脱氢醋酸钠致大鼠和家兔出血的初步研究

发布时间：2018-01-05 21:12

本文关键词：脱氢醋酸钠致大鼠和家兔出血的初步研究　出处：《扬州大学》2016年硕士论文　论文类型：学位论文

【摘要】：目的：脱氢醋酸钠(Sodium Dehydroacetate)抗菌谱广,具有较好的防霉效果,作为防腐剂广泛用于食品、饲料及化妆品等相关领域。高剂量应用脱氢醋酸钠可能引起大鼠有出血倾向。本课题组前期研究中也发现脱氢醋酸钠能引起大鼠多器官出血。本文通过给予大鼠和家兔不同剂量脱氢醋酸钠,检测凝血指标PT和APTT,测定血液中VK.1、脱氢醋酸钠血药浓度及VKORC1蛋白的表达,并对主要脏器进行HE染色组织病理学分析,研究脱氢醋酸钠应用于大鼠及家兔后引起的出血倾向。方法：Wistar大鼠,雌雄各半,分别灌胃给予脱氢醋酸钠50mg/kg、100mg/kg、150 mg/kg和200 mg/kg,生理盐水作为对照组,每组14只。每天一次,连续13d。家兔灌胃给予脱氢醋酸钠分为100 mg/kg组、200 mg/kg组和生理盐水对照组,每组14只。在给药后第3d、5d、7d、9d、11d和13d分别采集血液和主要脏器,每组大鼠或家兔4只。测定PT和APTT,对肝脏、脾脏、肺脏、心脏和肾脏进行组织病理学检查,HPLC法检测血液中的脱氢醋酸钠浓度,ELISA法检测大鼠血清VK1含量,免疫组化检测组织中VKORC1蛋白表达。HPLC法检测血清中脱氢醋酸钠的浓度,色谱条件为X BridgeC18色谱柱,甲醇-0.02 mol/L乙酸铵(35/65,v/v)为流动相,293 nm检测。血液样品经乙腈提取、浓缩后上机测定。结果：不同剂量脱氢醋酸钠给予大鼠后,与对照组比较PT和APTT值极显著延长(P0.001),且雄鼠的PT和APTT值显著高于雌鼠(P0.05)。对大鼠HE染色组织切片分析,200 mg/kg组心脏、肝脏、脾脏、肺脏和肾脏都有明显的出血；肝脏组织的出血程度与脱氢醋酸钠剂量有关。100-200 mg/kg剂量脱氢醋酸钠均可引起血清中VK1的含量显著低于生理盐水对照组(P0.01)。 VKORC1蛋白在大鼠肝脏、肺脏、脾脏、心脏和肾脏的表达水平均显著低于对照组(P0.01)。 HPLC法测定大鼠血清中脱氢醋酸钠的浓度在20-50 mg/L。相关性分析表明大鼠血清中VK1水平与PT和APTT的相关系数分别为0.719和0.599；大鼠血清中脱氢醋酸钠浓度与PT和APTT的相关系数分别为0.900-0.988和0.806-0.852。与对照组比较,100-200 mg/kg剂量脱氢醋酸钠给予家兔7d后,PT和APTT值极显著地延长(P0.001)；组织病理学分析200 mg/kg组除心脏外,肝脏,脾脏,肺脏和肾脏都有明显的出血。HPLC法测定脱氢醋酸钠在家兔的血药浓度为5-20 mg/L。相关性分析表明家兔血清中脱氢醋酸钠血药浓度与PT和APTT的相关系数分别为0.837和0.351。结论：较高剂量(150-200 mg/kg)脱氢醋酸钠可引起大鼠和兔凝血指标明显延长,主要脏器有明显的出血；引起大鼠血液中VK1水平明显降低,组织中VKORC1表达降低。脱氢醋酸钠致大鼠或家兔PT和APTT延长,与脱氢醋酸钠血药浓度以及血清中VK1水平高度相关。脱氢醋酸钠导致大鼠和家兔的出血,可能与抑制VKORC1,影响VK循环,导致血清中VK1的水平降低有关。
[Abstract]:Objective: sodium Dehydroacetate has a wide antibacterial spectrum and a good anti-mildew effect. It is widely used as a preservative in food. High dose of sodium dehydroacetate may cause bleeding tendency in rats. We also found that sodium dehydroacetate can cause multiple organ bleeding in rats. And different doses of sodium dehydroacetate in rabbits. Blood coagulation parameters PT and APTT, blood VK.1, blood concentration of sodium dehydroacetate and expression of VKORC1 protein were detected. The histopathological analysis of main organs was performed by HE staining. To study the bleeding tendency induced by sodium dehydroacetate in rats and rabbits. Methods Wistar rats, male and female, were given 50 mg / kg of sodium dehydroacetate and 100 mg / kg of sodium dehydroacetate respectively. 150 mg/kg and 200 mg / kg, normal saline as control group, 14 rats in each group, once a day. For 13 consecutive days, the rabbits were given sodium dehydroacetate intragastrically, which were divided into two groups: 100 mg/kg group (n = 14) and saline control group (n = 14). At the 3rd day, 5 days after administration, the rabbits received sodium dehydroacetate for 7 days. Blood and main organs were collected at 9 d and 13 d, respectively. PT and APTT were measured in each group of rats or rabbits. Histopathological examination of liver, spleen, lung, heart and kidney were carried out. HPLC method was used to detect the concentration of sodium