Cells in living tissues respond not only to biochemical signals but also to mechanical cues in their microenvironment. Through mechanosensing and mechanotransduction, they perceive stiffness of the extracellular matrix and externally applied forces, factors that can influence gene expression, protein synthesis, cell differentiation and proliferation. To investigate these effects, cells must be embedded in a 3D biocompatible matrix that supports viability and mechanotransduction as well as cell spreading. Hydrogels are currently the material of choice for such applications.

In this work, we present a novel soft microactuator (Figure 1) based on thermoresponsive poly(N isopropylacrylamide) (PNIPAM), fabricated using stop-flow lithography (SFL) and remotely actuated by a laser. The U-shaped design enables reversible and tunable cyclic tensile stimulation of embedded ligament cells at the cellular scale.

We evaluated the biocompatibility of several photocrosslinkable hydrogels such as methacrylated collagen (ColMA), hyaluronic acid (HAMA), dextran hydroxyethyl methacrylate (Dex-HEMA), and human platelet lysate (hPLMA), by encapsulating anterior cruciate ligament (ACL) cells in 3D. Cell viability and morphology were assessed over one week using fluorescence staining and confocal microscopy. Promising formulations were further tested in a microfluidic SFL setup to confirm their feasibility for microscale encapsulation and compatibility with soft actuation.

The developed system offers a controllable environment for studying the role of mechanical cues in ligament cell behaviour in both healthy and diseased tissue models.