研究目的
Investigating the intraoperative visualization of the tumor microenvironment and quantification of extracellular vesicles by label-free nonlinear imaging for cancer diagnosis and prognosis.
研究成果
The intraoperative imaging and visualization of the tumor microenvironment, and the quantification of EV density, demonstrate the strengths of the portable nonlinear optical imaging system in cancer research and clinical applications. The results suggest the feasibility and future potential for implementing intraoperative label-free nonlinear optical imaging to investigate the human breast tumor microenvironment and the spatial EV distribution, both to improve our fundamental understanding of carcinogenesis and to potentially provide new biomarkers for tumor invasiveness.
研究不足
The imaged tissue volumes for each breast cancer case are relatively small considering the heterogeneity of breast tissue specimens. A direct relationship between EV density and histologic grade has not been established with controlled tumor-to-margin distances due to the limited number of cases with similar tumor-to-margin distances and different cancer invasiveness grades.
1：Experimental Design and Method Selection:
A custom portable multimodal system for label-free nonlinear optical imaging was designed and built to visualize the unperturbed tumor microenvironment in real time. Four nonlinear optical imaging modalities were integrated into this system: SHG for visualizing collagen fiber reorganization, 2PF for visualizing elastin fibers and FAD-containing cell cytoplasm, THG for highlighting interface structures such as cell membranes, lipid boundaries, and EVs, and 3PF for mapping NADH in the lipids.
2：Sample Selection and Data Sources:
Tissue specimens from 29 human breast cancer cases and 7 cancer-free breast reduction cases were imaged and analyzed.
3：List of Experimental Equipment and Materials:
A portable imaging system integrating four nonlinear optical imaging modalities was used. The laser source provided transform-limited 55-fs laser pulses at a 70-MHz repetition rate.
4：Experimental Procedures and Operational Workflow:
The fresh human breast tissue specimens were imaged within 30 min after surgical excision. The sites imaged on the surgical margin were chosen to be closest to the tumor within the resected specimen.
5：Data Analysis Methods:
An automated segmentation algorithm was developed to extract the EV signal from the background and subsequently quantify EV density.
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