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研究生:楊奕玲
研究生(外文):Yang Yi-Ling
論文名稱:熱休克蛋白對大白鼠熱中暑之保護作用
論文名稱(外文):The Heat Shock Protein Protects against Heatstroke in Rats
指導教授:林茂村林茂村引用關係
指導教授(外文):Lin, M.T.
學位類別:博士
校院名稱:國立成功大學
系所名稱:基礎醫學研究所
學門:醫藥衛生學門
學類:醫學學類
論文種類:學術論文
論文出版年:1998
畢業學年度:86
語文別:中文
論文頁數:134
中文關鍵詞:熱休克蛋白熱中暑
外文關鍵詞:Heat Shock ProteinHeatstroke
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為了探究熱中暑生成之機制,利用高效液相層析法(high performance
liquid chromatography:HPLC)量測熱中暑生成過程中腦內單胺類(
monoamine)及麩胺酸(glutamate)的變化。發現熱中暑過程中,腦內部份
區域(包括大腦皮質、紋狀體、下視丘等),其多巴胺(dopamine)、血清
張力素(serotonin)與麩胺酸之濃度於熱中暑生成過程中會顯著增加。事
先抑制腦中多巴胺受器之作用,如D1, D2受器拮抗劑(如SCH23390及
haloperidol)卻無法保護動物免於熱中暑所引起之腦缺血及神經細胞死
亡。然而若事先抑制腦中血清張力素受器之作用或麩胺酸受器之作用,
如5-HT2受器拮抗劑:ketanserine;NMDA受器拮抗劑:MK-801,
ketamine等則可保護動物免於熱中暑所引起之腦缺血及神經細胞死亡。
由此可知腦中多巴胺並非藉由其受器而造成神經細胞死亡,血清張力素
藉由其受器(5-HT2)而麩胺酸藉由其NMDA受器造成神經細胞死亡。
利用西方墨漬法(Western blot)分析不同時間下,熱休克蛋白於腦中各
區域(包括大腦皮質、小腦、紋狀體、視丘、下視丘、橋腦、延腦等構
造)、心肌、腎臟、脾臟、肝臟及骨骼肌等器官的表現。結果發現,於
誘發處理後四小時開始可偵測到熱休克蛋白的表現,誘發處理後十六至
二十四小時間,熱休克蛋白表現量為最大,而誘發處理四十八小時後,
即無法偵測到熱休克蛋白的表現。故探討熱休克蛋白保護機制之研究,
乃採取誘發處理後十六小時之大白鼠進行熱中暑之誘發實驗。
利用高效液相層析法,量測熱中暑生成過程中,對照組及事先誘發熱休
克蛋白表現組之腦內單胺類及麩胺酸的變化。發現熱中暑過程中,大腦
皮質、紋狀體、下視丘其多巴胺、血清張力素與麩胺酸之濃度於熱中暑
生成過程中會顯著增加。事先誘發熱休克蛋白生成則可有效保護動物免
於熱中暑所引起之腦缺血及神經細胞死亡。並可顯著延長其存活時間自
對照組的十五分鐘至三百分鐘。由此可知熱休克蛋白可有效藉由降低腦
中多巴胺、血清張力素及麩胺酸之濃度而保護神經細胞免於熱中暑所造
成之腦缺血及細胞壞死。除此之外,本實驗亦評估熱休克蛋白對熱中暑
生成時腦內局部血流量與多巴胺受體的鍵結能力之影響。利用雷射都卜
勒血流測定儀(Laser-Doppler flowmeter)及自動顯影測定法
(autoradiography)二種方法証實熱中暑生成時腦內局部血流量急遽降低
,致使腦缺血生成而熱休克蛋白事先誘發可顯著提高腦內局部血流量。
除此之外,利用高效液相層析法量測熱中暑生成過程中,對照組及事先
誘發熱休克蛋白表現組之腦內興奮性胺基酸的變化。發現熱中暑過程中
,大腦皮質、紋狀體、下視丘等區域,其麩胺酸之濃度於熱中暑生成過
程中會顯著增加,而事先誘發熱休克蛋白可有效降低細胞外麩胺酸之累
積,保護動物免於熱中暑所引起之腦缺血及大量麩胺酸之累積所造成之
神經細胞死亡。
利用電磁血流記錄器測定下列血流動力參數:心收縮量、平均動脈血壓
、心跳、心輸出量(cardiac output)、周邊血管阻力指數(total
peripheral resistance index, TPRI)、心搏量指數(stroke index,
SI)等。實驗結果指出熱中暑生成時所產生之循環系統衰竭肇因於心肌收
縮力降低因而導致心搏量、心輸出量下降,而周邊血管阻力在熱中暑生
成時並無顯著的變化。若事先於大白鼠體內誘發熱休克蛋白,可有效提
升大白鼠之心肌收縮能力,因而導致心搏量、心輸出量維持於一較熱中
暑生成時為高之值,而避免循環系統衰竭。
To elucidate the protection effect of heat shock protein 72-kDa
(HSP 72) in rat heatstroke, the following experiments were
under taken. Rats at 0 h, 16 h, or 48 h after the heat shock
(42oC core temperature for 15 min) or chemical stress
(5 mg/kg sodium arsenite, i.p.) were exposed to a high ambient
temperature (43 oC) to induce heatstroke onset. Heatstroke was
induced by exposing the animals to a high ambient temperature.
