Decline In Endothelial Cell Density Research Articles

Evaluation of the safety of anterior capsule staining with trypan blue under air: a retrospective analysis.

To evaluate the safety of using 0.03% trypan blue under air for anterior capsule staining in cataract surgery. The current study involved a retrospective analysis of the medical records of 86 patients with vitreous hemorrhage, who underwent pars plana vitrectomy and cataract surgery. The patients were classified into two groups. The trypan blue group (n=45) comprised patients who underwent anterior capsule staining with 0.03% trypan blue under an air bubble. The control group (n=41) comprised of patients who underwent intracameral illuminator-assisted capsulorhexis. The status of endothelial cell density (ECD) in both the groups was analyzed. The trypan blue group displayed significant decline in ECD at 1mo (7.91% loss, P<0.001) and 3mo (9.65% loss, P<0.001) after the surgery, whereas no significant changes were observed in the control group. Moreover, the number of patients who did not display a postoperative decline in ECD was significantly higher in the control group (43.9%; 18 patients) than in the trypan blue group (17.1%; 7 patients, P=0.004). Anterior capsule staining with trypan blue under the air bubble would not be as safe as the intracameral illuminator. The ECD loss might be attributed to the air bubble rather than to the deleterious effects of 0.03% trypan blue. Further studies are required to clarify this.

Long-Term Graft Survival and Decline in Endothelial Cell Density Following Penetrating Keratoplasty with Organ-Cultured Corneas.

IntroductionEndothelial cell density (ECD) changes long after penetrating keratoplasty (PKP) of organ-cultured corneas have been little studied. We aim to calculate the point when ECD decline stabilises following PKP with organ culture stored corneas.MethodsThis is an observational study of first-ever PKPs and first-ever re-grafts, performed over 17 years under a single surgeon. ECDs were acquired at 3 and 6 months, 1 year post-graft and annually thereafter by specular microscopy. Time-dependent ECD data was fitted to a log-biexponential model.ResultsWe studied 465 first-ever grafts and 128 re-grafts. Mean recipient age was 59 years (range 0–96 years; SD 22). Median follow-up was 5.7 (range 0.2–17.1) years. Probability of ED at 5 years in first grafts and re-grafts was 4.4% (2.6–7.1%) and 14.8% (8.3–23.2%). In first grafts, ECD loss reached 0.6% per annum at 7.9 (6.2–9.6) years post-operatively. The half-lives of ECD loss during the immediate post-operative period for first grafts, re-grafts, dystrophies, ectasias, and previous ocular surgery are 20.1 (14.9–30.9), 12.8 (6.9–79.4), 19.5 (13.1–37.7), 26.2 (16.2–68), and 11.6 (6.7–41.3) months, respectively. The half-life during this rapid phase of ECD loss has an inverse correlation with graft survival at 10 years (r = − 0.89, p = 0.02).ConclusionsRate of endothelial decompensation is higher in first grafts than re-grafts. ECD decline stabilises 7.9 years post-operatively in first grafts but then becomes lower than the physiological loss expected. Further work is needed to verify whether organ-cultured grafts reach physiological levels of ECD loss faster than hypothermically stored grafts.

Preclinical testing of small diameter Descemet membrane endothelial keratoplasty grafts to increase tissue availability.

In this study, we describe a process of preparing, surgically manipulating, and validating a novel “small diameter” 4mm circular Descemet membrane endothelial keratoplasty (DMEK) graft in vitro. Three small diameter DMEK grafts can be prepared from a single donor endothelium and could, therefore, potentially expand the donor pool. Prior to clinical use, however, we aimed to examine each step of the process to determine the effect on the endothelial cell loss and whether or not cells retained their capacity to migrate uniformly. For this study, circular small diameter grafts, obtained from twelve corneas of ten donors deemed ineligible for transplantation, were included. Small diameter DMEK graft preparation was successful in all cases (n = 36). Endothelial cell density (ECD), determined in the eye bank on seventeen grafts, showed an average decrease from 2413 (±189) cells/mm2 before to 2240 (±413) cells/mm2 after preparation. Twenty-four grafts were used to simulate DMEK-surgery in vitro and were successfully stained with 0.06% trypan blue, loaded into a straight DMEK-injector, unfolded, positioned, and centered within the circular ~ 4mm descemetorhexis. The estimated % area populated by viable cells on the grafts decreased from on average 92 (±3) % before to 78 (±10) % (n = 4) after in vitro surgery. Cells displayed a capacity for uniform cell migration from all edges of the graft (n = 4) when embedded in the 3D hydrogel system. Our data show, that by using an in vitro model of DMEK-surgery it was possible to test the 4mm circular DMEK grafts from eye bank preparation to surgical implantation. The cell loss after in vitro surgery was comparable with the in vivo ECD decline early after DMEK and the capacity of the cells to migrate to potentially cover bare stroma indicates that these small diameter grafts may be a viable clinical option to treat central endothelial disease.

