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《中国兽医学报》2020,(3)
采用组织块贴壁法分离鸡成骨细胞并进行体外培养,细胞纯化后经姬姆萨、甲苯胺蓝、碱性磷酸酶(ALP)和茜素红染色鉴定;在成骨细胞培养液中分别添加0,10,50,100μmol/L褪黑素,连续14 d测定成骨细胞增殖情况;在第1,3,7,14天测定诱导分化培养的成骨细胞ALP活性,并在第14天检测成骨细胞BSP基因表达水平。甲苯胺蓝染色结果与小鼠胚胎成骨细胞前体细胞系MC3T3-E1一致;ALP染色呈阳性,可见大量黑色颗粒;茜素红染色后矿化结节呈深红色。成骨细胞ALP活性在第3天有所降低,但随培养时间延长,ALP活性逐渐升高;添加褪黑素对细胞增殖的影响差异不显著(P0.05),但在第1天和第14天显著提高成骨细胞ALP活性(P0.05);添加褪黑素提高了BSP基因的相对表达量(P=0.09)。结果表明,利用组织块贴壁法,分离得到鸡颅骨成骨细胞,确定了褪黑素促进成骨细胞分化,且具有时间和浓度依赖性,但不影响增殖。 相似文献
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骨是镉毒性作用的主要靶器官之一,但其对鸡骨髓基质细胞(bone marrow stromal cells,BMSCs)增殖和成骨分化的毒性作用仍不清楚。本研究利用差速贴壁纯化法获得鸡BMSCs,加入不同浓度镉处理不同时间,采用CCK-8法检测细胞增殖,碱性磷酸酶(alkaline phosphatase,ALP)和茜素红染色鉴定成骨分化,RT-PCR检测成骨相关基因(COL1、OSX、RUNX2、ALP、OCN、OPG、OPN、RANKL)表达变化。结果显示,1~10μmol/L的镉显著促进BMSCs增殖,20μmol/L的镉显著抑制其增殖(P<0.05);5μmol/L以上的镉可显著抑制ALP活性,并以浓度依赖方式极显著抑制成骨相关基因(COL1、OSX、RUNX2、ALP、OCN、OPG、OPN)mRNA表达,上调RANKL mRNA表达(P<0.01)。表明,一定浓度镉可抑制鸡BMSCs体外增殖及其向成骨细胞(OB)的分化。 相似文献
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利用不同浓度的醋酸铅溶液(0、20、40、80μmol/L)作用成骨细胞24 h,研究醋酸铅对体外培养新生SD大鼠成骨细胞增殖分化和钙调蛋白(Calmodulin,CaM)的影响。结果表明,成骨细胞的增殖功能和ALP活性随着醋酸铅浓度的增加而显著降低(P<0.05或P<0.01),呈现一定的剂量-效应关系;与对照组相比,染铅组成骨细胞ALP活性显著下降(P<0.05),而钙调蛋白mRNA和蛋白的表达逐渐增加,当铅浓度为40μmol/L时,差异显著或极显著(P<0.05或P<0.01)。表明醋酸铅通过影响成骨细胞的增殖分化活力、ALP活性和钙调蛋白的表达,发挥其毒性损伤作用。 相似文献
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《中国兽医学报》2020,(2)
脂联素受体激动剂(AdipoRon)是人工合成的与脂联素(ADPN)有相似作用的口服活性小分子,研究显示ADPN可调节成骨细胞生长和分化,但AdipoRon是否具有类似的功能目前鲜见报道。本试验主要研究AdipoRon对鸡成骨细胞的影响。从14日龄鸡胚额骨中分离获得成骨细胞,细胞培养第4日,分别添加100,200 mg/L AdipoRon处理成骨细胞,处理72 h后,MTT法检测细胞增殖活性,并进行碱性磷酸酶(ALP)染色。Real-time PCR检测脂联素受体1(AdipoR1)、脂联素受体2(AdipoR2)、成骨细胞成熟标志基因骨钙素(osteocalcin,OC)、Ⅰ型胶原α2链(alpa2 of type I collegen,COL1A2)、细胞凋亡相关基因Caspase-3、Bcl-2及Bax的基因表达量,计算Bcl-2与Bax基因表达量的比值。结果显示,100,200 mg/L AdipoRon处理后的成骨细胞正常形态消失,细胞数量减少,体积变小,细胞核明显固缩,细胞存活率极显著降低(P<0.01)。AdipoRon可增加成骨细胞中AdipoR1、AdipoR2、OC、Bcl-2、Bax和Caspase-3的表达量(P<0.05),并呈剂量依赖性,但对COL1A2和Bcl-2/Bax的表达无显著性影响。结果表明,100,200 mg/L AdipoRon均可促进鸡成骨细胞AdipoR1/2的表达,且能抑制成骨细胞的增殖,促进成骨细胞的成熟及凋亡。 相似文献
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《畜牧兽医学报》2018,(12)
成骨细胞在骨代谢过程中发挥成骨作用,笔者制备羊骨髓肽钙螯合物(MP-Ca),探究其对体外培养成骨细胞的增殖及活性的影响,并将其效果同羊骨髓肽(MP)进行比较。制备的MP-Ca经傅里叶红外光谱仪进行扫描鉴定;从新生SD大鼠颅骨中分离纯化的成骨细胞经碱性磷酸酶(ALP)和茜素红染色鉴定;在培养液中添加不同浓度的受试物,CCK-8法确定最佳作用浓度。以17-β雌二醇(10~2μg·mL~(-1))为阳性对照,通过测定各组细胞OD_(450 nm)和ALP、骨钙素(BGP)的含量,考察MP、MP-Ca在最适作用浓度(10~3μg·mL~(-1))下对成骨细胞增殖和活性的影响。结果表明,MP和MP-Ca的红外光谱图未完全重叠,吸收峰发生了明显的位移变化,说明MP与CaCl_2反应形成了MP-Ca。分离的成骨细胞经ALP染色呈阳性,胞内可见大量黑褐色颗粒;茜素红染色后细胞呈深红色,并形成钙化结节。MP-Ca处理组的OD_(450 nm)和ALP、BGP水平均显著(P 0. 05或P 0. 01)高于MP处理组,但均显著(P 0.05或P 0. 01)低于雌激素组。综上,尽管效果不及雌激素,MP-Ca和MP均可促进体外培养成骨细胞的增殖并增强其成骨活性,效果MP-Ca优于MP。 相似文献
