Junjian Chen

Antimicrobial Hyaluronic Acid/Poly(amidoamine) Dendrimer Multilayer on Poly(3-hydroxybutyrate-co-4-hydroxybutyrate) Prepared by a Layer-by-Layer Self-Assembly Method

In this article, we prepared hyaluronic acid/poly(amidoamine) dendrimer (HA/PAMAM) multilayers on a poly(3-hydroxybutyrate-co-4-hydroxybutyrate) [P(3HB-4HB)] substrate by a layer-by-layer self-assembly method for antimicrobial biomaterials. The results of ζ potential and quartz crystal microbalance with dissipation (QCM-D) showed that HA/PAMAM multilayers could be formed on the substrate layer by layer. We used QCM-D to show that both the HA outer layer and the PAMAM outer layer exhibited good protein-resistant activity to bovine serum albumin and bacterial antiadhesion activity to Escherichia coli. By a live/dead assay and the colony counting method, we found that the PAMAM outer layer could also exhibit bactericidal activity against E. coli, while the HA outer layer had no bactericidal activity. Both the bacterial antiadhesion activity and the bactericidal activity of the samples could be maintained even after storage in phosphate-buffered saline for up to 14 days. An in vitro MTT assay showed that the multilayers had no cytotoxicity to L929 cells, and HA molecules in the multilayers could improve the biocompatibility of the film.

Aggregation-Induced Emission Active Probe for Light-Up Detection of Anionic Surfactants and Wash-Free Bacterial Imaging.

Anionic surfactants are widely used in daily life and industries, but their residues can cause serious damage to the environment. The current detection methods for anionic surfactants suffer from various limitations and a new detection strategy is highly desirable. Based on 2-(2-hydroxyphenyl)benzothiazole fluorogen with aggregation-induced emission characteristics, we have developed a fluorescent probe HBT-C18 for selective and sensitive detection of anionic surfactants. By in situ formation of catanionic aggregates or micelles with anionic surfactants, the emission intensity of the HBT-C18 probe can increase with increasing keto/enol emission ratio through restriction of intramolecular motion and excited-state intramolecular proton-transfer mechanisms. The probe can also be used for wash-free imaging of bacteria enveloped by a negatively charged outer membrane. The results of this study provide a new strategy for sensitive detection of anionic surfactants and wash-free bacterial imaging.

Long-Term Tracking of the Osteogenic Differentiation of Mouse BMSCs by Aggregation-Induced Emission Nanoparticles

Bone marrow-derived mesenchymal stem cells (BMSCs) have shown great potential for bone repair due to their strong proliferation ability and osteogenic capacity. To evaluate and improve the stem cell-based therapy, long-term tracking of stem cell differentiation into bone-forming osteoblasts is required. However, conventional fluorescent trackers such as fluorescent proteins, quantum dots, and fluorophores with aggregation-caused quenching (ACQ) characteristics have intrinsic limitations of possible interference with stem cell differentiation, heavy metal cytotoxicity, and self-quenching at a high labeling intensity. Herein, we developed aggregation-induced emission nanoparticles decorated with the Tat peptide (AIE-Tat NPs) for long-term tracking of the osteogenic differentiation of mouse BMSCs without interference of cell viability and differentiation ability. Compared with the ability of the commercial Qtracker 655 for tracking of only 6 passages of mouse BMSCs, AIE-Tat NPs have shown a much superior performance in long-term tracking for over 12 passages. Moreover, long-term tracking of the osteogenic differentiation process of mouse BMSCs was successfully conducted on the biocompatible hydroxyapatite scaffold, which is widely used in bone tissue engineering. Thus, AIE-Tat NPs have promising applications in tracking stem cell fate for bone repair.

Preparation of an antimicrobial surface by direct assembly of antimicrobial peptide with its surface binding activity

Antimicrobial peptides (AMPs) are a broad prospect for clinical application against bacterial infections of biomaterials. However, a bottleneck exists as there is a lack of simple technology to prepare AMPs on biomaterials with sufficient activity, as the activity of AMP is dependent on the correct orientation on the biomaterial. In the present study, based on the conventional AMP (Tet213: KRWWKWWRRC) and surface binding peptide (SKHKGGKHKGGKHKG), we designed an Anchor-AMP that could be directly assembled onto the surface of the biomaterial and also showed excellent antimicrobial activity. By characterizing the surface using a quartz crystal microbalance with dissipation (QCM-D), contact angle, atom force microscopy (AFM) and X-ray photoelectron spectroscopy (XPS), we found that Anchor-AMP could adsorb onto the titanium surface with a strong affinity. Different from Tet213 peptide, Anchor-AMP exhibits excellent antimicrobial activity on the titanium surface being able to inhibit 95.33% of Escherichia coli and 96.67% of Staphylococcus aureus after 2.5 h. The improved antimicrobial activity is a result of improved orientation of Anchor-AMP on the biomaterial compared to that of the Tet213 peptide. In addition, the antimicrobial activity of Anchor-AMP was active for more than 24 h. The CCK-8 assay illustrated that the modified titanium surface showed negligible cytotoxicity to bone marrow mesenchymal stem cells. The in vivo results showed that it exhibited excellent antimicrobial activity after 5 and 7 days, inhibiting 89.32% and 99.78% of S. aureus, respectively. We also demonstrated that Anchor-AMP could be applied on a variety of surfaces including gold (Au), polymethyl methacrylate (PMMA) and hydroxyapatite (HA) with strong affinity and good antimicrobial activity.

