Alteration of Gene Expression in Pathological Keratinization of the Ocular Surface.

Purpose: To investigate the molecular mechanism of pathological keratinization in the chronic phase of ocular surface (OS) diseases. Methods: In this study, a comprehensive gene expression analysis was performed using oligonucleotide microarrays on OS epithelial cells obtained from three patients with pathological keratinization (Stevens-Johnson syndrome [n = 1 patient], ocular cicatricial pemphigoid [n = 1 patient], and anterior staphyloma [n = 1 patient]). The controls were three patients with conjunctivochalasis. The expression in some transcripts was confirmed using quantitative real-time PCR. Results: Compared to the controls, 3118 genes were significantly upregulated by a factor of 2 or more than one-half in the pathological keratinized epithelial cells (analysis of variance P < 0.05). Genes involved in keratinization, lipid metabolism, and oxidoreductase were upregulated, while genes involved in cellular response, as well as known transcription factors (TFs), were downregulated. Those genes were further analyzed with respect to TFs and retinoic acid (RA) through gene ontology analysis and known reports. The expression of TFs MYBL2, FOXM1, and SREBF2, was upregulated, and the TF ELF3 was significantly downregulated. The expression of AKR1B15, RDH12, and CRABP2 (i.e., genes related to RA, which is known to suppress keratinization) was increased more than twentyfold, whereas the expression of genes RARB and RARRES3 was decreased by 1/50. CRABP2, RARB, and RARRES3 expression changes were also confirmed by qRT-PCR. Conclusions: In pathological keratinized ocular surfaces, common transcript changes, including abnormalities in vitamin A metabolism, are involved in the mechanism of pathological keratinization.

Senescent characteristics of human corneal endothelial cells upon ultraviolet-A exposure.

PURPOSE: The objective of this study was to investigate the senescent phenotypes of human corneal endothelial cells (hCEnCs) upon treatment with ultraviolet (UV)-A. METHODS: We assessed cell morphology, senescence-associated ß-galactosidase (SA-ß-gal) activity, cell proliferation and expression of senescence markers (p16 and p21) in hCEnCs exposed to UV-A radiation, and senescent hCEnCs induced by ionizing radiation (IR) were used as positive controls. We performed RNA sequencing and proteomics analyses to compare gene and protein expression profiles between UV-A- and IR-induced senescent hCEnCs, and we also compared the results to non-senescent hCEnCs. RESULTS: Cells exposed to 5 J/cm2 of UV-A or to IR exhibited typical senescent phenotypes, including enlargement, increased SA-ß-gal activity, decreased cell proliferation and elevated expression of p16 and p21. RNA-Seq analysis revealed that 83.9% of the genes significantly upregulated and 82.6% of the genes significantly downregulated in UV-A-induced senescent hCEnCs overlapped with the genes regulated in IR-induced senescent hCEnCs. Proteomics also revealed that 93.8% of the proteins significantly upregulated in UV-A-induced senescent hCEnCs overlapped with those induced by IR. In proteomics analyses, senescent hCEnCs induced by UV-A exhibited elevated expression levels of several factors part of the senescence-associated secretory phenotype. CONCLUSIONS: In this study, where senescence was induced by UV-A, a more physiological stress for hCEnCs compared to IR, we determined that UV-A modulated the expression of many genes and proteins typically altered upon IR treatment, a more conventional method of senescence induction, even though UV-A also modulated specific pathways unrelated to IR.

Potential marker subset of blood-circulating cytokines on hematopoietic progenitor-to-Th1 pathway in COVID-19.

