The Effects of Resveratrol on the Quality of Frozen Sheep Semen

The effects of resveratrol on the quality of frozen-thawed sheep semen were studied in this study.Semen was collected from six Yunnan semi-fine wool sheep using artificial vagina.The semen was diluted with the Optidyl extender supplemented with resveratrol with a concentration of 0,0.1,1,10,or 20 μmol/L,followed by loading into plastic straws and equilibration at a low temperature.Then,the straws were pre-frozen in liquid nitrogen vapor,followed by storage in liquid nitrogen for 30 days.After thawing for 30 seconds in a 37 ℃ water bath,the parameters including sperm motility,acrosome integrity,membrane integrity,distribution of phosphatidylserine (PS) and ROS were measured.The results showed that the post-thaw sperm total motility,progressive motility and the rate of sperm with curved tail were 76.14%±0.97%,43.56%±0.91% and 43.24%±1.68% in the 10 μmol/L resveratrol group,respectively,which were significantly higher than those in the other groups (P<0.05).However,when the concentration of resveratrol in the freezing extender was 20 μmol/L,the post-thaw total motility,progressive motility and the rate of sperm with curved tail were 21.78%±0.79%,25.23%±1.34% and 4.84%±0.68%,respectively,which were significantly lower than those in the other groups (P<0.05).The acrosome integrity of sperm frozen in the 10 μmol/L resveratrol group was best,which was 50.47%±0.91% and significantly higher than the other treatments (P<0.05).The results of PS distribution showed that the percentage of viable sperm in the 10 μmol/L resveratrol group was 46.43%±2.95%,and significantly higher than that in the 20 μmol/L group (31.14%±3.56%,P<0.05),but there was no significant difference with the other groups (P>0.05).In addition,the PS labeling rate of sperm in the 20 μmol/L resveratrol group (39.82%±3.38%) was significantly higher than those of the other groups (P<0.05).In terms of ROS production,this study demonstrated that the rate of viable sperm (ROS and PI were negative) in the 10 μmol/L resveratrol group (63.57%±0.71%) was significantly higher than that of the other groups (P<0.05),while the rate of viable sperm (32.45%±1.42%) in the 20 μmol/L resveratrol group was significantly lower than those of the other groups (P<0.05).In conclusion,the post-thaw quality of sheep semen could be improved after the addition of resveratrol to the freezing extenders,which might be related to the antioxidant properties of resveratrol.But,the cryoprotective effects of resveratrol dependents on its used doses,and the optimal concentration was 10 μmol/L based on this present study.However,excessively high concentration of resveratrol could aggravate cryodamage on sperm.In addition,the protective effects of resveratrol on sheep sperm still needs to be verified by in vitro fertilization and artificial insemination.     

虎杖愈伤组织的诱导及其白藜芦醇形成初探

将虎杖P olygonum cusp id a tum不同外植体经不同消毒时间处理后,接种在添加不同激素种类和水平的相同基本培养基上或相同激素种类和水平基本培养基上进行诱导实验,同时对根和根茎芽、叶、韧皮诱导的愈伤组织进行白藜芦醇含量的测定.结果表明:基本培养基以M S较好,外植体叶对激素种类较为敏感,其中适当浓度的NAA诱导愈伤组织比2,4-D的效果好,KT比BA好,添加KT的培养基上诱导愈伤组织比较紧密,有利于分化,在M S+NAA 2 m g/L+KT 0.1 m g/L培养基上诱导愈伤组织较好,根茎芽的诱导率最高,为73%;愈伤组织的生长趋势从接种的第3天开始生长,到21 d时生长达到最高峰,干质量为0.461 2 g,以后生长速度减慢;对不同材料诱导的愈伤组织进行白藜芦醇含量的测定,其中根茎部芽的诱导的愈伤组织中白藜芦醇含量最高,其次是叶和根,最低的为韧皮.     