dehydroacetate in blood. Elisa was used to detect the content of serum VK1 in rats. The expression of VKORC1 protein in tissues was detected by immunohistochemistry. The concentration of sodium dehydroacetate in serum was detected by HPLC. The chromatographic condition was X BridgeC18 column. Methanol-0.02 mol/L ammonium acetate 35 / 65 v / v / v) was detected by mobile phase at 293nm. Blood samples were extracted by acetonitrile. Results: after different doses of sodium dehydroacetate were given to rats, PT and APTT values were significantly longer than those of control group (P 0.001). The PT and APTT values of male rats were significantly higher than that of female rats (P 0.05). The heart, liver and spleen of 200 mg/kg group were analyzed by HE staining. There were obvious bleeding in lung and kidney. The degree of hemorrhage in liver tissue was related to the dosage of sodium dehydroacetate. 100-200 mg/kg of sodium dehydroacetate could induce the content of VK1 in serum significantly lower than that in the control group (P < 0.05). VKORC1 protein was found in rat liver. Lungs, spleen. The expression levels in heart and kidney were significantly lower than those in control group (P 0.01). The concentration of sodium dehydroacetate in serum of rats was determined by HPLC method in the range of 20-50. The correlation analysis of mg / L showed that the correlation coefficients between VK1 level and PT and APTT were 0.719 and 0.599 respectively. The correlation coefficients between the concentration of sodium dehydroacetate and PT and APTT in serum of rats were 0.900-0.988 and 0.806-0.852.Compared with the control group. The values of PT and APTT in rabbits treated with sodium dehydroacetate for 100-200 mg/kg for 7 days were significantly prolonged (P 0.001). Histopathological analysis was performed in the 200 mg/kg group except the heart, liver and spleen. The blood concentration of sodium dehydroacetate in rabbits was 5-20 by HPLC. Correlation analysis showed that the correlation coefficients between serum concentration of sodium dehydroacetate and PT and APTT in rabbit serum were 0.837 and 0.351 respectively. Sodium dehydroacetate (150 ~ 200 mg / kg) significantly prolonged the coagulation parameters of rats and rabbits. There was obvious bleeding in the main organs. The levels of VK1 in blood and the expression of VKORC1 in tissues were significantly decreased, and PT and APTT were prolonged in rats or rabbits induced by sodium dehydroacetate. It was highly correlated with the concentration of sodium dehydroacetate in blood and the level of VK1 in serum. Sodium dehydroacetate caused hemorrhage in rats and rabbits, which may be related to the inhibition of VKORC1 and the influence of VK circulation. It is related to the decrease of serum VK1 level.
【学位授予单位】：扬州大学
【学位级别】：硕士
【学位授予年份】：2016
【分类号】：S852.3

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