Effect of Penetrating Keratoplasty and Keratoprosthesis Implantation on the Posterior Segment of the Eye

Purpose To compare the effects of post-penetrating keratoplasty (PK) and post-keratoprosthesis (KPro) surgery-related inflammation on the posterior segment of the eye and to assess inhibition of tumor necrosis factor alpha (TNFα) and interleukin-1 beta (IL-1β) on these effects. Methods: BALB/C (syngeneic) or C57BL/6 (allogeneic) corneas were transplanted onto BALB/C host beds as part of PK or miniature KPro (m-KPro) implantation. Intraocular pressure (IOP) was measured via an intracameral pressure sensor; tissues were harvested and analyzed 8 weeks after surgery. Expression of TNFα and IL-1β in the retina was analyzed using real-time quantitative (q)PCR. Optic nerve degeneration (axon count, circularity, and area) was assessed quantitatively using ImageJ software. After m-KPro implantation, mice were treated with saline, anti-TNFα, or anti-IL-1β antibody, and axonal loss was assessed after 10 weeks. Results: Mean IOP was within normal limits in the operated and fellow eyes in all groups. The mRNA expression of TNFα and IL-1β was highest in m-KPro groups with either syngeneic or an allogeneic carrier. We observed optic nerve degeneration in both allogeneic PK and m-KPro implanted eyes with an allogeneic carrier. However, TNFα blockade significantly reduced axonal loss by 35%. Conclusions: Allogeneic PK and m-KPro implants with an allogeneic carrier lead to chronic inflammation in the posterior segment of the eye, resulting in optic nerve degeneration. In addition, blockade of TNFα prevents axonal degeneration in this preclinical model of allogeneic m-KPro (alloKPro) implantation

Effect of Penetrating Keratoplasty and Keratoprosthesis Implantation on the Posterior Segment of the Eye

Citation:Črnej A, Omoto M, Dohlman TH, et al. Effect of penetrating keratoplasty and keratoprosthesis implantation on the posterior segment of the eye. Invest Ophthalmol Vis Sci. 2016;57:164357: -164857: . DOI:10.1167 PURPOSE. To compare the effects of post-penetrating keratoplasty (PK) and post-keratoprosthesis (KPro) surgery-related inflammation on the posterior segment of the eye and to assess inhibition of tumor necrosis factor alpha (TNFa) and interleukin-1 beta (IL-1b) on these effects. METHODS. BALB/C (syngeneic) or C57BL/6 (allogeneic) corneas were transplanted onto BALB/ C host beds as part of PK or miniature KPro (m-KPro) implantation. Intraocular pressure (IOP) was measured via an intracameral pressure sensor; tissues were harvested and analyzed 8 weeks after surgery. Expression of TNFa and IL-1b in the retina was analyzed using real-time quantitative (q)PCR. Optic nerve degeneration (axon count, circularity, and area) was assessed quantitatively using ImageJ software. After m-KPro implantation, mice were treated with saline, anti-TNFa, or anti-IL-1b antibody, and axonal loss was assessed after 10 weeks. RESULTS. Mean IOP was within normal limits in the operated and fellow eyes in all groups. The mRNA expression of TNFa and IL-1b was highest in m-KPro groups with either syngeneic or an allogeneic carrier. We observed optic nerve degeneration in both allogeneic PK and mKPro implanted eyes with an allogeneic carrier. However, TNFa blockade significantly reduced axonal loss by 35%. CONCLUSIONS. Allogeneic PK and m-KPro implants with an allogeneic carrier lead to chronic inflammation in the posterior segment of the eye, resulting in optic nerve degeneration. In addition, blockade of TNFa prevents axonal degeneration in this preclinical model of allogeneic m-KPro (alloKPro) implantation

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

Establishment of a novel in vitro model of stratified epithelial wound healing with barrier function

