BACKGROUND & AIMS: :Human embryonic and mesenchymal stem cell therapies may offer significant benefit to a large number of patients. Recently, however, human embryonic stem cell lines cultured on mouse feeder cells were reported to be contaminated by the xeno-carbohydrate N-glycolylneuraminic acid (Neu5Gc) and considered potentially unfit for human therapy. To determine the extent of the problem of Neu5Gc contamination for the development of stem cell therapies, we investigated whether it also occurs in cells cultured on human feeder cells and in mesenchymal stem cells, what are the sources of contamination, and whether the contamination is reversible. We found that N-glycolylneuraminic acid was present in embryonic stem cells cultured on human feeder cells, correlating with the presence of Neu5Gc in components of the commercial serum replacement culture medium. Similar contamination occurred in mesenchymal stem cells cultured in the presence of fetal bovine serum. The results suggest that the Neu5Gc is present in both glycoprotein and lipid-linked glycans, as detected by mass spectrometric analysis and monoclonal antibody staining, respectively. Significantly, the contamination was largely reversible in the progeny of both cell types, suggesting that decontaminated cells may be derived from existing stem cell lines. Although major complications have not been reported in the clinical trials with mesenchymal stem cells exposed to fetal bovine serum, the immunogenic contamination may potentially be reflected in the viability and efficacy of the transplanted cells and thus bias the published results. Definition of safe culture conditions for stem cells is essential for future development of cellular therapies.

背景与目标： : 人类胚胎和间充质干细胞疗法可能为大量患者提供显着益处。然而，最近，据报道，在小鼠饲养细胞上培养的人类胚胎干细胞系被异种碳水化合物N-糖基神经氨酸 (Neu5Gc) 污染，并被认为可能不适合人类治疗。为了确定Neu5Gc污染在干细胞疗法发展中的问题程度，我们调查了它是否也发生在人类饲养细胞和间充质干细胞上培养的细胞中，什么是污染源，以及污染是否可逆。我们发现N-甘氨酰神经氨酸存在于在人饲养细胞上培养的胚胎干细胞中，这与商业血清替代培养基成分中Neu5Gc的存在有关。在胎牛血清存在下培养的间充质干细胞中也发生了类似的污染。结果表明，分别通过质谱分析和单克隆抗体染色检测到，Neu5Gc存在于糖蛋白和脂质连接的聚糖中。值得注意的是，在两种细胞类型的后代中，污染在很大程度上是可逆的，这表明去污的细胞可能来自现有的干细胞系。尽管在暴露于胎牛血清的间充质干细胞的临床试验中尚未报道主要并发症，但免疫原性污染可能反映在移植细胞的活力和功效中，从而使已发表的结果产生偏差。干细胞安全培养条件的定义对于细胞疗法的未来发展至关重要。

BACKGROUND & AIMS: :The aim of this study was to assess the clinical value of serum oestradiol concentration 8 days after embryo transfer (D8E2) and beta-human chorionic gonadotrophin (HCG-beta) concentration 12 days after embryo transfer (D12HCG-beta) in the prediction of pregnancy and the outcome of pregnancy following assisted reproduction, taking into account the day of transfer, which was either day 3 (D3) or day 5 (D5). The objective was to improve patient counselling by giving quantitative and reliable predictive information instead of non-specific uncertainties. A total of 2035 embryo transfer cycles performed between January 2003 and June 2005 were analysed retrospectively. Biochemical pregnancy, ectopic pregnancy and first-trimester abortions were classified as non-viable pregnancies; pregnancies beyond 12 weeks gestation were classified as ongoing pregnancies (OP). Significantly higher D8E2 and D12HCG-beta were obtained in D5 transfers compared with D3 transfers with regard to pregnancy and OP (P

BACKGROUND & AIMS: :A panel of monoclonal antibodies which react with empty non-infectious type 3 poliovirus particles (C antigen) but not infectious virus (D antigen) were characterized for their reactivity with C antigen particles derived from neutralization-resistant virus strains which had single amino acid substitutions at each of the antigenic sites. Antibodies were identified which failed to bind to variant viruses with modifications at each of antigenic sites 2b, 3b and 4 indicating that the same amino acid sequences involved in the neutralization of infectious virus are also present on the surface of non-infectious particles but in different configurations.

