Absence of detectable SARS-CoV-2 replication in ex vivo cultured cornea and cornea-derived epithelial cells.

PURPOSE: To study the possibility of SARS-CoV-2 to infect human corneal cells and tissues under standard corneal culture conditions using explants of COVID-19 donors and primary cornea-derived epithelial cells. METHODS: Cornea isolated from deceased COVID-19 donors was cultured for 4 weeks, and SARS-CoV-2 replication was monitored by qRT-PCR. Furthermore, primary corneal epithelial cells from healthy donors were cultured ex vivo and infected with SARS-CoV-2 and human cytomegalovirus (HCMV) as a control. Infection status was assessed by western blotting and reporter gene expression using green fluorescent protein-expressing viral strains. ACE2 and TMPRSS2 receptor expression levels in cornea and epithelial cells were assessed by qRT-PCR. RESULTS: We did not detect SARS-CoV-2 replication in 10 corneas isolated from deceased COVID-19 patients and cultured for 4 weeks, indicating absence of infection under natural conditions. Furthermore, high-titer SARS-CoV-2 infection of ex vivo cultured cornea-derived epithelial cells did not result in productive virus replication. In contrast, the same cells were highly permissive for HCMV. This phenotype could potentially be explained by low ACE2 and TMPRSS2 transcriptional activity in cornea and cornea-derived epithelial cells. CONCLUSIONS: Our data suggest that cornea and limbal epithelial cells are refractory to productive SARS-CoV-2 infection. This could be due to the absence of robust receptor expression levels necessary for viral entry. This study adds further evidence to support the very low possibility of transmission of SARS-CoV-2 from an infected corneal transplant donor to a recipient in corneal organ cultures.

Outcomes after Descemet membrane endothelial keratoplasty over a period of 7 years at a tertiary referral center: endothelial cell density, central corneal thickness, and visual acuity.

PURPOSE: To better assess clinical trajectories of patients with or without ocular comorbidity after Descemet membrane endothelial keratoplasty. BACKGROUND: To report on the outcomes of eyes with differing starting conditions following surgery. DESIGN: Retrospective study at a University Eye Hospital. PARTICIPANTS: 361 eyes separated into group 1 (n=229; eyes with endothelial disease only) and group 2 (n=132; eyes with additional ocular comorbid conditions, such as herpetic eye disease 18/132 (13.6%), glaucoma 16/132 (12.1%), dry age-related macular degeneration 14/132 (10.6%), epiretinal membranes 10/132 (7.6%), and wet age-related macular degeneration 9/132 (6.8%)). METHODS: Consecutive eyes that underwent Descemet membrane endothelial keratoplasty over a follow-up period of up to 7 years at a tertiary referral center were reviewed. Main outcome measures were best-corrected visual acuity, postoperative complications, graft survival, central corneal thickness, and endothelial cell density. RESULTS: Postoperative best-corrected visual acuity at year 1 improved in both groups significantly (Wilcoxon signed rank test: group 1, p =.002; .63 to .23 logMAR; group 2, p <.001; 1.15 to .87 logMAR) with a group difference in favor of group 1 (p =.009, Mann-Whitney-Wilcoxon). A decrease of the endothelial cell density and central corneal thickness was noted at postoperative year 1 for both groups (paired t-tests (group 1, p <.001; group 2, p =.045) and paired t-tests (group 1, p <.001; group 2, p =.003). Complications were less common, and graft longevity was superior in group 1. CONCLUSION: Eyes with different starting conditions might experience a visual improvement and benefit from surgery. Descemet membrane endothelial keratoplasty is a valid treatment for endothelial disorders in manifold of eyes. Further long-term studies are required.

Can SARS-CoV-2 infect the eye? An overview of the receptor status in ocular tissue.

Is the new coronavirus SARS-CoV­2 able to infect ocular tissue and thus poses a risk of infection through the tissue in addition to the risk of contact? This is the question that has occupied ophthalmologists since the beginning of the outbreak. In order to infect a certain type of tissue specific receptors for each virus and sometimes also coreceptors or other proteins must be present. The aim of this review was to summarize and reflect the current state of research with the help of the currently available literature as of 28 May 2020. At the time of the research, angiotensin-converting enzyme 2 (ACE2) was clearly identified as the receptor and transmembrane serine protease 2 (TMPRSS2) as the necessary protease to enable the infection of human cells with SARS-CoV­2. In the eye both ACE2 and TMPRSS2 are expressed, although sometimes very weakly and with varying degrees in different tissues. It is noteworthy that very different results were obtained with different methods. Several reasons can account for this effect: Firstly, the method of detection or preservation of the tissue, secondly, the possibly different expression of the tested tissue samples and thirdly, a possibly rapid loss of receptor expression post-mortem. Therefore, an infection of the eye seems possible, which has already been reported in various publications. The amount of virus or receptor expression necessary to cause an infection is not known. According to current state of knowledge the eye is not considered to be a high-risk tissue due to the low ACE2 and TMPRSS2 expression. Nevertheless, appropriate protective measures are necessary for both medical personnel and patients. In cases of corneal transplantation an infection of the donor tissue with SARS-CoV­2 must be excluded.