The repair of wounds through collective movement of epithelial cells is a fundamental process in multicellular organisms. In stratified epithelia such as the cornea and skin, healing occurs in three steps that include a latent, migratory, and reconstruction phases. Several simple and inexpensive assays have been developed to study the biology of cell migration in vitro. However, these assays are mostly based on monolayer systems that fail to reproduce the differentiation processes associated to multilayered systems. Here, we describe a straightforward in vitro wound assay to evaluate the healing and restoration of barrier function in stratified human corneal epithelial cells. In this assay, circular punch injuries lead to the collective migration of the epithelium as coherent sheets. The closure of the wound was associated with the restoration of the transcellular barrier and the re-establishment of apical intercellular junctions. Altogether, this new model of wound healing provides an important research tool to study the mechanisms leading to barrier function in stratified epithelia and may facilitate the development of future therapeutic applications

Photographic-Based Optical Evaluation of Tissues and Biomaterials Used for Corneal Surface Repair: A New Easy-Applied Method.

Journal Article; Correction: doi: 10.1371/journal.pone.0146205.PURPOSE Tissues and biomaterials used for corneal surface repair require fulfilling specific optical standards prior to implantation in the patient. However, there is not a feasible evaluation method to be applied in clinical or Good Manufacturing Practice settings. In this study, we describe and assess an innovative easy-applied photographic-based method (PBM) for measuring functional optical blurring and transparency in corneal surface grafts. METHODS Plastic compressed collagen scaffolds (PCCS) and multilayered amniotic membranes (AM) samples were optically and histologically evaluated. Transparency and image blurring measures were obtained by PBM, analyzing photographic images of a standardized band pattern taken through the samples. These measures were compared and correlated to those obtained applying the Inverse Adding-Doubling (IAD) technique, which is the gold standard method. RESULTS All the samples used for optical evaluation by PBM or IAD were histological suitable. PCCS samples presented transmittance values higher than 60%, values that increased with increasing wavelength as determined by IAD. The PBM indicated that PCCS had a transparency ratio (TR) value of 80.3 ± 2.8%, with a blurring index (BI) of 50.6 ± 4.2%. TR and BI obtained from the PBM showed a high correlation (ρ>|0.6|) with the diffuse transmittance and the diffuse reflectance, both determined using the IAD (p<0.005). The AM optical properties showed that there was a largely linear relationship between the blurring and the number of amnion layers, with more layers producing greater blurring. CONCLUSIONS This innovative proposed method represents an easy-applied technique for evaluating transparency and blurriness of tissues and biomaterials used for corneal surface repair.This study was supported by the research project JA TEP-1136 from Junta de Andalucía and MAT2013-43946R from the Spanish Ministry of Economy and Competitiveness.Ye

Correction: Photographic-Based Optical Evaluation of Tissues and Biomaterials Used for Corneal Surface Repair: A New Easy-Applied Method.

[This corrects the article DOI: 10.1371/journal.pone.0142099.]

Covalent Functionalization of PMMA Surface with L-3,4-Dihydroxyphenylalanine (L-DOPA) to Enhance its Biocompatibility and Adhesion to Corneal Tissue

The Boston keratoprosthesis (B-KPro) is globally the most commonly implanted artificial cornea for patients with severe corneal diseases, particularly those with multiple allograft failures. Despite providing a good visual recovery, the poor adhesion between the poly(methyl methacrylate) (PMMA)-made stem and the donor tissue poses a challenge, impacting the clinical outcome of the B-KPro. Using single-molecule covalent bonding, PMMA surface is functionalized with l-3,4-dihydroxyphenylalanine (l-DOPA) and its chemical, optical, mechanical, and biological properties are studied. The functionalization process significantly improves biocompatibility of PMMA, without affecting its optical and mechanical properties. Human corneal fibroblasts (HCF) and human corneal epithelial cells (HCEp) seeded on l-DOPA surface both exhibit greater confluency and metabolic rate compared to those of PMMA during 7-day cell culture. Moreover, HCF cultured on l-DOPA demonstrates a higher expression of ALDH3A1, Ki67, Integrin 1, and FAK with no expression of alpha-SMA, compared to those of PMMA, which instead show greater expression of alpha-SMA. These suggest that l-DOPA surface fosters cellular adhesion, proliferation, and migration, without adversely impacting the phenotype of the cells. This study offers an inexpensive and efficient tactic to modify the surface of materials with l-DOPA to achieve the optimal biocompatibility and biointegration of medical devices

Torsional wave elastography to assess the mechanical properties of the cornea.

