A review of Bowman's layer structure, function, and transplantation.

Bowman's layer is an acellular corneal structure, which is considered to be a specially modified anterior stroma. It is presumed, that it forms as a result of ongoing epithelial-stromal interactions and no clear physiological purpose has been proven. Despite this fact, Bowman's layer has found its place in corneal transplantation. It has been performed for over a decade, mainly in treatment of advanced keratoconus with multiple modifications. Transplantation of Bowman's layer can be expected to become a widely used surgical procedure in the treatment of many corneal pathologies involving fragmentation and destruction of Bowman's layer. This article aims to summarize information available on its structure, possible function, and transplantation. A thorough literature search was performed in the PubMed database and Google Scholar using keywords: Bowman's layer, structure, function, preparation and corneal transplantation. All the relevant sources were used, which represent 77 peer-reviewed articles with information corcerning the topic of this article.

The new future perspective in corneal tissue utilisation - methods of preparation and preservation.

PURPOSE: The goal of our study is to find an optimal approach to the preparation and preservation of corneal stromal tissue. We want to compare different methods of corneal stromal tissue creation and storage to optimize the efficacy of this process under the conditions of an eye bank. After we find the most suitable method to create a safe high quality product, we want to prove the possibility of using a single donor cornea for more than one patient. We would also like to verify the feasibility of making more corneal lenticules after the removal of a corneal endothelium for DMEK transplantation. METHODS: We provided morphological (histology, scanning electron microscope) and microbiological analysis in order to compare different methods of corneal lenticule and corneal stromal lamellae preparation and preservation. We also tested the surgical handling of the tissue to secure a safe manipulation of the tissue for clinical use. We compared two methods of corneal lenticule preparation: microkeratome dissection and femtosecond laser. As methods of preservation, we tested hypothermia, cryopreservation at -80 degrees Celsius in DMSO (dimethyl sulfoxide) and storage at room temperature with glycerol. Some intrastromal lenticules and lamellae in each group were previously irradiated with gamma radiation of 25 kGy (KiloGray). RESULTS: Corneal stromal lamellae prepared with a microkeratome have a smoother cut - side surface compared to lamellae prepared with a femtosecond laser. Femtosecond laser preparation caused more irregularities on the surface and we detected more conglomerates of the fibrils, while lamellae made with microkeratome had more sparse network. Using femtosecond laser, we were able to make more than five lenticules from a single donor cornea. Gamma irradiation led to damage of collagen fibrils in corneal stroma and a loss of their regular arrangement. Corneal tissue stored in glycerol showed collagen fibril aggregates and empty spaces between fibrils caused by dehydration. Cryopreserved tissue without previous gamma irradiation showed the most regular structure of the fibrils comparable to storage in hypothermia. CONCLUSION: Our results suggest that formation of a corneal lenticule lamellae by microkeratome results in smoother corneal lenticules, while being much cheaper than formation by femtosecond laser. Gamma irradiation of 25 kGy caused damage of the collagen fibres as well as their network arrangement, which correlated with loss of transparency and stiffer structure. These changes impair possible surgical utilisation of gamma irradiated corneas. Storage in glycerol at room temperature and cryopreservation had similar outcomes and we believe that both methods are appropriate and safe for further clinical use .

Preservation of corneal stromal lenticule: review.

