肝癌热休克蛋白—抗原肽复合物的构建及诱导CTL反应的初步研究

目的：研究体外构建热休克蛋白-抗原肽复合的方法,观察其在体外的抗肿瘤作用。方法：用经43℃热处理1小时的人肝细胞悬液,经过裂解液裂解,60%-80%饱和硫酸铵沉淀,用SephadexG-100柱制备,取分子量为70KD组份,Westen blot进行性质鉴定。应用多肽解离液处理该组份,SephadexG-25柱过滤获得未结合多肽的蛋白分子,使其在体外与肝癌抗原肽SLIVHLNEV结合,构建成热休克蛋白-抗原肽复合物。应用此复合物树突状细胞,激活同源外周血T淋巴细胞产生肿瘤特异性杀伤T淋巴细胞（CTL）,应用MTT法检测其对T2细胞及肿瘤细胞的杀伤活性。结果：所得蛋白经电泳及Western blot进行蛋白分子量及性质鉴定为热休克蛋白70。SephadexG-25柱双分离法证实应用上述方法成功构建了肝癌热休克蛋白70-抗原肽复合物,用该复合物负荷树突状细胞在体外可以诱导出较强的肝癌抗原肽物特异性CTL,可以杀伤负荷有该肽的T2细胞及递呈该肽的肿瘤细胞系。结论：体外构建的热休克蛋白-抗原肽复合物可以增强抗原肽诱导CTL反应能力,热休克蛋白是良好的T细胞免疫佐剂,有可能在肿瘤疫苗治疗中发挥重要作用。     

不寐方组方配伍历史沿革

系统分析历代不寐方发展演变及组方选药的特点。汉唐时期组方以清热泻火药和滋阴养血药为主,宋金元时期以清心安神、补气养心、养血安神药为主,又出现气血双补、交通心肾、滋阴潜阳、补益肝肾类方剂,在治法上取得了突破性进展;明清时期以养血安神方、重镇安神方、祛痰化饮方、交通心肾方为主,和胃消食方、理气解郁方、活血化瘀方的出现进一步完善了不寐证的治疗思路。     

盐黄柏炮制历史沿革和机制的研究进展及其质量标志物（QMarker）预测分析

盐黄柏是黄柏Phellodendri Chinensis Cortex经盐水加工后的炮制品,具有滋阴降火的功效,在我国具有广泛且悠久的应用历史。通过查阅历代本草典籍及现代研究文献报道,梳理盐黄柏的历史沿革,并对其现代炮制机制的研究进展进行归纳。从药性、盐制后化学成分与功效变化、药动学及体内过程相关性等方面对盐黄柏的质量标志物(quality marker,Q-Marker)进行预测分析,初步确定小檗碱、小檗红碱、黄柏碱等为其可能的Q-Marker,以期为盐黄柏的质量评价、临床有效应用及相关基础研究提供科学依据。     

温郁金、温莪术、片姜黄饮片的炮制工艺沿革及现代研究

温郁金、温莪术、片姜黄均为道地药材。温郁金炮制方法主要有炒制、酒制、醋制、蒸制;温莪术炮制方法主要有酒制、醋制、蒸制;片姜黄炮制方法主要是切制。目前,三者的化学成分研究主要以莪术油、莪术二酮、榄香烯类和姜黄素类等物质为主,主要活性成分为挥发油和姜黄素类。温郁金具有抗肿瘤、抗炎、镇痛解热、抗氧化、抗血栓等作用;温莪术具有抗肿瘤、抗突变、抗病毒、抗炎、镇痛、抗血栓、提高免疫等作用;片姜黄具有抗肿瘤、抗炎、抗早孕、终止妊娠等作用。三者均具有抗肿瘤、抗炎、抗血栓等功效。综述温郁金、温莪术、片姜黄炮制工艺的发展历史及现代研究进展,为其炮制研究提供参考和依据。     

基于肺络理论论治肺结节病

随着肺结节病发病率及检出率的逐年上升,肺结节病逐渐引起各界的关注及重视,对肺结节病CT随访期进行有效的干预治疗,延缓进展,控制生长,以致消除结节,是现代医学面临的关键问题。从肺络论治肺结节病,着手于肺络之内涵、结构、生理特点等,进而探究肺络虚损是肺结节病的病理基础、肺络-气机失调是肺结节病发生的病理环节、肺络-气郁痰瘀停聚是肺结节病的重要病理演变、肺络郁闭是肺结节病的基本病机。最终,立足于中医络病理论的优势,提出通络法论治肺结节病,以期为临床论治肺结节打开新的诊疗思路,提供新的理论认识。     