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EGF和IGF-1对山羊卵母细胞体外成熟的影响 总被引：3，自引：0，他引：3
研究了在培养液中添加不同浓度的EGF、IGF-1以及EGF联合IGF-1对山羊有腔卵母细胞体外成熟的影响。结果表明: (1)10, 20, 30μg/LEGF对山羊卵母细胞体外成熟无显著影响(P>0.05)。(2) 10μg/L的IGF-1对山羊卵母细胞体外成熟无显著影响(P>0.05); 20, 30μg/L的IGF-1能显著提高山羊卵母细胞体外成熟率 (P<0.05)。(3)添加 10μg/LEGF+20μg/LIGF-1组卵母细胞体外成熟率显著高于添加 20μg/L的IGF-1组(P<0.05),极显著高于添加 10μg/LEGF组和对照组(P<0.01)。可见,EGF和IGF-1对山羊卵母细胞体外成熟有协同作用。 相似文献
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取新生奶牛肋骨,剪成1 mm3大小骨片,胰蛋白酶消化5次,D-Hank's液彻底冲洗后,骨片置于含15?S的DMEM中培养,获取单层细胞.采用倒置显微镜下观察细胞形态,改良钙-钴法染色,检测碱性磷酸酶(ALP)活性、骨钙素(OC)mRNA丰度及骨钙素蛋白浓度等方法,对细胞进行鉴定.通过MTT法绘制生长曲线,研究细胞增殖特性,茜素红染色观察细胞钙化功能.结果显示,采用本法所获得的细胞具有典型的成骨细胞形态和特性:ALP染色呈阳性,活性水平较高;培养7 d后可检测到骨钙素mRNA表达,12 d后细胞上清液中可检测到骨钙素蛋白;诱导培养2周后细胞可形成钙化结节.提示采用胰蛋白酶消化,组织块移行法所培养的细胞具有成骨细胞特性.本方法可获得高纯度、高活性的奶牛成骨细胞. 相似文献
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钙对体外培养大鼠成骨细胞增殖分化及细胞周期的影响 总被引：1，自引：1，他引：0
在体外培养大鼠成骨细胞的过程中,添加不同剂量的钙(0、1、2、4 mmol/L),通过检测细胞周期、细胞增殖能力、碱性磷酸酶(ALP)活性及骨桥蛋白表达,探讨了钙对成骨细胞增殖分化及细胞周期的影响.结果表明,2、4 mmol/L钙组在1～8 d均显著或极显著地促进成骨细胞增殖(P<0.05或P<0.01),1 mmol/L钙组则在5、6、8 d有显著或极显著的差异(P<0.05或P<0.01);添加不同浓度的钙除1 mmol/L组第2天外的不同时间均极显著地抑制细胞内ALP活性(P<0.01);1 mmol/L钙能极显著的使成骨细胞滞留在S期(P<0.01);添加不同浓度的钙均能使G2期显著或极显著的增加(P<0.05或P<0.01),G1期极显著的下降(P<0.01),并能极显著地诱导细胞内骨桥蛋白的表达(P<0.01).表明添加不同浓度的钙均能抑制其早期分化,促进成骨细胞的增殖,使细胞滞留在S期或G2期,诱导细胞在基质成熟期分化,有利于细胞的钙化. 相似文献
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《黑龙江畜牧兽医》2010,(21)
为了研究胰岛素样生长因子-1(insulin-like growth factor,IGF-1)对奶牛成骨细胞骨重建相关因子表达的影响,探讨IGF-1对奶牛成骨细胞形成-吸收的作用,试验采用体外分离培养奶牛成骨细胞,用改良Gomori钙钴法染色鉴定,再以0,1,10,50,100 ng/mL人重组胰岛素样生长因子-1(recombinated human insulin-like growth factor-1,rhIGF-1)干预培养5 d后,半定量RT-PCR法检测成骨细胞骨保护素(osteoprotegerin,OPG)和核因子κB受体活化因子配体(receptor activator of nu-clear factor-κB ligand,RANKL)mRNA的表达,ELISA检测细胞培养上清液OPG蛋白水平。结果表明:1~100 ng/mL rhIGF-1干预5 d,OPG、RNAKL mRNA表达均较未干预组增加,10,50,100 ng/mL组OPG/β-actin、RANKL/β-actin比值显著高于未干预组(P0.05),且在1~100 ng/mL浓度范围内,rhIGF-1呈剂量依赖方式上调OPG蛋白表达,其中10,50,100 ng/mL干预组与未干预组差异显著(P0.05)。说明IGF-1调控奶牛成骨-破骨细胞介导的形成-吸收偶联,促进骨形成,这可能是IGF-1参与奶牛骨代谢性疾病的重要机制之一。 相似文献
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骨碎补水提液对大鼠成骨细胞的影响 总被引：1，自引：0，他引：1
为观察骨碎补水提液对体外培养大鼠成骨细胞增殖、分化的影响,用体外培养新生SD大鼠颅盖骨分离的成骨细胞,将不同浓度的骨碎补水提液分别与第3代大鼠成骨细胞进行体外共同培养,采用噻唑蓝(MTT)法测定细胞增殖能力,对硝基苯磷酸盐法(PNPP法)测定细胞的碱性磷酸酶(ALP)活性.结果显示,与对照组相比,作用48 h、72 h,骨碎补水提液均能显著促进大鼠成骨细胞的增殖,且与时效有关;作用48 h ,10-3、10-2 mg/mL骨碎补水提液浓度组可显著升高细胞ALP活性(P<0.05).表明骨碎补水提液中存在较高活性的促大鼠成骨细胞增殖和分化的物质. 相似文献
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魏静 《四川畜牧兽医学院学报》2009,(4):28-32