Testing for SARS-CoV-2 seroprevalence: experiences of a tertiary eye centre.

INTRODUCTION: The actual prevalence of a SARS-CoV-2 infection and the individual assessment of being or having been infected may differ. Facing the great uncertainty-especially at the beginning of the pandemic-and the possibility of asymptomatic or mildly symptomatic, subclinical infections, we evaluate the experience of SARS-CoV-2 antibody screening at a tertiary clinical setting. METHODS AND ANALYSIS: All employees of a tertiary eye centre and a research institute of ophthalmology were offered antibody testing in May 2020, using a sequential combination of different validated assays/antigens and point-of-care (POC) testing for a subset (NCT04446338). Before taking blood, a systematic inquiry into past symptoms, known contacts and a subjective self-assessment was documented. The correlations between serostatus, patient contacts and demographic characteristics were analysed. Different tests were compared by Kappa statistics. RESULTS: Among 318 participants, SARS-CoV-2 antibodies were detected in 9 employees. Chemiluminescence assays (chemiluminescence immunoassay and electrochemiluminescence) showed superior specificity and high reproducibility, compared with ELISA and POC results.In contrast to the low seropositivity (2.8%) of healthcare workers, higher than that of the other departments of the hospital, a large proportion mistakenly assumed that they might have already been infected. Antiviral antibody titres increased and remained on a plateau for at least 3 months. CONCLUSIONS: The great demand and acceptance confirmed the benefit of highly sensitive testing methods in the early phase of the pandemic. The coincidence of low seroprevalence and anxious employees may have contributed to internalising the need of hygiene measures.

First results of investigations of SARS-CoV-2 RNA in human corneal tissue.

Preliminary investigations of human corneal tissues from coronavirus disease 2019 (COVID-19) cadaveric donors indicated that no severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) RNA is present. Current eye banking guidelines do not recommend any type of routine testing for SARS-CoV­2 RNA in post-mortem donor tissue. This is partly based on factors that can influence the test results of the reverse transcription polymerase chain reaction (RT-PCR).

Absence of Severe Acute Respiratory Syndrome-Coronavirus-2 RNA in Human Corneal Tissues.

PURPOSE: To examine corneal tissue for severe acute respiratory syndrome-coronavirus 2 (SARS-CoV-2) positivity regarding implications for tissue procurement, processing, corneal transplantation, and ocular surgery on healthy patients. We performed quantitative reverse transcription-polymerase chain reaction qRT-PCR-testing for SARS-CoV-2 RNA on corneal stroma and endothelium, bulbar conjunctiva, conjunctival fluid swabs, anterior chamber fluid, and corneal epithelium of coronavirus disease 2019 (COVID-19) postmortem donors. METHODS: Included in this study were 10 bulbi of 5 COVID-19 patients who died because of respiratory insufficiency. Informed consent and institutional review board approval was obtained before this study (241/2020BO2). SARS-CoV-2 was detected by using a pharyngeal swab and bronchoalveolar lavage. Tissue procurement and tissue preparation were performed with personal protective equipment (PPE) and the necessary protective measures. qRT-PCR-testing was performed for each of the abovementioned tissues and intraocular fluids. RESULTS: The qRT-PCRs yielded no viral RNA in the following ocular tissues and intraocular fluid: corneal stroma and endothelium, bulbar-limbal conjunctiva, conjunctival fluid swabs, anterior chamber fluid, and corneal epithelium. CONCLUSIONS: In this study, no SARS-CoV-2-RNA was detected in conjunctiva, anterior chamber fluid, and corneal tissues (endothelium, stroma, and epithelium) of COVID-19 donors. This implicates that the risk for SARS-CoV-2 infection using corneal or conjunctival tissue is very low. However, further studies on a higher number of COVID-19 patients are necessary to confirm these results. This might be of high importance for donor tissue procurement, processing, and corneal transplantation.