Corneal mechanical changes are believed to occur before any visible structural alterations observed during routine clinical evaluation. This study proposed developing an elastography technique based on torsional waves (TWE) adapted to the specificities of the cornea. By measuring the displacements in the propagation plane perpendicular to the axis of the emitter, the effect of guided waves in plate-like media was proven negligible. Ex vivo experiments were carried out on porcine corneal samples considering a group of control and one group of alkali burn treatment ([Formula: see text]OH) that modified the mechanical properties. Phase speed was recovered as a function of intraocular pressure (IOP), and a Kelvin-Voigt rheological model was fitted to the dispersion curves to estimate viscoelastic parameters. A comparison with uniaxial tensile testing with thin-walled assumptions was also performed. Both shear elasticity and viscosity correlated positively with IOP, being the elasticity lower and the viscosity higher for the treated group. The viscoelastic parameters ranged from 21.33 to 63.17 kPa, and from 2.82 to 5.30 Pa s, for shear elasticity and viscosity, respectively. As far as the authors know, no other investigations have studied this mechanical plane under low strain ratios, typical of dynamic elastography in corneal tissue. TWE reflected mechanical properties changes after treatment, showing a high potential for clinical diagnosis due to its rapid performance time and paving the way for future in vivo studies

Graphene-Lined Porous Gelatin Glycidyl Methacrylate Hydrogels: Implications for Tissue Engineering

Despite rigorous research, inferior mechanical properties and structural homogeneity are the main challenges constraining hydrogel's suturability to host tissue and limiting its clinical applications. To tackle those, we developed a reverse solvent interface trapping method, in which organized, graphene-coated microspherical cavities were introduced into a hydrogel to create heterogeneity and make it suturable. To generate those cavities, (i) graphite exfoliates to graphene sheets, which spread at the water/ heptane interfaces of the microemulsion, (ii) heptane fills the microspheres coated by graphene, and (iii) a cross-linkable hydrogel dissolved in water fills the voids. Cross-linking solidifies such microemulsion to a strong, suturable, permanent hybrid architecture, which has better mechanical properties, yet it is biocompatible and supports cell adhesion and proliferation. These properties along with the ease and biosafety of fabrication suggest the potential of this strategy to enhance tissue engineering outcomes by generating various suturable scaffolds for biomedical applications, such as donor cornea carriers for Boston keratoprosthesis (BK)

Golgi α1,2-mannosidase I induces clustering and compartmentalization of CD147 during epithelial cell migration.

CD147 is a widely expressed matrix metalloproteinase inducer involved in the regulation of cell migration. The high glycosylation and ability to undergo oligomerization have been linked to CD147 function, yet there is limited understanding on the molecular mechanisms behind these processes. The current study demonstrates that the expression of Golgi α1,2-mannosidase I is key to maintaining the cell surface organization of CD147 during cell migration. Using an in vitro model of stratified human corneal epithelial wound healing, we show that CD147 is clustered within lateral plasma membranes at the leading edge of adjacent migrating cells. This localization correlates with a surge in matrix metalloproteinase activity and an increase in the expression of α1,2-mannosidase subtype IC (MAN1C1). Global inhibition of α1,2-mannosidase I activity with deoxymannojirimycin markedly attenuates the glycosylation of CD147 and disrupts its surface distribution at the leading edge, concomitantly reducing the expression of matrix metalloproteinase-9. Likewise, treatment with deoxymannojirimycin or siRNA-mediated knockdown of MAN1C1 impairs the ability of the carbohydrate-binding protein galectin-3 to stimulate CD147 clustering in unwounded cells. We conclude that the mannose-trimming activity of α1,2-mannosidase I coordinates the clustering and compartmentalization of CD147 that follows an epithelial injury