在现代法律秩序中,商会自治规范是制定法的基础和必要的补充,甚至在某些方面替代了制定法;商会自治规范主要包括商会组织规范、行为规范、惩罚规范以及争端解决规范等;其效力仅及于其内部成员;商会自治规范和制定法之间存在冲突,但也存在整合的基础。 相似文献
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本文概述了猪的毛色类型、猪的毛色遗传模式,着重综述了猪毛色基因分子基础的研究进展,指出存在问题并就未来发展方向做了思考。 相似文献
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以国际标准强毒Ｒ株人工感染非免疫产蛋鸡,定时扑杀,分别从鼻窦、眶下孔、气管、肺、气囊、卵巢和输卵管分离ＭＧ,并收集感染鸡所产蛋分离ＭＧ。结果表明,人工感染４８小时后上、下呼吸道及肺已被全面感染,９６小时气囊已被感染,１２０小时输卵管已能分离到ＭＧ,卵巢始终分离不到ＭＧ。人工感染鸡自１４４小时便能在其所产蛋中分离出ＭＧ。药物治疗能在７２小时内消除感染,油乳剂苗则需２４天后逐渐降低蛋内ＭＧ分离率,药物卵内注射、种蛋药浴、高温处理均能杀死卵内ＭＧ,但以研制的种蛋浸泡剂药浴效果为最好。 相似文献
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REASONS FOR PERFORMING STUDY: Centesis of the bicipital bursa using an 8.9 cm long spinal needle has been reported but the alternative of employing a 3.8 cm long hypodermic needle requires validation. OBJECTIVE: To compare the efficacy of 2 different methods of centesis of the bicipital bursa and to evaluate the usefulness of ultrasonographic imaging to determine the location of solution administered when centesis of the bursa is attempted. METHODS: For Trial 1, 6 clinicians, who had no previous experience of centesis of the bicipital bursa, attempted to inject a solution composed of an aqueous radiopaque contrast medium and physiological saline solution (PSS) into the bicipital bursae of 2/12 horses using the previously described distal approach to inject one bursa and a proximal approach to inject the contralateral bursa. The bicipital tendon and bursa were examined ultrasonographically before and after injection; and both shoulders were examined radiographically to identify the location of the medium. In Trial 2, another 6 clinicians, also with no previous experience of centesis, repeated Trial 1, using 6 horses, but the radiopaque contrast medium was mixed with air instead of PSS. RESULTS: Accuracy of centesis using the proximal approach was 39% and that of the distal approach 28%. Ultrasonographic examination of the shoulder allowed the location of solution and air to be accurately predicted in all 12 shoulders examined. CONCLUSIONS: Clinicians who have had no previous experience performing centesis of the bicipital bursa are unlikely to be successful in centesis using either approach. Radiographic examination after injecting a radiopaque contrast medium may be necessary to assess the success of centesis especially if bursal fluid is not obtained during centesis. Injecting air along with the radiopaque contrast medium provides more accurate ultrasonographic confirmation of centesis and better radiographic definition than does injection without air. 相似文献