Local co-delivery and release of antimicrobial peptide and RGD using porous TiO2

We demonstrated for the first time the use of two kinds of porous TiO2 films to co-deliver peptide HHC36 (KRWWKWWRR) and RGD. The film co-delivering these two peptides exhibited excellent antimicrobial activity against S. aureus and E. coli and low cytotoxicity to rat bone mesenchymal stem cells (rBMSCs).

Immobilization of an antimicrobial peptide on silicon surface with stable activity by click chemistry

Infections associated with biomedical implants and devices pose a serious clinical challenge in hospitals worldwide. Antimicrobial peptides (AMPs) have become a great prospect to inhibit this type of infection due to their broad-spectrum antimicrobial activity and low cytotoxicity. However, it is still a challenge to apply AMPs on the biomaterial surface as the activity of AMPs is sensitive to salt or enzyme. In the present study, we prepared a spacer molecule, poly[2-(methacryloyloxy)ethyl]dimethyl-(3-sulfopropyl)ammonium hydroxide (polySBMA), on a model silicon surface via surface-initiated atom transfer radical polymerization (SI-ATRP). We then modified the antimicrobial peptide HHC36 (KRWWKWWRR) with l-propargylglycine (PraAMP) to improve its salt-tolerant activity and integrated PraAMP onto the spacer molecule using click chemistry. We employed X-ray photoelectron spectroscopy (XPS), contact angle goniometry, and atomic force microscopy (AFM) to confirm the success of the immobilization process. We also characterized the antimicrobial activity and stability of the surface with an antimicrobial assay. The results reveal that the modified surface exhibits good antimicrobial activity to inhibit 98.26% of E. coli, 83.72% of S. aureus, and 81.59% of P. aeruginosa. Furthermore, as compared to the control group without the polySBMA spacer, the modified surface improved its resistance to enzymolysis. An in vitro CCK-8 assay also illustrated that this surface showed negligible cytotoxicity to mouse bone mesenchymal stem cells.

Aggregation-Induced Emission Probe for Study of the Bactericidal Mechanism of Antimicrobial Peptides

Multidrug resistant bacterial infection has become one of the most serious threats to human health. Antimicrobial peptides (AMPs) have been identified as potential alternatives to antibiotics owing to their excellent bactericidal activity. However, the complicated bactericidal mechanism of AMPs is still poorly understood. Fluorescence imaging has many advantages in terms of dynamic monitoring, easy operation, and high sensitivity. In this study, we developed an aggregation-induced emission (AIE)-active probe AMP-2HBT by decorating the antimicrobial peptide HHC36 (KRWWKWWRR) with an AIEgen of 2-(2-hydroxyphenyl)benzothiazole (HBT). This AIE-active probe exhibited an excellent light-up fluorescence after binding with bacteria, enabling a real-time monitoring of the binding process. Moreover, a similar time-dependent bactericidal kinetics was observed for the AIE-active probe and HHC36 peptide, which indicated that the bactericidal activity of the peptide was not compromised by decorating with the AIEgen. The bactericidal mechanism of HHC36 peptide was further investigated by super-resolution fluorescence microscopy, transmission electron microscopy (TEM), and scanning electron microscopy (SEM), which suggested that the probe tended to accumulate on the bacterial membrane and efficiently disrupt the membrane structure to kill both Gram-positive and -negative bacteria. This AIE-active probe thus provided a convenient tool to investigate the bactericidal mechanism of AMPs.

Temperature-Controlled Reversible Exposure and Hiding of Antimicrobial Peptides on an Implant for Killing Bacteria at Room Temperature and Improving Biocompatibility in Vivo

Modification of implants by antimicrobial peptides (AMPs) can improve the antimicrobial activity of the implants. However, AMPs have some cytotoxicity in vivo when they are exposed at body temperature. To tackle this challenge, we propose to develop a new approach to generating a smart antimicrobial surface through exposure of AMPs on the surface. A polydopamine film was first formed on the substrates, followed by the conjugation of a temperature-sensitive polymer, poly( N-isopropylacrylamide) (pNIPAM), to the film through atom transfer radical polymerization (ATRP). Then, AMPs were conjugated to the NIPAM on the resultant pNIPAM-modified surface through a click chemistry reaction. Because of the temperature-sensitive property of pNIPAM, the AMPs motif was more exposed to the external environment at room temperature (25 °C) than at body temperature (37 °C), making the surface present a higher antimicrobial activity at room temperature than at body temperature. More importantly, such a smart behavior is accompanied with the increased biocompatibility of the surface at body temperature when compared to the substrates unmodified or modified by AMPs or pNIPAM alone. Our in vivo study further verified that pNIPAM-AMP dual modified bone implants showed increased biocompatibility even when they were challenged with the bacteria at room temperature before implantation. These results indicate that the implants are antibacterial at room temperature and can be safely employed during surgery, resulting in no infection after implantations. Our work represents a new promising strategy to fully explore the antimicrobial property of AMPs, while improving their biocompatibility in vivo. The higher exposure of AMPs at room temperature (the temperature for storing the implants before surgery) will help decrease the risk of bacterial infection, and the lower exposure of AMPs at body temperature (the temperature after the implants are placed into the body by surgery) will improve the biocompatibility of AMPs.