engineering-tendon-assembloids-to-probe-cellular-crosstalk-disease

Engineering Tendon Assembloids to Probe Cellular Crosstalk in Disease and Repair

Tendons enable locomotion by transferring muscle forces to bones. They rely on a tough tendon core comprising collagen fibers and stromal cell ...

Balgrist, University Hospital Zurich, University of Zurich

ETH Zurich

Tendinopathy is a widespread and unresolved clinical challenge, in which associated pain and hampered mobility present a major cause for work-related disability. Tendinopathy associates with a change from a healthy tissue with aligned extracellular matrix (ECM) and highly polarized cells that are connected head-to-tail, towards a diseased tissue with a disorganized ECM and randomly distributed cells, scar-like features that are commonly attributed to poor innate regenerative capacity of the tissue. A fundamental clinical dilemma with this scarring process is whether treatment strategies should focus on healing the affected (disorganized) tissue or strengthen the remaining healthy (anisotropic) tissue. The question was thus asked whether the intrinsic remodeling capacity of tendon-derived cells depends on the organization of the 3D extracellular matrix (isotropic vs anisotropic). Progress in this field is hampered by the lack of suitable in vitro tissue platforms. We aimed at filling this critical gap by creating and exploiting a next generation tissue platform that mimics aspects of the tendon scarring process; cellular response to a gradient in tissue organization from isotropic (scarred/non-aligned) to highly anisotropic (unscarred/aligned) was studied, as was a transient change from isotropic towards highly anisotropic. Strikingly, cells residing in an 'unscarred' anisotropic tissue indicated superior remodeling capacity (increased gene expression levels of collagen, matrix metalloproteinases MMPs, tissue inhibitors of MMPs), when compared to their 'scarred' isotropic counterparts. A numerical model then supported the hypothesis that cellular remodeling capacity may correlate to cellular alignment strength. This in turn may have improved cellular communication, and could thus relate to the more pronounced connexin43 gap junctions observed in anisotropic tissues. In conclusion, increased tissue anisotropy was observed to enhance the cellular potential for functional remodeling of the matrix. This may explain the poor regenerative capacity of tenocytes in chronic tendinopathy, where the pathological process has resulted in ECM disorganization. Additionally, it lends support to treatment strategies that focus on strengthening the remaining healthy tissue, rather than regenerating scarred tissue.

Tissue alignment enhances remodeling potential of tendon-derived cells - Lessons from a novel microtissue model of tendon scarring.

Healthy tendon tissue features a highly aligned extracellular matrix that becomes disorganized with disease. Recent evidence suggests that inflammation coexists with early degenerative changes in tendon, and that crosstalk between immune-cells and tendon fibroblasts (TFs) can contribute to poor tissue healing. We hypothesized that a disorganized tissue architecture may predispose tendon cells to degenerative extracellular matrix remodeling pathways, particularly within a pro-inflammatory niche. This hypothesis was tested by analyzing human TFs cultured on electrospun polycaprolactone (PCL) mats with either highly aligned or randomly oriented fiber structures. We confirmed that fibroblast morphology, phenotype, and markers of matrix turnover could be significantly affected by matrix topography. More strikingly, the TF response to paracrine signals from polarized macrophages or by stimulation with pro-inflammatory cytokines featured significant downregulation of signaling related to extracellular synthesis, with significant concomitant upregulation of gene and protein expression of matrix degrading enzymes. Critically, this tendency towards degenerative re-regulation was exacerbated on randomly oriented PCL substrates. These novel findings indicate that highly aligned tendon cell scaffolds not only promote tendon matrix synthesis, but also play a previously unappreciated role in mitigating adverse resident fibroblast response within an inflammatory milieu.

Substrate fiber alignment mediates tendon cell response to inflammatory signaling.