Accumulation and maintenance of information in evolution

Selection accumulates information in the genome—it guides stochastically evolving populations toward states (genotype frequencies) that would be unlikely under neutrality. This can be quantified as the Kullback–Leibler (KL) divergence between the actual distribution of genotype frequencies and the corresponding neutral distribution. First, we show that this population-level information sets an upper bound on the information at the level of genotype and phenotype, limiting how precisely they can be specified by selection. Next, we study how the accumulation and maintenance of information is limited by the cost of selection, measured as the genetic load or the relative fitness variance, both of which we connect to the control-theoretic KL cost of control. The information accumulation rate is upper bounded by the population size times the cost of selection. This bound is very general, and applies across models (Wright–Fisher, Moran, diffusion) and to arbitrary forms of selection, mutation, and recombination. Finally, the cost of maintaining information depends on how it is encoded: Specifying a single allele out of two is expensive, but one bit encoded among many weakly specified loci (as in a polygenic trait) is cheap.

Throughout evolution, selection accumulates information in the genome. It guides evolving populations toward fitter phenotypes, genotypes, and genotype frequencies, which would be highly unlikely to arise by chance. This information—the degree to which selection can control the stochastic process of evolution—has been a long-standing subject of research (1–7), and relates to basic questions in evolutionary biology and genetics.     

Comparing the development of cortex-wide gene expression patterns between two species in a common reference frame

Advances in sequencing techniques have made comparative studies of gene expression a current focus for understanding evolutionary and developmental processes. However, insights into the spatial expression of genes have been limited by a lack of robust methodology. To overcome this obstacle, we developed methods and software tools for quantifying and comparing tissue-wide spatial patterns of gene expression within and between species. Here, we compare cortex-wide expression of RZRβ and Id2 mRNA across early postnatal development in mice and voles. We show that patterns of RZRβ expression in neocortical layer 4 are highly conserved between species but develop rapidly in voles and much more gradually in mice, who show a marked expansion in the relative size of the putative primary visual area across the first postnatal week. Patterns of Id2 expression, by contrast, emerge in a dynamic and layer-specific sequence that is consistent between the two species. We suggest that these differences in the development of neocortical patterning reflect the independent evolution of brains, bodies, and sensory systems in the 35 million years since their last common ancestor.

Almost everything we know about the human brain comes from comparative studies of other animals: from genes involved in cortical development to system-level networks that generate complex behaviors. Comparative studies of living species provide a robust means by which to understand unknown forms, like humans, and even extinct forms like our early mammalian ancestors. Importantly, these types of studies are critical for identifying features of brain organization that are conserved between species and those that may have been derived in different lineages. They also allow us to determine how developmental programs and timing schedules may vary across species, and to better understand how phenotypic diversity can be generated over shorter and longer timescales. Finally, by making valid comparisons across species, we can begin to understand how complexity emerges in different nervous systems, the rules of brain construction, and the constraints imposed on developing and evolving nervous systems.Despite the importance of comparative studies in biology, most comparisons of anatomically reconstructed data are subjective, and most gene sequencing studies neglect the actual spatial patterns of gene expression across a structure, focusing instead on cell-type expression (1–3). Moreover, many current methods for making comparisons fail to capture the three-dimensional nature of the brain, which is composed of asymmetrical structures that can vary markedly in relative shape, size, and location across species and between developmental time-points. Despite the three-dimensional (3D) nature of the brain, most studies collapse data into two dimensions driven largely by the plane of section at which the brain is cut.As such, neurobiologists are faced with two challenges. First, attempting to understand 3D structures by analyzing two-dimensional (2D) images is inherently problematic because the loss of spatial information is unavoidable, especially in curved structures (4). 2D analysis often involves prespecifying regions of interest (ROIs) to quantify the presence of labeled cells or mRNA expression after in-situ hybridization (ISH), narrowing the focus and potentially missing overall differences across a structure, such as the neocortex. A second challenge, which arises when making comparisons between structures in different species and/or at different developmental time-points, or between different experimental conditions, is determining the extent to which 2D spatial patterning might be invariant to basic transformations in the size and shape of the 3D structure. To this end, it is important for comparisons to be made with respect to a common anatomical reference frame.In the current study, we overcame these challenges by developing a set of algorithms for brain slice registration in 3D, and for incorporating ISH data into a common reference frame to enable point-by-point comparisons between species or experimental conditions (see Materials and Methods). These tools, which we refer to collectively as Stalefish, the Spatial Analysis of Fluorescent (and nonfluorescent) In-Situ Hybridization, allowed for the laminar and spatial patterns of expression of genes involved in cortical development to be quantified and compared in two species of age matched rodents across early postnatal development. Our analysis of Id2 and RZRβ cortical expression patterns in mouse and vole brains reveals both a strong layer-specific conservation of the patterning of these genes, as well as area-specific differences in development that shed new light on the ontogeny and phylogeny of neocortical arealization.     

