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. 2018 Apr 23:13:2477-2489.
doi: 10.2147/IJN.S159785. eCollection 2018.

Amyloid-like staining property of RADA16-I nanofibers and its potential application in detecting and imaging the nanomaterial

Yongzhu Chen  1 , Yusi Hua  2 , Wensheng Zhang  3 , Chengkang Tang  4 , Yan Wang  4 , Yujun Zhang  3 , Feng Qiu  3
Affiliations

Amyloid-like staining property of RADA16-I nanofibers and its potential application in detecting and imaging the nanomaterial

Yongzhu Chen et al. Int J Nanomedicine. .

Abstract

Background: Designer self-assembling peptide nanofibers (SAPNFs) as a novel kind of emerging nanomaterial have received more and more attention in the field of nanomedicine in recent years. However, a simple method to monitor and image SAPNFs is still currently absent.

Methods: RADA16-I, a well-studied ionic complementary peptide was used as a model to check potential amyloid-like staining properties of SAPNFs. Thioflavin-T (ThT) and Congo red (CR) as specific dyes for amyloid-like fibrils were used to stain RADA16-I nanofibers in solution, combined with drugs or cells, or injected in vivo as hydrogels. Fluorescent spectrometry and fluorescent microscopy were used to check ThT-binding property, and polarized light microscopy was used to check CR-staining property.

Results: ThT binding with the nanofibers showed enhanced and blue-shifted fluorescence, and specific apple-green birefringence could be observed after the nanofibers were stained with CR. Based on these properties we further showed that ThT-binding fluorescence intensity could be used to monitor the forming and changing of nanofibers in solution, while fluorescent microscopy and polarized light microscopy could be used to image the nanofibers as material for drug delivery, 3D cell culture, and tissue regeneration.

Conclusion: Our results may provide convenient and reliable tools for detecting SAPNFs, which would be helpful for understanding their self-assembling process and exploring their applications.

Keywords: Congo red; amyloid fibrils; nanofibers; self-assembling peptides; thioflavin-T.

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Conflict of interest statement

Disclosure The authors report no conflicts of interest in this work.

Figures

Figure 1
Figure 1
ThT-binding fluorescence of RADA16-I nanofibers. (A) TEM image of RADA16-I nanofibers without ThT. (B) TEM image of RADA16-I nanofibers with ThT. (C) Fluorescence spectrum of ThT-binding RADA16-I. (D) Change of fluorescence intensity at 495 nm at different time points after ThT-binding. Abbreviations: AU, arbitrary units; TEM, transmission electron microscopy; ThT, Thioflavin-T.
Figure 2
Figure 2
Effect of peptide concentration on thioflavin-T-binding fluorescence. (A) Fluorescence spectra at different peptide concentrations. (B) Fluorescence intensity at 495 nm indicated a critical aggregating concentration value (broken red arrow). Abbreviation: AU, arbitrary units.
Figure 3
Figure 3
Thioflavin-T-binding fluorescence reveals the kinetic self-assembly, disassembly, and reassembly of RADA16-I. (A) Self-assembly kinetics of freshly prepared peptide solution. (B) Disassembly–reassembly process after ultrasound treatment. Abbreviation: AU, arbitrary units.
Figure 4
Figure 4
Effect of pH on self-assembly of RADA16-I. (A) Change of thioflavin-T-binding fluorescence of RADA16-I nanofibers incubated at different pH. (B) Circular dichroism spectra of RADA16-I at different pH. (C) Transmission electron microscopy images of nanostructures formed by RADA16-I at different pH. Abbreviation: AU, arbitrary units.
Figure 5
Figure 5
Microscope images of RADA16-I nanofibers based on amyloid-like staining properties. Thioflavin-T-binding nanofibers could be observed by fluorescent microscope (A, B). Congo red-stained nanofibers were sorrel under normal light and (C) apple-green under polarized light (D).
Figure 6
Figure 6
Fluorescent microscope images of RADA16-I nanofibers/hydrogel with pyrene (A) or Dox (B) embedded. Insets are photographs for a suspension with pyrene (A) and a hydrogel with Dox (B). Abbreviations: Dox, doxorubicin hydrochloride; ThT, thioflavin-T.
Figure 7
Figure 7
Fluorescent microscope images of PC12 cells embedded in RADA16 nanofiber scaffold. Arrows indicate cells beginning to stretch out and intrude into the surrounding scaffold. Abbreviations: RhoB, Rhodamine B; ThT, thioflavin-T.
Figure 8
Figure 8
Congo red staining revealed distribution of RADA16-I nanofibers in vivo. Left: under normal light. Right: under polarized light.
See this image and copyright information in PMC

References

    1. Pugliese R, Gelain F. Peptidic biomaterials: from self-assembling to regenerative medicine. Trends Biotechnol. 2017;35(2):145–158. - PubMed
    1. Acar H, Srivastava S, Chung EJ, et al. Self-assembling peptide-based building blocks in medical applications. Adv Drug Deliv Rev. 2017:110–111. 65–79. - PMC - PubMed
    1. Koutsopoulos S. Self-assembling peptide nanofiber hydrogels in tissue engineering and regenerative medicine: progress, design guidelines, and applications. J Biomed Mater Res A. 2016;104(4):1002–1016. - PubMed
    1. Wang TW, Chang KC, Chen LH, Liao SY, Yeh CW, Chuang YJ. Effects of an injectable functionalized self-assembling nanopeptide hydrogel on angiogenesis and neurogenesis for regeneration of the central nervous system. Nanoscale. 2017;9(42):16281–16292. - PubMed
    1. Li R, Pang Z, He H, et al. Drug depot-anchoring hydrogel: a self-assembling scaffold for localized drug release and enhanced stem cell differentiation. J Control Release. 2017;261:234–245. - PubMed

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