Introduction: The reconstruction of the abdominal wall after abdominal surgery [NN1] results in the loss of tissue biomechanical properties and the occurrence of surgical site infections (SSI). Current available solutions (i.e. implantation of surgical meshes) can restore the abdominal tissue wall biomechanics although inducing significant clinical complications. Bioengineered spider silk proteins offer great potential as drug-free biomaterials . The fusion with human-derived antimicrobial peptides (AMP) represents an innovative approach to render them with antimicrobial activity . Herein, we explore the antimicrobial potential of immobilized bioengineered spider silk proteins with AMP on nanofibrous meshes (NFM) aiming to simultaneously prevent infections and enable to restore the abdominal wall biomechanics.
Methodology: A facile functionalization method comprising the immobilization of bioengineered spider silk protein with antimicrobial peptide (6mer-HNP1), as well as bioengineered spider silk protein alone (6mer), on electrospun polycaprolactone (PCL) NFM was used. The surface of PCL NFM was activated by exposing both sides to ultraviolet ozone for 2 min each. The production of the bioengineered proteins 6mer and 6mer-HNP1 were induced and purified by an immobilized metal affinity chromatography (IMAC). It was determined the protein immobilization capacity over the activated NFM and the functionalized mesh was further characterized in terms of antibacterial activity and cytocompatibility .
Results: The maximum immobilization capacity of the bioengineered proteins 6mer and 6mer-HNP1 were 200 μg mL−1 and 250 μg mL−1, respectively. The immobilization of the proteins on the NFM showed no effect on the formation of beta-sheets by the spider silk domain. Functionalized meshes with 6mer-HNP1 inhibited significantly the adherence and formation of biofilm of Methicillin-Resistant Staphylococcus aureus (MRSA) and Escherichia coli (E. coli), demonstrating their anti-infective potential. The cytocompatibility of the functionalized meshes was validated in vitro in cell studies using a human umbilical vein cell line (EA.hy926) and supported the proliferation of muscle-related cells (Human aortic smooth muscle cells).
Conclusion(s): NFM functionalized with the 6mer-HNP1 showed a significant inhibitory effect against MRSA and E. coli, evidencing their anti-infective properties. The growth and proliferation of human endothelial and muscle cells further evidence their potential for biomedical applications. Overall, this study demonstrates that the use of functionalized meshes with bioengineered spider silk proteins can be a safe and effective alternative to the development of high-performance surgical meshes for challenging abdominal wall repair surgeries.
Acknowledgments: This work was supported by the Portuguese Foundation for Science and Technology (FCT) under the project PTDC/BII-BIO/28870/2017 and PTDC/BTM-MAT/2844/2021; A.R. Franco grant SFRH/BPD/100760/2014.
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