Identification of novel thiazolo[5,4-d]pyrimidine derivatives as human A1 and A2A adenosine receptor antagonists/inverse agonists

In this study a new set of thiazolo[5,4-d]pyrimidine derivatives was synthesized. These derivatives bear different substituents at positions 2 and 5 of the thiazolopyrimidine core while maintaining a free amino group at position-7. The new compounds were tested for their affinity and potency at human (h) A₁, A_2A, A_2B and A₃ adenosine receptors expressed in CHO cells. The results reveal that the higher affinity of these new set of thiazolopyrimidines is toward the hA₁ and hA_2A adenosine receptors subtypes and is tuned by the substitution pattern at both the 2 and 5 positions of the thiazolopyrimidine nucleus. Functional studies evidenced that the compounds behaved as dual A₁/A_2A antagonists/inverse agonists. Compound 3, bearing a 5-((2-methoxyphenyl) methylamino) group and a phenyl moiety at position 2, displayed the highest affinity (hA₁ K_i?=?10.2?nM; hA_2A K_i?=?4.72?nM) and behaved as a potent A₁/A_2A antagonist/inverse agonist (hA₁ IC₅₀?=?13.4?nM; hA_2A IC₅₀?=?5.34?nM).     

Immunoperoxidase Staining for Estrogen and Progesterone Receptors in Archival Formalin Fixed, Paraffin Embedded Breast Carcinomas after Microwave Antigen Retrieval

Immunoperoxidase staining was performed for estrogen and progesterone receptors in 93 cases of primary breast carcinoma. Breast tumor samples were fixed in formalin and embedded in paraffin. Antigen retrieval was performed by microwave heating in citrate buffer, pH 6.0, using precisely defined and reproducible conditions. The cases studied included material from the current year and from paraffin blocks retrieved from archival storage dating back to 1981. In all cases, estrogen and progesterone receptor values determined by biochemical assay were available for comparison with the immunohistochemical results. We found 94% agreement of results between the two methods.     

Effects of 2.45 GHz microwave fields on liposomes entrapping glycoenzyme ascorbate oxidase: evidence for oligosaccharide side chain involvement.

Previous observations reported by our group indicate that 2.45 GHz microwave fields at specific absorption rate (SAR) of 5.6 W/kg reduce the enzyme activity rate of ascorbate oxidase (AO) trapped in liposomes. In this study, we report dose-response studies on these AO containing liposomes irradiated at different SAR values (1.4, 2.8, 4.2, and 5.6 W/kg). No response was observed for SAR below 5.6 W/kg. Liposomes entrapping functional AO in its deglycated form (AO-D) were also used. In this case, no MW related enzyme activity changes were observed, demonstrating a direct involvement of oligosaccharide chains of AO. Furthermore, the catalytic properties of both AO and AO-D were not impaired by MW irradiation, neither in homogeneous solution nor loaded in liposomes, excluding possible changes in the conformation of enzyme as a mechanism. Our results suggest that the oligosaccharide chains of AO are critical to elicit the microwave observed effects on lipid membrane.     

Limiting racemization and aspartimide formation in microwave-enhanced Fmoc solid phase peptide synthesis.

Microwave energy represents an efficient manner to accelerate both the deprotection and coupling reactions in 9-fluorenylmethyloxycarbonyl (Fmoc) solid phase peptide synthesis (SPPS). Typical SPPS side reactions including racemization and aspartimide formation can occur with microwave energy but can easily be controlled by routine use of optimized methods. Cysteine, histidine, and aspartic acid were susceptible to racemization during microwave SPPS of a model 20mer peptide containing all 20 natural amino acids. Lowering the microwave coupling temperature from 80 degrees C to 50 degrees C limited racemization of histidine and cysteine. Additionally, coupling of both histidine and cysteine can be performed conventionally while the rest of the peptide is synthesized using microwave without any deleterious effect, as racemization during the coupling reaction was limited to the activated ester state of the amino acids up to 80 degrees C. Use of the hindered amine, collidine, in the coupling reaction also minimized formation of D-cysteine. Aspartimide formation and subsequent racemization of aspartic acid was reduced by the addition of HOBt to the deprotection solution and/or use of piperazine in place of piperidine.     

Microwave-assisted RCM for the synthesis of carbocyclic peptides.

Microwave irradiation dramatically improves the efficiency of ring closing metathesis (RCM) reactions of resin-attached peptides and the technology is illustrated by the highly selective synthesis of dicarba analogues of alpha-conotoxin IMI.     

基于微波遥感技术探测森林地表土壤含水率

森林地表土壤含水率是森林生态系统中的重要参数,使用微波遥感技术快速准确地估算区域尺度上的森林地表土壤含水率,对于森林生态系统研究具有重要的现实意义.本文利用TDR-300土壤含水率速测仪测得黑龙江大兴安岭地区塔河林业局盘古林场内120块样地的森林地表土壤含水率作为因变量,利用C波段全极化SAR数据的极化分解参数作为自变量,构造多元线性回归统计模型和BP神经网络模型,定量估测森林地表土壤含水率,通过模型反演获得区域尺度上森林地表土壤含水率的空间分布.结果表明: 多元线性回归统计模型的精度为86.0%,均方差根误差(RMSE)为3.0%;BP神经网络模型的精度为89.4%,RMSE为2.7%.说明利用BP神经网络模型定量估测森林地表土壤含水率优于多元线性回归模型,将全极化SAR数据通过BP神经网络模型进行仿真,最终得到研究区域的森林地表土壤含水率空间分布图.     

