Meniscus cell regional phenotypes: Dedifferentiation and reversal by biomaterial embedding

Abstract

Meniscus injuries are common and a major cause of long-term joint degeneration and disability. Current treatment options are limited, so novel regenerative therapies or tissue engineering strategies are urgently needed. The development of new therapies is hindered by a lack of knowledge regarding the cellular biology of the meniscus and a lack of well-established methods for studying meniscus cells in vitro.The goals of this study were to (1) establish baseline expression profiles and dedifferentiation patterns of inner and outer zone primary meniscus cells, and(2) evaluate the utility of poly(ethylene glycol) diacrylate (PEGDA) and gelatin methacrylate (GelMA) polymer hydrogels to reverse dedifferentiation trends for long-term meniscus cell culture. Using reverse transcription-quantitative polymerase chain reaction, we measured expression levels of putative meniscus phenotype marker genes in freshly isolated meniscus tissue, tissue explant culture, and monolayer culture of inner and outer zone meniscus cells from porcine knees to establish baseline dedifferentiation characteristics, and then compared these expression levels to PEGDA/GelMA embedded passaged meniscus cells. COL1A1 showed robust upregulation, while CHAD, CILP, and COMP showed downregulation with monolayer culture. Expression levels of COL2A1, ACAN, and SOX9 were surprisingly similar between inner and outer zone tissue and were found to be less sensitive as markers of dedifferentiation. When embedded in PEGDA/GelMA hydrogels, expression levels of meniscus cell phenotype genes were significantly modulated by varying the ratio of polymer components, allowing these materials to be tuned for phenotype restoration, meniscus cell culture, and tissue engineering applications.

Gene expression profiling of primary fibrochondrocyte cultures in traumatic and degenerative meniscus lesions

“…Genes such as COL1A1, COL2A1, aggrecan (ACAN) and SOX9 are commonly used to determine the phenotype of meniscal cells however, it was recently reported that COL2A1 and ACAN may not be suitable markers for meniscal cells since they have significantly low expression levels in the meniscus. 22,23 Some studies have found a dramatic downregulation of COL2A1 mRNA, 24,25 while some was unable to detect the expression in cell culture, 26 including ours. It is important to note that some conditions such as injury, degeneration, cell culture environment and extracellular matrix component that is used during culturing may change gene expression profile.…”

A single-cell transcriptional atlas reveals resident progenitor cell niche functions in TMJ disc development and injury

“…For example, in FB1 and FB2 clusters, Col5a1 , Col12a1 , Col14a1 and Fndc1 were highly expressed, which are genes associated with fibrillogenesis and fibrillary homeostasis in connective tissues 26 – 29 . In the FB5 and FB6 clusters, Chad , Comp and Cilp appeared to be more active ECM markers, which are genes associated with chondrogenesis and injury repair in fibrocartilage 30 , 31 . Co22a1 is especially active in the FB4 cluster, suggesting potential functions related to myotendinous junctions and vascular stability in this cell population 32 – 34 (Fig.…”

A Tale of Two Loads: Modulation of IL-1 Induced Inflammatory Responses of Meniscal Cells in Two Models of Dynamic Physiologic Loading

“…Cells were isolated from the inner third and outer two-thirds of porcine medial menisci from skeletally mature female pigs obtained from a local abattoir, as previously described ( Riera et al, 2011 ; McNulty et al, 2013 ; Andress et al, 2021 ). Briefly, menisci were separated into inner and outer regions and minced.…”

“…Notably, only a few studies have compared the responses of inner and outer zone cells to mechanical stimulation ( Furumatsu et al, 2012 ; Kanazawa et al, 2012 ). The meniscus inner and outer zones vary in regard to cellular phenotype ( Upton et al, 2006a ; Son and Levenston, 2012 ; Grogan et al, 2017 ; Andress et al, 2021 ), ECM composition, and mechanical properties ( Cheung, 1987 ; Scott et al, 1997 ; Sanchez-Adams et al, 2011 ), and experience different forces in vivo ( Upton et al, 2006b ; Freutel et al, 2014 ). Therefore, the inner and outer zone cells may exhibit differences in mechanoresponsiveness.…”