Endothelial Cell Density Changes in the Corneal Center Versus Paracentral Areas After Descemet Membrane Endothelial Keratoplasty.

To analyze whether endothelial cell density (ECD) differs between central and paracentral areas after Descemet membrane endothelial keratoplasty (DMEK) and to identify the locations of the highest and lowest ECD. In this retrospective cohort study, central and paracentral ECDs of 30 eyes of 30 patients who underwent DMEK for Fuchs endothelial corneal dystrophy were evaluated. Central, superonasal, superotemporal, and inferior specular microscopic images were analyzed at 1, 3, 6, 9, and 12 months after DMEK. Changes in ECD by location and over time and changes in location for the highest and lowest ECD were evaluated. When compared with the preoperative donor ECD, the central ECD decreased by 32 (±11)% at 12 months postoperatively. ECD decline between 1 and 12 months postoperatively in the central, inferior, superonasal, and superotemporal location were 7%, 12%, 16%, and 13%, respectively (P < 0.0001 for all locations). Mean ECD was the highest in the center between the 3- to 12-month follow-up, whereas mean ECD was the lowest in the paracentral superonasal area at all time points (P < 0.001). Mean ECD of the highest and lowest density locations differed at all follow-up time points (P < 0.003). Postoperative ECD was not found to be uniform across the graft. Contrary to the density distribution in the normal endothelium, paracentral area ECDs were found to be consistently lower than in the central area. These differences were most prominent in the superonasal area and persisted 12 months postoperatively. Central ECD after DMEK might, thus, not represent an accurate proxy for cell density of the entire graft.

Changes in Endothelial Cell Count Up to Three Years After Implantation of Toric Implantable Collamer Lenses.

To determine the temporal effect of toric implantable collamer lens (TICL) implantation and location on corneal endothelial cell density (ECD) over a period of 36 months after surgery. ECD [number of cells per square millimeter estimated using the Specular Microscope SP-1P (Topcon Europe Medical B.V., Netherlands)] data were collected from cases deemed suitable for the TICL (VTICMO, VTICM5; STAAR Surgical, Nidau, Switzerland). The preoperative refractive error (sphere and cylinder) ranged from -1.00 to -22.25 diopter sphere and from -0.50 to -5.50 diopter cylinder. ECD was evaluated at preoperative and all postoperative sessions. Key findings were as follows: the mean ECD (±SD, 95% confidence interval) was 2720 cells/mm (±272, 2620-2820 cells/mm) preoperatively, which was reduced to 2372 cells/mm (±325, 2250-2490 cells/mm) at 36 months postoperatively (P < 0.001). Linear regression revealed the following significant correlations between the (1) log of the change in ECD (y1) and log of preoperative ECD (x1) at 2 years postoperatively, y1 = 2.513x1-6.2816 (n = 62, r= 0.3503, P = 0.005); (2) mean ECD (y2) and log time (in months, x2), y2= 2543.7-36.997x2-38.99x2 (r=-0.9654, n = 7, P = 0.0004); and (3) mean axial distance between the front surface of the crystalline lens and the TICL back surface (y3) and time postoperatively (in months, x3), y3 = 0.1035x3-5.2808x3 +473.18 (r = 0.8512, n = 7, P = 0.015). Expected ECD loss after TICL implantation by 2 years postoperatively is predictable. On average, over 3 years after implantation, there is (1) an initial rapid decline in ECD, followed by a gradual fall in the rate of cell loss, and (2) a gradual fall in the distance between the TICL and the crystalline lens by 2 years postoperatively, followed by a reversal by the third year.

