0026mdash;a key morphologic feature distinguishing this entity from AIS and MIA. Furthermore, we observed that claudin18.2 is expressed in distal-type BAs with characteristic luminal surface positivity. This luminal localization contrasts with the absence of claudin18.2 in bronchiolar epithelium and its lateral\u0026ndash;basal distribution in alveolar epithelium. Claudin18.2 is essential for maintaining tight junctions between epithelial cells (\u003cspan citationid=\"CR17\" class=\"CitationRef\"\u003e17\u003c/span\u003e). Previous studies have shown that carcinogenesis disrupts tight junctions in epithelial cells, resulting in claudin18.2 exposure (\u003cspan citationid=\"CR16\" class=\"CitationRef\"\u003e16\u003c/span\u003e, \u003cspan citationid=\"CR17\" class=\"CitationRef\"\u003e17\u003c/span\u003e). As distal-type BAs are believed to arise from terminal bronchiolar epithelium, this luminal localization may reflect distinct tight junction architecture in BAs, possibly related to their terminal bronchiolar origin and compact cellular arrangement.\u003c/p\u003e \u003cp\u003eIn AIS, MIA, and ILA with acinar/papillary growth patterns, IHC staining revealed claudin18.2 expression patterns distinct from those in BAs. Claudin18.2 was expressed in AIS and MIA with lateral and basal membrane positivity, resembling that of alveolar epithelium; however, it was absent in ILA with acinar and papillary patterns. We hypothesize that AIS and MIA share tight junction characteristics with alveolar epithelium, as these lesions exhibit loosely arranged cells and uneven luminal surfaces. Conversely, in ILA, tight junctions may be disrupted or undergo significant alterations, leading to complete loss of claudin18.2 expression.\u003c/p\u003e \u003cp\u003eOur findings align with a recent report demonstrating absent claudin18.2 expression in non-mucinous ILA with solid, papillary, acinar, or micropapillary patterns, in contrast to weak expression in lepidic-predominant adenocarcinoma (\u003cspan citationid=\"CR15\" class=\"CitationRef\"\u003e15\u003c/span\u003e). Collectively, these findings establish a diagnostic algorithm based on claudin18.2 localization: luminal-surface expression defines distal-type BA (including atypical variants); lateral and basal membrane positivity indicates AIS or MIA; and absent expression supports ILA. In routine diagnostic practice, we propose a tiered IHC staining strategy: initial assessment using P40 to evaluate basal cell layer integrity; when P40 shows indeterminate results (discontinuous or rare basal cells), claudin18.2 staining is added\u0026mdash;luminal-surface positivity supports BA diagnosis, whereas lateral\u0026ndash;basal membrane positivity or negative staining suggests non-BA lesions (AIS/MIA/ILA).\u003c/p\u003e \u003cp\u003eThis study has several limitations. First, the sample size is limited, with claudin18.2 expression examined in only six distal-type BAs (including three atypical variants). Moreover, proximal-type BAs were not included in this analysis. Consequently, the characteristic luminal surface expression pattern requires validation in a larger cohort, particularly one encompassing the full morphologic spectrum of BA subtypes. Second, integrated molecular profiling was not performed. Whether the observed claudin18.2 expression pattern correlates with specific genetic alterations (e.g., BRAF V600E or EGFR mutations) remains to be determined in future studies combining morphological, immunophenotypic, and molecular analyses.