The moment at which the mean arterial pressure (MAP) and
striatal blood flow (SBF) decreased from their peak values was
taken as the time of heatstroke onset. HSP72 was detected by
western blot analysis. When compared with normothermic controls,
rats with heatstroke showed higher values of colonic
temperature, heart rate, but lower values of MAP, SBF, stroke
volume (SV), and cardiac output (CO). Prior heat shock or
chemical stress conferred significant protection against
heatstroke-induced arterial hypotension, decreased SV and CO,
cerebral ischemia, cerebral neuronal damage and death, and
correlated with expression of HSP72 in brain, heart, liver and
kidney at 16 h. However, at 48 h, when HSP72 expression
returned to basal values, the above responses that occurred
after the onset of heatstroke of two groups (0 h group VS 48 h
group) were indistinguishable.
Striatal neurons have been shown to be susceptible to cerebral
ischemia in rat heatstroke. Marked accumulation of monoamine
and glutamate in the corpus striatum has also been shown to
be associated with the heatstroke-induced striatal ischemic
insult. HSP72 induction protects neurons from ischemic insult.
This study has attempted to ascertain whether the ischemic
damage to striatal neurons and the augmented striatal
monoamine and glutamate release can be attenuated by HSP72
induction in rat heatstroke.
SBF, monoamines and glutamate release, and neuronal damage
score were assayed in rats with or without striatal HSP72
induction. Striatal monoamines and glutamate release were
determined by microdialysis combined with high-performance
liquid chromatography. During onset of heatstroke, rats
without HSP72 induction displayed higher values of colonic
temperature, striatal monoamines and glutamate release, and
striatal neuronal damage score but lower values of MAP and
SBF as compared with the normothermic control rats. The
diminished MAP and SBF, the accumulated striatal monoamine
and glutamate release and the elevated striatal neuronal
damage score during onset of heatstroke were significantly
attenuated by HSP72 induction. In addition, the survival
time (interval between the heatstroke onset and death) of the
rats with heatstroke was markedly extended by pretreatment
with HSP72 induction.
In another separate experiment when SCH23390 (2.5 mg/kg i.v.;
D1 receptor antagonist), haloperidol (5 mg/kg i.v.; D2
receptor antagonist) and NAN-190 (0.1 mg/kg i.v.; 5-HT1A
receptor antagonist) was administered 1h before the onset of
heatstroke, there is no significant change in the diminished
SBF, the augmented striatal monoamine release, the increased
striatal neuronal damage score and the shortened survival
time after the onset of heatstroke. However, pretreatment of
animal with MK-801 (1 mg/kg i.p.; NMDA receptor antagonist),
ketamine (50 mg/kg i.p.; NMDA receptor antagonist) or
ketanserine (50 mg/kg i.v.; 5-HT2 receptor antagonist) before
the onset of heatstroke, the diminished SBF, the augmented
striatal serotonin and glutamate release and the increased
striatal neuronal damage score during onset of heatstroke
were significantly attenuated. In addition, the survival
time (interval between onset of heatstroke and death) of
the rats with heatstroke was prolonged by pretreatment with
these receptor antagonists.
These results suggest that marked accumulation of monoamines
and glutamate in the corpus striatum is important for
developing ischemia to striatal neurons during heatstroke.
Pretreatment with MK-801, ketamine or ketanserine attenuates
the heatstroke formation resulting from striatal ischemic
damage via inhibiting NMDA and 5-HT2 receptor activity.
HSP72 synthesis attenuates the heatstroke-induced ischemic,
neuronal damage by reducing monoamines and glutamate
accumulations in the striatum.
封面
目錄
中文摘要
英文摘要
緒論
1-1熱中暑
1-2熱休克蛋白
1-3多巴胺
1-4血清張力素
1-5麩胺酸
1-6研究動機
實驗材料及方法
結果


討論
結論
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