Severe acute respiratory Syndrome-Coronavirus-2: Can it be detected in the retina?

BACKGROUND/OBJECTIVES: The systemic organ involvement of SARS-CoV-2 needs to be thoroughly investigated including the possibility of an ocular reservoir in humans. To examine retinal tissues and vitreous for histopathology and SARS-CoV-2 presence with regard to possible effects on the human retina and/ or vitreous. We performed histopathological analyses and quantitative (q)RT-PCR-testing for SARS-CoV-2 RNA on retinal tissues and vitreous of COVID-19 postmortem donors. SUBJECTS/METHODS: Included in this study were 10 eyes of 5 deceased COVID-19 patients. The diagnosis of SARS-CoV-2 infection was confirmed via pharyngeal swabs and broncho-alveolar fluids. The highest level of personal protective equipment (PPE) and measures was employed during fluid-tissue procurement and preparation. Histopathological examinations and qRT-PCR-testing were carried out for all retinal tissues and vitreous fluids. RESULTS: The histopathological examinations revealed no signs of morphologically identifiable retinal inflammation or vessel occlusions based on hematoxylin and eosin stains. By qRT-PCRs, we detected no significant level of viral RNA in human retina and vitreous. CONCLUSIONS: In this study, no significant level of SARS-CoV-2-RNA was detected in the human retinal and vitreous fluid samples of deceased COVID-19 patients. Histopathological examinations confirmed no morphological sign of damage to retinal vasculature or tissues. Further studies are needed to confirm or refute the results.

[Basic principles of replication and immunology of SARS-CoV-2]. / Grundlagen der Replikation und der Immunologie von SARS-CoV-2.

Coronaviruses are a genetically highly variable family of viruses that infect vertebrates and have succeeded in infecting humans many times by overcoming the species barrier. The severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2), which initially appeared in China at the end of 2019, exhibits a high infectivity and pathogenicity compared to other coronaviruses. As the viral coat and other viral components are recognized as being foreign by the immune system, this can lead to initial symptoms, which are induced by the very efficiently working immune defense system via the respiratory epithelium. During severe courses a systemically expressed proinflammatory cytokine storm and subsequent changes in the coagulation and complement systems can occur. Virus-specific antibodies, the long-term expression of which is ensured by the formation of B memory cell clones, generate a specific immune response that is also detectable in blood (seroconversion). Specifically effective cytotoxic CD8+ T­cell populations are also formed, which recognize viral epitopes as pathogen-specific patterns in combination with MHC presentation on the cell surface of virus-infected cells and destroy these cells. At the current point in time it is unclear how regular, robust and durable this immune status is constructed. Experiences with other coronavirus infections (SARS and Middle East respiratory syndrome, MERS) indicate that the immunity could persist for several years. Based on animal experiments, already acquired data on other coronavirus types and plausibility assumptions, it can be assumed that seroconverted patients have an immunity of limited duration and only a very low risk of reinfection. Knowledge of the molecular mechanisms of viral cycles and immunity is an important prerequisite for the development of vaccination strategies and development of effective drugs.

Descemet Membrane Endothelial Keratoplasty: Development of Contrast Sensitivity and Color Vision in Patients With Fuchs Endothelial Corneal Dystrophy.

PURPOSE: To assess the developments in contrast sensitivity, color vision, and subjective perception after Descemet membrane endothelial keratoplasty (DMEK) in patients with Fuchs endothelial corneal dystrophy (FECD). METHODS: Included in this study were pseudophakic, unilateral DMEK patients with bilateral FECD having a follow-up period of 6 months (n = 23). The mean age at surgery was 70 years (range: 52-81 years). Pseudophakic eyes without history of other ocular pathology or surgery served as a control (n = 10). Pelli-Robson contrast sensitivity and Panel-D15 color vision tests were used. Best-corrected visual acuity, modified visual functioning questionnaire-25, central corneal thickness, and endothelial cell density were assessed. We visualized the subjective impression of patients with bilateral FECD after unilateral DMEK in a subgroup using Photoshop CS6. RESULTS: Contrast sensitivity improved significantly from 1.35 ± 0.26 to 1.64 ± 0.17 (P = 0.002; control eyes: 1.92 ± 0.09). No difference in the color vision error score was observed for preoperative and postoperative eyes (P = 0.063). The best-corrected visual acuity improved significantly after surgery (P = 0.001). The average values in the Logarithm of the Minimum Angle of Resolution were 0.59 ± 0.42 preoperatively and 0.1 ± 0.10 postoperatively (control eyes: 0.01 ± 0.03). Examinations revealed a decrease of the central corneal thickness and endothelial cell density after surgery (P = 0.001; P = 0.001, respectively). Scores in the general and the driving questionnaire were significantly higher after surgery (P = 0.001; P = 0.005, respectively). CONCLUSIONS: This study showed significant improvement in subjective patient satisfaction and contrast sensitivity. Spontaneous subjective color vision improvement might be explained by significantly improved contrast sensitivity. Contrast sensitivity might be considered as a parameter in preoperative decision-making and evaluation of surgical outcome.