Effects of Perfluorobutylpentane (F4H5) on Corneal Endothelial Cells

ABSTRACTPurpose: To evaluate the effects of perfluorobutylpentane (F4H5) on corneal endothelial cell density (ECD) and morphology using a porcine corneal endothelial organ culture model.Materials and methods: “Split corneal buttons” were cultivated for 15 days (d) after incubation in F4H5 (15, 30, 60, and 120 min) or BSS (controls). ECD was assessed manually on d1, d8, and d15. After histological staining (trypan blue, alizarin red S) on d15 morphological changes (reformation figures, rosette formations, and alizarin red cells) were evaluated.Results: ECD was significantly reduced after incubation in F4H5 for 120 min (median ± 25%/75%-quartile; 3281 ± 43/222 cells/mm2; p = 0.046) on d15 compared to controls (3658 ± 129/296 cells/mm2), but not after shorter incubation times (15, 30, and 60 min). Morphological assessment supports these findings as reformation figures (F4H5 120 min: 10.5 ± 9.3/13.9/mm2 vs. controls: 5.2 ± 2.8/7.2/mm2; p = 0.010), rosette formations (F4H5 120 min 25.566 ± 17.044/36.219/mm2 vs. controls: 8.333 ± 0.000/15.667/mm2; p = 0.002), and alizarin red cells (F4H5 120 min: 38.350 ± 29.827/51.333/mm2 vs. controls: 20.833 ± 10.417/25.000/mm2; p = 0.049) were significantly more prevalent after incubation in F4H5 for 120 min compared to controls. Also, F4H5 60 min showed significantly more rosette formations (25.452 ± 16.968/36.057/mm2; p = 0.006) and alizarin red cells (46.662 ± 42.420/50.903/mm2; p = 0.007), but not reformation figures (7.0 ± 2.2/1.6 %; p = 0.953).Conclusion: Short exposure (≤30 min) of porcine corneal endothelial cells to F4H5 does not have significant effects on ECD or morphological characteristics. Longer exposure times (≥60–120 min) may cause ECD decline and/or induce morphological changes.

Long-Term Functional and Anatomical Outcome after Descemet Stripping Automated Endothelial Keratoplasty: A Prospective Single-Center Study.

Purpose To investigate the long-term anatomical and functional outcomes of Descemet stripping automated endothelial keratoplasty (DSAEK). Methods Prospective follow-up of 114 eyes (95 subjects) after DSAEK for endothelial dysfunction. Measurements included best spectacle-corrected visual acuity (BSCVA), straylight, endothelial cell density (ECD), and graft thickness. Results The mean follow-up time was 5.1 ± 1.5 years. Four grafts ultimately failed (after 5 to 7 years). From baseline up to 1 year after DSAEK, mean BSCVA improved by 0.30 logMAR. This beneficial effect remained until the last follow-up (LFU). After DSAEK, straylight was reduced. ECD sharply dropped by 900 cells/mm2 (33%) immediately after surgery and, thereafter, steadily decreased at a rate of 11 cells/mm2 per month. No significant correlation was observed between graft thickness at 3 years and BSCVA. Conclusions We observed a low graft failure rate and a normalization of graft thickness. Postoperative straylight remained elevated relative to the normal population. The sharp initial and the subsequent more gradual ECD decline are consistent with other studies. A significant and prolonged functional gain can be achieved by posterior lamellar grafting for endothelial dysfunction.

Long-Term Endothelial Cell Loss in Patients with Artisan Myopia and Artisan Toric Phakic Intraocular Lenses: 5- and 10-Year Results