白藜芦醇对玉米赤霉烯酮致小鼠肝脏氧化损伤及炎症的保护作用

为探究白藜芦醇（RSV）对玉米赤霉烯酮（ZEA）中毒小鼠肝脏损伤是否具有保护作用,试验将40只清洁级雄性BALB/c小鼠随机分为5组：对照组（生理盐水）、ZEA组（40 mg/kg）、不同浓度的RSV （50、100或200 mg/kg）与ZEA （40 mg/kg）共处理组,每组8只,各组均灌胃给药,试验期12 d。试验结束后采集小鼠肝脏样品,计算肝脏系数,采用苏木素-伊红（HE）染色和免疫组织化学染色法观察各组肝脏病理变化;检测肝脏NF-κB蛋白表达;比色法检测肝脏组织中丙二醛（MDA）含量、超氧化物歧化酶（SOD）和过氧化氢酶（CAT）活性及血清中谷丙转氨酶（ALT）、谷草转氨酶（AST）和乳酸脱氢酶（LDH）活性,ELISA法检测血清中白细胞介素-6（IL-6）、肿瘤坏死因子-α（TNF-α）和IL-1β含量。结果显示,与对照组相比,ZEA组小鼠肝组织发生明显病理学变化;与ZEA组相比,不同浓度RSV与ZEA共处理组小鼠肝脏组织病理学变化都有所减轻。免疫组织化学结果显示,与对照组相比,ZEA组肝脏NF-κB蛋白表达增多;与ZEA组相比,不同浓度的RSV与ZEA共处理组肝脏NF-κB蛋白表达均有下降。与对照组相比,ZEA组肝脏系数和肝脏组织MDA含量极显著升高（P<0.01）,肝脏组织SOD和CAT活性显著或极显著降低（P<0.05;P<0.01）,血清中AST、ALT和LDH活性,IL-6、TNF-α和IL-1β含量均极显著升高（P<0.01）。与ZEA组相比,不同浓度RSV与ZEA共处理组肝脏组织CAT活性显著或极显著升高（P<0.05;P<0.01）,血清中AST、ALT和LDH活性及IL-6和IL-1β含量显著或极显著降低（P<0.05;P<0.01）;50 mg/kg RSV与ZEA共处理组肝脏组织SOD活性和血清中TNF-α含量分别显著升高（P<0.05）和极显著降低（P<0.01）;100和200 mg/kg RSV与ZEA共处理组肝脏系数和肝脏组织MDA含量显著或极显著降低（P<0.05;P<0.01）。结果表明,ZEA对小鼠肝脏有严重的氧化及炎症损伤作用,RSV对ZEA中毒的肝损伤具有一定保护作用,尤其以100 mg/kg RSV保护效果最佳。     

成熟液中添加白藜芦醇对猪卵母细胞体外发育和抗冻能力的影响

为探究猪卵母细胞成熟培养液中添加白藜芦醇(Resveratrol,RES)对其体外发育与抗冻能力的影响。本试验通过添加不同浓度的RES,观察其对卵母细胞体外发育和相关基因的表达的影响;通过OPS(Open Pulled Straw)玻璃化冷冻和解冻,观察RES对卵母细胞冷冻前后线粒体功能、凋亡和发育能力的影响。结果表明：(1)成熟培养液中添加2 μmol/L RES有利于提高猪卵母细胞的体外成熟和发育能力,提高RES浓度至10 μmol/L时则表现为细胞毒性作用;(2)添加2 μmol/L RES可显著提高卵母细胞Sirt1,MATER,SOD1和Bcl2基因的表达量,并显著降低c-mos,MAPK和Mfn2的表达;当RES添加量增至10 μmol/L时,卵母细胞凋亡相关基因的表达呈现出促凋亡状态;(3)添加RES后,成熟卵母细胞在冷冻前后的ROS水平均要低于未添加组,ATP含量和线粒体膜电位值则高于未添加组;(4)成熟液中添加RES后,卵母细胞冻后FDA染色存活率和孤雌激活卵裂率均高于未添加组,,早期凋亡率则低于未添加组,其中FDA染色存活率差异显著。综上所述,2 μmol/L的RES在猪卵母细胞成熟液中添加,可通过改善其发育和凋亡相关基因的表达,提高线粒体功能,达到提高卵母细胞发育和抗冻能力的目的。     

