BACKGROUND & AIMS: OBJECTIVE:To determine the validity of intrinsic value and therapeutic utility. DESIGN:The medicines prescribed in a random sample of 50 cases of common cold from each health centre evaluated were classified according to their intrinsic value (ordinal classification using five categories) and therapeutic use (dichotomized classification based on their intrinsic value). Patients with immunodeficiency or underlying pathology (COPD, cardiopathy, diabetes) were excluded. Each of the generic and non-specific indicators were studied to determine sensitivity and specificity for detecting problems in the rational prescription of medicines. SETTING:A representative sample from 8 primary care centres in the Murcia region. MEASUREMENTS AND MAIN RESULTS:The operative replies curve for intrinsic value revealed that this indicator was of little use in evaluating the quality of prescription. CONCLUSIONS:The low validity of the generic prescription indicators suggests that systems to monitor quality based on a specific focus (linking the prescription to the disease treated) should be designed.

背景与目标： 目的：确定内在价值和治疗实用性的有效性。
设计：按照评估的内在价值（按五类常规分类）和治疗用途（根据内在价值二等分类）对来自每个评估中心的50例普通感冒的随机样本中指定的药物进行分类。具有免疫缺陷或基础病理（COPD，心脏病，糖尿病）的患者被排除在外。对每种通用和非特异性指标进行了研究，以确定敏感性和特异性，以检测药物合理处方中的问题。
地点：穆尔西亚地区8个初级保健中心的代表性样本。
测量和主要结果：内在价值的手术回复曲线表明，该指标在评估处方质量方面几乎没有用。
结论：通用处方指标的有效性较低，表明应设计基于特定重点（将处方与治疗的疾病联系起来）的质量监控系统。

BACKGROUND & AIMS: :Diffuse intrinsic pontine gliomas arise almost exclusively in children, and despite advances in treatment, the majority of patients die within 2 years after initial diagnosis. Because of their infiltrative nature and anatomic location in an eloquent area of the brain, most pontine gliomas are treated without a surgical biopsy. The corresponding lack of tissue samples has resulted in a limited understanding of the underlying genetic and molecular biologic abnormalities associated with pontine gliomas, and is a substantial obstacle for the preclinical testing of targeted therapeutic agents for these tumors. We have established a human glioma cell line that originated from surgical biopsy performed on a patient with a pontine glioma. To insure sustainable in vitro propagation, tumor cells were modified with hTERT (human telomerase ribonucleoprotein reverse transcriptase), and with a luciferase reporter to enable non-invasive bioluminescence imaging. The hTERT modified cells are tumorigenic in athymic rodents, and produce brainstem tumors that recapitulate the infiltrative growth of brainstem gliomas in patients.

背景与目标： ：弥漫性桥脑神经胶质瘤几乎仅在儿童中发生，尽管治疗有所进步，但大多数患者在初诊后2年内死亡。由于其浸润性和大脑雄辩区域的解剖位置，大多数桥脑神经胶质瘤无需手术活检即可治疗。组织样品的相应缺乏导致对与桥脑神经胶质瘤相关的潜在遗传和分子生​​物学异常的了解有限，并且是针对这些肿瘤的靶向治疗剂进行临床前测试的主要障碍。我们已经建立了人类脑胶质瘤细胞系，该细胞系源自对桥脑胶质瘤患者进行的手术活检。为了确保可持续的体外繁殖，使用hTERT（人端粒酶核糖核蛋白逆转录酶）和荧光素酶报道分子修饰肿瘤细胞，以实现非侵入性生物发光成像。 hTERT修饰的细胞在无胸腺啮齿动物中具有致瘤性，并产生脑干肿瘤，可概括患者脑干神经胶质瘤的浸润性生长。

BACKGROUND & AIMS: :Proteins are often classified in a binary fashion as either structured or disordered. However this approach has several deficits. Firstly, protein folding is always conditional on the physiochemical environment. A protein which is structured in some circumstances will be disordered in others. Secondly, it hides a fundamental asymmetry in behavior. While all structured proteins can be unfolded through a change in environment, not all disordered proteins have the capacity for folding. Failure to accommodate these complexities confuses the definition of both protein structural domains and intrinsically disordered regions. We illustrate these points with an experimental study of a family of small binding domains, drawn from the RNA polymerase of mumps virus and its closest relatives. Assessed at face value the domains fall on a structural continuum, with folded, partially folded, and near unstructured members. Yet the disorder present in the family is conditional, and these closely related polypeptides can access the same folded state under appropriate conditions. Any heuristic definition of the protein domain emphasizing conformational stability divides this domain family in two, in a way that makes no biological sense. Structural domains would be better defined by their ability to adopt a specific tertiary structure: a structure that may or may not be realized, dependent on the circumstances. This explicitly allows for the conditional nature of protein folding, and more clearly demarcates structural domains from intrinsically disordered regions that may function without folding.

