Hybrid derivative of cathelicidin and human beta defensin-2 against Gram-positive bacteria: A novel approach for the treatment of bacterial keratitis

AbstractBacterial keratitis (BK) is a major cause of corneal blindness globally. This study aimed to develop a novel class of antimicrobial therapy, based on human-derived hybrid host defense peptides (HyHDPs), for treating BK. HyHDPs were rationally designed through combination of functional amino acids in parent HDPs, including LL-37 and human beta-defensin (HBD)-1 to -3. Minimal inhibitory concentrations (MICs) and time-kill kinetics assay were performed to determine the concentration- and time-dependent antimicrobial activity and cytotoxicity was evaluated against human corneal epithelial cells and erythrocytes. In vivo safety and efficacy of the most promising peptide was examined in the corneal wound healing and Staphylococcus aureus (ATCC SA29213) keratitis murine models, respectively. A second-generation HyHDP (CaD23), based on rational hybridization of the middle residues of LL-37 and C-terminal of HBD-2, was developed and was shown to demonstrate good efficacy against methicillin-sensitive and methicillin-resistant S. aureus [MIC = 12.5–25.0 μg/ml (5.2–10.4 μM)] and S. epidermidis [MIC = 12.5 μg/ml (5.2 μM)], and moderate efficacy against P. aeruginosa [MIC = 25-50 μg/ml (10.4–20.8 μM)]. CaD23 (at 25 μg/ml or 2× MIC) killed all the bacteria within 30 min, which was 8 times faster than amikacin (25 μg/ml or 20× MIC). After 10 consecutive passages, S. aureus (ATCC SA29213) did not develop any antimicrobial resistance (AMR) against CaD23 whereas it developed significant AMR (i.e. a 32-fold increase in MIC) against amikacin, a commonly used treatment for BK. Pre-clinical murine studies showed that CaD23 (0.5 mg/ml) achieved a median reduction of S. aureus bioburden by 94% (or 1.2 log10 CFU/ml) while not impeding corneal epithelial wound healing. In conclusion, rational hybridization of human-derived HDPs has led to generation of a potentially efficacious and safe topical antimicrobial agent for treating Gram-positive BK, with no/minimal risk of developing AMR.

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Hybrid Derivative of Cathelicidin and Human Beta Defensin-2 Against Gram-Positive Bacteria: A Novel Approach for the Treatment of Bacterial Keratitis

10.1101/2021.04.22.440925 ◽

2021 ◽

Author(s):

Darren Shu Jeng Ting ◽

Eunice Tze Leng Goh ◽

Venkatesh Mayandi ◽

Joanna M. F. Busoy ◽

Thet Tun Aung ◽

...

Keyword(s):

Bacterial Keratitis ◽

Gram Positive ◽

Host Defense Peptides ◽

Corneal Epithelial ◽

Minimal Inhibitory Concentrations ◽

Hybrid Peptides ◽

Novel Approach ◽

Time Kill ◽

Topical Antimicrobial

ABSTRACTPurposeBacterial keratitis (BK) represents the leading cause of corneal blindness worldwide. This study aimed to generate potent hybridized human-derived host defense peptides (HDPs) as novel topical antimicrobial therapy for bacterial keratitis.MethodsHybrid peptides were rationally designed through combination of functional amino acids in parent HDPs, including LL-37 and human beta-defensin (HBD)-1 to −3. Minimal inhibitory concentrations (MICs) and time-kill kinetics assay were performed to determine the concentration- and time-dependent antimicrobial activity and cytotoxicity was evaluated against human corneal epithelial cells (HCE-2) and erythrocytes. In vivo safety and efficacy of the most promising peptide was examined in the corneal wound healing and Staphylococcus aureus (ATCC SA29213) keratitis murine models, respectively.ResultsA second-generation hybrid peptide (CaD23), based on rational hybridization of the middle residues of LL-37 and C-terminal of HBD-2, demonstrated good efficacy against methicillin-sensitive and methicillin-resistant S. aureus [MIC=12.5-25.0μg/ml (5.2-10.4μM)] and S. epidermidis [MIC=3.1-12.5μg/ml (1.3-5.2μM)], and moderate efficacy against P. aeruginosa [MIC=50μg/ml (20.9μM)]. CaD23 (at 25μg/ml or 2x MIC) killed all the bacteria within 30 mins, which was 8 times faster than amikacin (25μg/ml or 20x MIC). At 200μg/ml (16x MIC), CaD23 was shown to be relatively safe against HCE-2 (<30% toxicity) and erythrocytes (<10% toxicity). Pre-clinical murine studies showed that CaD23 0.05% (500μg/ml) achieved a median reduction of S. aureus bioburden by 94% (or 1.2 log10 CFU/ml) while not impeding corneal healing.ConclusionsRational hybridization of human-derived HDPs has led to generation of a potentially efficacious and safe topical antimicrobial agent for treating Gram-positive BK.

