熱休克前處置對血管通透度變化影響之探討

The effects of preconditioning heat shock treatment on vascular permeability alteration

呂宗禧(Tzong-Shi Lu)

指導教授 ： 楊瑞成

高雄醫學大學/醫學院/醫學研究所/博士(2004年)

摘要

體內環境恆定(Homeostasis)是生理系統中從細胞、組織到器官等各項不同作用所要統合協調及追求的目標，也是維持正常生理作用所需要的必要環境，其中，循環系統的恆定，是維持體內血液、養份、氣體、代謝產物及荷爾蒙等重要生理物質，是否能正確及有效的運送到其標的器官或細胞的一項重要影響因子；在影響循環系統恆定的因子中，血管通透度(vascular permeability)的恆定與否，即佔了一個舉足輕重的地位。熱休克效應是指先給一非致死性的熱休克前處置，可誘發耐受性的發生，可幫助生物體面對隨後而來的各種壓力所造成之傷害，利用熱休克處置誘發熱休克蛋白質產生，已被證實的確對繼發的致死性傷害具有保護作用。從近期的文獻資料中，我們發現血管通透度及熱休克蛋白質間具有相當程度的關聯，並且熱休克蛋白於血管通透度改變上所扮演的細胞保護性角色，確實是生物體內一項重要的生理性保護機制，但其二者之相關性，及其相關作用機制仍有待探討。而血管的特性及部位不同，在血管通透度上會有不同的影響因子。所以，我們選擇了三個不同的實驗動物及細胞培養研究模式，從不同面向針對熱休克蛋白在血管通透性變化中所扮演的保護性角色及作用機制加以探討。首先，我們採用過敏性休克大白鼠動物模式誘發之血管通透度改變來探討週邊循環系統中，熱休克前處置對其變化的影響。第二項，我們採用血腦障壁破壞大白鼠動物模式誘發之血管通透度改變來探討中樞循環系統中，熱休克前處置對其變化的影響。最後，我們採用組織氨誘發之血管內皮細胞完整性改變之體外研究模式，對熱休克前處置影響血管通透度的機制作一探討。由結果發現，熱休克處置可藉由保持大鼠之週邊循環系統血管通透度，以減低其微血管蛋白質的滲漏，而使過敏性休克大鼠的平均動脈壓迅速回升並維持其生命跡象。其次，熱休克處置可減低血腦障壁破壞大鼠腦部微血管蛋白質的滲漏，可能之機制為熱休克蛋白質72可以和腦部血管內皮細胞緊密接合蛋白質(Zo-1和occludin)有連結作用，並幫助其於遭受高張溶液破壞下，仍保持其生理功能及結構之穩固，而維持血腦障壁的完整，進而保持中樞循環系統血管通透度的維持。而在體外培養之血管內皮細胞，可以觀察到熱休克前處置可誘發大量熱休克蛋白質72的合成及表現，並可以連結在緊密接合蛋白Zo-1或Occludin，扮演分子伴護者的角色，使緊密接合蛋白質在遭受破壞或壓力的情況下，仍保有正常之構型及生理功能，進而可穩定血管內皮細胞之細胞骨架及穩固緊密接合蛋白質，進而保持血管內皮細胞之完整性。由我們以上結果可以得知: 熱休克處置可以成功的誘發生物體內細胞或培養的內皮細胞中熱休克蛋白72的合成；並且，熱休克蛋白72可以和緊密接合蛋白Zo-1或Occludin連結，並扮演分子伴護者的角色，保護內皮細胞的正常之構型及緊密結合的正常生理屏障功能，確實使生物體在血管通透度變化之病理情況威脅之下，仍能維持正常之循環系統血管通透度，維持體內環境恆定性，進而維持生物體內正常生理作用所需要的必要環境，免於血管通透度變化所產生之病理情況威脅所造成對生物體的傷害。

關鍵字

熱休克蛋白 ； 緊密結合蛋白 ； 血管通透度 ； 內皮細胞 ； 血腦障壁

並列摘要

The homeostasis of the internal microenvironment plays an important role since it is related to many systemic physiological functions and the homeostasis of the circulation system plays the most important part of the body. The stabilization of the vascular permeability acts as an important factor among all risk factors that influence the homeostasis of circulation system. For decades, heat shock proteins (HSPs), induced by previous heat shock treatment have been well documented. They are characterized as a family of highly conserved and universally inducible proteins found in almost all organisms and cultured cells in response to stress. These protective proteins act against subsequent damage by increasing the tolerance of the organisms, known as thermotolerance or cross-tolerance phenomena. The studies indicated that the HSPs could play a protective role during vascular permeability change but the real mechanism is still obscured. In the study, we investigated the protective effects and probably mechanisms of HSPs that induced by preconditioning heat shock treatment during vascular permeability change with three different models. First, the anaphylactic shock animal model was used to determine whether induction of HSPs leads to decreased protein leakage and attenuated hypotension in anaphylactic rats. Second, we investigated the effects of heat shock treatment on Blood-Brain Barrier (BBB) permeability by evaluating the extravasation of macromolecular tracer, Evans blue, in rats whose BBB was opened by hyperosmotic solution infusion and the possible mechanism was also studied. The last study is the human umbilical vein endothelial cells (HUVEC) monolayer model that used for investigate the potential protective effects of heat shock proteins, induced by heat shock treatment, on the histamine-induced HUVEC integrity alteration and the probably mechanism was also studied. The results show that the reduction of increased protein leakage and attenuation of hypotension during anaphylactic shock may result from the induction of HSPs by preconditioning heat shock treatment. The other result indicated that HSP72 induced by heat shock treatment plays a role in stabilizing the tight junction proteins of the endothelial cells and subsequently maintain the BBB integrity in BBB-opening condition caused by hyperosmotic stress. In HUVEC monolayer model, HSPs, induced after heat shock treatment, also play a role in stabilizing the tight junction proteins and cytoskeleton of the endothelial cells and subsequently preserving the HUVEC integrity during a adverse condition under histamine treatment. In conclusion, the homeostasis could be maintained by HSPs, induced by preconditioning heat shock treatment, work as a molecular chaperone that stabilize the tight junction proteins and cytoskeleton and preserve the integrity of BBB or HUVEC and subsequently the maintenance of vascular permeability during pathological condition.

並列關鍵字

heat shock protein ； vascular permeability ； blood brain barrier ； endothelial cell ； tight junction protein

參考文獻

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