Collagen fiber features and COL1A1: are they associated with elastic parameters in breast lesions and can COL1A1 predict axillary lymph node metastasis?

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Ying Jiang, Bo Wang, Jun Kang Li, Shi Yu Li, Rui Lan Niu, Nai Qin Fu, and 3 more This is a preprint; it has not been peer reviewed by a journal. https://doi.org/ 10.21203/rs.3.rs-1517751/v1 This work is licensed under a CC BY 4.0 License Status: Under Review Version 1 posted 3 You are reading this latest preprint version Abstract Purpose: This study aimed to explore whether collagen fiber features and collagen type I alpha 1 (COL1A1) are related to the stiffness of breast lesions and whether COL1A1 can predict axillary lymph node metastasis (LNM). Methods: A prospective analysis was performed in our hospital. A total of 94 breast lesions in 94 patients were enrolled between May 2021 to December 2021. Ultrasound (US) and shear wave elastography (SWE) examinations were performed for every lesion before surgery. Pathology results obtained by surgery were regarded as gold criteria. Sirius red staining and Immunohistochemical were used to examine the collagen fibers shape and arrangement features and COL1A1 expression of included tissue samples, and analyze the correlation between SWE parameters and them. To analyze the effectiveness of COL1A1 expression level in predicting axillary LNM. Results: The optimal cutoff values for Emax, Emean, and Eratio for diagnosis of benign group and malignant group, were 58.70 kPa, 52.50 kPa, and 3.05. The optimal cutoff values for diagnosing axillary LNM were 107.5 kPa, 85.15 kPa, and 3.90, respectively. In the present study, collagen fiber shape and arrangement features in breast lesions were classified into three categories. For all 94 lesions, one-way ANOVA showed that Emax, Emean and Eratio were different between categories 0, 1, and 2 (all P < 0.05). The correlation coefficient between Emax and collagen category was 0.318 ( P < 0.001), between Emean and collagen category was 0.261 ( P = 0.001). And between Eratio and collagen category was 0.349 ( P 0.145 was taken as the cut-off value, its efficacy in benign and malignant breast lesions was 0.808, with a sensitivity of 66%, and a specificity 90%. COL1A1 expression level >0.150 was taken as the cut-off value, its efficacy in predicting axillary LNM was 0.796, with a sensitivity of 96%, and a specificity 59%. Conclusions: The expression level of the collagen fiber features and COL1A1 were positively correlated with the elastic parameters of breast lesions. The expression of COL1A1 might be helpful to diagnose benign and malignant breast lesions and predict axillary LNM. SWE COL1A1 Breast lesions Lymph node metastasis Collagen Figures Figure 1 Figure 2 Figure 3 Figure 4 Introduction The incidence of female breast cancer in China is increasing year by year, which seriously affects women's quality of life[ 1 ]. The axillary lymph nodes are early metastatic sites of breast cancer, and their proper evaluation preoperatively is key to the formulation of rational treatment regimens. Stiffness is an important characteristic of tissues and organs and is the basis for clinicians to diagnose breast abnormalities. The advent of shear wave elastography (SWE) has provided the possibility of achieving quantitative judgment of tissue stiffness. Recently, several studies have reported that SWE showed good performance in assessing benign and malignant lesions [ 2 – 4 ]. The stromal reaction that occurs due to invasive behavior results in abnormal tumor-associated collagen deposition, potentially leading to increased elastic modulus value in breast cancer[ 5 ]. Studies have shown that the elastic parameters of SWE, could be useful for predicting the nodal status of invasive breast cancers (IBCs), but the pathological basis for the diagnosis of axillary lymph node metastasis is unclear [ 6 ]. Previous studies of SWE and breast tissue stroma focused on the relationship between quantity of collagen or fibrosis and elastic parameters. However, the influence of the shape and arrangement features of collagen fibers on the stiffness of extracellular matrix (ECM) is rarely considered. Malignant lesions are stiffer than benign ones because of the presence of desmoplastic ECM, with collagen type I being a major structural component [ 7 ]. The main function of collagen type I is to be the support of organs and tissues and make tissues form tension [ 8 ]. At the same time, collagen type I is also closely related to cell growth, proliferation, differentiation and inflammation [ 9 ]. Collagen type I consists of a triple helix structure consisting of α1(I) chains (COL1A1) and α2(I) chains (COL1A2).The two polypeptide chains are encoded by different genes and are generally synthesized in a ratio of 2:1[ 10 ]. Nevertheless, little is known about the functional role of COL1A1 in the elastography features differences and axillary LNM of breast lesions. The focus of this study was to explore the relationship between the shape and arrangement features of collagen fibers and the elastic parameters of breast lesions evaluated by SWE. To evaluate the role of COL1A1 expression in the differential diagnosis of benign and malignant breast lesions and to analyze whether COL1A1 can predict axillary LNM. Materials And Methods Patients From May 2021 to December 2021, 94 lesions in 94 patients were recruited to this prospective study, and were examined with conventional US and SWE prior to surgical resection. The inclusion criteria were newly hospitalized women diagnosed with breast lesions who underwent US and SWE examinations before biopsy. The exclusion criteria included the following: biopsy before US and SWE, undergoing neoadjuvant chemotherapy before US and SWE, and patients with pathologically multifocal IBCs. The pathology results obtained by surgery were regarded as gold criteria. The trial was conducted in accordance with the Declaration of Helsinki (as revised in 2013). The study was approved by the Ethics Committee of the Chinese PLA General Hospital (No. S2021-683-01), and informed consent was taken from all individual participants. According to the postoperative pathology, they were divided into benign group and malignant group. The malignant group was divided into LNM – (negative axillary lymph node metastasis) group and LNM + (positive axillary lymph node metastasis) group. Preoperative US and SWE examination Using AixPlorer US system (SuperSonic Imagine, Aix-en-Provence, France) with a L14-5 linear array probe operating at 4–15 MHz, SWE was performed after conventional US. The same sonographer with more than 10 years of clinical experience, performed all US and SWE examinations. The probe was applied as lightly as possible to avoid too much pressure and was maintained as steadily as possible for at least 10–20 seconds during elastic image acquisition. Participants were asked to hold their breath to prevent motion artefacts if needed. To enable a comparison with histopathology data, the maximum imaging section of the lesion was obtained to measure the stiffness of the lesion. Conventional ultrasound was used to obtain the maximum imaging section of the lesion, and then start the SWE check, freeze, and measure. Pay attention to adjusting ROI to include any halo or inelastic abnormal edge lesions, while minimizing any normal tissue. The Eratio was obtained by placing the ROI in normal breast tissue at the same depth as the lesion. The above process was repeated for three times, and the final value of Emax, Emean, Emin and Eratio was the average value of these three repetitions. Collagen staining and categorization Sirius red staining was used to assess collagen fibers shape and arrangement features and immunohistochemical staining was used to examine the expression of COL1A1. An OLYMPUS BX53P microscope equipped with a polarisedfilter. Observe the collagen fibers at 400x magnification. In this study, collagen fibers shape and arrangement feature in breast lesions were classified into three categories. The definitions are described according to the following descriptions: category 0: wavy collagen fibers similar to collagen fibers in normal breast tissue; category 1: taut parallel collagen fibers around tumor nests; category 2: straightened and aligned collagen fibers tending to be perpendicular to the tumor boundary[ 11 – 14 ]. Analysis and classification of ECM collagen fibers in each section by a physician majoring in histopathology for 5 years, unaware of SWE results and surgical pathology results. COL1A1 was detected by immunohistochemistry (IHC). The staining results of IHC were analyzed using Image-Pro Plus 6.0 software (Media Cybernetics, Rockville, MD, USA). A total of 5 areas of interest were taken from each slice under 200× field of view and photographed. After measuring the integrated optical density (IOD) and area, average optical density (AOD = IOD/Area) was used to evaluate the expression level of COL1A1. Statistical analysis All statistical analyses were performed with SPSS 25.0, standard version (SPSS, Chicago, IL, USA). The measurement data were expressed as mean ± SD and count data as percentage (%). Independent sample t-test was used to compare the COL1A1 expression level, Emax, Emean, Emin and Eratio of benign and malignant breast lesions. One-way analysis of variance (ANOVA)was used to compare the values of Emax, Emean and Eratio between different collagen fibers categories in 94 breast lesions. The chi-squared test was used to compare the rate between groups. Receiver operating Characteristic (ROC) curves were constructed for SWE values and the COL1A1 expression levels in differentiating benign from malignant breast lesions and predicting the axillary LNM. And the corresponding sensitivity and specificity were recorded (95% confidence interval). The correlation between COL1A1 expression level and elastic parameters in breast lesions were