1993 Fiscal Year Annual Research Report

SV40TS遺伝子導入による不死化カテコラミン産生細胞株の開発と脳内移植への応用

Research Project

Project/Area Number	04557005
Research Institution	Nagoya City University
Principal Investigator	西野仁雄名古屋市立大学, 医学部, 教授 (60073730)
Co-Investigator(Kenkyū-buntansha)	源良樹味の素, 中央研究所, 主任研究員小幡邦彦岡崎国立共同研究機構生理学研究所, 教授 (60013976) 古山富士弥名古屋市立大学, 医学部, 助手 (00080101)
Keywords	遺伝子導入 / 不死化 / カテコラミン細胞 / 神経移植 / 機能修復
Research Abstract	1.胎仔(E14-15)ラット脳幹及び新生仔ラット副腎より得た組織を酵素処理後細胞浮遊液とし、SV40T/TS遺伝子を導入されたpsi2細胞シート上あるいはpsi2細胞条件培養液添加培地で培養し、脳幹ドーパミン参謀及び副腎髄質細胞にSV40T/TS遺伝子を導入した。 2.SV40T/TS遺伝子導入後の増殖期(4週間)に、培地にEGF,FGFあるいは線条体細胞条件培養液を加えると、1の細胞の増殖が促進された。 3.遺伝子導入及び増殖後にG418でセレクッションし、G418耐性の脳幹細胞(MS-1,2,3)及び副腎細胞(AD-1,2)を得た。 4.これらの細胞は、33℃では3〜4週間でほぼ倍増した(doubling time3〜4週間)。また38℃では増殖不能であった(温度変異性)。 5.チロシン水酸化酵素(TH)免疫染色性は、MS-1とAD-1が陽性であった。MS-2,3はneurofilament及びGFAPともに陽性であった。 6.MS-1及びAD-1をパーキンソン病モデルラットの線条体に移植した。両細胞ともに移植後2週間迄は生着したが、4週間後にはTH免疫活性が低下し、運動機能の改善をもたらす迄には至らなかった。以上、脳幹ドーパミン細胞及び副腎髄質細胞にSV40T/TS遺伝子を導入し、in vitroで増殖能をもつ細胞を得ることができた。しかしこれらの細胞は脳内移植後は長期にわたり安定して生存することができず、機能修復を得る迄には至らなかった。これらの結果は、脳内移植後、機能改善をもたらすような細胞株を得るには、in vivoにおいて長期にわたり安定した遺伝子発現を可能とする内因性(native)のpromoterの選択が不可欠であることを意味する。現在、この点について探索中である。

Research Products

(18 results)

All Other

All Publications (18 results)

[Publications] Karadi,Z.: "Responses of lateral hypothalamic glucose-sensitive and glucose- insensitive neurons to chamical stimuli in behaving rhesus monkeys" J.Neurophysiol.76. 389-400 (1992)
[Publications] Furuyama,F.: "Overshoot and high equilibrium in body temperature responses of rats to ambient heat" J.Vet.Med.Sci.54. 915-921 (1992)
[Publications] Hashitani,T.: "Sialyl cholesteral enhances the development of grafted neurons and motor recovery" Brain Ros.Bull.29. 795-806 (1992)
[Publications] Hattori,S.: "Treadmill running test for evaluating locomotor activity after 6-OHDA lesions and dopaminergic cell grafts in the rat" Brain Res.Bull.31. 433-435 (1993)
[Publications] Nishino,H.: "Striatal grafts in the ischemic striatum improve pallidal GABA release and passive avoidance" Brain Res.Bull.32. 517-520 (1993)
[Publications] Koide,K.: "Improvement of passive avoidance task after grafting of fetal striatal cell suspensions in ischemic striatum in the rat" Restor.Neurol.Neurosci.5. 205-214 (1993)
[Publications] Nishino,H: "Reconstruction of GABA ergic transmission and behavior by striatel cell grafts in rats with ischemic infarcts in the middle cerebrel artery" J.Neural Transp.Plast.4. 147-155 (1993)
[Publications] Nishino,H.: "Intracerebral grafting of catecholanine producing cells and roconstruction of disturbed brain function" Neurosci.Res.16. 157-172 (1993)
[Publications] Hattori,S.: "Treadmill running combined with microdialysis can evaluate mator daficit and improvement following dopaminergic grafts in 6-OHDA lesioned rats" Restor.Neurol.Neurosci.6. 65-72 (1993)
[Publications] Czurko,A.: "‘Collapsed'(argyrophilic,dark)neurons in rat model of fransient focal cerbrol ischemia" Neurosci.Lett.162. 71-74 (1993)
[Publications] Furuyama,F.: "Genetic development of an inbred rat strain with increased resistance adaptation to a hot environment" Am.J.Physiol.265. R957-R962 (1993)
[Publications] Scott,T.R.: "Gustatory neural coding in the amygdala of the alert macaque monkey" J.Neurophysiol.69. 1810-1820 (1993)
[Publications] Hashimoto,N.: "Dopamine has inhibitory and accelerating effects on ischemia-induced neuronal cell damage in the hot striatum" Brain Res.Bull.33. 281-288 (1994)
[Publications] Czurko,A.: "Appearance of immunoglobulin G and complement factor C3 in the striatum after transient focal ischemia in the rat" Neurosci.Lett.166. 51-54 (1994)
[Publications] Aihara,N.: "Striatal grafts in infarct striatspallidum incresse GABA release,reorganize GABA_A receptor and improve water-maze learning in the rat" Brain Res.Bull.(in press) (1994)
[Publications] Nishino,H.: "Pathsphysislogical process after transient ischemia of the middle cerebral artery in the rat" Brain Res,Bull.(in press) (1994)
[Publications] Zhou,X.R.: "Amygdala kindling activates the phosphorylation of ca^〓/calmodulin-dependent protein kinase II in rat hippocampus" Neurosci.Lett.(in press) (1994)
[Publications] Hattorei,S.: "Striatal dopamine turnover during treadmill running in the rat:relation to the speed of running" Brain Res.Bull.(in press) (1994)