背景与目标： ：Protein通常以二进制方式分类为结构化或无序。但是，这种方法存在一些缺陷。首先，蛋白质折叠总是以物理化学环境为条件。在某些情况下结构化的蛋白质在其他情况下会变得无序。其次，它掩盖了行为的基本不对称性。虽然所有结构化蛋白质都可以通过环境变化来展开，但并非所有无序蛋白质都具有折叠能力。无法适应这些复杂性会混淆蛋白质结构域和内在无序区域的定义。我们通过对腮腺炎病毒及其近亲的RNA聚合酶绘制的一系列小结合域的实验研究说明了这些观点。按面值评估，这些域位于结构连续体上，具有折叠，部分折叠和接近非结构化的成员。然而，该家族中存在的病症是有条件的，并且这些紧密相关的多肽可以在适当的条件下进入相同的折叠状态。强调构象稳定性的蛋白质结构域的任何启发式定义都将该结构域家族分为两部分，这是没有生物学意义的。结构域可以采用特定的三级结构的能力来更好地定义：根据情况，该结构可能会实现，也可能不会实现。这明确地允许蛋白质折叠的条件性质，并且更清楚地从可能无折叠而起作用的内在无序的区域划定结构域。

BACKGROUND & AIMS: :Cooperation declines in repeated public good games because individuals behave as conditional cooperators. This is because individuals imitate the social behaviour of successful individuals when their payoff information is available. However, in human societies, individuals cooperate in many situations involving social dilemmas. We hypothesize that humans are sensitive to both success (payoffs) and how that success was obtained, by cheating (not socially sanctioned) or good behaviour (socially sanctioned and adds to prestige or reputation), when information is available about payoffs and prestige. We propose and model a repeated public good game with heterogeneous conditional cooperators where an agent's donation in a public goods game depends on comparing the number of donations in the population in the previous round and with the agent's arbitrary chosen conditional cooperative criterion. Such individuals imitate the social behaviour of role models based on their payoffs and prestige. The dependence is modelled by two population-level parameters: affinity towards payoff and affinity towards prestige. These affinities influence the degree to which agents value the payoff and prestige of role models. Agents update their conditional strategies by considering both parameters. The simulations in this study show that high levels of cooperation are established in a population consisting of heterogeneous conditional cooperators for a certain range of affinity parameters in repeated public good games. The results show that social value (prestige) is important in establishing cooperation.

背景与目标： ：在反复的公益活动中，合作减少了，因为个人表现为有条件的合作者。这是因为，当可获得他们的回报信息时，个人会模仿成功人士的社交行为。但是，在人类社会中，个人在许多涉及社会困境的情况下进行合作。我们假设人们对成功（收益）以及成功与否（无论是通过作弊（未受到社会制裁）还是良好行为（受到社会认可并会增加声望或声誉））都非常敏感，只要他们可以获得有关收益和声望的信息。我们提出了一个具有异类条件合作者的重复公共物品博弈并对其进行建模，其中代理在公共物品博弈中的捐赠取决于将前一轮中人口捐赠的数量与代理的任意选择的条件合作标准进行比较。这些人根据他们的回报和声望来模仿榜样的社会行为。依赖关系由两个总体级别的参数建模：对收益的亲和力和对声望的亲和力。这些亲和力会影响代理人重视榜样的回报和声望的程度。代理通过考虑两个参数来更新其条件策略。本研究中的模拟显示，在重复的公益游戏中，对于特定范围的亲和力参数，在由异质条件合作者组成的群体中建立了高水平的合作。结果表明，社会价值（信誉）在建立合作中很重要。

BACKGROUND & AIMS: OBJECTIVES:The aim of this study was to derive longitudinal reference values of fetal growth (estimated fetal weight [EFW] and abdominal circumference [AC]) during the third trimester and to develop coefficients for conditional growth assessment. PATIENTS AND METHODS:A prospective cohort study was conducted involving consecutive singleton pregnancies in a low-risk population for a routine third-trimester scan at 30+0-34+6 weeks and follow-up at 37+0-38+6 weeks for an additional ultrasound. Statistical analysis was based on multilevel modeling using MLwiN software. Unconditional centiles were calculated from z-values at each gestational age, and conditional centiles were calculated from z-values at a given measurement (30-34 weeks) and the expected measurement (37-38 weeks). RESULTS:At 30-34 weeks, 8 and 9.3% of the fetuses had an unconditional EFW below the 10th and above the 90th centile, respectively. At 37-38 weeks, these figures were 10.3 and 9.3%, respectively. Regarding the unconditional AC, at the first scan, 8.9 and 9.6% had values below the 10th and above the 90th centile, while at the second scan 10.5 and 10.5% had values below the 10th and above the 90th centile, respectively. The proportion with a conditional EFW below the 10th and above the 90th centile was 10.2 and 9.4% at the second scan, respectively. For conditional AC, these figures were 10.7 and 10.3%, respectively. CONCLUSION:We have produced reference centiles for EFW and AC growth during the third trimester as a useful tool for quantifying growth.