evaluated by Pearson correlation analysis, and the coefficients were defined as follows: 0.75 indicated excellent reliability[ 15 ]. Kendall's tau-b correlation coefficient was used to determine the correlation between different collagen fiber features and SWE parameters in 94 cases of breast lesions. A value of P < 0.05 was considered statistically significant. Results Clinicopathological characteristics A total of 94 breast tumors were surgically removed and histopathologically approved, and included 30 benign lesions and 64 malignant lesions. Benign lesions consisted of fibroadenoma (n = 17), sclerosing adenosis(n = 5), inflammatory lesions(n = 3), intraductal papilloma(n = 4) and benign phyllode tumor(n = 1). Malignant lesions consisted of invasive cancers(n = 55), Intraductal cancers(n = 5), and mucinous cancers(n = 4). The presence of axillary lymph nodes was determined according to postoperative pathology, and breast malignant tumors were divided into LNM + and LNM - groups. Of the 64 malignant tumors, 25(39%) were positive axillary lymphatic metastases, and 39(61%) were clinically negative. There were no statistically significant differences in age and tumor size between groups (Table 1 ). Elastic parameters, collagen fiber features and COL1A1 expression Ultrasonic elastography, Sirius red staining and COL1A1 immunohistochemical staining of included 94 samples are shown in Fig. 3 . The Emax, Emean and Eratio of malignant breast lesions were significantly higher than those of benign lesions ( P < 0.001; Table 1 ), while the Emin of malignant breast lesions and benign lesions was not significantly different ( P = 0.135). For the diagnosis of malignant group and benign group, the ROC curves of the Emax, Emean and Eratio elasticity are shown in Fig. 2 . The optimal cutoff values, yielding the maximum of sensitivity and specificity, were greater than 58.70 kPa, 52.50 kPa, and 3.05. The areas under the ROC curves were 0.853(0.779–0.928), 0.823(0.737–0.910), and 0.882(0.813–0.951), respectively. The best cutoff values of SWE for diagnosing lymphatic metastasis were as follows: 107.5 kPa for Emax, 85.15 kPa for Emean, and 3.90 for Eratio. Eratio showed the best diagnostic performance according to the area under the curve (AUC), with a 95% confidence interval (CI) of 0.845 (0.754–0.937). Pairwise comparison of SWE parameters of benign group, LNM- group and LNM + group showed that the difference was statistically significant (Fig. 1 ). In the present study, collagen fiber shape and arrangement features in breast lesions were classified into three categories. For all 94 lesions, one-way ANOVA showed that Emax, Emean and Eratio were different between categories 0, 1, and 2 (all P < 0.05; Table 2 ). This shows that Emax, Emean and Eratio increase when collagen fiber class is increased from 0 to 2. Kendall's tau-b correlation coefficient calculation shows that the correlation coefficient between Emax and collagen category was 0.318, between Emean and collagen category was 0.261, and between Eratio and collagen category was 0.349 (all P ≤ 0.001; Table 3 ). The mean expression level of COL1A1 in benign breast lesions was 0.105 ± 0.323 and malignant breast lesions mean expression level was 0.159 ± 0.047 ( P = 0.012). COL1A1 expression level was used in the differential diagnosis of benign and malignant breast lesions. The average optical density > 0.145 was the cut-off value, the effective rate was 0.808 (0.723–0.894), and the sensitivity and specificity were 66% and 90% respectively. When the expression level of COL1A1 was used to diagnose LNM - and LNM + malignant breast lesions, the average optical density > 0.150 was taken as the cut-off value, the effectiveness was 0.80 (0.69–0.91), and the sensitivity and specificity were 96% and 59% respectively (Fig. 2 ). Pairwise comparison of COL1A1 expression levels of benign group, LNM- group and LNM + group showed that the difference was statistically significant (Fig. 1 ). The Pearson correlation analysis showed a positive correlation between Emax, Emean, Eratio of breast lesions and the expression level of COL1A1 (r = 0.406, 0.362, 0.425, respectively. P < 0.001; Fig. 4 ). Table 1 Basic information, elastic parameters and COL1A1 expression of enrolled cases. Parameters Benign(n = 30) Malignant(n = 64) t/χ2 P value Age(years) 44.230 ± 8.947 47.700 ± 9.146 0.018 0.894 Size(cm) 2.003 ± 0.615 2.233 ± 0.679 0.847 0.360 Elastic parameters Emax(kPa) 44.450 ± 20.960 106.709 ± 55.490 20.853 < 0.001 Emean(kPa) 33.687 ± 18.466 68.155 ± 33.830 9.803 0.002 Emin(kPa) 20.320 ± 14.422 26.342 ± 18.469 2.272 0.135 Eratio 1.927 ± 0.516 4.492 ± 2.307 20.540 < 0.001 Expression of COL1A1(AOD) 0.105 ± 0.323 0.159 ± 0.047 6.564 0.012 Emax, maximum elasticity; Emean, mean elasticity; Emin, minimum elasticity; Eratio, the elasticity ratio of the lesions to the peripheral tissue; COL1A1, Collagen type I alpha 1; AOD, average optical density. Table 2 Emax, Emean and Eratio between different collagen fibers categories in 94 breast lesions. SWE values Category of collagen fibers P value 0 1 2 Emax(kPa) 64.670 ± 51.160 93.100 ± 54.654 112.010 ± 51.427 0.002 Emean(kPa) 47.280 ± 32.654 55.550 ± 31.357 73.440 ± 33.236 0.008 Eratio 2.710 ± 1.480 3.450 ± 1.617 5.340 ± 2.910 < 0.001 Emax, maximum elasticity; Emean, mean elasticity; Eratio, the elasticity ratio of the lesions to the peripheral tissue. Table 3 Correlation between different collagen fibers categories and SWE parameters in 94 breast lesions. N Kendall’s tau-b correlation coefficient P value Emax vs. collagen fibers categories 94 0.318 < 0.001 Emean vs. collagen fibers categories 94 0.261 0.001 Eratio vs. collagen fibers categories 94 0.349 < 0.001 Emax, maximum elasticity; Emean, mean elasticity; Eratio, the elasticity ratio of the lesions to the peripheral tissue. Discussion Shear wave elastography is a new ultrasound diagnosis technology. It uses shear wave of the tissue induced by acoustic radiation pulse to visualize and quantify the stiffness of tissue in a real-time, reliable, and reproducible manner[ 16 ]. Tissue stiffness has become an important parameter in diagnosing potential malignancies or other diseases[ 17 ]. For breast lesions, elastic parameters play an important role in the differential diagnosis of benign and malignant breast lesions. Our study showed that compared with benign breast lesions, malignant lesions had higher Emax, Emean, Eratio, which was consistent with the results of several previous studies[ 18 – 20 ]. Considering that preoperative knowledge of axillary lymph node status is crucial for the use of neoadjuvant chemotherapy and appropriate surgical treatment, it is necessary to develop imaging tools for LNM diagnosis[ 21 , 22 ]. Our study showed that the best cutoff values of the SWE parameters for diagnosing lymphatic metastasis were calculated as follows: 107.5 kPa for Emax, 85.15 kPa for Emean, and 3.9 for Eratio. According to ROC analysis, Eratio exhibited the best diagnostic performance with an AUC of 0.85 (0.75–0.94). A study[ 23 ] shows that lesion stiffness is a predictor of axillary lymph node metastasis in breast cancer, which combined with Breast Imaging Reporting and Data System (BI-RADS) scores reported AUC for predicting axillary lymph node metastasis status, Emean, Emax and Emin were 0.73 (0.67–0.80), 0.75 (0.69–0.81) and 0.69 (0.62–0.76), respectively. Xin Wen et al. [ 24 ] reported that the best cutoff values of the SWE parameters for diagnosing lymphatic metastasis were calculated as follows: 111.05 kPa for Emax, 79.80 kPa for Emean, and 6.89 for EmeanR. According to ROC analysis, Emax exhibited the best diagnostic performance with an AUC of 0.82 (0.76–0.87). This difference may be caused by the following three factors: fewer cases were included in our study; the setting of the instrument and how to obtain Eratio (there is no relevant information in Xin Wen's report); operator dependent changes. The present study demonstrated that the shape and arrangement of ECM collagen fibers are divided into three categories, which positively correlated with Emax, Emean and Eratio of breast lesion stiffness assessed by SWE. According to the results of all 94 lesions, the Emax, Emean and Eratio showed an increasing tendency from category 0 to category 2 (All P < 0.05). In normal breast tissue, collagen fibers showed wavy or sinusoidal waves[ 11 , 12 ]. Thus, this type of collagen fibers was assigned to category 0. Brabrand et al. reported that collagen in the peritumoural region was arranged in a more parallel alignment, Aligned or linearized collagen correlated with increased stiffness of breast tissue or lesions[ 8 , 25 ]. We classify this type as class 1. The type 2 is characterized by bundles of straightened collagen fibers perpendicular to the tumor boundary, which can change in a honeycomb shape and show high hardness[ 14 ].These results indicated that breast lesion stiffness will increase when ECM collagen changes into taut, straightened, and arranged parallel fibers. Additionally, a more complicated structure, collagen fibers that was connected in a honeycomb arrangement, indicated the most stiffness. It indicated that ECM collagen fibers shape and arrangement may have also contributed to stiffness variance in malignant breast lesions. Survival of breast cancer patients is correlated with stromal biology including the reorganization of the ECM to promote cancer invasion and migration[ 26 ]. Nazim Uddin's team suggests that the expression of stromal genes COL1A1 has an association with the progression of breast cancers and recurrence-free survival in breast cancer patients[ 27 ]. In our cohort, there were 30 tissue samples in the benign group, 64 tissue samples in the malignant group consecutively included for COL1A1 immunohistochemical staining. The Pearson correlation analysis showed a positive correlation between Emax, Emean, Eratio of breast lesions and the expression level of COL1A1 (r = 0.406, 