P-糖蛋白在多药耐药的K562细胞转运苯妥英纳与卡马西平中的作用

研究证实,多药转运体与难治性癫痫耐药机制密切相关,P-糖蛋白在其中起重要作用．主要研究P-糖蛋白拮抗剂维拉帕米对P-糖蛋白过表达的K562细胞耐药性及细胞内苯妥英纳与卡马西平浓度的影响．首先建立了P-糖蛋白高表达的K562/Dox(阿霉素诱导)耐药细胞株,比较耐药细胞株和P-糖蛋白表达阴性的K562细胞株对苯妥英纳和卡马西平的耐药性,并观察给予维拉帕米后,耐药细胞内抗癫痫药物的浓度变化．结果发现,苯妥英纳和卡马西平对K562/Dox细胞株的半数抑制浓度(IC₅₀)明显高于K562细胞株,加入维拉帕米后,苯妥英纳和卡马西平对K562/Dox 细胞的IC₅₀明显下降,逆转倍数分别为2.5和1.5．进一步研究发现,K562/Dox细胞内苯妥英纳和卡马西平的浓度均显著少于其药敏K562细胞,仅分别为正常K562细胞的23.6%和32.2%．当加入维拉帕米后,K562/Dox细胞内抗癫痫药物浓度明显升高(P < 0.05)．由此证明,高表达的P-糖蛋白参与了细胞的药物转运,在难治性癫痫的耐药机制中扮演重要角色．     

酸弗林蛋白酶酸性氨基酸簇分选蛋白-1的原核表达、纯化及多克隆抗体制备

目的：原核表达和纯化PACS-1,并制备其多克隆抗体。方法：通过RT-PCR扩增出PACS-1的编码基因,测序正确后克隆入原核表达载体pGEX4T-1,转化大肠杆菌BL21（DE3）,以IPTG诱导PACS-1与GST融合蛋白的表达并经Glutathione Sepharose 4B纯化;经SDS-PAGE和Western blot鉴定,应用纯化的蛋白免疫家兔制备多克隆抗体,用ELISA测定抗体的效价。结果：表达和纯化了PACS-1,并获得了较高效价的抗血清。结论：获得纯化的PACS-1及其多克隆抗体,为进一步研究PACS-1的功能奠定了基础。     

可表达随机12肽库的逆转录病毒表达系统的建立及评价

目的：建立可表达随机12肽库的逆转录病毒表达系统。方法：体外合成编码随机12肽的DNA片段;在最优化的实验参数和反应条件下将DNA片段克隆入带有EGFP标记的逆转录病毒载体后分批次电击转化大肠杆菌,合并转化所得菌液即为可表达随机12肽库的逆转录病毒原始载体库;半固体扩增法扩增该原始载体库,提取质粒并转染GP2-293包装细胞,在EGFP表达最强的时间点收集细胞培养上清,即为可表达随机12肽库的逆转录病毒库。结果：可表达随机12肽库的逆转录病毒原始载体库的库容量为3.14×10^6cfu;扩增后的逆转录病毒载体库滴度为5.2×109cfu/mL,库容量为2.34×1011cfu;转染了已扩增的载体库质粒后的GP2-293包装细胞可以成功地表达随机12肽库。结论：建立了可表达随机12肽库的逆转录病毒表达系统,为抗病毒寡肽的筛选以及进一步的深入研究奠定了良好的基础。     

A tumor-specific kinase activity regulates the viral death protein Apoptin

Apoptin, a chicken anemia virus-encoded protein, is thought to be activated by a general tumor-specific pathway, because it induces apoptosis in a large number of human tumor or transformed cells but not in their normal, healthy counterparts. Here, we show that Apoptin is phosphorylated robustly both in vitro and in vivo in tumor cells but negligibly in normal cells, and we map the site to threonine 108. A gain-of-function point mutation (T108E) conferred upon Apoptin the ability to accumulate in the nucleus and kill normal cells, implying that phosphorylation is a key regulator of the tumor-specific properties of Apoptin. An activity that could phosphorylate Apoptin on threonine 108 was found specifically in tumor and transformed cells from a variety of tissue origins, suggesting that activation of this kinase is generally associated with the cancerous or pre-cancerous state. Moreover, analyses of human tissue samples confirm that Apoptin kinase activity is detectable in primary malignancies but not in tissue derived from healthy individuals. Taken together, our results support a model whereby the dysregulation of the cellular pathway leading to the phosphorylation of Apoptin contributes to human tumorigenesis.     

新疆地区绿僵菌菌株筛选及抗逆性研究

本文对新疆地区土壤中分离得到的17株绿僵菌Metarhizium菌株以东亚飞蝗Locusta migratoria manilensis为供试昆虫进行毒力测定,筛选获得高毒力菌株,并对筛选后的高毒力绿僵菌菌株进行耐短时高温能力、抗紫外线能力和耐干旱能力的测试,分析高毒力绿僵菌菌株的抗逆性,以期获得致病力高且抗逆性好的菌株,为下一步绿僵菌生物农药的开发提供依据。研究发现,M1-17、M1-13、M1-09、M1-16、M1-05五株菌株为对东亚飞蝗高致病力的菌株,平均僵虫率在80.00%~96.67%之间,LT50在2.92~3.65之间。对高温的抗性效果较好的菌株为M1-17和M1-05;对紫外线的抗性效果较好的菌株为M1-17和M1-16;而菌株M1-09、M1-17和M1-16抗旱能力较好。菌株M1-17较其它菌株具有更好的抗逆性,具有很好的开发利用价值。     

APOE and KLF14 genetic variants are sex-specific for low high-density lipoprotein cholesterol identified by a genome-wide association study

