The hiPSC-derived corneal endothelial progenitor-like cell recovers the rabbit model of corneal endothelial dystrophy.

INTRODUCTION: Corneal endothelial dysfunction results in cornea opacity, damaging sightedness, and affecting quality of life. A corneal transplant is the current effective intervention. Due to the scarcity of donated cornea, such an unmet medical need requires a novel therapeutic modality. OBJECTIVES: Customizing patients' corneal endothelial progenitor cells with proliferative activity and lineage restriction properties shall offer sufficient therapeutic cells for corneal endothelial dystrophy. METHODS: The customized induced human corneal endothelial progenitor-like cell (iHCEPLC) was obtained through cell fate conversions starting from PBMC (peripheral blood mononuclear cell), hiPSC (human induced pluripotent stem cell), and hNCC (human neural crest cell), while it finally reached the iHCEPLC state via a series of induction. Several molecular diagnoses were applied to depict its progenitor state, including RNAseq, FlowCytometer, immunostainings, and rtPCR. Significantly, it can be induced to gain differentiation maturity through contact inhibition. In addition, a BAK-mediated rabbit model of corneal endothelial dystrophy was established in the present study to test the therapeutic effectiveness of the iHCEPLC. RESULTS: After inducing cell fate conversion, the specific HCEC markers were detected by rtPCR and immunostaining in iHCEPLC. Further, RNAseq was applied to distinguish its progenitor-like cell fate from primary human corneal endothelial cells (HECE). FlowCytometry profiled the heterogeneity subpopulation, consistently displaying a subtle difference from primary HCEC. A terminal differentiation can be induced in iHCEPLC, addressing its progenitor-like fate. iHCEPLC can restore the BAK-based rabbit model of corneal endothelial dystrophy. Immunohistochemistry verified that such acuity restoration of the BAK-treated cornea is due to the introduced iHCEPLC, and such therapeutic effectiveness is observed in the long term. CONCLUSION: Here, we demonstrated that customized iHCEPLC has long-term therapeutic efficacy. As a progenitor cell, our iHCEPLC has a restricted cell lineage nature and can proliferate in vitro, supporting sufficient therapeutic candidate cells. Due to the immune-privileged nature of the cornea, our iHCEPLC proves the principle of therapeutical feasibility in both autogenic and allogeneic modalities.

Peripheral ulcerative keratitis in a patient with granulomatous rosacea.

A 24-year-old woman visited our emergency department due to intermittent dull pain in the right eye, blurred vision, foreign body sensation for 3 weeks, and progressive facial rash with pustules for 3 months. She had a history of recurring skin rash on her face and extremities since early adolescence. Peripheral ulcerative keratitis (PUK) was diagnosed based on slit-lamp examination and corneal topography and then granulomatous rosacea (GR) based on clinical manifestations and skin pathology. Topical prednisolone, artificial tears, oral doxycycline, oral prednisolone, and topical clindamycin were administered. After 1 month, PUK progressed to corneal perforation probably due to eye rubbing. The corneal lesion was repaired with a glycerol-preserved corneal graft. A dermatologist prescribed oral isotretinoin for 2 months in conjunction with topical betamethasone gradually tapered for 14 months. After 34 months of follow-up, no signs of skin and ocular recurrence were noted, and the cornea graft was intact. In conclusion, PUK may present with GR, and oral isotretinoin may be an effective therapy for PUK in the setting of GR.

Amniotic membrane transplantation in a patient with impending perforated corneal ulcer caused by Streptococcus mitis: A case report and review of literature.

BACKGROUND: Streptococcus mitis (S. mitis) is an opportunistic pathogen that can lead to severe ocular infections. In previous reports, penetrating keratoplasty (PK) was usually adopted for the treatment of persistent corneal ulcers. This report describes an unusual case of nonhealing descemetocele caused by S. mitis treated by antibiotics plus amniotic membrane transplantation (AMT). CASE SUMMARY: A 63-year-old woman presented with a right persistent corneal ulcer that she had suffered from for the past 9 mo. The culture of a corneal scraping yielded S. mitis. The right eye descemetocele decreased in diameter from 3 to 0.8 mm after the continuous administration of topical vancomycin and ceftriaxone for 2 wk. Due to the slow healing, AMT was performed. Her corneal erosion healed and gradually became clear. Her visual acuity recovered from initially counting fingers to 100/200 at the last follow-up, 67 mo after AMT. CONCLUSION: Antibiotics plus AMT may be an effective alternative treatment other than PK to promote epithelialization and to reduce inflammation in the corneas complicated by S. mitis keratitis.

Accelerated Corneal Endothelial Cell Loss after Phacoemulsification in Patients with Mildly Low Endothelial Cell Density.

By evaluating preoperative endothelial cell density (ECD), ECD loss after phacoemulsification can be predicted. In this retrospective cross-sectional study, we compared outcomes of phacoemulsification with different levels of preoperative ECD. Three-hundred-and-fifty-three patients aged between 18 and 90 years received phacoemulsification at Chang Gung Memorial Hospital. Age (p = 0.003), preoperative logMAR (p = 0.048), cataract grade (p = 0.005), preoperative ECD (p < 0.001), operation time (p = 0.043), phacoemulsification time (p = 0.001), and phacoemulsification energy (p < 0.001) were significantly associated with postoperative ECD change (%). Patients were divided into three groups according to preoperative ECD levels. Level of ECD, coefficient of variation (CV), cell hexagonality (HEX), central corneal thickness (CCT), visual acuity, underlying diseases, and complications were analyzed. With regard to groups, 29, 71, and 252 patients were respectively allocated into the markedly low (group A; ECD below 1000 cells/mm2), mildly low (group B; ECD between 1000 to 2000 cells/mm2), and normal (group C; ECD above 2000 cells/mm2) ECD level groups. The highest CV (40.8 ± 13.9%; p < 0.001) and lowest HEX (58.4 ± 14.6%; p < 0.001) were found in group A. Significant ECD loss was found in group B (28.9 ± 9.2%) as compared to group A (19.9 ± 5.4%) and C (15.0 ± 12.0%) (p < 0.001). No significant differences were found with regard to changes in CV (p = 0.941), HEX (p = 0.937), CCT (p = 0.346), and logMAR (p = 0.557) among the three groups. In conclusion, preoperative ECD level could be a novel predictive value for postoperative cell loss, which was the most prominent in mildly low ECD level group. Less phacoemulsification energy, earlier surgical intervention, or novel topical medications could be suggested for patients with an ECD range from 1000 to 2000 cells/mm2.