In this study, we analyzed a relatively large subset of proteins, including 109 kinds of blood-circulating cytokines, and precisely described a cytokine storm in the expression level and the range of fluctuations during hospitalization for COVID-19. Of the proteins analyzed in COVID-19, approximately 70% were detected with Bonferroni-corrected significant differences in comparison with disease severity, clinical outcome, long-term hospitalization, and disease progression and recovery. Specifically, IP-10, sTNF-R1, sTNF-R2, sCD30, sCD163, HGF, SCYB16, IL-16, MIG, SDF-1, and fractalkine were found to be major components of the COVID-19 cytokine storm. Moreover, the 11 cytokines (i.e., SDF-1, SCYB16, sCD30, IL-11, IL-18, IL-8, IFN-γ, TNF-α, sTNF-R2, M-CSF, and I-309) were associated with the infection, mortality, disease progression and recovery, and long-term hospitalization. Increased expression of these cytokines could be explained in sequential pathways from hematopoietic progenitor cell differentiation to Th1-derived hyperinflammation in COVID-19, which might also develop a novel strategy for COVID-19 therapy with recombinant interleukins and anti-chemokine drugs.

A new surgical approach for the treatment of a refractory foveal microaneurysm: A case report.

Purpose: To report a case of a refractory foveal microaneurysm (MA) that was successfully treated by use of a new surgical procedure. Observations: This study involved a 79-year-old female with an active foveal MA associated with branch retinal vein occlusion in her left eye. Despite anti-vascular endothelial growth factor treatments, the MA remained active without closure, and best-corrected visual acuity (VA) gradually decreased from 20/20 to 20/200. After our new surgical procedure was explained in detail to the patient, written informed consent was obtained from the patient and the surgery was performed. Briefly, following pars plana vitrectomy, the internal limiting membrane in her left eye was peeled and the retina of the external wall of the MA was then gently incised. The exposed MA was then directly grabbed and pulled up onto the retina using 27-gauge microforceps, and photocoagulation was performed. At 3-months postoperative, closure of the MA and improvement in the retinal findings were observed, and best-corrected VA improved to 20/67. Conclusions and importance: We report a case of a refractory foveal MA that was successfully treated with a novel surgical technique that closed the MA, avoided thermal damage to the surrounding tissue, and resulted in improved postoperative VA.

Donor Corneal Endothelial Cell Maturity and Its Impact on Graft Survival in Glaucoma Patients Undergoing Corneal Transplantation.

PURPOSE: To examine corneal graft survival via corneal endothelial cell density (ECD) and corneal endothelial cell loss (ECL) at 5 years post-transplantation in the eyes of patients with and without a history of undergoing glaucoma surgery according to the maturity of the donor corneal endothelial cells. DESIGN: Prospective cohort study. METHODS: This prospective cohort study included 17 patients with glaucoma and 51 patients without glaucoma who underwent Descemet's stripping automated endothelial keratoplasty or penetrating keratoplasty at the Baptist Eye Institute, Kyoto, Japan, between October 2014 and October 2016. Human corneal endothelial cells were cultured from residual peripheral donor cornea tissue, and the maturity of the cells was evaluated by cell surface markers (ie, CD166+, CD44-/dull, CD24-, and CD105-) using fluorescence-activated cell sorting. Kaplan-Meier analysis or the chi-square test was used to assess the rate of successful corneal graft survival post-transplantation. RESULTS: At 36 months postoperatively, the mean ECD and ECL in the glaucoma-bleb eyes were 1197 ± 352 cells/mm2 and 55.5% ± 13.9% in the high-maturity group and 853 ± 430 cells/mm2 and 67.7% ± 18.1% in the low-maturity group, respectively. Kaplan-Meier analysis revealed that at 5 years postoperatively, the overall rate of survival was 45%, that is, 100% in the high-maturity group and 25% in the low-maturity group (P < .05). CONCLUSIONS: The findings in this prospective cohort study revealed that the use of donor corneal grafts containing mature-differentiated corneal endothelial cells could maintain the survival of the transplanted graft for a long-term period, even in patients with a history of undergoing glaucoma surgery.

The existence of senescent cells in conjunctival epithelium from elderly individuals.