背景与目标： : 一组与空的非感染性3型脊髓灰质炎病毒颗粒 (C抗原) 反应但不与感染性病毒 (D抗原) 反应的单克隆抗体的特征在于它们与源自中和抗性病毒株的C抗原颗粒的反应性在每个抗原位点具有单个氨基酸取代。鉴定出的抗体未能与变异病毒结合，并在每个抗原位点2b，3b和4处进行了修饰，这表明参与中和传染性病毒的相同氨基酸序列也存在于非感染性颗粒的表面上，但处于不同的构型。

BACKGROUND & AIMS: UNLABELLED:Radiolabeled benzamides have recently been introduced for the detection of melanoma. We evaluated the potential clinical applicability of 123I-N-(2-diethylaminoethyl) 4-iodobenzamide ([123I]IDAB) for SPECT imaging of ocular melanoma.

METHODS:Fourteen patients were studied, 10 with or suspected of malignant ocular melanoma and four with ocular naevi. All patients underwent SPECT imaging of the head and whole-body scintigraphy 4-5 hr after injection of 170 MBq [123I]IDAB.

RESULTS:A definite tracer hyperfixation was observed in the pathological eye in 9 of 10 (90%) patients with ocular melanoma. The pathological-to-normal eye ratio averaged 1.46 (range 1.07-2.86). The melanoma nature of the scintigraphic lesions was confirmed after enucleation in eight cases and by clinical evolution in two. A false-negative scan was reported in a patient with a small and hypochromic lesion. In patients with ocular naevi, no false-positive scintigrams were documented.

CONCLUSION:Iodine-123-IDAB scintigraphy may contribute significantly to decide about enucleation in cases where some doubt persists with conventional techniques.

背景与目标： 未标记 : 最近已引入放射性标记的苯甲酰胺用于检测黑色素瘤。我们评估了123I-N-(2-二乙氨基乙基) 4-碘苯甲酰胺 ([123I]IDAB) 在眼部黑色素瘤SPECT成像中的潜在临床适用性。
方法 : 研究了14名患者，10例患有或怀疑患有恶性眼部黑色素瘤，4例患有眼部恶性黑色素瘤。注射170 MBq [123I]IDAB后4-5小时，所有患者均接受了头部SPECT成像和全身闪烁显像。
结果 : 在10例 (90% 例) 眼部黑色素瘤患者中，有9例在病理眼中观察到了明确的示踪剂超固定。病理与正常眼的比率平均为1.46 (范围1.07-2.86)。闪烁显像病变的黑色素瘤性质在8例摘除后得到证实，两例通过临床进化得到证实。在一名患有小的低色度病变的患者中报告了假阴性扫描。在患有眼痣的患者中，没有记录假阳性闪烁图。
结论 : 在常规技术仍然存在疑问的情况下，Iodine-123-IDAB闪烁显像可能会大大有助于决定摘除。

BACKGROUND & AIMS: :The aim of this study was to examine the different patterns of cerebellar and/or brainstem atrophy in spinocerebellar ataxia (SCA) type 3 and 6. Eighteen patients (SCA3 n=9, SCA6 n=9) and 15 healthy volunteers were studied. Voxel-based morphometry (VBM) was applied to segmented grey matter (GM) and white matter (WM) of high-resolution T1-weighted brain volumes of each group. We found reduction of grey matter in the pons as well as in the vermis in SCA3 as compared to control subjects. In SCA6 significant grey matter loss was found in hemispheric lobules bilaterally as well as in the vermis. White matter analysis revealed significant changes in SCA3, especially in the pons, in the white matter surrounding the dentate nucleus (DN) and in the cerebellar peduncles, whereas no significant white matter reduction was found in SCA6 patients. Our results demonstrate different patterns of grey and white matter affection detected by magnetic resonance imaging (MRI) in SCA3 and SCA6 patients, confirming the pathological concept of cortical cerebellar atrophy in SCA6. In contrast, SCA3 represents a form of ponto-cerebellar atrophy with predominant affection of pontine nuclei and fibre tracts.

背景与目标： : 这项研究的目的是检查3型和6型脊髓小脑共济失调 (SCA) 的小脑和/或脑干萎缩的不同模式。研究了18名患者 (SCA3 n = 9，SCA6 n = 9) 和15名健康志愿者。将基于体素的形态计量学 (VBM) 应用于每组高分辨率T1-weighted脑体积的分段灰质 (GM) 和白质 (WM)。与对照组相比，我们发现SCA3中的脑桥和ver中的灰质减少。在SCA6中，双侧半球小叶以及ver骨中都发现了明显的灰质损失。白质分析显示SCA3发生了显着变化，尤其是在脑桥，齿状核 (DN) 周围的白质和小脑梗中，而在SCA6患者中未发现明显的白质减少。我们的结果表明，通过磁共振成像 (MRI) 在SCA3和SCA6患者中检测到的灰质和白质的不同模式，证实了SCA6中皮质小脑萎缩的病理概念。相反，SCA3代表一种桥小脑萎缩的形式，主要影响桥脑核和纤维束。

