In Vivo and In Vitro Mechanical Loading of Mouse Achilles Tendons and Tenocytes-A Pilot Study.

Viviane Fleischhacker,Britt Wildemann,Franka Klatte-Schulz,Maximilian Rummler,Bettina M Willie,Aysha Schmock,Anne Seliger,Susann Minkwitz

doi:10.3390/ijms21041313

Abstract

Mechanical force is a key factor for the maintenance, adaptation, and function of tendons. Investigating the impact of mechanical loading in tenocytes and tendons might provide important information on in vivo tendon mechanobiology. Therefore, the study aimed at understanding if an in vitro loading set up of tenocytes leads to similar regulations of cell shape and gene expression, as loading of the Achilles tendon in an in vivo mouse model. In vivo: The left tibiae of mice (n = 12) were subject to axial cyclic compressive loading for 3 weeks, and the Achilles tendons were harvested. The right tibiae served as the internal non-loaded control. In vitro: tenocytes were isolated from mice Achilles tendons and were loaded for 4 h or 5 days (n = 6 per group) based on the in vivo protocol. Histology showed significant differences in the cell shape between in vivo and in vitro loading. On the molecular level, quantitative real-time PCR revealed significant differences in the gene expression of collagen type I and III and of the matrix metalloproteinases (MMP). Tendon-associated markers showed a similar expression profile. This study showed that the gene expression of tendon markers was similar, whereas significant changes in the expression of extracellular matrix (ECM) related genes were detected between in vivo and in vitro loading. This first pilot study is important for understanding to which extent in vitro stimulation set-ups of tenocytes can mimic in vivo characteristics.

Highlights

Tendons play a key role in the musculoskeletal system by transmitting forces from muscle to bone, allowing joint movement
To investigate the impact mechanical loading has on the Achilles tendon in vivo, and on mouse tenocytes in vitro, the loading protocols were chosen to be similar
The Hematoxylin and Eosin staining showed the typical picture of a tendon with elongated tenocytes laying between the parallel-aligned collagen bundles

Summary

Introduction

Tendons play a key role in the musculoskeletal system by transmitting forces from muscle to bone, allowing joint movement. Mechanical forces include tension, hydrostatic pressure, fluid shear stress, and compression, and are transduced by cells to evoke a biochemical response. These responses affect processes such as proliferation, differentiation, tissue development, and skeletal maintenance [1]. Loading within a physiological range (4–8%) induces increased collagen production and has therapeutic effects in ex vivo tendon models [6,7,8]. Compared to ex vivo tendon models, physiological loading led to elevated collagen type I production, decorin expression, and tenogenic differentiation; did not activate inflammatory responses during bi-and uniaxial stretching [11,12,13,14,15]. A few studies exist that directly compare in vitro and in vivo mechanical loading [22,23]

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