0.362, 0.425, respectively. P < 0.001). The average expression level of COL1A1 in benign breast lesions was significantly lower than that in malignant breast lesions. COL1A1 expression level can be used to diagnose lymph node metastasis. The average optical density > 0.150 was taken as the cut-off value, the effectiveness was 0.80 (0.69–0.91), and the sensitivity and specificity were 96% and 59% respectively. Many studies[ 28 , 29 ] have also proved the correlation between collagen fiber content and SWE in breast lesions, and proved that the change of collagen fiber content leads to the change of tumor tissue hardness. F F Li [ 30 ] et al. studied 148 triple-negative breast cancers and found that the high expression of COL1A1 in triple-negative breast cancer was an independent prognostic factor. Protein-protein interaction network analysis confirmed that COL1A1 is a prognostic matrix gene in breast cancer, and its expression is related to the progression of breast cancer[ 27 ].These results suggest that COL1A1 may regulate tumor metastasis through some signal pathways; we have begun to investigate the possible signaling mechanisms using in vivo and in vitro methods. There was one limitation in this study that should be addressed. Correlations between stiffness and other components of the ECM, such as elastic fibers, laminin, and fibronectinwere not included; however, collagen is the most dominant structural protein in the ECM and its correlation with the elastic modulus could be the most important. Conclusion The expression level of the collagen fiber features and COL1A1 were positively correlated with the elastic parameters of breast lesions. The expression of COL1A1 might be helpful to diagnose benign and malignant breast lesions and predict axillary lymph node metastasis. Declarations Ethical approval and consent to participants The study was approved by the Ethics Committee of the Chinese PLA General Hospital (No. S2021-683-01), and informed consent was taken from all individual participants. Consent for publication Not applicable. Data availability All data generated or analyzed during this study are included in this published article. Author contributions : Conception and design: Ying Jiang, Zhi Li Wang; (II) Acquisition of study materials or patients: All authors; (III) Data analysis and interpretation: Ying Jiang, Bo Wang; (IV) Drafting the article: All authors. (V) Final approval of the version to be published: Gang Liu, Zhi Li Wang. Conflict of interest Conflict of interest statement: All authors disclosed no relevant relationships. Funding This study was supported by grants from the National Natural Science Foundation of China (No.82071925). Acknowledgments Not applicable. References Li, T., C. Mello-Thoms, and P.C. Brennan, Descriptive epidemiology of breast cancer in China: incidence, mortality, survival and prevalence. Breast Cancer Res Treat, 2016. 159 (3): p. 395-406. Suvannarerg, V., et al., Diagnostic performance of qualitative and quantitative shear wave elastography in differentiating malignant from benign breast masses, and association with the histological prognostic factors. Quant Imaging Med Surg, 2019. 9 (3): p. 386-398. Wang, Z.L., et al., Study of quantitative elastography with supersonic shear imaging in the diagnosis of breast tumours. Radiol Med, 2013. 118 (4): p. 583-90. 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A premise for tumour invasiveness? Apmis, 2015. 123 (1): p. 1-8. Conklin, M.W. and P.J. Keely, Why the stroma matters in breast cancer: insights into breast cancer patient outcomes through the examination of stromal biomarkers. Cell Adh Migr, 2012. 6 (3): p. 249-60. Uddin, M.N. and X. Wang, Identification of key tumor stroma-associated transcriptional signatures correlated with survival prognosis and tumor progression in breast cancer. Breast Cancer, 2022. Liu, G., et al., Shear wave elasticity of breast lesions: would it be correlated with the extracellular matrix components? Gland Surg, 2019. 8 (4): p. 399-406. Wang, Z.L., et al., Relationship between elasticity and collagen fiber content in breast disease: a preliminary report. Ultrasonics, 2015. 57 : p. 44-9. Li, F.F., J.J. Fan, and B.L. Ma, [Effect of high expression of type Ⅰ collagen α1 chain protein on the prognosis of triple negative breast cancer]. Zhonghua Zhong Liu Za Zhi, 2020. 42 (2): p. 122-126. Additional Declarations No competing interests reported. Cite Share Download PDF Status: Under Review Version 1 posted Editor invited by journal 13 May, 2022 Submission checks completed at journal 13 May, 2022 First submitted to journal 02 Apr, 2022 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-1517751","acceptedTermsAndConditions":true,"allowDirectSubmit":false,"archivedVersions":[],"articleType":"Research Article","associatedPublications":[],"authors":[{"id":105924679,"identity":"075f3c56-05ad-45a4-8f1e-76273321df75","order_by":0,"name":"Ying Jiang","email":"","orcid":"","institution":"Nankai University School of Medicine","correspondingAuthor":false,"submittingAuthor":false,"prefix":"","firstName":"Ying","middleName":"","lastName":"Jiang","suffix":""},{"id":105924680,"identity":"26d27c58-e511-4017-be89-0d4fe9cb020c","order_by":1,"name":"Bo Wang","email":"","orcid":"","institution":"Chinese PLA General Hospital","correspondingAuthor":false,"submittingAuthor":false,"prefix":"","firstName":"Bo","middleName":"","lastName":"Wang","suffix":""},{"id":105924681,"identity":"9122c0f5-81fd-481e-a9d5-e823a9c9977c","order_by":2,"name":"Jun Kang Li","email":"","orcid":"","institution":"Chinese PLA General Hospital","correspondingAuthor":false,"submittingAuthor":false,"prefix":"","firstName":"Jun","middleName":"Kang","lastName":"Li","suffix":""},{"id":105924682,"identity":"4b4e542d-aa9b-439a-9ce9-552153182146","order_by":3,"name":"Shi Yu Li","email":"","orcid":"","institution":"Chinese PLA General Hospital","correspondingAuthor":false,"submittingAuthor":false,"prefix":"","firstName":"Shi","middleName":"Yu","lastName":"Li","suffix":""},{"id":105924683,"identity":"d144f74c-e41c-4570-8ecc-eb8210346220","order_by":4,"name":"Rui Lan Niu","email":"","orcid":"","institution":"Chinese PLA General Hospital","correspondingAuthor":false,"submittingAuthor":false,"prefix":"","firstName":"Rui","middleName":"Lan","lastName":"Niu","suffix":""},{"id":105924684,"identity":"8994bdf2-307e-431d-abea-aa0d0cc17c3e","order_by":5,"name":"Nai Qin Fu","email":"","orcid":"","institution":"Chinese PLA General Hospital","correspondingAuthor":false,"submittingAuthor":false,"prefix":"","firstName":"Nai","middleName":"Qin","lastName":"Fu","suffix":""},{"id":105924685,"identity":"d0a41635-3bee-4b7d-a987-2f970d0cc2d0","order_by":6,"name":"Jiao Jiao Zheng","email":"","orcid":"","institution":"Chinese PLA General Hospital","correspondingAuthor":false,"submittingAuthor":false,"prefix":"","firstName":"Jiao","middleName":"Jiao","lastName":"Zheng","suffix":""},{"id":105924686,"identity":"a9d9e0bd-ea39-4538-b935-7c7d0b1fec30","order_by":7,"name":"Gang Liu","email":"","orcid":"","institution":"Chinese PLA General Hospital","correspondingAuthor":false,"submittingAuthor":false,"prefix":"","firstName":"Gang","middleName":"","lastName":"Liu","suffix":""},{"id":105924687,"identity":"b10c9053-a3a2-4827-9a03-0fae99a4869e","order_by":8,"name":"Zhi Li Wang","email":"data:image/png;base64,iVBORw0KGgoAAAANSUhEUgAAAZAAAAAyAQMAAABI0h/eAAAABlBMVEX///8AAABVwtN+AAAACXBIWXMAAA7EAAAOxAGVKw4bAAAArklEQVRIiWNgGAWjYFAC5sYDDBUMPKRoYWw4wHCGZC2MbaRoMDh/sOHg13l1MubsBxg/fMwhRsuBgw2HZbcd5rHsSWCWnLmNCC1mBxsbDktuO8BjcCCBjZmXKC2HGYFa5tTxGJx/QKyWY4wNBz82MPMY3CDWFvszQFsYjh0GannYTJxfJPsPH3z4o6bO3uB88sEPH4nRAgLMkHhkbCBSPUjtD+LVjoJRMApGwUgEAOzHOw5Nn4HdAAAAAElFTkSuQmCC","orcid":"","institution":"Nankai University School of Medicine","correspondingAuthor":true,"submittingAuthor":false,"prefix":"","firstName":"Zhi","middleName":"Li","lastName":"Wang","suffix":""}],"badges":[],"createdAt":"2022-04-03 01:44:08","currentVersionCode":1,"declarations":"","doi":"10.21203/rs.3.rs-1517751/v1","doiUrl":"https://doi.org/10.21203/rs.3.rs-1517751/v1","draftVersion":[],"editorialEvents":[],"editorialNote":"","failedWorkflow":false,"files":[{"id":21522454,"identity":"0416b3d6-848f-4d67-92b8-70296db0658b","added_by":"auto","created_at":"2022-05-16 15:57:54","extension":"png","order_by":1,"title":"Figure 1","display":"","copyAsset":false,"role":"figure","size":57816,"visible":true,"origin":"","legend":"\u003cp\u003ePairwise comparison of elasticity parameters and COL1A1 expression levels in benign, LNM- and LNM+ breast lesions.\u003c/p\u003e\u003cp\u003e****, \u003cem\u003eP\u003c/em\u003e\u0026lt;0.0001; ***, \u003cem\u003eP\u003c/em\u003e\u0026lt;0.001; **, \u003cem\u003eP\u003c/em\u003e\u0026lt;0.05; LNM, lymph node metastasis; COL1A1, Collagen type I alpha 1; Emax, maximum elasticity; Emean, mean elasticity; Eratio, the elasticity ratio of the lesions to the peripheral tissue.\u003c/p\u003e\u003cp\u003e\u003cbr\u003e\u003c/p\u003e","description":"","filename":"Figure1.png","url":"https://assets-eu.researchsquare.com/files/rs-1517751/v1/e3bc15b9df82913a6b8ead6f.png"},{"id":21523110,"identity":"16919dae-7b7e-4d14-9d36-8e9883a119d5","added_by":"auto","created_at":"2022-05-16 16:02:54","extension":"png","order_by":2,"title":"Figure 2","display":"","copyAsset":false,"role":"figure","size":40771,"visible":true,"origin":"","legend":"\u003cp\u003eThe ROC curve of Emax, Emean, Eratio and the COL1A1 expression level in differential diagnosis of benign and LNM- breast lesions(A). The ROC curve of Emax, Emean, Eratio and the COL1A1 expression level in differential diagnosis of LNM- and LNM+ breast lesions(B).\u003c/p\u003e\u003cp\u003eROC, receiver operating characteristic; COL1A1, Collagen type I alpha 1; Emax, maximum elasticity; Emean, mean elasticity; Eratio, the elasticity ratio of the lesions to the peripheral tissue;\u003c/p\u003e\u003cp\u003eLNM, lymph node metastasis.