上皮间质转化促乳腺癌机制及药物干预治疗研究进展

乳腺癌是犬、猫等伴侣动物与人类常发疾病,作为人类及动物常患恶性肿瘤和主要致死肿瘤之一,其疾病负担仍呈逐步加重趋势,乳腺癌的预防及治疗形势愈加严峻。上皮间质转化(EMT)是乳腺癌发生发展中重要的生物学过程。EMT还可促进恶性肿瘤的侵袭、扩散及耐药,因此它在肿瘤的研究中日益受到关注,靶向于EMT是治疗乳腺癌的重要研究方向与热点。文章就EMT发生过程中细胞形态功能及标志物的变化、EMT分类及EMT与乳腺癌的关系分别展开论述,详细解析了EMT相关TGF-β/Smad、NF-κB及Wnt信号通路转导途径;随后对乳腺癌治疗药物研究进展,包括TGF-β/Smad通路抑制剂开发,相关药物、基因及细胞因子治疗前景、NF-κB通路与Wnt通路抑制剂的动物试验研究结果进行了详细论述;最后对乳腺癌的治疗发展与趋势进行了展望。深入认识信号通路调控乳腺癌EMT的生物学过程,明确其发生发展机制,寻找关键靶点及开发靶向药物,将为乳腺癌的精准治疗带来曙光。     

猪TGFβ1基因及其受体组织表达特性的研究

以妊娠母猪为研究材料,采用RT-PCR半定量方法对猪转化生长因子β1(TGFβ1)及其3种受体组织表达特点进行研究。结果表明:TGFβ1在11种组织中的表达量,由高到低依次为肾上腺、下丘脑、肌肉、卵巢、脾、大脑、肾、子宫、肺、肝和心脏。3种受体的表达,总体上来说在子宫中相对较高。TGFβ1及其受体在肾上腺和子宫等组织中有相对较高的表达量,提示其在繁殖过程中可能发挥作用。     

可变剪接体WNT4-β对山羊卵泡颗粒细胞增殖和激素分泌的影响

旨在研究WNT4的一个可变剪接体（WNT4-β）对山羊卵泡颗粒细胞增殖的影响。本研究选取4~6月龄健康母羊20只,采集双侧卵巢,体外分离卵泡颗粒细胞进行培养。通过免疫荧光染色技术确定WNT4-β的表达位置;在山羊颗粒细胞中过表达或干扰WNT4-β后,利用RT-qPCR、Western blot检测WNT4-β和WNT信号通路中关键标记因子ROA1、RHOA及颗粒细胞增殖标记基因cyclin-D2、CDK4的表达变化;CCK-8技术检测颗粒细胞增殖情况;并通过ELISA分析颗粒细胞中生殖激素水平的变化。免疫荧光染色结果显示,WNT4-β只在山羊卵泡颗粒细胞中表达,在卵母细胞不表达;过表达WNT4-β后,WNT4-β和颗粒细胞增殖因子cyclin-D2、CDK4的mRNA相对表达量极显著增加（P<0.01）,蛋白表达水平显著增加（P<0.05）;WNT信号通路标记因子ROA1、RHOA mRNA表达水平显著增加（P<0.05）,β-catenin蛋白表达水平显著增加（P<0.05）;干扰WNT4-β后,WNT4-β、cyclin-D2、CDK4、ROA1和RHOA 的mRNA表达显著降低（P<0.05）,WNT4-β、cyclin-D2、CDK4及β-catenin蛋白表达显著降低（P<0.05）。CCK-8结果显示,过表达WNT4-β促进颗粒细胞增殖（P<0.05）;ELISA结果显示,过表达WNT4-β后,颗粒细胞中雌二醇（estradiol,E₂）水平显著增加（P<0.05）,孕酮（progesterone,P₄）水平升高但不显著（P>0.05）;干扰WNT4-β后则结果相反,颗粒细胞增殖受到抑制（P<0.05）,E₂和P₄的水平显著降低（P<0.05）。综上所述,WNT4可变剪接体WNT4-β通过调控WNT信号通路促进山羊卵泡颗粒细胞增殖及类固醇激素分泌,本研究为解析WNT4调控山羊颗粒细胞增殖的潜在分子机制提供理论基础。     