Although the molecular mechanisms behind tendon disease remain obscure, aberrant stromal matrix turnover and tissue hypervascularity are known hallmarks of advanced tendinopathy. We harness a tendon explant model to unwind complex cross-talk between the stromal and vascular tissue compartments. We identify the hypervascular tendon niche as a state-switch that gates degenerative matrix remodeling within the tissue stroma. Here pathological conditions resembling hypervascular tendon disease provoke rapid cell-mediated tissue breakdown upon mechanical unloading, in contrast to unloaded tendons that remain functionally stable in physiological low-oxygen/-temperature niches. Analyses of the stromal tissue transcriptome and secretome reveal that a stromal niche with elevated tissue oxygenation and temperature drives a ROS mediated cellular stress response that leads to adoption of an immune-modulatory phenotype within the degrading stromal tissue. Degradomic analysis further reveals a surprisingly rich set of active matrix proteases behind the progressive loss of tissue mechanics. We conclude that the tendon stromal compartment responds to aberrant mechanical unloading in a manner that is highly dependent on the vascular niche, with ROS gating a complex proteolytic breakdown of the functional collagen backbone.

Tendon response to matrix unloading is determined by the patho-physiological niche.

(1) To study the reliability of quantifying rear-chain stabilization capacity during deadbug bridging (DBB), (2) to provide reference values for competitive alpine skiers, and (3) to study associations with age, anthropometrics, maturation, skiing performance and back overuse complaints.

Biomechanical quantification of deadbug bridging performance in competitive alpine skiers: Reliability, reference values, and associations with skiing performance and back overuse complaints.

Tendon extracellular matrix (ECM) mechanical unloading results in tissue degradation and breakdown, with niche-dependent cellular stress directing proteolytic degradation of tendon. Here, we show that the extracellular-signal regulated kinase (ERK) pathway is central in tendon degradation of load-deprived tissue explants. We show that ERK 1/2 are highly phosphorylated in mechanically unloaded tendon fascicles in a vascular niche-dependent manner. Pharmacological inhibition of ERK 1/2 abolishes the induction of ECM catabolic gene expression (MMPs) and fully prevents loss of mechanical properties. Moreover, ERK 1/2 inhibition in unloaded tendon fascicles suppresses features of pathological tissue remodeling such as collagen type 3 matrix switch and the induction of the pro-fibrotic cytokine interleukin 11. This work demonstrates ERK signaling as a central checkpoint to trigger tendon matrix degradation and remodeling using load-deprived tissue explants.

Inhibition of ERK 1/2 kinases prevents tendon matrix breakdown.

(1) To evaluate the reliability of quantifying thigh muscle activation patterns and dynamic knee valgus during drop jump landings, (2) to provide reference values for female and male youth alpine skiers, and (3) to study their associations with age, anthropometrics, biological maturation and knee overuse complaints.

Thigh muscle activation patterns and dynamic knee valgus at peak ground reaction force during drop jump landings: Reliability, youth competitive alpine skiing-specific reference values and relation to knee overuse complaints.

Article	Year
Tissue alignment enhances remodeling potential of tendon-derived cells - Lessons from a novel microtissue model of tendon scarring. Matrix biology : journal of the International Society for Matrix Biology\| PubMed ID: 28636876	2018
Substrate fiber alignment mediates tendon cell response to inflammatory signaling. Acta biomaterialia\| PubMed ID: 29530822	2018
Tendon response to matrix unloading is determined by the patho-physiological niche. Matrix biology : journal of the International Society for Matrix Biology\| PubMed ID: 31917255	2020
Biomechanical quantification of deadbug bridging performance in competitive alpine skiers: Reliability, reference values, and associations with skiing performance and back overuse complaints... Physical therapy in sport : official journal of the Association of Chartered Physiotherapists in Sports Medicine\| PubMed ID: 32629419	2020
Inhibition of ERK 1/2 kinases prevents tendon matrix breakdown. Scientific reports\| PubMed ID: 33767224	2021
Thigh muscle activation patterns and dynamic knee valgus at peak ground reaction force during drop jump landings: Reliability, youth competitive alpine skiing-specific reference values and r... Journal of science and medicine in sport\| PubMed ID: 34238661	2021

Jess G. Snedeker

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