Fig. 4

Tissue-matched comparison of OA medial vs. OA lateral tibial plateau. a Safranin-O staining of an OA patient (P1) showed cartilage loss in the medial joint (right) with intact lateral cartilage (left). b Combined MALDI-MSI spectra from medial and lateral sections revealed higher peak intensities in the medial region. c Hierarchical segmentation of the top 500 m/z features identified cartilage (blue) and bone subtypes: healthy trabecular bone (yellow), transitional layer (green), and diseased bone beneath cartilage loss (magenta sclerotic bone). d Pairwise comparison of non-arthritic vs. diseased bone regions identified features with AUROC ≥ 0.85. A representative Col1a1 peptide (G⁶²⁶PAGERGEQ⁶³⁴, m/z 900.40) localized to diseased bone had a predicative AUROC value of 0.872 using only automated segmentation protocols. e In patient P1, 34 discriminatory features were identified between non-arthritic and diseased bone. f Manual segmentation of the same molecular data was completed to demonstrate that segmentation guided by both automated hierarchical segmentation and physiologic insights produced improved AUROC performance. This novel manual segmentation method outlining the diseased subchondral bone specifically and only below the area of cartilage loss (magenta outline) vs. the healthy intact lateral bone (yellow outline) increased AUROC performance from 0.872 (d) to 0.95 for the representative Col1a1 peptide at m/z 900.40