Label-Free Identification of Early Stages of Breast Ductal Carcinoma via Multiphoton Microscopy

Abstract

Breast cancer can be cured by early diagnosis. Appropriate and effective clinical treatment benefits from accurate pathological diagnosis. However, due to the lack of effective screening and diagnostic imaging methods, early stages of breast cancer often progress to malignant breast cancer. In this study, multiphoton microscopy (MPM) via two-photon excited fluorescence combined with second-harmonic generation was used for identifying the early stages of breast ductal carcinoma. The results showed differences in both cytological features and collagen distribution among normal breast tissue, atypical ductal hyperplasia, low-grade ductal carcinoma in situ, and high-grade ductal carcinoma in situ with microinvasion. Furthermore, three features extracted from the MPM images were used to describe differences in cytological features, collagen density, and basement membrane circumference in the early stages of breast ductal carcinoma. They revealed that MPM has the ability to identify early stages of breast ductal carcinoma label-free, which would contribute to the early diagnosis and treatment of breast cancer. This study may provide the groundwork for the further application of MPM in the clinic.

Multiphoton microscopy for monitoring the occurrence, metastasis and therapy of breast cancer

“…91 Chen et al extracted feature information from MPM images to identify ductal carcinoma in situ. 67 It could be seen from the MPM images that tumor cells had strong TPEF signals. The tumor cells with larger nucleus lacked polarity and were confined to the inside of the duct, not breaking through the basement membrane.…”

“…More importantly, this two‐layer cell structure is vital for breast pathology diagnosis, and is also one of the indicators to distinguish benign and malignant lesions 66 . The stroma out of the lobules is mainly composed of bundles of connective tissue—collagen bundles 65,67 . Due to their non‐centrally symmetric structure and endogenous fluorescent substances, 68–70 they have both SHG signals and TPEF signals.…”

“…It is present in most invasive breast cancers and may be a precursor of invasive cancer, 86–91 accounting for 25% to 30% of all diagnosed breast cancer 91 . Chen et al extracted feature information from MPM images to identify ductal carcinoma in situ 67 . It could be seen from the MPM images that tumor cells had strong TPEF signals.…”

“…66 The stroma out of the lobules is mainly composed of bundles of connective tissuecollagen bundles. 65,67 Due to their non-centrally symmetric structure and endogenous fluorescent substances, [68][69][70] they have both SHG signals and TPEF signals. The mammary ducts are composed of many epithelial cells 70 and are surrounded by collagen fibers that can be seen more clearly in the magnification multiphoton image (Figure 2(E)).…”

Intratumoral heterogeneity of second-harmonic generation scattering from tumor collagen and its effects on metastatic risk prediction

“…In SHG images of excised primary breast tumor, the presence of collagen fibers oriented perpendicular to the tumor border has been shown to be prognostic of breast cancer progression and to enable tumor cell invasion [19,27,28], and in in vivo models of breast cancer, tumor cells can be observed locomoting along SHG+ collagen fibers [29]. Recent studies have demonstrated that SHG can be used, alone and in conjunction with two-photon excited fluorescence, to identify early stages of breast ductal carcinoma, collagen morphological changes during tumor progression, and to provide prognostic information on patient survival [30][31][32].…”