| 摘要: | 背景：創傷出血休克常造成死亡及器官衰竭。休克造成組織及細胞缺氧，誘發組織缺氧及再灌流傷害，產生過氧化物自由基，引起組織進一步傷害及發炎反應。休克常導致肝臟衰竭。
近期吾等以大白鼠之出血性模式應用DNA微陣列研究,發現休克與輸液復甦其基因體變化甚為相似，僅有葡萄糖-六-磷酸去氫酶 (glucose-6-phosphate dehydrogenase, G6PD) 於出血休克及輸液治療後24小時呈現有意義減少。G6PD為細胞關鍵調節酶，於細胞內氧化還原反應提供還原NADPH以維細胞恆定。麩氨基硫(glutathione)能清除過氧化基，是肝細胞抗過氧化劑(antioxidant)能力的指標。除此之外，G6PD產生之NADPH經由Nox途徑產生自由基。因此吾等推論G6PD 在出血性休克後與輸液復甦中扮演一個重要角色，值得進一步探討。
方法：雄性大白鼠共分三組，第一組：控制組，第二組：休克組，第三組：休克及復甦組。每組分為6次組(實驗開始0小時、1.5小時、4小時、8小時、12小時、24小時)。出血性休克採用大白鼠出血固定血壓模式，維持固定平均動脈壓為40±5毫米汞柱達一小時。復甦組給予10毫升血液/公斤體重之乳酸林格液使穩定。實驗結束給予安樂死，同時將大白鼠由心臟抽血並且將肝臟組織取下。檢體保存於-80℃冰箱直到進一步實驗分析。血中細胞激IL-6，IL-10及G6PD活性，麩氨基硫(氧化態及還原態)及肝組織麩氨基硫麩氨基硫(氧化態及還原態) 及G6PD予以檢測。
結果與討論：血液G6PD活性在休克組及復甦組早期即增加;並維持較控制組為高之活性。復甦液給與並未在24小時讓血液內G6PD活性增加的情形有所改變或消失。創傷出血休克後肝臟組織G6PD定量先增加後下降。復甦介入降低肝臟組織之氧化壓力;也抑制肝臟組織G6PD在出血休克後增加之反應。創傷出血休克改變大白鼠血液內氧化壓力狀態；血液還原型麩氨基硫(GSH)及血液GSH/GSSG 比值在休克後不久即走低並持續12-24小時。復甦液給與改善血中部分氧化壓力。創傷出血休克亦改變肝臟組織氧化壓力，但肝臟組織對於氧化壓力變化之反應殊於血液；肝臟組織的還原型麩氨基硫(GSH)及GSH/GSSG 比值先下降;但在8-12小時點快速增加。然而復甦組並未出現此情形。發炎細胞激素IL6及抗發炎細胞激素IL10在創傷出血休克之後，最早的幾個小時內顯著增加，但只維持8-12小時。復甦液給與抑制發炎細胞激素IL6增加；然而復甦液給與只部份抑制抗發炎細胞激素IL10增加。
結論：創傷出血休克改變血液及肝臟組織的氧化壓力。也改變並調節血液中及肝臟G6PD之反應。但血液及肝臟之G6PD反應是有所不同。創傷出血休克後,血液中氧化壓力大增;血液中G6PD活性也增加;血液中發炎細胞激素IL6及抗發炎細胞激素IL10也增加。林格氏液復甦介入, 抑制血液中發炎細胞激素IL6上升之反應及抑制部份血液中抗發炎細胞激素IL10上升之反應;也降低血液中部份氧化壓力。但並未明顯降低血液G6PD活性增加之情形。相對地,出血休克後, 肝臟組織氧化壓力先增後降; 肝臟組織G6PD也是先增後降。林格氏液復甦介入降低肝臟組織之氧化壓力;也抑制肝臟組織G6PD對出血休克後之增加反應;這個反應異於血液中之情形。
Background:
Trauma-induced massive hemorrhage lead to development of tissue hypoxemia and ischemia/reperfusion injury. Cellular injury and oxidative stress initiate post-injury priming of neutrophils and subsequent severe SIRS response. During the hemorrhage shock with or without immediate resuscitation, rat animal model, we had observed that the up-regulation of G6PD gene expression disappeared after immediate resuscitation compared with no immediate resuscitation group. We hypothesize that G6PD provides reducing power, NADPH for reduction of GSSG to GSH to eliminate free radicals and reduce oxidative stress. However, G6PD also provide NADPH to generate more free radicals via Nox pathway. So G6PD may play an important role in the early shock with ischemia and reperfusion injury and subsequent immune response, development of multiple organ failure (MOF).
Methods:
108 mature SD male rats will be randomized into three groups ( Gr.I: control group; Gr.II : hemorrhagic shock without resuscitation(HS); Gr.III: hemorrhagic shock with resuscitation(HS+R)). Rat hemorrhagic shock model /c fixed pressure model would be performed . The animals will be sacrificed by euthanasia and the whole blood and liver tissue will be collected at 0, 1.5, 4, 8, 12, and 24 hours after the initiation of experiment. The whole blood sample and liver tissue will be collected and frozen until further studies. G6PD activity, GSH and GSSG will be analyzed in blood and liver tissues. The plasma levels of inflammation cytokine IL6 and anti-inflammation cytokine IL10 would be tested.
Result and Discussion:
Blood G6PD activity increased early in HS group and HS+R group rats, compared with control group rats. There is significant difference of blood G6PD activity between shock group and control group and between HS+R group and control group. Resuscitation dose not alter the increasing response of blood G6PD activity. Liver tissue G6PD measured by Western blot method increased early and sharply in HS group and lasted for about 4 hours. Then it dropped markedly and it was below the level of control group latter. Resuscitation suppressed the increasing response of liver tissue G6PD markedly and it made obvious different response between the blood and liver tissue.
Shock episode induced change of blood oxidative stress status; whole blood GSH and whole blood GSH/GSSG ratio responded and went down early; it lasted for 12-24 hours. Resuscitation improved partial oxidative stress in blood. Oxidative stress condition also changed by shock episode in liver tissue and its response to the oxidative stress was differed from the whole blood. Increase of liver tissue GSH noted but delayed till the time-8-12-hour point. Resuscitation did not alter or change the liver tissue GSH in the early 12 hours, compared with the control group.
Inflammation cytokine IL6 and anti-inflammation cytokineIL10 increased markedly after shock episode in the early several hours but they disappeared after the time-12-hour point. Resuscitation made the increase of IL6 disappeared after shock episode in the early several hours. But IL10 still increased but less obvious than shock group after resuscitation given in the early several hours.
Conclusion:
The traumatic hemorrhagic shock episode changed the oxidative stress condition in blood and liver tissue. It also changed the blood G6PD enzyme activity and liver tissue G6PD. However, the G6PD response of liver tissue was different from the response of blood while confronting with the traumatic hemorrhagic shock episode. Resuscitation dose not alter the change of blood G6PD activity. But it dose alter the G6PD response in liver tissue. Therefore, we supposed that the response and the mechanism of G6PD and GSH were different between the blood and liver tissue while confronting with the traumatic hemorrhagic shock episode. But it needs more studies to prove it. |
