성장인자수용체의 발현 ered solution(ph 7.5, 이하 PBS)과 4% paraform-alde- the first 28 N-Terminal residues of the mature human hyde를 각각 100 ml를 좌심실을 통해 관류 고정하였다. 단

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1 KISEP Otology Korean J Otolaryngol 1998;418: 기니픽와우에서성장인자수용체의분포 이병돈 박영진 안희영 장혁순 강주원 Expression of Receptor Tyrosine Kinases in the Guinea Pig Cochlea Byung Don Lee, MD, Young Jin Park, MD, Hyuck Soon Chang, MD, Hee Young Ahn, MD and Ju Won Kang, MD Department of Otolaryngology, College of Medicine, Soonchunhyang University, Seoul, Korea ABSTRACT Background and ObjectivesDamage in the auditory sensory epithelia is irreversible and the resulting hearing loss is permanent in mammals. Nevertheless, mammals like birds can recover their auditory function anatomically and physiologically after the sensorineural hearing loss. One of the functions of a receptor tyrosine kinase is to controll the cellular metabolisms like motility, growth, differentiation, regeneration, and ion transport via various pathways. There are no reports on the distribution of receptor tyrosine kinases in the guinea pig cochlea. The purpose of this study is to evaluate the expression of the fibroblast growth factor receptor FGFR and transforming growth factor receptor TGFR-, type II in the cochlear sensory epithelia of guinea pigs. Materials and MethodsWe investigated the immunoreactivity and functional roles of the receptor tyrosine kinases-fgfr and TGFR-type II - by using surface preparation technique in the cochlea of the Preyer s positive, pigmented guinea pig. ResultsThe results showed that receptor tyrosine kinases were expressed in the cytoplasm of Deiter s and Hensen s cells to FGFR and TGFR- type II, and the stereocillia of inner and outer hair cells to FGFR by surface preparation technique. ConclusionThe findings suggest that receptor tyrosine kinases are involved in the cellular growth, differentiation and regeneration in the cochlear supporting cells, and ion transport in the hair cells of the mammals. We found that receptor tyrone kinases are not related to outer hair cells participating in the PLC1-IP3 second messenger system. Korean J Otolaryngol 1998;418: KEY WORDSReceptor tyrosine kinase Signal transduction Auditory regeneration. 971

성장인자수용체의 발현 ered solution(ph 7.5, 이하 PBS)과 4% paraform-alde- the first 28 N-Terminal residues of the mature human hyde를 각각 100 ml를 좌심실을 통해 관류 고정하였다.

A)를 두 후 otic capsule을 열고 등골을 적출하고 정원창과 와우의 각각 1%의 BSA로 1 50과 1 5로 희석하여 4 에서 12 apical turn의 골편을 제거하여 24시간 동안 4% paraformaldehyde로 와우를 재고정하였다.

비특이적 결합을 방지하기 위 해 8% bovine serum albumin(이하 BSA)으로 2시간 반응 시킨 후, 일차 항체는 다클론성 IgG rabbit anti-chicken 섬 유아세포 성장인자 수용체 항체(rabbit polyclonal IgG, An ti-chicken FGF Receptor, residues 119-114

A) 및 rabbit anti-human 형질변환인자 수용체 항체(rabbit polyclonal IgG, Anti-Human TGF-β Receptor, type II, Fig. 1. Control. Surface preparation of cochlear sensory epithelia.

Strong immunoreactive staining was seen in Deiter s cells (arrow), not outer hair cells. IHC inner hair cell, OHC outer hair cell (mid-turn, ABC method, 1,000). 972 Fig. 3.