\u003c/p\u003e\u003cp\u003e\u003cbr\u003e\u003c/p\u003e","description":"","filename":"Figure2.png","url":"https://assets-eu.researchsquare.com/files/rs-1517751/v1/41e5c63330e1e29fdff0835b.png"},{"id":21523281,"identity":"3cb4819b-f9d8-44b5-b78f-e6ae6ba5f2de","added_by":"auto","created_at":"2022-05-16 16:07:54","extension":"png","order_by":3,"title":"Figure 3","display":"","copyAsset":false,"role":"figure","size":431984,"visible":true,"origin":"","legend":"\u003cp\u003eThe comparison of SWE (A, D, G), COL1A1 immunohistochemical staining (B, E, H: ×200) and collagen Sirius staining (C, F, I: ×400) of breast lesions between the benign (A, B, C), LNM– (D, E, F) and LNM+ (G, H, I) groups.\u003c/p\u003e\u003cp\u003eSWE, shear wave elastography; COL1A1, Collagen type I alpha 1; LNM, lymph node metastasis.\u003c/p\u003e\u003cp\u003e\u003cbr\u003e\u003c/p\u003e","description":"","filename":"Figure3.png","url":"https://assets-eu.researchsquare.com/files/rs-1517751/v1/672901cb538ea7126568c686.png"},{"id":21522455,"identity":"161614ba-b5a8-408b-ab45-719071ddec59","added_by":"auto","created_at":"2022-05-16 15:57:54","extension":"png","order_by":4,"title":"Figure 4","display":"","copyAsset":false,"role":"figure","size":28419,"visible":true,"origin":"","legend":"\u003cp\u003eThe expression level of COL1A1 was positively correlated with Emax(A), Emean(B), Eratio(C), (r=0.406, 0.362, 0.425, respectively. P\u0026lt;0.001).\u003c/p\u003e\u003cp\u003eCOL1A1, Collagen type I alpha 1; Emax, maximum elasticity; Emean, mean elasticity; Eratio, the elasticity ratio of the lesions to the peripheral tissue.\u003c/p\u003e","description":"","filename":"Figure4.png","url":"https://assets-eu.researchsquare.com/files/rs-1517751/v1/673c213c3524177001c9a1f3.png"},{"id":21523284,"identity":"02edca07-a031-46eb-9a4d-2c7f2d29df44","added_by":"auto","created_at":"2022-05-16 16:07:57","extension":"pdf","order_by":0,"title":"","display":"","copyAsset":false,"role":"manuscript-pdf","size":907361,"visible":true,"origin":"","legend":"","description":"","filename":"manuscript.pdf","url":"https://assets-eu.researchsquare.com/files/rs-1517751/v1/9673df99-757f-4fcb-8109-95124f338609.pdf"}],"financialInterests":"No competing interests reported.","formattedTitle":"\u003cp\u003eCollagen fiber features and COL1A1: are they associated with elastic parameters in breast lesions and can COL1A1 predict axillary lymph node metastasis?\u003c/p\u003e","fulltext":[{"header":"Introduction","content":"\u003cp\u003eThe incidence of female breast cancer in China is increasing year by year, which seriously affects women's quality of life[\u003cspan citationid=\"CR1\" class=\"CitationRef\"\u003e1\u003c/span\u003e]. The axillary lymph nodes are early metastatic sites of breast cancer, and their proper evaluation preoperatively is key to the formulation of rational treatment regimens. Stiffness is an important characteristic of tissues and organs and is the basis for clinicians to diagnose breast abnormalities. The advent of shear wave elastography (SWE) has provided the possibility of achieving quantitative judgment of tissue stiffness. Recently, several studies have reported that SWE showed good performance in assessing benign and malignant lesions [\u003cspan additionalcitationids=\"CR3\" citationid=\"CR2\" class=\"CitationRef\"\u003e2\u003c/span\u003e\u0026ndash;\u003cspan citationid=\"CR4\" class=\"CitationRef\"\u003e4\u003c/span\u003e]. The stromal reaction that occurs due to invasive behavior results in abnormal tumor-associated collagen deposition, potentially leading to increased elastic modulus value in breast cancer[\u003cspan citationid=\"CR5\" class=\"CitationRef\"\u003e5\u003c/span\u003e]. Studies have shown that the elastic parameters of SWE, could be useful for predicting the nodal status of invasive breast cancers (IBCs), but the pathological basis for the diagnosis of axillary lymph node metastasis is unclear [\u003cspan citationid=\"CR6\" class=\"CitationRef\"\u003e6\u003c/span\u003e].\u003c/p\u003e \u003cp\u003ePrevious studies of SWE and breast tissue stroma focused on the relationship between quantity of collagen or fibrosis and elastic parameters. However, the influence of the shape and arrangement features of collagen fibers on the stiffness of extracellular matrix (ECM) is rarely considered. Malignant lesions are stiffer than benign ones because of the presence of desmoplastic ECM, with collagen type I being a major structural component [\u003cspan citationid=\"CR7\" class=\"CitationRef\"\u003e7\u003c/span\u003e]. The main function of collagen type I is to be the support of organs and tissues and make tissues form tension [\u003cspan citationid=\"CR8\" class=\"CitationRef\"\u003e8\u003c/span\u003e]. At the same time, collagen type I is also closely related to cell growth, proliferation, differentiation and inflammation [\u003cspan citationid=\"CR9\" class=\"CitationRef\"\u003e9\u003c/span\u003e]. Collagen type I consists of a triple helix structure consisting of α1(I) chains (COL1A1) and α2(I) chains (COL1A2).The two polypeptide chains are encoded by different genes and are generally synthesized in a ratio of 2:1[\u003cspan citationid=\"CR10\" class=\"CitationRef\"\u003e10\u003c/span\u003e]. Nevertheless, little is known about the functional role of COL1A1 in the elastography features differences and axillary LNM of breast lesions. The focus of this study was to explore the relationship between the shape and arrangement features of collagen fibers and the elastic parameters of breast lesions evaluated by SWE. To evaluate the role of COL1A1 expression in the differential diagnosis of benign and malignant breast lesions and to analyze whether COL1A1 can predict axillary LNM.\u003c/p\u003e"},{"header":"Materials And Methods","content":"\u003cdiv id=\"Sec3\" class=\"Section2\"\u003e \u003ch2\u003ePatients\u003c/h2\u003e \u003cp\u003eFrom May 2021 to December 2021, 94 lesions in 94 patients were recruited to this prospective study, and were examined with conventional US and SWE prior to surgical resection. The inclusion criteria were newly hospitalized women diagnosed with breast lesions who underwent US and SWE examinations before biopsy. The exclusion criteria included the following: biopsy before US and SWE, undergoing neoadjuvant chemotherapy before US and SWE, and patients with pathologically multifocal IBCs.\u003c/p\u003e \u003cp\u003eThe pathology results obtained by surgery were regarded as gold criteria. The trial was conducted in accordance with the Declaration of Helsinki (as revised in 2013). The study was approved by the Ethics Committee of the Chinese PLA General Hospital (No. S2021-683-01), and informed consent was taken from all individual participants. According to the postoperative pathology, they were divided into benign group and malignant group. The malignant group was divided into LNM \u0026ndash; (negative axillary lymph node metastasis) group and LNM + (positive axillary lymph node metastasis) group.\u003c/p\u003e \u003c/div\u003e \u003cdiv id=\"Sec4\" class=\"Section2\"\u003e \u003ch2\u003ePreoperative US and SWE examination\u003c/h2\u003e \u003cp\u003eUsing AixPlorer US system (SuperSonic Imagine, Aix-en-Provence, France) with a L14-5 linear array probe operating at 4\u0026ndash;15 MHz, SWE was performed after conventional US. The same sonographer with more than 10 years of clinical experience, performed all US and SWE examinations. The probe was applied as lightly as possible to avoid too much pressure and was maintained as steadily as possible for at least 10\u0026ndash;20 seconds during elastic image acquisition. Participants were asked to hold their breath to prevent motion artefacts if needed. To enable a comparison with histopathology data, the maximum imaging section of the lesion was obtained to measure the stiffness of the lesion. Conventional ultrasound was used to obtain the maximum imaging section of the lesion, and then start the SWE check, freeze, and measure. Pay attention to adjusting ROI to include any halo or inelastic abnormal edge lesions, while minimizing any normal tissue. The Eratio was obtained by placing the ROI in normal breast tissue at the same depth as the lesion. The above process was repeated for three times, and the final value of Emax, Emean, Emin and Eratio was the average value of these three repetitions.\u003c/p\u003e \u003c/div\u003e \u003cdiv id=\"Sec5\" class=\"Section2\"\u003e \u003ch2\u003eCollagen staining and categorization\u003c/h2\u003e \u003cp\u003eSirius red staining was used to assess collagen fibers shape and arrangement features and immunohistochemical staining was used to examine the expression of COL1A1.\u003c/p\u003e \u003cp\u003eAn OLYMPUS BX53P microscope equipped with a polarisedfilter. Observe the collagen fibers at 400x magnification. In this study, collagen fibers shape and arrangement feature in breast lesions were classified into three categories. The definitions are described according to the following descriptions: category 0: wavy collagen fibers similar to collagen fibers in normal breast tissue; category 1: taut parallel collagen fibers around tumor nests; category 2: straightened and aligned collagen fibers tending to be perpendicular to the tumor boundary[\u003cspan additionalcitationids=\"CR12 CR13\" citationid=\"CR11\" class=\"CitationRef\"\u003e11\u003c/span\u003e\u0026ndash;\u003cspan citationid=\"CR14\" class=\"CitationRef\"\u003e14\u003c/span\u003e]. Analysis and classification of ECM collagen fibers in each section by a physician majoring in histopathology for 5 years, unaware of SWE results and surgical pathology results.\u003c/p\u003e \u003cp\u003eCOL1A1 was detected by immunohistochemistry (IHC). The staining results of IHC were analyzed using Image-Pro Plus 6.0 software (Media Cybernetics, Rockville, MD, USA). A total of 5 areas of interest were taken from each slice under 200\u0026times; field of view and photographed. After measuring the integrated optical density (IOD) and area, average optical density (AOD\u0026thinsp;=\u0026thinsp;IOD/Area) was used to evaluate the expression level of COL1A1.