白藜芦醇的生物学功能及其应用前景

本文综述白藜芦醇在抗肿瘤、抗氧化、抗炎、抗组织损伤、抗流感、心血管保护、免疫调节等方面的作用及其在动物生产中的应用。     

Effects of dietary resveratrol supplementation during gestation and lactation of sows on milk composition of sows and fat metabolism of sucking piglets

The purpose of this article was to investigate the effects of dietary resveratrol supplementation during gestation and lactation of sows on the milk composition of sows and the fat metabolism of sucking piglets. Forty sows were allotted to two experimental treatment groups that included the following: (a) control sows (CON treatment, n = 20) fed with a corn–soybean meal control diet and (b) treatment sows (RES treatment, n = 20) fed with a control diet with addition of 300 mg/kg resveratrol. The results showed that the content of lactose in the colostrum was increased (p < 0.05) and the content of fat in 21‐day milk was increased (p < 0.05) by dietary resveratrol supplementation. In the RES treatment group, the concentrations of high‐density lipoprotein‐cholesterol (HDL‐C), low‐density lipoprotein‐cholesterol (LDL‐C), lipase activity and insulin (INS) in plasma of sucking piglets were increased (p < 0.05). In the adipose tissue, the enzyme activities of hormone‐sensitive lipase (HSL), acetyl‐CoA carboxylase (ACC) and lipoprotein lipase (LPL) increased significantly by RES treatment (p < 0.05), and the mRNA levels of acetyl coenzyme A‐alpha (ACCα), LPL, fatty acid transport protein (FATP1) and CCAAT–enhancer‐binding protein gene (C/EBPα) were higher in the RES treatment group (p < 0.05). In conclusion, resveratrol supplementation on gestational and lactating sows improved the content of lactose in the colostrum and the content of fat in milk at day 21 of lactation. In addition, resveratrol supplementation on sows increased HDL and LDL in the plasma of piglets. In piglet adipose tissue, the enzyme activity and mRNA level related to lipolysis, fatty acid uptake from circulating triacylglycerols and lipogenesis are partially improved by resveratrol supplementation on sows. These aspects affect fat metabolism in piglets.     

Effects of resveratrol on in vitro maturation of porcine oocytes and subsequent early embryonic development following somatic cell nuclear transfer

As a natural plant‐derived antitoxin, resveratrol possesses several pharmacological activities. This study aimed to evaluate the effects of resveratrol addition on nuclear maturation, oocyte quality during in vitro maturation (IVM) of porcine oocytes and subsequent early embryonic development following somatic cell nuclear transfer (SCNT). Our experiments showed that the treatment of porcine oocytes with 5 µM resveratrol during IVM resulted in the highest rate of the first polar body extrusion. Treatment of oocytes with resveratrol had no influence on cytoskeletal dynamics, whereas it significantly increased glucose uptake ability compared to the control oocytes. Oocytes matured with 5 μM resveratrol displayed significantly lower intracellular reactive oxygen species (ROS) levels and higher relative mRNA expression levels of the genes encoding such antioxidant enzymes as catalase (CAT) and superoxide dismutase 1 (SOD1). In addition, resveratrol also prevented onset and progression of programmed cell death in porcine oocytes, which was confirmed by significant upregulation of the anti‐apoptotic B‐cell lymphoma 2 (BCL‐2) gene and significant downregulation of the pro‐apoptotic BCL2‐associated X (BAX) gene. Furthermore, the blastocyst rates and the blastocyst cell numbers in cloned embryos derived from the oocytes that had matured in the presence of 5 μM resveratrol were significantly increased. In conclusion, supplementation of IVM medium with 5 μM resveratrol improves the quality of porcine oocytes by protecting them from oxidative damage and apoptosis, which leads to the production of meiotically matured oocytes exhibiting enhanced developmental potential following SCNT.     

