It is therefore evident that this gradual escalation in the guidance of MC3T3-E1 cells by the grating patterns is due to the increase in etch depth. when compared to flat surfaces. The study revealed that an increase in etch depth from 150?nm to 4.5?m enhanced cell alignment and elongation along the grating patterns. In the presence of discontinuous elements, cell migration velocity was accelerated when compared to gratings of the same etch depth. These results indicated that cell directionality preference was influenced by a high level of pattern discontinuity. On patterns with bends, cells were more inclined to reverse on 45 bends, with 69% of cells reversing at least once, compared to 54% on 135 bends. These results are attributed to cell morphology and motility mechanisms that are associated with surface topography, where actin filament structures such as BMS-536924 filopodia and lamellipodia are essential in sensing the surrounding environment and controlling cell displacement. Knowledge of geometric guidance cues could provide a better understanding on how cell migration is usually influenced by extracellular matrix topography in vivo. Subject terms: Biomedical engineering, Biotechnology Introduction Cell migration is usually a tightly regulated and essential process for normal development, wound healing, and tissue regeneration, as well as a key driver for BMS-536924 the metastasis of cancer1C4. These biological processes are mediated by the extracellular matrix (ECM), an active component of living tissue that facilitates cell adhesion, cell to cell communication, and cell proliferation, to name a few5,6. Importantly, the ECM is known to influence cell migration track and velocity through its topography and physical properties. During cancer development, cells have the ability to degrade the ECM and migrate away from the primary tumour, thus making cell migration a highly profound area of research7. The guidance of cells through contact with their surroundings was found to be important as cells were observed to BMS-536924 sense surface topographies at the microscale and subsequently, the nanoscale8,9. There is a plethora of evidence demonstrating the guidance of cells in two-dimensional (2D) microenvironments10,11. However, a growing number of studies have successfully exhibited cell guidance within a three-dimensional (3D) microenvironment12,13. Studies using 3D platforms are on the rise as they closely mimic the ECM, therefore producing a more accurate and reliable representation of cell migration in vivo. Additionally, studies have manipulated feature dimensions such as width, etch depth, and spacing, as well as different patterns, as a means to identify the best form of topographical guidance. Other characteristics such as biochemicals and BMS-536924 nano or micro scaled topographies, have also been shown to influence cell guidance14,15. It is long established that cells on flat surfaces have a tendency to move randomly and at a slower velocity compared to patterned topographies16,17. Comparatively, gratings, the most commonly used topographical guiding pattern, have been shown to induce cell alignment in actin rich structures known as lamellipodia and filopodia18. Lamellipodia are large, sheet-like projections associated with cell displacement, whereas HHIP filipodia are spiky cytoplasmic projections which acts as a sensor and explores the microenvironment19. Various cellular structures including integrins are a part of a larger complex known as focal adhesions (FAs) and also play a role in sensing the environment. These structures facilitate the conversation between the cytoskeleton and intracellular components within the ECM through a number of signalling pathways, ultimately resulting in changes in the cytoskeleton and subsequently, cell function20,21. Given the vast range of topographies and features existed in living tissue, continuous topographies and structures may not accurate representations of the ECM as a whole. It is therefore important to investigate guiding patterns other than continuous gratings in order to fully understand cell migration. In this study, engineered platforms comprising of various surface topographies and altered feature characterisations were used to investigate the different guiding effects on MC3T3-E1 osteoblast cell migration. In this systematic study, cells were sensitive.