Photoprotective effects of cranberry juice and its various fractions against blue light-induced impairment in human retinal pigment epithelial cells

Context: Cranberry has numerous biological activities, including antioxidation, anticancer, cardioprotection, as well as treatment of urinary tract infection (UTI), attributed to abundant phenolic contents.

Objective: The current study focused on the effect of cranberry juice (CJ) on blue light exposed human retinal pigment epithelial (ARPE-19) cells which mimic age-related macular degeneration (AMD).

Materials and methods: Preliminary phytochemical and HPLC analysis, as well as total antioxidant capacity and scavenging activity of cranberry ethyl acetate extract and different CJ fractions (condensed tannins containing fraction), were evaluated. In cell line model, ARPE-19 were irradiated with blue light at 450?nm wavelength for 10?h (mimic AMD) and treated with different fractions of CJ extract at different doses (5–50?μg/mL) by assessing the cell viability or proliferation rate using MTT assay (repairing efficacy).

Results: Phytochemical and HPLC analysis reveals the presence of several phenolic compounds (flavonoids, proanthocyanidin, quercetin) in ethyl acetate extract and different fractions of CJ. However, the condensed tannin containing fraction of ethyl acetate extract of CJ displayed the greater (p?p?p?Discussion and conclusion: In conclusion, this study distinctly proved that condensed tannin containing fraction of CJ probably exhibits better free radicals scavenging activity and thereby effectively protected the ARPE-19 cells and thus, hampers the progress of AMD.     

Induction of vascular endothelial growth factor by 4-hydroxynonenal and its prevention by glutathione precursors in retinal pigment epithelial cells

Although 4-hydroxynonenal, a highly reactive lipid peroxidation product, is implicated in several age-related disorders such as Alzheimer's and Parkinson's diseases, its role in age-related macular degeneration is not known. The purpose of this study was to determine whether 4-hydroxynonenal increases vascular endothelial growth factor (VEGF) expression in human retinal pigment epithelial cells (ARPE-19), a source of VEGF in choroidal neovascularization observed in age-related macular degeneration. In addition, it was the purpose of this study to assess whether glutathione (GSH) and GSH precursors can inhibit the effects of 4-hydroxynonenal. At 1 micro M, 4-hydroxynonenal did not alter cell viability, but elevated VEGF secretion and mRNA expression by 35% (p<0.05) and 1.9-fold (p<0.05), respectively. However, at concentrations 5 microM and above, 4-hydroxynonenal reduced VEGF secretion as well as cell viability. At 1 and 10 microM, 4-hydroxynonenal did not induce apoptosis in ARPE-19 cells. 4-Hydroxynonenal (1 microM) reduced intracellular GSH by 25% (p<0.05) and increased oxidative stress by 50% (p<0.05). GSH precursor pretreatment for 1 h, which increased intracellular GSH levels by 50% (p<0.05), as well as GSH co-treatment, inhibited the VEGF-inductive and cytotoxic effects of 4-hydroxynonenal. Thus, 4-hydroxynonenal (1 microM) induces VEGF expression and secretion in ARPE-19 cells. This effect is likely due to GSH depletion and an associated increase in intracellular oxidative stress, resulting in increased VEGF mRNA levels. 4-Hydroxynonenal-mediated VEGF secretion as well as cytotoxicity can be reversed with GSH precursor pretreatment or GSH co-treatment.     

Bisphenol A induced apoptosis via oxidative stress generation involved Nrf2/HO-1 pathway and mitochondrial dependent pathways in human retinal pigment epithelium (ARPE-19) cells

Bisphenol A (BPA) is an estrogen-like compound, and an environmental hormone, that is commonly used in daily life. Therefore, it may enter the human body through food or direct contact, causing BPA residues in blood and urine. Because most studies focused on the analysis of BPA in reproductive cells or tissues, regarding evidence the effect of BPA on human retinal pigment epithelium (ARPE-19) cells unavailable. Accordingly, the present study explored the cytotoxicity of BPA on ARPE-19 cells. After BPA treatment, the expression of Bcl-XL an antiapoptotic protein, in the mitochondria decreased, and the expression of Bax, a proapoptotic protein increased. Then the mitochondrial membrane potential was affected. BPA changed in mitochondrial membrane potential led to the release of cytochrome C, which activated caspase-9 to promote downstream caspase-3 leading to cytotoxicity. The nuclear factor (erythroid-derived 2)-like 2 (Nrf2) and heme oxygenase 1 (HO-1) pathway play a major role in age-related macular degeneration. Our results showed that expression of HO-1 and Nrf2 suppressed by BPA. Superoxide dismutase and catalase, which Nrf2 downstream antioxidants, were degraded by BPA. AMP-activated kinase (AMPK), which can regulate the phosphorylation of Nrf2, and the phosphorylation of AMPK expression was reduced by BPA. Finally, BPA-induced ROS generation and cytotoxicity were reduced by N-acetyl-l -cysteine. Taken together, these results suggest that BPA induced ARPE-19 cells via oxidative stress, which was associated with down regulated Nrf2/HO-1 pathway, and the mitochondria dependent apoptotic signaling pathway.     