背景与目标： 目的：本研究的目的是得出妊娠中期胎儿生长的纵向参考值（估计的胎儿体重[EFW]和腹围[AC]），并建立条件生长评估的系数。
患者和方法：进行了一项前瞻性队列研究，涉及低危人群中连续单胎妊娠，在30 0-34 6周进行常规的孕中期扫描，并在37 0-38 6周进行随访，以进行额外的超声检查。统计分析基于使用MLwiN软件的多级建模。根据每个胎龄的z值计算无条件的百分位数，根据给定的测量值（30-34周）和预期的测量值（37-38周）的z值计算条件的百分位。
结果：在30-34周时，分别有8％和9.3％的胎儿在第10个百分位以下和第90个百分位以上有无条件的EFW。在第37-38周时，这些数字分别为10.3和9.3％。对于无条件AC，在第一次扫描时，8.9和9.6％的值低于第10个百分位，在第90个百分点之上，而在第二次扫描中，10.5和10.5％的值分别在第10个百分位数和第90个百分率以上。在第二次扫描时，条件EFW低于第10个百分位和高于第90个百分位的比例分别为10.2％和9.4％。对于有条件的AC，这些数字分别为10.7％和10.3％。
结论：我们为妊娠晚期的EFW和AC增长绘制了参考百分位，作为量化增长的有用工具。

BACKGROUND & AIMS: :Age-related changes in primary lymphoid organs are well described. Less is known about age-related changes affecting peripheral lymphoid organs, although defects in old peripheral lymph nodes (pLNs) were recently described in both steady state and during viral infection. To address whether such pLN defects were intrinsic to old T cells or extrinsic (due to aging microenvironment), we employed heterochronic parabiosis. We found no age-related intrinsic or extrinsic barriers to T cell circulation and seeding of pLN, spleen, and bone marrow. However, heterochronic parabiosis failed to improve cellularity of old pLN, suggesting an environment-based limit on pLN cellularity. Furthermore, upon parabiosis, pLN of the adult partner exhibited reduced, old-like stromal and T cell cellularity, which was restored following separation of parabionts. Decay measurement of adult and old T cell subsets following separation of heterochronic parabionts delineated both T cell-intrinsic and environmental changes in T cell maintenance. Moreover, parabiotic separation revealed differences between CD4 and CD8 T cell subset maintenance with aging, the basis of which will require further investigation. Reasons for this asymmetric and subset-specific pattern of differential maintenance are discussed in light of possible age-related changes in lymph nodes as the key sites for peripheral T cell maintenance.

背景与目标： ：与淋巴管原发性器官的年龄相关的变化已被很好地描述。关于年龄相关变化影响外周淋巴器官的知之甚少，尽管最近已在稳定状态和病毒感染期间描述了旧的外周淋巴结（pLNs）的缺陷。为了解决此类pLN缺陷是旧T细胞固有的还是外部T细胞固有的（由于老化的微环境），我们采用了异时共生。我们没有发现年龄相关的内在或外在障碍对T细胞循环和pLN，脾脏和骨髓的播种。但是，异时生物共生不能改善旧pLN的细胞性，这表明基于环境的pLN细胞性受到限制。此外，在发生共生关系时，成年伴侣的pLN表现出减少的，老样的基质和T细胞细胞性，这在分离代谢物后得以恢复。异时生物体分离后，成年和老T细胞亚群的衰变测量描述了T细胞内在性和T细胞维持环境的变化。此外，共生生物分离揭示了CD4和CD8 T细胞亚群维持与衰老之间的差异，其基础将需要进一步研究。鉴于可能与年龄相关的淋巴结变化是外周T细胞维持的关键部位，讨论了这种差异维护的非对称和子集特定模式的原因。

BACKGROUND & AIMS: OBJECTIVE:To investigate the functions of Fibroblast Growth Factor Receptor-2 (FGFR2) at different stages of cell differentiation. The engineered murine embryonic stem (ES) cells with conditional knockout of FGFR2 were developed depending on Cre-loxP. METHODS:Cre-loxP system was used in a conditional targeting vector. The competent AM-1 bacteria, which expressed Cre-recombinase, was used to confirm the Cre-mediated deletion of the floxed exons 7 and 8 of FGFR2. The targeting vector was electroporated into the ES cells, and the transfected ES cells were screened with G418 and Ganciclovir. Finally, the ES clones with correct targeting events were identified by Southern Blot and PCR. RESULTS:The targeting vector with conditional knockout of murine FGFR2 was successfully constructed and confirmed by PCR and digestion analysis in bacteria. 86 ES clones were collected by selective culture with G418 and Ganciclovir. Four of the 86 ES clones were found containing the targeting gene sequence in genomic DNA proved by Southern Blot with a 5'-end flank probe. Two of the four ES clones had the correct targeting events that included the insertion of the targeting gene sequence in genomic DNA and were checked by Southern Blot with a 3'-end flanking probe. Finally, the insertion of loxP (loxP3) between exons 8 and 9 in genomic DNA was identified in one of the two ES clones by Southern Blot and PCR. CONCLUSION:FGFR2 conditional knockout depending on Cre-loxP can be successfully used in ES cells.

