Researchers study the mechanisms and cell signaling pathways that control proliferation and migration of smooth muscle cells (SMCs) after vascular injury and during the development of atherosclerosis [1,2]. These cells line the vasculature and are found in every tissue type throughout the body, including but not limited to lungs, intestines, and brain. Published studies report evidence for SMC proliferation and migration in cancer, injury to the eye, and the development of atherosclerosis [1,2]. It is shown here how wound healing of cultured smooth muscle cells in multiwell plates is investigated with less effort and reliably over time using a Leica inverted microscope and on-stage incubator.
When a wound is created in a confluent monolayer of cells, the regrowth or “healing” of the wound must be monitored over time to measure the kinetics of the recovery . This requires a reliable imaging solution that can: 1) keep the cells alive for a prolonged period of time and 2) maintain focus on the cells the entire time even when they are imaged in multiwell plates.
For this study a “wound” was generated in vitro with a confluent monolayer of adherent SMCs in a multiwell plate. The wound was inflicted via a p200 pipette tip. To investigate wound healing, cell growth was then monitored for over 19 hours with a DMi8 S inverted microscope using a 10x/0.32 NA (numerical aperture) objective and phase contrast. The Quantum Stage and a variety of on-stage incubators were used to keep the cell cultures stable . The reproducibility of the stage positioning allows for multiple positions to be captured without losing the location after repeated passes. To maintain focus, the Adaptative Focus Control (