Label‐free imaging diagnosis and collagen‐optical evaluation of endometrioid adenocarcinoma with multiphoton microscopy

The assessment of tumor grade and pathological stage plays a pivotal role in determining the treatment strategy and predicting the prognosis of endometrial cancer. In this study, we employed multiphoton microscopy (MPM) to establish distinctive optical pathological signatures specific to endometrioid adenocarcinoma (EAC), while also assessing the diagnostic sensitivity, specificity, and accuracy of MPM for this particular malignancy. The MPM technique exhibits robust capability in discriminating between benign hyperplasia and various grades of cancer tissue, with statistically significant differences observed in nucleocytoplasmic ratio and second harmonic generation/two‐photon excited fluorescence intensity. Moreover, by utilizing semi‐automated image analysis, we identified notable disparities in six collagen signatures between benign and malignant endometrial stroma. Our study demonstrates that MPM can differentiate between benign endometrial hyperplasia and EAC without labels, while also quantitatively assessing changes in the tumor microenvironment by analyzing collagen signatures in the endometrial stromal tissue. Our study employed multiphoton microscopy to establish the optical pathological characteristics of endometrioid adenocarcinoma, demonstrating its label‐free ability to differentiate between benign endometrial hyperplasia and cancerosis. Furthermore, it enabled quantitative assessment of tumor microenvironment alterations by analyzing collagen signatures in endometrial stromal tissue.