黑种草子提取物对小鼠酒精性肝损伤的修复作用及机制研究

【目的】验证黑种草子提取物对小鼠酒精性肝损伤的修复作用并探究其机制,为其临床开发研究提供理论依据。【方法】将56只3周龄昆明小鼠随机均分为7组：空白对照组,模型组,黑种草子提取物高、中、低剂量组,黑种草子粉剂组及水飞蓟宾阳性对照组,每组8只。除空白对照组外,其余各组均以10 mL/kg 60%酒精灌胃,每天1次,连续15 d。第16天,黑种草子提取物高、中、低剂量组分别给予8、4、2 mL/kg黑种草子提取物,每天1次,连续10 d后处死小鼠。计算各组小鼠肝脏指数;检测血清中谷丙转氨酶（alanine aminotransferase,ALT）、谷草转氨酶（aspartate aminotransferase,AST）、过氧化氢酶（catalase,CAT）、超氧化物歧化酶（superoxide dismutase,SOD）和谷胱甘肽过氧化物酶（glutathione peroxidase,GSH-Px）活力;制备肝脏石蜡切片观察其病理变化;Western blotting检测NOD样受体热蛋白结构域相关蛋白3（NOD-like receptor thermal protein domain associated protein 3,NLRP3）、含半胱氨酸的天冬氨酸蛋白水解酶1（cysteinyl aspartate specific proteinase 1,Caspase-1）和白细胞介素1β（interleukin-1β,IL-1β）的蛋白表达水平。【结果】与空白对照组相比,模型组小鼠肝脏指数显著上升（P<0.05）,肝细胞出现明显颗粒变性,水泡变性,核溶解,界限区分不清,肝索消失;血清ALT、AST活力显著上升（P<0.05）,SOD、CAT、GSH-Px活力显著下降（P<0.05）,说明肝损伤模型建立成功,且肝脏组织中NLRP3、Caspase-1和IL-1β蛋白表达水平显著升高（P<0.05）。与模型组相比,黑种草子提取物各剂量组血清ALT、AST活力均呈下降趋势,SOD、CAT、GSH-Px活力均呈上升趋势,其中高剂量组上述各指标均呈显著差异（P<0.05）,肝脏组织结构明显改善,NLRP3、Caspase-1和IL-1β蛋白表达水平均显著下降（P<0.05）。【结论】高剂量（8 mL/kg）黑种草子提取物可显著提高肝损伤小鼠的抗氧化水平,且可以通过抑制NLRP3炎性小体的表达而减少IL-1β的合成,从而改善肝脏功能和修复受损的肝脏组织结构。     

肉桂醛催化选择加氢制氢化肉桂醛

采用煤质活性炭作载体,分别用HNO3、H2O2和(NH4)2S2O8进行预处理,然后用浸渍法负载PdCl2,在H2流中还原得到Pd/C催化剂,用于肉桂醛(CAL)选择加氢制氢化肉桂醛(HCAL)的反应。考查了Pd负载量,反应温度和压力对加氢反应的影响,优化了反应条件。进一步考查了助剂Fe、Co、Ni[1]对反应的影响。最终,CAL转化率为98%,HCAL选择性为88%。     

Wnt/β-catenin信号通路上游基因在山羊绒周期性再生及着色过程中的表达分析

为初步探究Wnt/β-catenin信号通路是否参与山羊绒周期性再生及着色过程,本试验采用实时荧光定量技术对Wnt/β-catenin信号通路上游基因β-catenin、Lef1/Tcf3及该通路拮抗基因Dkk1mRNA在白色绒山羊绒毛生长不同时期及黑、白色绒山羊绒毛生长旺盛期时体侧部皮肤组织中的相对表达量进行研究。结果显示,上述通路中β-catenin、Lef1/Tcf3基因mRNA在白色绒山羊不同时期体侧部皮肤组织中的相对表达量具有明显变化规律即生长前期缓慢上调,旺盛期时表达量最高,生长后期及退行期逐渐下调,休止期表达量最低;Dkk1基因mRNA在白色绒山羊不同时期体侧部皮肤组织中的相对表达量则无明显规律性。上述各基因mRNA在黑、白色绒山羊生长旺盛期时体侧部皮肤组织中的相对表达量并无显著差异(P0.05)。结果提示,Wnt/β-catenin信号通路参与山羊绒周期性再生过程,但该通路并不涉及生长旺盛期时绒毛的着色过程。     

