OB cells metabolism in direct contact with pure Mg, Mg-2Ag, and Mg-10Gd.

<p>a): Cell viability by Live/Dead staining. Control is cells on tissue culture plates. b): SEM images of cell morphology and coverage. Arrows refer to blebs on the cell surface. c): LDH release from cells induced by the degradation. The measurement was performed after 1, 2, 3, 7, and 14 days of immersion. LDH values at 7 and 14 days (not shown) were approximately 0%. Scale bars in white, grey, and black represent 500, 100, and 50 μm; respectively.</p

Saturation level index for different Ca-PO₄ phases according to the immersion medium parameters.

<p>Saturation level index for different Ca-PO₄ phases according to the immersion medium parameters.</p

The chemical composition of the studied materials.

<p>The chemical composition of the studied materials.</p

Degradation parameters after 4 and 14 days of immersion with and without cells.

<p>a): Mean degradation rate [mm/year] according to the weight loss method for pure Mg, Mg-2Ag and Mg-10Gd discs. Significant differences between the Mg-10Gd and pure Mg were detected using a one way analysis of variance (ANOVA) on ranks with Dunn’s multiple comparison post hoc test at a significance level of p < 0.05. Differences between the conditions of Mg-2Ag were analysed with an ANOVA and the Holm-Sidak post hoc test at a significance level of p < 0.001 (*). b) Online pH measurements during 14 days of immersion. i): controls on tissue culture plastic, ii): pure Mg, iii): Mg-2Ag, and iv): Mg-10Gd. Arrows in the Fig refer to medium exchange and the end point of the immersion test. c): Osmolality changes [Osmole/Kg] during immersion with pure Mg, Mg-2Ag, and Mg-10Gd.</p

Biominerliazation detection by osteo-Image.

<p>a): Fluorescent images of the stained hydroxyapatite (HA) on the different materials in monochromatic mode (i.e., mineralization appears white) after 14 days of immersion; without cells, with primary human osteoblasts, and with L929 mouse fibroblast cell line. The stained area is estimated in % of the shown field of view (values represent the mean of triplicate measurements for each condition). Scale bar represent 2 mm b): Fluorescent images at 20x magnification after 4 and 14 days of immersion. Cells stained with DAPI are shown in white/grey, and the mineralized matrix stained with Osteo-Image is shown in green. Scale bars represent 100 μm.</p

Weight % of O, C, P and Mg quantified out of the EDX mapping with 10 keV on Mg, Mg2Ag, Mg10Gd and Mg4Y3Re after 72 h of pre-incubation in culture medium and in cell culture conditions.

<p>Weight % of O, C, P and Mg quantified out of the EDX mapping with 10 keV on Mg, Mg2Ag, Mg10Gd and Mg4Y3Re after 72 h of pre-incubation in culture medium and in cell culture conditions.</p

Confocal fluorescence microscopy of focal adhesion and actin cytoskeleton in HUCPV cells cultured for 24 h on Mg2Ag, Mg10Gd, Mg4Y3RE and Mg on 24 h (a) and 48 h (b) pre-incubated samples.

<p>Focal contacts were stained with vinculin monoclonal antibody (green); actin filaments were stained with a mixture of anti-mouse secondary antibody (FITC) and TRITC-conjugated phalloidin (red); nuclei were stained with DAPI (blue). Images at 20x were merged displaying the triple labeling (yellow areas obtained from the overlapping of red and green labelling).</p

FITC intensity quantification for living HUCPV cultured for 24 h on Mg, Mg2Ag, Mg10Gd and Mg4Y3RE at 24 h, 48 h and 72 h pre-incubation states.

<p>Measurements were obtained from images taken form 3 samples of each material at each pre-incubation condition after LIVE/DEAD assay. Significance level was set at p < 0.05; n = 3 per group.</p

SEM images and the corresponding EDX analysis of magnesium alloys and hp Mg.

<p>Analysis was performed after 72 h pre-incubation time applying 10 KeV accelerating voltage.</p

Confocal fluorescence microscopy of focal adhesion and actin cytoskeleton in HUCPV cells cultured for 24 h on Mg2Ag, Mg10Gd, Mg4Y3RE and Mg on 48 h pre-incubated.

<p>Focal contacts were stained with vinculin monoclonal antibody (green); actin filaments were stained with a mixture of anti-mouse secondary antibody (FITC) and TRITC-conjugated phalloidin (red); nuclei were stained with DAPI (blue). Images at 40x were merged displaying the triple labeling (yellow areas obtained from the overlapping of red and green labelling).</p