缺氧誘導(dǎo)因子1α對休克血管反應(yīng)性的調(diào)控作用及其機制
發(fā)布時間：2018-09-09 18:16
【摘要】： 嚴(yán)重創(chuàng)傷、休克、膿毒癥病人在經(jīng)歷缺血缺氧、再灌注損傷以及腸道菌群移位、內(nèi)毒素釋放等的“多重打擊”后,常常在失代償期出現(xiàn)不可逆的組織細胞損傷,出現(xiàn)全身炎癥反應(yīng)綜合癥(SIRS)、急性呼吸窘迫綜合癥(ARDS)乃至多器官功能衰竭(MOF)。究其原因,除了全身炎癥失控外,休克后血管反應(yīng)性降低是另一重要原因。血管低反應(yīng)性表現(xiàn)為全身血管對縮血管物質(zhì)和舒血管物質(zhì)的反應(yīng)降低或反應(yīng)麻痹,導(dǎo)致血壓不能有效提升、組織灌注難以改善,細胞缺氧和損傷進行性加重。因此,血管低反應(yīng)性的發(fā)生一方面影響休克的發(fā)生和發(fā)展,另一方面嚴(yán)重地影響著創(chuàng)傷/休克的治療和轉(zhuǎn)歸。研究發(fā)現(xiàn)血管低反應(yīng)性的發(fā)生與腎上腺素能受體失敏、血管平滑肌細胞(VSMC)鉀、鈣通道功能失常及細胞膜超極化有關(guān)。此外,本實驗室前期研究也發(fā)現(xiàn)Rho激酶可通過調(diào)節(jié)肌球蛋白輕鏈磷酸酶(MLCP)的活性和肌球蛋白輕鏈(MLC20)磷酸化水平及鈣敏感性變化參與休克后血管反應(yīng)性的調(diào)節(jié)。 HIF-1α通過調(diào)節(jié)多種基因的表達來調(diào)節(jié)細胞對低氧的適應(yīng)性反應(yīng),是迄今為止發(fā)現(xiàn)的唯一的一個在缺氧狀態(tài)下發(fā)揮特異性活性的調(diào)節(jié)分子,被認(rèn)為是調(diào)節(jié)缺氧相關(guān)基因表達的關(guān)鍵分子,它同激活蛋白(activated protein 1, AP-1),核轉(zhuǎn)錄因子(nuclear factorκB, NF-κB)和p53等協(xié)同作用調(diào)節(jié)機體對缺氧的反應(yīng),其功能涉及能量代謝、細胞增殖、造血作用、血管形成、重塑與收縮等諸多方面。 缺血缺氧是各種類型休克的最基本的病理生理變化,故HIF-1α應(yīng)該是休克后表達和活性變化最顯著的分子之一。大量研究表明:失血性休克后期可激發(fā)明顯的失控性系統(tǒng)炎癥和缺血再灌注損傷, HIF-1α通過多種途徑參與了上述兩種損傷效應(yīng)的形成和發(fā)展。而有關(guān)HIF-1α與血管舒縮活性及反應(yīng)性的相關(guān)研究目前鮮有文獻報道。由此,本實驗選擇缺氧誘導(dǎo)因子1α(HIF-1α)及其一系列下游分子作為研究靶點,以HIF-1α特異性阻斷劑寡霉素為工具藥,探討HIF-1α對休克血管反應(yīng)性的調(diào)控作用。具體內(nèi)容包括兩大部分:⑴HIF-1α在失血性休克后血管反應(yīng)性變化中的調(diào)節(jié)作用;⑵失血性休克后HIF-1α調(diào)節(jié)血管反應(yīng)性的機制。 主要實驗方法: 第一部分觀察HIF-1α在失血性休克后血管反應(yīng)性變化中的調(diào)節(jié)作用,包括:建立大鼠重癥失血性休克模型,制備腸系膜上動脈(SMA)血管環(huán),利用離體血管環(huán)張力測定技術(shù),觀察休克后不同時相點(休克0.5h,休克1.0h,休克2.0h,休克3.0h,休克4.0h,休克6.0h)SMA對梯度濃度去甲腎上腺素(NE)的收縮反應(yīng)性。同時取腸系膜動脈,利用RT-PCR技術(shù)分析測定休克后各時相點HIF-1αmRNA表達變化。同時利用以HIF-1α特異性阻斷劑寡霉素為工具藥,觀察給藥后休克各時相點血管環(huán)張力和HIF-1αmRNA表達變化,進而分析HIF-1α基因轉(zhuǎn)錄水平對休克血管反應(yīng)性的影響。第二部分,通過在體和離體實驗,探討失血性休克后HIF-1α調(diào)節(jié)血管反應(yīng)性的機制。