\u003c/p\u003e"},{"header":"Conclusions","content":"\u003cp\u003eThis study identifies characteristic luminal-surface claudin18.2 expression as a diagnostic marker for distal-type BAs, including atypical variants with incomplete or absent basal cell layers. When integrated with P40 immunostaining, claudin18.2 enables a tiered diagnostic approach to distinguish BA from AIS, MIA, and ILA with acinar and papillary growth patterns.\u003c/p\u003e"},{"header":"Declarations","content":"\u003cp\u003e\u003cstrong\u003e1. Ethics approval and consent to participate\u003c/strong\u003e\u003c/p\u003e\n\u003cp\u003eThe study was conducted in accordance with the Declaration of Helsinki and was approved by the Institutional Review Board (IRB) of the Seventh Affiliated Hospital of Sun Yat-sen University. The requirement for informed consent was waived due to the retrospective nature by the IRB of the Seventh Affiliated Hospital (application ID: KY-2025-399-01).\u003c/p\u003e\n\u003cp\u003e\u003cstrong\u003e2. Consent for publication\u003c/strong\u003e\u003c/p\u003e\n\u003cp\u003eWritten informed consent for the publication of potentially identifiable information (age, sex, and pathological images, but excluding photographs with facial features, ID numbers, and hospital admission numbers) was obtained from all patients.\u003c/p\u003e\n\u003cp\u003e\u003cstrong\u003e3. Availability of data and materials\u003c/strong\u003e\u003c/p\u003e\n\u003cp\u003eAggregated data are available from the corresponding author upon reasonable request.\u003c/p\u003e\n\u003cp\u003e\u003cstrong\u003e4. Competing interests\u003c/strong\u003e\u003c/p\u003e\n\u003cp\u003eThe authors declare no competing interests.\u003c/p\u003e\n\u003cp\u003e\u003cstrong\u003e5. Funding\u003c/strong\u003e\u003c/p\u003e\n\u003cp\u003eNo funding was involved in this study.\u003c/p\u003e\n\u003cp\u003e\u003cstrong\u003e6. Authors\u0026apos; contributions\u003c/strong\u003e\u003c/p\u003e\n\u003cp\u003eQCF, LXY and XCY: Literature search, study design and concept drafting, references and writing of the manuscript.\u003c/p\u003e\n\u003cp\u003eLXL and ZTZ: Literature search, study design, writing of manuscript and interpretation of results.\u003c/p\u003e\n\u003cp\u003eLDZ: IHC staining and review of the final manuscript.\u003c/p\u003e\n\u003cp\u003e\u003cstrong\u003e7. Acknowledgment\u003c/strong\u003e\u003c/p\u003e\n\u003cp\u003eNone \u0026nbsp;\u003c/p\u003e"},{"header":"References","content":"\u003col\u003e\u003cli\u003e\u003cspan\u003eKamata T, Yoshida A, Kosuge T, Watanabe S, Asamura H, Tsuta K. Ciliated muconodular papillary tumors of the lung: a clinicopathologic analysis of 10 cases. Am J Surg Pathol. 2015;39(6):753\u0026ndash;60.\u003c/span\u003e\u003c/li\u003e \u003cli\u003e\u003cspan\u003eChang JC, Montecalvo J, Borsu L, Lu S, Larsen BT, Wallace WD, et al. Bronchiolar Adenoma: Expansion of the Concept of Ciliated Muconodular Papillary Tumors With Proposal for Revised Terminology Based on Morphologic, Immunophenotypic, and Genomic Analysis of 25 Cases. Am J Surg Pathol. 2018;42(8):1010\u0026ndash;26.\u003c/span\u003e\u003c/li\u003e \u003cli\u003e\u003cspan\u003eNicholson AG, Tsao MS, Beasley MB, Borczuk AC, Brambilla E, Cooper WA, et al. The 2021 WHO Classification of Lung Tumors: Impact of Advances Since 2015. J Thorac oncology: official publication Int Association Study Lung Cancer. 2022;17(3):362\u0026ndash;87.\u003c/span\u003e\u003c/li\u003e \u003cli\u003e\u003cspan\u003eZhu M, Yang Q, Zhan S, Liu W, Liu W, Guo L, et al. Clinicopathological analysis of bronchiolar adenoma combined with lung adenocarcinoma: Report of eight cases and literature review. Histol Histopathol. 