鹿角盘水提物对大鼠乳腺增生的作用效果及其机制研究

为了探索鹿角盘(DHAB)水提物对大鼠乳腺增生的治疗作用,初步探讨其作用机理和用药剂量,试验利用苯甲酸雌二醇(0.5mg/(kg·d))联合黄体酮(5.0mg/(kg·d))对SD大鼠进行肌内注射建立乳腺增生模型。试验共分7个组,每组10只大鼠,分别为空白组、乳腺增生症(HMG)模型组、阳性药三苯氧胺组(0.036mg/(kg·d))、鹿角盘水提物4个剂量组(高剂量,2.625g/(kg·d);中高剂量,1.575g/(kg·d);中低剂量,1.05g/(kg·d);低剂量,0.63g/(kg·d))。按组别连续灌胃45d,每周固定时间测定大鼠乳头直径、高度及体重。试验结束后采血取样,称量大鼠胸腺、脾脏、子宫和卵巢质量,取乳腺组织做病理学切片、HE染色,用放射免疫法检测大鼠血清中雌二醇(E2)及孕酮(P)的含量。结果表明,使用不同剂量鹿角盘水提物均能减小乳腺增生大鼠乳头直径、高度,提高胸腺、脾脏指数,降低子宫指数,减轻乳腺小叶数、腺泡数和分泌物,降低血清E2含量,升高血清P的含量,与模型组相比有统计学差异(P0.01;P0.05))。综合试验结果,鹿角盘水提物对乳腺增生大鼠有治疗作用,效果显著,且以中高剂量组变化最为明显。其作用机制可能通过增强机体免疫力、调节血清中E2、P水平实现。     

鹿角盘不同提取部位对环磷酰胺致小鼠骨质疏松的防治作用

建立了环磷酰胺所致小鼠骨质疏松模型,利用空白对照组和葡萄糖酸钙阳性组做对比,以小鼠免疫器官脏器指数、股骨计量学、骨钙、磷、镁、骨羟脯氨酸含量为指标观察鹿角盘提取物对小鼠免疫功能、骨量的影响.结果:20mg/(kg·d)环磷酰胺可引起小鼠胸腺指数下降(P＜0.01),股骨干重、股骨横径减少(P＜0.01),股骨长减少(P＜0.05),骨钙、羟脯氨酸含量同时下降(P＜0.01),引起小鼠骨丢失;鹿角盘水提物和70％乙醇提取物均可增加小鼠股骨的干重和骨钙的含量,水提物高剂量组和70％乙醇提取物高低、高剂量组使骨羟脯氨酸含量显著提高.结果表明鹿角盘对以低转换型骨丢失为特征的骨质疏松具有一定的预防作用.     

螺旋藻多糖与银杏叶提取物复方对四氧嘧啶模型小鼠的降血糖作用

为研究复合螺旋藻多糖与银杏有效成分的降血糖作用,试验选用SPF级昆明种雄性小鼠,腹腔注射0.2 mL 200 mg/（kg·d）四氧嘧啶,连续5 d,断尾采血测定空腹血糖,其值大于11.1 mmol/L时表明造模成功。试验小鼠随机分为空白对照组（C组）、模型组（M组）、阳性对照组（CY组）、单一用药组（螺旋藻多糖组（P组）、银杏黄酮组（F组）、银杏内酯组（L组）,复合用药组将螺旋藻多糖（PSP）与银杏叶有效成分（GBE）配制成1：1（CP₁组）、2：1（CP₂组）、1：2（CP₃组）的复合组,每组10只,共9组。对照和CY组每只小鼠灌服2.5 mg/（kg·d）格列本脲,其余各组按200 mg/（kg·d）灌服相应药物,每天1次,连续灌服30 d,测定小鼠血糖值及体重、脾脏指数、胸腺指数及肝糖原等指标。结果显示,与M组相比,用药组小鼠体重均极显著增加（P<0.01）,复合用药组药效高于单一用药组;各用药组脾脏、胸腺指数均极显著升高（P<0.01）;肝糖原含量极显著增多（P<0.01）;血糖值极显著下降（P<0.01）;其中CP₂组的小鼠血糖值降幅最大,其降糖率为52.14%;肝糖原增加为61.88%。综上所述,螺旋藻多糖和银杏叶有效成分发生了协同增效作用,对由四氧嘧啶引起的高血糖小鼠有明显的降血糖作用。     