Corneal stromal lenticule is a part of corneal stroma, which can be created by manual dissection, by femtosecond laser from the donor cornea, but chiefly it is a waste product of a refractive procedure ReLex SMILE (Small Incision Lenticule Extraction). Corneal lenticule has a huge potential in corneal surgery. In recent years, many studies have been published to show the possibility to use this tissue to treat corneal defects, as well as in refractive surgery. Thanks to the quantity of lenticules which arise every day during SMILE operations, this tissue is much more accessible than any other kind of corneal tissue. According to the experience with lenticule implantation in animal models, or even human patients, lenticule implantation is considered safe, reversible method, which is not associated with immune rejection or other severe complications. However, the crucial step before the process of lenticule implantation, is proper preservation of this tissue. Donor corneal tissue containing endothelium is usually preserved in hypothermia and then usable maximally for two weeks. Newer methods such as organ culture storage and use of a sterile cornea prolong the time of usability of the tissue. The possibilities for corneal lenticule storage are theoretically wider thanks to the fact, that we do not need to preserve fragile cellular structures. Besides the storage in hypothermia, other preserving methods such as cryopreservation and storage after decellularization have been tested. This review aimed to examine the current literature that describes possible methods of corneal lenticule preservation. A comprehensive search was created based on articles published in English on PubMed.gov, Cochranelibrary.com and Scopus.com using following keywords: corneal lenticule preservation, corneal lenticule storage, cold storage corneal lenticule, corneal lenticule cryopreservation till 2020.

Scleral Grafts in Ophthalmic Surgery. A Review.

AIM: To summarize the history and current trends in the use of scleral grafts in ophthalmology. MATERIALS AND METHODS: We conducted a review of the literature through the MEDLINE and Cochrane Library databases. The search terms were "sclera", "graft", and "surgery". The search resulted in 1596 articles, of which we evaluated 192 as relevant. The relevant articles were sorted chronologically and according to the method of using scleral grafts, which enabled the development of a review article. RESULTS: The sclera has been routinely used in ophthalmology since the 1950s in many different indications. Some of these indications have become practically obsolete over time (for example, use in the surgical management of retinal detachment), but a large number still find application today (especially use in glaucoma or oculoplastic surgery, or as a patch for a defect in the sclera or cornea). CONCLUSION: Even though allogeneic sclera is currently used less frequently in ophthalmology compared to other tissue banking products and the range of its indications has partially narrowed, it remains a useful material due to its availability and properties.

Optimizing Corneal Transplant Safety: The Impact of Mandatory Prion Protein Testing on Iatrogenic Creutzfeldt-Jakob Disease Prevention in the Czech Republic.

PURPOSE: The purpose of this study was to assess the effectiveness of legally mandated testing for pathogenic prion proteins in corneal tissue donors in the Czech Republic, considering its impact on safety, financial, and temporal costs. METHODS: Between January 2007 and December 2023, standardized brain regions were collected from all corneal tissue donors in the Czech Republic. Tissue samples were tested for the presence of pathogenic prion proteins by the Czech Reference Laboratory for Human Prion Diseases. The testing used a Western blot analysis, using 2 distinct monoclonal anti-PrP antibodies. RESULTS: A total of 8030 donors were tested. Four tested samples were initially weakly positive. Subsequent testing conclusively determined these samples to be negative. The remaining 8026 tests yielded negative results confirming the safety of donor screening. CONCLUSIONS: We did not observe any cases of proven transmissible spongiform encephalopathies (TSEs). TSE testing has consistently confirmed that no patients with TSE have been selected into the corneal donor pool, solidifying the effectiveness of the active surveillance program and exclusion criteria. We propose that these mechanisms effectively prevent patients with TSEs from being included in the corneal tissue donor pool. However, the substantial financial costs and a 2-day delay in processing pose challenges, contributing to graft nonutilization and potential negative impacts on patients in acute need. Moreover, the unique requirement for pathogenic prion testing in the Czech Republic also makes importing any corneal grafts from other countries impossible, as those tissues do not fulfill Czech legal requirements.

Chemical, Physical, and Biological Corneal Decellularization Methods: A Review of Literature.

The cornea is one of the most commonly transplanted tissues worldwide. It is used to restore vision when severe visual impairment or blindness occurs in patients with corneal diseases or after trauma. Due to the global shortage of healthy donor corneas, decellularized corneal tissue has significant potential as an alternative to corneal transplantation. It preserves the native and biological ultrastructure of the cornea and, therefore, represents the most promising scaffold. This article discusses different methods of corneal decellularization based on the current literature. We searched PubMed.gov for articles from January 2009 to December 2023 using the following keywords: corneal decellularization, decellularization methods, and corneal transplantation. Although several methods of decellularization of corneal tissue have been reported, a universal standardised protocol of corneal decellularization has not yet been introduced. In general, a combination of decellularization methods has been used for efficient decellularization while preserving the optimal properties of the corneal tissue.