BACKGROUND & AIMS: The present study was designed to elucidate the neurotransmitters involved in activation of the noradrenergic nucleus, locus coeruleus, by distention of the distal colon. Locus coeruleus spontaneous discharge rate was recorded from halothane-anesthetized rats before, during and after distention of the colon produced by inflation of a balloon catheter with varying volumes of water. Locus coeruleus activation by colon distention was volume-dependent and reversible. Activation of cortical electroencephalographic activity was temporally correlated with locus coeruleus activation during colon distention and prolonged distention (greater than 2 min) resulted in tachyphalaxis to both locus coeruleus and cortical electroencephalographic activation. The corticotropin-releasing factor antagonist, DPheCRF(12-41), administered intracerebroventricularly (3 microg) or microinfused into the locus coeruleus (10 ng) significantly attenuated locus coeruleus activation produced by lower, but not higher magnitudes of colon distention, implicating corticotropin-releasing factor afferents to the locus coeruleus in this response. Consistent with this, prior exposure to 30 min of footshock stress, which desensitizes locus coeruleus neurons to corticotropin-releasing factor, produced a similar attenuation of locus coeruleus activation by low, but not high magnitudes of distention. Kynurenic acid, administered intracerebroventricularly (5 micromol), significantly antagonized locus coeruleus activation by all magnitudes of colon distention. However, this excitatory amino acid antagonist was ineffective when administered directly into the locus coeruleus (0.3 nmol). Together, these findings suggest that low magnitudes of colon distention activate the locus coeruleus-noradrenergic system via corticotropin-releasing factor release within the locus coeruleus and that excitatory amino acid neurotransmission at a site distal to the locus coeruleus is necessary for this response. Activation of the locus coeruleus-noradrenergic system during colon distention may serve as a cognitive limb of the peripheral parasympathetic response. This activation may also play a role in disorders characterized by comorbidity of colonic and psychiatric symptoms, such as irritable bowel syndrome.

背景与目标： 本研究旨在阐明通过远端结肠扩张而激活去甲肾上腺素能核蓝斑的神经递质。记录了氟烷麻醉大鼠的蓝斑自然放电速率，该速率是在通过气囊导管充满不同体积的水而产生的结肠扩张之前，期间和之后。结肠扩张对蓝斑的激活是体积依赖性和可逆的。在结肠扩张期间，皮质脑电图活动的激活与蓝斑轨迹的激活在时间上相关，而长时间的扩张 (大于2分钟) 导致蓝斑轨迹和皮质脑电图激活均出现心动过速。促肾上腺皮质激素释放因子拮抗剂DPheCRF(12-41) 在脑室内给药 (3 microg) 或微注入蓝斑 (10 ng) 显着减弱了由较低但不是较高程度的结肠扩张产生的蓝斑激活，在这种反应中，促肾上腺皮质激素释放因子传入蓝斑。与此一致的是，先前暴露于30分钟的脚休克应激，使蓝斑基因座神经元对促肾上腺皮质激素释放因子脱敏，通过低但不高的扩张幅度产生了类似的蓝斑基因座激活衰减。脑室内给药 (5 micromol) 的犬尿酸可通过所有程度的结肠扩张显着拮抗蓝斑的激活。然而，当直接给予蓝斑 (0.3 nmol) 时，这种兴奋性氨基酸拮抗剂是无效的。总之，这些发现表明，低程度的结肠扩张通过在蓝斑内释放促肾上腺皮质激素释放因子来激活蓝斑-去甲肾上腺素能系统，并且在蓝斑远端的兴奋性氨基酸神经传递是这种反应所必需的。结肠扩张期间蓝斑-去甲肾上腺素能系统的激活可能是周围副交感神经反应的认知肢体。这种激活也可能在以结肠和精神症状合并症为特征的疾病中发挥作用，例如肠易激综合征。

