Activation of dendritic cells by crosslinked collagen hydrogels (artificial corneas) varies with their composition

ACKNOWLEDGEMENTS We are grateful to the Iain Fraser Flow Cytometry Centre, the Microscopy and Histology Facility, and the Medical Research Facility at the University of Aberdeen. This work was supported by the Royal College of Surgeons of Edinburgh, UK, and Saving Sight in Grampian/Development Trust of the University of Aberdeen, UK. Funding Information: Saving Sight in Grampian/Development Trust of the University of Aberdeen, UK Royal College of Surgeons of Edinburgh, UKPeer reviewedPostprin

Biomaterials-enabled cornea regeneration in patients at high risk for rejection of donor tissue transplantation

The severe worldwide shortage of donor organs, and severe pathologies placing patients at high risk for rejecting conventional cornea transplantation, have left many corneal blind patients untreated. Following successful pre-clinical evaluation in mini-pigs, we tested a biomaterials-enabled pro-regeneration strategy to restore corneal integrity in an open-label observational study of six patients. Cell-free corneal implants comprising recombinant human collagen and phosphorylcholine were grafted by anterior lamellar keratoplasty into corneas of unilaterally blind patients diagnosed at high-risk for rejecting donor allografts. They were followed-up for a mean of 24 months. Patients with acute disease (ulceration) were relieved of pain and discomfort within 1-2 weeks post-operation. Patients with scarred or ulcerated corneas from severe infection showed better vision improvement, followed by corneas with burns. Corneas with immune or degenerative conditions transplanted for symptom relief only showed no vision improvement overall. However, grafting promoted nerve regeneration as observed by improved touch sensitivity to normal levels in all patients tested, even for those with little/no sensitivity before treatment. Overall, three out of six patients showed significant vision improvement. Others were sufficiently stabilized to allow follow-on surgery to restore vision. Grafting outcomes in mini-pig corneas were superior to those in human subjects, emphasizing that animal models are only predictive for patients with non-severely pathological corneas; however, for establishing parameters such as stable corneal regeneration and nerve regeneration, our pig model is satisfactory. While further testing is merited, we have nevertheless shown that cell-free implants are potentially safe, efficacious options for treating high-risk patients

Collagen analogs with phosphorylcholine are inflammation-suppressing scaffolds for corneal regeneration from alkali burns in mini-pigs

The long-term survival of biomaterial implants is often hampered by surgery-induced inflammation that can lead to graft failure. Considering that most corneas receiving grafts are either pathological or inflamed before implantation, the risk of rejection is heightened. Here, we show that bioengineered, fully synthetic, and robust corneal implants can be manufactured from a collagen analog (collagen-like peptide-polyethylene glycol hybrid, CLP-PEG) and inflammation-suppressing polymeric 2-methacryloyloxyethyl phosphorylcholine (MPC) when stabilized with the triazine-based crosslinker 4-(4,6-Dimethoxy-1,3,5-triazin-2-yl)-4-methylmorpholinium chloride. The resulting CLP-PEG-MPC implants led to reduced corneal swelling, haze, and neovascularization in comparison to CLP-PEG only implants when grafted into a mini-pig cornea alkali burn model of inflammation over 12 months. Implants incorporating MPC allowed for faster nerve regeneration and recovery of corneal sensation. CLP-PEG-MPC implants appear to be at a more advanced stage of regeneration than the CLP-PEG only implants, as evidenced by the presence of higher amounts of cornea-specific type V collagen, and a corresponding decrease in the presence of extracellular vesicles and exosomes in the corneal stroma, in keeping with the amounts present in healthy, unoperated corneas

Phytochemical Investigations of Campsis Radicans L.

Petroleum ether, dichloromethane and ethyl acetate soluble fractions were obtained through partitioning the crude methanolic extract of the leaves of Campsis radicans L. (Family. Bignoniaceae) followed by the chromatographic separation of secondary metabolites from them. A total of five triterpene compounds i.e., corosolic acid methyl ester (1), β-amyrin (2), arjunolic acid (3), maslinic acid (4) and 28-O-[β-D-glucopyranosyl-(1→6)-β-D-glucopyranosyl]-2α,3α,19α-trihydroxy-12-en-28-ursolic acid (5) were isolated from the dichloromethane fractions and their structures were characterized by 1H NMR spectroscopy and compared the NMR data with published values

The structural and optical properties of type III human collagen biosynthetic corneal substitutes

The structural and optical properties of clinically biocompatible, cell-free hydrogels comprised of synthetically cross-linked and moulded recombinant human collagen type III (RHCIII) with and without the incorporation of 2-methacryloyloxyethyl phosphorylcholine (MPC) were assessed using transmission electron microscopy (TEM), X-ray scattering, spectroscopy and refractometry. These findings were examined alongside similarly obtained data from 21 human donor corneas. TEM demonstrated the presence of loosely bundled aggregates of fine collagen filaments within both RHCIII and RHCIII-MPC implants, which X-ray scattering showed to lack D-banding and be preferentially aligned in a uniaxial orientation throughout. This arrangement differs from the predominantly biaxial alignment of collagen fibrils that exists in the human cornea. By virtue of their high water content (90%), very fine collagen filaments (2-9 nm) and lack of cells, the collagen hydrogels were found to transmit almost all incident light in the visible spectrum. They also transmitted a large proportion of UV light compared to the cornea which acts as an effective UV filter. Patients implanted with these hydrogels should be cautious about UV exposure prior to regrowth of the epithelium and in-growth of corneal cells into the implants. (C) 2015 Acta Materialia Inc. Published by Elsevier Ltd.Funding Agencies|MRC [503626]; Swedish Research Council [521-2012-5706]</p