1993 Fiscal Year Annual Research Report

SV40TS遺伝子導入による不死化カテコラミン産生細胞株の開発と脳内移植への応用

Principal Investigator

西野 仁雄 名古屋市立大学, 医学部, 教授 (60073730)

Research Products

[Publications] Karadi,Z.: "Responses of lateral hypothalamic glucose-sensitive and glucose- insensitive neurons to chamical stimuli in behaving rhesus monkeys" J.Neurophysiol.76. 389-400 (1992)

[Publications] Furuyama,F.: "Overshoot and high equilibrium in body temperature responses of rats to ambient heat" J.Vet.Med.Sci.54. 915-921 (1992)

[Publications] Hashitani,T.: "Sialyl cholesteral enhances the development of grafted neurons and motor recovery" Brain Ros.Bull.29. 795-806 (1992)

[Publications] Hattori,S.: "Treadmill running test for evaluating locomotor activity after 6-OHDA lesions and dopaminergic cell grafts in the rat" Brain Res.Bull.31. 433-435 (1993)

[Publications] Nishino,H.: "Striatal grafts in the ischemic striatum improve pallidal GABA release and passive avoidance" Brain Res.Bull.32. 517-520 (1993)

[Publications] Koide,K.: "Improvement of passive avoidance task after grafting of fetal striatal cell suspensions in ischemic striatum in the rat" Restor.Neurol.Neurosci.5. 205-214 (1993)

[Publications] Nishino,H: "Reconstruction of GABA ergic transmission and behavior by striatel cell grafts in rats with ischemic infarcts in the middle cerebrel artery" J.Neural Transp.Plast.4. 147-155 (1993)

[Publications] Nishino,H.: "Intracerebral grafting of catecholanine producing cells and roconstruction of disturbed brain function" Neurosci.Res.16. 157-172 (1993)

[Publications] Hattori,S.: "Treadmill running combined with microdialysis can evaluate mator daficit and improvement following dopaminergic grafts in 6-OHDA lesioned rats" Restor.Neurol.Neurosci.6. 65-72 (1993)

[Publications] Czurko,A.: "‘Collapsed'(argyrophilic,dark)neurons in rat model of fransient focal cerbrol ischemia" Neurosci.Lett.162. 71-74 (1993)

[Publications] Furuyama,F.: "Genetic development of an inbred rat strain with increased resistance adaptation to a hot environment" Am.J.Physiol.265. R957-R962 (1993)

[Publications] Scott,T.R.: "Gustatory neural coding in the amygdala of the alert macaque monkey" J.Neurophysiol.69. 1810-1820 (1993)

[Publications] Hashimoto,N.: "Dopamine has inhibitory and accelerating effects on ischemia-induced neuronal cell damage in the hot striatum" Brain Res.Bull.33. 281-288 (1994)

[Publications] Czurko,A.: "Appearance of immunoglobulin G and complement factor C3 in the striatum after transient focal ischemia in the rat" Neurosci.Lett.166. 51-54 (1994)

[Publications] Aihara,N.: "Striatal grafts in infarct striatspallidum incresse GABA release,reorganize GABA_A receptor and improve water-maze learning in the rat" Brain Res.Bull.(in press) (1994)

[Publications] Nishino,H.: "Pathsphysislogical process after transient ischemia of the middle cerebral artery in the rat" Brain Res,Bull.(in press) (1994)

[Publications] Zhou,X.R.: "Amygdala kindling activates the phosphorylation of ca^〓/calmodulin-dependent protein kinase II in rat hippocampus" Neurosci.Lett.(in press) (1994)

[Publications] Hattorei,S.: "Striatal dopamine turnover during treadmill running in the rat:relation to the speed of running" Brain Res.Bull.(in press) (1994)

西野仁雄名古屋市立大学, 医学部, 教授 (60073730)