BACKGROUND & AIMS: INTRODUCTION:Adverse drug reactions (ADRs) occur frequently in hospitals and increase costs of health care; however, few studies have quantified the clinical and economic impact of ADRs in Colombia. OBJECTIVES:These impacts were evaluated by calculating costs associated with ADRs in patients hospitalized in the internal medicine ward of a Level 3 hospital located in Bogotá, Colombia. In addition, salient clinical features of ADRs were identified and characterized. MATERIAL AND METHODS:Intensive follow-ups for a cohort of patients were conducted for a five month period in order to detect ADRs; different ways to classify them, according to literature, were considered as well. Information was collected using the INVIMA reporting format, and causal probability was evaluated with the Naranjo algorithm. Direct costs were calculated from the perspective of payer, based on the following costs: additional hospital stay, medications, paraclinical tests, additional procedures, patient displacement to intermediate or intensive care units, and other costs. RESULTS:Of 836 patients admitted to the service, 268 adverse drug reactions were detected in 208 patients (incidence proportion 25.1%, occurence rate 0.32). About the ADRs found, 74.3% were classified as probable, 92.5% were type A, and 81.3% were moderate. The body system most often affected was the circulatory system (33.9%). Drugs acting on the blood were most frequently those ones associated with adverse reactions (37.6%). The costs resulting from medical care of adverse drug reactions varied from COL dollar 93,633,422 (USD dollar 35,014.92) to COL dollar 122,155,406 (USD dollar 45,680.94), according to insurance type, during the study period. CONCLUSIONS:Adverse drug reactions have a significant negative health and financial impact on patient welfare. Because of the substantial resources required for their medical care and the significant proportion of preventable adverse reactions, active programs of institutional pharmacovigilance are highly recommended.

背景与目标：

BACKGROUND & AIMS: The GATA family of vertebrate DNA binding regulatory proteins are expressed in diverse tissues and at different times of development. However, the DNA binding regions of these proteins possess considerable homology and recognize a rather similar range of DNA sequence motifs. DNA binding is mediated through two domains, each containing a zinc finger. Previous results have led to the conclusion that although in some cases the N-terminal finger can contribute to specificity and strength of binding, it does not bind independently, whereas the C-terminal finger is both necessary and sufficient for binding. Here we show that although this is true for the N-terminal finger of GATA-1, those of GATA-2 and GATA-3 are capable of strong independent binding with a preference for the motif GATC. Binding requires the presence of two basic regions located on either side of the N-terminal finger. The absence of one of these near the GATA-1 N-terminal finger probably accounts for its inability to bind. The combination of a single finger and two basic regions is a new variant of a motif that has been previously found in the binding domains of other finger proteins. Our results suggest that the DNA binding properties of the N-terminal finger may help distinguish GATA-2 and GATA-3 from GATA-1 and the other GATA family members in their selective regulatory roles in vivo.

背景与目标： 脊椎动物DNA结合调节蛋白的GATA家族在不同的组织和发育的不同时间表达。但是，这些蛋白质的DNA结合区域具有相当大的同源性，并且可以识别相当相似范围的DNA序列基序。DNA结合通过两个结构域介导，每个结构域都包含一个锌指。先前的结果得出的结论是，尽管在某些情况下，N末端手指可以促进特异性和结合强度，但它不会独立结合，而C末端手指对于结合既必要又足够。在这里，我们表明，尽管对于GATA-1的N末端手指是正确的，但GATA-2和GATA-3的手指能够强烈独立结合，并且偏爱基序GATC。结合需要存在位于N末端手指两侧的两个基本区域。在GATA-1的N末端手指附近没有这些手指之一可能是其无法结合的原因。单个手指和两个基本区域的组合是基序的新变体，以前已在其他手指蛋白的结合域中发现。我们的结果表明，N末端手指的DNA结合特性可能有助于将GATA-2和GATA-3与GATA-1和其他GATA家族成员在体内的选择性调节作用区分开。

BACKGROUND & AIMS: :In Escherichia coli at least five enzyme activities required for the beta-oxidation of fatty acids are associated with a multienzyme complex composed of two subunits in alpha 2 beta 2 conformation (A. Pramanik et al., J. Bacteriol. 137:469-473, 1979). In the present work, the DNA sequence of the genes encoding these two subunits, fadB and fadA, has been determined. The direction of transcription was from fadB to fadA rather than from fadA to fadB, as suggested previously (S. K. Spratt et al., J. Bacteriol. 158:535-542, 1984). Only 10 nucleotides separated the coding sequences for the two peptides, confirming the suggestion that these genes form an operon. The peptides encoded by fadB and fadA were 729 amino acids and 387 amino acids, respectively, in length. The larger and smaller peptides had predicted molecular masses of 79,678 and 40,876 Da, respectively. Recently, the sequence of the fadA gene was published in a separate report (Yang et al., J. Biol. Chem. 265:10424-10429, 1990). In this work, most of the DNA sequence for fadA was confirmed, and 10 errors were corrected. Three of these nucleotide changes resulted in five amino acid residue changes predicted in the carboxy terminus of the fadA-encoded peptide. By comparison to other peptide sequences, the alpha subunit encoded within fadB had 31% perfect identity with the rat peroxisomal enoyl-coenzyme A:hydratase-3-hydroxyacyl-coenzyme A dehydrogenase trifunctional enzyme over the entire length of the two peptides. In agreement with the work of Yang et al., the beta subunit encoded within fadA had 35 to 45% perfect identity with five thiolase genes from different eucaryotic sources over the entire length of the peptide.