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A nano-phytochemical ophthalmic solution for marked improvement of corneal wound healing in healthy or diabetic mice

Nanomedicine ◽

10.2217/nnm-2021-0417 ◽

2021 ◽

Author(s):

Qiqi Li ◽

Meng Xin ◽

Xianggen Wu ◽

Bo Lei

Keyword(s):

Wound Healing ◽

Ophthalmic Solution ◽

Diabetic Mice ◽

Promoting Effect ◽

Related Factors ◽

Corneal Wound Healing ◽

Corneal Epithelial ◽

Trigeminal Neurons ◽

Corneal Wound

Aim: To formulate a novel nano-phytochemical ophthalmic solution to promote corneal wound healing. Methods: Dipotassium glycyrrhizinate (DG) and palmatine (PAL) were used to formulate this formulation marked as DG-PAL, and its efficacy and mechanisms for promoting corneal wound healing were evaluated in mice. Results: DG-PAL was easily fabricated with excellent physical profiles. In in vivo efficiency evaluations, DG-PAL demonstrated an excellent promoting effect on corneal epithelial/nerve wound healing in both healthy and diabetic mice. These effects were involved in the DG-PAL-induced decreased expression levels of HMGB1 and its signaling-related factors in the corneas and trigeminal neurons of the healthy or diabetic mice. Conclusion: DG-PAL possibly represents a promising ophthalmic solution for promoting corneal wound healing.

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Daptomycin and AgNP co-loaded rGO nanocomposites for specific treatment of Gram-positive bacterial infection in vitro and in vivo

Biomaterials Science ◽

10.1039/c9bm01229j ◽

2019 ◽

Vol 7 (12) ◽

pp. 5097-5111 ◽

Cited By ~ 5

Author(s):

Chunyi Tong ◽

Li Li ◽

Feng Xiao ◽

Jialong Fan ◽

Xianghua Zhong ◽

...

Keyword(s):

Wound Healing ◽

Bacterial Infection ◽

Specific Treatment ◽

Gram Positive ◽

Gram Positive Bacteria ◽

Bacteria Killing

rGO was used for simultaneously anchoring AgNPs and Daptomycin to prepare rGO@Ag@Dap for anti-bacterium. The new nanomaterial showed strong Gram-positive bacteria killing ability in vitro and enhanced wound healing infected with S. aureus in vivo.

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Fibronectin Enhancement of Corneal Epithelial Wound Healing of Rabbits In Vivo

Archives of Ophthalmology ◽

10.1001/archopht.1984.01040030369040 ◽

1984 ◽

Vol 102 (3) ◽

pp. 455-456 ◽

Cited By ~ 76

Author(s):

T. Nishida ◽

S. Nakagawa ◽

C. Nishibayashi ◽

H. Tanaka ◽

R. Manabe

Keyword(s):

Wound Healing ◽

Corneal Epithelial ◽

Epithelial Wound Healing

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Effect of Ofloxacin on Corneal Epithelial Wound Healing Evaluated by In Vitro and In Vivo Methods

Drug Investigation ◽

10.1007/bf03258459 ◽

1993 ◽

Vol 6 (2) ◽

pp. 96-103 ◽

Cited By ~ 2

Author(s):

Steven S. Matsumoto ◽

Michael E. Stern ◽

Roger M. Oda ◽

Corine R. Ghosn ◽

Josephine W. Cheng ◽

...

