A strategy for mechanically integrating robust hydrogel-tissue hybrid to promote the anti-calcification and endothelialization of bioprosthetic heart valve

Figures

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  Figure 1.. (A) Synthesis route of the rhCOLIII. (B) Preparation process of the mechanically robust integrated hydrogel-tissue embedded anticoagulant rhCOLIII and anti-inflammatory TA. (C) Structure of the BHVs modified with PVA/rhCOLIII-TA coating.

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  Figure 2.. (A) SEM Images of GLUT-PP, PVA-TA and PVA/rhCOLIII-TA modified biological valve leaflets. Scale bars = 20 μm. (B) AFM images of GLUT-PP, PVA-TA and PVA/rhCOLIII-TA. (C) ATR-FTIR spectra of different samples. (D) Enlarged view of ATR-FTIR spectra. (E) Wide scan XPS spectra before and after modified BHVs. (F–H) Corresponding high resolution of the C1s spectrum of different samples. (I) UTS, (J) elongation and (K) tangent modulus of different samples. n = 6.

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  Figure 3.. (A) Platelet adhesion and activation on GLUT-PP, PVA-TA and PVA/rhCOLIII-TA-modified heart valve specimens, observed by SEM. (B) The relative adhesion of platelets was measured by LDH kit on the different sample surfaces. (C) Corresponding quantitative analysis of platelet counts on different samples. (D) Illustration of the arteriovenous shunt rabbit model ex vivo. (E) Photographs of thrombus on valve leaflets surfaces. (F) SEM images and (G) weights of the surface thrombus before and after BHVs modification after 1 h of cycling.

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  Figure 4.. (A) The live/dead cells were labeled with calcein and PI after incubation with GLUT-PP, PVA-TA and PVA/rhCOLIII-TA extracts for 3 days. (B) Fluorescent staining of the HUVECs on GLUT-PP, PVA-TA and PVA/rhCOLIII-TA-modified heart valve leaflets after incubation for 3 and 7 days at 37°C, respectively. (C) Cell viability of L929 after 3 days of culturing. Cell viability of HUVECs on the different valve leaflets for (D) 3 and (E) 7 days. (F) Characteristic WB bands of HUVECs-related proteins and (G) quantitative analysis of 1L-1β, vWF, eNOS and CD31.

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  Figure 5.. (A) Fluorescent staining of the RAW 264.7 on GLUT-PP, PVA-TA and PVA/rhCOLIII-TA-modified heart valve leaflets for 3 and 7 days at 37°C, respectively. (B) Cell viability of different samples. (C) Representative flow cytometry plots of M2 phenotypic RAW 264.7 after different sample treatments. (D) Quantification of M2 phenotypic RAW 264.7. (E) TNF-α and (F) IL-10 levels expressed by RAW 264.7 detected by ELISA.

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  Figure 6.. Stability of coatings modified biological valve leaflets after immersion in PBS at designed times. (A) Illustration of the arteriovenous shunt rabbit model ex vivo. (B) Photographs of thrombus on valve leaflets surfaces. (C) SEM images and (D) weights of thrombus formed on GLUT, PVA-TA and PVA/rhCOLIII-TA-modified biological valve leaflets after immersion for 30 days in PBS at 37°C after 1 h of cycling. (E) Relative quantification adhesion of platelets after 3D, 7D, 15D and 30 D of immersion in PBS solution at 37°C, determined by the LDH kit. (F) Fluorescent staining of different samples cultured with HUVEC for 3 days at 37°C after 30 days of immersion in PBS solution at 37°C. (G) Cell viability of different valve leaflets cultured with HUVEC for 3 days after 3D, 7D, 15D, and 30D of immersion in PBS solution at 37°C.

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  Figure 7.. (A) Immunohistochemistry CD68 and CD3 staining of biological valve leaflets before and after modifications after implantation at 15 and 30 days. Quantitative analysis of the counts of (B) CD68-positive and (C) CD3-positive cells in different samples. (D) Alizarin red staining of GLUT-PP, PVA-TA and PVA/rhCOLIII-TA group slices. (E) Calcium contents of different samples after implantation for 90 days.