Changes And Organ Diversity Of Vascular Reactivity And The Roles Of INOS And ET-1 Following Endtoxic Shock In Rabbits

It was shown that there is vascular hyporeactivity in the late period of various shock including hemorrhagic shock and septic shock. Vascular hyporeactivity is that blood vessels have lower or no reaction to vasoconstrictors or vasodilatators, and contribute to the failure of boosting pressure and improving tissue perfusion, then lead to progressive cell hypoxia and damage. Researches showed that vascular hyporeactivity is related to receptor desensitization, vascular smooth muscle cell hyperpolarization and calcium desensitization, and so on.Endtoxic shock is the main complication of Gram negative bacteremia infection, and characterized with systemic inflammation reaction syndrome (SIRS). The uncontrolled expression of inflammatory factors lead to microcirculation disorder, vascular hyporeactivity, systemic persistent hypotension and organ hyporeperfusion, even multiple organ dysfunction syndrome(MODS) eventually. At present, it is generally considered that vascular hyporeactivity is due to uncontrolled expression of various cytokies induced by lipopolysaccharide(LPS), such as overproduction of nitric oxide (NO), endothelin-1 (ET-1), tumor necrosis factorα(TNF-α), interleukin-1(IL-1), nuclear factor-κB (NF-κB), migration inhibitory factor(MIF), platelet activating factor(PAF), and so on, which lead to damage of vascular endothelium, vascular constriction and dilation function.Our previous researches showed that there was biphasic change of vascular reactivity following hemorrhagic shock, that is to say, hyperreactivity in early stage and hyporeactivity in late stage. And there were organ diversity of vascular reactivity following hemorrhagic shock, which perhaps related to the different expression of various inflammatory factors and vasoactive factors in organs. There is vascular hyporeactivity following endtoxic shock too, but we know less about the changes and organ diversity of vascular activity and its related mechanism following endtoxic shock. So we used rabbit model of endtoxic shock, took animals, superior mesenteric artery(SMA), celiac artery(CA) and left renal artery(LRA) as research object, took aminoguanidine(AG, specific inhibitor of inducible nitric oxide synthase) and PD-142893(nonselective inhibitor of ET-1 receptors) as tool drugs, observed the changes and organ diversity of vascular activity and its related mechanism following endtoxic shock via monitoring animals'hemodynamic changes, the tension of vascular rings and gene expression of inducible nitric oxide synthase (iNOS) and ET-1. The experiments were conducted in two parts as follows: (1) the changes and organ diversity of vascular reactivity following endtoxic shock; (2) the roles of iNOS and ET-1 in the changes of vascular reactivity following endtoxic shock.Methods:In the first part, we observed the changes and organ diversity of vascular reactivity following endtoxic shock. We established rabbit model of endtoxic shock via LPS intravenous injection. The changes of hemodynamic indexes following LPS injection were observed. Then SMA, CA and LRA from normal rabbits and rabbits at different time after LPS injection (0.5h, 1h, 2h, 4h and 6h after LPS injection) were adopted to analyze the vascular reactivity via observing the contraction initiated by norepinephrine(NE) and the relaxation initiated by acetylcholine(Ach) in isolated organ perfusion system.In the second part, we explored the roles of iNOS and ET-1 in the changes of vascular reactivity following endtoxic shock. We adopted SMA, CA and LRA tissues from normal rabbits and rabbits at different time after LPS injection (0.5h, 1h, 2h, 4h and 6h after LPS injection). Then iNOS and ET-1 mRNA expression of SMA, CA and LRA were semi-quantitated by RT-PCR. Meanwhile vascular reactivity of SMA, CA and LRA were tested after AG or PD-142893 administration. In vivo experiments, we observed the changes of hemodynamic indexes following endtoxic shock after AG or PD-142893 adminstration.Results:1. Selecting LPS 0.6, 0.8, 1.0 mg/kg body weight as candidate shock doses, we found that mean artery pressure (MAP) of rabbits descended quickly, and it decreased more than 30% at 0.5h after LPS injection, going to shock state, then it kept on descending steadily. It showed that rabbit model of endtoxic shock had been established successfully. There were the changes of hemodynamic indexes as follows: MAP descended obviously at 0.5h after LPS injection (P<0.05), then descended steadily until 6h after LPS injection. Left ventricular systolic pressure (LVSP) and±dp/dt max decreased significantly at 0.5h after LPS injection (P<0.05), while increased slightly at 1h after LPS injection, then decreased steadily until 6h after LPS injection and was significantly lower than normal level (P<0.05). While left ventricular end-diastolic pressure(LVEDP) didn't have significant change after LPS injection (P>0.05).2. SMA reactivity to NE slightly increased in the early period, but decreased obviously in the late period; their reactivity to Ach was increased significantly in the early period, but showed significant hyporeactivity in the late period. CA reactivity to NE increased in the early period, and then decreased obviously; their reactivity to Ach was consistent hyperreactivity, the peak appeared at 2h after LPS injection. LRA reactivity to NE increased quickly in the early period, and then decreased obviously; their reactivity to Ach increased quickly in the early period , while had no significantly change in the late period.3. iNOS gene expression of SMA increased significantly in the late period, which had negative relation with constriction or relaxation reaction in the late period. ET-1 gene expression of SMA also increased significantly in the late period, which had negative relation with constriction or relaxation reaction in the late period too. After AG or PD-142893 administration, the constriction hyporeactivity in the late period was partly improved significantly, while the relaxation hyporeactivity in the late period was not improved. iNOS gene expression of CA increased significantly at 1h after LPS injection, and lasted to 4h after LPS injection, and then decreased at 6h after LPS injection, which had positive relation with constriction hyperreactivity in the early period and relaxation hyperreactivity after LPS injection. After AG administration, constriction and relaxation hyperreactivity in the early period and constriction hyporeactivity in the late period were all partly improved. ET-1 gene expression of CA increased significantly in the late period, which had negative relation with constriction hyporeactivity in the late period. After PD-142893 adminstration, constriction hyporeactivity in the late period didn't have improvement. iNOS gene expression of LRA increased significantly in the late period, which had negative relation with constriction reaction in the late period. After AG administration, the constriction hyporeactivity in the late period was partly improved significantly. ET-1 gene expression of LRA increased significantly in the late period, which had negative relation with constriction reaction in the late period. After PD-142893 administration, the constriction hyporeactivity in the late period was partly improved significantly.4. In the in vivo experiments, we found that AG administration could improve hemodynamic indexes including MAP, LVSP,±dp/dt max quickly. PD-142893 administration could improve hemodynamic indexes including MAP, LVSP,±dp/dt max slowly.Conclusion:1. Rabbit model of endtoxic shock had been established successfully in the present study. There was consistent hypotension following endtoxic shock, MAP decreased steadily, and hemodynamic indexes including LVSP,±dp/dt max had similar changes.2. There are biphase change and organ diversity of vascular reactivity following endotoxic shock. Vascular reactivity increased in the early period and decreased in the late period. Comparing with SMA and LRA, vasoconstriction reaction of CA increased later in the early period and lost less in the late period, vasodilatation reaction of increased more in the early period and remained hyperreactivity in the late period. Organ diversity of vascular reactivity following endtoxic shock may relate with blood redistribution of various organs following shock.3. iNOS and ET-1 gene expression have relations with the changes of vascular reactivity following endtoxic shock. We found vascular reactivity and hemodynamics indexes were improved significantly after inhibiting iNOS and ET-1. It was suggested that iNOS and ET-1 play important role in the changes of vascular reactivity following endtoxic shock.