芦丁促进大鼠泌乳性能的研究

通过研究天然植物提取物芦丁对大鼠泌乳性能、内分泌激素、脏器指数的影响,考察芦丁调节大鼠泌乳的效果.试验选择18只Wistar受孕母鼠,随机分为3组,每组6只,分别为对照组、芦丁组(每日灌服芦丁60 mg/kg BW)、雌二醇组(每周肌肉注射雌二醇60 μg/kg BW),从哺乳第4天开始连续给药2周,基础饲粮相同.测定指标包括:大鼠泌乳量,仔鼠平均体增重,乳腺器官指数,胸腺指数,脾脏指数,血浆与乳腺组织中雌激素(E2)、孕激素(P)、催乳素(PRL)、生长激素(GH)的含量.试验结果显示:芦丁组大鼠泌乳量显著高于对照组(P＜0.05),与雌二醇组差异不显著(P＞0.05);仔鼠体增重芦丁组与雌二醇组差异不显著(P＞0.05),但显著大于对照组(P＜0.05);大鼠血浆与乳腺组织中E2、PRL、GH的水平,芦丁组显著高于对照组(P＜0.05),低于雌二醇组(P＜0.05),P水平各组间差异不显著(P＞0.05);大鼠乳腺、胸腺、脾脏器官指数雌二醇组显著高于芦丁组和对照组(P＜0.05).由此得出,芦丁能够显著提高大鼠泌乳量,促进大鼠胸腺、脾脏等器官的发育.     

植物甾醇对去卵巢KM小鼠血清生殖激素及乳腺组织孕激素受体的影响

本试验旨在研究植物甾醇对去卵巢小鼠血清激素及乳腺组织中孕激素受体的影响。50只雌性小鼠,随机选40只进行去卵巢手术,恢复7 d。分组如下：正常组、去卵巢对照组、植物甾醇低、中、高3个剂量组（每日灌胃20、80和320 mg/kg植物甾醇）。连续灌胃3周,第22天采集血样和乳腺组织,对小鼠血清中的雌二醇、孕酮和催乳素水平进行检测,对乳腺组织的孕激素受体基因表达进行荧光定量检测。结果表明,去卵巢对照组小鼠雌二醇水平显著低于正常组（P〈0.05）。植物甾醇组小鼠血清雌二醇水平持续升高,其中,高剂量组小鼠血清雌二醇水平与正常组最接近;与去卵巢对照组相比,去卵巢植物甾醇组小鼠血清孕酮水平均有提高,但差异不显著（P〉0.05）;与去卵巢对照组相比,去卵巢植物甾醇组小鼠催乳素水平先下降后上升,但他们之间无显著差异（P〉0.05）。与去卵巢对照组相比,植物甾醇各剂量组小鼠孕激素受体基因相对表达量呈上升趋势,低、中、高剂量组小鼠孕激素受体基因相对表达量分别是去卵巢对照组的1.05、1.13和2.78倍。植物甾醇可改善去卵巢小鼠内源激素不平衡的状况,促进乳腺组织PR基因的表达。     