Interlaminar Microstructure and Mechanical Properties of Narrow Gap Laser Welding of 40-mm-Thick Ti-6Al-4V Alloy

Narrow gap laser welding (NGLW) is a common solution for the welding of thick structures. NGLW was carried out on narrow-gap butt joints of 40 mm-thick Ti-6Al-4V alloy plates with a U-shaped groove. The distribution characteristics of the interlaminar microstructure in different height ranges of the joint were investigated, and the evolution behavior and formation mechanism of the interlaminar microstructure of the joint were also revealed. This showed that a large amount of short needle martensite nucleated and grew up near the fusion line and the upper boundary of the remelting zone. The “softening” phenomenon occurred in all welds except the cover layer weld. The microstructure evolution and defect migration, induced by multiple welding thermal cycles in the upper weld forming process, were the main reasons for the “softening” of the lower weld. The tensile strength of each sample changed in the range of 920~990 MPa; the fracture mode of the sample belongs to a transgranular ductile fracture. In addition, compared with the upper part of the joint, the plasticity and toughness of the weld area in the lower part of the joint was improved.     

Apprehending the NAD+–ADPr-Dependent Systems in the Virus World

NAD⁺ and ADP-ribose (ADPr)-containing molecules are at the interface of virus–host conflicts across life encompassing RNA processing, restriction, lysogeny/dormancy and functional hijacking. We objectively defined the central components of the NAD⁺–ADPr networks involved in these conflicts and systematically surveyed 21,191 completely sequenced viral proteomes representative of all publicly available branches of the viral world to reconstruct a comprehensive picture of the viral NAD⁺–ADPr systems. These systems have been widely and repeatedly exploited by positive-strand RNA and DNA viruses, especially those with larger genomes and more intricate life-history strategies. We present evidence that ADP-ribosyltransferases (ARTs), ADPr-targeting Macro, NADAR and Nudix proteins are frequently packaged into virions, particularly in phages with contractile tails (Myoviruses), and deployed during infection to modify host macromolecules and counter NAD⁺-derived signals involved in viral restriction. Genes encoding NAD⁺–ADPr-utilizing domains were repeatedly exchanged between distantly related viruses, hosts and endo-parasites/symbionts, suggesting selection for them across the virus world. Contextual analysis indicates that the bacteriophage versions of ADPr-targeting domains are more likely to counter soluble ADPr derivatives, while the eukaryotic RNA viral versions might prefer macromolecular ADPr adducts. Finally, we also use comparative genomics to predict host systems involved in countering viral ADP ribosylation of host molecules.     

Evolution of Swine Influenza Virus H3N2 in Vaccinated and Nonvaccinated Pigs after Previous Natural H1N1 Infection

Swine influenza viruses (SIV) produce a highly contagious and worldwide distributed disease that can cause important economic losses to the pig industry. Currently, this virus is endemic in farms and, although used limitedly, trivalent vaccine application is the most extended strategy to control SIV. The presence of pre-existing immunity against SIV may modulate the evolutionary dynamic of this virus. To better understand these dynamics, the viral variants generated in vaccinated and nonvaccinated H3N2 challenged pigs after recovery from a natural A(H1N1) pdm09 infection were determined and analyzed. In total, seventeen whole SIV genomes were determined, 6 from vaccinated, and 10 from nonvaccinated animals and their inoculum, by NGS. Herein, 214 de novo substitutions were found along all SIV segments, 44 of them being nonsynonymous ones with an allele frequency greater than 5%. Nonsynonymous substitutions were not found in NP; meanwhile, many of these were allocated in PB2, PB1, and NS1 proteins. Regarding HA and NA proteins, higher nucleotide diversity, proportionally more nonsynonymous substitutions with an allele frequency greater than 5%, and different domain allocations of mutants, were observed in vaccinated animals, indicating different evolutionary dynamics. This study highlights the rapid adaptability of SIV in different environments.