\u003c/p\u003e \u003c/div\u003e \u003cdiv id=\"Sec6\" class=\"Section2\"\u003e \u003ch2\u003eStatistical analysis\u003c/h2\u003e \u003cp\u003eAll statistical analyses were performed with SPSS 25.0, standard version (SPSS, Chicago, IL, USA). The measurement data were expressed as mean\u0026thinsp;\u0026plusmn;\u0026thinsp;SD and count data as percentage (%). Independent sample t-test was used to compare the COL1A1 expression level, Emax, Emean, Emin and Eratio of benign and malignant breast lesions. One-way analysis of variance (ANOVA)was used to compare the values of Emax, Emean and Eratio between different collagen fibers categories in 94 breast lesions. The chi-squared test was used to compare the rate between groups. Receiver operating Characteristic (ROC) curves were constructed for SWE values and the COL1A1 expression levels in differentiating benign from malignant breast lesions and predicting the axillary LNM. And the corresponding sensitivity and specificity were recorded (95% confidence interval). The correlation between COL1A1 expression level and elastic parameters in breast lesions were evaluated by Pearson correlation analysis, and the coefficients were defined as follows: \u0026lt;0.40 indicated poor reliability; 0.40\u0026ndash;0.75 indicated good reliability, and \u0026gt;\u0026thinsp;0.75 indicated excellent reliability[\u003cspan citationid=\"CR15\" class=\"CitationRef\"\u003e15\u003c/span\u003e]. Kendall's tau-b correlation coefficient was used to determine the correlation between different collagen fiber features and SWE parameters in 94 cases of breast lesions. A value of \u003cem\u003eP\u003c/em\u003e\u0026thinsp;\u0026lt;\u0026thinsp;0.05 was considered statistically significant.\u003c/p\u003e \u003c/div\u003e"},{"header":"Results","content":"\u003cdiv class=\"Section2\" id=\"Sec8\"\u003e\n \u003ch2\u003eClinicopathological characteristics\u003c/h2\u003e\n \u003cp\u003eA total of 94 breast tumors were surgically removed and histopathologically approved, and included 30 benign lesions and 64 malignant lesions. Benign lesions consisted of fibroadenoma (n\u0026thinsp;=\u0026thinsp;17), sclerosing adenosis(n\u0026thinsp;=\u0026thinsp;5), inflammatory lesions(n\u0026thinsp;=\u0026thinsp;3), intraductal papilloma(n\u0026thinsp;=\u0026thinsp;4) and benign phyllode tumor(n\u0026thinsp;=\u0026thinsp;1). Malignant lesions consisted of invasive cancers(n\u0026thinsp;=\u0026thinsp;55), Intraductal cancers(n\u0026thinsp;=\u0026thinsp;5), and mucinous cancers(n\u0026thinsp;=\u0026thinsp;4). The presence of axillary lymph nodes was determined according to postoperative pathology, and breast malignant tumors were divided into LNM\u0026thinsp;+\u0026thinsp;and LNM - groups. Of the 64 malignant tumors, 25(39%) were positive axillary lymphatic metastases, and 39(61%) were clinically negative. There were no statistically significant differences in age and tumor size between groups (Table \u003cspan class=\"InternalRef\"\u003e1\u003c/span\u003e).\u003c/p\u003e\n\u003c/div\u003e\n\u003cdiv class=\"Section2\" id=\"Sec9\"\u003e\n \u003ch2\u003eElastic parameters, collagen fiber features and COL1A1 expression\u003c/h2\u003e\n \u003cp\u003eUltrasonic elastography, Sirius red staining and COL1A1 immunohistochemical staining of included 94 samples are shown in Fig. \u003cspan class=\"InternalRef\"\u003e3\u003c/span\u003e. The Emax, Emean and Eratio of malignant breast lesions were significantly higher than those of benign lesions (\u003cem\u003eP\u003c/em\u003e\u0026thinsp;\u0026lt;\u0026thinsp;0.001; Table \u003cspan class=\"InternalRef\"\u003e1\u003c/span\u003e), while the Emin of malignant breast lesions and benign lesions was not significantly different (\u003cem\u003eP\u003c/em\u003e\u0026thinsp;=\u0026thinsp;0.135). For the diagnosis of malignant group and benign group, the ROC curves of the Emax, Emean and Eratio elasticity are shown in Fig. \u003cspan class=\"InternalRef\"\u003e2\u003c/span\u003e. The optimal cutoff values, yielding the maximum of sensitivity and specificity, were greater than 58.70 kPa, 52.50 kPa, and 3.05. The areas under the ROC curves were 0.853(0.779\u0026ndash;0.928), 0.823(0.737\u0026ndash;0.910), and 0.882(0.813\u0026ndash;0.951), respectively. The best cutoff values of SWE for diagnosing lymphatic metastasis were as follows: 107.5 kPa for Emax, 85.15 kPa for Emean, and 3.90 for Eratio. Eratio showed the best diagnostic performance according to the area under the curve (AUC), with a 95% confidence interval (CI) of 0.845 (0.754\u0026ndash;0.937). Pairwise comparison of SWE parameters of benign group, LNM- group and LNM\u0026thinsp;+\u0026thinsp;group showed that the difference was statistically significant (Fig. \u003cspan class=\"InternalRef\"\u003e1\u003c/span\u003e).\u003c/p\u003e\n \u003cp\u003eIn the present study, collagen fiber shape and arrangement features in breast lesions were classified into three categories. For all 94 lesions, one-way ANOVA showed that Emax, Emean and Eratio were different between categories 0, 1, and 2 (all \u003cem\u003eP\u003c/em\u003e\u0026thinsp;\u0026lt;\u0026thinsp;0.05; Table \u003cspan class=\"InternalRef\"\u003e2\u003c/span\u003e). This shows that Emax, Emean and Eratio increase when collagen fiber class is increased from 0 to 2. Kendall\u0026apos;s tau-b correlation coefficient calculation shows that the correlation coefficient between Emax and collagen category was 0.318, between Emean and collagen category was 0.261, and between Eratio and collagen category was 0.349 (all \u003cem\u003eP\u003c/em\u003e\u0026thinsp;\u0026le;\u0026thinsp;0.001; Table \u003cspan class=\"InternalRef\"\u003e3\u003c/span\u003e).\u003c/p\u003e\n \u003cp\u003eThe mean expression level of COL1A1 in benign breast lesions was 0.105\u0026thinsp;\u0026plusmn;\u0026thinsp;0.323 and malignant breast lesions mean expression level was 0.159\u0026thinsp;\u0026plusmn;\u0026thinsp;0.047 (\u003cem\u003eP\u003c/em\u003e\u0026thinsp;=\u0026thinsp;0.012). COL1A1 expression level was used in the differential diagnosis of benign and malignant breast lesions. The average optical density\u0026thinsp;\u0026gt;\u0026thinsp;0.145 was the cut-off value, the effective rate was 0.808 (0.723\u0026ndash;0.894), and the sensitivity and specificity were 66% and 90% respectively. When the expression level of COL1A1 was used to diagnose LNM - and LNM\u0026thinsp;+\u0026thinsp;malignant breast lesions, the average optical density\u0026thinsp;\u0026gt;\u0026thinsp;0.150 was taken as the cut-off value, the effectiveness was 0.80 (0.69\u0026ndash;0.91), and the sensitivity and specificity were 96% and 59% respectively (Fig. \u003cspan class=\"InternalRef\"\u003e2\u003c/span\u003e). Pairwise comparison of COL1A1 expression levels of benign group, LNM- group and LNM\u0026thinsp;+\u0026thinsp;group showed that the difference was statistically significant (Fig. \u003cspan class=\"InternalRef\"\u003e1\u003c/span\u003e). The Pearson correlation analysis showed a positive correlation between Emax, Emean, Eratio of breast lesions and the expression level of COL1A1 (r\u0026thinsp;=\u0026thinsp;0.406, 0.362, 0.425, respectively. \u003cem\u003eP\u003c/em\u003e\u0026thinsp;\u0026lt;\u0026thinsp;0.001; Fig.\u0026nbsp;\u003cspan class=\"InternalRef\"\u003e4\u003c/span\u003e).\u003c/p\u003e\n \u003cdiv class=\"gridtable\"\u003e\n \u003cdiv align=\"char\" class=\"colspec\"\u003e\u003cbr\u003e\u003c/div\u003e\u0026nbsp;\u003cbr\u003e\n \u003ctable border=\"1\" id=\"Tab1\"\u003e\n \u003ccaption language=\"En\"\u003e\n \u003cdiv class=\"CaptionNumber\"\u003eTable 1\u003c/div\u003e\n \u003cdiv class=\"CaptionContent\"\u003e\n \u003cp\u003eBasic information, elastic parameters and COL1A1 expression of enrolled cases.\u003c/p\u003e\n \u003c/div\u003e\n \u003c/caption\u003e\n \u003ccolgroup cols=\"5\"\u003e\u003c/colgroup\u003e\n \u003cthead\u003e\n \u003ctr\u003e\n \u003cth align=\"left\"\u003e\n \u003cp\u003eParameters\u003c/p\u003e\n \u003c/th\u003e\n \u003cth align=\"left\"\u003e\n \u003cp\u003eBenign(n\u0026thinsp;=\u0026thinsp;30)\u003c/p\u003e\n \u003c/th\u003e\n \u003cth align=\"left\"\u003e\n \u003cp\u003eMalignant(n\u0026thinsp;=\u0026thinsp;64)\u003c/p\u003e\n \u003c/th\u003e\n \u003cth align=\"left\"\u003e\n \u003cp\u003et/\u0026chi;2\u003c/p\u003e\n \u003c/th\u003e\n \u003cth align=\"left\"\u003e\n \u003cp\u003e\u003cem\u003eP\u003c/em\u003e value\u003c/p\u003e\n \u003c/th\u003e\n \u003c/tr\u003e\n \u003c/thead\u003e\n \u003ctbody\u003e\n \u003ctr\u003e\n \u003ctd align=\"left\"\u003e\n \u003cp\u003eAge(years)\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd align=\"char\"\u003e\n \u003cp\u003e44.230\u0026thinsp;\u0026plusmn;\u0026thinsp;8.947\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd align=\"char\"\u003e\n \u003cp\u003e47.700\u0026thinsp;\u0026plusmn;\u0026thinsp;9.146\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd align=\"char\"\u003e\n \u003cp\u003e0.018\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd align=\"char\"\u003e\n \u003cp\u003e0.894\u003c/p\u003e\n \u003c/td\u003e\n \u003c/tr\u003e\n \u003ctr\u003e\n \u003ctd align=\"left\"\u003e\n \u003cp\u003eSize(cm)\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd align=\"char\"\u003e\n \u003cp\u003e2.003\u0026thinsp;\u0026plusmn;\u0026thinsp;0.615\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd align=\"char\"\u003e\n \u003cp\u003e2.233\u0026thinsp;\u0026plusmn;\u0026thinsp;0.679\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd align=\"char\"\u003e\n \u003cp\u003e0.847\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd align=\"char\"\u003e\n \u003cp\u003e0.360\u003c/p\u003e\n \u003c/td\u003e\n \u003c/tr\u003e\n \u003ctr\u003e\n \u003ctd align=\"left\"\u003e\n \u003cp\u003eElastic parameters\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd align=\"left\"\u003e\u0026nbsp;\u003c/td\u003e\n \u003ctd align=\"left\"\u003e\u0026nbsp;\u003c/td\u003e\n \u003ctd align=\"left\"\u003e\u0026nbsp;\u003c/td\u003e\n \u003ctd align=\"left\"\u003e\u0026nbsp;\u003c/td\u003e\n \u003c/tr\u003e\n \u003ctr\u003e\n \u003ctd align=\"left\"\u003e\n \u003cp\u003eEmax(kPa)\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd align=\"char\"\u003e\n \u003cp\u003e44.450\u0026thinsp;\u0026plusmn;\u0026thinsp;20.960\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd align=\"char\"\u003e\n \u003cp\u003e106.709\u0026thinsp;\u0026plusmn;\u0026thinsp;55.490\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd align=\"char\"\u003e\n \u003cp\u003e20.853\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd align=\"char\"\u003e\n \u003cp\u003e\u0026lt;\u0026thinsp;0.001\u003c/p\u003e\n \u003c/td\u003e\n \u003c/tr\u003e\n \u003ctr\u003e\n \u003ctd align=\"left\"\u003e\n \u003cp\u003eEmean(kPa)\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd align=\"char\"\u003e\n \u003cp\u003e33.687\u0026thinsp;\u0026plusmn;\u0026thinsp;18.466\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd align=\"char\"\u003e\n \u003cp\u003e68.155\u0026thinsp;\u0026plusmn;\u0026thinsp;33.830\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd align=\"char\"\u003e\n \u003cp\u003e9.803\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd align=\"char\"\u003e\n \u003cp\u003e0.002\u003c/p\u003e\n \u003c/td\u003e\n \u003c/tr\u003e\n \u003ctr\u003e\n \u003ctd align=\"left\"\u003e\n \u003cp\u003eEmin(kPa)\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd align=\"char\"\u003e\n \u003cp\u003e20.320\u0026thinsp;\u0026plusmn;\u0026thinsp;14.422\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd align=\"char\"\u003e\n \u003cp\u003e26.342\u0026thinsp;\u0026plusmn;\u0026thinsp;18.469\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd align=\"char\"\u003e\n \u003cp\u003e2.272\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd align=\"char\"\u003e\n \u003cp\u003e0.135\u003c/p\u003e\n \u003c/td\u003e\n \u003c/tr\u003e\n \u003ctr\u003e\n \u003ctd align=\"left\"\u003e\n \u003cp\u003eEratio\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd align=\"char\"\u003e\n \u003cp\u003e1.927\u0026thinsp;\u0026plusmn;\u0026thinsp;0.516\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd align=\"char\"\u003e\n \u003cp\u003e4.492\u0026thinsp;\u0026plusmn;\u0026thinsp;2.307\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd align=\"char\"\u003e\n \u003cp\u003e20.540\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd align=\"char\"\u003e\n \u003cp\u003e\u0026lt;\u0026thinsp;0.001\u003c/p\u003e\n \u003c/td\u003e\n \u003c/tr\u003e\n \u003ctr\u003e\n \u003ctd align=\"left\"\u003e\n \u003cp\u003eExpression of COL1A1(AOD)\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd align=\"char\"\u003e\n \u003cp\u003e0.105\u0026thinsp;\u0026plusmn;\u0026thinsp;0.323\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd align=\"char\"\u003e\n \u003cp\u003e0.159\u0026thinsp;\u0026plusmn;\u0026thinsp;0.047\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd align=\"char\"\u003e\n \u003cp\u003e6.564\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd align=\"char\"\u003e\n \u003cp\u003e0.012\u003c/p\u003e\n \u003c/td\u003e\n \u003c/tr\u003e\n \u003c/tbody\u003e\n \u003ctfoot\u003e\n \u003ctr\u003e\n \u003ctd colspan=\"5\"\u003eEmax, maximum elasticity; Emean, mean elasticity; Emin, minimum elasticity; Eratio, the elasticity ratio of the lesions to the peripheral tissue; COL1A1, Collagen type I alpha 1; AOD, average optical density.\u003c/td\u003e\n \u003c/tr\u003e\n \u003c/tfoot\u003e\n \u003c/table\u003e\n \u003c/div\u003e\n \u003cdiv class=\"gridtable\"\u003e\n \u003cdiv align=\"char\" class=\"colspec\"\u003e\u003cbr\u003e\u003c/div\u003e\u0026nbsp;\u003ctable border=\"1\" id=\"Tab2\"\u003e\n \u003ccaption language=\"En\"\u003e\n \u003cdiv class=\"CaptionNumber\"\u003eTable 2\u003c/div\u003e\n \u003cdiv class=\"CaptionContent\"\u003e\n \u003cp\u003eEmax, Emean and Eratio between different collagen fibers categories in 94 breast lesions.\u003c/p\u003e\n \u003c/div\u003e\n \u003c/caption\u003e\n \u003ccolgroup cols=\"5\"\u003e\u003c/colgroup\u003e\n \u003cthead\u003e\n \u003ctr\u003e\n \u003cth align=\"left\" rowspan=\"2\"\u003e\n \u003cp\u003eSWE values\u003c/p\u003e\n \u003c/th\u003e\n \u003cth align=\"left\" colspan=\"3\"\u003e\n \u003cp\u003eCategory of collagen fibers\u003c/p\u003e\n \u003c/th\u003e\n \u003cth align=\"left\" rowspan=\"2\"\u003e\n \u003cp\u003e\u003cem\u003eP\u003c/em\u003e value\u003c/p\u003e\n \u003c/th\u003e\n \u003c/tr\u003e\n \u003ctr\u003e\n \u003cth align=\"left\"\u003e\n \u003cp\u003e0\u003c/p\u003e\n \u003c/th\u003e\n \u003cth align=\"left\"\u003e\n \u003cp\u003e1\u003c/p\u003e\n \u003c/th\u003e\n \u003cth align=\"left\"\u003e\n \u003cp\u003e2\u003c/p\u003e\n \u003c/th\u003e\n \u003c/tr\u003e\n \u003c/thead\u003e\n \u003ctbody\u003e\n \u003ctr\u003e\n \u003ctd align=\"left\"\u003e\n \u003cp\u003eEmax(kPa)\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd align=\"char\"\u003e\n \u003cp\u003e64.670\u0026thinsp;\u0026plusmn;\u0026thinsp;51.160\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd align=\"char\"\u003e\n \u003cp\u003e93.100\u0026thinsp;\u0026plusmn;\u0026thinsp;54.654\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd align=\"char\"\u003e\n \u003cp\u003e112.010\u0026thinsp;\u0026plusmn;\u0026thinsp;51.427\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd align=\"char\"\u003e\n \u003cp\u003e0.002\u003c/p\u003e\n \u003c/td\u003e\n \u003c/tr\u003e\n \u003ctr\u003e\n \u003ctd align=\"left\"\u003e\n \u003cp\u003eEmean(kPa)\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd align=\"char\"\u003e\n \u003cp\u003e47.280\u0026thinsp;\u0026plusmn;\u0026thinsp;32.654\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd align=\"char\"\u003e\n \u003cp\u003e55.550\u0026thinsp;\u0026plusmn;\u0026thinsp;31.357\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd align=\"char\"\u003e\n \u003cp\u003e73.440\u0026thinsp;\u0026plusmn;\u0026thinsp;33.236\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd align=\"char\"\u003e\n \u003cp\u003e0.008\u003c/p\u003e\n \u003c/td\u003e\n \u003c/tr\u003e\n \u003ctr\u003e\n \u003ctd align=\"left\"\u003e\n \u003cp\u003eEratio\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd align=\"char\"\u003e\n \u003cp\u003e2.710\u0026thinsp;\u0026plusmn;\u0026thinsp;1.480\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd align=\"char\"\u003e\n \u003cp\u003e3.450\u0026thinsp;\u0026plusmn;\u0026thinsp;1.617\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd align=\"char\"\u003e\n \u003cp\u003e5.340\u0026thinsp;\u0026plusmn;\u0026thinsp;2.910\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd align=\"char\"\u003e\n \u003cp\u003e\u0026lt;\u0026thinsp;0.001\u003c/p\u003e\n \u003c/td\u003e\n \u003c/tr\u003e\n \u003c/tbody\u003e\n \u003ctfoot\u003e\n \u003ctr\u003e\n \u003ctd colspan=\"5\"\u003eEmax, maximum elasticity; Emean, mean elasticity; Eratio, the elasticity ratio of the lesions to the peripheral tissue.\u003c/td\u003e\n \u003c/tr\u003e\n \u003c/tfoot\u003e\n \u003c/table\u003e\n \u003c/div\u003e\n \u003cdiv class=\"gridtable\"\u003e\n \u003cdiv align=\"char\" class=\"colspec\"\u003e\u003cbr\u003e\u003c/div\u003e\u0026nbsp;\u003ctable border=\"1\" id=\"Tab3\"\u003e\n \u003ccaption language=\"En\"\u003e\n \u003cdiv class=\"CaptionNumber\"\u003eTable 3\u003c/div\u003e\n \u003cdiv class=\"CaptionContent\"\u003e\n \u003cp\u003eCorrelation between different collagen fibers categories and SWE parameters in 94 breast lesions.\u003c/p\u003e\n \u003c/div\u003e\n \u003c/caption\u003e\n \u003ccolgroup cols=\"4\"\u003e\u003c/colgroup\u003e\n \u003cthead\u003e\n \u003ctr\u003e\n \u003cth align=\"left\"\u003e\u0026nbsp;\u003c/th\u003e\n \u003cth align=\"left\"\u003e\n \u003cp\u003eN\u003c/p\u003e\n \u003c/th\u003e\n \u003cth align=\"left\"\u003e\n \u003cp\u003eKendall\u0026rsquo;s tau-b correlation coefficient\u003c/p\u003e\n \u003c/th\u003e\n \u003cth align=\"left\"\u003e\n \u003cp\u003e\u003cem\u003eP\u003c/em\u003e value\u003c/p\u003e\n \u003c/th\u003e\n \u003c/tr\u003e\n \u003c/thead\u003e\n \u003ctbody\u003e\n \u003ctr\u003e\n \u003ctd align=\"left\"\u003e\n \u003cp\u003eEmax vs. collagen fibers categories\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd align=\"char\"\u003e\n \u003cp\u003e94\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd align=\"char\"\u003e\n \u003cp\u003e0.318\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd align=\"char\"\u003e\n \u003cp\u003e\u0026lt;\u0026thinsp;0.001\u003c/p\u003e\n \u003c/td\u003e\n \u003c/tr\u003e\n \u003ctr\u003e\n \u003ctd align=\"left\"\u003e\n \u003cp\u003eEmean vs. collagen fibers categories\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd align=\"char\"\u003e\n \u003cp\u003e94\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd align=\"char\"\u003e\n \u003cp\u003e0.261\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd align=\"char\"\u003e\n \u003cp\u003e0.001\u003c/p\u003e\n \u003c/td\u003e\n \u003c/tr\u003e\n \u003ctr\u003e\n \u003ctd align=\"left\"\u003e\n \u003cp\u003eEratio vs. collagen fibers categories\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd align=\"char\"\u003e\n \u003cp\u003e94\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd align=\"char\"\u003e\n \u003cp\u003e0.349\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd align=\"char\"\u003e\n \u003cp\u003e\u0026lt;\u0026thinsp;0.001\u003c/p\u003e\n \u003c/td\u003e\n \u003c/tr\u003e\n \u003c/tbody\u003e\n \u003ctfoot\u003e\n \u003ctr\u003e\n \u003ctd colspan=\"4\"\u003eEmax, maximum elasticity; Emean, mean elasticity; Eratio, the elasticity ratio of the lesions to the peripheral tissue.