D7200

A total of 45 petals were selected to measure their length as previously described (Li et al., 2015 (link)). Petals were imaged with a Nikon camera D7200 (Japan) and measured using ImageJ software. To measure the petal cell length and number, the top, middle and basal region of each petal were stained with propidium iodide (0.1 mg mL^–1) for 5 min. Next, images of adaxial epidermal cells were captured using a confocal laser scanning microscope (LSM710, Carl Zeiss, Germany) and more than 50 cells were analyzed using ImageJ software. At least three biological replicates were used for each observation. The elongation rate was calculated according Han et al. (2017) (link).

The photographs showing the plant morphology of Suaeda salsa were taken at 10, 20, 40, 80,100 days after germination (DAG). The leaves and lateral stems of Suaeda salsa from 100 DAG were dissected and photographed using a Nikon D7200 digital camera. The flower buds at different developmental stages were picked up with a tweezer. The floral organs were dissected with 0.1 mm syringes under an anatomical microscope and then placed on agar plates (0.8%) for photographing. The images were taken through a Leica DFC550 microscope, and the measurements were performed using ImageJ software (NIH).

To observe swimming behaviors, 2-day old larvae to 5-day old competent larvae were observed using a dissecting microscope. The light microscopy images and videos were acquired using an inverted compound microscope (OLYMPUS, IX 71) with a mounted camera (Nikon, D7200) examining animals in small petri dishes with coverslip bottom or mounted on slides in filtered ASW, sometimes adding 7% MgCl₂ solution for further immobilization.

Matrigel (Corning, NY, USA) was thawed on ice overnight and diluted in DMEM at a ratio of 2:3. Each well of the 96-well plates was coated with 50 µl of diluted Matrigel and incubated at 37°C in 5% CO₂ for 1 h. HUAECs were cultured in ECM supplemented with 0.2% FBS for 24 h before being used for tube formation. Cells were counted and resuspended to a final concentration of 7.5 × 10⁶ cells/ml in DMEM supplemented with 10% FBS. The resuspended ECs were treated with vehicle, RGMa (2 µg/ml), VEGF (50 ng/ml), or VEGF plus RGMa. The cells were then seeded in 96-well plates coated with Matrigel and further incubated for 3–4 h in an incubator. Images were acquired with a microscope (magnification 40×) connected to a digital camera (D7200, Nikon, Tokyo, Japan). Photographs of 10 representative fields were used for quantification analysis using Image J. Three images were taken randomized from each group for analysis. The tube-formation assays were performed in triplicate using three sets of cells.

Bilayer strips were put into DI water at room temperature until reaching stable configurations. A camera (D7200, Nikon, Tokyo, Japan) was used to take photos of the hydrated hydrogels, and the curvature analysis was done using the digital imaging processing software Fiji and the plugin Kappa. Kappa measured the curvature of each point on the bilayer spline, then the average of the curvature was used to represent the curvature of each bilayer strip. For imaging the morphology and adhesion of the bilayer interface, bilayer strips were pre-frozen in liquid nitrogen for 5 min and then freeze-dried for 24 h. The samples were imaged by scanning electron microscopy (SEM; TESCAN RISE, TESCAN, Brno, Czech Republic) at Michigan Center for Materials Characterization (MC2) at the University of Michigan. To get the thickness input for the computational simulation, the bilayer strips were dried using ethanol with the method mentioned above. Then, the fully dehydrated bilayer strips were imaged with SEM to determine the thickness of PAAm hydrogel and PDMS. The input PDMS thickness was set as 300 $\mathsf{\mu }\mathrm{m}$ , and the PAAm hydrogel thickness was normalized according to the actual PDMS thickness as: 74.5 $\mathsf{\mu }\mathrm{m}$ (5 wt%), 77.9 $\mathsf{\mu }\mathrm{m}$ (7 wt%), 87.7 $\mathsf{\mu }\mathrm{m}$ (10 wt%), 96.4 $\mathsf{\mu }\mathrm{m}$ (15 wt%), and 196.1 $\mathsf{\mu }\mathrm{m}$ (20 wt%).