【作者】 唐建霞；

【导师】 黄克和；

【作者基本信息】 南京农业大学 ， 临床兽医学， 2003， 硕士

【摘要】 为探讨高钙致痛风的机理和为痛风的防制提供理论基础，本文对高钙日粮引起鸡痛风进行了体内和体外实验研究。本研究分为三部分： 试验一探讨了高钙导致鸡尿液及尿沉渣的变化情况。将100羽35日龄健康伊沙蛋雏鸡随机均分为对照组和高钙组。对照组饲喂全价雏蛋鸡料(含钙1.0％)，高钙组在此基础上添加石粉使饲料中实际含钙量达3.85％。两组鸡均进行结肠造口术，收集尿液，研究鸡尿液及尿沉渣的变化。结果表明，高钙日粮导致严重的肾脏病变，尿量增加，与对照组比较差异显著(p＜0.01)；尿液pH值升高，与对照组比较差异不显著；高钙组鸡尿液中钾、钙(p＜0.01)、钠、镁(P＜0.05)的相对含量均显著低于对照组；尿液尿酸低于对照组，但统计学差异不显著；无机磷的相对含量与对照组相近。高钙组鸡尿液中钙(p＜0.01)、尿酸、镁、及无机磷(p＜0.05)的总量显著高于对照组，钠(p＜0.01)和钾(p＜0.05)的总量低于对照组，统计学差异显著。扫描电镜下，高钙组尿沉渣晶体中针状较多。能谱分析表明，尿沉渣中钙、磷含量升高，而钾、钠下降。 试验二用高钙与高蛋白日粮对鸡痛风进行了人工发病实验研究。120只35日龄健康蛋仔鸡随机均分为3组，对照组饲喂基础日粮，高钙组(high calcium，HC)在饲喂基础日粮的同时添加石粉(含钙3.80％)，高钙高蛋白组(high calcium and high protein，HCHP)在饲喂基础日粮的同时添加石粉和豆粕(含钙3.55％，粗蛋白24.58％)，进行30天饲养试验。结果显示：HC组鸡出现肾脏损害，未见尿酸盐在内脏器官的沉积；与对照组相比，血清无机磷显著降低(p＜0.01)，血清尿酸、钠(P＜0.05)显著升高；尿液的检查显示尿液中尿酸、钙(p＜0.01)、无机磷、镁和钾离子(P＜0.05)总量显著升高，钠离子总量显著降低(P＜0.05)。HCHP组鸡发生典型内脏型痛风，尿酸盐在内脏器官表面沉积；血清尿酸(p＜0.01)、钙(P＜0.05)显著升高；尿液中尿酸、钙、镁(p＜0.01)、无机磷和钾(P＜0.05)总量与对照组相比显著升高，钠离子总量与对照组相比显著降低(p＜0.05)。尿沉渣扫描电镜观察，高钙和高蛋白日粮均可造成尿沉渣晶体形态结构的改变，其中HC组鸡尿沉渣中以针状晶体较多，球形晶体较少；HCHP组鸡尿沉渣中针状与球状晶体含量相当。能谱分析结果表明，HC、HCHP组鸡尿沉渣钾离子、钠离子均下降；无机磷均升高；HCHP组磷含量最高；HC组钙含量最高。 实验三，建立了鸡原代肾小管上皮细胞的培养方法，作为高钙对肾小管损害的体外研究模型。在此模型上应用不同浓度的钙进行处理，通过MTT比色法和乳酸脱氢酶(LDH)活性测定法观察钙对细胞活性和细胞膜的损伤情况。结果表明，分别用2.5mmol／L(试验组Ⅰ)，4.0mmol／L(试验组Ⅱ)，5.5mmol／L(试验组Ⅲ)，7.0mmol／L 高钙日粮引起鸡痛风的体内和体外研究（试验组 IV），8．5 auno ／L（试验组 V）和 10．0 uuno几（试验组 VI）Ca‘”处理细胞，各组的细胞活性与对照组相比均显著降低，细胞外钙浓度越高，细胞活性降低越明显。不同浓度的Ca‘“处理细胞，对培养液中LDH的活性影响不明显。更多还原

【Abstract】 In order to study the mechanism of avian gout caused by high dietary calcium and provide a theory basis for preventing of gout, studies on avian gout induced by high dietary calcium in vivo and in vitro were conducted. The present study included three series as follows:In experimental 1, 1 00 pullets of 35 days old were selected for the experiment. The pullets were divided randomly into the control group and the experimental group. The pullets in the control group were fed with a normal diet and the pullets in the experimental group were fed with a high calcium diet (3.85%). The pullets in both the experimental group and the control group were colostomized, and the urine was collected to quantify high dietary calcium effects on urine and urinary deposit composition. The results showed that excessive dietary calcium during the pullets growing phase could cause several damage of kidney. The pullets raised on the high calcium diet had higher urine pH, but not significantly when compared with the control, and significantly larger volume of urine when compared with the control. The high dietary calcium significantly decreased the relative contents of potassium, calcium (p<0.01), sodium and magnesium (p<0.05) in urine compared with the control. The high dietary calcium increased the total content of calcium (p<0.01), uric acid, magnesium, inorganic phosphorous (p<0.05) in urine, but decreased the total contents of sodium (p<0.01), potassium (p<0.05) in urine. Scanning electron micrograph of urinary deposit showed that high dietary calcium could change the crystal structure. The results of X-ray energy dispersive spectrometer revealed that calcium, phosphorus increased and sodium, potassium decreased in the urinary deposit in the experimental group compared with the control group.In experimental 2, one hundred and twenty 35 days of age, healthy pullets were divided randomly into three groups. A basic diet was fed to the control pullets. The pullets in the other two experimental groups were fed with a high calcium (3.80%), normal protein (17.83%) diet (HC), and a high calcium (3.55%), high protein (24.58%) diet (HCHP) respectively. All the pullets were fed for 30 days. The results were as follows: the HC diet could cause severel damage of kidney, typical visceral gout was not found in pullets raisedon it. The HC diet significantly increased plasma uric acid and sodium (p<0.05), but decreased plasma inorganic P (P<0.01) of pullets when compared with the control group. Pullets raised on HC diet had higher fractional of calcium (p<0.01), magnesium, Inorganic P and potassium (p<0.05) significantly, and had significantly lower fractional sodium (p<0.05) in urine when compared with pullets raised on normal diet. Most of the pullets in HCHP group developed visceral gout. The HCHP increased plasma uric acid (p<0.01) and calcium (p<0.05) significantly. The pullets raised on HCHP diet had significantly higher fractional of calcium, magnesium (p<0.01) and inorganic P, potassium (p<0.05), significantly lower fractional sodium (P<0.05) in urine when compared with pullets raised on normal diet. Scanning electron micrograph of urinary deposit showed that HC and HCHP could change the construction of crystals in urinary deposit. The results of X-ray energy dispersive spectrometer revealed that calcium, phosphorus increased and sodium, potassium decreased in the urinary deposit of both the group of HC and HCHP compared with the control group, the urinary deposit of pullets in the group of HCHP had the highest level of inorganic P of all, and the urinary deposit of pullets in the HC had the highest level of calcium in the three groups.In experimental 3, the method of new-born chicken’s renal tubule cells primary culture in vitro was established and used as the model of studying the mechanism of avian renal tubules cells’ injury caused by high extracellular calcium. The viability of the cells was examined by MTT colorimetric analysis, and the injury of cellular membrane was detected through the levels of LDH released in cult更多还原