背景与目标： 目的：探讨成纤维细胞生长因子受体2（FGFR2）在不同分化阶段的功能。根据Cre-loxP，开发了具有条件敲除FGFR2的工程鼠胚胎干（ES）细胞。
方法：将Cre-loxP系统用于条件靶向载体。表达Cre重组酶的感受态AM-1细菌用于确认Cre介导的FGFR2外显子7和8的缺失。将靶向载体电穿孔到ES细胞中，并用G418和更昔洛韦筛选转染的ES细胞。最后，通过Southern印迹和PCR鉴定具有正确靶向事件的ES克隆。
结果：成功构建了有条件敲除鼠FGFR2的靶向载体，并通过PCR和酶切分析鉴定。通过用G418和更昔洛韦选择性培养收集了86个ES克隆。发现86个ES克隆中有四个在基因组DNA中包含靶向基因序列，该DNA由Southern Blot用5'端侧翼探针证实。四个ES克隆中的两个具有正确的靶向事件，包括在基因组DNA中插入了靶向基因序列，并使用3'端侧翼探针通过Southern Blot检测。最后，通过Southern印迹和PCR在两个ES克隆之一中鉴定出loxP（loxP3）在基因组DNA的外显子8和9之间的插入。
结论：依赖于Cre-loxP的FGFR2条件敲除可成功用于ES细胞。

BACKGROUND & AIMS: :Mouse mutants heterozygously deficient for the myelin protein P0 (P0+/-) resemble certain forms of human hereditary neuropathies. Endoneurial macrophages of intrinsic origin are intimately involved in the pathogenesis of the demyelinating neuropathy in these mutants. We have previously shown that deficiency for macrophage colony stimulating factor (M-CSF) prevents an increase of the number of endoneurial macrophages and alleviates the mutants' demyelinating phenotype. The aim of this study was to investigate which population of endoneurial macrophages - long-term resident macrophages or recently infiltrated macrophages - is affected by M-CSF deficiency. For this purpose, we generated bone marrow chimeric mice by transplanting GFP+ bone marrow into P0 mutants (P0+/-) and P0 mutants that lack M-CSF (P0+/- mcsf-op). This enabled us to discriminate recently infiltrated short-term resident GFP+ macrophages from long-term resident GFP- macrophages. Three months after bone marrow transplantation, P0+/- mice expressing M-CSF showed a substantial upregulation and activation of both GFP- and GFP+ macrophages in femoral nerves when compared to P0+/+ mice. In contrast, in P0+/- mcsf-op mutants, both GFP- and GFP+ macrophages did not substantially increase. Only small numbers of GFP+ but no GFP- macrophages were activated and phagocytosed myelin in chimeric P0+/- mcsf-op mutants, possibly reflecting recent activation outside the endoneurium before entering the nerve. Our findings demonstrate that M-CSF is crucial for the activation, in situ increase and myelin phagocytosis of both long-term and short-term resident endoneurial macrophages in P0+/- myelin mutants. M-CSF is, therefore, considered as a target candidate for therapeutic strategies to treat human demyelinating neuropathies.

背景与目标： 杂合子缺乏髓磷脂蛋白P0（P0 /-）的小鼠突变体类似于人类遗传性神经病的某些形式。在这些突变体中，固有来源的神经内膜巨噬细胞与脱髓鞘性神经病的发病机理密切相关。先前我们已经表明，巨噬细胞集落刺激因子（M-CSF）的缺乏会阻止神经内膜巨噬细胞数量的增加，并减轻突变体的脱髓鞘表型。这项研究的目的是调查哪些神经内膜巨噬细胞群-长期驻留巨噬细胞或最近渗透的巨噬细胞-受M-CSF缺乏症的影响。为此，我们通过将GFP骨髓移植到P0突变体（P0 /-）和缺少M-CSF的P0突变体（P0 /-mcsf-op）中，生成了骨髓嵌合小鼠。这使我们能够将近期浸润的短期驻留GFP巨噬细胞与长期驻留GFP-巨噬细胞区分开来。与P0 /小鼠相比，骨髓移植后三个月，表达M-CSF的P0 /-小鼠显示股神经中GFP-和GFP巨噬细胞均显着上调和激活。相反，在P 0 --mcsf-op突变体中，GFP-和GFP巨噬细胞均没有显着增加。在嵌合的P0-mcsf-op突变体中，只有少量的GFP激活，但没有GFP-巨噬细胞被吞噬的髓磷脂，可能反映了进入神经之前在神经内膜外的最近激活。我们的发现表明，M-CSF对于P0 /-髓磷脂突变体中长期和短期驻留的神经内膜巨噬细胞的活化，原位增加和髓磷脂吞噬至关重要。因此，M-CSF被认为是治疗人脱髓鞘性神经病的治疗策略的靶标候选物。

