稻瘟菌糖原合成酶激酶MoGSK3功能探究

【目的】观测稻瘟菌（Magnaporthe oryzae）糖原合成酶激酶基因MoGSK3敲除突变体表型,明确MoGSK3在稻瘟菌中的潜在生物学功能,为挖掘防治稻瘟菌新型药剂的潜在靶标提供参考。【方法】基于同源重组原理,用split-PCR方法获得稻瘟菌MoGSK3敲除突变体菌株,将MoGSK3基因克隆到pFL2载体上得到MoGSK3-C融合载体,并将其通过PEG介导的原生质体转化法导入MoGSK3突变体中得到回补菌株。培养观察野生型菌株Guy11、突变体菌株G3-9及回补菌株GC-1的菌落形态和生长状况,采用实时荧光定量PCR(qRT-PCR)检测稻瘟菌产孢相关基因的表达量;观察稻瘟菌不同菌株的分生孢子形态,通过附着胞洋葱表皮穿透试验及接种水稻叶片,研究其致病力;通过KI-I-2染色对野生型菌株Guy11及突变体菌株G3-9分生孢子和附着胞中的糖原运输能力进行观测。【结果】稻瘟菌MoGSK3突变体存在多个表型缺陷,与野生型菌株Guy11相比,MoGSK3突变体菌株G3-9菌落直径显著变小,生长缓慢,产孢相关基因表达量下降且分生孢子出现末端伸长畸形的状态。MoGSK3基因缺失还会导致稻瘟菌菌丝末端无法形成正常的分生孢子梗,附着胞无法穿透洋葱表皮形成侵染菌丝,接种划伤水稻叶片也无法形成褐色病斑。对MoGSK3突变体菌株G3-9分生孢子及附着胞进行糖原染色后发现,突变体菌株G3-9在糖原转运能力方面存在明显缺陷。【结论】MoGSK3基因参与稻瘟菌的生长、分生孢子形成及形态建成、侵染、糖原转运等过程,是稻瘟菌重要的毒力因子。     

铜对肉鸡原代肝细胞活性及糖原含量的影响

为了探讨铜对肉鸡原代肝细胞活性及糖原含量的影响,试验分别用10,50,100μmol/L Cu2+在体外孵育原代肝细胞,PAS染色后测定30分钟和1,2,4,24,48,72小时各时间点的原代肝细胞活性及24小时时的糖原含量。结果表明:50μmol/L和100μmol/L Cu2+仅分别在72小时和4小时之后明显降低细胞活性;随着铜浓度的升高,糖原含量减少。说明高浓度铜可降低原代肝细胞活性,促进肝糖原分解。     

Lysine limitation alters the storage pattern of protein,lipid and glycogen in on‐growing Atlantic salmon

This experiment aimed to test the interaction of lysine limitation with nutrient accretion and muscle carnitine depot in Atlantic salmon. Fish were fed adequate or low‐lysine diets for 3 months. Lysine intake was significantly less (48%) in fish fed the low‐lysine diet as compared with that fed the adequate one. There was no difference in dietary amino acids between treatments, with the exception of lysine. The lower lysine intake was reflected in plasma free lysine being 52% less while the free lysine concentration in the liver and muscle were unaffected. Although there was no significant difference between voluntary feed intakes among treatments, fish fed the low‐lysine diet had reduced growth, protein and energy deposition as compared with fish fed the adequate lysine diet. White trunk muscle contained more glycogen and less protein in fish fed the low‐lysine diet while no difference in lipid was observed. The livers from fish fed the low‐lysine diet contained less glycogen and slightly more fat and protein than the livers from fish fed the adequate lysine diet. Lysine limitation reduced carnitine in the liver without affecting muscle carnitine depot. Thus, low‐lysine diets did not likely affect the fatty acid oxidation capacity. This fact was supported by unaffected fatty acid profiles and lipid classes between treatments during the 3‐month study. In conclusion, lysine limitation does not deplete the muscle carnitine depot during the on‐growing seawater phase of Atlantic salmon, but affects the deposition pattern of nutrients.     

