Simulation of <i>in utero</i> electroporation (IUE).

(A) Opening of the mouse abdominal cavity. (B) Handling of embryos outside of the abdominal cavity. (C) Injection of the DNA-containing solution in the lateral ventricle. (D) Application of voltage through forceps-type platinum electrodes.</p

Questionnaire for the <i>in utero</i> electroporation simulation model.

Questionnaire for the in utero electroporation simulation model.</p

Steps of mold creation from the digital uterus model.

The mold creation for the uterus model followed the same steps as for the embryo model, but used a three-part design to accommodate the anatomical tube shape of a single uterus horn. (A) and (B) Import of the uterus model into Netfabb and Meshmixer to clean up the model. (C) An additional uterus shape was created within the original, with an offset of 0.5 mm. (D)The offset uterus shape was equipped with cylinders at both ends and a cube at the lower end. (E) The original uterus model and the cube were subtracted from a surrounding box. The printed model consists of the two outer parts that outline the original uterus shape and the merged offset uterus model combined with the cylinders and the box to sit inside the two outer parts.</p

Evaluation of the IUE simulator.

(A-E) The user responses to each of our questions to evaluate the IUE simulator. The X-axis indicates percentage points, while the Y-axis indicates the level of user experience. Statistical comparisons were made according to the ’ ’user’s level of expertise (Expert in white and beginner/intermediate in black). Student’s t-test was used for statistical comparison (n = 3). Significance was set as P-value less than 0.05. Bars in the graph represent SD. (F) indicates the statistical comparison between the questions. The X axes indicate percentage points, whereas the Y axis indicates the respective question. The one-way ANOVA followed by Tukey’s HSD (honestly significant difference) test was used for statistical comparison. Significance was set as P-value less than 0.05. Different letters indicate significant differences. Data are depicted in percentage units.</p

Steps of mold creation from the digital embryo model.

(A) Importation of embryo model into Netfabb. (B) Importation of the posterior ventricle into Netfabb. (C) Creation of a box from the part library; it was scaled to cover the embryo model. (D) The plane cut through the box, the embryo model, and the lateral ventricle model. (E) Merging of the upper and lower parts of the lateral ventricle after the plane cut. (F) Subtraction of the head part of the embryo after the plane was cut from the surrounding box with the "Boolean difference" function. (G) Subtraction of the embryo body model after the plane is cut from w the surrounding box with the "Boolean difference" function. (H) Separation of the embryo’s body mold into two halves to create the final silicon embryo.</p

Assembly of the silicone embryo model.

(A) The upper part of the head of the embryo model. The silicone was poured into the mold and hardened overnight. Due to the partial shape of the lateral ventricle on the mold cover on the right, the silicone contains two cavities representing the respective part of the lateral ventricle. (B) The model of the embryo body as seen above. Using a cover, the remaining shape of the lateral ventricle is represented as two cavities in the embryo model. (C) The upper and lower parts of the embryo model are assembled so that the two parts of the lateral ventricle meet each other. (D) The silicone embryo model was assembled with the upper and lower parts glued. The two parts of the lateral ventricle meet each other due to automatic registration in the design process, forming the complete shape of the lateral ventricle.</p

Comparison of the mechanical properties from biosamples and their models.

(A) Statistical comparison between actual embryos and the simulator to evaluate the load [N] per displacement [mm] as a measure of hardness. The two-way ANOVA with Sidak multiple comparison was used to compare the statistics (Embryo’s (E15): n = 19, Embryo model: n = 3). The bars in the graph represent the mean and the SD. For displacement of 1 mm: mean difference = -0.1981, 95% CI -0.3992 to 0.002982. For 1.5 mm displacement: mean difference = -0.3966, 95% CI: -0.5977 to -0.1956, P-valueex vivo uteri as well (uterus(mean = 0.02574) vs. simulator Sh00-20(mean = 0.09674): P-value = 0.0079; uterus versus simulator Sh00-30(mean = 0.1349): P-value<0.0001). Significance was set as P-value less than 0.05. Bars in the graph represent the SD.</p

Formation of minerals onto SaOS-2 cells.

<p>(<b>A</b>) Influence of the tested polymers, polyP·Ca²⁺-complex (polyP) (100 µmoles/L), silica (50 µmoles/L), biosilica (50 µmoles/L), and polyP·Ca²⁺-complex together with biosilica (polyP + biosilica) on the extent of biomineralization, measured by binding of Alizarin Red S to the inorganic deposits. The color reaction was followed spectroscopically at 570 nm; the values are correlated with the DNA content in the respective sample to allow a direct correlation with the cell numbers. The assays were run in the absence (minus) or presence of 5 mg/ml of bioglass (plus bioglass) in the incubation medium for the organic, printed scaffold. The results shown come from eight experiments; the means ± SD are indicated. Significance between the values from the controls (no polymer added) and one series of a given experimental set, in the absence or presence of bioglass, had been calculated and is marked, either with * <i>P</i><0.01 or * <i>P</i><0.001 (Student's <i>t</i>-test). The statistical evaluation by two-way ANOVA test is given under “Results”. (<b>B</b> to <b>D</b>) Element mapping by EDS of the surface of SaOS-2 cells comprising mineral nodules. One nodule is marked (no). Element mapping was performed for O (<u>B</u>), P (<u>C</u>) and C (<u>D</u>). The regions of brighter pseudocolor represent larger accumulations of the respective elements.</p

Morphology of the bioglass preparation used.

<p>(<b>A</b> and <b>B</b>) SEM micrograph. The sintering temperature was 900°C.</p

3D cell bioprinting.

<p>(<b>A</b>) Sketch of the procedure. The alginate/gelatin/SaOS-2 cell suspension is filled into a cartridge, fixed to the printing head (ph). The suspension is pressed through a needle into a culture dish filled with 0.4% CaCl₂ as cross-linking solution. (<b>B</b>) A bioprinted stack of 13 mm in diameter and 1.5 mm in height, placed in a 24-well plate. (<b>C</b>) A bioprinted stack after being incubated in medium/FCS for 3 d. (<b>D</b>) SEM image of mineral nodules (no) on the surface of SaOS-2 cells embedded in alginate/gelatin and 100 µmoles/L polyP·Ca²⁺-complex and incubated for 3 d in the absence and then for 5 d in the presence of the osteogenic cocktail. The specimen was then inspected by SEM.</p