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Protective Effect of Hexane and Ethanol Extract of Piper Longum L. on Gentamicin-Induced Hair Cell Loss in Neonatal Cultures

최준|송재준|yadav mukesh kumar
대한이비인후과학회 (Korean Society of Otorhinolaryngology-Head and Neck Surgery)|2014.03|Clinical and Experimental Otorhinolaryngology|제7권 제1호|pp. 13-18|DOI http://dx.doi.org/10.3342/ceo.2014.7.1.13

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키워드

AntioxidantApoptosisOuter hair cellGentamicinInner hair cellAutotoxicityPiper longum

초록

Objectives. Gentamicin (GM) is a commonly used aminoglycoside antibiotic that generates free oxygen radicals within the inner ear, which can cause vestibulo-cochlear toxicity and permanent damage to the sensory hair cells and neurons. Piper longum L. (PL) is a well-known spice and traditional medicine in Asia and Pacific islands, which has been re- ported to exhibit a wide spectrum of activity, including antioxidant activity. In this study, we evaluated the effect of hexane:ethanol (2:8) PL extract (subfraction of PL [SPL] extract) on GM-induced hair cell loss in basal, middle and apical regions in a neonatal cochlea cultures. Methods. The protective effects of SPL extract were measured by phalloidin staining of cultures from postnatal day 2-3 mice with GM-induced hair cell loss. The anti-apoptosis activity of SPL extract was measured using double labeling by terminal deoxynucleotidyl transferase dUTP nick end labeling (TUNEL) and myosin-7a staining. The radical-scav- enging activity of SPL extract was assessed using the 1,1-diphenyl-2-picrylhydrazyl (DPPH) assay. Results. SPL extract at a concentration of 1 µg/mL significantly inhibited GM-induced hair cell loss at basal and middle re- gion of cochlea, while 5 µg/mL was effective against apical region hair cell loss. The protective effect of SPL extract was concentration dependent and hair cells retained their stereocilia in explants treated with SPL extract prior to treat- ment with 0.3 mM GM. SPL extract decreased GM-induced apoptosis of hair cells as assessed by TUNEL staining. The outer hair and inner hair counts were not decreased in SPL extract treated groups in compare to GM treated ex- plants. Additionally, SPL extract showed concentration dependent radical scavenging activity in a DPPH assay. Conclusion. An anti-apoptosis effect and potent radical scavenger activity of SPL extract protects from GM-induced hair cell loss at basal, middle and apical regions in neonatal cochlea cultures.

Abstract

Objectives. Gentamicin (GM) is a commonly used aminoglycoside antibiotic that generates free oxygen radicals within the inner ear, which can cause vestibulo-cochlear toxicity and permanent damage to the sensory hair cells and neurons. Piper longum L. (PL) is a well-known spice and traditional medicine in Asia and Pacific islands, which has been re- ported to exhibit a wide spectrum of activity, including antioxidant activity. In this study, we evaluated the effect of hexane:ethanol (2:8) PL extract (subfraction of PL [SPL] extract) on GM-induced hair cell loss in basal, middle and apical regions in a neonatal cochlea cultures. Methods. The protective effects of SPL extract were measured by phalloidin staining of cultures from postnatal day 2-3 mice with GM-induced hair cell loss. The anti-apoptosis activity of SPL extract was measured using double labeling by terminal deoxynucleotidyl transferase dUTP nick end labeling (TUNEL) and myosin-7a staining. The radical-scav- enging activity of SPL extract was assessed using the 1,1-diphenyl-2-picrylhydrazyl (DPPH) assay. Results. SPL extract at a concentration of 1 µg/mL significantly inhibited GM-induced hair cell loss at basal and middle re- gion of cochlea, while 5 µg/mL was effective against apical region hair cell loss. The protective effect of SPL extract was concentration dependent and hair cells retained their stereocilia in explants treated with SPL extract prior to treat- ment with 0.3 mM GM. SPL extract decreased GM-induced apoptosis of hair cells as assessed by TUNEL staining. The outer hair and inner hair counts were not decreased in SPL extract treated groups in compare to GM treated ex- plants. Additionally, SPL extract showed concentration dependent radical scavenging activity in a DPPH assay. Conclusion. An anti-apoptosis effect and potent radical scavenger activity of SPL extract protects from GM-induced hair cell loss at basal, middle and apical regions in neonatal cochlea cultures.

참고문헌 (30)

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