Cochlear delivery, of fibroblast growth factor 1 and its effects on apoptosis and cell cycling in noise-exposed guinea pig ears
ABSTRACT Acidic fibroblast growth factor 1 (FGF-1) is a mitogen and antiapoptotic factor synthesized by cochlear neurons and transported to the organ of Corti. The objectives of this investigation were threefold: (1) to develop an animal model to study the cochlear effects of intratympanic delivery of FGF-1; (2) to determine the distribution, in the mature mammalian cochlea, of FGF-1 and the receptor, FGFR3, to which it binds with high affinity; and (3) to examine the effect of exogenous FGF-1 on cochlear apoptotic and cell-cycling markers in noise and non-noise-exposed guinea pigs ears.
Fifteen adult Hartley guinea pigs were divided into three groups. Group 1 animals (n = 5) underwent direct placement of FGF-1 in phosphate buffered saline (PBS) (20 pg/mL) soaked Gelfoam pledgets to the right round window membrane. Phosphate buffered saline-soaked Gelfoam pledgets were placed on the left round window membrane as a control. In group 2 animals (n = 5), surgical placement of either FGF-1 or PBS was followed by exposure to 120 dB of white noise for 2 hours. Group 3 animals (n = 5) were subjected to identical noise conditions prior to undergoing round window application of either FGF-1 or PBS. All groups were allowed to recover in a noise-controlled environment for 12 hours following surgery. Anti-FGF-1-stained Western blots and optical densitometry analyses were used to quantitate passage of FGF-1 into cochlear perilymph. Standard in situ immunohistochemical techniques were used to stain each cochlea for FGF-1 and FGFR3, apoptotic markers p53 and p21, Bcl-2, and the cell-cycling marker proliferating cell nuclear antigen (PCNA). Tissue sections were subjected to the terminal deoxynucleotidyl transferase-mediated deoxyuridine triphosphate nick end-labelling technique (TUNEL) for apoptosis.
Western blot and optical densitometry analyses of cochlear perilymph showed increased concentrations of FGF-1 in 10 of 14 experimental cochleas. Cochlear perilymph FGF-1 was consistently bound to heparan sulphate proteoglycan (HSPG). Immunoreactivity of both FGF-1 and FGFR3 was observed in spiral ganglion neurons, inner and outer hair cells, pillar cells, and Dieter and Hensen's cells. Specific FGF-1 immunostaining to the distal portion of the pars pectinata of the basilar membrane was noted in noise-exposed animals only. Bcl-2 and PCNA immunostaining was not detected in any group. There was no significant nuclear immunoreactivity to proapoptotic markers, p53 and p21, in any group. Semiquantitative analysis of TUNEL staining in block sections of all cochleas demonstrated a 340% increase in nuclear immunoreactivity of noise-exposed outer hair cells and organ of Corti cells. There was no difference between FGF-1 treated and control ears subjected to TUNEL staining.
Exogenous FGF-1 crosses the round window membrane and is bound to HSPG in cochlear perilymph. The specific immunoreactivity of the pars pectinata to FGF-1 may represent a unique reservoir for cochlear FGF-1 in noise-exposed ears of the guinea pig. Noise induces apoptosis of organ of Corti cells as demonstrated with the TUNEL technique. PCNA, Bcl-2, p53, and p21 in noise-exposed and non-noise-exposed guinea pig cochleas are not affected by exogenous FGF-1. Noise-induced hair cell apoptosis appears to be independent of the p53 pathway. Lack of immunoreactivity to Bcl-2 supports the concept that the apoptotic mechanism is likely to involve C-Jun-N-terminal kinase- or caspase-dependent pathways. Exogenous FGF-1 does not alter apoptosis or cell cycling in the mature guinea pig cochlea within 12 hours of acute acoustic trauma.