中草药复合微生态制剂对急性肝脏损伤蛋鸡肝脏保护作用的研究

试验旨在研究中草药复合微生态制剂（CHCP）对急性肝脏损伤蛋鸡肝脏的保护作用。试验分两部分：①用四氯化碳豆油（SCCl₄）建立蛋鸡急性肝脏损伤模型。选取1 d健康蛋雏鸡108只随机分为4组,模型Ⅰ~Ⅲ组分别按1、2、4 mL/kg体重灌胃10%（V/V）SCCl₄,对照组按2 mL/kg体重灌胃豆油,分别在14、28及35 d时各灌胃一次;②CHCP对蛋鸡急性肝脏损伤的影响。选择1 d 健康蛋雏鸡60只随机分为5组：对照组（豆油）、模型对照组（SCCl₄）及低、中、高剂量CHCP组（SCCl₄+1‰ CHCP、SCCl₄+2‰ CHCP、SCCl₄+4‰ CHCP）,7 d起饮水使用CHCP,14、28 d按2 mL/kg体重剂量给鸡灌胃10% SCCl₄。结果表明,按2 mL/kg体重剂量在14、28 d分别灌胃10% SCCl₄即可造成蛋鸡肝脏损伤,引起肝细胞脂肪变性,空泡样变严重,核固缩,坏死;与模型对照组相比,低、中、高剂量CHCP组蛋鸡血清谷草转氨酶（AST）含量分别降低4.35%（P > 0.05）、7.57%（P > 0.05）及9.79%（P < 0.05）,中、高剂量CHCP组血清谷丙转氨酶（ALT）含量分别降低34.92%（P < 0.01）、36.51%（P < 0.01）,血清总胆红素（TB）含量分别降低25.49%（P < 0.01）、27.45%（P < 0.01）;蛋鸡肝细胞切片显示CHCP不同剂量组肝细胞充血减轻,未见脂滴空泡,排列整齐,细胞质丰富而均匀。结果表明,本试验所用的CHCP按2‰、4‰饮水使用均能减少肉鸡肝细胞坏死,降低血清转氨酶活性及TB水平,对SCCl₄诱导的肝脏损伤有保护作用。     

虫草素预防肥胖及通过ADORA1介导GH3细胞中泌乳素分泌研究

为了研究虫草素是否具有预防肥胖的作用及其可能机制,试验利用高脂饮食诱导肥胖型高脂血症大鼠动物模型,并给予高、中、低不同剂量（50、25、12.5 mg/（kg·d））的虫草素,连续灌胃35 d后观察大鼠体重、脂肪系数、血脂水平的变化;体外培养大鼠脑垂体瘤细胞（GH3）,通过CCK8、Western blotting、ELISA等方法检测虫草素对GH3细胞的增殖毒性,以及腺苷受体A₁（ADORA₁）的表达和对泌乳素分泌量的影响。体内试验结果显示,与正常对照组相比,模型组大鼠体重、脂肪系数和血脂水平（高密度脂蛋白（HDL）除外）均极显著升高（P<0.01）;虫草素高、中、低各剂量组均能极显著或显著降低大鼠体重、脂肪系数及总胆固醇（TC）、甘油三酯（TG）、低密度脂蛋白（LDL）水平（P<0.01;P<0.05）,但HDL水平仅在虫草素高剂量组中显著升高（P<0.05）,而虫草素中、低剂量组均无显著变化（P>0.05）;体外试验结果显示,虫草素浓度低于25 μg/mL时对GH3细胞增殖无影响;虫草素能够诱导GH3细胞ADORA₁表达升高;虫草素和ADORA₁激动剂（R-PIA）单独作用能显著或极显著降低泌乳素的分泌量（P<0.05;P<0.01）,虫草素与ADORA₁抑制剂（DPCPX）联合作用时能够在一定程度上阻断虫草素对泌乳素的抑制作用。结果表明,虫草素具有预防肥胖和降血脂的作用,其可能的作用机制是通过诱导ADORA₁表达抑制泌乳素的分泌,该研究结果将为虫草素作为一种潜在的减肥降脂药物的开发提供一定依据。     

促卵泡素和促黄体生成素对儋州鸡生殖激素调控规律的研究

为研究促卵泡素（FSH）和促黄体生成素（LH）对儋州鸡体内其他生殖激素的调控规律,本试验通过改变FSH和LH在儋州鸡血液中的浓度,并采用双抗体一步夹心法酶联免疫吸附试验（ELISA）对处理前后儋州鸡血液中FSH、LH、催乳素（PRL）、孕酮（P）、雌二醇（E₂）的浓度进行测定。结果发现,注射外源性FSH和LH分别能提高儋州鸡血液中FSH和LH浓度;当儋州鸡血液中FSH或LH浓度显著升高时则均能引起PRL浓度显著降低（P < 0.05）,但当FSH和LH浓度同时显著升高时,PRL浓度显著升高（P < 0.05）;当儋州鸡血液中FSH浓度显著升高时,E₂及P浓度显著提升（P < 0.05）,且在高浓度LH的协同下提升幅度更大;当儋州鸡血液中LH浓度显著升高时E₂及P浓度升高但不显著（P > 0.05）。本研究结果表明,儋州鸡血液中FSH或LH浓度的提高均能降低PRL的浓度,并能不同程度的提升E₂及P的浓度,但FSH与LH浓度同时提高则能通过协同作用刺激E₂及P浓度的大幅提升,当E₂及P浓度过高时能通过刺激PRL的释放,负反馈调节血液中FSH与LH,并恢复血液中E₂及P浓度。     