Scleral Grafts in Ophthalmic Surgery. A Review.

AIM: To summarize the history and current trends in the use of scleral grafts in ophthalmology. MATERIALS AND METHODS: We conducted a review of the literature through the MEDLINE and Cochrane Library databases. The search terms were "sclera", "graft", and "surgery". The search resulted in 1596 articles, of which we evaluated 192 as relevant. The relevant articles were sorted chronologically and according to the method of using scleral grafts, which enabled the development of a review article. RESULTS: The sclera has been routinely used in ophthalmology since the 1950s in many different indications. Some of these indications have become practically obsolete over time (for example, use in the surgical management of retinal detachment), but a large number still find application today (especially use in glaucoma or oculoplastic surgery, or as a patch for a defect in the sclera or cornea). CONCLUSION: Even though allogeneic sclera is currently used less frequently in ophthalmology compared to other tissue banking products and the range of its indications has partially narrowed, it remains a useful material due to its availability and properties.

Corneal stromal lenticule transplantation for the treatment of corneal ulcers.

PURPOSE: To evaluate the safety and efficacy of using corneal stromal lenticules (CSLs) obtained during refractive surgery Refractive Lenticule Extraction (ReLEx) with the Small Incision Lenticule Extraction (SMILE) procedure for the treatment of corneal ulcers. METHODS: This retrospective study included 12 eyes of 12 patients, 7 men and 5 women with varying degrees of corneal ulcer. The mean age was 64 ± 18 (range 34 to 95 years). The monitoring included corrected distance visual acuity (CDVA), slit-lamp biomicroscopy examination, a Seidel test, stability of the graft and anterior segment optical coherence tomography (AS-OCT) inspection. Patients were closely monitored for possible postoperative complications for at least 6 months. RESULTS: In 7/12 (58%) eyes, the corneal ulcer was successfully sealed with CSL and amniotic membrane (AM) without the need for any additional surgical intervention. In 3 eyes, penetrating keratoplasty (PK) was needed in addition to CSL transplantation and in 2 eyes the scleral patch was used to fully seal after CSL transplantation. During the follow-up period no signs of rejection or infection were detected in any patient. CONCLUSION: The use of CSLs from ReLEx SMILE may be considered as an alternative method for the treatment of corneal ulcers before a more extensive and definitive solution - PK - is used. Our preliminary findings suggest that properly performed CSL transplantation using cryopreserved lenticules is a safe and effective method to temporarily cover the corneal partial-thickness defect or even perforation.

Novel Biomaterials in Glaucoma Treatment.

Glaucoma is a significant cause of blindness worldwide, and its treatment remains challenging. The disease progressively leads to damage to the optic disc and thus loss of visual acuity and visual field. High intraocular pressure (IOP) is a common risk factor. There are three major methods to treat this disease: topical, laser, and surgical. None of these are completely satisfactory; therefore, alternatives using new biomaterials are being sought. Since biomaterial engineering has experienced significant growth in recent decades, its products are gradually being introduced to various branches of medicine, with the exception of ophthalmology. Biomaterials, such as glaucoma drainage implants, have been successfully used to treat glaucoma. There is significant ongoing research on biomaterials as drug delivery systems that could overcome the disadvantages of topical glaucoma treatment, such as poor intraocular penetration or frequent drug administration. This article summarizes the use of novel biomaterials for glaucoma treatment presented in the literature. The literature search was based on articles published in English on PubMed.gov, Cochranelibrary.com, and Scopus.com between 2018 and 2023 using the following term "biomaterials in glaucoma." A total of 103 published articles, including twenty-two reviews, were included. Fifty-nine articles were excluded on the basis of their titles and abstracts.

The Efficacy of Fibrin Sealant in Sutureless Deep Sclerectomy-Preliminary Outcomes.

