Article

Hsp70 inhibits aminoglycoside-induced hair cell death and is necessary for the protective effect of heat shock

Pathology and Laboratory Medicine, Medical University of South Carolina, Charleston, SC 29403, USA.
Journal of the Association for Research in Otolaryngology (Impact Factor: 2.55). 06/2008; 9(3):277-89. DOI: 10.1007/s10162-008-0122-2
Source: PubMed

ABSTRACT Sensory hair cells of the inner ear are sensitive to death from aging, noise trauma, and ototoxic drugs. Ototoxic drugs include the aminoglycoside antibiotics and the antineoplastic agent cisplatin. Exposure to aminoglycosides results in hair cell death that is mediated by specific apoptotic proteins, including c-Jun N-terminal kinase (JNK) and caspases. Induction of heat shock proteins (Hsps) is a highly conserved stress response that can inhibit JNK- and caspase-dependent apoptosis in a variety of systems. We have previously shown that heat shock results in a robust upregulation of Hsps in the hair cells of the adult mouse utricle in vitro. In addition, heat shock results in significant inhibition of both cisplatin- and aminoglycoside-induced hair cell death. In our system, Hsp70 is the most strongly induced Hsp, which is upregulated over 250-fold at the level of mRNA 2 h after heat shock. Therefore, we have begun to examine the role of Hsp70 in mediating the protective effect of heat shock. To determine whether Hsp70 is necessary for the protective effect of heat shock against aminoglycoside-induced hair cell death, we utilized utricles from Hsp70.1/3 (-/-) mice. While heat shock inhibited gentamicin-induced hair cell death in wild-type utricles, utricles from Hsp70.1/3 (-/-) mice were not protected. In addition, we have examined the role of the major heat shock transcription factor, Hsf1, in mediating the protective effect of heat shock. Utricles from Hsf1 (-/-) mice and wild-type littermates were exposed to heat shock followed by gentamicin. The protective effect of heat shock on aminoglycoside-induced hair cell death was only observed in wild-type mice and not in Hsf1 (-/-) mice. To determine whether Hsp70 is sufficient to protect hair cells, we have utilized transgenic mice that constitutively overexpress Hsp70. Utricles from Hsp70-overexpressing mice and wild-type littermates were cultured in the presence of varying neomycin concentrations for 24 h. The Hsp70-overexpressing utricles were significantly protected against neomycin-induced hair cell death at moderate to high doses of neomycin. This protective effect was achieved without a heat shock. Taken together, these data indicate that Hsp70 and Hsf1 are each necessary for the protective effect of heat shock against aminoglycoside-induced death. Furthermore, overexpression of Hsp70 alone significantly inhibits aminoglycoside-induced hair cell death.

