Semi-quantitative analysis of cytokine mRNA expression induced by the herbal medicine Sho-saiko-to (TJ-9) using a Gel Doc system.

The RT-PCR method was employed to determine the cytokine mRNA expression of human peripheral lymphocytes induced by the Japanese herbal medicine Sho-saiko-to (TJ-9). The results showed that the mRNA expression of IL-12, IL-1beta, IL-10, TNF-alpha, G-CSF, and IFN-gamma increased after 6 hr in culture. This is the first reported finding that TJ-9 is an IFN-gamma inducer. Next, cytokine mRNA expression was semi-quantitatively measured using the Gel Doc system with a CCD camera and then statistically analyzed in order to determine which component of TJ-9 was the true cytokine inducer. The results showed that the scutellaria root is the main component inducing the cytokines, while the glycyrrhiza root is the secondary component. When the cytokine concentrations in the supernatants of cell cultures were measured by ELISA, the levels of IL-12, IL-1beta, IL-10, TNF-alpha, and G-CSF reflected mRNA expression levels in the cell fraction. However, the level of IFN-gamma was below the detectable limit. The effects of various reagents on many different kinds of cytokine mRNA expression could be analyzed objectively in a short time using the Gel Doc system. Many important findings could be demonstrated by this simple, easy, sensitive, and cheap method. After the clinical significance of cytokine analysis is confirmed, this method may become a useful clinical examination tool.     

Flow Cytometry Panel-Reactive Antibody Screening of Anti-HLA Antibodies in the Waiting List Significantly Reduces the Occurrence of Acute Rejection After Kidney Transplantation

Introduction

The presence of preformed HLA-reactive antibodies in recipient serum before transplantation has long been recognized as a prominent risk factor for a generally worse graft outcome. Screening and identification of HLA antibodies can be used to stratify patients into high- and low-risk categories.

过表达miR-520d 通过抑制自噬蛋白Beclin1 逆转三阴性乳腺癌细胞化疗耐药性

[摘要] 目的：探讨miR-520d 通过调控自噬逆转三阴性乳腺癌（TNBC）细胞化疗耐药的作用及分子机制。方法：以人TNBC细胞系MDA-MB-231 和MDA-MB-468 为亲本株细胞构建多西他赛（Doc）耐药细胞株MDA-MB-231/Doc 和MDA-MB-468/Doc,实验分为空白组（亲本细胞）、对照组（耐药细胞组）和过表达miR-520d 组。用qPCR检测空白组和耐药细胞组细胞中miR-520d 的表达水平,MTT实验检测过表达miR-520d 的耐药细胞对Doc 的敏感性,MDC染色后荧光显微镜观察细胞中自噬小体的发生情况、共聚焦显微镜观察过表达miR-520d 的耐药细胞中自噬相关蛋白LC3 阳性的细胞数。用荧光素酶报告基因实验验证miR-520d 与Beclin1 的靶向关系。用WB实验检测过表达miR-520 对细胞中自噬相关蛋白Beclin1 和LC3Ⅰ、LC3Ⅱ表达的影响。结果：TNBC耐药细胞中miR-520d 的表达水平明显低于空白组细胞（P<0.01）。过表达miR-520d 的耐药细胞对Doc的敏感性显著提高（P<0.01）、细胞的自噬活性明显降低（P<0.01）。荧光素酶报告基因实验证明Beclin1 是miR-520d 可能的靶分子。Doc 与miR-520d mimics 联合使用可降低TNBC耐药细胞中LC3-Ⅱ/Ⅰ比值和自噬蛋白Beclin1 的表达水平（均P<0.05）。结论：通过调控miR-520d 水平可能改变自噬蛋白Beclin1 表达,从而逆转TNBC细胞Doc化疗耐药性。     

应用基因组步移克隆小鼠Doc-1R基因组序列

目的：获得小鼠Doc-1R基因组序列。方法：根据小鼠Doc-1R基因的cDNA序列设计、合成特异引物,应用基因组步移策略对小鼠基因组步移文库进行扩增。以含有特殊接头（adaptor)的小鼠基因组步移文库作为模板,用巢式PCR法扩增目的片段。结果：应用此方法得到约1．5kb的目的片段,经测序分析证实Doc-1R基因克隆成功。此基因含有4个外显子、3个内含子,外显子与内含子接头符合GT－AG法则。结论：基因组步移方法简单、可靠、有效,是一种比较理想的克隆基因组片段的方法。     