PURPOSE: The ocular surface microenvironment changes with aging. However, it remains unclear if cellular senescence influences the ocular surface. We investigated the presence of p16INK4a-expressing senescent cells in healthy human conjunctiva. STUDY DESIGN: Clinical and experimental. METHODS: Healthy conjunctival tissue samples were obtained from middle-aged and elderly subjects. RT-qPCR was performed to assess the expression of senescence markers CDKN2A (p16INK4a) and CDKN1A (p21CIP1/WAF1) and immunostaining was performed to examine the expression of the senescence marker p16INK4a, stem cell markers Ki67 and p63, tight-junction marker ZO-1. RESULTS: Our study involved 19 conjunctival tissue samples (10 elderly and 9 middle-aged), mean age [elderly: 75.8 ± 3.7 years (72-81), middle-aged: 52.7 ± 7 years (38-59)], sex (elderly: 3 men, 7 women; middle-aged: 3 men, 6 women). The expression of p16INK4a was significantly increased at the RNA level in the elderly compared to middle-aged (p < 0.05). Positivity rate of p16INK4a was significantly elevated in the elderly (15.0 ± 7.8%) compared to middle-aged (0.2 ± 0.6%) (p < 0.05). Positivity rate of Ki67and p63 was significantly reduced in the elderly (1.7 ± 1.7% and 16.5 ± 9.5%) compared to middle-aged (3.9 ± 1.8% and 24.7 ± 5.7%) (p < 0.05). ZO-1 expression was reduced in tissue samples showing p16INK4a-positivity but retained in tissue samples in which p16INK4a was undetectable. CONCLUSIONS: Senescent cells accumulate with age in the conjunctival epithelium, accompanied by a decrease in Ki67, p63 and ZO-1 expressing cells.

Dematiaceous fungal keratitis caused by Cladophialophora boppii - A case report.

Purpose: To report a rare case of dematiaceous fungal keratitis caused by Cladophialophora boppii (C. boppii) in an immunocompromised patient. Observations: An 83-year-old male with chronic renal failure was referred to the Department of Ophthalmology, Kyoto Prefectural University of Medicine, Kyoto, Japan due to persistent corneal epithelial defects (PEDs) in his left eye. Initial examination revealed decreased central corneal sensitivity and decreased tear secretion in that eye, both thought to be associated with herpetic keratitis. Permanent punctal-plug surgery combined with therapeutic soft contact lens wear was performed to treat the PED, which initially healed, yet recurred. Follow-up examination revealed a 1.0-mm-diameter black lesion consistent with the PED site, which subsequently increased in size, so treatment with miconazole solution eye drops, natamycin ophthalmic ointment, and systemic itraconazole was initially performed. Since the region of the lesion had progressed to corneal perforation, corneal transplantation surgery under general anesthesia was scheduled, yet the patient refused to undergo surgery. Mycological testing via DNA sequencing of the internal transcribed spacer of ribosomal DNA regions revealed that the isolate or pathogen was C. boppii. Mycotic keratitis caused by C. boppii was found to be resistant to antifungal drugs. Conclusion and importance: This is a rare case of fungal keratitis caused by C. boppii in an elderly immunocompromised patient.

Identification and Analysis of Primary Cilia in the Corneal Endothelial Cells of Patients with Bullous Keratopathy.

PURPOSE: To identify primary cilia in human corneal endothelial cells (CECs) obtained from patients with bullous keratopathy (BK). METHODS: This study involved CEC specimens obtained from 10 eyes of 10 consecutive patients (three males and seven females; mean age: 74.5 years, range: 68-90 years) with BK who underwent Descemet's stripping automated endothelial keratoplasty at Baptist Eye Institute, Kyoto, Japan between August 2019 and September 2020. Three corneal buttons obtained from 3 patients who underwent penetrating keratoplasty for keratoconus were used as 'non-BK' controls. All specimens were evaluated with immunofluorescence staining using an antibody against acetylated α-tubulin. RESULTS: Ciliary expression was observed in six of the 10 CEC specimens; i.e. in two specimens obtained from BK patients after glaucoma surgery (trabeculectomy), in two specimens obtained from patients with Fuchs endothelial corneal dystrophy, and in two specimens obtained from a patient with BK after laser iridotomy for primary angle closure. There was acetylated α-tubulin staining but no hair-like structures in two specimens, and ciliary expression was unknown in two specimens due to the absence of cells. The length of the primary cilia varied between all specimens. In contrast, no primary cilia were observed in the corneal buttons obtained from the three keratoconus patients. CONCLUSION: The findings in this study clearly demonstrate the expression of primary cilia in the CECs of patients afflicted with BK.

