Histologic and molecular features in pathologic human menisci from knees with and without osteoarthritis

Abstract

The objective of this study was to evaluate histologic and molecular features of meniscus degeneration in cohorts of patients with and without osteoarthritis (OA) of the knee. Menisci were obtained from patients undergoing total knee arthroplasty for OA (TKA) or arthroscopic partial meniscectomy (APM) for a torn knee meniscus. Degenerative meniscal tears were among the most common tear type in the APM group based on the pattern. Using an integrative workflow for molecular evaluation of formalin‐fixed and paraffin‐embedded tissues, human menisci underwent blinded histologic evaluation and NanoString gene expression analyses. Histology revealed increased proteoglycan content in TKA menisci compared to APM menisci, but otherwise no significant differences in the total pathology score or sub‐scores between patients based on age or cohort. NanoString analyses revealed differential expression of genes primarily associated with the PI3K‐AKT signaling pathway, cell cycle, and apoptosis. These data provide new insights into histological and molecular features of meniscus degeneration in patients with and without knee OA. Histologic assessment of menisci showed similar severity of overall degeneration between cohorts, but there were differences at the molecular level. The dysregulated pathways identified in this study could contribute to early‐onset meniscus degeneration, or to a predisposition to meniscus tears and subsequent knee OA. Further studies that validate genes and pathways uncovered in this study will allow us to evaluate novel approaches to assess and treat meniscal degeneration.

Targeted transcriptomic analyses of RNA isolated from formalin‐fixed and paraffin‐embedded human menisci

“…Furthermore, the molecular approach used in this study permits the use of degraded RNA obtained from FFPE sections, as we 13,14 and others 44–50 have shown. However, NanoString analyses are limited because they require the use of panels with preselected probes and therefore do not permit an unbiased evaluation of changes in gene expression.…”

“…A corrected input of 50 ng of RNA from each sample was hybridized with the nCounter® CodeSet using the nCounter® MAX analysis system. Analysis of the expression profiles for more than 700 genes was performed using the nanoString nCounter® PanCancer Pathways Panel and analyzed with the accompanying nSolver TM 4.0 software as previously reported (nanoString Technologies) 13,14 …”

“…Analysis of the expression profiles for more than 700 genes was performed using the nanoString nCounter® PanCancer Pathways Panel and analyzed with the accompanying nSolver TM 4.0 software as previously reported (nanoString Technologies). 13,14 2.4 | Histology Histologic evaluation of menisci was performed in serial adjacent sections to the samples used for RNA isolation, to assess cellularity and the extent of degenerative changes in the microstructure and composition of the extracellular matrix (ECM). Histological assessments were done in parallel with the molecular evaluation of the same sample.…”

“…It was expected that the results of this study would provide further insight into pathophysiological mechanisms in the injured meniscus using a technically simple, point-of-care approach, that we have utilized for other studies at our institution. [13][14][15] 2 | METHODS…”

“…It was hypothesized that (i) degraded RNA could be effectively isolated from injured FFPE human menisci and used for multiplex gene expression analyses using the NanoString nCounter® platform regardless of storage time; and (ii) the resulting targeted transcriptomic data would allow development of hypotheses relating the molecular and histologic data to clinical parameters for testing in subsequent experiments. It was expected that the results of this study would provide further insight into pathophysiological mechanisms in the injured meniscus using a technically simple, point‐of‐care approach, that we have utilized for other studies at our institution 13–15 …”