不同剂量硒蛋白粉对大鼠血常规指标和血液生化指标的影响

本试验旨在研究不同剂量硒蛋白粉对大鼠血常规指标及血液生化指标的影响。选择平均体重为70 g的健康清洁级SD大鼠80只,随机分为4组,每组2个重复,每个重复10只,雌、雄各半。对照组饲喂基础日粮,低、中和高剂量组分别饲喂添加3.5、7和10.5 mg/kg硒蛋白粉的日粮（其含量以硒计）,试验期30 d。结果表明,低、中和高剂量组大鼠血常规指标与对照组相比差异均不显著（P＞0.05）;低、中和高剂量组大鼠血清中血糖、总蛋白含量与对照组相比差异均不显著（P＞0.05）,低剂量组大鼠血清中甘油三酯、低密度脂蛋白含量均显著低于对照组（P＜0.05）;中剂量组大鼠血清中白蛋白/球蛋白及甘油三酯、低密度脂蛋白含量均显著低于对照组（P＜0.05）,而谷丙转氨酶和谷草转氨酶水平均显著高于对照组（P＜0.05）;高剂量组大鼠血清中白蛋白、白蛋白/球蛋白、甘油三酯、低密度脂蛋白含量均显著低于对照组（P<0.05）,而球蛋白、谷丙转氨酶、谷草转氨酶、尿素氮、肌酐、高密度脂蛋白含量均显著升高（P＜0.05）。综上所述,不同剂量硒蛋白粉对大鼠血常规指标无不良影响,低剂量硒蛋白粉除具有降低大鼠血脂作用外对大鼠其他血液生化指标均无不良影响,中、高剂量硒蛋白粉对大鼠的部分血液生化指标有一定影响。     

Study on Liver-protective Effect of Chinese Herbal Compound Probiotics on Acute Liver Injury Layers

This experiment was conducted to investigate the liver-protective effect of Chinese herbal compound probiotics (CHCP) on acute liver injury layers.One hundred and eight 1 day old hens were divided into 4 groups with 3 replicates in each group and 9 layers per replicate in trial 1.The layers in model groups Ⅰ to Ⅲ were gavaged with 10% (V/V) soybean oil solution of carbon tetrachloride (SCCl₄) according to 1,2 and 4 mL/(kg·BW) at 14,28 and 35 d,respectively.The layers in control group were gavaged with 2 mL/(kg·BW) soybean oil.In trial 2,sixty 1 d layers were divided into 5 groups:Control group (soybean oil),model control group(SCCl₄)and low-dose,middle-dose and high-dose CHCP group (SCCl₄+1‰ CHCP,SCCl₄+2‰ CHCP and SCCl₄+4‰ CHCP respectively).CHCP were used by drinking water since 7 days.SCCl₄ were gavaged according to 2 mL/(kg·BW) at 14 and 28 d.The results showed as follows:The model of layers liver damage could be built by intragastric administration of 2 mL/(kg·BW) 10%(V/V) SCCl₄ at 14,28 d respectively,with the signs of hepatic steatosis,severe vacuolar degeneration,nuclear condensation and necrosis.Compared to the model control group,the serum AST levels in low,medium and high dose CHCP groups were decreased by 4.35% (P > 0.05),7.57% (P > 0.05) and 9.79% (P < 0.05),the serum ALT levels in medium and high dose CHCP groups were decreased by 34.92% (P < 0.01),36.51% (P < 0.01),the serum total bilirubin content in medium and high dose CHCP groups were decreased by 25.49% (P < 0.01),27.45% (P < 0.01).The liver cell congestion was reduced to varying degrees in different dose CHCP groups,and the liver cell had no vacuoles,arranged in neat rows,abundant cytoplasm and uniform in different dose CHCP groups.In conclusion,2‰,4‰ CHCP could reduce hepatocyte necrosis,decrease the serum activities of ALT,AST and total bilirubin levels,and had protective effect on hepatic injury induced by SCCl₄.     

