ABSTRACT
BACKGROUND: The traditional pathologic grading for human renal cell carcinoma (RCC) has low concordance between biopsy and surgical specimen. There is a need to investigate adjunctive pathology technique that does not rely on the nuclear morphology that defines the traditional grading. Changes in collagen organization in the extracellular matrix have been linked to prognosis or grade in breast, ovarian, and pancreatic cancers, but collagen organization has never been correlated with RCC grade. In this study, we used Second Harmonic Generation (SHG) based imaging to quantify possible differences in collagen organization between high and low grades of human RCC. METHODS: A tissue microarray (TMA) was constructed from RCC tumor specimens. Each TMA core represents an individual patient. A 5 µm section from the TMA tissue was stained with standard hematoxylin and eosin (H&E). Bright field images of the H&E stained TMA were used to annotate representative RCC regions. In this study, 70 grade 1 cores and 51 grade 4 cores were imaged on a custom-built forward SHG microscope, and images were analyzed using established software tools to automatically extract and quantify collagen fibers for alignment and density assessment. A linear mixed-effects model with random intercepts to account for the within-patient correlation was created to compare grade 1 vs. grade 4 measurements and the statistical tests were two-sided. RESULTS: Both collagen density and alignment differed significantly between RCC grade 1 and RCC grade 4. Specifically, collagen fiber density was greater in grade 4 than in grade 1 RCC (p < 0.001). Collagen fibers were also more aligned in grade 4 compared to grade 1 (p < 0.001). CONCLUSIONS: Collagen density and alignment were shown to be significantly higher in RCC grade 4 vs. grade 1. This technique of biopsy sampling by SHG could complement classical tumor grading approaches. Furthermore it might allow biopsies to be more clinically relevant by informing diagnostics. Future studies are required to investigate the functional role of collagen organization in RCC.
