Imageable AuNP-ECM Hydrogel Tissue Implants for Regenerative Medicine

Figures

• 

  Figure 1. Schematic illustration of the technology. AuNPs are conjugated with Gd and encapsulated within an ECM-based hydrogel. The hydrogel is injected into the left ventricle of mice post ischemia–reperfusion injury, allowing to determine the location of the treatment in relation to scar tissue in the heart.

• 

  Figure 2. Gd-AuNP characterization. (a) TEM image of the pristine AuNPs; scale bar = 200 nm. (b) Size distribution of the AuNPs. (c) Absorbance spectrum of the AuNPs. (d) Scheme of the Gd-AuNPs. (e) FTIR analysis of the different steps in Gd-AuNP synthesis: pristine AuNPs in orange, DTPA complex in green, GdL complex in pink, and Gd-AuNPs in purple. (f) Diameter of the conjugated AuNPs as indicated by DLS. (g) Charge of the conjugated AuNPs as indicated by zeta potential measurements. (h–j) TEM images of (h) pristine AuNPs and (i,j) Gd-AuNPs after negative staining; scale bars = 100, 50, and 25 nm, respectively.

• 

  Figure 3. Gd-AuNP nanocomposite hydrogel characterization. (a) An image of the nanocomposite hydrogel after gelation at 37 °C. (b,c) HRSEM images of the nanocomposite hydrogel. The NPs appear in white. Scale bars (b) = 1 μm, (c) = 300 nm. (d) EDX analysis of the composite hydrogel. (e,f) Rheological properties of the composite hydrogel over time (e) and 15 min post gelation in 37 °C (f).

• 

  Figure 4. Engineered cardiac implants within the nanocomposite hydrogel. (a,b) Viability test within the nanocomposite (a) and pristine hydrogels (b), 1 week post encapsulation; scale bar = 100 μm. (c,d) Cardiac sarcomeric actinin immunostaining of the nanocomposite implants on day 7 (c) and day 14 (d). Actinin in pink, nuclei in blue; scale bar = 25 μm. (e) Contraction rate of the nanocomposite implant before and after the addition of 1 µM noradrenaline.

• 

  Figure 5. Detection of the nanocomposite hydrogel by MRI. (a) MRI imaging of AuNP-Gd, AuNP, and water solutions. (b) MRI imaging of droplets of the nanocomposite hydrogel, pristine hydrogel, and water. (c–f) Images of the heart slice (c,e) and ex vivo MRI imaging (d,f) of the treated (c,d) and untreated (e,f) hearts, 6 weeks post IRI surgery; scale bar = 1 mm.

• 

  Figure 6. MRI imaging of the heart. (a–d) MRI images for (a) untreated, (b) pristine hydrogel-, (c) AuNP composite hydrogel-, and (d) Gd-AuNP composite hydrogel-treated mice, 6 weeks post IRI. The white arrow in (d) indicates the location of the Gd-AuNP composite hydrogel (black area), which was not observed in all other treatments. (e,f) MRI analysis monitoring the location of Gd-AuNP composite hydrogel (dashed area) over time. (e) SAX (short axis slices) and (f) LAX (long axis slices) at (I) 1 week, (II) 4 weeks, and (III) 6 weeks post IRI; scale bar = 1 mm.

• 

  Figure 7. Imaging the coverage area of the treatment in relation to scar area. (a–d) Segmentation process for detection of the Gd-AuNP nanocomposite hydrogel (yellow). (a) LV wall in MRI imaging. (b) K-means algorithm, k = 4. (c) The location of the Gd-AuNPs composite hydrogel as detected by the algorithm. (d) Final MRI image with the location of the hydrogel. (e–h) Subtraction process for detection of the scar tissue (red). (e) LV wall post systemic injection of Gd. (f) LV wall before systemic injection of Gd. (g) Subtraction MRI image. (h) Final MRI image with the location of the scar tissue after noise filtering. (i) Merged image of (d,h). The nanocomposite hydrogel (yellow) with respect to the scar tissue (red), 45 min post systemic injection of Gd-DTPA solution; scale bars = 1 mm.