2024;39(6):783\u0026ndash;94.\u003c/span\u003e\u003c/li\u003e \u003cli\u003e\u003cspan\u003eShirsat H, Zhou F, Chang JC, Rekhtman N, Saqi A, Argyropoulos K, et al. Bronchiolar Adenoma/Pulmonary Ciliated Muconodular Papillary Tumor. Am J Clin Pathol. 2021;155(6):832\u0026ndash;44.\u003c/span\u003e\u003c/li\u003e \u003cli\u003e\u003cspan\u003eLiu S, Cai X, Pan J, Liu S, Lin J, Yue X. Bronchiole adenoma/pulmonary ciliated mucinous nodular papillary tumor: Case series and literature review. Medicine. 2023;102(50):e36559.\u003c/span\u003e\u003c/li\u003e \u003cli\u003e\u003cspan\u003eShao J, Yin JC, Bao H, Zhao R, Han Y, Zhu L, et al. Morphological, immunohistochemical, and genetic analyses of bronchiolar adenoma and its putative variants. J Pathol Clin Res. 2021;7(3):287\u0026ndash;300.\u003c/span\u003e\u003c/li\u003e \u003cli\u003e\u003cspan\u003eBo J, Chen X, Zhang T, Zhu X, Zhang L, Liu Y, et al. Clinicopathological features and genomic analysis of bronchiolar adenoma. Histol Histopathol. 2023;38(12):1465\u0026ndash;74.\u003c/span\u003e\u003c/li\u003e \u003cli\u003e\u003cspan\u003eDing B, Shang Z, Xiang Z, Han Y. Clinicopathologic Features and Frozen Diagnostic Pitfalls of Bronchiolar Adenoma/Ciliated Muconodular Papillary Tumors (BA/CMPTs). Am J Surg Pathol. 2023;47(4):431\u0026ndash;9.\u003c/span\u003e\u003c/li\u003e \u003cli\u003e\u003cspan\u003eLin DL, Ding L, Shao SH, Xin FJ, Zhang LX, Li GQ, et al. 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Pores in the wall: claudins constitute tight junction strands containing aqueous pores. J Cell Biol. 2000;149(1):13\u0026ndash;6.\u003c/span\u003e\u003c/li\u003e \u003cli\u003e\u003cspan\u003eZhang J, Dong R, Shen L. Evaluation and reflection on claudin 18.2 targeting therapy in advanced gastric cancer. Chin J cancer Res = Chung-kuo yen cheng yen chiu. 2020;32(2):263\u0026ndash;70.\u003c/span\u003e\u003c/li\u003e \u003cli\u003e\u003cspan\u003eYan P, Dong Y, Zhang F, Zhen T, Liang J, Shi H, et al. Claudin18.2 expression and its clinicopathological feature in adenocarcinoma from various parts. J Clin Pathol. 2025;78(12):815\u0026ndash;21.\u003c/span\u003e\u003c/li\u003e \u003cli\u003e\u003cspan\u003eSahin U, Koslowski M, Dhaene K, Usener D, Brandenburg G, Seitz G, et al. Claudin-18 splice variant 2 is a pan-cancer target suitable for therapeutic antibody development. Clin cancer research: official J Am Association Cancer Res. 2008;14(23):7624\u0026ndash;34.\u003c/span\u003e\u003c/li\u003e \u003cli\u003e\u003cspan\u003ePak S, Simon AG, Lyu SI, Tolkach Y, Alakus H, Zander T, et al. The expression of the tight junction protein and therapeutical target Claudin 18.2 is heterogeneously distributed within esophageal and gastric adenocarcinoma. Sci Rep. 2025;15(1):28958.\u003c/span\u003e\u003c/li\u003e\u003c/ol\u003e"}],"fulltextSource":"","fullText":"","funders":[],"hasAdminPriorityOnWorkflow":false,"hasManuscriptDocX":true,"hasOptedInToPreprint":true,"hasPassedJournalQc":"","hasAnyPriority":false,"hideJournal":true,"highlight":"","institution":"","isAcceptedByJournal":false,"isAuthorSuppliedPdf":false,"isDeskRejected":"","isHiddenFromSearch":false,"isInQc":false,"isInWorkflow":false,"isPdf":false,"isPdfUpToDate":true,"isWithdrawnOrRetracted":false,"journal":{"display":true,"email":"[email protected]","identity":"researchsquare","isNatureJournal":false,"hasQc":true,"allowDirectSubmit":true,"externalIdentity":"","sideBox":"","snPcode":"","submissionUrl":"/submission","title":"Research