Journal
INTERNATIONAL JOURNAL OF NANOMEDICINE
Volume 17, Issue -, Pages 537-551Publisher
DOVE MEDICAL PRESS LTD
DOI: 10.2147/IJN.S338974
Keywords
nanobody; Aspergillus fumigatus; dectin 1; inflammation; cornea
Funding
- National Natural Science Foundation of China [81870632]
- Natural Science Foundation of Shandong Province [ZR2017BH025]
- Shandong Province Key Research and Development Plan [2019GSF107022]
- Taishan Scholar Project of Shandong Province [tsqn201812151]
- Youth National Natural Science Foundation of China [81500695]
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In this study, a nanobody specific to dectin 1 was successfully prepared and showed high-affinity binding to recombinant dectin 1. The nanobody also exhibited specific binding to dectin 1 expressed on the surfaces of cells and recognized dectin 1 in mouse corneas. Importantly, it demonstrated anti-inflammatory effects on Aspergillus fumigatus keratitis.
Objective: To prepare a nanobody specific to dectin 1 and verify its specificity and anti-inflammatory effects on Aspergillus fumigatus keratitis. Methods: The nanobody was selected from a high-quality shark-antibody library constructed with phage-display technology. The nanobody was developed in the expression systems of Escherichia coli. Indirect ELISA was used to determine the specificity of the nanobody to recombinant dectin 1 protein. The potential of the nanobody to be recognized and expressed on the surfaces of cells and corneas was detected by immunofluorescence, and its anti-inflammatory effect on A. fumigatus keratitis was further verified. After infection with A. fumigatus, eyes of C57B L/6 mice were treated with nanobodies. Human corneal epithelial cells (HCECs) were pretreated with nanobodies and then incubated with A. fumigatus. Clinical scores and slit-lamp photography were used to assess disease response in mouse corneas. RT-PCR and ELISA were used to evaluate mRNA and protein expression of IL1 beta and IL6 in both mouse corneas and HCECs. Results: The nanobody was successfully expressed through microbial system and showed specific high-affinity binding to recombinant dectin 1. Furthermore, it exhibited specific binding to dectin 1 expressed on the surfaces of cells and recognized dectin 1 in mouse corneas. Importantly, it reduced clinical scores of A. fumigatus keratitis in mice compared with a PBS-treatment group. In addition, it decreased mRNA and protein expression of IL1 beta and IL6 in infected corneas and HCECs stimulated with A. fumigatus. Conclusion: These results suggest that this nanobody can bring about anti-inflammatory effects. This highlights the potential of these nanobodies as innovative therapeutic agents in A. fumigatus.
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