在體實驗:取大鼠失血性休克SMA組織,RT-PCR觀察休克后eNOS、iNOS、HO-1、COX-2 mRNA時相表達變化,同時分別采用連二亞硫酸鈉(Na2S2O4)還原法、硝酸還原酶法和放射免疫技術(shù)(RIA)測定休克后各時相血NO、CO、PGI含量變化,進而分析失血性休克后HIF-1α對eNOS / iNOS—NO、HO-1—CO和COX-2—PGI通路的影響。離體實驗:利用Transwell培養(yǎng)小室缺氧共培養(yǎng)血管平滑肌細胞(VSMC)和內(nèi)皮細胞(VEC),測定Transwell下腔熒光滲透率反映VSMC對去甲腎上腺素(NE)的收縮反應(yīng)性;同時利用RT-PCR半定量分析HIF-1α及其相關(guān)分子的mRNA表達變化規(guī)律:內(nèi)皮型一氧化氮合酶(eNOS)、誘生型一氧化氮合酶(iNOS)、環(huán)氧合酶-2(COX-2)、血紅素氧合酶-1(HO-1),進一步分析缺氧后VSMC收縮反應(yīng)的影響與上述分子的影響的關(guān)系。 主要研究結(jié)果: 1.失血性休克后HIF-1αmRNA表達逐漸增強,并于4h出現(xiàn)表達峰值(P 0.01)。血管反應(yīng)性呈現(xiàn)雙相變化,即早期(0.0h-1.0h)血管反應(yīng)性增高,量-效曲線左移、最大收縮力(Emax)顯著性增大和pD2減小(P 0.01)。至休克中、后期,血管反應(yīng)性進行性下降,表現(xiàn)為量-效曲線右移、Emax降低和pD2增大,至休克后4h血管反應(yīng)性即低于正常水平(P 0.01)。休克后給藥組血管反應(yīng)性亦呈現(xiàn)雙相變化,但其幅度明顯降低。表現(xiàn)為在早期(0.5-1.0h)血管反應(yīng)性的抬高趨勢被部分抑制(P 0.01),至休克晚期(4.0-6.0h)可使血管反應(yīng)性得以輕度回升(P 0.05或P 0.01)。 2.失血性休克后eNOS、iNOS、HO-1、COX-2 mRNA表達均隨時相的延長而逐漸增強,分別于1.0h、2.0h、3.0h和4.0h到達高峰(P 0.05或P 0.01)。給藥組eNOS、HO-1、COX-2 mRNA表達量基本波動于正常范圍。而iNOS則表現(xiàn)為休克早、中期(1.0h-3.0h)表達受抑制,而在晚期(4.0h-6.0h)較休克同時相有顯著增加(P 0.01)。 3.血漿NO、PGI和全血CO濃度均隨著休克時相進展而顯著升高(P 0.05或P0.01)。HIF-1α特異性阻斷劑寡霉素可使CO則穩(wěn)定于正常水平,而NO和PGI表達被部分抑制,隨著休克時相的延長有增高趨勢,但其幅度顯著低于休克同時相組(P 0.01)。 4.缺氧后VSMC收縮反應(yīng)性呈現(xiàn)雙相變化,即缺氧早期(0.0-1.0h) VSMC收縮反應(yīng)性顯著性增高且于0.5h達到峰值(P 0.01)。至缺氧后期(4.0-6.0h)VSMC收縮反應(yīng)性進行性下降,至4h血管反應(yīng)性即低于正常水平,至6h僅為正常對照的70% (P0.05)。給藥組則表現(xiàn)為缺氧后早期反應(yīng)性的增高趨勢被完全抑制(P 0.05或P 0.01),而4.0h-6.0h VSMC的收縮反應(yīng)性可有部分提高(P 0.05)。 5.VSMC+VEC混合培養(yǎng)后,隨著缺氧時相的延長, HIF-1α、iNOS、HO-1、COX-2 mRNA表達均逐漸增強,分別于4.0h、2.0h、3.0h和4.0h到達高峰(P 0.01或P 0.05)。給藥組各時相表現(xiàn)為上述mRNA表達增強趨勢被完全抑制,基本均波動于正常范圍內(nèi)。eNOS mRNA表達受缺氧和給藥處理方式的影響輕微,缺氧組和給藥組各時相表達量與正常對照相比均無顯著變化。 結(jié)論: 1.失血性休克后大鼠SMA收縮反應(yīng)呈雙相變化,即休克早期血管反應(yīng)性增高,晚期血管反應(yīng)性降低。HIF-1αmRNA表達隨休克時相進展逐漸上調(diào),至晚期進行性下降。HIF-1α阻斷劑寡霉素處理可對休克后血管反應(yīng)性的雙相變化產(chǎn)生雙相調(diào)節(jié)作用:在早期0.5-1h可部分抑制血管反應(yīng)性的抬高趨勢,至休克晚期可使血管反應(yīng)性得以輕度回升。