Ocular cicatricial pemphigoid following Dipeptidyl Peptidase-4 inhibitor use: A case report.

Purpose: To report a rare Ocular Cicatricial Pemphigoid (OCP) case in a patient taking a Dipeptidyl Peptidase-4 Inhibitor (DPP-4 inhibitor), a medication used for the management of type 2 diabetes, for at least six years. Observations: A 64-year-old male presented with refractory bilateral conjunctival inflammation and ocular discharge that had persisted for two months, despite multiple prior therapies for presumed bacterial conjunctivitis. Upon initial examination, clinical findings strongly suggested OCP, and he had elevated levels of anti-BP180 antibodies. Despite receiving systemic treatments such as steroid pulse therapy and therapeutic plasma exchange after discontinuing DPP-4 inhibitors, his condition progressively worsened, with manifestations such as forniceal shortening in his left eye. Consequently, the patient required keratoepithelioplasty, amniotic membrane transplantation in his left eye, and bilateral eyelid entropion surgery. His condition initially worsened for a time after discontinuing the DPP-4 inhibitor, but it gradually improved over time, and ocular surface surgical intervention was not required in the right eye. Conclusions and Importance: The findings in this study demonstrate that severe refractory OCP may occur while taking the DPP-4 inhibitor, thus indicating that a detailed interview regarding medications is essential for patients with ocular pemphigoid, especially those with type 2 diabetes.

Strategic combination of cultivated oral mucosal epithelial transplantation and postoperative limbal-rigid contact lens-wear for end-stage ocular surface disease: a retrospective cohort study.

PURPOSE: To provide the long-term outcome of patients with end-stage severe ocular surface disease (OSD) consecutively treated with cultivated oral mucosal epithelial transplantation (COMET) followed by limbal-rigid contact lens (CL)-wear therapy. DESIGN: Retrospective cohort. METHODS: In 23 eyes of 18 patients with severe OSD who underwent COMET surgery between 2002 and 2019 and who were followed with limbal-rigid CL-wear therapy for at least 1 year postoperative, patient demographics, best-corrected visual acuity (BCVA, logMAR), Ocular Surface Grading Scores (OSGS), surgical indication and adverse events were reviewed. Primary and secondary outcomes were BCVA and OSGS changes at baseline and final examination, respectively. RESULTS: This study involved 16 patients with Stevens-Johnson syndrome and 2 patients with mucous membrane pemphigoid (mean age: 59±15 years). The indications for COMET were as follows: corneal reconstruction for vision improvement (10 eyes (43.5%)), corneal reconstruction for persistent epithelial defect (4 eyes (17.4%)) and conjunctival (fornix) reconstruction for symblepharon release (9 eyes (39.1%)). The mean duration of CL-wear postsurgery was 6.4±3.9 years (range: 1.4 to 13.3 years). The mean BCVA at baseline and at final follow-up was logMAR 1.9±0.5 and 1.3±0.7, respectively (p<0.05). Compared with those at baseline, the OSGSs for symblepharon and upper and lower fornix shortening showed significant improvement at each follow-up time point post treatment initiation. No serious intraoperative or postoperative adverse events were observed. CONCLUSION: In patients afflicted with severe OSD, COMET combined with limbal-rigid CL-wear therapy postsurgery was found effective for vision improvement and ocular surface stabilisation.

Gene expression signatures of human senescent corneal and conjunctival epithelial cells.