To evaluate the long-term change in endothelial cell density (ECD) after the implantation of 2 types of rigid iris-fixated phakic intraocular lenses (pIOLs) for the treatment of myopia and astigmatism. Prospective, clinical cohort study. A total of 507 eyes of 289 patients receiving the Artisan Myopia or Artisan Toric (Ophtec B.V., Groningen, The Netherlands) iris-fixated pIOL for the treatment of myopia or astigmatism at the University Eye Clinic Maastricht as of January1998. A total of 381 myopic and 126 toric pIOLs were implanted. Five- and 10-year follow-ups were completed by 193 and 127 eyes implanted with the myopic pIOL and by 40 and 20 eyes implanted with the toric pIOL, respectively. Chronic endothelial cell (EC) loss, percentage of eyes with a decrease of ≥25% in ECD, and percentage of eyes with an ECD <1500 cells/mm2. Chronic EC loss was calculated from 6 months postoperatively to the end of follow-up and showed an annual ECD decline of 48 cells/mm2 (standard error, 3.14) and 61 cells/mm2 (standard error, 6.30) in the myopic (P < 0.001) and toric (P < 0.001) groups, respectively, resulting in a total EC loss of 16.6% and 21.5% from 6 months to 10 years postoperatively, respectively. Ten years after implantation, ECD had decreased by ≥25% in 7.9% and 6.3%, whereas ECD was <1500 cells/mm2 in 3.9% and 4.0% in the myopic and toric groups, respectively. Explantation of the pIOL occurred in 6.0% in the myopic group and 4.8% in the toric group. Risk factors for increased EC loss were a shallow anterior chamber depth (ACD) (P ≤ 0.005) and a smaller distance between the central and peripheral pIOL edge to the endothelium (P ≤ 0.005). A significant linear chronic EC loss was reported after implantation with myopic or toric iris-fixated pIOLs. A smaller ACD and smaller distance between pIOL edge and endothelium were risk factors for EC loss. Modification of preoperative age-related ECD thresholds is indicated to maintain an ECD that warrants safe future combined pIOL explantation and cataract surgery.

Identification of Prdm genes in human corneal endothelium

Corneal endothelial cells (CECs) are essential for maintaining corneal stromal hydration and ensuring its transparency, which is necessary for normal vision. Dysfunction of CECs leads to stromal decompensation, loss of transparency and corneal blindness. Corneal endothelium has low proliferative potential compared to surface epithelial cells leading to poor regeneration of CEC following injury. Additionally, the tissue exhibits age related decline in endothelial cell density with re-organisation of the cell layer, but no regeneration. The mechanisms which control proliferation and differentiation of neural crest derived CEC progenitors are yet to be clearly elucidated.Prdm (Positive regulatory domain) family of transcriptional regulators and chromatin modifiers are important for driving differentiation of a variety of cellular types. Many Prdm proteins are expressed in specific precursor cell populations and are necessary for their progression to a fully differentiated phenotype. In the present work, we sought to identify members of the Prdm gene family which are specifically expressed in human (h) CECs with a view to begin addressing their potential roles in CEC biology, focussing especially on Prdm 4 and 5 genes. By performing semi-quantitative reverse transcription coupled to PCR amplification we found that in addition to Prdm4 and Prdm5, Prdm2 and Prdm10 genes are expressed in hCECs. We further found that cultured primary hCECs or immortalised HCEC-12 cells express all of the Prdm genes found in CECs, but also express additional Prdm transcripts. This difference is most pronounced between Prdm gene expression patterns of CECs isolated from healthy human corneas and immortalised HCEC-12 cells. We further investigated Prdm 4 and Prdm 5 protein expression in cultured primary hCECs and HCEC-12 cells as well as in a human cadaveric whole cornea. Both Prdm 4 and Prdm 5 are expressed in human corneal endothelium, primary hCECs and in HCECs-12 cells, characterised by expression of the Na+/K+-ATPase. We observed that both proteins exhibit cytosolic (intracellular, but non-nuclear and distinct from extracellular fluid) as well as nuclear localisation within the endothelial layer, with Prdm 5 being more concentrated in the nuclei of the endothelial cells than Prdm 4. Thus, our work identifies novel Prdm genes specifically expressed in corneal endothelial cells which may be important in the control of CEC differentiation and proliferation.

Corneal Endothelial Cell Density in Children: Normative Data From Birth to 5 Years Old