背景与目标： 在大肠杆菌中，脂肪酸的 β-氧化所需的至少五种酶活性与由 α2β2构象中的两个亚基组成的多酶复合物相关 (a.Pramanik等人，J. Bacteriol. 137:469-473，1979)。在本工作中，已经确定了编码这两个亚基fadB和fadA的基因的DNA序列。转录的方向是从fadB到fadA，而不是从fadA到fadB，如前所述 (S. K. Spratt等人，J. Bacteriol. 158:535-542，1984)。只有10个核苷酸分离了两个肽的编码序列，证实了这些基因形成操纵子的暗示。fadB和fadA编码的肽长度分别为729个氨基酸和387个氨基酸。较大和较小的肽分别预测79,678和40,876 Da的分子量。最近，fadA基因的序列发表在单独的报告中 (Yang等人，J. Biol. Chem. 265:10424-10429，1990)。在这项工作中，fadA的大部分DNA序列得到了确认，并纠正了10个错误。这些核苷酸变化中的三个导致fadA编码肽的羧基末端预测的五个氨基酸残基变化。与其他肽序列相比，fadB内编码的 α 亚基与大鼠过氧化物酶体烯酰辅酶a具有31% 完美的同一性: 在两个肽的整个长度上hydratase-3-hydroxyacyl-coenzyme脱氢酶三官能酶。与Yang等人的工作一致，在fadA内编码的 β 亚基在肽的整个长度上与来自不同真核来源的五个硫解酶基因具有35至45% 完美的同一性。

BACKGROUND & AIMS: :Neurons within each layer of cerebral cortex express multiple members of the neurotrophin family and their corresponding receptors. This multiplicity could provide functional redundancy; alternatively, different neurotrophins may direct distinct aspects of cortical neuronal growth and differentiation. By neutralizing endogenous neurotrophins in organotypic slices of developing cortex with Trk receptor bodies (Trk-IgGs), we found that BDNF and NT-3 oppose one another in regulating the dendritic growth of pyramidal neurons. In layer 4, both endogenous and exogenous NT-3 inhibited the dendritic growth stimulated by BDNF. In contrast, in layer 6 both endogenous and exogenous BDNF inhibited dendritic growth stimulated by NT-3. These antagonistic actions of endogenous BDNF and NT-3 provide a mechanism by which dendritic growth and retraction can be dynamically regulated during cortical development, and suggest that the multiple neurotrophins expressed in developing cortex represent distinct components of an extracellular signaling system for regulating dendritic growth.

背景与目标： : 大脑皮层各层内的神经元表达神经营养蛋白家族的多个成员及其相应的受体。这种多样性可以提供功能冗余; 或者，不同的神经营养蛋白可能会指导皮质神经元生长和分化的不同方面。通过用Trk受体体 (Trk-igg) 中和发育中的皮层器官型切片中的内源性神经营养蛋白，我们发现BDNF和NT-3在调节锥体神经元的树突状生长方面相互对抗。在第4层中，内源性和外源性NT-3均抑制BDNF刺激的树突状生长。相反，在第6层中，内源性和外源性BDNF均抑制NT-3刺激的树突生长。内源性BDNF和NT-3的这些拮抗作用提供了一种机制，通过该机制可以在皮质发育过程中动态调节树突生长和收缩，并表明在发育中的皮质中表达的多种神经营养蛋白代表了调节树突生长的细胞外信号系统的不同成分。

BACKGROUND & AIMS: :Matrix metalloproteinases (MMPs) have been the subject of intense research because of their roles in tumor metastasis and in the rise and spread of degenerative diseases such as osteo- and rheumatoid arthritis. A preliminary class of 140 druglike, small-molecule matrix metalloproteinase-3 inhibitors, intended as starting scaffolds for optimization and synthesis, has been designed in silico using a series of highly predictive three-dimensional quantitative structure-activity relationship models, including comparative molecular field analysis and comparative molecular similarity indices analysis, with docking and scoring. Thalidomide was chosen as the skeleton on which to base the new lead series, as it moderately inhibits MMP-3, is antiangiogenic, and lends itself easily to structural modifications. Most of the new compounds demonstrate medium to high predicted biological activity and good bioavailability as estimated by the octanol-water partition coefficient ClogP. Compound 102 in particular exhibits extremely favorable predicted activity against MMP-3; is moderately bioavailable; satisfies Lipinski's Rule of Five; and shows promise for further optimization, synthesis, and experimental evaluation as a potential adjunct anticancer or antirheumatic therapeutic.