[Can SARS-CoV-2 infect the eye?-An overview of the receptor status in ocular tissue]. / Kann SARS-CoV-2 das Auge infizieren? ­ Ein Überblick über den Rezeptorstatus in okularem Gewebe.

Is the new coronavirus SARS-CoV­2 able to infect ocular tissue and thus poses a risk of infection through the tissue in addition to the risk of contact? This is the question that has occupied ophthalmologists since the beginning of the outbreak. In order to infect a certain type of tissue specific receptors for each virus and sometimes also coreceptors or other proteins must be present. The aim of this review was to summarize and reflect the current state of research with the help of the currently available literature as of 28 May 2020. At the time of the research, angiotensin-converting enzyme 2 (ACE2) was clearly identified as the receptor and transmembrane serine protease 2 (TMPRSS2) as the necessary protease to enable the infection of human cells with SARS-CoV­2. In the eye both ACE2 and TMPRSS2 are expressed, although sometimes very weakly and with varying degrees in different tissues. It is noteworthy that very different results were obtained with different methods. Several reasons can account for this effect: Firstly, the method of detection or preservation of the tissue, secondly, the possibly different expression of the tested tissue samples and thirdly, a possibly rapid loss of receptor expression post-mortem. Therefore, an infection of the eye seems possible, which has already been reported in various publications. The amount of virus or receptor expression necessary to cause an infection is not known. According to current state of knowledge the eye is not considered to be a high-risk tissue due to the low ACE2 and TMPRSS2 expression. Nevertheless, appropriate protective measures are necessary for both medical personnel and patients. In cases of corneal transplantation an infection of the donor tissue with SARS-CoV­2 must be excluded.

[Seroprevalence and SARS-CoV-2 testing in healthcare occupations]. / Seroprävalenz und SARS-CoV-2-Testung in Gesundheitsberufen.

The SARS-CoV­2 causes a disease spectrum that includes asymptomatic and mildly symptomatic infections with subclinical manifestations but which can nevertheless still be potentially contagious. Evidence from SARS-CoV­2 infected macaque monkeys and from studies with seasonal coronaviruses suggests that the infection is likely to produce an immunity that is protective for a certain period of time. Available test methods enable a high degree of reliability, e.g. if high-quality serological methods are combined. Although individual test results have to be interpreted with caution, serosurveillance in a tertiary eye care center and large eye research institute can reduce anxiety and provide clarity regarding the actual number of (unreported) SARS-CoV­2 infections.

[Importance of corneal organ culture in donors with possible SARS-CoV-2 infections]. / Bedeutung der Hornhautorgankultur bei Spendern mit möglicher SARS-CoV-2-Infektion.

The appearance of the novel severe acute respiratory syndrome-coronavirus-2 (SARS-CoV­2) poses challenges in ophthalmology particularly for eye banks. A valid risk assessment for the removal and processing of donor corneas is difficult due to the lack of data. The risk to infect transplant recipients with SARS-CoV­2 still appears very unlikely due to the experience with severe acute respiratory syndrome -coronavirus(­1) (SARS-CoV(­1)) and Middle East respiratory syndrome-coronavirus (MERS-CoV); however, due to the occurrence of angiotensin-converting enzyme 2 (ACE2) receptors in the cornea an infection of this tissue with SARS-CoV­2 cannot be completely excluded. Therefore, routine testing of the organ culture medium used for donor corneas for SARS-CoV­2 prior to transplantation during the coronavirus disease 2019 (COVID­19) pandemic should be considered.

Absence of Severe Acute Respiratory Syndrome-Coronavirus-2 RNA in ocular tissues.