To establish a normative database of endothelial cell density (ECD) using invivo specular microscopy in children under 5 years old. Cross-sectional study. Specular microscopy was performed during a clinic visit in cooperative children in the standard upright position. In uncooperative children, specular microscopy was performed in the lateral decubitus position under general anesthesia, before surgery for other reasons. Corneal diameter (CD) was measured in children undergoing general anesthesia and was stratified according to age. One hundred and eighteen eyes of 118 patients were included in the study. The mean patient age was 2.6 ± 1.4 years (range 0.1-5 years) and the mean ECD was 3746 ± 370 cells/mm2 (range 3145-5013 cells/mm2). The mean CD under 2 years of age was 11.85 ± 0.57mm (n= 40, range 10.50-12.75mm). Up to 2 years of age, ECD was more inversely correlated with CD than with age (r=-0.61, P < .0001; r=-0.38, P= .01, respectively). In contrast, after the age of 2 years, the ECD was inversely correlated with age but not with CD (r=-0.27, P= .02; r=-0.24, P= .2). Between the first and second year of life, the rate of ECD decrease was significantly higher than between 2 and 5 years of age (8.2%, 334 cells/mm2 vs 2.7%, 100 cells/mm2 a year, respectively). In the first 2 years of life there is a rapid decline in ECD, which is likely related to growth in CD and hence surface area. After the cornea reaches adult size, the ECD decreases at a rate similar to that reported in adults.

Corneal endothelial cell loss after pars plana vitrectomy and combined phacoemulsification–vitrectomy surgeries

ObjectiveTo compare postoperative corneal endothelial cell density (ECD) loss in eyes undergoing pars plana vitrectomy (PPV), or combination of cataract extraction (using phacoemulsification) and intraocular lens implantation with vitrectomy (CE/IOL-PPV) surgeries. MethodsInstitutional setting. Best-corrected visual acuity (BCVA) and ECD by specular microscopy were measured preoperatively and 3 months postoperatively in both groups. Relative postoperative ECD loss was the primary outcome measure. Change in BCVA was the secondary outcome measure. ResultsForty eyes of 40 patients undergoing PPV and 46 eyes of 46 patients undergoing CE/IOL-PPV were included in the final analysis. Postoperative ECD was decreased slightly more in the CE/IOL-PPV group compared with the PPV group (13.9% ± 15.5% vs 9.0% ± 14.6%), although this was not statistically significant (p = 0.10). The improvement in the logMAR BCVA was, however, statistically more significant in the CE/IOL-PPV group compared with the PPV group (−56.6% ± 24.3% vs −38.6% ± 45.5%, p = 0.04). ConclusionsPPV and the combination CE/IOL-PPV surgeries lead to modest and statistically similar postoperative decline in ECD. The combination surgery may lead to slightly more postoperative cells loss, but also more improvement in visual acuity.

Descemet Stripping Endothelial Keratoplasty: Ten-Year Endothelial Cell Loss Compared with Penetrating Keratoplasty

To assess endothelial cell density (ECD) and model 10-year cell loss after Descemet stripping endothelial keratoplasty (DSEK), and to compare with results after penetrating keratoplasty (PK) performed for the same indications in the Cornea Donor Study (CDS). Retrospective, longitudinal study. Five hundred ninety DSEK recipients, 21 to 96 years of age, treated for Fuchs' dystrophy (n= 498; 84%), pseudophakic or aphakic corneal edema (n= 89; 15%), or other endothelial dysfunction (n= 3; <1%). Review of 1005 consecutive primary DSEK procedures by 6 surgeons identified 752 grafts (75%) in 590 recipients with ECD measurements at 1 or more postoperative examinations between 6 months and 10 years. Four statistical models applied previously to the CDS PK data were considered. The preferred model (Bayesian Markov chain Monte Carlo) accounted for missing data, selective dropout from graft failure, correlations between fellow eyes, and correlations between longitudinal repeated measures. Central corneal ECD. The median donor corneal ECD (25th-75th percentiles) was 3005 cells/mm(2) (2852-3203 cells/mm(2)) at baseline, 2077 cells/mm(2) (1660-2411 cells/mm(2)) at 6 months, and 926 cells/mm(2) (617-1433 cells/mm(2)), with a range from 408 to 2538 cells/mm(2) at 10 years. After median cell loss of 32% in the perioperative period, ECD declined at a linear rate of approximately 110 cells/mm(2) per year between 6 months and 10years after DSEK. At 10 years, the median cell loss was 71% (53%-80%) relative to the baseline donor ECD, comparable with the 76% (70%-82%) median cell loss after PK by 68 surgeons in the CDS. The 10-year ECD correlated with the 6-month postoperative ECD (r= 0.56; P < 0.001), but not with the baseline donor ECD(r=-0.04; P= 0.78). Selective dropout from graft failure did not affect the cell loss model significantly. The linear decline in ECD after DSEK was consistent with shorter-term endothelial keratoplasty studies and was distinct from the biexponential cell loss trend characteristic of PK. By 10 years, cell loss was comparable in surviving clear grafts for both DSEK and PK.