Accelerated corneal endothelial cell loss in two patients with granulomatosis with polyangiitis following phacoemulsification.

BACKGROUND: Generally, the loss rate of human endothelial cells (HCEC) in routine cataract surgery is 8.5%. When the corneal endothelial cells density (ECD) drops, the HCEC may decompensate to keep cornea dehydration which leads to corneal edema. Granulomatosis with polyangiitis (GPA) is an uncommon autoimmune disease involving multiple organs including eyes such as conjunctivitis, scleritis, uveitis, and corneal ulcer. In this study, we report two cases of GPA whose corneal ECD decreased significantly after phacoemulsification cataract surgery. CASE PRESENTATION: In the first case of 69-year-old male with GPA, the ECD dropped 39.6% (OD) four months after phacoemulsification and 38.1% (OS) six months postoperatively respectively. At the final follow-up, the residual ECD was only 55% in the right eye in the 49th month, and 56% remained in the left eye in the 39th month. In the second case of 54-year old female, left ECD dropped 63.9% at the 4th month after surgery and 69.6% ECD remained at the 15th month postoperatively while similar ECD of right eye before and after left eye surgery. CONCLUSION: Extensive preoperative ophthalmic evaluation and meticulous postoperative inflammation control should be applied to prevent severe loss of HCEC in GPA patients.

Clinical Outcomes of Penetrating Keratoplasty and Descemet Stripping Automated Endothelial Keratoplasty in Asian Population with American Corneas.

To determine the comparative efficacy and safety of penetrating keratoplasty (PK) and Descemet stripping automated endothelial keratoplasty (DSAEK) in the Asian population receiving imported donor corneas, our single-center retrospective study provides analysis supporting the transition from PK to DSAEK in the Asian population using imported American donor corneas. We analyzed 259 patients with 241 and 57 cases of PK and DSAEK respectively during 2008 to 2017 using imported corneas at Chang Gung Memorial Hospital, Linkou, Taiwan. In terms of long-term graft survival analysis, there was no difference between PK and DSAEK (log-rank p = 0.386, HR = 0.920, 95% CI: [0.641-1.380]). However, Cox proportional regression analysis revealed that corneal survival rate of DSAEK group in the first 100 days after transplantation was inferior than that of PK group (log-rank p < 0.001, HR = 2.733, 95% CI: [1.501-4.977])]. Despite the inferior survival rate, there were significantly less neovascularization and Descemet membrane folds in the DSAEK group. Importantly, the non-complication rate of DSAEK was much higher than that of PK with significant difference (PK, 25.7% vs DSAEK 42.0%, p = 0.022). Collectively, DSAEK is suggested as an alternative surgical modality in Asian patients using imported American donor corneas because of less complication, and no difference in long-term corneal graft survival rates between PK and DSAEK.

Optimization of reprogramming culture conditions for the generation of induced pluripotent stem cells from Col1a1 4F2A-Oct4-GFP mice with high efficiency.

A reprogrammable transgenic mouse strain, called Col1a1 4F2A-Oct4-GFP, was bred for the present study. Because the somatic cells of this mouse strain contain only two copies of each Yamanaka factor, these animals are inefficient at producing induced pluripotent stem cells (iPSCs; approx. 0.005%) under traditional culture conditions. With an optimized culture condition, the iPSC production rate of mouse embryonic fibroblasts (MEFs) of Col1a1 4F2A-Oct4-GFP mice (MEFCol1a14F2A-Oct4-GFP ) was increased to approximately 8%. Further, promotion of cell proliferation by serum supplementation did not enhance iPSC production. Inhibition of transforming growth factor ß (TGF-ß) in the serum by SB431542 neither affected the growth rate of MEFCol1a14F2A-Oct4-GFP nor promoted iPSC production. However, the use of the gamma-irradiated STO-NEO-LIF (γSNL) cells to serve as feeders for iPSC production resulted in a 5-fold higher rate of iPSC production than the use of γMEFICR feeders. Interestingly, the use of SB431542 with the γMEFICR -adopted system could eliminate this difference. RT-PCR-based comparative analysis further demonstrated that TGF-ß expression was 10-fold higher in γMEFICR than in γSNL cells. Consistent with previous reports, mesenchymal to epithelial transition was found to participate in the initial steps of reprogramming in the specific context of MEFCol1a14F2A-Oct4-GFP . Moreover, we found that the initial seeding density is one of the pivotal factors for determining a high efficiency of iPSC generation. The iPSCs efficiently generated from our MEFCol1a14F2A-Oct4-GFP resembled mouse embryonic stem cells (mESCs) in aspects of teratoma formation and germline transmission. Depending on the culture conditions, our Col1a1 4F2A-Oct4-GFP mouse system can generate bona fide iPSCs with variable efficiencies, which can serve as a tool for interrogating the route taken by cells during somatic reprogramming.