PURPOSE: This study aimed to investigate the senescent phenotypes of human corneal and conjunctival epithelial cells. METHODS: We examined cell morphology, senescence-associated ß-galactosidase (SA-ß-gal) activity, cell proliferation, and expression of senescence markers (p16 and p21). RNA sequencing analysis was conducted to compare gene expression profiles between senescent and non-senescent cells. Finally, the potential involvement of senescent cells in the pathogenesis of ocular surface diseases was investigated. RESULTS: X-irradiated corneal and conjunctival epithelial cells exhibited typical senescence phenotypes, i.e., flattened morphologies, increased SA-ß-gal activity, decreased cell proliferation, and increased expression of senescence markers, p16 and p21. RNA-seq analysis revealed substantial differences in gene expression profiles between senescent corneal (SCo) and conjunctival epithelial cells (SCj). Moreover, SCj were detected in pathological conjunctival tissues associated with limbal stem cell deficiency (LSCD) due to Stevens-Johnson syndrome or chemical burns, potentially being involved in abnormal differentiation. CONCLUSION: This study highlights the cellular and molecular characteristics of senescent ocular surface cells, particularly in SCj that show abnormal keratin expression, and their potential roles in severe ocular surface diseases and pathology.

Development and Validation of a Novel Score to Predict Mortality in Stevens-Johnson Syndrome and Toxic Epidermal Necrolysis: CRISTEN.

BACKGROUND: Stevens-Johnson syndrome (SJS) and toxic epidermal necrolysis (TEN) are life-threatening, severe mucocutaneous adverse reactions. Severity prediction at early onset is urgently required for treatment. However, previous prediction scores have been based on data of blood tests. OBJECTIVE: This study aimed to present a novel score that predicts mortality in patients with SJS/TEN in the early stages based on only clinical information. METHODS: We retrospectively evaluated 382 patients with SJS/TEN in a development study. A clinical risk score for TEN (CRISTEN) was created according to the association of potential risk factors with death. We calculated the sum of these risk factors using CRISTEN, and this was validated in a multinational survey of 416 patients and was compared with previous scoring systems. RESULTS: The significant risk factors for death in SJS/TEN comprised 10 items, including patients' age of ≥65 years, ≥10% body surface area involvement, the use of antibiotics as culprit drugs, the use of systemic corticosteroid therapy before the onset, and mucosal damage affecting the ocular, buccal, and genital mucosa. Renal impairment, diabetes, cardiovascular disease, malignant neoplasm, and bacterial infection were included as underlying diseases. The CRISTEN model showed good discrimination (area under the curve [AUC] = 0.884) and calibration. In the validation study, the AUC was 0.827, which was statistically comparable to those of previous systems. CONCLUSION: A scoring system based on only clinical information was developed to predict mortality in SJS/TEN and was validated in an independent multinational study. CRISTEN may predict individual survival probabilities and direct the management and therapy of patients with SJS/TEN.

Spatiotemporal Coordination of RPE Cell Quality by Extracellular Vesicle miR-494-3p Via Competitive Interplays With SIRT3 or PTEN.