Association Between Graft Storage Time and Donor Age With Endothelial Cell Density and Graft Adherence After Descemet Membrane Endothelial Keratoplasty.

After retrospectively evaluating the clinical outcome of 500 consecutive cases after Descemet membrane endothelial keratoplasty (DMEK), we extended the analysis in this study by assessing the effect of donor-related parameters on endothelial cell density (ECD) decline and detachment rate in this group. This retrospective case series included 500 cases who had undergone DMEK from October 2007 to September 2012 at the Netherlands Institute for Innovative Ocular Surgery (NIIOS), Rotterdam, the Netherlands. Logistic regression analysis (n = 332 eyes) showed that donor age might be associated with a 3% increase in the risk for a detachment (odds ratio, 0.97; 95% CI, 0.94-1.00; P = .049) (ie, higher donor age seems to be associated with lower chances of a detachment). In addition, linear regression analysis indicated that graft storage time in medium was associated with ECD decrease (ie, the longer the storage time, the larger the decrease at 6 months after DMEK) (P = .01). We showed an association between graft storage time and ECD decline after DMEK and possibly between donor age and graft detachment. Therefore, donor storage times should be kept as short as possible to improve short-term ECDs. More research is needed to draw definite conclusions on the possible effect of donor age on the chance of a detachment after DMEK.

How Much Progress Has Been Made in Corneal Transplantation?

To assess the benefits of component lamellar corneal surgery, we investigated the surgical outcomes of deep anterior lamellar keratoplasty (DALK) for keratoconus and Descemet stripping automated endothelial keratoplasty (DSAEK) for laser iridotomy-induced bullous keratopathy (LI-BK). The results were compared with the surgical outcomes of penetrating keratoplasty (PKP). Seventy-nine eyes with keratoconus treated with DALK and 81 eyes with LI-BK treated with DSAEK were studied, and the graft clarity rate, best spectacle-corrected visual acuity, endothelial cell density (ECD), and surgical complications were compared with 273 eyes with keratoconus and 98 eyes with LI-BK all treated with PKP. Keratoconus eyes showed excellent visual outcomes with DALK and PKP, but graft survival was higher for PKP (100%, 98.9%, and 96.3% vs. 98.6%, 90.6%, and 86.1%, at 1, 3, and 5 years after surgery, respectively; P < 0.0001). There were no significant differences in the ECD up to 1 year after surgery. Greater ECD decline was observed for PKP at 3 and 5 years after surgery. Descemet membrane microperforation was observed in 49 eyes treated with DALK. Postoperative complications in eyes treated with PKP were increased incidence of immunological rejection and secondary glaucoma compared with eyes treated with DALK. LI-BK graft survival rates were excellent in both groups (DSAEK: 89.6% and 86.1%; PKP: 100% and 98.6%, 1- and 2-year rates, respectively; P = 0.17). Best spectacle-corrected visual acuity was better in the DSAEK group, and ECD showed a higher rate of decrease with DSAEK than with PKP. DALK and DSAEK considerably improved surgical outcomes in eyes with keratoconus and those with LI-BK, respectively. However, several key areas require further work, including development of safe surgical methods in DALK and prevention of early postoperative ECD loss in DSAEK.

WNT10B Enhances Proliferation through β-Catenin and RAC1 GTPase in Human Corneal Endothelial Cells

The cornea is the anterior, transparent tissue of the human eye that serves as its main refractive element. Corneal endothelial cells are arranged as a monolayer on the posterior surface of the cornea and function as a pump to counteract the leakiness of its basement membrane. Maintaining the cornea in a slightly dehydrated state is critical for the maintenance of corneal transparency. Adult human corneal endothelial cells are G1-arrested, even in response to injury, leading to an age-dependent decline in endothelial cell density. Corneal edema and subsequent vision loss ensues when endothelial cell density decreases below a critical threshold. Vision loss secondary to corneal endothelial dysfunction is a common indication for transplantation in developed nations. An impending increase in demand for and a current global shortage of donor corneas will necessitate the development of treatments for vision loss because of endothelial dysfunction that do not rely on donor corneas. Wnt ligands regulate many critical cellular functions, such as proliferation, making them attractive candidates for modulation in corneal endothelial dysfunction. We show that WNT10B causes nuclear transport and binding of RAC1 and β-catenin in human corneal endothelial cells, leading to the activation of Cyclin D1 expression and proliferation. Our findings indicate that WNT10B promotes proliferation in human corneal endothelial cells by simultaneously utilizing both β-catenin-dependent and -independent pathways and suggest that its modulation could be used to treat vision loss secondary to corneal endothelial dysfunction.

