晚期糖基化终末产物受体基因Gly82Ser多态性与结肠癌相关性的研究

目的 探讨结肠癌患者晚期糖基化终末产物受体(receptor for advanced glycation endproducts,RAGE) Gly82Ser基因多态性与结肠癌发生发展的关系.方法 选取90例结肠癌患者为实验组和78例年龄、性别相匹配的结肠良性息肉患者为对照组,应用聚合酶链反应-限制性片段长度多态性技术检测RAGE Gly82Ser基因多态性的分布频率.结果 RAGE Gly82Ser基因型GG(野生型纯合子)、GS(杂合子)和SS(突变型纯合子)在肿瘤组的分布频率分别为53.3％、36.7％、10.0％,在对照组的分布频率分别为76.9％、19.2％、3.8％,各基因型在肿瘤组和对照组的分布频率符合Hardy-Weinberg平衡(P＞0.05).与GG基因型相比,杂合子GS基因型和S等位基因携带者(GS+ SS)的OR值分别为2.037(95％ CI:1.207～3.438)和2.022(95％CI:1.275 ～3.208),这种差异在TNM分期Ⅲ期+Ⅳ期和低分化的结肠癌患者中尤为显著,OR值分别为3.575 (95％CI:1.495 ～8.550)和3.580(95％ CI:1.390 ～9.217).结论 RAGE Gly82Ser基因多态性不仅能增加罹患结肠癌的风险,并且与结肠癌的恶性程度密切相关.     

聚合酶链反应限制性内切酶消化法检测人载脂蛋白E基因多态性

采用聚合酶链反应与限制性核酸内切酶ＨｈａＩ消化相结合的方法建立一种简便。实用的人载脂蛋白Ｅ基因多态性检测方法。ＰＣＲ扩增含有编码１１２和１５８位氨基酸的基因片断序列，产物经ＨｈａⅠ酶消化，经聚丙烯酸 胺凝胶电泳可见２－４条特定长度的ＤＮＡ条带易于判定ａｐｏＥ基因型。     

抗甲胎蛋白单链抗体与阿霉素联合对人肝癌细胞Huh7增殖的抑制作用

探讨抗人甲胎蛋白(AFP)单链抗体和阿霉素(DOX)单用或联用对人肝癌细胞Huh7细胞增殖的抑制作用。采用MTT法检测抗AFP单链抗体、阿霉素单用及联合作用对Huh7细胞增殖的影响;采用Annexin V/PI荧光双染法检测抗AFP单链抗体、阿霉素单用及联合作用对Huh7细胞凋亡的影响;采用PI单染法检测抗AFP单链抗体、阿霉素单用及联合作用对Huh7细胞周期分布的影响。结果表明,抗AFP单链抗体、阿霉素单药组与联用组均能有效抑制Huh7细胞增殖,呈剂量依赖性,且两药联用具有协同效应;抗AFP单链抗体、阿霉素单用及联用均可诱导细胞凋亡且抗AFP单链抗体(40 μg/mL)与阿霉素(0.25 μg/mL)联合组凋亡率明显高于单独给药组,具有统计学意义(P<0.05);抗AFP单链抗体(40 μg/mL)单独作用后,Huh7的细胞周期阻滞于G₀/G₁期,阿霉素(0.25 μg/mL)单独作用Huh7的细胞周期阻滞于S期,而当两种药物联用时,Huh7的细胞周期被阻滞于G₂/M期,抑制细胞增殖。     

Detection of Mycoplasma species in cell culture by PCR and RFLP based method: effect of BM-cyclin to cure infections

PURPOSE: A two-stage nested polymerase chain reaction (PCR) assay system was described that amplifies the 16S-23S rRNA spacer region sequences of Mycoplasma and Acholeplasma infections in cell cultures and virus stocks. METHODS: Established cell lines and virus stocks were screened for the presence of Mycoplasma by using nested PCR using two sets of outer and inner primers, amplifies 16S-23S rRNA. PCR and restriction fragment length polymorphism (RFLP) assay was used to detect and identify most of the species-specific Mycoplasmas involved in cell cultures and virus stock contaminants. Infected cultures detected by PCR-RFLP were further treated with BM-cyclin (5 microg/mL) and passaged for three times and tested for Mycoplasma infections by PCR-RFLP. RESULTS: Mycoplasma pirum and Mycoplasma orale infections were detected by nested PCR. Species specificity was identified by using RFLP of Vsp I, Cla I and Hin dIII restriction enzymes. Mycoplasma infections were cured by treatment with BM-cyclin. This was further confirmed by non-amplification of PCR amplimers in BM-cyclin treated vs. non-treated cultures. CONCLUSIONS: Regular monitoring of cell cultures for Mycoplasma infections and identification of species-specific Mollicutes will identify the source of contaminations. This approach can be used for quality control of the biological reagents used in cell culture and virology laboratories.     

