Introduction

PIEZO mechanoreceptors are increasingly recognized to play critical roles in fundamental physiological processes like proprioception, touch, or tendon biomechanics. However, their gating mechanisms and downstream signaling are still not completely understood, mainly due to the lack of effective tools to probe these processes. Here, we developed new tailor-made nanoswitches enabling wireless targeted actuation on PIEZO1 by combining molecular imprinting concepts with magnetic systems.

Extracellular matrix (ECM) mechanical cues guide healing in tendons. Yet, the molecular mechanisms orchestrating the healing processes remain elusive. Appropriate tissue tension is essential for tendon homeostasis and tissue health. By mapping the attainment of tensional homeostasis, we aim to understand how ECM tension regulates healing. We hypothesize that diseased tendon returns to homeostasis only after the cells reach a mechanically gated exit from wound healing.

We engineered a 3D mechano-culture system to create tendon-like constructs by embedding patient-derived tendon cells into a collagen I hydrogel. Casting the hydrogel between posts anchored in silicone allowed adjusting the post stiffness. Under this static mechanical stimulation, cells remodel the (unorganized) collagen representing wound healing mechanisms. We quantified tissue-level forces using post deflection measurements. Secreted ECM was visualized by metabolic labelling with non-canonical amino acids, click chemistry and confocal microscopy. We blocked cell-mediated actin-myosin contractility using a ROCK inhibitor (Y27632) to explore the involvement of the Rho/ROCK pathway in tension regulation.

Tissue tension forces reached the same homeostatic level at day 21 independent of post compliance (p = 0.9456). While minimal matrix was synthesized in early phases of tissue formation (d3-d5), cell-deposited ECM was present in later stages (d7-d9). More ECM was deposited by tendon constructs cultured on compliant (1Nm) compared to rigid posts (p = 0.0017). Matrix synthesized by constructs cultured on compliant posts was less aligned (greater fiber dispersion, p = 0.0021). ROCK inhibition significantly decreased tissue-level tensional forces (p < 0.0001).

Our results indicate that tendon cells balance matrix remodeling and synthesis during tissue repair to reach an intrinsically defined “mechanostat setpoint” guiding tension-mediated exit from wound healing towards homeostasis. We are identifying specific molecular mechanosensors governing tension-regulated healing in tendon and investigate the Rho/ROCK system as their possible downstream pathway.

Tendon tissue equilibrium very heavily depends on appropriate mechanical loading within a narrow, and still poorly defined, physiological range. We will present an overview of our recent work on the tendon cell-matrix interactions that drive tissue homeostasis, matrix remodelling and eventual tissue degeneration, and discuss a roadmap for unravelling these mechanically regulated signalling pathways for the development of effective treatment strategies. Our data suggest that tissue damage accumulates in the tendon until “intrinsic repair mechanisms” are overwhelmed. At this point, the metabolic cost of extracellular matrix remodeling exceeds the locally available nutrient supply. We hypothesize that upon reach S43.1 ing this “Metabolic Tipping Point”, the vascular system is recruited along with accompanying nerve supply (and pain) and the tissue enters into a chronic disease state characterized by high matrix turnover and increasingly poor tissue quality. In this paradigm, a delicate mechanically regulated balance exists between recruitment and suppression of the extrinsic vascular system by the resident tendon core cells. Upon injury or damage, this regulation in turn steers the tissue towards either functional remodeling or chronic tendon disease.

Tear pattern and tendon involvement are risk factors for the development of a pseudoparalytic shoulder. However, some patients have similar tendon involvement but significantly different active forward flexion. In these cases, it remains unclear why some patients suffer from pseudoparalysis and others with the same tear pattern show good active range of motion. Moment arms (MA) and force vectors of the RC and the deltoid muscle play an important role in the muscular equilibrium to stabilize the glenohumeral joint. Biomechanical and clinical analyses were conducted calculating different MA-ratios of the RC and the deltoid muscle using computer rigid body simulation and a retrospective radiographic investigation of two cohorts with and without pseudoparalysis and massive RC tears. Idealized MAs were represented by two spheres concentric to the joints centre of rotation either spanning to the humeral head or deltoid origin of the acromion. Individual ratios of the RC /deltoid MAs on antero-posterior radiographs using the newly introduced Shoulder Abduction Moment (SAM) Index was compared between the pseudoparalytic and non-pseudoparalytic patients.

Decrease of RC activity and improved glenohumeral stability (+14%) was found in simulations for MA ratios with larger diameters of the humeral head which also were consequently beneficial for the (remaining) RC. Clinical investigation of the MA-ratio showed significant risk of having pseudoparalysis in patients with massive tears and a SAM Index <0.77 (OR=11). The SAM index, representing individual biomechanical characteristics of shoulder morphology has an impact on the presence or absence of pseudoparalysis in shoulders with massive RC tears.