PRCIS: Deep sclerectomy (DS) with fibrin adhesive can constitute a safe alternative to the classic procedure using sutures, providing nonallergenic, nontoxic, and secure adhesion with no sign of aqueous humor outflow obstruction postoperatively. OBJECTIVE: To evaluate short and medium-term postoperative results of DS with a fibrin sealant. PATIENTS AND METHODS: This prospective, noncomparative, interventional case series involves 12 eyes of 12 patients with uncontrolled open angle glaucoma who underwent DS with Esnoper (Clip or V2000) implant between February 2021 and March 2022. A novel method of wound closure (sclera, Tenon fascia, and conjunctiva) employing fibrin glue was used instead of classic sutures. Surgical outcomes assessed include: intraocular pressure and glaucoma therapy reduction, best-corrected visual acuity changes, and number of complications registered peri and postoperatively. All measurements were performed preoperatively, as well as at 1 day, at 1 and 2 weeks, and at 1, 2, 3, 6, 9, and 12 months after surgery. RESULTS: The mean intraocular pressure decreased from 24.0 ± 9.1 mm Hg to 13.8 ± 6.3 mm Hg at 1 year postoperatively ( P < 0.001). Kaplan-Meier survival analysis revealed complete and qualified success rates of 83.3% and 91.7%. The mean glaucoma therapy decreased from 3.2 ± 1.1 to 0.8 ± 1.3 drugs 12 months after surgery ( P < 0.001). Nd:YAG goniopunture was performed in 2 eyes at 1 and 12 months postoperatively. No significant best-corrected visual acuity changes were registered. Perioperatively, we noted a trabeculo-descemet microperforation in 1 eye, transient hypotony in 5 eyes, and mild hyphema in 2 eyes. CONCLUSIONS: Fibrin adhesive provided an effective closure in sutureless DS in the patients included in our study. This modification of classical DS may simplify the surgical technique, ensure secure wound adaptation, optimize healing, and lower the risk of inflammation and fibrosis postoperatively.

P37-A124†Smile bank - corneal stromal lenticules from relex smile - the new eye bank product.

PURPOSE: The aim of our presentation is to introduce future eye bank product - corneal stromal lenticule from living donors, which can be used for allotransplantation. METHODS: ReLEx (refractive lenticule extraction) SMILE (small incision lenticule extraction) is a common approach in laser eye surgery. It is minimally invasive and flap-free procedure. During this procedure part of corneal stroma (lenticule) is created by femtosecond laser and consequently removed through small incision. The lenticule is basically waste material of the ReLEx SMILE procedure. In the International Eye Bank of Prague, we decided to establish new protocol for lenticule withdrawal, storage and release for transplantation. RESULTS: All donors signed an informed consent, and their serum was tested for the presence of infectious diseases. After ReLEx SMILE procedure the lenticule was stored in container with cryopreservation solution and frozen in the eye bank using the same protocol for frozen amniotic membrane. After 6 months in -80°C tissues were defrosted and examined histologically, using conventional light histology staining and electron microscopy. CONCLUSION: We believe, that lenticule from living donor is a safe and effective tissue, that can be used for many indications and in particular situations represents good alternative to whole donor cornea and amniotic membrane.

Refractive Outcome After Corneal Lenticule Implantation Ex Vivo Non-Human Study.

