5. Conclusions and Future Work
5.2. Future Work
Although we showed some promising results for GO/elastin membranes towards bone regeneration, there are several aspects worth exploring in the future:
• The application of GO-based materials in biomedical fields is very limited due to insufficient knowledge of their long-term stability. We showed that the free-standing GO/elastin membrane was stable in water for 28 days. One possible way to check the membrane stability is to evaluate its mechanical properties after water immersion. We also suggest testing the membrane stability by long-term degradation studies with enzymes.
• Along the same lines, we showed that the TCP-supported GO/elastin membrane was stable in cell culture for up to 19 days without having negative effects on the BMSCs. However, this timeframe is insufficient for potential clinical applications. Long-term cell culture experiments will be necessary to better understand cytocompatibility.
• As mentioned in the method section, we used TCP-supported membranes for cell culture experiments because we were not able to sterilize free-standing membranes without reducing GO and denaturing elastin after autoclaving, UV radiation, and ethanol immersion.
To be able to use the proposed membranes in biomedical applications, it will be necessary to find a way to sterilize them without altering their properties. Alternatives worth exploring can be chemical sterilization with ethylene oxide and gamma radiation. We suggest characterizing the membranes after sterilization to make sure these techniques will not damage the membranes. Another option is to sterilize GO and elastin prior to mixing.
Then prepare the membrane in a sterilized condition, such as drying inside a biosafety cabinet.
• We showed that the 80GO composite membrane can induce mineralization in SBF and osteogenic differentiation of BMSCs. However, these in vitro results cannot predict what would happen in vivo. We suggest evaluating the bone-bonding ability of the composite membrane in a mouse or rat model to understand its bone regeneration potential better. For example, the membrane could be implanted in a mouse or rat calvarial defect as a guided bone regeneration membrane. Alternatively, the membrane could be rolled up to form a
Conclusions and Future Works
65 cylindrical 3D structure, which could be used as an implant in the mouse or rat femoral defect. Also, as we showed for the TCP-supported membranes, it could be possible to apply the GO/elastin composite as a coating on the bone-implant material such as PEEK to increase the bone-bonding ability of PEEK.
• Beyond the application in bone regeneration, we foresee the potential application of this GO/elastin composite membrane in other biomedical applications. An example we are currently exploring is tympanic membrane restoration. The high stiffness indicates the membranes may have good sound conduction. The good biocompatibility with BMSCs suggests that the membranes may also support the adhesion and proliferation of epithelial cells present in the native tympanic membrane to facilitate the restoration process.
66
Supporting Information
Figure S1. Cross-sectional EDS mapping of free-standing membranes
Supporting Information
67 Table S1. Elemental compositions of 100GO, 90GO, 80GO, 70GO, elastin powder, and GO powder from
XPS survey (n = 6)
at% 100GO 90GO 80GO 70GO Elastin GO powder
C1s 67.0±0.8 67.3±1.0 64.5±1.9 66.5±0.8 67.8±0.6 67.4±0.9 O1s 29.8±0.5 27.4±0.2 26.5±1.2 26.3±0.1 17.8±0.3 31.6±1.2 N1s 1.9±0.2 4.1±0.5 5.5±0.4 6.4±0.3 14.4±0.3 N/A S2p* 0.84±0.09 0.67±0.13 0.55±0.03 0.52±0.11 N/A 0.97±0.14
Si2p** 1.1±0.5 1.3±0.3 2.7±0.3 1.1±0.4 N/A N/A
* S derives from traces of H2SO4 used to prepare GO
** Si derives from the silicone mold used during evaporation process
Table S2. List of calcium phosphate compounds and their Ca/P atomic ratios[216]
Name Formula Ca/P ratio
Amorphous calcium phosphate
(ACP) Ca3(PO4)2·3H2O 1.2-2.2
Octacalcium phosphate (OCP) Ca8H2(PO4)6·5H2O 1.33 Calcium-deficient
hydroxyapatite (CDHA)
Ca10-x(HPO4)x(PO4)6-x(OH)2-x
(0<x<1) 1.3-1.67
Hydroxyapatite (HA) Ca10(PO4)6(OH)2 1.67
Carbonated hydroxyapatite (CHA)
Ca10-p(PO4)6-p(OH)2-p(CO3)p
(0<p<1) >1.67
Supporting Information
68 Figure S2. Representative SEM images of (a) 80GO and (b) 100GO membrane surface before SBF
incubation
Figure S3. XRD patterns of 80GO membranes before and after 2× SBF incubation for 10 and 14 days
10 20 30 40
Intensity (a.u.)
2θ (˚)
80GO D10 80GO D14
80GO
Supporting Information
69 Figure S4. EDS mapping of Ca and P for 100GO and 80GO membranes after 10 days and 14 days of
immersion in 2× SBF.
70
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