To demonstrate the loci that relate to high-density lipoprotein cholesterol (HDL-C) levels and genetic sex heterogeneity, we enrolled 41,526 participants aged between 30 and 70 years old from the Taiwan Biobank in a genome-wide association study. We applied the Manhattan plot to display the p-values estimated for the relationships between loci and low HDL-C. A total of 160 variants were significantly associated with low HDL-C. The genotype TT of rs1364422 located in the KLF14 gene has 1.30 (95% CI=1.20 - 1.42) times the risk for low-HDL compared to genotype CC in females (log(-p) =8.98). Moreover, the genes APOC1, APOE, PVRL2, and TOMM40 were associated significantly with low-HDL-C in males only. Excluding the variants with high linkage disequilibrium, we revealed the rs429358 located in APOE as the major genetic variant for lowering HDL-C, in which genotype CT has 1.24 (95% CI= 1.16 - 1.32) times the risk. In addition, we also examine 12 genes related to HDL-C in both sexes, including LPL, ABCA1, APOA5, BUD13, ZPR1, ALDH1A2, LIPC, CETP, HERPUD1, LIPG, ANGPTL8, and DOCK6. In conclusion, low-HDL-C is a genetic and sex-specific phenotype, and we discovered that the APOE and KLF14 are specific to low-HDL-C for men and women, respectively.     

A Meta-Analysis of Core Stability Exercise versus General Exercise for Chronic Low Back Pain

Objective

To review the effects of core stability exercise or general exercise for patients with chronic low back pain (LBP).

Summary of Background Data

Exercise therapy appears to be effective at decreasing pain and improving function for patients with chronic LBP in practice guidelines. Core stability exercise is becoming increasingly popular for LBP. However, it is currently unknown whether core stability exercise produces more beneficial effects than general exercise in patients with chronic LBP.

Methods

Published articles from 1970 to October 2011 were identified using electronic searches. For this meta-analysis, two reviewers independently selected relevant randomized controlled trials (RCTs) investigating core stability exercise versus general exercise for the treatment of patients with chronic LBP. Data were extracted independently by the same two individuals who selected the studies.

Results

From the 28 potentially relevant trials, a total of 5 trials involving 414 participants were included in the current analysis. The pooling revealed that core stability exercise was better than general exercise for reducing pain [mean difference (−1.29); 95% confidence interval (−2.47, −0.11); P = 0.003] and disability [mean difference (−7.14); 95% confidence interval (−11.64, −2.65); P = 0.002] at the time of the short-term follow-up. However, no significant differences were observed between core stability exercise and general exercise in reducing pain at 6 months [mean difference (−0.50); 95% confidence interval (−1.36, 0.36); P = 0.26] and 12 months [mean difference (−0.32); 95% confidence interval (−0.87, 0.23); P = 0.25].

Conclusions

Compared to general exercise, core stability exercise is more effective in decreasing pain and may improve physical function in patients with chronic LBP in the short term. However, no significant long-term differences in pain severity were observed between patients who engaged in core stability exercise versus those who engaged in general exercise.

Systematic Review Registration

http://www.crd.york.ac.uk/PROSPERO PROSPERO registration number: CRD42011001717.     

Heat shock protein 104 inhibited the fibrillization of prion peptide 106-126 and disassembled prion peptide 106-126 fibrils in vitro

Amyloid-like fibrils have been associated with the pathogenesis of human prion diseases. Prion peptide of aa 106-126 (PrP106-126) exhibits many PrP(Sc)-like biochemical features, forming amyloid-like fibrils in vitro. Here, we found that the recombinant yeast-derived molecular chaperon Hsp104 inhibited significantly the fibril assembly of the synthetic PrP106-126 peptide by dynamic ThT assays in vitro. EM assays revealed almost no fibril-like structure after incubation of the synthetic PrP106-126 peptides with Hsp104 for 12h. Circular dichroism assays identified that treatment of Hsp104 shifted the secondary structure of PrP106-126 fibrils from β-sheet to a random coil. MTT tests confirmed that interaction of PrP106-126 with Hsp104 maintained the toxicity of PrP106-126 on human neuroblastoma cell line SK-N-SH. Additionally, Hsp104 was able to disassemble the mature PrP106-126 fibrils in vitro, leading to recovering the cytotoxicity of PrP106-126 on SK-N-SH cells. Our study provides the molecular evidences that the yeast-derived Hsp104 can interfere in the fibril assembly and disassembly of human PrP106-126 segment.     

火木层孔菌发酵菌粉及其组分对小鼠移植性肝癌H22的体内生长作用

采用肝癌H22荷瘤小鼠的肿瘤模型,对火木层孔菌(桑黄)Phellinus igniarius发酵菌粉及其各提取物组分的体内抗肿瘤活性进行评价。结果表明,火木层孔菌发酵菌粉及其各提取物组分对荷瘤小鼠肝癌H22都具有一定的抗肿瘤作用并能延长荷瘤小鼠生存期,其中,火木层孔菌发酵菌粉(1,000mg/kg/d)及其组分I(I多糖组分)(360mg/kg/d)具有较为显著的抗肿瘤作用,抑瘤率分别为33.5%和40.3%。组织病理学研究结果表明在菌粉及其多糖组分作用后,肿瘤细胞坏死细胞明显增多,免疫组化检测表明火木层孔菌发酵菌粉及其多糖组分能明显的降低瘤组织中Bcl-2基因蛋白的表达,提高小鼠瘤组织中的Bax基因蛋白的表达。火木层孔菌发酵菌粉及其多糖组分具有较好的体内抗肿瘤活性。