黄曲霉毒素B_1与M_1对小鼠肠道细菌多样性的影响

试验旨在研究黄曲霉毒素B_1与M_1(AFB_1与AFM_1)对小鼠肠道细菌多样性的影响。选取体况良好的4周龄ICR(CD-1)雄性小鼠40只,随机分成4组,每组10个重复,每个重复1只小鼠,其中AFB_1组小鼠每只每天灌胃0.3mg/kg体重AFB_1,AFM_1组小鼠每只每天灌胃3.0mg/kg体重AFM_1,AFB_1+AFM_1组每只每天灌胃AFB_1与AFM_1的混合溶液,其中AFB_1终浓度0.3mg/kg体重,AFM_1终浓度3.0mg/kg体重,以上3组毒素溶剂均为1.0%二甲基亚砜水溶液。对照组小鼠每只每天灌胃1.0%二甲基亚砜水溶液。每只灌胃剂量均为200μL,每天09∶00灌胃一次,连续灌胃28d。灌胃结束后,处死并解剖小鼠,收集结肠内容物,采用16SrRNA测序的方法对肠内容物细菌多样性进行测序分析。结果显示:在细菌群落的门水平、科水平及属水平,与对照组相比,3个毒素处理组小鼠肠道内容物细菌优势菌群均未发生明显排序变化(P>0.05),但不同的毒素处理仍造成了不同分类水平下细菌菌群丰度的显著变化:与对照组相比,AFB_1组及联合灌胃组小鼠肠内致病菌或条件致病菌,如Facklamia、Staphylococcus、Corynebacterium属细菌丰度显著升高(P<0.05);而AFM_1组与对照组相比未见显著差异(P>0.05)。综合试验结果,AFB_1单独作用或与AFM_1联合作用可诱导小鼠肠道内致病菌或条件致病菌细菌增殖,改变肠道健康微生物区系,损伤肠道微生物屏障功能。     

Study on Colloidal Gold Immunochromatographic Assay for Rapid Detection of Estradiol in Milk

In order to develop a sensitive and specific method for the detection of estradiol residues (E₂) in milk, a colloidal gold immunochromatographic method was established. Colloidal gold particles were prepared by trisodium citrate reduction method and labeled with rabbit polyclonal antibodies (PcAb) by physical adsorption method. After optimization of reaction conditions such as the amount of coating antigen and labeled antibody to assemble test strip, the sensitivity, specificity, repeatability and accelerated preservation of the test strip were determined. The estrogen analogue cross reaction and milk matrix interference reaction were also measured. The results showed that the optimized concentration of E₂-OVA antigen was 2 mg/mL and the optimized antibody concentration was 20 μg/mL, visual detection limits of E₂ was 10 μg/L in PBS within 10 min. The cross reaction rate of this method with estriol was 40%, there were no negligible cross-reactivities with other estrogen compounds including estradiol valerate, estradiol benzoate, estrone, diethylstilbestrol, quinestrol, ethinyloestradiol and nonylphenol. Milk samples only needed two times diluted before analysis, and the results could judge by naked eye after 10 min with cut-off of 20 μg/L. The results demonstrated that the developed method was suitable for rapid on-site screening of E₂ residues in milk samples.     

牛奶中雌二醇胶体金试纸条快速检测技术研究

为了建立快速、敏感、特异的雌二醇（E₂）残留免疫检测方法,本研究应用胶体金免疫层析技术,研究制备一种快速检测牛奶中E₂的方法。采用柠檬酸三钠还原法制备胶体金颗粒,采用物理吸附法将E₂兔多克隆抗体偶联至胶体金,经过优化抗原包被量和多克隆抗体标记量等反应条件组装成检测试纸条,测定检测试纸的灵敏度、特异性、重复性和加速保存等参数,并对E₂类似物交叉反应和牛奶基质干扰反应进行了测定。结果表明,试纸条最优包被抗原浓度为2 mg/mL,抗体浓度为20 μg/mL。采用消线法判定结果,检测时间10 min,PBS缓冲液中E₂检测限为10 μg/L,该方法与雌三醇的交叉反应率为40%,与戊酸雌二醇、苯甲酸雌二醇、雌酮、己烯雌酚、炔雌醚、乙炔雌二醇、壬基酚等类似物均无可见交叉反应,牛奶样品经2倍稀释消除基质干扰直接用于检测,该方法在牛奶样品中的判定值为20 μg/L。本研究建立的E₂胶体金试纸条使用简单方便,适合现场快速检测牛奶中E₂残留。     