PURPOSE: To evaluate changes in corneal refractive parameters after implantation of a stromal lenticule of different thickness. We assume that the refractive outcome depends on the optical power of the used lenticule. METHODS: We conducted an ex-vivo non-human study on 33 normotonic porcine eyeballs divided into two groups, for 4D and 8D human lenticule implantation. Corneal stromal lenticules were obtained as a by-product from a laser procedure ReLEx SMILE. We evaluated corneal refractive parameters measured on Oculus Pentacam© device before and immediately after the intrastromal lenticule implantation. RESULTS: There was no statistically significant difference in corneal refractive parameters between the eyeball groups before lenticule implantation. In both groups, the intrastromal implantation in the depth of 300um led to a significant increase of central corneal pachymetry and corneal anterior steepening. In the 4D group the average central corneal pachymetry increased from 903 ± 124.59 to 1230 ± 148.99 (p = 0.0022) and in 8D group from 733.35 ± 69.60 to 1109 ± 161.64 (p = 0.0008). Induced changes in other studied parameters were not statistically significant, Kmax changed from 45.57 ± 2.78 to 72.07 ± 16.83 (p = 0.0094) and Km front from 40.72 ± 1.60 to 48.87 ± 5.83 (p = 0.0037) in 4D group and in the 8D group average Kmax increased from 42.22 ± 1.54 to 62.95 ± 12.67 (p = 0.0001) and K2 front 40.46 ± 1.64 to 51.51 ± 9.63 (p = 0.0037). There were no significant differences in refractive changes between the 4D and 8D groups after lenticule implantation. CONCLUSION: Intrastromal corneal lenticule implantation induces changes in corneal refractive parameters. In both groups, the implantation induced a significant increase of an anterior corneal steepening without any significant influence on posterior corneal flattening. Corneal lenticule implantation did not lead to any significant change of corneal astigmatism. However, in order to have more precise data for future clinical applications we need to continue with the experiments and verify the results on human corneas.

Phenotype and genotype of concurrent keratoconus and Fuchs endothelial corneal dystrophy.

PURPOSE: To characterise the phenotype and genotype of concurrent keratoconus and Fuchs endothelial corneal dystrophy (KC + FECD). METHODS: We recruited 20 patients with concurrent KC + FECD for a retrospective observational case series from the United Kingdom and the Czech Republic. We compared eight parameters of corneal shape (Pentacam, Oculus) with two groups of age-matched controls who had either isolated keratoconus (KC) or isolated FECD. We genotyped probands for an intronic triplet TCF4 repeat expansion (CTG18.1) and the ZEB1 variant c.1920G >T p.(Gln640His). RESULTS: The median age at diagnosis of patients with KC + FECD was 54 (interquartile range 46 to 66) years, with no evidence of KC progression (median follow-up 84 months, range 12 to 120 months). The mean (standard deviation (SD)) of the minimum corneal thickness, 493 (62.7) µm, was greater than eyes with KC, 458 (51.1) µm, but less than eyes with FECD, 590 (55.6) µm. Seven other parameters of corneal shape were more like KC than FECD. Seven (35%) probands with KC + FECD had a TCF4 repeat expansion of ≥50 compared to five controls with isolated FECD. The average of the largest TCF4 expansion in cases with KC + FECD (46 repeats, SD 36 repeats) was similar to the age-matched controls with isolated FECD (36 repeats, SD 28 repeats; p = 0.299). No patient with KC + FECD harboured the ZEB1 variant. CONCLUSIONS: The KC + FECD phenotype is consistent with KC but with superimposed stromal swelling from endothelial disease. The proportion of cases with a TCF4 expansion is similar in concurrent KC + FECD and age-matched controls with isolated FECD.

Effect of the position of the corneal lamella on the frequency of its detachment.

AIM: To determine the influence of gaps [places where neither the donor's nor the recipient's Descemet's membrane (DM) is present] and overlaps (places where the recipient's DM is covered by the donor's DM) on the frequency of postoperative detachment of DM endothelial keratoplasty (DMEK) lamellae. METHODS: Totally 64 eyes of 64 patients with Fuchs' endothelial dystrophy or bullous keratopathy indicated for DMEK were randomly divided in two groups. The diameter of the implanted DMEK lamella was the same in both groups (8 mm), but we changed the diameter of the removed recipient DM. In the first group (32 eyes), the circular area was approximately 8.5 mm (gaps); in the second group (32 eyes), the diameter was 7.5 mm (overlaps). Postoperatively we noted all cases of detachment visible on the slit lamp and these cases we indicated for rebubbling. We also measured the uncorrected distance visual acuity (UDVA) as well as corrected distance visual acuity (CDVA) in decimal and postoperative endothelial cell density (ECD). The minimum follow-up time was 6mo. RESULTS: The number of rebubbling procedures in the entire group of patients was 13, i.e., 20.3%, with 6 eyes (18.7%) in the gap group, and 7 eyes (21.9%) in the overlap group. Lamella replacement (re-DMEK) was required in 3 (gap group) and 2 patients (overlap group), respectively. The difference between the groups was statistically insignificant. The UDVA was 0.54±0.21 in the gap group and 0.58±0.24 in the overlap group. The CDVA was 0.74±0.22 and 0.80±0.16, respectively. ECD was 1920±491 and 2149±570 cells/mm2. The small differences between both groups were not statistically significant. CONCLUSION: We do not notice any difference in the group of patients with overlaps or gaps of DM. The presence of small areas of gaps or overlaps does not affect the frequency of detachment of the DMEK lamellae.