Personalising docetaxel and G-CSF schedules in cancer patients by a clinically validated computational model

Background:

This study was aimed to develop a new method for personalising chemotherapeutic and granulocyte colony-stimulating factor (G-CSF) combined schedules, and use it for suggesting efficacious chemotherapy with reduced neutropenia.

Methods:

Clinical data from 38 docetaxel (Doc)-treated metastatic breast cancer patients were employed for validating a new pharmacokinetic/pharmacodynamics model for Doc, combined with a mathematical model for granulopoiesis. An optimisation procedure was constructed and used for selecting improved treatment schedules.

Results:

The combined model accurately predicted observed nadir timing (r=0.99), grade 3 or 4 neutropenia (86% success) and neutrophil counts over time in individual patients (r=0.63), and showed robustness to CYP3A-induced variability in Doc clearance. For average patients, the predicted optimal support for the standard chemotherapy regimen, Doc 100 μg m⁻² tri-weekly, is G-CSF, 300 μg, Q1D × 3, starting day 7 post-Doc. This regimen largely moderates chemotherapy-induced neutrophil nadir and neutropenia duration. The more intensive Doc dose, 150 mg m⁻², is optimally supported by the slightly less cost-effective G-CSF 300 μg, Q1D × 4, 5 days post-Doc. The latter regimen is optimal for borderline patients (2000 neutrophils per μl) under Doc, 100–150 mg m⁻² tri-weekly.

Conclusions:

The new computational method can serve for tailoring efficacious cytotoxic and supportive treatments, minimising side effects to individual patients. Prospective clinical validation is warranted.     

肺癌细胞株中NVB和Doc药物敏感性关联基因的研究

目的:筛选小细胞肺癌(SCLC)和非小细胞肺癌(NSCLC)细胞株中与诺维苯(NVB)、泰索帝(Doc)药物感受性相关的基因.方法:采用MTT比色分析法测定4株SCLC细胞株和6株NSCLC细胞株对NVB、Doc的药物感受性,同时应用cDNA macroarray技术检测10株肺癌细胞株中1291个药物感受性关联基因的表达状态,分析二者之间的相关性.结果:与NVB和Doc药物敏感性关联(r≥±0.4)的基因共有56个.1)与NVB药物敏感性相关联的基因有36个,SCLC NSCLC组与NSCLC组共同表达的基因有20个,其中,SCLC NSCLC组表达的基因有27个,特异表达的有7个,NSCLC组为29个基因,9个基因特异表达.2)与Doc药物敏感性相关联的基因有50个,其中,SCLC NSCLC组与NSCLC组共同表达的基因有12个,SCLC NSCLC组表达的基因有24个,12个基因特异表达,NSCLC组为38个基因,26个基因特异表达.3)肺癌细胞株中与NVB、Doc药物感受性相关联基因的分类主要分布于:信号转导分子、代谢酶类及其抑制剂、细胞因子和转录因子等4类.结论:SCLC和NSCLC细胞株中NVB、Doc药物敏感性相关联基因存在明显差异,但分类基本相同.     

Doc McStuffins and Dora the Explorer: representations of gender,race, and class in US animation

Children are exposed to animated cartoons at an early age, and the gender, class, and race-related messages are likely to be influential in the development of children’s beliefs and attitudes about gender roles, class, and racial groups. Although representations of females and minority children in animated cartoons have improved over the years, stereotypical and racially biased portrayals of females and racial minority groups remain. Using an intersectionality theoretical lens, a textual analysis was conducted to examine the representation of the two current leading, minority female characters in children’s animated cartoons: Doc McStuffins and Dora the Explorer. Character portrayals, stereotypes, challenges to traditional stereotypes, and the intersecting role of race, gender, and class on the representations of the characters are explored in this analysis.