BACKGROUND & AIMS: :Clustered regularly interspaced short palindromic repeats (CRISPRs) and CRISPR-associated (Cas) proteins provide adaptive immunity to prokaryotes against invading phages and plasmids. As a countermeasure, phages have evolved anti-CRISPR (Acr) proteins that neutralize the CRISPR immunity. AcrIIA5, isolated from a virulent phage of Streptococcus thermophilus, strongly inhibits diverse Cas9 homologs, but the molecular mechanism underlying the Cas9 inhibition remains unknown. Here, we report the solution structure of AcrIIA5, which features a novel α/β fold connected to an N-terminal intrinsically disordered region (IDR). Remarkably, truncation of the N-terminal IDR abrogates the inhibitory activity against Cas9, revealing that the IDR is essential for Cas9 inhibition by AcrIIA5. Progressive truncations and mutations of the IDR illustrate that the disordered region not only modulates the association between AcrIIA5 and Cas9-sgRNA, but also alters the catalytic efficiency of the inhibitory complex. The length of IDR is critical for the Cas9-sgRNA recognition by AcrIIA5, whereas the charge content of IDR dictates the inhibitory activity. Conformational plasticity of IDR may be linked to the broad-spectrum inhibition of Cas9 homologs by AcrIIA5. Identification of the IDR as the main determinant for Cas9 inhibition expands the inventory of phage anti-CRISPR mechanisms.

背景与目标： ：聚簇的规则间隔的短回文重复序列（CRISPR）和CRISPR相关的（Cas）蛋白为原核生物提供了针对入侵的噬菌体和质粒的适应性免疫力。作为对策，噬菌体已经进化出可中和CRISPR免疫力的抗CRISPR（Acr）蛋白。从嗜热链球菌的强毒噬菌体中分离出的AcrIIA5可以强烈抑制各种Cas9同源物，但仍不知道抑制Cas9的分子机制。在这里，我们报告AcrIIA5的解决方案结构，其特征是连接到N端固有无序区（IDR）的新型α/β折叠。值得注意的是，N末端IDR的截短取消了对Cas9的抑制活性，这表明IDR对于AcrIIA5抑制Cas9是必不可少的。 IDR的逐步截短和突变表明，无序区域不仅调节AcrIIA5与Cas9-sgRNA之间的结合，而且还改变了抑制性复合物的催化效率。 IDR的长度对于AcrIIA5识别Cas9-sgRNA至关重要，而IDR的电荷含量决定了其抑制活性。 IDR的构象可塑性可能与AcrIIA5对Cas9同源物的广谱抑制有关。 IDR作为Cas9抑制的主要决定因素的鉴定扩大了噬菌体抗CRISPR机制的库存。

BACKGROUND & AIMS: BACKGROUND:Breast (BCa) and prostate (PCa) cancers are hormone receptor (HR)-driven cancers. Thus, BCa and PCa patients are given therapies that reduce hormone levels or directly block HR activity; but most patients eventually develop treatment resistance. We have previously reported that interleukin-1 (IL-1) inflammatory cytokine downregulates ERα and AR mRNA in HR-positive (HR+) BCa and PCa cell lines, yet the cells can remain viable. Additionally, we identified pro-survival proteins and processes upregulated by IL-1 in HR+ BCa and PCa cells, that are basally high in HR- BCa and PCa cells. Therefore, we hypothesize that IL-1 confers a conserved gene expression pattern in HR+ BCa and PCa cells that mimics conserved basal gene expression patterns in HR- BCa and PCa cells to promote HR-independent survival and tumorigenicity. METHODS:We performed RNA sequencing (RNA-seq) for HR+ BCa and PCa cell lines exposed to IL-1 and for untreated HR- BCa and PCa cell lines. We confirmed expression patterns of select genes by RT-qPCR and used siRNA and/or drug inhibition to silence select genes in the BCa and PCa cell lines. Finally, we performed Ingenuity Pathway Analysis (IPA) and used the gene ontology web-based tool, GOrilla, to identify signaling pathways encoded by our RNA-seq data set. RESULTS:We identified 350 genes in common between BCa and PCa cells that are induced or repressed by IL-1 in HR+ cells that are, respectively, basally high or low in HR- cells. Among these genes, we identified Sequestome-1 (SQSTM1/p62) and SRY (Sex-Determining Region Y)-Box 9 (SOX9) to be essential for survival of HR- BCa and PCa cell lines. Analysis of publicly available data indicates that p62 and SOX9 expression are elevated in HR-independent BCa and PCa sublines generated in vitro, suggesting that p62 and SOX9 have a role in acquired hormone receptor independence and treatment resistance. We also assessed HR- cell line viability in response to the p62-targeting drug, verteporfin, and found that verteporfin is cytotoxic for HR- cell lines. CONCLUSIONS:Our 350 gene set can be used to identify novel therapeutic targets and/or biomarkers conserved among acquired (e.g. due to inflammation) or intrinsic HR-independent BCa and PCa.