Purpose: To reveal the molecular mechanism underlying degeneration in human retinal pigment epithelial (hRPE) cells with dysfunctional mitochondrial homeostasis. Methods: The expression of recently identified miR-494-3p in extracellular vesicles (EV) released from induced-pluripotential-stem-cell-derived human RPE (iPS-hRPE), during coculture with macrophages (Mps) was investigated in iPS-hRPE and ARPE cells differentiated in the presence of nicotinamide (Nic-ARPE). The expression of phosphatase and tensin homolog (PTEN), sirtuin3 (SIRT3), and mitochondrial marker proteins before and after the transfection of miR-494-3p inhibitor and mimic, and the changes in mitochondrial metabolism, membrane potential, and oxidative phosphorylation (OXPHOS) were monitored. Results: Compared with senescent dedifferentiated ARPE19 cells, iPS-hRPE and Nic-ARPE cells expressed elevated levels of mitochondrial marker proteins but a repressed cellular miR-494-3p level. The expression of target proteins of miR-494-3p, PTEN, and SIRT3 was upregulated along with the differentiation disposition of these RPE cells. The ratio of PTEN/SIRT3 in de-differentiated ARPE19 cells was surprisingly elevated by around 20 times compared with that in iPS-hRPE and Nic-ARPE cells. The novel molecular interplay of EV miR-494-3p either with mitochondria selective SIRT3 or organelle nonselective PTEN was found to participate in the degeneration of hRPE cells by inducing mitochondrial dysfunctions and repressed OXPHOS, mitochondrial membrane potential, and ATP and NAD+ production. Conclusions: Our results demonstrate a clear causal link between miR-494-3p and hRPE cell degeneration via the regulation of mitochondrial integrity. EV miR-494-3p may play a pivotal role in pathogenic spreading of degenerated hRPE cells from the local perifovea throughout the macula.

Differences in gene regulation by TLR3 and IPS-1 signaling in murine corneal epithelial cells.

Toll-like receptor 3 (TLR3) and interferon-beta promoter stimulator-1 (IPS-1) are associated with antiviral responses to double-stranded RNA viruses and contribute to innate immunity. We previously reported that conjunctival epithelial cell (CEC) TLR3 and IPS-1 pathways respond to the common ligand polyinosinic:polycytidylic acid (polyI:C) to regulate different gene expression patterns as well as CD11c + cell migration in murine-model corneas. However, the differences in the functions and the roles of TLR3 and IPS-1 remain unclear. In this study, we investigated the differences of TLR3 or IPS-1-induced gene expression in corneal epithelial cells (CECs) in response to polyI:C stimulation using cultured murine primary CECs (mPCECs) derived from TLR3 and IPS-1 knockout mice via comprehensive analysis. The genes associated with viral responses were upregulated in the wild-type mice mPCECs after polyI:C stimulation. Among these genes, Neurl3, Irg1, and LIPG were dominantly regulated by TLR3, while interleukin (IL)-6 and IL-15 were dominantly regulated by IPS-1. CCL5, CXCL10, OAS2, Slfn4, TRIM30α, and Gbp9 were complementarily regulated by both TLR3 and IPS-1. Our findings suggest that CECs may contribute to immune responses and that TLR3 and IPS-1 possibly have different functions in the corneal innate immune response.

Corneal Electrolysis for Granular Corneal Dystrophy Type 2 (Avellino Corneal Dystrophy) Exacerbation After LASIK.

PURPOSE: To report two cases in which exacerbation of granular corneal dystrophy type 2 (GCD2; Avellino corneal dystrophy) after laser in situ keratomileusis (LASIK) was successfully removed by corneal electrolysis. METHODS: This study involved a 66-year-old man and a 43-year-old man with GCD2 who had undergone bilateral LASIK for myopia 10 or more years prior to presentation. In both patients, GCD2 corneal opacity gradually developed postoperatively at the LASIK flap interface, thus resulting in a decrease of visual acuity. For treatment, the LASIK flaps in both patients were surgically lifted to directly remove the opacity. Corneal electrolysis was then applied to the back of each LASIK flap and stromal bed. RESULTS: Postoperatively, the ocular symptoms and corneal opacities related to GCD exacerbation disappeared, with improvement of corrected and uncorrected distance visual acuity and almost no change of refractive error. CONCLUSIONS: The findings reveal that corneal electrolysis is safe and effective for treating exacerbations of GCD2 following LASIK when applied to a surgically lifted flap, and that it successfully removes GCD2-related LASIK flap interface opacities with almost no change of refractive error postoperatively. [J Refract Surg. 2023;39(1):61-65.].

Strengthening of the barrier function in human telomerase reverse transcription (hTERT) immortalized corneal and conjunctival epithelium by double-stranded RNA.