p53 Arg72Pro多态性与新疆维吾尔族和汉族妇女宫颈癌发生的相关性

目的探讨p53 Arg72Pro多态性与新疆维吾尔族、汉族妇女宫颈癌发生及与HPV感染宫颈癌的相关性。方法采用聚合酶链反应-限制性片段长度多态性分析（PCR—RFLP）方法检测152例维吾尔族妇女宫颈癌组织及110例非癌宫颈组织、120例汉族妇女宫颈癌组织及122例非癌宫颈组织中p53 Arg／Arg、Pro／Pro、Arg／Pro三种基因型的分布。结果比较维吾尔族妇女宫颈癌和非癌对照组中p53基因型总构成比,差异有统计学意义（χ^2=7．196,P〈0．05）,Arg／Arg所占比例在维吾尔族妇女宫颈癌组高于非癌对照组。比较汉族妇女宫颈癌和非癌对照组中p53基因型总构成比,差异有统计学意义（χ^2=8．231,P〈0．025）,Pro／Pro所占比例在汉族妇女宫颈癌组高于对照组。比较维吾尔族妇女宫颈癌中HPV16阳性组和阴性组p53基因型构成比,差异有统计学意义（χ^2=7．177,P〈0．05）,Arg／Arg所占比例在HPV16阳性组高于阴性组。结论p53 Arg／Arg基因型可能是新疆维吾尔族妇女宫颈癌发生的遗传易感因素,且与维吾尔族妇女HPV相关宫颈癌的发生有关。p53 Pro／Pro基因型可能是新疆汉族妇女宫颈癌发生的遗传易感因素。新疆维吾尔族妇女与汉族妇女宫颈癌的发生可能存在遗传易感性方面的差异。     

High throughput ranking of recombinant avian scFv antibody fragments from crude lysates using the Biacore A100

Advances in molecular evolution strategies have made it possible to identify antibodies with exquisite specificities and also to fine-tune their biophysical properties for practically any specified application. Depending on the desired function, antibody/antigen interactions can be long-lived or short-lived and, therefore, particular attention is needed when seeking to identify antibodies with specific reaction-rate and affinity properties. Surface plasmon resonance (SPR) biosensors routinely generate sensitive and reliable kinetic data from antibody/antigen interactions for both therapeutic and diagnostic applications. However, many kinetic-based screening assays require rigorous sample preparation and purification prior to analysis. To ameliorate this problem, we developed a rapid and reliable assay for characterising recombinant scFv antibody fragments, directly from crude bacterial lysates. Ninety-six scFv antibodies derived from chickens immunised with C-reactive protein (CRP) were selected by phage display and evaluated using the Biacore A100 protein interaction array system. Antibodies were captured from crude bacterial extracts on the sensor chip surface and ranked based on the percentage of the complex left (% left) after dissociation in buffer. Kinetic rate constants (k(a) and k(d)) and affinity (K(D)) data were obtained for six clones that bound monomeric CRP across a broad affinity range (2.54 x 10(-8) to 3.53 x 10(-10) M). Using this assay format the A100 biosensor yielded high quality kinetic data, permitting the screening of nearly 400 antibody clones per day.     

酒石酸美托洛尔片联合左卡尼汀注射剂治疗慢性心力衰竭的临床研究

目的观察酒石酸美托洛尔片联合左卡尼汀注射剂治疗慢性心力衰竭的临床疗效及安全性。方法将98例慢性心力衰竭患者随机分为对照组和试验组,每组49例。对照组予以口服酒石酸美托洛尔片6.25 mg,bid;试验组在对照组的基础上,予以静脉滴注左卡尼汀注射剂2 g,qd,2组均持续治疗10 d。比较2组患者的临床疗效,血清半乳糖凝集素-3(Gal-3)、糖类抗原125(CA125)和N末端脑钠肽前体(NT-proB NP)水平和药物不良反应发生率。结果试验组和对照组总有效率分别为93.87%(46例/49例)和71.42%(35例/49例),差异有统计学意义(P<0.05)。治疗后,试验组和对照组血清Gal-3分别为(4.19±0.59),(5.64±0.73)μg·L^-1;CA125分别为(48.16±5.54),(73.47±8.02)U·L^-1;NT-proB NP分别为(291.93±36.30),(389.61±45.60)μg·L^-1,差异均有统计学意义(均P<0.05)。试验组和对照组的药物不良反应主要有转氨酶上升和头痛,药物不良反应发生率分别为22.44%(11例/49例)和18.37%(9例/49例),差异无统计学意义(P>0.05)。结果酒石酸美托洛尔片联合左卡尼汀注射剂治疗慢性心力衰竭的临床疗效肯定,且安全性高。     