背景与目标： 背景：乳腺癌（BCa）和前列腺癌（PCa）是激素受体（HR）驱动的癌症。因此，为BCa和PCa患者提供降低激素水平或直接阻断HR活动的疗法；但是大多数患者最终都会产生治疗抵抗力。我们以前曾报道过白介素1（IL-1）炎性细胞因子下调HR阳性（HR）BCa和PCa细胞系中的ERα和AR mRNA，但这些细胞仍可以保持活力。另外，我们在HR Bca和PCa细胞中基本较高的HR BCa和PCa细胞中鉴定了促存活蛋白和IL-1上调的过程。因此，我们假设IL-1赋予HR BCa和PCa细胞保守的基因表达模式，该模式模仿HR-BCa和PCa细胞中保守的基础基因表达模式，从而促进HR非依赖性生存和致瘤性。
方法：我们对暴露于IL-1的HR BCa和PCa细胞系以及未经处理的HR- BCa和PCa细胞系进行了RNA测序（RNA-seq）。我们通过RT-qPCR证实了选择基因的表达模式，并使用siRNA和/或药物抑制使BCa和PCa细胞系中的选择基因沉默。最后，我们进行了创新途径分析（IPA），并使用了基于基因本体网络的工具GOrilla来识别由RNA-seq数据集编码的信号传导途径。
结果：我们确定了BCa和PCa细胞之间共有350个基因，这些基因分别由HR细胞中基本高或低的HR细胞中的IL-1诱导或抑制。在这些基因中，我们确定了Sequestome-1（SQSTM1 / p62）和SRY（性决定区Y）-Box 9（SOX9）对HR-BCa和PCa细胞系的存活至关重要。公开数据的分析表明，在体外产生的HR非依赖性BCa和PCa亚系中p62和SOX9表达升高，表明p62和SOX9在获得性激素受体独立性和治疗耐药性中起作用。我们还评估了针对p62靶向药物verteporfin的HR细胞系活力，并发现verteporfin对HR细胞系具有细胞毒性。
结论：我们的350个基因集可用于鉴定在获得性（例如由于炎症）或内源性HR无关的BCa和PCa中保守的新治疗靶标和/或生物标志物。

BACKGROUND & AIMS: :The optimization of aqueous solubility is an important step along the route to bringing a new therapeutic to market. We describe the development of an empirical computational model to rank the pH-dependent aqueous solubility of drug candidates. The model consists of three core components to describe aqueous solubility. The first is a multivariate QSAR model for the prediction of the intrinsic solubility of the neutral solute. The second facet of the approach is the consideration of ionization using a predicted pKa and the Henderson-Hasselbalch equation. The third aspect of the model is a novel method for assessing the effects of crystal packing on solubility through a series of short molecular dynamics simulations of an actual or hypothetical small molecule crystal structure at escalating temperatures. The model also includes a Monte Carlo error function that considers the variability of each of the underlying components of the model to estimate the 90% confidence interval of estimation.

背景与目标： ：水溶性的优化是将新疗法推向市场的重要一步。我们描述了经验计算模型的发展，以对候选药物的pH依赖性水溶解度进行排名。该模型由三个核心成分组成，用于描述水溶性。第一个是用于预测中性溶质固有溶解度的多元QSAR模型。该方法的第二个方面是考虑使用预测的pKa和Henderson-Hasselbalch方程进行电离。该模型的第三方面是一种新颖的方法，可通过在升高的温度下对实际或假设的小分子晶体结构进行一系列短分子动力学模拟来评估晶体堆积对溶解度的影响。该模型还包括蒙特卡洛误差函数，该函数考虑模型的每个基本组件的可变性以估计90％的估计置信区间。

BACKGROUND & AIMS: OBJECTIVES:Rifampicin, a potent first-line TB drug of the rifamycin group, shows only little activity against the emerging pathogen Mycobacterium abscessus. Reportedly, bacterial resistance to rifampicin is associated with polymorphisms in the target gene rpoB or the presence of enzymes that modify and thereby inactivate rifampicin. The aim of this study was to investigate the role of the MAB_0591 (arrMab)-encoded rifampicin ADP-ribosyltransferase (Arr_Mab) in innate high-level rifampicin resistance in M. abscessus. METHODS:Recombinant Escherichia coli and Mycobacterium tuberculosis strains expressing MAB_0591 were generated, as was an M. abscessus deletion mutant deficient for MAB_0591. MIC assays were used to study susceptibility to rifampicin and C25 carbamate-modified rifamycin derivatives. RESULTS:Heterologous expression of MAB_0591 conferred rifampicin resistance to E. coli and M. tuberculosis Rifamycin MIC values were consistently lower for the M. abscessus ΔarrMab mutant as compared with the M. abscessus ATCC 19977 parental type strain. The rifamycin WT phenotype was restored after complementation of the M. abscessus ΔarrMab mutant with arrMab Further MIC data demonstrated that a C25 modification increases rifamycin activity in WT M. abscessus However, MIC studies in the M. abscessus ΔarrMab mutant suggest that C25 modified rifamycins are still subject to modification by Arr_Mab CONCLUSIONS: Our findings identify Arr_Mab as the major innate rifamycin resistance determinant of M. abscessus. Our data also indicate that Arr_Mab-mediated rifamycin resistance in M. abscessus can only in part be overcome by C25 carbamate modification.

