Microcaliper

Lamin-shRNA-transduced or HMGB1-shRNA-transduced B16 cells (1 × 10⁵) were injected subcutaneously in 100 μl PBS into the flank of wild-type mice. Where indicated, mice were treated with 50 μg BoxA (HMGBiotech, Milano, Italy), one of the highly conserved DNA-binding domains of HMGB1, which antagonizes HMGB1 binding to its receptor RAGE. In independent sets of experiments, 50 μg anti-IL-10 antibody or control isotype (Biolegend, San Diego, CA) was injected intraperitoneally every 3 days in 100 μl PBS. Mice were monitored every other day and tumour size was measured using a microcaliper (Mitutoyo, Kawasaki, Japan). Tumour volume is expressed as (long diameter × shorter diameter²) × π/6. Lung metastases were generated by tail vein intravenous injection of 1 × 10⁵ B16 cells in 100 μl of PBS. Mice were sacrificed 13 days after tumour cell inoculation and the number of macroscopically visible melanoma metastases on the surface of the lungs was counted in a blinded manner by two different investigators.

The tensile test was performed on TA. XTplus Texture analyser (Stable Micro Systems, Surrey, UK) with a load cell of 10 kg to identify the tensile properties of the printed scaffolds following ASTM D882 standard. Samples of blank and curcumin-loaded PCL scaffolds (7.0% w/w) with different thicknesses (0.1 mm, 0.2 mm, 0.4 mm) and pore shapes (honeycomb, square, and triangle) were tested (n = 5). The scaffolds were cut into rectangular specimens (10 mm × 60 mm) and the thickness of each specimen was measured at 5 points (Microcaliper, Mitutoyo, Japan). The specimen was gripped at each end for 10 mm and stretched at a strain rate of 5 mm/s until breakage. The force and distance were recorded during the extension of the specimen. The stress-strain curve was automatically generated by the Texture analyser software. The tensile strength was calculated as F/(W·D), where F is force at break, w is width of the specimen at the point of break, and D is specimen thickness. The elongation at break (%) was calculated by E/L·100, where E is the distance of rupture and L is the distance onset of separation. The toughness was calculated by the area underneath the stress-strain curve and the tensile modulus was calculated by the inclination of stress-strain curves in the linear elastic region.

All experimental protocols (CBNU2016–085) were approved by the Committee on the Care of Laboratory Animal Resources, Chonbuk National University and were conducted in accordance with the Guide for the Care and Use of Laboratory Animals. Male BALB/c mice (6 weeks old) were obtained from SAMTAKO (Osan, South Korea). They were individually housed in polycarbonate cages and maintained under constant temperature (25–27 °C) with a 12 h light-dark cycle. They were provided free access to standard diet and tap water. These animals were allowed to acclimate to these conditions for at least 7 days before the experiment.
These mice were randomly divided into 4 different groups (4 mice/group) as follows: B: non-treatment, PA: Live P. acnes (1.34 × 10⁹ CFU/ 20 μl PBS) was intradermally injected into the left ear. The right ear was received an equal amount of PBS. PA/ES 1 mg and PA/ES 10 mg with live P. acnes were intradermal injected into both the left and right ears. At 24 h after the injection, ES (1 or 10 mg/ml in PBS) was applied to the surface of the right ear skin of each group. At the end of each treatment period, these animals were sacrificed by cervical dislocation and their ears were measured using a micro-caliper (Mitutoyo, Kanagawa, Japan).

Film thickness was measured with average measurement from five random locations on the films by using a microcaliper (Mitutoyo, Japan). The mean thickness values were used to calculate the mechanical properties of films including tensile strength, percentage elongation and water vapor transmission rate.

The electrospun membranes (P, G, P25, P50, P75)
were crosslinked
for 6 days by soaking in 0.2% (w/v) genipin/ethanol solution. Following
crosslinking, these scaffolds (P, G, P25, P50, and P75) were washed
with adequate ethanol 3 × 10 min to remove residual genipin.
The crosslinked membranes were dried in vacuum for 3 days to eliminate
any leftover solvent. All of the scaffolds were stored in a vacuum
desiccator covered with foil until use to prevent degradation. The
crosslinking mechanism of genipin (Scheme 1A,B) includes two modes of action with different
sites on the genipin.^{29 (link)} The first mechanism
of action involves nucleophilic substitution reaction between the
−NH₂ and −COOCH₃, creating a secondary
amide bond and replacing the ester group. The second action involves
nucleophilic attack of the primary amine on gelatin with the C3 carbon
on genipin, generating an intermediate aldehyde functional group.
A heterocyclic compound is then formed by the interaction of the newly
formed secondary amine with the aldehyde group.
The electrospun PCL/gelatin membranes had thickness
measurement
carried out using a microcaliper (Mitutoyo, Japan). Apparent density
and porosity measurements were calculated as follows^{30 (link)}

We performed an anatomical study on four sides from two fresh, frozen, and thawed adult cadavers (one male, 81 years at death; one female, 73 years at death) in a surgical training facility (Seattle Science Foundation, Seattle, Washington, USA). The dissections were initially performed in the prone position between the 12th rib and highest point of the iliac crest on each side. The latissimus dorsi muscle and the thoracolumbar fascia were dissected. The retroperitoneum was opened and the fat tissue was removed. Next, the cadavers were positioned in the direct lateral position (90°) and K-wires were placed into the intervertebral discs. The placement was confirmed using anteroposterior (Figure 1) and lateral fluoroscopy (Figure 2).
All wires were placed by fellowship-trained spine surgeons. The wires were positioned at L1/L2, L2/L3, L3/L4, and L4/L5 levels. After this, the distances from the wires to the most posterior aspect of the adjacent ascending or descending colon were measured by two different surgeons and the average of the measurements taken (Figure 3, 4).
Measurements were made using microcalipers (Mitutoyo, Kanagawa, Japan) with a resolution of 0.01 mm and an accuracy of ±0.025mm.