J Korean Ophthalmol Soc.  2016 Jan;57(1):113-119. 10.3341/jkos.2016.57.1.113.

Neuroprotective Effects of Betaxolol Mediated by Heme Oxygenase-1 Induction in RGC-5

• Cha JB
• Kwon MY
• Chung SW
• Woo JM

• KMID: 2213854
• DOI: http://doi.org/10.3341/jkos.2016.57.1.113

Abstract

PURPOSE
To evaluate the neuroprotective effects of betaxolol (betaxolol hydrochloride) under hypoxic conditions using retinal ganglion cells (RGC-5) and determine whether heme oxygenase-1 (HO-1) expression exerts cytoprotective effects.
METHODS
In this study, cultured RGC-5 cells were incubated with different concentrations of betaxolol hydrochloride (0.1 microM, 1 microM or 5 microM) and with 10 microM zinc protoporphyrin (ZnPP), in a hypoxia incubator (1% O2, 5% CO2, 94% N2) for 48 hours and the cell viability of each group was determined. Additionally, cell viability was measured after RGC-5 cells were incubated with 5 microM of brinzolamide (Azopt(R)), brimonidine tartrate (Alphagan(R)) or travoprost (Travatan(R)). RGC-5 cells were divided into three groups and incubated under three different conditions, normoxia group (20% O2, 5% CO2), hypoxia group (1% O2, 5% CO2) and the group with 5 microM of Betoptic S(R) treated under hypoxic conditions (hypoxia, Betoptic S(R)). After incubation for 4, 8, 12 and 24 hours, HO-1 expression was analyzed using Western blotting.
RESULTS
Cell viability significantly increased in RGC-5 cells treated with Betoptic S(R) compared with other antiglaucoma agents. Increased levels of HO-1 expression indicate its relevance in cell viability. Furthermore, increased RGC-5 cell viability by Betoptic S(R) was significantly reduced in the HO-1 inhibitor ZnPP-treated group.
CONCLUSIONS
We reaffirmed the known cytoprotective effects of Betoptic S(R) and the results suggests that HO-1 expression exerts cytoprotective effects against hypoxia.

Keyword

Betaxolol; Heme oxygenage-1; Hypoxia; Retinal ganglion cells

MeSH Terms

Anoxia
Betaxolol*
Blotting, Western
Cell Survival
Heme Oxygenase-1*
Heme*
Incubators
Neuroprotective Agents*
Retinal Ganglion Cells
Zinc
Brimonidine Tartrate
Travoprost
Betaxolol
Heme
Heme Oxygenase-1
Neuroprotective Agents
Zinc

Figure

• 

  Figure 1. Comparison of retinal ganglion cell (RGC-5) surviv-al rates with anti-glaucomatous agent after 48 hours. Addition of 5 μ M of betaxolol hydrochloride increased cellular survival (-STD: before preconditioning; +STD: after precondition-ing). STD = stauroporine; Azopt® = brinzolamide; Alphagan®= brimonidine tartrate; Travatan® = travopost; Betoptic S® = betaxolol hydrochloride. * p < 0.05 by student’s paired t-test.

• 

  Figure 2. Survival rates and microscopic finding of retinal gan-glion cells compared to BET concentraion. (A) Control and BET 5 μ M-treated group cultured for 48 hours and then taken by an op-tical microscope magnification 40 times without stain (-STD: be-fore preconditioning; +STD: after preconditioning). (B) Effect of BET on the survival of retinal ganglion cell (RGC-5) compared to BET concentration (Hypoxia: 5% CO2, 1% O2). STD = staur-oporine; BET = Betoptic S®. * p < 0.05 by student’s paired t-test.

• 

  Figure 3. Expression of HO-1 in accordance with the period in control group, hypoxia group, hypoxia and BET-treated group. Histogram presents the ratio of HO-1/β-actin expression. Western blotting was analyzed by the Image J software (Normoxia: 5% CO2, 20% O2; Hypoxia: 5% CO2, 1% O2). STD = stauroporine; BET = Betoptic S® 5 μ M; HO-1 = heme oxygenase-1. * p < 0.05 by student’s unpaired t-test.

• 

  Figure 4. Effect of HO-1 inhibitor on cell survival rates after ad-dition of BET. When compared to non-addition of HO-1 in-hibitor (ZnPP), Increased retinal ganglion cell (RGC-5) due to BET neuroprotective effect was reduced after addition of HO-1 inhibitor (ZnPP; -STD, – Hypoxia, -BET and – ZnPP: before preconditioning; +STD, +Hypoxia, -BET and -ZnPP: just af-ter the preconditioning). STD = stauroporine; BET = Betoptic S®; ZnPP = zinc protoporphyrin 10 μ M; HO-1 = heme oxy-genase-1. * p < 0.05 by student’s paired t-test.

Reference

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