一种新型免疫层析技术在临床乳腺癌Her-2蛋白定量检测中的应用

建立一种靶点蛋白质快速定量检测方法。在原有侧向流动免疫层析技术的基础上,通过优化层析材料和纳米微球的均一性、改进检测区的检测方法,经逐点扫描技术,建立标准浓度曲线,以达到对临床靶点蛋白质的定量检测。以乳腺癌组织中的Her2表达为例,通过对已知浓度样品的检测,验证本技术方法的准确度大于96%。另外,以蛋白质免疫印迹作为组织中特定蛋白质检测金标准,分析临床肿瘤组织中Her2蛋白的含量,其准确率也达到95.5%,而免疫组织化学方法检测准确率仅为69.58%。新型免疫层析法检测结果与靶向治疗患者的愈后密切相关(P<0.01)。改进后的新型免疫层析方法能够准确地对临床靶点蛋白质进行定量检测,而且结合侧向流动技术的简单、快速和易用性,这种新型检测方法可以广泛应用于临床组织标本、血液标本和体液标本中靶点蛋白质的临场定量检测,在一定程度上可以替代免疫组化技术。     

EPA,not DHA,prevents fibrosis in pressure overload-induced heart failure: potential role of free fatty acid receptor 4

Heart failure with preserved ejection fraction (HFpEF) is half of all HF, but standard HF therapies are ineffective. Diastolic dysfunction, often secondary to interstitial fibrosis, is common in HFpEF. Previously, we found that supra-physiologic levels of ω3-PUFAs produced by 12 weeks of ω3-dietary supplementation prevented fibrosis and contractile dysfunction following pressure overload [transverse aortic constriction (TAC)], a model that resembles aspects of remodeling in HFpEF. This raised several questions regarding ω3-concentration-dependent cardioprotection, the specific role of EPA and DHA, and the relationship between prevention of fibrosis and contractile dysfunction. To achieve more clinically relevant ω3-levels and test individual ω3-PUFAs, we shortened the ω3-diet regimen and used EPA- and DHA-specific diets to examine remodeling following TAC. The shorter diet regimen produced ω3-PUFA levels closer to Western clinics. Further, EPA, but not DHA, prevented fibrosis following TAC. However, neither ω3-PUFA prevented contractile dysfunction, perhaps due to reduced uptake of ω3-PUFA. Interestingly, EPA did not accumulate in cardiac fibroblasts. However, FFA receptor 4, a G protein-coupled receptor for ω3-PUFAs, was sufficient and required to block transforming growth factor β1-fibrotic signaling in cultured cardiac fibroblasts, suggesting a novel mechanism for EPA. In summary, EPA-mediated prevention of fibrosis could represent a novel therapy for HFpEF.     

Physical limits on kinesin-5–mediated chromosome congression in the smallest mitotic spindles

A characteristic feature of mitotic spindles is the congression of chromosomes near the spindle equator, a process mediated by dynamic kinetochore microtubules. A major challenge is to understand how precise, submicrometer-scale control of kinetochore micro­tubule dynamics is achieved in the smallest mitotic spindles, where the noisiness of microtubule assembly/disassembly will potentially act to overwhelm the spatial information that controls microtubule plus end–tip positioning to mediate congression. To better understand this fundamental limit, we conducted an integrated live fluorescence, electron microscopy, and modeling analysis of the polymorphic fungal pathogen Candida albicans, which contains one of the smallest known mitotic spindles (<1 μm). Previously, ScCin8p (kinesin-5 in Saccharomyces cerevisiae) was shown to mediate chromosome congression by promoting catastrophe of long kinetochore microtubules (kMTs). Using C. albicans yeast and hyphal kinesin-5 (Kip1p) heterozygotes (KIP1/kip1∆), we found that mutant spindles have longer kMTs than wild-type spindles, consistent with a less-organized spindle. By contrast, kinesin-8 heterozygous mutant (KIP3/kip3∆) spindles exhibited the same spindle organization as wild type. Of interest, spindle organization in the yeast and hyphal states was indistinguishable, even though yeast and hyphal cell lengths differ by two- to fivefold, demonstrating that spindle length regulation and chromosome congression are intrinsic to the spindle and largely independent of cell size. Together these results are consistent with a kinesin-5–mediated, length-dependent depolymerase activity that organizes chromosomes at the spindle equator in C. albicans to overcome fundamental noisiness in microtubule self-assembly. More generally, we define a dimensionless number that sets a fundamental physical limit for maintaining congression in small spindles in the face of assembly noise and find that C. albicans operates very close to this limit, which may explain why it has the smallest known mitotic spindle that still manifests the classic congression architecture.     

Role of 5-aminolevulinic acid in the formation of winter rape resistance to sulfonylurea herbicides

Growing of winter rape (Brassica napus L.) plants for 7–8 days in the presence of a sulfonylurea herbicide Magnum (methsulfuron methyl; 200 and 500 mg/L) exerted an organ-specific influence on the seedlings: suppressed hypocotyl and root growth and increased the accumulation of fresh weight of cotyledonary leaves. Exogenous 5-aminolevulinic acid (ALA; 0.1, 1.0, and 10 mg/L) partially negated the adverse effect of Magnum on the length and fresh weight of roots and hypocotyls and stimulated the development of cotyledons. The herbicide suppressed the activity of ascorbate peroxidase (APX). The addition of ALA to the 200 mg/L herbicide solution caused a steady activation of APX as compared with the effect of herbicide alone. In plants grown on Magnum solutions, the activity of glutathione reductase (GR) rose. Positive effect of exogenous ALA was only observed when 500 mg/L herbicide was used (Magnum-500 type of treatment). In plants grown on the Magnum solution, the content of reduced and oxidized forms of glutathione rose. In contrasty, exogenous ALA reduced the total content of glutathione but in this case the ratio between its reduced and oxidized forms rose. In the presence of the herbicide, the content of anthocyans considerably decreased and upon the addition of exogenous ALA their levels rose. In rape plants treated with Magnum, the ability to generate superoxide anion radical was essentially the same as in control plants; however, the content of hydrogen peroxide (H₂O₂) therein rose. The addition of ALA to the herbicide reduced the ability to generate superoxide anion radical and the level of H₂O₂. The activity of acetolactate synthase remained on the control level when 200 mg/L Magnum was used, decreased in the plants of Magnum-500 treatment, and rose upon the addition of exogenous ALA to this type of treatment. It was concluded that ALA had a positive effect on the development of winter rape resistance to Magnum via stimulation of growth processes, activation of APX and GR, predominant elevation of the content of reduced glutathione and anthocyans, and partial recovery of acetolactate synthase activity, which on the whole caused a decrease in the H₂O₂ level and in the ability of plants to generate superoxide anion radical.