HIF-1α在失血性休克大鼠血管收縮反應(yīng)性的雙相變化的形成中發(fā)揮了重要的調(diào)節(jié)作用:其轉(zhuǎn)錄水平在休克早期的輕度上調(diào)與血管收縮反應(yīng)性呈正相關(guān),至休克晚期則呈負相關(guān)。 2.休克后eNOS、iNOS、HO-1和COX-2 mRNA表達依次激活、上調(diào)并達峰值,在阻斷HIF-1α表達后其轉(zhuǎn)錄水平亦不同程度地受到明顯抑制。HIF-1α調(diào)節(jié)血管反應(yīng)性的途徑包括eNOS/iNOS—NO、HO-1—CO、COX-2—PGI等通路,通過影響具有血管舒縮功能調(diào)節(jié)作用分子的基因表達,從而引起血管舒縮活性物質(zhì)的生成和釋放的相應(yīng)變化:HS早期HIF-1α及其下游分子代償性增加,其輕度表達有利于血管反應(yīng)性的適應(yīng)性保護,為正相關(guān)因素。至休克失代償期轉(zhuǎn)化為負相關(guān):即HIF-1α過度激活、表達并蓄積,引起NO、CO、PGI等血管活性物質(zhì)的產(chǎn)生失調(diào)、釋放失控和作用過度,造成組織損傷。 3.缺氧刺激可引起VSMC對NE收縮反應(yīng)性類似在體實驗的雙相變化,阻斷HIF-1α表達可明顯削弱該雙相變化的幅度。HIF-1α參與了VSMC收縮反應(yīng)性的調(diào)節(jié),其機制主要與iNOS—NO、HO-1—CO、COX-2—PGI通路有關(guān)。
[Abstract]:Severe trauma, shock, sepsis patients often suffer from irreversible cell injury, systemic inflammatory response syndrome (SIRS), acute respiratory distress syndrome (ARDS) and even multiple organ failure (MOF) during decompensation after multiple blows such as ischemia and hypoxia, reperfusion injury, intestinal flora translocation, and endotoxin release. In addition to systemic inflammatory disorders, the decrease in vascular responsiveness after shock is another important cause. Vascular hyporesponsiveness is manifested by decreased or paralyzed responses of systemic vessels to vasoconstrictor and vasodilator substances, resulting in an ineffective increase in blood pressure, difficulty in improving tissue perfusion, and progressive increase in cellular hypoxia and injury. It has been found that the occurrence of vascular hyporeactivity is related to adrenergic receptor desensitization, vascular smooth muscle cell (VSMC) potassium, calcium channel dysfunction and membrane hyperpolarization. Pre-laboratory studies have also shown that Rho kinase can regulate vascular responsiveness after shock by regulating myosin light chain phosphatase (MLCP) activity, myosin light chain (MLC20) phosphorylation and calcium sensitivity.