Study on Regulation of Reproductive Hormones in Danzhou Chicken by FSH and LH

To investigate the regulation of reproductive hormones in Danzhou chicken by follicle-stimulating hormone (FSH) and luteotropic hormone (LH),the concentration of FSH and LH in Danzhou chicken blood were changed by different treatments and the concentration of FSH,LH, prolactin (PRL), progesterone (P), and estradiol (E₂) in the blood were determined using the enzyme-linked immunosorbent assay (ELISA).The results showed that exogenous FSH and LH could improve the concentrations of FSH and LH,respectively. When one of FSH and LH increased alone,the concentration of PRL was significantly decreased (P < 0.05),while it was significantly increased when FSH and LH were both increased (P < 0.05).The high level of the FSH could result the significant increase of E₂ and P (P < 0.05),and it would expand the increasing effect in cooperation with the high concentration of LH. While the high level of the LH could elevate the concentration of E₂ and P,but effect was not significant (P > 0.05).In conclusion,the study suggested that the increase level of FSH or LH could reduce the concentration of PRL in Danzhou chicken,meanwhile,it could increase the concentration of E₂ and P in varying degrees. However,both of the FSH and LH increase in the blood could result the significant increase of E₂ and P as the synergy. When the concentration of E₂ and P were too high,it could stimulate the release of PRL which could adjust the concentration of FSH and LH,and rebalance the concentrations of E₂ and P in the blood.     

Effect of Human Menopausal Gonadotropin,17β-estradiol and Epidermal Growth Factor on the Quality of in vitro Maturation Bovine Oocytes

For optimizing in vitro maturation system of bovine oocytes,we firstly examined the influence of four different hormonal regimes(FSH+LH,HMG,FSH+LH+E₂ and HMG+E₂) on oocyte maturation rates.Then we studied the effects of epidermal growth factor (EGF) in the above defined medium on bovine oocyte maturation,in vitro development and quality of parthenogenetic embryos.The cell apoptotic index of parthenogenetic blastocysts was detected by TUNEL.No significant difference was observed in maturation rates in four groups supplemented with different hormones.However,human menopausal gonadotropin (HMG) provided steady maturation results in replicates.Maturation of oocytes was promoted by supplementation with 17β-estradiol (E₂).Combination of HMG and E₂ gave rise to steady and efficient mature results.The presence of EGF at 30 ng/mL concentration significantly increased maturation rate and blastocyst rate and reduced apoptotic cells in parthenogenetic blastocysts.Therefore,the optimal oocyte maturation solution could be supplemented with 0.075 IU/mL HMG,1 μg/mL E₂ and 30 ng/mL EGF.     

人尿促性腺激素、雌二醇及表皮生长因子对牛卵母细胞体外成熟质量的影响

试验旨在研究不同激素配比及表皮生长因子(EGF)浓度对牛卵母细胞体外成熟及卵母细胞质量的影响。将随机分组的卵丘-卵母细胞复合体于添加FSH+LH、HMG、FSH+LH+E₂、HMG+E₂ 4种不同激素组合配比的成熟基础液中培养,对比其体外成熟率,比较了EGF对牛卵母细胞体外成熟率和孤雌胚胎体外发育的影响,并采用TUNEL法检测添加不同浓度EGF的牛孤雌激活囊胚细胞凋亡情况。结果表明,添加HMG的成熟试验结果稳定,E₂对牛卵母细胞成熟有一定的促进作用,HMG+E₂联合使用可以得到高效稳定的成熟结果;在此基础上,在成熟液中添加30 ng/mL EGF对牛卵母细胞的成熟质量、胚胎发育及降低胚胎细胞凋亡都有明显的促进作用。因此,在体外成熟培养液中添加0.075 IU/mL HMG、1 μg/mL E₂和30 ng/mL EGF对牛卵母细胞的成熟和质量较为有益。