背景与目标： : 基质金属蛋白酶 (MMPs) 由于其在肿瘤转移以及退行性疾病 (如骨关节炎和类风湿性关节炎) 的兴起和传播中的作用而成为深入研究的主题。初步设计了140类药物小分子基质metalloproteinase-3抑制剂，作为优化和合成的起始支架，使用一系列高度预测性的三维定量构效关系模型，包括比较分子场分析和比较分子相似性指数分析，对接和得分。沙利度胺被选为新铅系列的骨架，因为它适度抑制MMP-3，具有抗血管生成作用，并且易于进行结构修饰。根据辛醇-水分配系数ClogP估计，大多数新化合物表现出中等至高的预测生物活性和良好的生物利用度。化合物102尤其表现出对MMP-3极其有利的预测活性; 具有适度的生物利用度; 满足Lipinski的五法则; 并显示出有望进一步优化，合成和实验评估作为潜在的辅助抗癌或抗风湿治疗剂。

BACKGROUND & AIMS: :Apicomplexan parasites are the cause of numerous important human diseases including malaria and AIDS-associated opportunistic infections. Drug treatment for these diseases is not satisfactory and is threatened by resistance. The discovery of the apicoplast, a chloroplast-like organelle, presents drug targets unique to these parasites. The apicoplast-localized fatty acid synthesis (FAS II) pathway, a metabolic process fundamentally divergent from the analogous FAS I pathway in humans, represents one such target. However, the specific biological roles of apicoplast FAS II remain elusive. Furthermore, the parasite genome encodes additional and potentially redundant pathways for the synthesis of fatty acids. We have constructed a conditional null mutant of acyl carrier protein, a central component of the FAS II pathway in Toxoplasma gondii. Loss of FAS II severely compromises parasite growth in culture. We show FAS II to be required for the activation of pyruvate dehydrogenase, an important source of the metabolic precursor acetyl-CoA. Interestingly, acyl carrier protein knockout also leads to defects in apicoplast biogenesis and a consequent loss of the organelle. Most importantly, in vivo knockdown of apicoplast FAS II in a mouse model results in cure from a lethal challenge infection. In conclusion, our study demonstrates a direct link between apicoplast FAS II functions and parasite survival and pathogenesis. Our genetic model also offers a platform to dissect the integration of the apicoplast into parasite metabolism, especially its postulated interaction with the mitochondrion.

背景与目标： : Apicomplexan寄生虫是许多重要人类疾病的病因，包括疟疾和与艾滋病相关的机会性感染。这些疾病的药物治疗并不令人满意，并受到耐药性的威胁。叶绿体样细胞器apicoplast的发现提出了这些寄生虫特有的药物靶标。apicoplast定位脂肪酸合成 (FAS II) 途径是一种与人类类似FAS I途径根本不同的代谢过程，它代表了一个这样的目标。然而，apicoplast FAS II的特定生物学作用仍然难以捉摸。此外，寄生虫基因组还编码用于合成脂肪酸的其他且潜在的冗余途径。我们已经构建了酰基载体蛋白的条件无效突变体，这是弓形虫中FAS II途径的核心组成部分。FAS II的丢失严重损害了寄生虫在培养中的生长。我们显示FAS II是激活丙酮酸脱氢酶所必需的，丙酮酸脱氢酶是代谢前体乙酰辅酶a的重要来源。有趣的是，酰基载体蛋白敲除还导致apicoplast生物发生缺陷，并导致细胞器丢失。最重要的是，在小鼠模型中体内敲除apicoplast FAS II可从致命的攻击感染中治愈。总之，我们的研究证明了apicoplast FAS II功能与寄生虫存活和发病机理之间的直接联系。我们的遗传模型还提供了一个平台，可以将apicoplast整合到寄生虫代谢中，尤其是其与线粒体的假定相互作用。

BACKGROUND & AIMS: The effect of epinephrine on leucine and phenylalanine kinetics was measured by using the stable isotope amino acid tracers L-[1-(13)C]leucine and L-[phenyl-2H5]-phenylalanine in the postabsorptive state and during the intravenous administration of a standard amino acid solution with respect to the amino acid load. Infusion of epinephrine (plasma concentrationapproximately 3600 pmol/L) decreased leucine and phenylalanine and increased ketoisocaproate plasma concentrations and increased the metabolic clearance rate of leucine and phenylalanine. Epinephrine neither influenced leucine or phenylalanine flux nor leucine oxidation or leucine net balance. Hyperaminoacidemia from amino acid infusion reduced endogenous leucine release and stimulated leucine oxidation and nonoxidative disposal of leucine, resulting in a dose-dependent increase in leucine net balance. Epinephrine did not influence any changes in amino acid kinetics during parenteral amino acid administration. Therefore, we conclude that epinephrine had no catabolic effects on amino acid metabolism and no negative effect on the utilization of a parenterally offered amino acid solution in healthy humans.