[Serological findings of Chlamydial pneumoniae in the Czech Republic--control group of patients examined in the study: Chlamydia pneumoniae in the aetiology of keratoconjunctivitis sicca]. / Sérologické nálezy Chlamydiových protilátek v Ceské republice--kontrolní soubor pacientu vysetrených v rámci studie: Chlamydia pneumoniae v etiologii keratokonjunktivitis sicca.

BACKGROUND: One of the aims of IGA NS-10016 Project "Chlamydia pneumoniae in the etiology of keratoconjunctivitis sicca" was to evaluate the serologic results regarding chlamydia infections in patients affected by keratoconjunctivitis sicca, and to compare these findings with serology tests of the control group. Serologic examination comprised anti-LPS (genus specific) antibodies detection, Chlamydia pneumoniae and Chlamydia trachomatis species specific antibodies and antibodies against chlamydia heat-shock protein cHSP60 IgG. METHODS AND RESULTS: The control group was formed by FNKV Ophthalmology Clinic patients who were coming for pre-operative investigation. At the time of laboratory sample acquisition or clinical investigation no acute ocular or general disease was found in these persons. During the last six moths, they had neither antibiotic therapy, nor therapy of respiratory tract chronic inflammation, asthma, joint or cardiovascular disease in their personal history. The control group consisted of 103 subjects. CONCLUSIONS: We detected anamnestic antibodies against CP in 69% of subjects in the control group, in 31% of them IgA and/or IgM were also detected in our study. In 45% of them anti-cHSP60 IgG was positive even in subjects without symptoms of acute infection or inflammatory laboratory markers. It is obvious that this finding can be misleading during serologic investigation and interpretation, and it is always necessary to evaluate results in relation to the clinical condition.

Long-term Results of a Combined Procedure of Cataract Surgery and Descemet Membrane Endothelial Keratoplasty With Stromal Rim.

PURPOSE: To evaluate the long-term results of combining cataract surgery, intraocular lens (IOL) implantation, and Descemet membrane endothelial keratoplasty with peripheral stromal support (DMEK-S). METHODS: The outcomes of 107 eyes of 37 patients who had undergone a combination of cataract surgery, IOL implantation, and DMEK-S between October 2007 and February 2015 were retrospectively evaluated. The average follow-up duration was 18.2 ± 18.3 months (range, 6-84 months). Changes in uncorrected and corrected distance visual acuity, spherical equivalent, and corneal endothelial cell density were analyzed. Intraoperative and postoperative complications were also recorded. RESULTS: The mean logMAR uncorrected distance visual acuity improved from 0.92 ± 0.60 preoperatively to 0.33 ± 0.33, 0.28 ± 0.25, 0.24 ± 0.20, and 0.27 ± 0.27 at 6, 12, 24, and 36 months postoperatively (P < 0.001), respectively. The mean logMAR corrected distance visual acuity improved from 0.62 ± 0.65 preoperatively to 0.17 ± 0.29, 0.13 ± 0.21, 0.07 ± 0.16, and 0.11 ± 0.23 at 6, 12, 24, and 36 months postoperatively (P < 0.001), respectively. The mean endothelial cell density decreases were 50.7% ± 21.1%, 50.8% ± 19.8%, 60.7% ± 16.6%, 62.2% ± 11.8%, and 61.8% ± 20.0% at 6, 12, 24, 36, and 48 months after surgery, respectively. Similar trends, although with slightly better visual outcomes, were found in the subgroup of patients without other eye diseases. The mean final deviation from intended spherical equivalent was 0.77 ± 1.19 D, representing a hyperopic shift. CONCLUSIONS: A combined procedure of cataract surgery, IOL implantation, and DMEK-S is an effective method of treatment in patients with combined corneal disease and cataract, maintaining visual rehabilitation in the long term. Future developments should be performed to minimize the incidence of complications.