To investigate the response to polyinosinic:polycytidylic acid [poly(I:C)], a double-stranded RNA Toll-like receptor 3 agonist that mimics viral infection, in the barrier function of two established human telomerase reverse transcriptase-immortalized cell lines, termed HCLE for the human corneal-limbal epithelial line and HCjE for the human conjunctival-epithelial line. In this study, HCLE and HCjE cells were used to evaluate the underlying mechanism of epithelial-cell barrier function regulation. Briefly, HCLE and HCjE cells were first cultured on 12-well Transwell® (Corning®) filter-plates, and reverse transcription-polymerase chain reaction, western blotting, and immunohistochemical examinations were then performed to assess tight junction (TJ)-related protein expression and cellular distribution. Next, the barrier function of the cells was measured via transepithelial electrical resistance (TEER) and paracellular molecular flux. The cells were then stimulated with poly(I:C) and the TEER and TJ-related protein expressions were analyzed. Similar to that in in vivo epithelium, the expression of claudin (CLDN) subtypes CLDN-1, -4, and -7 was observed in the HCLE and HCjE cells, and the barrier function in the HCLE cells was tighter than that in the HCjE cells. Post stimulation with poly(I:C), TEER of the HCLE and HCjE cells increased in a dose- and time-dependent manner, the production of TJ-related protein mRNA and CLDN-4 protein were elevated, and the barrier function of the HCLE and HCjE cells increased, thus possibly indicating that the increased barrier function is a defense mechanism against viral infection.

Watchful waiting is an acceptable treatment option for asymptomatic primary ocular adnexal mucosa-associated lymphoid tissue lymphoma: A retrospective study.

BACKGROUND: Primary ocular adnexal mucosa-associated lymphoid tissue (MALT) lymphoma (POAML) is the most common subtype of indolent ocular adnexal lymphomas. Although radiotherapy (RT) is the standard of care for localized POAML, it can occasionally lead to permanent side effects. Other treatment strategies, such as rituximab (R) monotherapy and immunochemotherapy, have been used for POAML treatment, but their long-term benefits and relative merits remain unclear. While watchful waiting (WW) is a potential option for some indolent lymphomas, the benefits of WW for POAML patients are also unclear. METHODS: We here retrospectively analyzed 75 patients who were diagnosed with POAML between 2008 and 2019 in the institutions of the Kyoto Clinical Hematology Study Group. RESULTS: Commonly involved sites were conjunctiva (42.7%), orbit (36.0%), and lacrimal gland (12.0%), and most patients (92.0%) presented with Ann Arbor stage IE disease. The treatment strategy was selected at the physicians' discretion. More patients without subjective symptoms by tumor mass were subjected to WW (29 patients), while more patients with tumor-derived subjective symptoms were treated by tumor-directed therapy (24 received focal RT, and 19 received R monotherapy). Complete response rates were 79.2% and 42.1% in the RT and R groups, respectively. At 60 months of follow-up, the estimated proportions of POAML patients not requiring new treatment were 69.4%, 85.2%, and 53.8% in the WW, RT, and R groups, respectively. There were no significant differences in the time to start a new treatment between WW and RT groups (median: both not reached [NR], p = 0.187) and between WW and R groups (median: NR vs. 69.0 months, p = 0.554). No specific predictive factor for the future need of treatment was identified in the WW group. CONCLUSION: Our results demonstrate WW may be an acceptable treatment option for POAML, especially for asymptomatic patients.

Socket expansion with conformers in congenital anophthalmia and microphthalmia.

