Nutr Res Pract.  2020 Jun;14(3):203-217. 10.4162/nrp.2020.14.3.203.

Centella asiatica extract prevents visual impairment by promoting the production of rhodopsin in the retina

• Park DW
• Jeon H
• So R
• Kang SC

• KMID: 2502481
• DOI: http://doi.org/10.4162/nrp.2020.14.3.203

Abstract

BACKGROUND
/OBJECTIVE: Centella asiatica, also known as Gotu kola, is a tropical medicinal plant native to Madagascar, Southeast Asia, and South Africa. It is well known to have biological activities, including wound healing, anti-inflammatory, antidiabetic, cytotoxic, and antioxidant effects. The purpose of this study was to determine the efficacy of extracts of C. asiatica against age-related eye degeneration and to examine their physiological activities.
MATERIALS/METHODS
To determine the effects of CA-HE50 (C. asiatica 50% EtOH extract) on retinal pigment cells, we assessed the cytotoxicity of CoCl2 and oxidized-A2E in ARPE- 19 cells and observed the protective effects of CA-HE50 against N-methyl-N-nitrosourea (MNU)-induced retinal damage in C57BL/6 mice. In particular, we measured factors related to apoptosis and anti-oxidation and the protein levels of rhodopsin/opsin. We also measured glucose uptake to characterize glucose metabolism, a major factor in cell protection.
RESULTS
Induction of cytotoxicity with CoCl2 and oxidized-A2E inhibited decreases in the viability of ARPE-19 cells when CA-HE50 was administered, and promoted glucose uptake under normal conditions (P < 0.05). In addition, CA-HE50 inhibited degeneration/apoptosis of the retina in the context of MNU-induced toxicity (P < 0.05). In particular, CA-HE50 at 200 mg/kg inhibited the cleavage of pro-caspase-3 and pro-poly (ADP-ribose)-polymerase and maintained the expressions of nuclear factor erythroid 2-related factor 2 and heme oxygenase-1 similar to normal control levels. Rhodopsin/opsin expression was maintained at a higher level than in normal controls.
CONCLUSION
A series of experiments confirmed that CA-HE50 was effective for inhibiting or preventing age-related eye damage/degeneration. Based on these results, we believe it is worthwhile to develop drugs or functional foods related to age-related eye degeneration using CA-HE50.

Keyword

Centella asiatica; etinal degeneration; oxidative stress; cytoprotection

Figure

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  Fig. 1 Cytoprotective effects of CA-HE50 against CoCl2 in ARPE-19 cells. (A) Cytotoxicity by CoCl2 in ARPE-19 cells. Cytoprotective effects of (B) CA30, (C) CA50, and (D) CA70 against 500 µM CoCl2. Representative data are expressed as mean ± standard error of the mean of three different experiments (n = 3) for each group.*P < 0.05 compared with the normal control group. †P < 0.05 compared with the CoCl2 control (vehicle) group.

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  Fig. 2 High-performance liquid chromatography (HPLC-PDA) chromatograms of C. asiatica 50% EtOH extract (CA-HE50), asiaticoside, and asiatic acid. (A) HPLC chromatograms of asiaticoside and asiatic acid (standard). (B) HPLC chromatogram of CA-HE50.

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  Fig. 3 Cytoprotective effects of asiaticoside and asiatic acid on CoCl2 in ARPE-19 cells. Cytotoxicity of (A) AS and (B) AA in ARPE-19 cells. Cytoprotective effects of (C) AS and (D) AA on CoCl2. Representative data are expressed as mean ± standard error of the mean of 3 different experiments (n = 3) for each group.*P < 0.05 compared with the normal control group. †P < 0.05 compared with the CoCl2 control (vehicle) group.

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  Fig. 4 Cytoprotective effects of CA-HE50 on oxidized-A2E in ARPE-19 cells. (A) Production of oxidized-A2E depends on UV-irradiation time. (B) Inhibitory effect of CA-HE50 on oxidized-A2E production. (C) Cytoprotective effect of CA-HE50 on oxidized-A2E in ARPE-19 cells. Representative data are expressed as mean ± standard error of the mean of three different experiments (n = 3) for each group.A2E, a lipofuscin fluorophore; UV, ultraviolet.*P < 0.05 compared with the normal control group. †P < 0.05 compared with the oxidized-A2E control (vehicle) group.

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  Fig. 5 Effects of CA-HE50, asiaticoside, and asiatic acid on glucose uptake in ARPE-19 cells. Effects of (A) CA-HE50, (B) AS, and (C) AA on glucose uptake in ARPE-19 cells. Representative data are expressed as mean ± standard error of the mean of 3 different experiments (n = 3) for each group.*P < 0.05, **P < 0.01, and ***P< 0.001 compared with the control group.

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  Fig. 6 Hepatoprotective effect of CA-HE50 against MNU-induced hepatotoxicity. (A) AST and (B) ALT release the inhibitory effect of CA-HE50. Representative data are expressed as mean ± standard error of the mean (n = 6) for each group.MNU, N-methyl-N-nitrosourea; AST, aspartate aminotransferase; ALT, alanine aminotransferase.*P < 0.001 compared with the normal control group. †P < 0.001 compared with the MNU (vehicle) control group.

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  Fig. 7 Histopathology of retinas according to H&E staining from 5 groups of mice at 24 h post-injection. (A) Representative retinal images of H&E-stained retinal sections from normal (MNU-uninjected) eyes. (B) Representative retinal image from the MNU-injection group (vehicle group). (C) Representative retinal image for mice administered 50 mg/kg CA-HE50 for 7 days and then injected with MNU. (D) Representative retinal image for mice administered 100 mg/kg CA-HE50 for 7 days and then injected with MNU. (E) Representative retinal image for mice administered 200 mg/kg CA-HE50 for 7 days and then injected with MNU. Data are representative of 6 independent experiments.MNU, N-methyl-N-nitrosourea; GCL, ganglion cell layer; IPL, inner plexiform layer; INL, inner nuclear layer; OPL, outer plexiform layer; ONL, outer nuclear layer; IS, inner segments; OS, outer segments; RPE, retinal pigment epithelium.

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  Fig. 8 Western blot analysis for expressions of proteins associated with retinal damage expressed in the eyes of C57BL/6 mice that induced retinal damage with MNU after 7 days of CA-HE50. (A) Effects of CA-HE50 on NRF2, HO-1, pro-PARP, and pro-caspase-3 at the protein level. (B) Effects of CA-HE50 on rhodopsin and opsin at the protein level. β-actin was detected as an internal standard. Data are representative of 6 independent experiments.MNU, N-methyl-N-nitrosourea; NRF2, Nuclear factor erythroid 2-related factor 2; HO-1, heme oxygenase 1; PARP, Poly (ADP-ribose) polymerase.

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