Induction of defense response of Oryza sativa L. against Pyricularia grisea (Cooke) Sacc. by treating seeds with chitosan and hydrolyzed chitosan

Seeds of rice (Oryza sativa L.) were treated with chitosan and hydrolyzed chitosan at 100, 500 and 1000 mg L⁻¹. After 18 days of germination, spore suspension of Pyricularia grisea was applied. The enzyme activity of phenylalanine ammonia-lyase, β-1-3-glucanase, chitinase and chitosanase in leaves of rice seedlings was evaluated after 24, 72, 120 and 168 h of inoculation. Blast affected area (%) was evaluated 7 and 14 days after spraying spore suspension. Chitosan performance to elicit defense response induction was associated with the concentration and type of chitosan. The activity of most of the enzymes tested was induced in leaves of treated seeds with chitosan and hydrolyzed chitosan at 1000 and 500 mg L⁻¹, respectively. The highest enzyme activities were observed with hydrolyzed chitosan after 72 h however, compared to chitosan, the activity was not maintained during the entire post-inoculation period. The highest control (0 = no lesions) of P. grisea in rice seedlings was observed at 1000 mg L⁻¹ in both chitosan and hydrolyzed chitosan treated leaves. Symptoms of infection by P. grisea were evident after 14 days evaluation date, but according to the standard scale proposed by the International Rice Research Institute, these symptoms fell into the resistance category of blast diseases.     

Identification and quantification of benzimidazole resistance polymorphisms in Haemonchus contortus isolated in Northeastern Brazil

Haemonchus contortus is the most prevalent nematode in Brazil. The objective of this study was to select 6 populations of H. contortus of known or suspected benzimidazole resistance status and characterize these using quantitative real-time polymerase chain reaction (qPCR) for single nucleotide polymorphisms (SNPs) F200Y, F167Y and E198A in the β-tubulin isotype 1 gene. qPCR was performed using DNA from a pool of 10 adult male H. contortus from a single animal per farm. Faecal egg count reduction test (FECRT) and egg hatch test (EHT) were used to determine the resistance status. Samples were obtained from 6 farms located in 5 counties in the Ceará State: Tauá, Boa Viagem, Quixadá, Santa Quitéria and Solonópole. The inbred-susceptible-Edinburgh (ISE) isolate was used as reference for comparative purposes in the qPCR. Benzimidazole resistance was detected by FECRT on all farms with efficacy values ranging from 0 to 51%. EC50 values as determined by EHT were all above 1.49 μg/ml. High frequencies of the resistant SNPs F200Y and F167Y alleles were detected but no resistance was detected at SNP E198A. Our results suggest that the SNPs F167Y and F200Y are both important for benzimidazole resistance in the studied populations.     

糖对长白猪精液NAGaseⅡ的抑制机理

以长白猪精液为材料,采用经典的纯化方法获得N-乙酰-β-D-氨基葡萄糖苷酶（EC 3.3.1.52,NAGase）的2种同工酶,命名为：NAGaseⅠ和NAGaseⅡ。其中NAGaseⅡ的比活力为358.21U/mg,纯化倍数为6.37倍。以海藻糖、D-甘露糖、蔗糖、葡萄糖为效应物,研究其对该酶活性的影响。结果表明：在特定浓度范围内,海藻糖对酶活力基本没有影响;D-甘露糖、蔗糖、葡萄糖对该酶均有一定的抑制作用;D-甘露糖对酶的抑制作用表现为非竞争型,抑制常数KI为11.94mmo/L;蔗糖对酶的抑制作用表现为竞争型,抑制常数KI为0.56mmol/L;葡萄糖对酶的抑制作用表现为混合型,抑制常数KI和KIS分别为0.35mol/L和64.29mmol/L。