Indocyanine green loaded hyaluronan-derived nanoparticles for fluorescence-enhanced surgical imaging of pancreatic cancer

Pancreatic ductal adenocarcinoma is highly lethal and surgical resection is the only potential curative treatment for the disease. In this study, hyaluronic acid derived nanoparticles with physico-chemically entrapped indocyanine green, termed NanoICG, were utilized for intraoperative near infrared fluorescence detection of pancreatic cancer. NanoICG was not cytotoxic to healthy pancreatic epithelial cells and did not induce chemotaxis or phagocytosis, it accumulated significantly within the pancreas in an orthotopic pancreatic ductal adenocarcinoma model, and demonstrated contrast-enhancement for pancreatic lesions relative to non-diseased portions of the pancreas. Fluorescence microscopy showed higher fluorescence intensity in pancreatic lesions and splenic metastases due to NanoICG compared to ICG alone. The in vivo safety profile of NanoICG, including, biochemical, hematological, and pathological analysis of NanoICG-treated healthy mice, indicates negligible toxicity. These results suggest that NanoICG is a promising contrast agent for intraoperative detection of pancreatic tumors.     

Clinical translation of immunoliposomes for cancer therapy: recent perspectives

Introduction: Liposomes have been extensively investigated as drug delivery vehicles. Immunoliposomes (ILs) are antibody-conjugated liposomes designed to selectively target antigen-expressing cells. ILs can be used to deliver drugs to tumor cells for improving efficacy and reducing toxicity. In addition, ILs can be used in immunoassays, immunotherapy, and imaging. Although there has been extensive coverage on ILs in the literature, only a limited number of clinical trials have been reported and no IL drug has been approved by the FDA.

Areas covered: Factors to consider in developing ILs are discussed, including the choice of antibody or antibody fragment, the formulation of liposomes, and the conjugation chemistry. In addition, challenges and opportunities in clinical development of ILs are discussed. The purpose of this review is to provide an overview on the state of the art of ILs and to discuss potential future developments.

Expert opinion: IL research has had a lengthy history and numerous preclinical studies have yielded encouraging results. However, there are a number of obstacles to clinical translation of ILs. Given the unique capabilities of ILs, its potential for clinical application is underexplored. There is great potential for expanded role for ILs in the clinic and further efforts to this end are warranted.

Abbreviations: Ab: antibody; ADCs: antibody-drug conjugates; API: active pharmaceutical ingredient; ADCC: antibody-dependent cellular cytotoxicity; CR: complete remission; cGMP: current good manufacturing practice; DSPE: distearoyl phosphatidylethanolamine; EGF: epidermal growth factor; EGFR: epidermal growth factor receptor; EPR: enhanced permeability and retention; Fc: fragment crystalline; Tf: transferrin; HACA: human-anti-chimeric antibody; HAHA: human-anti-human antibody; HAMA: human-anti-mouse antibody; HER2: human epidermal growth factor 2; IL: immunoliposome; LNPs: lipid nanoparticles; MRI: magnetic resonance imaging; MTD: maximum tolerated dose; PEG: polyethylene glycol; PET: positron emission tomography; PR: partial response; PSMA: prostate-specific membrane antigen; scFv: single-chain variable fragment; SPECT: single photon emission computed tomography; TTR: transthyretin     

Characteristics of intercalary fragment in posterior malleolus fractures

BackgroundThe purpose of the study is to investigatethe prevalence of posterior malleolus intercalary fragment (ICF) in adult ankle fractures; to analyze the morphological characteristics of ICF; and its mechanism.MethodsRetrospectively, 247 cases of posterior malleolus fractures (PMF) were included in 369 cases of adult ankle fractures. The prevalence of ICF, relationship between type of ankle fracture, posterior malleolar fracture and the ICF was studied.ResultsAmong 369 cases, the ICF was found in 43% of the PMF and 29% of the hospitalized unstable ankle fractures. Position of ICF is posterolateral in 68 (64%), mid posterior in 20 (19%) and posteromedial in 18 (17%) cases. The prevalence of ICF was 70% in Bartoníček classification type III fractures, which was significantly higher than type I, II and IV.ConclusionsThe highest incidence occurs in Bartoníček III type fracture. The study helps in further understanding and treatment of the PMF.Level of evidenceLevel IV, case series.