日本血吸虫GSK3蛋白编码cDNA的克隆、表达及其重组蛋白的免疫保护研究

本研究利用生物信息学分析了日本血吸虫(Schistosoma japonicum,sj)Glycogen synthase kinase 3(GSK3)蛋白,并对其中一个GSK3蛋白的编码cDNA进行了克隆和原核表达,制备了特异性的多克隆抗体.同时,还初步评估了重组蛋白的免疫保护效果.生物信息学分析表明在日本血吸虫数据库存在两种GSK3蛋白,且其中一个SjGSK3在日本血吸虫不同发育时期均有转录.Western blot结果表明本研究制备的抗体能特异性识别日本血吸虫SjGSK3重组蛋白,表明该重组蛋白具有良好的免疫原性.动物实验表明免疫SjGSK3重组蛋白的动物与佐剂对照组比较分别获得了平均10.6％减虫率和40.5％肝脏减卵率.     

Sarcoplasmic Reticulum Ca2+-ATPase Activity and Glycogen Content in Various Fiber Types after Intensive Exercise in Thoroughbred Horses

To find a new parameter indicating muscle fitness in Thoroughbred horses, we examined time-dependent recovery of glycogen content and sarcoplasmic reticulum (SR) Ca²⁺-ATPase activity of skeletal muscle after intensive treadmill running. Two repeated 50-sec running sessions (13 m/sec) were performed on a flat treadmill (approximately 90%VO₂max). Muscle samples of the middle gluteal muscle were taken before exercise (pre) and 1 min, 20 min, 60 min, and 24 hr after exercise. Muscle fiber type composition was determined in the pre muscle samples by immunohistochemical staining with monoclonal antibody to myosin heavy chain. SR Ca²⁺-ATPase activity of the muscle and glycogen content of each muscle fiber type were determined with biochemical analysis and quantitative histochemical staining, respectively. As compared to the pre value, the glycogen content of each muscle fiber type was reduced by 15–27% at 1 min, 20 min, and 60 min after the exercise and recovered to the pre value at 24 hr after exercise test. These results indicate that 24 hr is enough time to recover glycogen content after short-term intensive exercise. The mean value of the SR Ca²⁺-ATPase activity showed a slight decrease (not significant) immediately after exercise, and complete recovery at 60 min after exercise. There were no significant relationship between the changes in glycogen content of each muscle fiber type and SR Ca²⁺-ATPase. Although further studies are needed, SR Ca²⁺-ATPase is not a useful parameter to detect muscle fitness, at least in Thoroughbred horses.     

三种淡水鱼肌肉的糖元、乳酸含量和pH值及在冷藏中的变化

本文测定了鳙，团头鲂和乌鳢三种养殖淡水鱼即杀死后在5℃贮藏过程中糖元，乳酸含量和pH值的变化。结果表明：鳙，团头鲂和乌鳢的糖元含量比海水鱼的低，并在24h内全部分解完毕。乳酸含量在死后24－60h达到且维持在高位。三种淡水鱼即杀的pH值与海水鱼的接近，但死后量低的pH值比海水鱼的高，pH值的低值阶段出现在死后36－60h间。     

Metabolic function of hepatic hexokinase in perch, Perca fluviatilis

Feeding previously fasted perch with a carbohydrate‐rich pelleted diet resulted in a rapid transient insulin response followed by a transient increase in the activity of the hepatic low‐affinity hexokinase (glucokinase, GK). The activity of the high‐affinity hexokinase (hexokinase, HK) increased more slowly and steadily to about seven times the fasting value on the 10th day after start of feeding, in parallel with an increase in the hepatic glycogen content. An insulin response was also observed when fasted perch were fed a diet consisting of boiled fillet of cod (a high‐protein, very low‐fat and practically carbohydrate‐free diet). In this case no GK activity was observed, whereas the glycogen content increased steadily from the start of feeding to about the same level as when fish were given the carbohydrate‐rich diet. There was a delayed increase in the activity of HK, starting on about the sixth day (after the glycogen stores were filled), concomitant with increased activities of lipogenic enzymes (glucose‐6‐phosphate dehydrogenase, ATP citrate lyase and malic enzyme). The results are in line with the hypothesis that the mitochondria‐bound hepatic HK has a particular anabolic function in glycogen and fat synthesis.     