2 성장인자수용체의 발현 ered solution(ph 7.5, 이하 PBS)과 4% paraform-alde- the first 28 N-Terminal residues of the mature human hyde를 각각 100 ml를 좌심실을 통해 관류 고정하였다. 단 TGF-β receptor, Upstate Biotechnology. Inc., U.S.A)를 두 후 otic capsule을 열고 등골을 적출하고 정원창과 와우의 각각 1%의 BSA로 1 50과 1 5로 희석하여 4 에서 12 apical turn의 골편을 제거하여 24시간 동안 4% paraformaldehyde로 와우를 재고정하였다. 와우 고정 후 면역조직 화학 염색을 위해 실체 줌현미경과 드릴을 사용하여 와우의 외측벽과 혈관조를 박리하였다. 비특이적 결합을 방지하기 위 해 8% bovine serum albumin(이하 BSA)으로 2시간 반응 시킨 후, 일차 항체는 다클론성 IgG rabbit anti-chicken 섬 유아세포 성장인자 수용체 항체(rabbit polyclonal IgG, An ti-chicken FGF Receptor, residues of the deduced cdna sequence of the extracellular domain of the chicken FGF receptor, Upstate Biotechnology. Inc., U.S. A) 및 rabbit anti-human 형질변환인자 수용체 항체(rabbit polyclonal IgG, Anti-Human TGF-β Receptor, type II, Fig. 1. Control. Surface preparation of cochlear sensory epithelia. OHC outer hair cell, SC supporting cell (mid-turn, ABC method, 1,000). Fig. 2. Surface preparation of cochlear sensory epithelia stained with Anti-chicken FGF receptor. Strong immunoreactive staining was seen in Deiter s cells (arrow), not outer hair cells. IHC inner hair cell, OHC outer hair cell (mid-turn, ABC method, 1,000). 972 Fig. 3. Surface preparation of cochlear sensory epithelia stained with Anti-chicken FGF receptor. Strong immunoreactive staini-g was seen in Hensen s cells (arrow) (mid-turn, ABC method, 1,000). Fig. 4. Surface preparation of cochlear sensory epithelia stained with Anti-human TGF receptor. Strong immunoreactive staining was seen in Deiter s cells (arrow), not outer hair cells. OHC outer hair cell (mid-turn, ABC method, 1,000). Fig. 5. Surface preparation of cochlear sensory epithelia stained with Anti-human TGF receptor. Strong immunoreactive staining wa s seen in Hensen s cells (arrow) (mid-turn, ABC method, 1,000). Korean J Otolaryngol 1998;41(8):

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5 REFERENCES 1) Corwin JT. Postembryonic production and aging of inner ear hair cells in sharks. J Comp Neurol ) Corwin JT. Postembryonic growth of the macula neglect auditory detector in the Ray, Raja clavata continual increases in hair cell number, neural convergence, and physiological sensitivity. J Comp Neurol ) Cruz RM, Lambert PR, Rubel EW. Light microscopic evidence of hair cell regeneration after gentamycin toxicity in chick cochlea. Arch Otolar-yngol ) Lefebvre PP, Malgrange B, Staeker H, Moonen G, Van De Water TR. Retinoic acid stimulates regeneration of mammalian auditory hair cells. Science ) Ullrich A, Schlessinger J. Signal transduction by receptors with tyrosine kinase activity. Cell ) Hollenberg MD. Tyrosine kinase-mediated signal transduction pathways and the actions of polypeptide growth factors and G- protein-coupled agonists in the smooth muscle. Mol Cell Biochem / ) Bertorello AM, Katz AI. Short-term regulation of renal Na-K- ATPase activity Physiological relevance and cellular mechanisms. Am J Physiol ) Pardee AB. G1 Events and regulation of cell proliferation. Science ) Lefebvre PP, Van De Water TR, Staeker H, Weber T, Galinovic Schwartz V, Moonen G, et al. Nerve growth factor NGF stimulates neurite regeneration but not survival of adult auditory neurons in vitro. Acta Otolaryngol ) Moscatelli D, Presta M, Rifkin DB. Purification of a factor from human placenta that stimulates capillary endothelial cell protease production, DNA synthesis, and migration. Proc Natl Acad Sci USA ) Assoian RK, Komoriya A, Meyers CA, Miller DM, Sporn MB. Transforming growth factor- in human platelets. J Biol Chem ) Massague J. The transforming growth factor- family. Annu Rev Cell Biol ) Laiho M, De Caprio JA, Ludlow JW, Livingston DM, Massague J. Growth inhibition by TGF- linked to suppression of retinoblastoma protein phosphorylation. Cell ) Like B, Massague J. The anti-proliferative effect of type transforming growth factor occurs at a level distal from receptors for growth-activating factors. J Biol Chem ) Daniel TO, Gibbs VC, Milfay DF, Williams LT. Agents that increase camp accumulation block endothelial c-sis induction by thrombin and transforming growth factor-. J Biol Chem ) Oba T, Ogawa K, Lee BD, Kanzaki J. Heterogeneity of PLC in the cochlea of guinea pig. Proc Sendai Ear Symposium ) Corwin JT, Cotanch DA. Regeneration of sensory hair cells after acoustic trauma. Science ) Saunders JC, Tilney LG. Species differnces in susceptibility to noise exposure. In Hamernik RP, Henderson D, Salvi R. New York p ) Cousillas H, Rebillard G. Age-dependent effects of pure-tone trauma in the chick basilar papilla Evidence for development of the tonotopic organization. Hear Res ) Lee KH, Cotanche DA. Potential role of bfgf and retinoic acid in the regeneration of chicken cochlear hair cells. Hear Res

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169 KISEP Otology Korean J Otolaryngol 1998;412:168-173 기니픽에서 Cisplatin 이독성에대한 Fosfomycin 의예방적효과 김춘동 2 변성완 2 정원호 1 김종선 1 Preventive Effect of Fosfomycin on Cisplatin Induced Ototoxicity in Cochlea of Guinea