PURPOSE: To report the outcomes of acrylic conformer-assisted socket expansion in congenital anophthalmia and microphthalmia. METHODS: In this noncomparative, interventional case series, the medical records of 24 eyes of 18 consecutive patients with congenital anophthalmia (n = 3), clinical anophthalmia (n = 8), and microphthalmia (n = 13) were reviewed retrospectively. Twelve cases were unilateral; 6 were cases bilateral (3 clinical anophthalmia and 3 microphthalmia). Serial socket expansion with progressively larger acrylic conformers was managed in clinic. Horizontal palpebral fissure (HPF) width was graded as good (final HPF ≥20 mm, or interocular difference ≤2 mm for unilateral cases), fair (17-19 mm, or 3 mm interocular difference), or poor (<17 mm, or ≥4 mm difference). RESULTS: The mean initial lid lengths in anophthalmia, clinical anophthalmia, and microphthalmia were 11.0, 12.4, and 16.9, increasing to 21.0, 19.9, and 22.2, respectively, over a mean period of 51 months. Mean age at the initiation of treatment was 19 months (range, 1-78). Percentage increases in lid length were 90.9%, 61.2%, and 31.3% in anophthalmia, clinical anophthalmia, and microphthalmia, respectively, with an average 7 conformer exchanges. For unilateral cases, the mean final lid length of involved eyes was 22.3 mm, comparable to 23.5 mm in normal contralateral eyes. Good outcomes were achieved in 18 orbits (75%); fair outcomes, in 6 (25%) cases. None of the sockets had poor expansion at final follow-up. All cases had good cosmesis with acceptable prosthesis wear at last visit. CONCLUSIONS: In our patient cohort, good socket expansion was achieved with acrylic conformers alone in congenital anophthalmia and microphthalmia, with acceptable prosthesis wear.

Efficient DNA/RNA extraction from tarsal plates by SK mill, a freeze-crush apparatus.

The tarsal plate is an eyelid tissue that maintains lid structure from inside the upper/lower eyelids, and it surrounds the meibomian glands and supports their unique secretion mechanism. Sebaceous carcinoma, a malignant eyelid tumour, can sometimes develop from the meibomian glands and is usually excised together with the tarsal plate during surgery, so the tarsal plate serves as a control research tissue. However, since the plate is thick, hard and heterogeneous with few cells, obtaining enough genomic DNA and/or total RNA is often difficult. Therefore, we attempted to establish an efficient protocol to obtain DNA and RNA simultaneously by comparing the combinations of homogenization (mortar/pestle, pellet pestle or SK mill) and purification (organic solvent or spin column) methods using rabbit tarsal plates. Based on the yield, quality and hands-on time, the SK mill and spin column was found to be the most efficient combination. We then applied the established protocol to extract DNA/RNA from six human tarsal-plate samples and succeeded in generating high-quality exome and transcriptome datasets via a next-generation sequencer with sufficient coverage and meibomian gland-specific expression of representative genes, respectively. Our new findings will provide ideal reference data for future genetic and gene-expression studies of sebaceous carcinoma.

Limbal-Rigid Contact Lens Wear for the Treatment of Ocular Surface Disorders: A Review.

ABSTRACT: Recently, the prescription of large-diameter rigid gas-permeable contact lenses (CLs), also known as "scleral lenses," "corneoscleral lenses," and "limbal-rigid CLs," is on the rise for the treatment of both moderate and severe ocular surface disorders (OSDs). Compared with scleral lenses, the diameter of limbal-rigid CLs is generally smaller, that is, a diameter ranging from 13.0 to 14.0 mm, and they are designed so that the peripheral edge bears on the limbus. The Suncon Kyoto-CS (Sun Contact Lens Co., Ltd.) is a novel limbal-rigid CL design with multistep curves on the peripheral edge for easy tear exchange during blinking that removes debris and prevents lens clouding or fogging, thus allowing patients to enjoy a longer daily duration of CL wear. In severe OSD cases, limbal-rigid CL wear after surgery is a noninvasive therapeutic approach that can neutralize corneal irregularities, decrease dry eye symptoms, prevent the progression or recurrence of symblepharon, and improve the patient's visual acuity and overall quality of life. Thus, surgeries such as amniotic membrane transplantation and cultivated oral mucosal epithelial transplantation, as well as limbal-rigid CL wear, which is noninvasive, are valuable and effective treatment strategies that can now be applied for the management of patients afflicted with severe OSDs.