白藜芦醇对猪卵母细胞体外老化的抑制作用

旨在研究白藜芦醇(RES)对猪卵母细胞体外老化的影响及机制。添加不同浓度RES(0、1、2、4μmol/L),体外培养猪卵母细胞44 h后,通过卵丘扩散和第一极体排出率筛选出最佳有效浓度为2μmol/L。卵丘-卵母细胞复合体(cumulus-oocyte complexes, COCs)成熟培养44 h作为对照组,连续培养68 h作为体外老化组,置于含2μmol/L白藜芦醇成熟培养液中培养68 h,为白藜芦醇处理组。激光扫描共焦显微镜检测染色体和纺锤体,Western blot检测凋亡相关蛋白Caspase-3、Bcl-2和Bax表达水平。结果表明:RES处理组和对照组的猪卵母细胞染色体与纺锤体异常率均较老化组显著降低(P<0.01)。用2μmol/L RES处理后,猪卵母细胞Caspase-3和Bax蛋白表达量均有下降,显著低于老化组(P<0.001);Bcl-2蛋白表达量有所升高,但差异不显著。试验表明,RES可明显抑制猪卵母细胞的老化,这一作用可能主要通过抑制线粒体凋亡途径来实现的。     

白藜芦醇的生物学功能及其在动物生产中的应用

白藜芦醇是一种含有茋类结构的天然非黄酮类多酚类化合物,具有多种生物学功能：可抑制和清除体内过量的自由基,激活机体抗氧化酶系统,发挥抗氧化功能;通过调控一系列炎症或抗炎通路,降低机体内炎性因子表达,降低炎症反应。白藜芦醇还具有调节糖脂代谢、保护心血管、抗癌、抗病毒等重要生物学功能,且具有安全高效、无残留、不产生耐药性的优点,在动物生产中的应用具有广阔的应用前景。动物试验表明,白藜芦醇能提高肉鸡平均日增重、平均采食量,降低料重比;提高育肥猪肉品质,增加背最长肌pH_{24 h}、红度、粗蛋白质和肌红蛋白含量,还可降低24 h后肌肉亮度、剪切力、滴水损失、背膘厚度;提高动物机体内抗氧化酶活性;降低炎性因子的产生,增强机体免疫;改善肠道菌群组成和肠道黏膜形态;抑制癌细胞的生成。本试验针对白藜芦醇生物学特性、生理功能及其在畜禽生产中的应用现状进行综述,旨在为其在畜禽生产中的应用提供参考依据。     

Effect of dietary supplementation with resveratrol on nutrient digestibility,methanogenesis and ruminal microbial flora in sheep

Two experiments were conducted to evaluate the effect of resveratrol on methanogenesis and microbial flora in Dorper × thin‐tailed Han cross‐bred ewes. In experiment 1, ten ewes (67.2 ± 2.24 kg BW) were assigned to two dietary treatments, a basal diet and a basal diet supplemented with resveratrol (0.25 g/head·day), to investigate the effect of resveratrol on nutrient digestibility and nitrogen balance. In experiment 2, six ewes (64.0 ± 1.85 kg BW) with ruminal cannulae were assigned to the identical dietary treatments used in experiment 1 to investigate supplementary resveratrol on ruminal fermentation and microbial flora using qPCR. The results showed that supplementary resveratrol improved the digestibility of organic matter (OM; p < 0.001), nitrogen (N; p = 0.007), neutral detergent fibre (NDF; p < 0.001) and acid detergent fibre (ADF; p < 0.001). The excretion of faecal N was reduced (p = 0.007), whereas that of urinary N increased (p = 0.002), which led to an unchanged N retention (p = 0.157). Both CO₂ and CH₄ output scaled to digestible dry matter (DM) intake decreased from 602.5 to 518.7 (p = 0.039) and 68.2 to 56.6 (p < 0.001) respectively. Ruminal pH (p = 0.341), ammonia (p = 0.512) and total volatile fatty acid (VFA) (p = 0.249) were unaffected by resveratrol. The molar proportion of propionate increased from 13.1 to 17.5% (p < 0.001) while that of butyrate decreased from 11.0 to 9.55% (p < 0.001). The ratio of acetate to propionate (A/P) decreased from 5.44 to 3.96 (p < 0.001). Supplementary resveratrol increased ruminal population of Fibrobacter succinogenes, Ruminococcus albus and Butyrivibrio fibrisolvens (p < 0.001) while decreased protozoa and methanogens. In conclusion, dietary resveratrol inhibited methanogenesis without adversely affecting ruminal fermentation.