PURPOSE: To evaluate the status of ocular donor tissues of a COVID-19 postmortem donor. METHODS: SARS-CoV-2 was detected via a pharyngeal swab and broncho-alveolar lavage in the COVID-19 suspect. Postmortem tissue procurement and preparation were performed with personal protective equipment (PPE) and the necessary protective measures. qRT-PCR-testing was performed for the following ocular tissues and fluids: conjunctival fluid swabs, bulbar conjunctiva, corneal epithelium, corneal stroma, corneal endothelium, anterior chamber fluid, lens, iris, vitreous, retina, uvea, sclera, and optic nerve. Informed consent and Institutional Review Board approval was obtained prior to this study (196/2020BO2; Date of approval: 03/26/2020; Ethics Committee of the University of Tuebingen). RESULTS: In all ocular tissue and fluid samples no SARS-CoV-2 RNA was detected via qRT-PCR of the confirmed COVID-19 postmortem donor. CONCLUSIONS AND IMPORTANCE: Late-stage COVID-19 patients might not harbor an ocular reservoir of SARS-CoV-2. The risk of transmitting SARS-CoV-2 via ocular tissues and fluids might be low. This may bear future implications for patient management in ophthalmological practice, surgery and transplantation.

[First results of investigations of SARS-CoV­2 RNA in human corneal tissue]. / Erste Ergebnisse zu Untersuchungen der menschlichen Hornhaut auf SARS-CoV­2-RNA.

Preliminary investigations of human corneal tissues from coronavirus disease 2019 (COVID-19) cadaveric donors indicated that no severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) RNA is present. Current eye banking guidelines do not recommend any type of routine testing for SARS-CoV­2 RNA in post-mortem donor tissue. This is partly based on factors that can influence the test results of the reverse transcription polymerase chain reaction (RT-PCR).

Histopathology of a retrocorneal membrane after Descemet membrane endothelial keratoplasty: a case report.

BACKGROUND: We report the first histopathologically proven occurrence of a retrocorneal membrane after Descemet's membrane endothelial keratoplasty. CASE PRESENTATION: A white Caucasian 76-year-old woman received penetrating keratoplasty on her right eye 2 years after Descemet's membrane endothelial keratoplasty surgery with combined cataract extraction and intraocular lens implantation for Fuchs' endothelial corneal dystrophy due to an allograft rejection with ensuing graft failure. Her preoperative vision was counting fingers (20/2000) caused by immunological debris, corneal edema, and secondary membrane formation. Her postoperative vision at 3 months was 20/125. The histopathological evaluation showed a membranous structure overlying the denuded Descemet membrane. CONCLUSIONS: We report a case of a histopathologically proven retrocorneal membrane after Descemet's membrane endothelial keratoplasty surgery.

Long-term Follow-up of Patients with Tubulointerstitial Nephritis and Uveitis (TINU) Syndrome.

PURPOSE: To evaluate the response to treatment in patients with tubulointerstitial nephritis and uveitis (TINU) syndrome over a long-term follow-up period. METHODS: Nine patients with TINU syndrome were retrospectively reviewed. The mean follow-up was 54.8 months (range: 24-133 months). RESULTS: The mean number of recurrences per year declined from 1.7 in the 1st year to 0.66 in the 2nd year of treatment. The ocular inflammation responded to local corticosteroids in two patients, systemic corticosteroids in two patients, immunosuppressive therapy in four patients, and anti-TNF-α blocking agent in one patient. The therapy could be discontinued in six (67%) patients after a mean treatment period of 29.5 months. In five patients, remission with the recurrence-free period of 12.8 months was achieved. CONCLUSIONS: TINU syndrome was characterized by limited responsiveness to corticosteroid therapy and less by severe complications. A long-term course of immunosuppressants or biologics was necessary to control the uveitis and led to induction of remission.

Descemet membrane endothelial keratoplasty in vitrectomized eyes: clinical results.