猪RN基因的研究

RN基因是影响猪肉质性状的一个主效基因,已被定位在猪的15号染色体上(15q25),包含显性的不利等位基因(RN)为影响猪肉的工艺产量和隐性正常的等位基因(rn)。最近发现的与肌肉过量糖原含量相关的PRKAG3基因的非保守替换(R200Q),被认为是引起酸肉的主要原因。     

Hypophosphatemia Induced by Dietary Aluminium Hydroxide Supplementation in Growing Pigs: Effects on Erythrocytes,Myocardium, Skeletal Muscle and Liver

Håglin, L., B. Essén-Gustavsson and A. Lindholm: Hypophosphatemia induced by dietary aluminium hydroxide supplementation in growing pigs: Effects on erythrocytes, myocardium, skeletal muscle and liver. Acta vet. scand. 1994, 35, 263-271.– Three groups of pigs were studied during and after 10 weeks of treatment with either Al(OH)₃ (Al[OH]₃-group, n=8) to induce hypophosphatemia, A1P0₄ (AlP0₄-group, n=8, aluminium control without hypophosphatemia) or no addition to the feed (control group, n=8). Blood samples were taken at the start of the experiment and after 3, 6 and 10 weeks and were analyzed for phosphate, calcium and 2,3-diphosphoglycerate (2,3-DPG). Samples from myocardium, skeletal muscle and liver were obtained in connection with exsanguination and analyzed for glycogen, adenosine-tri-phosphate (ATP), creatine phosphate (CP), glucose-6-phosphate (G-6-P) and lactate. The Al(OH)₃-group became hypophosphatemic and hypercalcémie with low levels of 2,3-DPG in erythrocytes within 3 weeks and showed a retarded growth rate. After 10 weeks the Al(OH)₃-group had low levels of ATP in myocardium as compared with the control-group and low levels of G-6-P as compared with the AlP0₄-group. No disturbances on electro-cardiograms registered at rest could be documented. G-6-P concentration was low in the biceps muscle in the Al(OH)₃-group as compared with the AlP0₄-group and in the liver low G-6-P concentration was seen in addition to high lactate concentration. The fibre type composition in M. Longissimus did not differ between groups, but the Al(OH)₃-group had, due to retardation in growth, smaller mean fibre-areas than pigs in the AlP0₄-group. Hypophosphatemia gave rise to high serum calcium levels, low concentration of 2,3-DPG in erythrocytes and influenced G-6-P concentration in skeletal muscle, G-6-P and ATP in myocardium, G-6-P and lactate in liver. Retarded growth was one serious consequence of hypophosphatemia and the disturbed energy metabolism.     

谷氨酰胺的抗疲劳生化机制研究

用两批各 48只小鼠分 4组 (n =12 )每天分别灌喂生理盐水和 3 4mmol·kg-1的谷氨酰胺、谷氨酸和天冬酰胺 ,连续 10d。结果显示 ,三者都可提高小鼠血清、肝和骨骼肌中的肌酸磷酸激酶活性 ,降低乳酸脱氢酶活性 ,并显著增加肝、骨骼肌糖原积累。其中有相同碳架结构的谷氨酰胺、谷氨酸能极显著提高小鼠的游泳耐力 ,但与谷氨酰胺有相等酰胺氮的天冬酰胺作用不强。结果提示 ,谷氨酰胺提高机体抗疲劳作用与其碳架结构有关