背景与目标： 目的：利福平是利福霉素组的一种有效的一线结核病药物，对新兴的病原性脓杆菌分枝杆菌几乎没有活性。据报道，细菌对利福平的抗性与靶基因rpoB中的多态性或修饰或使利福平失活的酶的存在有关。这项研究的目的是调查MAB_0591（arrMab）编码的利福平ADP-核糖基转移酶（Arr_Mab）在脓毒症先天性高水平利福平耐药中的作用。
方法：产生表达MAB_0591的重组大肠杆菌和结核分枝杆菌菌株，以及缺失MAB_0591的脓肿分支杆菌缺失突变体。 MIC分析用于研究对利福平和C25氨基甲酸酯修饰的利福霉素衍生物的敏感性。
结果：MAB_0591的异源表达赋予了利福平对大肠埃希菌和结核分枝杆菌的利福霉素耐药性。MIC值与脓毒症ΔarrMab突变体相比，在脓毒症中的ATCC 19977亲本型菌株均较低。脓肿分枝杆菌ΔarrMab突变体与arrMab互补后，利福霉素WT表型得以恢复。进一步的MIC数据表明，C25修饰增加了WT脓肿中利福霉素的活性。结论：我们的发现确定Arr_Mab是脓肿分支杆菌的主要先天利福霉素耐药性决定因素。我们的数据还表明，脓肿支原体中Arr_Mab介导的利福霉素耐药性只能通过C25氨基甲酸酯修饰来部分克服。

BACKGROUND & AIMS: :G protein-coupled receptors (GPCRs) coupled to activation of Gs, such as the PTH1 receptor (PTH1R), have long been known to regulate skeletal function and homeostasis. However, the role of GPCRs coupled to other G proteins such as Gi is not well established. We used the tet-off system to regulate the expression of an activated Gi-coupled GPCR (Ro1) in osteoblasts in vivo. Skeletal phenotypes were assessed in mice expressing Ro1 from conception, from late stages of embryogenesis, and after weaning. Long bones were assessed histologically and by microcomputed tomography. Expression of Ro1 from conception resulted in neonatal lethality that was associated with reduced bone mineralization. Expression of Ro1 starting at late embryogenesis resulted in a severe trabecular bone deficit at 12 wk of age (>51% reduction in trabecular bone volume fraction in the proximal tibia compared with sex-matched control littermates; n = 11; P < 0.01). Ro1 expression for 8 wk beginning at 4 wk of age resulted in a more than 20% reduction in trabecular bone volume fraction compared with sex-matched control littermates (n = 16; P < 0.01). Bone histomorphometry revealed that Ro1 expression is associated with reduced rates of bone formation and mineral apposition without a significant change in osteoblast or osteoclast surface. Our results indicate that signaling by a Gi-coupled GPCR in osteoblasts leads to osteopenia resulting from a reduction in trabecular bone formation. The severity of the phenotype is related to the timing and duration of Ro1 expression during growth and development. The skeletal phenotype in Ro1 mice bears some similarity to that produced by knockout of Gs-alpha expression in osteoblasts and thus may be due at least in part to Gi-mediated inhibition of adenylyl cyclase.

背景与目标： 长期以来，已知与Gs激活偶联的：G蛋白偶联受体（GPCR）可调节骨骼功能和体内平衡。但是，与其他G蛋白（例如Gi）偶联的GPCR的作用尚不明确。我们使用tet-off系统来调节体内成骨细胞中激活的Gi偶联GPCR（Ro1）的表达。从受孕，胚胎发生的后期以及断奶后评估表达Ro1的小鼠的骨骼表型。通过组织学和微计算机断层扫描对长骨进行评估。从受孕中表达Ro1会导致新生儿致死性，这与减少的骨矿化有关。 Ro1的表达始于胚胎晚期，导致在12周龄时出现严重的小梁骨缺损（胫骨近端的小梁骨体积分数与性别匹配的对照同窝仔相比减少了51％以上； n = 11； P <0.01）。与性别匹配的对照同窝仔相比，从4周龄开始的8周Ro1表达导致小梁骨体积分数减少了20％以上（n = 16； P <0.01）。骨组织形态计量学表明，Ro1表达与成骨率和矿物质沉积率降低相关，而成骨细胞或破骨细胞表面无明显变化。我们的结果表明，成骨细胞中通过Gi偶联GPCR进行的信号传导导致骨小梁减少，其原因是小梁骨形成减少。表型的严重性与Ro1表达在生长发育过程中的时机和持续时间有关。 Ro1小鼠的骨骼表型与成骨细胞中Gs-α表达的敲除产生的相似，因此可能至少部分是由于Gi介导的对腺苷酸环化酶的抑制。