HIF-1a regulates the adaptive response of cells to hypoxia by regulating the expression of a variety of genes. It is the only regulatory molecule that has been found to play a specific role in hypoxia. It is considered to be the key molecule to regulate the expression of hypoxia-related genes. It is associated with activated protein 1 (AP-1), nuclear transcription factor (nu). Clear factor kappa B, NF-kappa B) and p53 regulate the body's response to hypoxia. Their functions involve energy metabolism, cell proliferation, hematopoiesis, angiogenesis, remodeling and contraction.
Ischemia and hypoxia are the most basic pathophysiological changes in various types of shock, so HIF-1a should be one of the most significant changes in expression and activity after shock. A large number of studies have shown that late hemorrhagic shock can trigger obvious uncontrolled systemic inflammation and ischemia-reperfusion injury, and HIF-1a participates in the above two kinds of injury through a variety of pathways. In this study, hypoxia-inducible factor-1a (HIF-1a) and a series of its downstream molecules were selected as research targets, and the specific blocker of HIF-1a, oligomycin, was used as a tool drug to explore the vascular responsiveness of HIF-1a to shock. Specific contents include two major parts: _HIF-1a in the regulation of vascular reactivity after hemorrhagic shock; _after hemorrhagic shock HIF-1a regulation of vascular reactivity mechanism.
Main experimental methods:
In the first part, we observed the role of HIF-1 alpha in the regulation of vascular reactivity after hemorrhagic shock, including the establishment of severe hemorrhagic shock model in rats, the preparation of superior mesenteric artery (SMA) vascular ring, and the use of in vitro vascular ring tension measurement technique to observe the different phases of shock (shock 0.5 h, shock 1.0 h, shock 2.0 h, shock 3.0 h, shock 4.0 h, shock). Meanwhile, the mesenteric artery was harvested and the expression of HIF-1 alpha mRNA was detected by RT-PCR. Meanwhile, the tension of vascular rings and HIF-1 alpha mRNA expression were observed by using oligomycin, a specific blocker of HIF-1 alpha, as a tool drug. In the second part, the mechanism of HIF-1a regulating vascular reactivity after hemorrhagic shock was studied by in vitro and in vivo experiments. In vivo, SMA tissues from rats with hemorrhagic shock were harvested and the expression of eNOS, iNOS, HO-1, COX-2 mRNA was observed by RT-PCR. Sodium dithionite (Na2S2O4) reduction method, nitrate reductase method and radioimmunoassay (RIA) were used to determine the changes of NO, CO and PGI in blood at different stages after shock. The effects of HIF-1a on eNOS/iNOS-NO, HO-1-CO and COX-2-PGI pathway after hemorrhagic shock were analyzed. Vascular smooth muscle cells (VSMC) and endothelial cells (VEC) were used to measure the fluorescence permeability of the Transwell inferior cavity to reflect the contractile response of VSMC to norepinephrine (NE), and the mRNA expression of HIV-1 alpha and its related molecules was semi-quantitatively analyzed by RT-PCR: endothelial nitric oxide synthase (eNOS), inducible nitric oxide synthase (iNOS), cyclooxygenase (COX). - 2 (COX-2), heme oxygenase-1 (HO-1), further analyzed the relationship between the effects of hypoxia on the contractile response of VSMC and the effects of these molecules.