Hypothermic storage or organ culture?

AbstractCurrently there are two storage practices for the cornea, both use liquids based on cell culture medium: the hypothermic storage at 2‐6°C, and organ culture at 30‐37°C. Hypothermic storage seems to offer donor tissues of good quality comparable to that obtained by organ culture, provided that the storage time is kept short. Indeed, according to the literature, the risk of primary graft failure increases significantly after storage longer than 7 days. Furthermore, corneas stored longer than 7 days display epithelial alterations that may hinder the surgical procedure or delay the full recovery of the graft. The organ culture storage method consists of a storage period in culture medium at 30‐37C°, and a shorter de‐swelling and transportation phase at 30‐37C° and room temperature in the same medium supplemented with 4‐8% dextran. Organ culture solutions contain fetal calf serum as a source of growth factors to limit endothelial cell loss. The serum must be obtained from prion‐free countries. A storage period of 30 days can be achieved without significant loss of endothelial cells. The endothelium shows reparative phenomena during storage. Organ culture offers a longer storage time, a less restricted donor supply on beforehand, corneal endothelium with a better defined quality, and a pre‐operative microbiologic control. Cultured corneas always display an epithelium made up of 2‐3 layers of viable cells. The 30 days storage period allows an efficient use of valuable donor tissue: planning of operations is easier, allowing sufficient time for the allocation. The disadvantages of this method are the relative technical complexity and the need for qualified staff to perform tissue culture and selection of the corneas. The studies comparing the effect of the storage methods on outcome demonstrate similar graft survival and post‐operative decline in endothelial cell density. Corneas stored by organ culture are at least comparable with those stored hypothermically for shorter periods.

Graft Rejection After Descemet's Stripping Automated Endothelial Keratoplasty: Graft Survival and Endothelial Cell Loss

To evaluate the effects of graft rejection episodes after Descemet's stripping automated endothelial keratoplasty surgery (DSAEK) on long-term endothelial cell density (ECD) decline and graft survival. Retrospective, comparative analysis of an interventional case series. We included 615 eyes of 415 Fuchs' dystrophy patients at a single institution with ≥ 6 months follow-up and without comorbidities known to influence postoperative ECD. All patients were enrolled as part of an ongoing, institutional review board-approved clinical protocol for a long-term, prospective study of endothelial keratoplasty in patients with endothelial dysfunction. Preoperative specular microscopy of donor corneal tissue was performed. Postoperative specular microscopy measurements were recorded at 6 and 12 months, and yearly thereafter. The percentages of endothelial cell loss recorded at 1, 2, 3, and 4 years were compared with the Mann-Whitney U test. Percentage ECD declines were calculated at each time point from the results of the preoperative and postoperative specular microscopy. Patients with graft rejection episodes and late endothelial failure were identified. Graft rejection was defined as findings of keratic precipitates with or without corneal edema, or anterior chamber cell and flare with or without corneal edema after the initial resolution of perioperative inflammation. We identified 45 cases of graft rejection. The greatest number of rejections occurred between postoperative months 12 and 18. Eyes with a graft rejection episode had a higher median percentage decline in ECD at all time points compared with eyes without graft rejection episodes. This was statistically significant at 2 and 3 years postoperatively. There is a trend toward a greater percentage of ECD loss with time in eyes experiencing graft rejection after DSAEK surgery. Proprietary or commercial disclosure may be found after the references.