BACKGROUND & AIMS: :Modifications of RNA, collectively termed as the epitranscriptome, are widespread, evolutionarily conserved and contribute to gene regulation and protein diversity in healthy and disease states. There are >160 RNA modifications described, greatly exceeding the number of modifications to DNA. Of these, adenosine-to-inosine (A-to-I) RNA editing is one of the most common. There are tens of thousands of A-to-I editing sites in mouse, and millions in humans. Upon translation or sequencing an inosine base is decoded as guanosine, leading to A-to-G mismatches between the RNA and DNA. Inosine has different base pairing properties to adenosine and as a result editing not only alters the RNA code but can also change the RNA structure. In mammals A-to-I editing is performed by ADAR1 and ADAR2. A feature of murine loss of function ADAR1 alleles is cell death and a failure to survive embryogenesis. Adar1-/- and editing deficient (Adar1E861A/E861A) mice die between E11.75-13.5 of failed hematopoiesis. Strikingly this phenotype is rescued by the deletion of the cytosolic dsRNA sensor MDA5 or its downstream adaptor MAVS, a mechanism conserved in human and mouse. Current literature indicates that the loss of ADAR1 leads to cell death via apoptosis, yet this has not been genetically established. We report that blockade of the intrinsic (mitochondrial) apoptosis pathway, through the loss of both BAK and BAX, does not rescue or modify the cellular phenotype of the fetal liver or extend the lifespan of ADAR1 editing deficient embryos. We had anticipated that the loss of BAK and BAX would rescue, or at least significantly extend, the gestational viability of Adar1E861A/E861A embryos. However, the triple mutant Adar1E861A/E861A Bak-/- Bax-/- embryos that were recovered at E13.5 were indistinguishable from the Adar1E861A/E861A embryos with BAK and BAX. The results indicate that cell death processes not requiring the intrinsic apoptosis pathway are triggered by MDA5 following the loss of ADAR1.

背景与目标： ：RNA的修饰，统称为表观转录组，是广泛存在的，在进化上保守的，并且有助于健康和疾病状态下的基因调节和蛋白质多样性。描述了超过160种RNA修饰，大大超过了DNA修饰的数量。其中，腺苷到肌苷（A到I）的RNA编辑是最常见的一种。鼠标中有成千上万个A-to-I编辑站点，而人类中则有数百万个。翻译或测序后，肌苷碱基被解码为鸟苷，导致RNA和DNA之间的A到G错配。肌苷与腺苷具有不同的碱基配对特性，因此编辑不仅会改变RNA编码，而且还会改变RNA结构。在哺乳动物中，ADAR1和ADAR2进行A对I编辑。小鼠功能ADAR1等位基因丧失的特征是细胞死亡和无法存活的胚胎发生。 Adar1-/-和编辑缺陷型（Adar1E861A / E861A）小鼠在造血功能衰竭的E11.75-13.5之间死亡。令人惊讶的是，该表型通过胞质dsRNA传感器MDA5或其下游衔接子MAVS的缺失而得以挽救，这种机制在人和小鼠中都得到了保留。当前文献表明，ADAR1的丢失通过细胞凋亡导致细胞死亡，但是这尚未在遗传学上得到证实。我们报告说，内在的（线粒体）凋亡途径的阻断通过BAK和BAX的丢失，不会挽救或改变胎儿肝脏的细胞表型，也不会延长ADAR1编辑缺陷胚胎的寿命。我们已经预料到，BAK和BAX的丧失将挽救或至少显着延长Adar1E861A / E861A胚胎的妊娠能力。但是，在E13.5恢复的三重突变型Adar1E861A / E861A Bak-/-Bax-/-胚胎与带有BAK和BAX的Adar1E861A / E861A胚胎没有区别。结果表明，不需要内在凋亡途径的细胞死亡过程是由ADAR1丢失后的MDA5触发的。

BACKGROUND & AIMS: :Gene expression noise refers to the variation of the expression level of a gene among isogenic cells in the same environment, and has two sources: extrinsic noise arising from the disparity of the cell state and intrinsic noise arising from the stochastic process of gene expression in the same cell state. Due to the low throughput of the existing method for measuring the two noise components, the architectures of intrinsic and extrinsic expression noises remain elusive. Using allele-specific single-cell RNA sequencing, we here estimate the two noise components of 3975 genes in mouse fibroblast cells. Our analyses verify predicted influences of several factors such as the TATA-box and microRNA targeting on intrinsic or extrinsic noises and reveal gene function-associated noise trends implicating the action of natural selection. These findings unravel differential regulations, optimizations, and biological consequences of intrinsic and extrinsic noises and can aid the construction of desired synthetic circuits.

背景与目标： ：基因表达噪声是指同一环境中同基因细胞之间基因表达水平的变化，有两种来源：细胞状态差异引起的外在噪声和基因表达随机过程引起的内在噪声。相同的细胞状态。由于用于测量两个噪声分量的现有方法的吞吐量较低，因此固有和非固有表达噪声的体系结构仍然难以捉摸。使用等位基因特异性单细胞RNA测序，我们在这里估计了小鼠成纤维细胞中3975个基因的两个噪声成分。我们的分析验证了诸如TATA-box和microRNA等针对内在或外在噪声的因素的预测影响，并揭示了与基因功能相关的噪声趋势，暗示了自然选择的作用。这些发现揭示了内在和外在噪声的差异调节，优化和生物学后果，并且可以帮助构建所需的合成电路。