Keyword(s):

Wound Healing ◽

Corneal Epithelial ◽

Epithelial Wound Healing

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Hyaluronic Acid Combined with Serum Rich in Growth Factors in Corneal Epithelial Defects

International Journal of Molecular Sciences ◽

10.3390/ijms20071655 ◽

2019 ◽

Vol 20 (7) ◽

pp. 1655 ◽

Cited By ~ 9

Author(s):

Carlota Suárez-Barrio ◽

Jaime Etxebarria ◽

Raquel Hernáez-Moya ◽

Marina del Val-Alonso ◽

Maddalen Rodriguez-Astigarraga ◽

...

Keyword(s):

Wound Healing ◽

Growth Factors ◽

Synergistic Effects ◽

Sodium Hyaluronate ◽

In Vitro Proliferation ◽

Corneal Epithelial ◽

Equivalent Efficacy ◽

And Migration

The aim of this study is to assess if an adhesive biopolymer, sodium hyaluronate (NaHA), has synergistic effects with s-PRGF (a serum derived from plasma rich in growth factors and a blood derivative that has already shown efficacy in corneal epithelial wound healing), to reduce time of healing or posology. In vitro proliferation and migration studies, both in human corneal epithelial (HCE) cells and in rabbit primary corneal epithelial (RPCE) cultures, were carried out. In addition, we performed studies of corneal wound healing in vivo in rabbits treated with s-PRGF, NaHA, or the combination of both. We performed immunohistochemistry techniques (CK3, CK15, Ki67, ß4 integrin, ZO-1, α-SMA) in rabbit corneas 7 and 30 days after a surgically induced epithelial defect. In vitro results show that the combination of NaHA and s-PRGF offers the worst proliferation rates in both HCE and RPCE cells. Addition of NaHA to s-PRGF diminishes the re-epithelializing capability of s-PRGF. In vivo, all treatments, given twice a day, showed equivalent efficacy in corneal epithelial healing. We conclude that the combined use of s-PRGF and HaNA as an adhesive biopolymer does not improve the efficacy of s-PRGF alone in the wound healing of corneal epithelial defects.

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Heparin-Modified Amniotic Membrane Combined With Growth Factors for Promoting Corneal Wound Healing After Alkali Burn

Frontiers in Bioengineering and Biotechnology ◽

10.3389/fbioe.2020.599800 ◽

2020 ◽

Vol 8 ◽

Author(s):

Xuan Zhao ◽

Xin Zuo ◽

Jing Zhong ◽

Bowen Wang ◽

Saiqun Li ◽

...

Keyword(s):

Wound Healing ◽

Growth Factors ◽

Sustained Release ◽

Corneal Epithelial Cell ◽

Control Group ◽

Therapeutic Effects ◽

Cell Growth And Migration ◽

Corneal Epithelial

Ocular chemical burns are potentially blinding ocular injuries and require urgent management. Amniotic membrane (AM) transplantation is an effective surgical treatment, one of the reasons is because AM is a rich source of growth factors that can promote epithelialization and wound healing. However, growth factors will be gradually lost and insufficient after preparation process and long-time storage, leading to unsatisfactory therapeutic effects. Herein, we present a modified AM (AM-HEP) for the supplement and sustained release of growth factor by surface grafting heparin for treatment of ocular chemical burns. Heparin grafting rate and stability, microstructure, physical property, and sustained release of epithelial growth factor (EGF) of AM-HEP were characterized. Biocompatibility and ability to promote corneal epithelial cell growth and migration were evaluated and compared with a biological amnion, which is available on the market in vitro. The therapeutic effects of AM-HEP combined with EGF (AM-HEP@EGF) in vivo had been evaluated in a model of mouse corneal alkali burn. The results indicated that heparin was introduced into AM and maintain stability over 3 weeks at 37°C. The modification process of AM-HEP did not affect microstructure and physical property after comparing with non-modified AM. EGF could be combined quickly and effectively with AM-HEP; the sustained release could last for more than 14 days. AM-HEP@EGF could significantly promote corneal epithelial cell growth and migration, compared with non-modified AM and control group. Faster corneal epithelialization was observed with the transplantation of AM-HEP@EGF in vivo, compared with the untreated control group. The corneas in the AM-HEP@EGF group have less inflammation and were more transparent than those in the control group. The results from in vitro and in vivo experiments demonstrated that AM-HEP@EGF could significantly enhance the therapeutic effects. Taken together, AM-HEP@EGF is exhibited to be a potent clinical application in corneal alkali burns through accelerating corneal epithelial wound healing.

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