Main findings:
1. After hemorrhagic shock, the expression of HIF-1 alpha mRNA increased gradually and reached its peak at 4 h (P 0.01). Vascular responsiveness showed two-phase change, i.e. early (0.0h-1.0h) vascular responsiveness increased, dose-response curve shifted to the left, maximum contractility (Emax) increased significantly and pD2 decreased (P 0.01). Vascular responsiveness decreased progressively in the middle and late stages of the shock. The dose-response curve shifted to the right, Emax decreased and pD2 increased. Vascular responsiveness was lower than the normal level at 4 hours after shock (P 0.01). Vascular responsiveness also showed biphasic changes, but the amplitude was significantly decreased in the treatment group after shock. The reactivity of the tube was slightly increased (P 0.05 or P 0.01).
2. After hemorrhagic shock, the expression of eNOS, iNOS, HO-1, COX-2 mRNA increased gradually with the prolongation of the phase, reaching the peak at 1.0 h, 2.0 h, 3.0 h and 4.0 h respectively (P Compared with shock, there was a significant increase in phase (P 0.01).
3. Plasma NO, PGI and whole blood CO concentrations increased significantly with the progression of shock phase (P 0.05 or P 0.01). HIF-1a specific blocker oligomycin could stabilize CO at normal level, while NO and PGI expression were partially inhibited. With the prolongation of shock phase, NO and PGI expression increased, but the amplitude was significantly lower than that of shock phase group (P 0.01).
4. After hypoxia, the contractile reactivity of VSMC showed a biphasic change, that is, the contractile reactivity of VSMC increased significantly in the early stage of hypoxia (0.0-1.0 h) and reached a peak at 0.5 h (P The increase trend of early reactivity after hypoxia was completely inhibited (P 0.05 or P 0.01), while the contractile reactivity of VSMC at 4.0h-6.0h was partly increased (P 0.05).
5. After mixed culture of VSMC and VEC, the expression of HIF-1a, iNOS, HO-1, COX-2 mRNA increased gradually with the prolongation of hypoxia, reaching the peak at 4.0 h, 2.0 h, 3.0 h and 4.0 h respectively (P 0.01 or P 0.05). The enhanced expression of eNOS mRNA was completely inhibited and fluctuated within the normal range in the administration group. There was no significant difference between hypoxia group and administration group.
Conclusion:
1. After hemorrhagic shock, the contractile response of SMA in rats showed a biphasic change, that is, the vascular reactivity increased in the early stage of shock and decreased in the late stage of shock. HIF-1a partially inhibits the elevation of vasoconstrictive responsiveness at early 0.5-1 h and slightly increases vasoconstrictive responsiveness at late stage of shock. HIF-1a plays an important regulatory role in the formation of biphasic changes in vasoconstrictive responsiveness in hemorrhagic shock rats: its transcriptional level is slightly up-regulated at early stage of shock and exhibits vasoconstrictive responsiveness. Positive correlation was negative in late shock stage.
2. After shock, the expression of eNOS, iNOS, HO-1 and COX-2 mRNA was activated in turn, up-regulated and peaked. The transcriptional level of HIF-1 alpha was also significantly inhibited after blocking the expression of HIF-1 alpha. The expression of HIF-1a and its downstream molecules increased compensatively in the early stage of HS. The slight expression of HIF-1a was beneficial to the adaptive protection of vascular reactivity and was a positive correlation factor. The production of vasoactive substances such as PGI is out of control, releasing out of control and overacting, resulting in tissue damage.
3. Hypoxic stimulation can induce a biphasic change in the contractile response of VSMC to NE similar to in vivo experiment. Blocking the expression of HIF-1a can significantly weaken the biphasic change. HIF-1a participates in the regulation of contractile response of VSMC, and its mechanism is mainly related to iNOS-NO, HO-1-CO, COX-2-PGI pathway.