The structural and optical properties of type III human collagen biosynthetic corneal substitutes

The structural and optical properties of clinically biocompatible, cell-free hydrogels comprised of synthetically cross-linked and moulded recombinant human collagen type III (RHCIII) with and without the incorporation of 2-methacryloyloxyethyl phosphorylcholine (MPC) were assessed using transmission electron microscopy (TEM), X-ray scattering, spectroscopy and refractometry. These findings were examined alongside similarly obtained data from 21 human donor corneas. TEM demonstrated the presence of loosely bundled aggregates of fine collagen filaments within both RHCIII and RHCIII-MPC implants, which X-ray scattering showed to lack D-banding and be preferentially aligned in a uniaxial orientation throughout. This arrangement differs from the predominantly biaxial alignment of collagen fibrils that exists in the human cornea. By virtue of their high water content (90%), very fine collagen filaments (2-9 nm) and lack of cells, the collagen hydrogels were found to transmit almost all incident light in the visible spectrum. They also transmitted a large proportion of UV light compared to the cornea which acts as an effective UV filter. Patients implanted with these hydrogels should be cautious about UV exposure prior to regrowth of the epithelium and in-growth of corneal cells into the implants. (C) 2015 Acta Materialia Inc. Published by Elsevier Ltd.Funding Agencies|MRC [503626]; Swedish Research Council [521-2012-5706]</p

Electron-Beam Irradiated Recombinant Human Collagen-Phosphorylcholine Corneal Implants Retain Pro-Regeneration Capacity

Sterilization of biodegradable, collagen-based implants is challenging as irradiation sterilization methods can alter their mechanical properties. Electron beam (EB) irradiation is a terminal sterilization method that has been used for biologically-derived implants. Here, recombinant human collagen type III-phosphorylcholine (RHCIII-MPC) hydrogels were irradiated with EB doses of 17, 19, or 21 kGy and their subsequent biocompatibility and ability to promote regeneration in rabbit corneas was evaluated. Unirradiated hydrogels stored in 1% chloroform in phosphate-buffered saline (C-PBS) were the controls. There were no significant differences between irradiated and non-irradiated samples in optical or physical properties (tensile strength, modulus, elasticity), or the ability to support cell growth. However, irradiated implants were more sensitive to high levels of collagenase than unirradiated controls and the C-PBS implants had increased cell growth compared to EB and controls at 72 h. Corneal implants e-beamed at 17 kGy or e-beamed and subsequently frozen (EB-F) to increase shelf-life showed no adverse biological effects of the irradiation. EB, EB-F, and C-PBS implanted corneas all rapidly re-epithelialized but showed mild neovascularization that resolved over 6 months. The regenerated neo-corneas were transparent at 6 months post-operation. In vivo confocal microscopy confirmed normal morphology for the epithelium, stroma, sub-basal nerves and unoperated endothelium. Histology showed that all the regenerated corneas were morphologically similar to the normal. Immunohistochemistry indicated the presence of a differentiated corneal epithelium and functional tear film. In conclusion, the e-beamed corneal implants performed as well as non-irradiated control implants, resulting in fully regenerated neo-corneas with new nerves and without blood vessels or inflammation that may impede vision or corneal function. Therefore, a complete validation study to establish EB irradiation as an effective means for corneal implant sterilization prior to clinical application is necessary as a next step

Correction: Short peptide analogs as alternatives to collagen in pro-regenerative corneal implants (vol 69, pg 120, 2018)

n/

Short peptide analogs as alternatives to collagen in pro-regenerative corneal implants

Short collagen-like peptides (CLPs) are being proposed as alternatives to full-length collagen for use in tissue engineering, on their own as soft hydrogels, or conjugated to synthetic polymer for mechanical strength. However, despite intended clinical use, little is known about their safety and efficacy, mechanism of action or degree of similarity to the full-length counterparts they mimic. Here, we show the functional equivalence of a CLP conjugated to polyethylene glycol (CLP-PEG) to full-length recombinant human collagen in vitro and in promoting stable regeneration of corneal tissue and nerves in a preclinical mini-pig model. We also show that these peptide analogs exerted their pro-regeneration effects through stimulating extracellular vesicle production by host cells. Our results support future use of CLP-PEG implants for corneal regeneration, suggesting the feasibility of these or similar peptide analogs in clinical application in the eye and other tissues.Correction in: Acta Biomaterialia, Vol. 81, Pages 330-331.DOI: 10.1016/j.actbio.2018.10.009Correction in: Acta Biomaterialia, Vol.88, Pages 556-557.DOI: 10.1016/j.actbio.2019.01.046</p