PURPOSE: The aim of this study was to evaluate clinical outcomes of Descemet membrane endothelial keratoplasty (DMEK) in vitrectomized eyes. METHODS: Medical records were reviewed for demographics, baseline and follow-up best spectacle-corrected visual acuities (BCVAs), endothelial cell densities, and indication for DMEK. Operative notes were analyzed to identify any specific intraoperative event. RESULTS: A total of 281 DMEK cases were reviewed. Twenty cases were considered eligible for the study. Seven eyes had a history of anterior vitrectomy, and 13 eyes had a history of complete removal of the vitreous body. Ages ranged from 37 to 78 years (mean, 62.9 years) and mean follow-up after DMEK was 11.2 months (range, 3-29 months). The preoperative mean BCVA was 1.4 ± 0.5 logarithm of minimum angle of resolution (logMAR) and increased to 1.0 ± 0.5 logMAR after 4 weeks (P = 0.0290). After 6-month and 12-month follow-up, BCVA was 0.8 ± 0.6 logMAR (P = 0.0055) and 0.6 ± 0.3 logMAR (P = 0.0001), respectively. All cases had ocular comorbidities affecting the BCVA. In 13 cases, significant intraoperative complications were experienced. In the immediate postoperative period, iatrogenic primary graft failure was reported in 2 eyes (2/20). In 4 eyes, late graft failure was observed. Two eyes had exacerbation of preexisting glaucoma. Mean preoperative endothelial cell density was 2301 ± 159 cells per square millimeter; postoperative follow-up visits at 6 and 12 months showed a decrease to 1398 ± 161 cells per square millimeter and 1241 ± 155 cells per square millimeter, respectively. CONCLUSIONS: DMEK seems to be successful in restoring visual acuity even in vitrectomized eyes. Nevertheless, the overall complication rate was higher than that in standard DMEK.

Cytotoxic properties of sunitinib and sorafenib on human corneal epithelial cells.

PURPOSE: To generate toxicology profiles of individual drugs on human corneal epithelial cells (HCEC) and compare their in vitro cytotoxicity. METHODS: Monolayer cultures of HCEC were harvested from two human donor eyes. Sunitinib (0.3-10 µg/mL) and Sorafenib (0.3-100 µg/mL), diluted in culture medium (CnT-BM.1, CELLnTEC Advanced Cell Systems AG, Bern, Switzerland), 1% Penicillin and 1% Streptomycin were added to cells that were being grown in cell culture dishes. The 3-(4,5-dimethylthiazol-2-yl)-2,5-diphenyltetrazolium bromide (MTT) was performed 24 hours, three days and five days after incubation. Live/dead viability/cytotoxicity assay (Live/dead assay) was performed and analyzed using fluorescence microscopy after 24 hours of incubation. The expression of p63, ABCG2 and PDGFRß was evaluated by immunocytochemistry prior to exposure. Cell morphology was assessed with a phase contrast microscope after 24 hours of exposure. RESULTS: Significant toxicity of Sunitinib was seen at concentrations of >3.3 µg/mL and of Sorafenib at concentrations of >1.0 µg/mL after 24 hours of incubation. Both drugs exhibited increasing toxicities over time. HCEC stained positively for p63, ABCG2 and PDGFRß. In comparison, the IC50 (inhibitory concentration 50) of Sorafenib was 2.26 times the IC50 of Sunitinib using Live/dead assay after 24 hours and 2.39, 1.29 and 0.78 times the IC50 of Sunitinib using the MTT test after 24 hours, three days and five days, respectively. CONCLUSIONS: These in vitro experimental findings support the safety of Sunitinib and Sorafenib on HCEC when used at a concentration of <3.3 µg/mL and <1.0 µg/mL, respectively, after 24 hours of exposure. The in vitro cytotoxicity of Sorafenib on HCEC was higher than Sunitinib.

Novel maneuver facilitating Descemet membrane unfolding in the anterior chamber.

PURPOSE: To describe a new technique for unfolding of the Descemet membrane (DM) during Descemet membrane endothelial keratoplasty. METHODS: DM and endothelium were stripped from donor corneas submerged in corneal storage solution. Donor DM diameters were 8.5 mm. The central 8 mm of DM was stripped from the recipient cornea. After staining with trypan blue to improve visualization, donor DM was inserted through a 2.75-mm incision. Unfolding was achieved with simultaneous digital pressure in the equatorial region and tapping of the corneal surface. This technique was performed in Descemet membrane endothelial keratoplasty patients with Fuchs endothelial dystrophy (n = 6) and bullous keratopathy (n = 4). RESULTS: In all cases, a fast attachment of the DM was achieved. The recurling was successfully hindered because of the simultaneous equatorial pressurization and induced fluid currents inside the anterior chamber by appropriate corneal tapping. In all cases, a rapid corneal clearance was achieved with significantly increased visual acuity (logarithm of the minimal angle of resolution visual acuity preoperative = 0.90, logarithm of the minimal angle of resolution visual acuity final assessment = 0.19, P < 0.001; follow-up 14 ± 5 weeks). CONCLUSION: Simultaneous digital pressure with corneal tapping is a helpful maneuver to unfold the DM inside the anterior chamber. This technique may reduce the need for unnecessary instrumental and noninstrumental manipulation of the graft.