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Effects of size of ingestively masticated fragments of plant tissues on kinetics of digestion of NDF
Ingestively masticated fragments were collected and sized via sieving. Different sizes of esophageal masticate and ruminal digesta fragments, and ground fragments of larger masticated pieces were incubated in vitro, and undigested NDF remaining at intervals of up to 168 h of incubation was determined. The ruminal age-dependent time delay (tau) for onset of digestion of NDF was positively correlated (P < 0.004) with the mean sieve aperture estimated to retain 50% of the fragments between successive sieve apertures (MRA). Degradation rate of potentially degradable NDF (PDF) and level of indigestible NDF were not related (P > 0.10) to MRA of masticated and ground fragments. Estimates of tau were positively related to MRA, with slopes of bermudagrass < corn silage < ruminal fragments of corn silage. It was concluded that fragment size-, and consequently, ruminal age-dependent onset of PDF degradation of a mixture of different fragment sizes results in an age-dependent rate of degradation of the more rapidly degrading of two subentities of PDF. Models are proposed that assume a tau before onset of simultaneous degradation of PDF from two pools characterized as having gamma-modeled age-dependency and age-constant rates. The ruminal age-dependent pool seems to be associated with the faster-degrading pool, and its rate parameter increases with range in MRA in the population of fragments. Conceptually, the ruminal age-dependent rate parameter for PDF degradation seems to represent a composite of several effects: 1) effects of the size-dependent tau; 2) range in MRA of the population of ingestively masticated fragments; and 3) subentities of PDF that degrade via more rapid age-dependent rates compared with subentities of PDF that degrade via age-constant rates. The estimated fractional rates of ruminative comminution of ingestively masticated fragments (0.060 to 0.075/h) were of a magnitude similar to the mean fractional rates of PDF digestion (0.030 to 0.085/h), which implies that ruminative comminution may be first-limiting to fractional rate of PDF digestion. The in vivo roles of ingestive and ruminative mastication of fragments on PDF degradation must be considered in any kinetic system for estimating PDF digestion in the rumen. These results and others in the literature suggest that the rate of surface area exposure rather than intrinsic chemical attributes of PDF may be first-limiting to degradation rate of PDF in vivo. 相似文献