背景与目标： 肾上腺素对亮氨酸和苯丙氨酸动力学的影响是通过使用稳定同位素氨基酸示踪剂L-[1-(13)C] 亮氨酸和L-[phenyl-2H5]-苯丙氨酸在后吸收状态和静脉内给药期间测量的。标准氨基酸溶液相对于氨基酸负荷输注肾上腺素 (血浆浓度约3600 pmol/L) 降低亮氨酸和苯丙氨酸，增加酮异己酸血浆浓度，增加亮氨酸和苯丙氨酸的代谢清除率。肾上腺素既不影响亮氨酸或苯丙氨酸通量，也不影响亮氨酸氧化或亮氨酸净平衡。氨基酸输注的高氨基酸血症降低了内源性亮氨酸释放并刺激亮氨酸氧化和亮氨酸的非氧化处理，导致亮氨酸净平衡的剂量依赖性增加。肾上腺素在肠胃外给药期间不影响氨基酸动力学的任何变化。因此，我们得出的结论是，肾上腺素对健康人的氨基酸代谢没有分解代谢作用，对肠胃外提供的氨基酸溶液的利用也没有负面影响。

BACKGROUND & AIMS: MyristoylCoA:protein N-myristoyltransferase (NMT) catalyzes the cotranslational covalent attachment of a rare cellular fatty acid, myristate, to the N-terminal Gly residue of a variety of eukaryotic proteins. The myristoyl moiety is often essential for expression of the biological functions for these proteins.

Attachment of C14:0 alone provides barely enough hydrophobicity to allow stable association with membranes. The partitioning of N-myrisotylproteins is therefore often modulated by "switches" that function through additional covalent or noncovalent modifications. Candida albicans, the principal cause of systemic fungal infection in immunocompromised humans, contains a single NMT gene that is essential for its viability. The functional properties of the acylCoA binding site of human and C. albicans NMT are very similar. However, there are distinct differences in their peptide binding sites. An ADP ribosylation factor (Arf) is included among the few cellular protein substrates of the fungal enzyme. Alanine scanning mutagenesis of an octapeptide derived from an N-terminal Arf sequence (GLYASKLS-NH2) disclosed that Gly1, Ser5, and Lys6 play predominant roles in binding. ALYASKLS-NH2 is an inhibitor competitive for peptide [Ki(app) = 15.3 +/- 6.4 microM] and noncompetitive for myristoylCoA. Remarkably, replacement of the N-terminal tetrapeptide with an 11-aminoundecanoyl group results in a competitive inhibitor (11-aminoundecanoyl-SKLS-NH2) that is approximately 40-fold more potent [Ki(app) = 0.40 +/- 0.03 microM] than the starting octapeptide. Removal of Leu-Ser from the C-terminus generates a competitive dipeptide inhibitor (11-aminoundecanoyl-SK-NH2) with a Ki(app) of 11.7 +/- 0.4 microM, equivalent to that of the starting octapeptide. A derivative dipeptide inhibitor containing a C-terminal N-cyclohexylethyl lysinamide moiety has the advantage of being more potent (IC50 = 0.11 +/- 0.03 microM) and resistant to digestion by cellular carboxypeptidases. Rigidifying the flexible aminoundecanoyl chain results in very potent general NMT inhibitors (IC50 = 40-50 nM). Substituting a 2-methylimidazole for the N-terminal amine and adding a benzylic alpha-methyl group with R stereochemistry to the rigidifying element produces even more potent inhibitors (IC50 = 20-50 nM) that are up to 500-fold selective for the fungal compared to human enzyme. A related less potent member of this series of compounds is fungistatic. Its growth inhibitory effects are associated with a reduction in cellular protein N-myristoylation, judged using cellular Arf as a reporter. These studies establish that NMT is a new antifungal target.