Frequency of Complications During Preparation of Corneal Lamellae Used in Posterior Lamellar Keratoplasty Using the Pneumodissection Technique (Big Bubble).

PURPOSE: To determine the frequency of formation of various types of bubbles and the potential impact of donor and lamella parameters on this frequency, and to identify possible risk factors of unsuccessful "big-bubble" creation in preparation of pre-Descemet endothelial keratoplasty and Descemet membrane endothelial keratoplasty with peripheral stromal support. METHODS: Donor age and sex, death to preservation time (DPT), storage time, presence of corneal scars (mainly a condition after cataract surgery), and endothelial cell density of 256 donor corneas were assessed before Descemet membrane endothelial keratoplasty with peripheral stromal support or pre-Descemet endothelial keratoplasty lamella preparation using the big-bubble technique. RESULTS: Mean donor age was 62.3 ± 8.5 years (28.3% women and 71.7% men). Mean endothelial cell density of the donor graft was 2866 ± 255 cells/mm. Mean DPT was 10.12 ± 4.88 hours, and mean storage time of the transplant before surgery was 6.5 ± 4.8 days. Corneal scars were present in 17 donor grafts (6.6%) after cataract surgery. Eleven corneas were devalued because of Descemet membrane rupture during preparation (4.3%). In 182 corneas, standard bubble type I was created (71.7%); in 27 corneas, bubble type II was created; eventually, both types of bubbles formed simultaneously (10.5%); in 47 corneas, no bubble was created (18.4%). CONCLUSIONS: We identified higher endothelial cell density, shorter DPT, and the presence of corneal scars after cataract surgery as risk factors threatening successful bubble formation. The only risk factor for creating type II bubbles was higher donor age in our study.

The suprabasal layer of corneal epithelial cells represents the major barrier site to the passive movement of small molecules and trafficking leukocytes.

AIMS: To investigate the site of barrier function to the passive diffusion of a small molecule (phalloidin) in the corneal epithelium in the mouse. METHODS: Penetration of phalloidin (molecular weight 1115 daltons) into the cornea was evaluated by studying fluorescent binding of phalloidin to actin in tissue sections, in whole mount preparations, and in the fixed intact globe by confocal microscopy. In addition, the location of tight junction proteins in the individual layers of the corneal epithelium was determined by immunohistochemistry. RESULTS: Phalloidin staining of corneal sections was positive in all corneal layers in tissue sections and in all layers of the corneal epithelium except the suprabasal layer in excised fixed whole mounts of the cornea. However, when phalloidin staining was attempted in intact fixed globes, before excision of the cornea for whole mount preparation, only the most superficial layer of cells was stained indicating that phalloidin could not penetrate the tissue beyond the suprabasal epithelial layer. Detergent (Triton X-100) treatment of the excised cornea and the intact fixed globe, allowed penetration of phalloidin into the suprabasal epithelial layer. Tight junction proteins occludin, ZO-1 and claudin were present in most layers of the cornea but while ZO-1 and occludin were distributed in a typical pericellular pattern, claudin seemed to be particularly prominent in the suprabasal layer and appeared only as a discontinuous punctate pericellular pattern in the superficial layer. Intraepithelial leukocytes were detected in the superficial epithelium and the basal epithelium but not in the suprabasal epithelium. CONCLUSION: The suprabasal epithelium cell layer appears to represent the main barrier site to the passage of small molecules and cells in the mouse cornea and this property may be attributable to prominent claudin expression in this layer.