- SourceAvailable from: Shujiro B Minami
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- "Acidic fibroblast growth factor (aFGF, or FGF 1 ) was similarly effective; delivered via osmotic mini-pump, it reduced PTS in guinea pigs (although TTS deficits were not reduced, an effect similar to that of antioxidant agents) (Sugahara et al., 2001). While the protective effects of FGF 1 /FGF 2 were not found in all studies (see Yamasoba et al., 2001;David et al., 2002), the effects of other NTF have generally been protective, with glial cell line derived neurotrophic factor (GDNF) (Ylikoski et al., 1998;Yamasoba et al., 1999) and NT3 (Shoji et al., 2000) shown to prevent noise-induced deficits in guinea pigs when infused chronically using osmotic mini-pumps. That BDNF (Shoji et al., 2000), and in some studies FGF 1 and FGF 2 (Yamasoba et al., 2001), did not reduce noise-induced injury may suggest the effect is growth factor specific, which could be a consequence of different NTF receptors on the hair cells (Ylikoski et al., 1993;Pirvola et al., 1997). "
ABSTRACT: Recent research has shown the essential role of reduced blood flow and free radical formation in the cochlea in noise-induced hearing loss (NIHL). The amount, distribution, and time course of free radical formation have been defined, including a clinically significant late formation 7-10 days following noise exposure, and one mechanism underlying noise-induced reduction in cochlear blood flow has finally been identified. These new insights have led to the formulation of new hypotheses regarding the molecular mechanisms of NIHL; and, from these, we have identified interventions that prevent NIHL, even with treatment onset delayed up to 3 days post-noise. It is essential to now assess the additive effects of agents intervening at different points in the cell death pathway to optimize treatment efficacy. Finding safe and effective interventions that attenuate NIHL will provide a compelling scientific rationale to justify human trials to eliminate this single major cause of acquired hearing loss.Hearing Research 05/2007; 226(1-2):22-43. DOI:10.1016/j.heares.2006.10.006 · 2.85 Impact Factor
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ABSTRACT: Lärmbelastungen in der Freizeit und dadurch bedingte Hörschäden haben bei Kindern, Jugendlichen und jungen Erwachsenen in den letzten Jahren deutlich zugenommen. Meist handelt es sich um Innenohrschäden nach Knalltraumata mit Spielzeugpistolen und Impulsschallbelastungen mit Rockmusik. Das Ziel der vorliegenden Arbeit war, die initialen und permanenten funktionellen und morphologischen Schäden nach solchen Schalltraumata erstmals systematisch an Meerschweinchen zu erarbeiten.Die Knalltraumata haben wir mit Spielzeugpistolen (163 bis 188 dB[lin], 8 Schüsse, 1/min, 10 cm Abstand zum linken Ohr, n=25) erzeugt, für die Impulsschallbelastungen verwendeten wir Rockmusik (Mittelungspegel 106 dB[A], 2×2,5 h, n=17). Zum Vergleich wurden auch Schallbelastungen mit einem Breitbandrauschen (115 dB[A], 2×2,5 h, n=18) durchgeführt. Die Hörschwellen haben wir mit der Hirnstammaudiometrie (f-BERA) bei 1,5; 2; 3; 4; 6; 8; 12 und 16 kHz vor und unmittelbar nach den Schallexpositionen sowie am 1., 2., 3., 5., 7. und 21. Tag bestimmt. Zudem wurden Zytokochleogramme mit Hilfe der Fluoreszenzmikroskopie erstellt, um die Haarzellschäden in 8 Frequenzregionen (26 kHz) unmittelbar nach den Schallbelastungen und am 1., 7. und 21. Tag zu quantifizieren. Nach den Knalltraumata waren jeweils 55% der äußeren Haarzellen (ÄHZ) in allen drei Reihen und 15–18% der inneren Haarzellen (IHZ) in der 3–5 kHz-Region irreversibel geschädigt. Bis zum 3. Tag kam es zu einer partiellen Erholung des Gehörs, danach persisierte ein permanenter maximaler Hörverlust um 32–34 dB bei 3–4 kHz. Nach den Beschallungen mit Breitbandrauschen kam es nur innerhalb von 24 Stunden zu einer partiellen Hörerholung, danach verblieb ein permanenter Hörverlust um 24–32 dB bei 1,5–8 kHz, ohne dass in diesem Frequenzbereich morphologische Schäden sichtbar waren. Auch nach den Impuls-Schallbelastungen mit Rockmusik kam es nur innerhalb von 24 Stunden zu einer partiellen Hörerholung, danach verblieb ein permanenter Hörverlust um 28–42 dB bei 6–16 kHz, wiederum ohne morphologische Schäden. Alle Versuchsergebnisse waren hochsignifikant (pAb dem 3. Tag nach einem Knalltrauma und ab dem 1. Tag nach den Beschallungen mit Breitbandrauschen oder Rockmusik kam es zu keiner weiteren spontanen Erholung des Gehörs. Den permanenten Hörverlusten nach Schallbelastungen mit Breitbandrauschen und Rockmusik liegen keine morphologisch sichtbaren Zilien- und Haarzellschäden zugrunde, was auf neue Aspekte permanenter zellulärer Funktionsstörungen im Hörsystem deutet.HNO 01/2004; 52(4). · 0.54 Impact Factor
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ABSTRACT: The organ of Corti in the mammalian inner ear is comprised of mechanosensory hair cells (HCs) and nonsensory supporting cells (SCs), both of which are believed to be terminally post-mitotic beyond late embryonic ages. Consequently, regeneration of HCs and SCs does not occur naturally in the adult mammalian cochlea, though recent evidence suggests that these cells may not be completely or irreversibly quiescent in at earlier postnatal ages. Furthermore, regenerative processes can be induced by genetic and pharmacological manipulations, but, more and more reports suggest that regenerative potential declines as the organ of Corti continues to age. In numerous mammalian systems, such effects of aging on regenerative potential are well established. However, in the cochlea, the problem of regeneration has not been traditionally viewed as one of aging. This is an important consideration as current models are unable to elicit widespread regeneration or full recovery of function at adult ages yet regenerative therapies will need to be developed specifically for adult populations. Still, the advent of gene targeting and other genetic manipulations has established mice as critically important models for the study of cochlear development and HC regeneration and suggests that auditory HC regeneration in adult mammals may indeed be possible. Thus, this review will focus on the pursuit of regeneration in the postnatal and adult mouse cochlea and highlight processes that occur during postnatal development, maturation, and aging that could contribute to an age-related decline in regenerative potential. Second, we will draw upon the wealth of knowledge pertaining to age related senescence in tissues outside of the ear to synthesize new insights and potentially guide future research aimed at promoting HC regeneration in the adult cochlea.Hearing research 11/2012; 297. DOI:10.1016/j.heares.2012.11.009 · 2.85 Impact Factor