\u003c/td\u003e\n \u003c/tr\u003e\n \u003c/tfoot\u003e\n \u003c/table\u003e\n \u003c/div\u003e\n \n\u003c/div\u003e"},{"header":"Discussion","content":"\u003cp\u003eShear wave elastography is a new ultrasound diagnosis technology. It uses shear wave of the tissue induced by acoustic radiation pulse to visualize and quantify the stiffness of tissue in a real-time, reliable, and reproducible manner[\u003cspan citationid=\"CR16\" class=\"CitationRef\"\u003e16\u003c/span\u003e]. Tissue stiffness has become an important parameter in diagnosing potential malignancies or other diseases[\u003cspan citationid=\"CR17\" class=\"CitationRef\"\u003e17\u003c/span\u003e]. For breast lesions, elastic parameters play an important role in the differential diagnosis of benign and malignant breast lesions. Our study showed that compared with benign breast lesions, malignant lesions had higher Emax, Emean, Eratio, which was consistent with the results of several previous studies[\u003cspan additionalcitationids=\"CR19\" citationid=\"CR18\" class=\"CitationRef\"\u003e18\u003c/span\u003e\u0026ndash;\u003cspan citationid=\"CR20\" class=\"CitationRef\"\u003e20\u003c/span\u003e].\u003c/p\u003e \u003cp\u003eConsidering that preoperative knowledge of axillary lymph node status is crucial for the use of neoadjuvant chemotherapy and appropriate surgical treatment, it is necessary to develop imaging tools for LNM diagnosis[\u003cspan citationid=\"CR21\" class=\"CitationRef\"\u003e21\u003c/span\u003e, \u003cspan citationid=\"CR22\" class=\"CitationRef\"\u003e22\u003c/span\u003e]. Our study showed that the best cutoff values of the SWE parameters for diagnosing lymphatic metastasis were calculated as follows: 107.5 kPa for Emax, 85.15 kPa for Emean, and 3.9 for Eratio. According to ROC analysis, Eratio exhibited the best diagnostic performance with an AUC of 0.85 (0.75\u0026ndash;0.94). A study[\u003cspan citationid=\"CR23\" class=\"CitationRef\"\u003e23\u003c/span\u003e] shows that lesion stiffness is a predictor of axillary lymph node metastasis in breast cancer, which combined with Breast Imaging Reporting and Data System (BI-RADS) scores reported AUC for predicting axillary lymph node metastasis status, Emean, Emax and Emin were 0.73 (0.67\u0026ndash;0.80), 0.75 (0.69\u0026ndash;0.81) and 0.69 (0.62\u0026ndash;0.76), respectively. Xin Wen et al. [\u003cspan citationid=\"CR24\" class=\"CitationRef\"\u003e24\u003c/span\u003e] reported that the best cutoff values of the SWE parameters for diagnosing lymphatic metastasis were calculated as follows: 111.05 kPa for Emax, 79.80 kPa for Emean, and 6.89 for EmeanR. According to ROC analysis, Emax exhibited the best diagnostic performance with an AUC of 0.82 (0.76\u0026ndash;0.87). This difference may be caused by the following three factors: fewer cases were included in our study; the setting of the instrument and how to obtain Eratio (there is no relevant information in Xin Wen's report); operator dependent changes.\u003c/p\u003e \u003cp\u003eThe present study demonstrated that the shape and arrangement of ECM collagen fibers are divided into three categories, which positively correlated with Emax, Emean and Eratio of breast lesion stiffness assessed by SWE. According to the results of all 94 lesions, the Emax, Emean and Eratio showed an increasing tendency from category 0 to category 2 (All \u003cem\u003eP\u003c/em\u003e\u0026thinsp;\u0026lt;\u0026thinsp;0.05). In normal breast tissue, collagen fibers showed wavy or sinusoidal waves[\u003cspan citationid=\"CR11\" class=\"CitationRef\"\u003e11\u003c/span\u003e, \u003cspan citationid=\"CR12\" class=\"CitationRef\"\u003e12\u003c/span\u003e]. Thus, this type of collagen fibers was assigned to category 0. Brabrand et al. reported that collagen in the peritumoural region was arranged in a more parallel alignment, Aligned or linearized collagen correlated with increased stiffness of breast tissue or lesions[\u003cspan citationid=\"CR8\" class=\"CitationRef\"\u003e8\u003c/span\u003e, \u003cspan citationid=\"CR25\" class=\"CitationRef\"\u003e25\u003c/span\u003e]. We classify this type as class 1. The type 2 is characterized by bundles of straightened collagen fibers perpendicular to the tumor boundary, which can change in a honeycomb shape and show high hardness[\u003cspan citationid=\"CR14\" class=\"CitationRef\"\u003e14\u003c/span\u003e].These results indicated that breast lesion stiffness will increase when ECM collagen changes into taut, straightened, and arranged parallel fibers. Additionally, a more complicated structure, collagen fibers that was connected in a honeycomb arrangement, indicated the most stiffness. It indicated that ECM collagen fibers shape and arrangement may have also contributed to stiffness variance in malignant breast lesions.\u003c/p\u003e \u003cp\u003eSurvival of breast cancer patients is correlated with stromal biology including the reorganization of the ECM to promote cancer invasion and migration[\u003cspan citationid=\"CR26\" class=\"CitationRef\"\u003e26\u003c/span\u003e]. Nazim Uddin's team suggests that the expression of stromal genes COL1A1 has an association with the progression of breast cancers and recurrence-free survival in breast cancer patients[\u003cspan citationid=\"CR27\" class=\"CitationRef\"\u003e27\u003c/span\u003e]. In our cohort, there were 30 tissue samples in the benign group, 64 tissue samples in the malignant group consecutively included for COL1A1 immunohistochemical staining. The Pearson correlation analysis showed a positive correlation between Emax, Emean, Eratio of breast lesions and the expression level of COL1A1 (r\u0026thinsp;=\u0026thinsp;0.406, 0.362, 0.425, respectively. \u003cem\u003eP\u003c/em\u003e\u0026thinsp;\u0026lt;\u0026thinsp;0.001). The average expression level of COL1A1 in benign breast lesions was significantly lower than that in malignant breast lesions. COL1A1 expression level can be used to diagnose lymph node metastasis. The average optical density\u0026thinsp;\u0026gt;\u0026thinsp;0.150 was taken as the cut-off value, the effectiveness was 0.80 (0.69\u0026ndash;0.91), and the sensitivity and specificity were 96% and 59% respectively. Many studies[\u003cspan citationid=\"CR28\" class=\"CitationRef\"\u003e28\u003c/span\u003e, \u003cspan citationid=\"CR29\" class=\"CitationRef\"\u003e29\u003c/span\u003e] have also proved the correlation between collagen fiber content and SWE in breast lesions, and proved that the change of collagen fiber content leads to the change of tumor tissue hardness. F F Li [\u003cspan citationid=\"CR30\" class=\"CitationRef\"\u003e30\u003c/span\u003e] et al. studied 148 triple-negative breast cancers and found that the high expression of COL1A1 in triple-negative breast cancer was an independent prognostic factor. Protein-protein interaction network analysis confirmed that COL1A1 is a prognostic matrix gene in breast cancer, and its expression is related to the progression of breast cancer[\u003cspan citationid=\"CR27\" class=\"CitationRef\"\u003e27\u003c/span\u003e].These results suggest that COL1A1 may regulate tumor metastasis through some signal pathways; we have begun to investigate the possible signaling mechanisms using in vivo and in vitro methods.\u003c/p\u003e \u003cp\u003eThere was one limitation in this study that should be addressed. Correlations between stiffness and other components of the ECM, such as elastic fibers, laminin, and fibronectinwere not included; however, collagen is the most dominant structural protein in the ECM and its correlation with the elastic modulus could be the most important.\u003c/p\u003e"},{"header":"Conclusion","content":"\u003cp\u003eThe expression level of the collagen fiber features and COL1A1 were positively correlated with the elastic parameters of breast lesions. The expression of COL1A1 might be helpful to diagnose benign and malignant breast lesions and predict axillary lymph node metastasis. \u003c/p\u003e\n"},{"header":"Declarations","content":"\u003cp\u003e\u003cstrong\u003eEthical approval and consent to participants\u003c/strong\u003e\u003c/p\u003e\n\u003cp\u003eThe study was approved by the Ethics Committee of the Chinese PLA General Hospital (No. S2021-683-01), and informed consent was taken from all individual participants.\u003c/p\u003e\n\u003cp\u003e\u003cstrong\u003eConsent for publication\u003c/strong\u003e\u003c/p\u003e\n\u003cp\u003eNot applicable.\u003c/p\u003e\n\u003cp\u003e\u003cstrong\u003eData availability \u0026nbsp;\u003c/strong\u003e\u003c/p\u003e\n\u003cp\u003eAll data generated or analyzed during this study are included in this published article.\u003c/p\u003e\n\u003cp\u003e\u003cstrong\u003eAuthor contributions\u003c/strong\u003e:\u003c/p\u003e\n\u003cp\u003eConception and design: Ying Jiang, Zhi Li Wang; (II)\u0026nbsp;Acquisition of study materials or patients: All authors; (III) Data analysis and interpretation: Ying Jiang, Bo Wang; (IV) Drafting the article: All authors. (V) Final approval of the version to be published: Gang Liu, Zhi Li Wang.\u003c/p\u003e\n\u003cp\u003e\u003cstrong\u003eConflict of \u0026nbsp;interest\u003c/strong\u003e\u003c/p\u003e\n\u003cp\u003eConflict of interest statement: All authors disclosed no relevant relationships.\u003cstrong\u003e\u0026nbsp;\u003c/strong\u003e\u003c/p\u003e\n\u003cp\u003e\u003cstrong\u003eFunding\u003c/strong\u003e\u003c/p\u003e\n\u003cp\u003eThis study was supported by grants from the National Natural Science Foundation of China (No.82071925).\u003c/p\u003e\n\u003cp\u003e\u003cstrong\u003eAcknowledgments\u003c/strong\u003e\u003c/p\u003e\n\u003cp\u003eNot applicable.\u003c/p\u003e"},{"header":"References","content":"\u003col\u003e\n \u003cli\u003eLi, T., C. Mello-Thoms, and P.C. 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Samir, \u003cem\u003eEssentials of Statistical Methods for Assessing Reliability and Agreement in Quantitative Imaging.\u003c/em\u003e Acad Radiol, 2018. \u003cstrong\u003e25\u003c/strong\u003e(3): p. 391-396.\u003c/li\u003e\n \u003cli\u003eBamber, J., et al., \u003cem\u003eEFSUMB guidelines and recommendations on the clinical use of ultrasound elastography. Part 1: Basic principles and technology.