Endothelial Cell Survival and Graft Profile Analysis in Descemet Stripping Endothelial Keratoplasty

To report the results of descemet stripping endothelial keratoplasty (DSEK) for a single-surgeon, consecutive case series. All patients undergoing DSEK at our institution from 2006 to 2007 under the care of a single consultant ophthalmologist were enrolled. This was a nonrandomized, retrospective case series. Grafts were dissected manually using an artificial anterior chamber and a Morlet lamellar dissector. Data were collected for refractive error, visual acuity, endothelial cell density, graft thickness, and graft profile. Twenty-one eyes of 20 patients were included. The average age at surgery was 69 ± 11 years (range, 37-88 years). The main indication for DSEK was Fuchs endothelial dystrophy. The mean preoperative best spectacle-corrected visual acuity was 1.2 logarithm of the minimum angle of resolution, improving to 0.48 logarithm of the minimum angle of resolution postoperatively (P = 0.001). Endothelial cell density decreased postoperatively compared with preoperatively, but the rate of cell loss decreased over time. No significant correlation was observed between mean graft thickness and final visual acuity or between graft profile and refractive shift. DSEK is a positive alternative to PK in the treatment of endothelial dysfunction. Visual function improves and the associated refractive change is negligible. Complication rates are low, and graft survival over 2 years is high. Manual preparation of tissue provides grafts of suitable thickness and profile, and endothelial cell density decline is comparable with that of donor's tissue cut via microkeratome or after PK.

Organ culture, but not hypothermic storage, facilitates the repair of the corneal endothelium following mechanical damage

To evaluate the reparative capacity of the mechanically injured endothelium of corneas stored under organ culture (OC) or hypothermic conditions. The central endothelium of 12 pairs of human corneas with similar endothelial parameters was damaged to create a 1 mm(2) lesion. One cornea from each pair was stored under OC and one under hypothermic conditions. The endothelial cell density (ECD), coefficient of variation, hexagonality and percentage of dead cells were assessed before and after damage and on days 7, 14, 21 and 28 of storage. The mean ECD of corneas subsequently stored under OC or hypothermic conditions was 2764/mm(2). Immediately after damage, a denuded Descemet's membrane with a few remaining dead cells was observed at the injured area. After 7 days of storage under OC conditions, almost no dead cells were observed at the place of injury. A non-significant worsening of the qualitative parameters (polymegatism and pleomorphism) was found. After 14 days, ECD was 1933/mm(2) and 2478/mm(2) centrally and pericentrally, respectively. Similar values were found after 21 and 28 days of storage. The lesions with remnant dead cells persisted throughout hypothermic preservation. From day 14 the corneas became cloudy and in poor condition, while the pericentral ECD was 2523/mm(2). The reparative capacity of the cornea is maintained under OC but not under hypothermic conditions. For corneas containing dead endothelial cells, OC is therefore the method of choice because it may improve the quality of the stored tissue.

The influence of donor age and cause of death on corneal endothelial cell density

To evaluate the joint effects of donor age and cause of death on the endothelial cell density of fresh human corneas. The corneal endothelial cell density of 946 eyes from 946 donors was analysed. The donors were divided into three age groups: group 1 comprised those aged 50 years and less; group 2 those aged 51-75 years, and group 3 those aged over 75 years. They were also grouped according to the cause of death: group A included those who had died of cancer; group B those who had died of acute myocardial infarction, pulmonary embolism or intracerebral haemorrhage, and group C those who had died as a result of other diseases. There was a statistically significant negative correlation between donor age and mean corneal endothelial cell density, which was 2868 +/- 309 cells/mm(2) in group 1, 2716 +/- 335 cells/mm(2) in group 2, and 2670 +/- 368 cells/mm(2) in group 3. Among the various causes of death, the mean endothelial cell density was 2677 +/- 343 cells/mm(2) in group A, 2741 +/- 367 cells/mm(2) in group B, and 2713 +/- 337 cells/mm(2) in group C. The endothelial cell density was significantly lower in group A than in group B (p = 0.02). When the donors in the oldest age group (group 3) were analysed according to the various causes of death, the difference between group A (2603 +/- 342 cells/mm(2)) and group B (2729 +/- 338 cells/mm(2)) was even more significant (p = 0.0067). Longlasting, severe diseases like cancer, leading to cachexia and catabolism, reduce the number of endothelial cells to a greater degree than diseases that cause a more rapid death. This negative effect of cancer on endothelial cell density is aggravated by the general decline in endothelial cell density with advancing age.