背景与目标： 肉豆蔻酰辅酶a : 蛋白质N-肉豆蔻酰转移酶 (NMT) 催化稀有细胞脂肪酸肉豆蔻酸酯与多种真核蛋白质的N末端Gly残基的共翻译共价连接。肉豆蔻酰部分对于表达这些蛋白质的生物学功能通常是必不可少的。
C14的附着 :0仅提供了足够的疏水性，无法与膜稳定结合。因此，N-肉豆蔻基蛋白的分配通常由 “开关” 调节，该开关通过其他共价或非共价修饰起作用。白色念珠菌是免疫功能低下的人体全身性真菌感染的主要原因，它包含一个对其生存能力至关重要的NMT基因。人与白色念珠菌NMT的酰基辅酶a结合位点的功能特性非常相似。然而，它们的肽结合位点存在明显差异。真菌酶的少数细胞蛋白底物中包括ADP核糖基化因子 (Arf)。衍生自N-末端Arf序列 (GLYASKLS-NH2) 的八肽的丙氨酸扫描诱变揭示Gly1、Ser5和Lys6在结合中起主要作用。ALYASKLS-NH2是一种对肽 [Ki(app) = 15.3 +/- 6.4 microM] 有竞争力的抑制剂，对肉豆蔻酰辅酶a无竞争力。值得注意的是，用11-氨基十一酰取代N-末端四肽导致竞争性抑制剂 (11-氨基十一酰-skls-nh2) 的效力比起始八肽高约40倍 [Ki(app) = 0.40 +/- 0.03微米]。从C末端去除Leu-Ser会产生竞争性二肽抑制剂 (11-氨基十一酰-sk-nh2)，Ki(app) 为11.7 +/- 0.4微米，相当于起始八肽。含有C-末端N-环己基乙基赖氨酰胺部分的衍生物二肽抑制剂具有更有效 (IC50 = 0.11 +/- 0.03微米) 和对细胞羧肽酶消化的抗性的优点。硬化柔性氨基十一酰链会产生非常有效的通用NMT抑制剂 (IC50 = 40-50nm)。用2-甲基咪唑代替N-末端胺并将具有R立体化学的苄基 α-甲基添加到硬化元件中，产生甚至更有效的抑制剂 (IC50 = 20-50nm)，其对真菌的选择性与人酶相比高达500倍。该系列化合物中一个相关的效力较低的成员是真菌抑制剂。使用细胞Arf作为报告基因，其生长抑制作用与细胞蛋白N-肉豆蔻酰化的减少有关。这些研究表明，NMT是一种新的抗真菌靶标。

BACKGROUND & AIMS: Assembly of fibronectin fibrils occurs at the surface of substrate-attached cells and is mediated by the first to the fifth type I modules in the N-terminal 70 kDa portion of the molecule. The first type III module (III1) of fibronectin, not present in the 70 kDa portion, contains a conformation-dependent binding site for the 70 kDa N-terminal region of fibronectin, suggesting that the III1 module on cell-surface fibronectin may serve as a binding site for fibronectin's N-terminus on substrate-attached cells. To explore this possiblility, we compared the ability of mutant recombinant 70 kDa proteins containing deletions of one or several of the first five type I modules to bind to fibroblasts and to III1. Proteins containing the fourth and fiftBiomolecular Chemistry and Medicine, University of Wisconsin, Madison, WI 53706U.S.A. Assembly of fibronectin fibrils occurs at the surface of substrate-attached cells and is mediated by the first to the fifth type I modules in the N-terminal 70 kDa portion of the molecule. The first type III module (III1) of fibronectin, not present in the 70 kDa portion, contains a conh as 70 kDa deletion mutants lacking I4 and I5 also bound to the cell surface, and deletion mutants lacking I1-3 and I4-5 both competed only partially for binding of 125I-labelled fibronectin or 70 kDa protein. These data indicate that the N-terminal part of fibronectin binds to III1 via I4 and I5 and that interactions in addition to that of I4 and I5 with III1 are important for cell-surface-mediated fibronectin polymerization.

背景与目标： 纤连蛋白原纤维的组装发生在附着于底物的细胞的表面，并由分子的N末端70 kDa部分中的第一至第五种I型模块介导。纤连蛋白的第一个III型模块 (III1) 不存在于70 kDa部分中，包含纤连蛋白70 kDa N末端区域的构象依赖性结合位点，提示细胞表面纤连蛋白上的III1模块可以用作底物附着细胞上纤连蛋白N末端的结合位点。为了探索这种可能性，我们比较了含有前五个I型模块中一个或几个缺失的突变重组70 kDa蛋白与成纤维细胞和iii1结合的能力。含有第四和第五生物分子化学和医学的蛋白质，威斯康星大学麦迪逊分校，WI 53706u.S.A.纤连蛋白原纤维的组装发生在附着于底物的细胞的表面，并由分子的N末端70 kDa部分中的第一至第五种I型模块介导。纤连蛋白的第一个III型模块 (III1) 不存在于70 kda部分中，包含也与细胞表面结合的缺乏I4和I5的conh as 70 kda缺失突变体，并且缺乏I1-3和I4-5的缺失突变体都仅部分竞争125i标记的纤连蛋白或70 kda蛋白的结合。这些数据表明，纤连蛋白的N端部分通过I4和I5与III1结合，并且除了I4和I5与III1的相互作用外，还对细胞表面介导的纤连蛋白聚合很重要。