High glucose causes apoptosis of rabbit corneal epithelial cells involving activation of PERK-eIF2α-CHOP-caspase-12 signaling pathway.

AIM: To investigate the effect of high concentration of glucose (HCG) on double stranded RNA-activated protein kinase-like ER kinase (PERK)-eukaryotic initiation factor-2α (eIF2α)-transcription factor C/EBP homologous protein (CHOP)-cysteine aspartate specific proteinase (caspase-12) signaling pathway activation and apoptosis in rabbit corneal epithelial cells (RCECs). METHODS: RCECs were treated by different concentrations of glucose for 0-48h. The expressions of PERK, p-PERK, eIF2α, p-eIF2α, 78 kDa glucose-regulated protein 78 (GRP78), CHOP, B-cell lymphoma 2 (Bcl-2), B-cell lymphoma-2-associated X protein (Bax) and caspase-12 were determined by Western blot. Apoptosis was detected by TUNEL assay. Meanwhile, the function of PERK-eIF2α-CHOP-caspase-12 signaling pathway activation in high glucose-induced apoptosis was evaluated using PERK inhibitor, GSK2606414. RESULTS: HCG significantly promoted the expression of p-PERK, p-eIF2α, GRP78, CHOP, Bax and cleaved caspase-12 in RCECs (P<0.05), while remarkably decreased the expression of Bcl-2 and caspase-12 (P<0.05), and the alterations caused by glucose were in concentration- and time-dependent manners. Meanwhile, PERK and eIF2α expressions were not affected in all groups (P>0.05). TUNEL assay showed that the apoptosis rate of RCECs in the HCG group increased significantly in contrast with that in the normal concentration of glucose or osmotic pressure control group (P<0.05), and the apoptosis rate increased with the increase of glucose concentration within limits (P<0.05). GSK2606414 down-regulated the expression of p-PERK and p-eIF2α in the HCG group (P<0.05), while still did not affect the expression of PERK and eIF2α among groups (P>0.05). Correspondingly, GSK2606414 also significantly reduced the apoptosis rate induced by high glucose (P<0.05). CONCLUSION: HCG activates PERK-eIF2α-CHOP-caspase-12 signaling pathway and promotes apoptosis of RCECs.

Effects of asymmetric dimethylarginine on bovine retinal capillary endothelial cell proliferation, reactive oxygen species production, permeability, intercellular adhesion molecule-1, and occludin expression.

PURPOSE: Asymmetric dimethylarginine (ADMA), an endogenous competitive inhibitor of nitric oxide synthase, is associated with impaired endothelial dysfunction, such as chronic heart failure, hypertension, diabetes, and pulmonary hypertension. The effects of ADMA on cell proliferation, reactive oxygen species (ROS) production, cell permeability, intercellular adhesion molecule-1 (ICAM-1), and tight-junction protein occludin levels in bovine retinal capillary endothelial cells (BRCECs) were investigated. METHODS: A cell proliferation assay was performed using the novel tetrazolium compound 3-(4,5-dimethylthiazol-2-yl)-5-(3-carboxymethoxyphenyl)-2-(4-sulfophenyl)-2H-tetrazolium and an electron coupling reagent. Intracellular ROS levels were determined using the fluorescent probe CM-H(2)DCFDA. Horseradish peroxidase was used for a permeability assay. ICAM-1 and tight-junction protein occludin were assessed by western blotting and quantitative real-time PCR. RESULTS: Cell proliferation was significantly inhibited by ADMA. ADMA increased intracellular ROS generation in BRCECs. The increased ROS production induced by ADMA was markedly inhibited by the angiotensin II receptor-blocker telmisartan, the angiotensin-converting enzyme inhibitor benazepril, the reduced form of nicotinamide-adenine dinucleotide phosphate (NADPH) oxidase inhibitor diphenyliodonium (DPI), or the antioxidant and free-radical scavenger N-acetyl-L-cysteine (NAC). ADMA significantly increased horseradish peroxidase (HRP) permeability in BRCECs. Benazepril, telmisartan, DPI, and NAC downregulated cell permeability. ADMA markedly upregulated ICAM-1 expression in BRCECs, which were downregulated by telmisartan, DPI, and NAC. ADMA significantly downregulated occludin expression in BRCECs. Benazepril and telmisartan upregulated occludin expression in BRCECs exposed to ADMA. CONCLUSIONS: Our results provide the first reported evidence that ADMA has potent adverse effects on cell proliferation, intracellular ROS generation, cell permeability, levels of ICAM-1, and the tight-junction protein occludin. Angiotensin-converting enzyme inhibitors, angiotensin II receptor blockers, and antioxidants are effective inhibitors of the adverse effects of ADMA.

Pediatric penetrating keratoplasty in Shanghai: a retrospective multiple centre study from 2003 to 2007.

BACKGROUND: With advances in pediatric keratoplasty over the last two decades, the success rate of surgery has increased remarkably. However, few epidemiological studies in this field have been performed in China. The present study investigated the indications and characteristics of pediatric penetrating keratoplasty in Shanghai. METHODS: All records of pediatric keratoplasty performed on 156 eyes in 149 children (< 14 years) at four ophthalmic units in Shanghai during a 5-year period (2003 - 2007) were used for this retrospective study. Patients were from the Eye, Ear, Nose & Throat Hospital of Fudan University, Shanghai No.10 People's Hospital, Shanghai No.1 People's Hospital, and Shanghai Peace Ophthalmology Hospital; and included mostly school-age children (97 boys and 59 girls). RESULTS: The median age at surgery was 9 years with an interval quartile range of 6-12 years. Scarring after keratitis (29.5%, 46/156) and traumatic corneal scar (19.2%, 30/156) were the most common indications. Best Corrected Snellen visual acuity (BCSVA) was reported only in 72% (112/156) of cases. Visual acuity outcomes were significantly better for keratoplasty 1 year postoperatively compared to preoperative visual acuity (P = 0.001). Of all patients, 13% (14/112) achieved a BCSVA of 6/18 or better. None of the indications was associated with a higher rate of failure compared to after 1 year follow-up. CONCLUSION: We obtained valuable information on pediatric keratoplasty in Shanghai. There was no significant difference in graft survival rate among the two indications. Vision outcomes after corneal transplantation in Chinese children were significantly better compared with pre-operation status. In conclusion, corneal graft survival can be achieved in childhood.