In vitro cytotoxicity of eight β-blockers in human corneal epithelial and retinal pigment epithelial cell lines: Comparison with epidermal keratinocytes and dermal fibroblasts

β-Blockers are a class of agents that have been used extensively in topical preparations for the treatment of glaucoma. Recent evidence indicates that they may also be useful in a number of retinal diseases. Because biocompatibility is of utmost importance in the treatment of ocular-related diseases, we compared the in vitro cytotoxicity, using the MTT assay, of eight clinically available β-blockers (propranolol, alprenolol, atenolol, labetalol, metoprolol, pindolol, timolol, and bisoprolol) on human corneal epithelial and retinal pigment epithelial cell lines. Primary and immortalized corneal and retinal cell lines were compared for their susceptibility to the cytotoxic effect of the drugs. The cytotoxicity of β-blockers was also evaluated on human skin keratinocytes and fibroblasts in order to investigate susceptibility differences as a function of the tissue of origin. Results demonstrated large differences in cytotoxicity (about 60-fold) for these closely related drugs on the same cell line. Conversely, only relatively small differences in cytotoxicity were observed between the different cell lines for the same drug, indicating that the mechanism of cytotoxicity is not cell-specific. Calculation of the ratio between the cytotoxicity of β-blockers and their β-blocking constant is presented as a potential tool to help identify the least irritating, most potent drug.     

Synergistic effects of hydrogen peroxide and ethanol on cell viability loss in PC12 cells by increase in mitochondrial permeability transition

The promoting effect of ethanol against the cytotoxicity of hydrogen peroxide (H2O2) in differentiated PC12 cells was assessed by measuring the effect on the mitochondrial membrane permeability. Treatment of PC12 cells with H2O2 resulted in the nuclear damage, decrease in the mitochondrial transmembrane potential, cytosolic accumulation of cytochrome c, activation of caspase-3, increase in the formation of reactive oxygen species (ROS) and depletion of GSH. In PC12 cells and dopaminergic neuroblastoma SH-SY5Y cells, the promoting effect of ethanol on the H2O2-induced cell death was increased with exposure time. Ethanol promoted the nuclear damage, change in the mitochondrial membrane permeability, ROS formation and decrease in GSH contents due to H2O2 in PC12 cells. Catalase, carboxy-PTIO, Mn-TBAP, N-acetylcysteine, cyclosporin A and trifluoperazine inhibited the H2O2 and ethanol-induced mitochondrial dysfunction and cell injury. The results show that the ethanol treatment promotes the cytotoxicity of H2O2 against PC12 cells. Ethanol may enhance the H2O2-induced viability loss in PC12 cells by promoting the mitochondrial membrane permeability change, release of cytochrome c and subsequent activation of caspase-3, which is associated with the increased formation of ROS and depletion of GSH. The findings suggest that ethanol as a promoting agent for the formation of mitochondrial permeability transition may enhance the neuronal cell injury caused by oxidants.     

Neurotoxic injury pathways in differentiated mouse motor neuron-neuroblastoma hybrid (NSC-34D) cells in vitro--limited effect of riluzole on thapsigargin, but not staurosporine, hydrogen peroxide and homocysteine neurotoxicity

The neuroblastoma-spinal motor neuron fusion cell line, NSC-34, in its differentiated form, NSC-34D, permits examining the effects of riluzole, a proven treatment for amyotrophic lateral sclerosis (ALS) on cell death induction by staurosporine (STS), thapsigargin (Thaps), hydrogen peroxide (H₂O₂) and homocysteine (HCy). These neurotoxins, applied exogenously, have mechanisms of action related to the various proposed molecular pathogenetic pathways in ALS and are differentiated from endogenous cell death that is associated with cytoplasmic aggregate formation in motor neurons. Nuclear morphology, caspase-3/7 activation and high content imaging were used to assess toxicity of these neurotoxins with and without co-treatment with riluzole, a benzothiazole compound with multiple pharmacological actions. STS was the most potent neurotoxin at killing NSC-34D cells with a toxic concentration at which 50% of maximal cell death is achieved (TC₅₀ = 0.01 μM), followed by Thaps (TC₅₀ = 0.9 μM) and H₂O₂ (TC₅₀ = 15 μM) with HCy requiring higher concentrations to kill at the same level (TC₅₀ = 2200 μM). Riluzole provided neurorescue with a 20% absolute reduction (47.6% relative reduction) in apoptotic cell death against Thaps-induced NSC-34D cell (p ≤ 0.05), but had no effect on STS-, H₂O₂- and HCy-induced NSC-34D cell death. This effect of riluzole on Thaps induction of cell death was independent of caspase-3/7 activation. Riluzole mitigated a toxin that can cause intracellular calcium dysregulation associated with endoplasmic reticulum (ER) stress but not toxins associated with other cell death mechanisms.