【學(xué)位授予單位】：第三軍醫(yī)大學(xué)
【學(xué)位級別】：碩士
【學(xué)位授予年份】：2007
【分類號】：R363
本文編號：2233184
[Abstract]:Severe trauma, shock, sepsis patients often suffer from irreversible cell injury, systemic inflammatory response syndrome (SIRS), acute respiratory distress syndrome (ARDS) and even multiple organ failure (MOF) during decompensation after multiple blows such as ischemia and hypoxia, reperfusion injury, intestinal flora translocation, and endotoxin release. In addition to systemic inflammatory disorders, the decrease in vascular responsiveness after shock is another important cause. Vascular hyporesponsiveness is manifested by decreased or paralyzed responses of systemic vessels to vasoconstrictor and vasodilator substances, resulting in an ineffective increase in blood pressure, difficulty in improving tissue perfusion, and progressive increase in cellular hypoxia and injury. It has been found that the occurrence of vascular hyporeactivity is related to adrenergic receptor desensitization, vascular smooth muscle cell (VSMC) potassium, calcium channel dysfunction and membrane hyperpolarization. Pre-laboratory studies have also shown that Rho kinase can regulate vascular responsiveness after shock by regulating myosin light chain phosphatase (MLCP) activity, myosin light chain (MLC20) phosphorylation and calcium sensitivity.
HIF-1a regulates the adaptive response of cells to hypoxia by regulating the expression of a variety of genes. It is the only regulatory molecule that has been found to play a specific role in hypoxia. It is considered to be the key molecule to regulate the expression of hypoxia-related genes. It is associated with activated protein 1 (AP-1), nuclear transcription factor (nu). Clear factor kappa B, NF-kappa B) and p53 regulate the body's response to hypoxia. Their functions involve energy metabolism, cell proliferation, hematopoiesis, angiogenesis, remodeling and contraction.
Ischemia and hypoxia are the most basic pathophysiological changes in various types of shock, so HIF-1a should be one of the most significant changes in expression and activity after shock. A large number of studies have shown that late hemorrhagic shock can trigger obvious uncontrolled systemic inflammation and ischemia-reperfusion injury, and HIF-1a participates in the above two kinds of injury through a variety of pathways. In this study, hypoxia-inducible factor-1a (HIF-1a) and a series of its downstream molecules were selected as research targets, and the specific blocker of HIF-1a, oligomycin, was used as a tool drug to explore the vascular responsiveness of HIF-1a to shock. Specific contents include two major parts: _HIF-1a in the regulation of vascular reactivity after hemorrhagic shock; _after hemorrhagic shock HIF-1a regulation of vascular reactivity mechanism.
Main experimental methods:
In the first part, we observed the role of HIF-1 alpha in the regulation of vascular reactivity after hemorrhagic shock, including the establishment of severe hemorrhagic shock model in rats, the preparation of superior mesenteric artery (SMA) vascular ring, and the use of in vitro vascular ring tension measurement technique to observe the different phases of shock (shock 0.5 h, shock 1.0 h, shock 2.0 h, shock 3.0 h, shock 4.0 h, shock). Meanwhile, the mesenteric artery was harvested and the expression of HIF-1 alpha mRNA was detected by RT-PCR. Meanwhile, the tension of vascular rings and HIF-1 alpha mRNA expression were observed by using oligomycin, a specific blocker of HIF-1 alpha, as a tool drug. In the second part, the mechanism of HIF-1a regulating vascular reactivity after hemorrhagic shock was studied by in vitro and in vivo experiments. In vivo, SMA tissues from rats with hemorrhagic shock were harvested and the expression of eNOS, iNOS, HO-1, COX-2 mRNA was observed by RT-PCR. Sodium dithionite (Na2S2O4) reduction method, nitrate reductase method and radioimmunoassay (RIA) were used to determine the changes of NO, CO and PGI in blood at different stages after shock. The effects of HIF-1a on eNOS/iNOS-NO, HO-1-CO and COX-2-PGI pathway after hemorrhagic shock were analyzed. Vascular smooth muscle cells (VSMC) and endothelial cells (VEC) were used to measure the fluorescence permeability of the Transwell inferior cavity to reflect the contractile response of VSMC to norepinephrine (NE), and the mRNA expression of HIV-1 alpha and its related molecules was semi-quantitatively analyzed by RT-PCR: endothelial nitric oxide synthase (eNOS), inducible nitric oxide synthase (iNOS), cyclooxygenase (COX). - 2 (COX-2), heme oxygenase-1 (HO-1), further analyzed the relationship between the effects of hypoxia on the contractile response of VSMC and the effects of these molecules.