Square","twitterHandle":"researchsquare","acdcEnabled":true,"dfaEnabled":false,"editorialSystem":"","reportingPortfolio":"","inReviewEnabled":false,"inReviewRevisionsEnabled":true},"keywords":"Distal-type Bronchiolar Adenoma, Atypical distal-type Bronchiolar Adenoma, Adenocarcinoma, Claudin18.2, Immunohistochemical staining","lastPublishedDoi":"10.21203/rs.3.rs-9167542/v1","lastPublishedDoiUrl":"https://doi.org/10.21203/rs.3.rs-9167542/v1","license":{"name":"CC BY 4.0","url":"https://creativecommons.org/licenses/by/4.0/"},"manuscriptAbstract":"\u003ch2\u003eBackground\u003c/h2\u003e \u003cp\u003eDistal-type bronchiolar adenoma (BA) is a benign lung neoplasm characterized by a bilayered structure. However, atypical variants lacking continuous basal cell layers pose diagnostic challenges, particularly in distinguishing them from adenocarcinoma in situ (AIS), minimally invasive adenocarcinoma (MIA), and invasive lung adenocarcinoma (ILA) with acinar and papillary growth patterns.\u003c/p\u003e\u003ch2\u003eAim\u003c/h2\u003e \u003cp\u003eTo investigate the immunohistochemical (IHC) expression of claudin18.2 in distal-type BA and assess its diagnostic utility in differentiating BA from histologic mimics.\u003c/p\u003e\u003ch2\u003eMethods\u003c/h2\u003e \u003cp\u003eDistal-type BA, atypical distal-type BA, AIS, MIA, and ILAs with acinar and papillary growth patterns were collected and subjected to IHC staining for claudin18.2.\u003c/p\u003e\u003ch2\u003eResults\u003c/h2\u003e \u003cp\u003eClaudin18.2 was expressed in all distal-type BAs (100%, n\u0026thinsp;=\u0026thinsp;6), including 3 atypical variants, with characteristic weak-to-moderate luminal-surface positivity. In contrast, AIS and MIA cases showed lateral and basal membrane staining (100%, n\u0026thinsp;=\u0026thinsp;6), whereas the ILA cases were negative (0%, n\u0026thinsp;=\u0026thinsp;3).\u003c/p\u003e\u003ch2\u003eConclusion\u003c/h2\u003e \u003cp\u003eClaudin18.2 may serve as a valuable IHC marker for diagnosing distal-type BAs and distinguishing atypical variants from their histologic mimics.\u003c/p\u003e","manuscriptTitle":"Luminal-Surface Claudin18.2 Expression Distinguishes Distal-Type Bronchiolar Adenoma from Mimicking Lesions: An Immunohistochemical Study","msid":"","msnumber":"","nonDraftVersions":[{"code":1,"date":"2026-04-23 15:09:31","doi":"10.21203/rs.3.rs-9167542/v1","editorialEvents":[{"type":"communityComments","content":0}],"status":"published","journal":{"display":true,"email":"[email protected]","identity":"researchsquare","isNatureJournal":false,"hasQc":true,"allowDirectSubmit":true,"externalIdentity":"","sideBox":"","snPcode":"","submissionUrl":"/submission","title":"Research Square","twitterHandle":"researchsquare","acdcEnabled":true,"dfaEnabled":false,"editorialSystem":"","reportingPortfolio":"","inReviewEnabled":false,"inReviewRevisionsEnabled":true}}],"origin":"","ownerIdentity":"d8ee0d82-1927-4f8a-8acc-df8072a0a16d","owner":[],"postedDate":"April 23rd, 2026","published":true,"recentEditorialEvents":[],"rejectedJournal":[],"revision":"","amendment":"","status":"posted","subjectAreas":[],"tags":[],"updatedAt":"2026-04-26T12:09:43+00:00","versionOfRecord":[],"versionCreatedAt":"2026-04-23 15:09:31","video":"","vorDoi":"","vorDoiUrl":"","workflowStages":[]},"version":"v1","identity":"rs-9167542","journalConfig":"researchsquare"},"__N_SSP":true},"page":"/article/[identity]/[[...version]]","query":{"redirect":"/article/rs-9167542","identity":"rs-9167542","version":["v1"]},"buildId":"XKTyCvWXoU3ODBz1xrDgd","isFallback":false,"isExperimentalCompile":false,"dynamicIds":[84888],"gssp":true,"scriptLoader":[]}