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    • "JNK activation occurs following aminoglycoside exposure (Pirvola et al., 2000; Cheng et al., 2005). Inhibitors of the JNK pathway (Pirvola et al., 2000; Bodmer et al., 2002; Wang et al., 2003; Matsui et al., 2004; Sugahara et al., 2006) or inducers of the heat shock pathway (Cunningham and Brandon, 2006; Taleb et al., 2008) are partially protective against aminoglycoside-induced hair cell death. Overexpression of the anti-apoptotic factor, Bcl-2, and inhibition of intrinsic caspase cascades (cas9, cas3) attenuates aminoglycoside-induced hair cell death in chick and rodent (Cunningham et al., 2002; Cheng et al., 2003; Cunningham et al., 2004), whereas inhibiting inhibitors of this path accentuates hair cell death (Tabuchi et al., 2007). "
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    ABSTRACT: We report a series of experiments investigating the kinetics of hair cell loss in lateral line neuromasts of zebrafish larvae following exposure to aminoglycoside antibiotics. Comparisons of the rate of hair cell loss and the differential effects of acute versus chronic exposure to gentamicin and neomycin revealed markedly different results. Neomycin induced rapid and dramatic concentration-dependent hair cell loss that is essentially complete within 90 min, regardless of concentration or exposure time. Gentamicin-induced loss of half of the hair cells within 90 min and substantial additional loss, which was prolonged and cumulative over exposure times up to at least 24h. Small molecules and genetic mutations that inhibit neomycin-induced hair cell loss were ineffective against prolonged gentamicin exposure supporting the hypothesis that these two drugs are revealing at least two cellular pathways. The mechanosensory channel blocker amiloride blocked both neomycin and gentamicin-induced hair cell death acutely and chronically indicating that these aminoglycosides share a common entry route. Further tests with additional aminoglycosides revealed a spectrum of differential responses to acute and chronic exposure. The distinctions between the times of action of these aminoglycosides indicate that these drugs induce multiple cell death pathways.
    Hearing research 04/2009; 253(1-2):32-41. DOI:10.1016/j.heares.2009.03.001 · 2.85 Impact Factor
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    • "In vitro and in vivo studies in birds and mammals have uncovered some of the key molecular players in these apoptotic cascades (reviewed in Van de Water et al., 2004; Cheng et al., 2005). Aminoglycoside-induced cell death is thought to act through the so-called ''intrinsic mitochondrial cell death pathway " although multiple overlapping death and protective factors are probably involved including oxidative stress, JNK signaling , and heat shock proteins (Hirose et al., 1997; Pirvola et al., 2000; Sha and Schacht, 2000; Dehne et al., 2002; Wang et al., 2003; Cunningham et al., 2004; Cunningham and Brandon, 2006; Sugahara et al., 2006; Taleb et al., 2008). Mitochondrial swelling and release of cytochrome c into the cytoplasm, hallmarks of classical apoptosis, are early signs of aminoglycoside ototoxicity (Hirose et al., 1999; Mangiardi et al., 2004; Matsui et al., 2004; Owens et al., 2007). "
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    ABSTRACT: Aminoglycoside antibiotics cause death of sensory hair cells. Research over the past decade has identified several key players in the intracellular cascade. However, the role of the extracellular environment in aminoglycoside ototoxicity has received comparatively little attention. The present study uses the zebrafish lateral line to demonstrate that extracellular calcium and magnesium ions modulate hair cell death from neomycin and gentamicin in vivo, with high levels of either divalent cation providing significant protection. Imaging experiments with fluorescently-tagged gentamicin show that drug uptake is reduced under high calcium conditions. Treating fish with the hair cell transduction blocker amiloride also reduces aminoglycoside uptake, preventing the toxicity, and experiments with variable calcium and amiloride concentrations suggest complementary effects between the two protectants. Elevated magnesium, in contrast, does not appear to significantly attenuate drug uptake, suggesting that the two divalent cations may protect hair cells from aminoglycoside damage through different mechanisms. These results provide additional evidence for calcium- and transduction-dependent aminoglycoside uptake. Divalent cations provided differential protection from neomycin and gentamicin, with high cation concentrations almost completely protecting hair cells from neomycin and acute gentamicin toxicity, but offering reduced protection from continuous (6 h) gentamicin exposure. These experiments lend further support to the hypothesis that aminoglycoside toxicity occurs via multiple pathways in a both a drug and time course-specific manner.
    Hearing research 04/2009; 253(1-2):42-51. DOI:10.1016/j.heares.2009.03.004 · 2.85 Impact Factor
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    • "OHC1–3 outer hair cells rows 1–3, IHC inner hair cells, Hsp70 OE Hsp70- overexpressing mice. Scale bar in l=25 μm and applies to all panels Hsp70 and aminoglycoside-induced hearing loss 433 induced hair cell death in vitro (Taleb et al. 2008). Therefore, we examined the utricles from the kanamycininjected mice in the current study. "
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    ABSTRACT: Sensory hair cells of the inner ear are sensitive to death from aging, noise trauma, and ototoxic drugs. Ototoxic drugs include the aminoglycoside antibiotics and the antineoplastic agent cisplatin. Exposure to aminoglycosides results in hair cell death that is mediated by specific apoptotic proteins, including c-Jun N-terminal kinase (JNK) and caspases. Induction of heat shock proteins (Hsps) can inhibit JNK- and caspase-dependent apoptosis in a variety of systems. We have previously shown that heat shock results in robust upregulation of Hsps in the hair cells of the adult mouse utricle in vitro. In addition, heat shock results in significant inhibition of both cisplatin- and aminoglycoside-induced hair cell death. In this system, Hsp70 is the most strongly induced Hsp, which is upregulated over 250-fold at the level of mRNA 2 h after heat shock. Hsp70 overexpression inhibits aminoglycoside-induced hair cell death in vitro. In this study, we utilized Hsp70-overexpressing mice to determine whether Hsp70 is protective in vivo. Both Hsp70-overexpressing mice and their wild-type littermates were treated with systemic kanamycin (700 mg/kg body weight) twice daily for 14 days. While kanamycin treatment resulted in significant hearing loss and hair cell death in wild-type mice, Hsp70-overexpressing mice were significantly protected against aminoglycoside-induced hearing loss and hair cell death. These data indicate that Hsp70 is protective against aminoglycoside-induced ototoxicity in vivo.
    Cell Stress and Chaperones 02/2009; 14(4):427-37. DOI:10.1007/s12192-008-0097-2 · 2.54 Impact Factor
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