Mesenchymal stem cell therapy for retro-corneal membrane - A clinical challenge in full-thickness transplantation of biosynthetic corneal equivalents.

Artificial corneas (keratoprostheses) and biosynthetic collagen-based corneal equivalents are surgical implants designed to ease the global burden of corneal blindness. However, keratoprostheses in many cases fail due to development of fibrous retro-corneal membranes (RCM). Fibrous membranes which develop in the anterior chamber after prosthesis implantation do so on a matrix of fibrin. This study investigated fibrin deposition and RCM formation after full-thickness collagen-based hydrogel implants and compared them with syngeneic and allogeneic corneal grafts in mice. Fibrin cleared from the anterior chamber within 14 days in both allo- and syn-grafts but, persisted in hydrogel implants and developed into dense retro-corneal membrane (RCM) which were heavily infiltrated by activated myofibroblasts. In contrast, the number of CD11b+ macrophages infiltrating the initial deposition of fibrin in the anterior chamber (AC) after hydrogel implantation was markedly reduced compared to syn- and allo-grafts. Inoculation of mesenchymal stem cells prior to collagen gel implant promoted clearance of gel-associated fibrin from the anterior chamber. We propose that a failure of macrophage-mediated clearance of fibrin may be the cause of RCM formation after collagen-based hydrogel implants and that mesenchymal stem cell therapy promotes clearance of fibrin and prevents RCM formation. STATEMENT OF SIGNIFICANCE: The manuscript addresses the potential value of bone marrow-derived mesenchymal stem cell therapy for retro-corneal membrane (RCM) formation in full-thickness transplantation of biosynthetic corneal equivalents. This work reports the pathophysiological changes in the anterior chamber of the mouse eye following full-thickness recombinant human cross-linked collagen-based hydrogel implants in which persistent fibrin promotes the development of dense RCM. Furthermore, pre-treatment with mesenchymal stem cells reduces RCM formation and enhances corneal transparency.

The role of Chlamydia pneumoniae in the etiology of keratoconjunctivitis sicca (KCS).

AIM: The authors aimed to show the possible relationship between keratoconjunctivitis sicca (KCS) and Chlamydia pneumonia from the point of view of clinical and microbiological diagnostics. MATERIAL AND METHODS: 94 adult patients were treated for follicular conjunctivitis with symptoms of KCS with possible Chlamydia pneumoniae etiology. The diagnosis of a chlamydial infection is based on the serological positivity of chlamydia antibodies and is further based on the antigen positivity in conjunctival imprint preparations. Patients were treated with azithromycin for a period of 12 days. RESULTS: The reciprocal relationship between chlamydial infection and ocular symptoms was proved at 21 patients (22%). Ninety% of patients showed positive anti-Chlamydia pneumoniae IgA and/or IgM with positivity in 80%, including anti-LSP IgA and/or IgM antibodies. This finding was in correlation with the medium to strongly positive finding of anti-cHSP60 IgG. In two patients, this infection was confirmed by the positivity of Chlamydia pneumoniae DNA in peripheral leucocytes. The test group (100 healthy persons) showed 69% negative finding of anti-Chlamydia pneumoniae antibodies or only positive anamnestic antibodies (IgG) and 31% positive antibodies IgA or IgM without clinical sings. CONCLUSION: This study indicated the possible relationship between KCS and Chlamydia pneumoniae in the course of simultaneous clinical signs of follicular conjunctivitis. KCS is a consequence of the action of local infection at the surface of the conjunctiva. It also indicated the necessity of simultaneous evaluation of microbiological findings and the clinical picture in consideration of overall antibiotic treatment in view of the high antibody background of Chlamydia pneumoniae in the adult population in the Czech Republic. The authors aimed to show the possible relationship between the keratoconjunctivitis sicca and Chlamydia pneumoniae based on results of the two studies. Some patents on conjunctivitis are also briefly described in this article.