Epitope mapping of anti‐myelin oligodendrocyte glycoprotein (MOG) antibodies in a mouse model of multiple sclerosis: microwave‐assisted synthesis of the peptide antigens and ELISA screening

The role of pathologic auto‐antibodies against myelin oligodendrocyte glycoprotein (MOG) in multiple sclerosis is a highly controversial matter. As the use of animal models may enable to unravel the molecular mechanisms of the human disorder, numerous studies on multiple sclerosis are carried out using experimental autoimmune encephalomyelitis (EAE). In particular, the most extensively used EAE model is obtained by immunizing C57BL/6 mice with the immunodominant peptide MOG(35–55). In this scenario, we analyzed the anti‐MOG antibody response in this model using the recombinant refolded extracellular domain of the protein, MOG(1–117). To assess the presence of a B‐cell intramolecular epitope spreading mechanism, we tested also five synthetic peptides mapping the 1–117 sequence of MOG, including MOG(35–55). For this purpose, we cloned, expressed in Escherichia coli and on‐column refolded MOG(1–117), and we applied an optimized microwave‐assisted solid‐phase synthetic strategy to obtain the designed peptide sequences. Subsequently, we set up a solid‐phase immunoenzymatic assay testing both naïve and EAE mice sera and using MOG protein and peptides as antigenic probes. The results obtained disclose an intense IgG antibody response against both the recombinant protein and the immunizing peptide, while no response was observed against the other synthetic fragments, thus excluding the presence of an intramolecular epitope spreading mechanism. Furthermore, as the properly refolded recombinant probe is able to bind antibodies with greater efficiency compared with MOG(35–55), we hypothesize the presence of both linear and conformational epitopes on MOG(35–55) sequence. Copyright © 2015 European Peptide Society and John Wiley & Sons, Ltd.     

遥感技术在鸟类生态学研究中的应用

获取鸟类活动及生境信息是鸟类生态学研究的基础, 而遥感技术弥补了传统野外调查方法的缺陷, 提供了获取多种信息的新途径。应用遥感技术的鸟类生态学研究热点从最初的种群行为观察, 到栖息地选择, 再到生境适宜性、破碎化及人为干扰探究等, 随着技术的不断发展也在扩展和变化。不同波段或组合下的遥感技术各有所长。光学遥感应用广泛, 尤其是信息量较大的红外波段图像和作为野外鸟巢及物种活动监测常用工具的红外相机; 多光谱图像常用于栖息地制图以及地物识别, 高空间分辨率的数据甚至可对鸟类种群进行直接计数; 高光谱数据则可对光谱特征相似的地物进行更为精确的区分和反演; 激光雷达遥感主要用于栖息地植被结构的三维探测, 为了解鸟类栖息地选择提供更好的依据。微波遥感在飞鸟探测上应用颇多, 近年来多极化数据在复杂栖息地精确制图上也具有优势, 但成本较高、解译复杂且推广度较低。在实际应用中, 遥感数据时空尺度的选择会影响研究结果, 部分遥感反演参数也缺乏生态学意义。多源遥感数据的结合应用能够提升制图分类的精度, 实现数据的时空分辨率互补, 优化鸟类生态研究所需参数。未来的遥感技术在鸟类生态学中的应用应致力于提供更加明确的光谱信息、相对简便的解译方法, 以及更为合理的多源数据组合方式等。     

以松毛虫为基质培养的冬虫夏草菌丝体中多糖的提取研究

研究了不同方法对冬虫夏草发酵菌丝体（以枯叶蛾科昆虫马尾松毛虫为基质发酵所得）中多糖提取的影响。结果表明,采用微波辅助水提法所得多糖的产率最高,影响提取的关键因素为液料比、微波提取时间、微波功率;采用Box-Behnken设计及响应面分析法对这3个因素进行优化,并通过回归拟合,建立了预测虫草多糖提取的多项式模型Y=6.87+0.058A+0.085B+0.075C+0.032AB+0.046AC+0.069BC-0.16A²-0.37B²-0.11C²。经响应面最优化分析,获得冬虫夏草发酵菌丝体中多糖的最优提取工艺参数为：液料比（mL/g）6.3:1、微波功率520W、微波提取时间326s,此工艺提取验证后的提取率达到6.76%。     

微波辐射对肾小管上皮细胞的影响以及金雀异黄素对其保护作用

目的：观察微波辐照对人近端肾小管上皮细胞（HKC）的影响及金雀异黄素对其的保护作用。方法：HKC分为对照组、微波辐照组、金雀异黄素组（n=6）。金雀异黄素组在辐照前2 h用含30μmol/L金雀异黄素的DMEM培养基进行预培养。辐照后24 h留取上清进行乳酸脱氢酶（LDH）、β-N-乙酰氨基葡萄糖苷酶（NAG）活性及丙二醛（MDA）、超氧化物歧化酶（SOD）活性检测。Hoechst 33258染色观察细胞凋亡情况。结果：与对照组比较,微波辐照组上清NAG、LDH活性明显增加（P < 0.01）,金雀异黄素预处理组则较微波辐照组明显下降（P < 0.01）;微波辐照组上清活性也较对照组明显增加（P < 0.01）。Hoechst 33258染色显示,微波辐照可导致较多量的细胞凋亡,而应用金雀异黄素预处理后细胞凋亡的比例均大大减少。微波辐照可大大提高HKC细胞中的MDA含量,SOD活性降低（P< 0.01）,应用金雀异黄素预处理后MDA的含量无明显降低,SOD的活性明显增大（P < 0.01）。结论：微波辐照可导致人近端肾小管上皮细胞出现功能损伤,金雀异黄素对其具有一定的保护作用,可能与其减轻氧化应激损伤、减少细胞凋亡有关。