Main findings:
1. After hemorrhagic shock, the expression of HIF-1 alpha mRNA increased gradually and reached its peak at 4 h (P 0.01). Vascular responsiveness showed two-phase change, i.e. early (0.0h-1.0h) vascular responsiveness increased, dose-response curve shifted to the left, maximum contractility (Emax) increased significantly and pD2 decreased (P 0.01). Vascular responsiveness decreased progressively in the middle and late stages of the shock. The dose-response curve shifted to the right, Emax decreased and pD2 increased. Vascular responsiveness was lower than the normal level at 4 hours after shock (P 0.01). Vascular responsiveness also showed biphasic changes, but the amplitude was significantly decreased in the treatment group after shock. The reactivity of the tube was slightly increased (P 0.05 or P 0.01).
2. After hemorrhagic shock, the expression of eNOS, iNOS, HO-1, COX-2 mRNA increased gradually with the prolongation of the phase, reaching the peak at 1.0 h, 2.0 h, 3.0 h and 4.0 h respectively (P Compared with shock, there was a significant increase in phase (P 0.01).
3. Plasma NO, PGI and whole blood CO concentrations increased significantly with the progression of shock phase (P 0.05 or P 0.01). HIF-1a specific blocker oligomycin could stabilize CO at normal level, while NO and PGI expression were partially inhibited. With the prolongation of shock phase, NO and PGI expression increased, but the amplitude was significantly lower than that of shock phase group (P 0.01).
4. After hypoxia, the contractile reactivity of VSMC showed a biphasic change, that is, the contractile reactivity of VSMC increased significantly in the early stage of hypoxia (0.0-1.0 h) and reached a peak at 0.5 h (P The increase trend of early reactivity after hypoxia was completely inhibited (P 0.05 or P 0.01), while the contractile reactivity of VSMC at 4.0h-6.0h was partly increased (P 0.05).
5. After mixed culture of VSMC and VEC, the expression of HIF-1a, iNOS, HO-1, COX-2 mRNA increased gradually with the prolongation of hypoxia, reaching the peak at 4.0 h, 2.0 h, 3.0 h and 4.0 h respectively (P 0.01 or P 0.05). The enhanced expression of eNOS mRNA was completely inhibited and fluctuated within the normal range in the administration group. There was no significant difference between hypoxia group and administration group.
Conclusion:
1. After hemorrhagic shock, the contractile response of SMA in rats showed a biphasic change, that is, the vascular reactivity increased in the early stage of shock and decreased in the late stage of shock. HIF-1a partially inhibits the elevation of vasoconstrictive responsiveness at early 0.5-1 h and slightly increases vasoconstrictive responsiveness at late stage of shock. HIF-1a plays an important regulatory role in the formation of biphasic changes in vasoconstrictive responsiveness in hemorrhagic shock rats: its transcriptional level is slightly up-regulated at early stage of shock and exhibits vasoconstrictive responsiveness. Positive correlation was negative in late shock stage.
2. After shock, the expression of eNOS, iNOS, HO-1 and COX-2 mRNA was activated in turn, up-regulated and peaked. The transcriptional level of HIF-1 alpha was also significantly inhibited after blocking the expression of HIF-1 alpha. The expression of HIF-1a and its downstream molecules increased compensatively in the early stage of HS. The slight expression of HIF-1a was beneficial to the adaptive protection of vascular reactivity and was a positive correlation factor. The production of vasoactive substances such as PGI is out of control, releasing out of control and overacting, resulting in tissue damage.
3. Hypoxic stimulation can induce a biphasic change in the contractile response of VSMC to NE similar to in vivo experiment. Blocking the expression of HIF-1a can significantly weaken the biphasic change. HIF-1a participates in the regulation of contractile response of VSMC, and its mechanism is mainly related to iNOS-NO, HO-1-CO, COX-2-PGI pathway.
【學(xué)位授予單位】：第三軍醫(yī)大學(xué)
【學(xué)位級別】：碩士
【學(xué)位授予年份】：2007
【分類號】：R363
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