\u003c/em\u003e Ultraschall Med, 2013. \u003cstrong\u003e34\u003c/strong\u003e(2): p. 169-84.\u003c/li\u003e\n \u003cli\u003eRiching, K.M., et al., \u003cem\u003e3D collagen alignment limits protrusions to enhance breast cancer cell persistence.\u003c/em\u003e Biophys J, 2014. \u003cstrong\u003e107\u003c/strong\u003e(11): p. 2546-58.\u003c/li\u003e\n \u003cli\u003eQu, S., et al., \u003cem\u003eOsterix promotes the migration and angiogenesis of breast cancer by upregulation of S100A4 expression.\u003c/em\u003e J Cell Mol Med, 2019. \u003cstrong\u003e23\u003c/strong\u003e(2): p. 1116-1127.\u003c/li\u003e\n \u003cli\u003eLink, T., et al., \u003cem\u003eClinical relevance of circulating MACC1 and S100A4 transcripts for ovarian cancer.\u003c/em\u003e Mol Oncol, 2019. \u003cstrong\u003e13\u003c/strong\u003e(5): p. 1268-1279.\u003c/li\u003e\n \u003cli\u003eJia, W., et al., \u003cem\u003eShear wave elastography and pulsed doppler for breast lesions: Similar diagnostic performance and positively correlated stiffness and blood flow resistance.\u003c/em\u003e Eur J Radiol, 2022. \u003cstrong\u003e147\u003c/strong\u003e: p. 110149.\u003c/li\u003e\n \u003cli\u003eMeng, L., et al., \u003cem\u003eDevelopment of a prediction model based on LASSO regression to evaluate the risk of non-sentinel lymph node metastasis in Chinese breast cancer patients with 1-2 positive sentinel lymph nodes.\u003c/em\u003e Sci Rep, 2021. \u003cstrong\u003e11\u003c/strong\u003e(1): p. 19972.\u003c/li\u003e\n \u003cli\u003eHe, Z., et al., \u003cem\u003eIdentification of Risk Factors Associated with Axillary Lymph Node Metastasis for Sentinel Lymph Node-Positive Breast Cancer Patients.\u003c/em\u003e J Oncol, 2020. \u003cstrong\u003e2020\u003c/strong\u003e: p. 8884337.\u003c/li\u003e\n \u003cli\u003eGu, J., et al., \u003cem\u003ePrediction of Invasive Breast Cancer Using Mass Characteristic Frequency and Elasticity in Correlation with Prognostic Histologic Features and Immunohistochemical Biomarkers.\u003c/em\u003e Ultrasound Med Biol, 2021. \u003cstrong\u003e47\u003c/strong\u003e(8): p. 2193-2201.\u003c/li\u003e\n \u003cli\u003eWen, X., et al., \u003cem\u003eQuantitative shear wave elastography in primary invasive breast cancers, based on collagen-S100A4 pathology, indicates axillary lymph node metastasis.\u003c/em\u003e Quant Imaging Med Surg, 2020. \u003cstrong\u003e10\u003c/strong\u003e(3): p. 624-633.\u003c/li\u003e\n \u003cli\u003eBrabrand, A., et al., \u003cem\u003eAlterations in collagen fibre patterns in breast cancer. A premise for tumour invasiveness?\u003c/em\u003e Apmis, 2015. \u003cstrong\u003e123\u003c/strong\u003e(1): p. 1-8.\u003c/li\u003e\n \u003cli\u003eConklin, M.W. and P.J. Keely, \u003cem\u003eWhy the stroma matters in breast cancer: insights into breast cancer patient outcomes through the examination of stromal biomarkers.\u003c/em\u003e Cell Adh Migr, 2012. \u003cstrong\u003e6\u003c/strong\u003e(3): p. 249-60.\u003c/li\u003e\n \u003cli\u003eUddin, M.N. and X. Wang, \u003cem\u003eIdentification of key tumor stroma-associated transcriptional signatures correlated with survival prognosis and tumor progression in breast cancer.\u003c/em\u003e Breast Cancer, 2022.\u003c/li\u003e\n \u003cli\u003eLiu, G., et al., \u003cem\u003eShear wave elasticity of breast lesions: would it be correlated with the extracellular matrix components?\u003c/em\u003e Gland Surg, 2019. \u003cstrong\u003e8\u003c/strong\u003e(4): p. 399-406.\u003c/li\u003e\n \u003cli\u003eWang, Z.L., et al., \u003cem\u003eRelationship between elasticity and collagen fiber content in breast disease: a preliminary report.\u003c/em\u003e Ultrasonics, 2015. \u003cstrong\u003e57\u003c/strong\u003e: p. 44-9.\u003c/li\u003e\n \u003cli\u003eLi, F.F., J.J. Fan, and B.L. Ma, \u003cem\u003e[Effect of high expression of type Ⅰ collagen \u0026alpha;1 chain protein on the prognosis of triple negative breast cancer].\u003c/em\u003e Zhonghua Zhong Liu Za Zhi, 2020. \u003cstrong\u003e42\u003c/strong\u003e(2): p. 122-126.\u003c/li\u003e\n\u003c/ol\u003e"}],"fulltextSource":"","fullText":"","funders":[],"hasAdminPriorityOnWorkflow":false,"hasManuscriptDocX":true,"hasOptedInToPreprint":true,"hasPassedJournalQc":"","hasAnyPriority":false,"hideJournal":false,"highlight":"","institution":"","isAcceptedByJournal":true,"isAuthorSuppliedPdf":false,"isDeskRejected":"","isHiddenFromSearch":false,"isInQc":false,"isInWorkflow":false,"isPdf":false,"isPdfUpToDate":true,"isWithdrawnOrRetracted":false,"journal":{"display":true,"email":"[email protected]","identity":"bmc-cancer","isNatureJournal":false,"hasQc":true,"allowDirectSubmit":false,"externalIdentity":"bcan","sideBox":"Learn more about [BMC Cancer](http://bmccancer.biomedcentral.com/)","snPcode":"","submissionUrl":"https://www.editorialmanager.com/bcan/default.aspx","title":"BMC Cancer","twitterHandle":"BMC_series","acdcEnabled":true,"dfaEnabled":false,"editorialSystem":"em","reportingPortfolio":"BMC Series","inReviewEnabled":true,"inReviewRevisionsEnabled":true},"keywords":"SWE, COL1A1, Breast lesions, Lymph node metastasis, Collagen","lastPublishedDoi":"10.21203/rs.3.rs-1517751/v1","lastPublishedDoiUrl":"https://doi.org/10.21203/rs.3.rs-1517751/v1","license":{"name":"CC BY 4.0","url":"https://creativecommons.org/licenses/by/4.0/"},"manuscriptAbstract":"\u003cp\u003e\u003cstrong\u003ePurpose:\u003c/strong\u003e This study aimed to explore whether collagen fiber features and collagen type I alpha 1 (COL1A1) are related to the stiffness of breast lesions and whether COL1A1 can predict axillary lymph node metastasis (LNM).\u003c/p\u003e\u003cp\u003e\u003cstrong\u003eMethods:\u003c/strong\u003e A prospective analysis was performed in our hospital. A total of 94 breast lesions in 94 patients were enrolled between May 2021 to December 2021. Ultrasound (US) and shear wave elastography (SWE) examinations were performed for every lesion before surgery. Pathology results obtained by surgery were regarded as gold criteria. Sirius red staining and Immunohistochemical were used to examine the collagen fibers shape and arrangement features and COL1A1 expression of included tissue samples, and analyze the correlation between SWE parameters and them. To analyze the effectiveness of COL1A1 expression level in predicting axillary LNM. \u003c/p\u003e\u003cp\u003e\u003cstrong\u003eResults:\u003c/strong\u003e The optimal cutoff values for Emax, Emean, and Eratio for diagnosis of benign group and malignant group, were 58.70 kPa, 52.50 kPa, and 3.05. The optimal cutoff values for diagnosing axillary LNM were 107.5 kPa, 85.15 kPa, and 3.90, respectively. In the present study, collagen fiber shape and arrangement features in breast lesions were classified into three categories. For all 94 lesions, one-way ANOVA showed that Emax, Emean and Eratio were different between categories 0, 1, and 2 (all \u003cem\u003eP\u003c/em\u003e\u0026lt; 0.05). The correlation coefficient between Emax and collagen category was 0.318 (\u003cem\u003eP\u003c/em\u003e\u0026lt; 0.001), between Emean and collagen category was 0.261 (\u003cem\u003eP\u003c/em\u003e= 0.001). And between Eratio and collagen category was 0.349 (\u003cem\u003eP\u003c/em\u003e \u0026lt; 0.001). Emax, Emean and Eratio were all positively correlated with COL1A1 expression level (r=0.406, 0.362, 0.425, respectively). COL1A1 expression level \u0026gt;0.145 was taken as the cut-off value, its efficacy in benign and malignant breast lesions was 0.808, with a sensitivity of 66%, and a specificity 90%. COL1A1 expression level \u0026gt;0.150 was taken as the cut-off value, its efficacy in predicting axillary LNM was 0.796, with a sensitivity of 96%, and a specificity 59%. \u003c/p\u003e\u003cp\u003e\u003cstrong\u003eConclusions:\u003c/strong\u003e The expression level of the collagen fiber features and COL1A1 were positively correlated with the elastic parameters of breast lesions. The expression of COL1A1 might be helpful to diagnose benign and malignant breast lesions and predict axillary LNM. \u003c/p\u003e","manuscriptTitle":"Collagen fiber features and COL1A1: are they associated with elastic parameters in breast lesions and can COL1A1 predict axillary lymph node metastasis?","msid":"","msnumber":"","nonDraftVersions":[{"code":1,"date":"2022-05-16 15:57:53","doi":"10.21203/rs.3.rs-1517751/v1","editorialEvents":[{"type":"communityComments","content":0},{"type":"editorInvited","content":"","date":"2022-05-14T02:45:00+00:00","index":"","fulltext":""},{"type":"checksComplete","content":"","date":"2022-05-14T02:42:43+00:00","index":"","fulltext":""},{"type":"submitted","content":"BMC Cancer","date":"2022-04-03T01:41:58+00:00","index":"","fulltext":""}],"status":"published","journal":{"display":true,"email":"[email protected]","identity":"bmc-cancer","isNatureJournal":false,"hasQc":true,"allowDirectSubmit":false,"externalIdentity":"bcan","sideBox":"Learn more about [BMC Cancer](http://bmccancer.biomedcentral.com/)","snPcode":"","submissionUrl":"https://www.editorialmanager.com/bcan/default.aspx","title":"BMC Cancer","twitterHandle":"BMC_series","acdcEnabled":true,"dfaEnabled":false,"editorialSystem":"em","reportingPortfolio":"BMC Series","inReviewEnabled":true,"inReviewRevisionsEnabled":true}}],"origin":"","ownerIdentity":"e7d758e0-341b-4099-a630-dd9ffe1c0041","owner":[],"postedDate":"May 16th, 2022","published":true,"recentEditorialEvents":[],"rejectedJournal":[],"revision":"","amendment":"","status":"under-review","subjectAreas":[],"tags":[],"updatedAt":"2022-09-14T09:59:25+00:00","versionOfRecord":[],"versionCreatedAt":"2022-05-16 15:57:53","video":"","vorDoi":"","vorDoiUrl":"","workflowStages":[]},"version":"v1","identity":"rs-1517751","journalConfig":"researchsquare"},"__N_SSP":true},"page":"/article/[identity]/[[...version]]","query":{"redirect":"/article/rs-1517751","identity":"rs-1517751","version":["v1"]},"buildId":"cBFmMYwuxLRRLfASyISRj","isFallback":false,"isExperimentalCompile":false,"dynamicIds":[84888],"gssp":true,"scriptLoader":[]}