Fragile X syndrome,the Fragile X related proteins,and animal models

The Fragile X syndrome (FraX), which is characterized among other physical and neurologic impairments by mental retardation, is caused by the absence of the product of the FMR1 gene. The Fragile X Mental Retardation Protein (FMRP) is a member of a novel family of RNA-binding proteins. The latter includes two other proteins highly homologous with FMRP: the fragile X related proteins 1 and 2 (FXRP1 and FXRP2). Characterization of FXRPs, including their interaction with FMRP, will provide critical information about the mechanisms of action of FMRP and the role of this group of proteins in FMRP-deficient conditions such as FraX. Genetic manipulations of FMRP and the FXRPs should also provide valuable tools for investigating pathophysiology and gene therapies in FraX. The present review summarizes the strategies used for identifying the FXRPs, their chromosomal localization, molecular structure, and tissue distribution. It also reviews interactions between different members of this family of RNA-binding proteins. Animal models, both knockout and transgenic, of FMRP and the FXRPs are discussed. Phenotypic features of the FMR1 knockout mouse, the FMR1 transgenic rescue mouse, and other novel strategies for manipulating and delivering FMRP and FXRPs to the brain and other tissues are described.     

Effects of Fragile X syndrome and an FMR1 knockout mouse model on forebrain neuronal cell biology

The neurological deficits exhibited by patients with Fragile X syndrome (FraX) have been attributed to the absence of the Fragile X Mental Retardation Protein (FMRP), the product of the FMR1 gene, which is nonfunctional in these individuals. While a great deal has been learned about FraX using non-invasive techniques and autopsy tissue from humans, the limited availability of subjects and specimens severely restricts the rate at which such data can be collected and the types of experimental questions posed. In view of these limitations, a transgenic mouse model of FraX has been constructed in which the FMR1 gene is selectively knocked out (KO) [Bakker et al. (1994) Cell 78:23-33]. These mice show molecular, morphological, and behavioral alterations consistent with phenotypes observed in FraX patients, making them good models to study the absence of FMRP expression.     

Fragile X mental retardation: Misregulation of protein synthesis in the developing brain?

Fragile X mental retardation results from the absence of a selective RNA-binding protein, FMRP. Previous studies demonstrated that FMRP forms messenger ribonucleoprotein (mRNP) complexes to associate with translating polyribosomes, suggesting that FMRP is involved in regulating protein synthesis. We are now facing the changing questions: How does FMRP influence protein synthesis in the brain? What is the target for FMRP in learning and memory? How does the absence of FMRP cause misregulation of protein synthesis, which in turn leads to mental impairment in fragile X syndrome? Models for abnormal neuronal function as a result of misregulated translation due to the absence of FMRP are discussed. Microsc. Res. Tech. 57:145–147, 2002. © 2002 Wiley-Liss, Inc.     

Influence of stimulants on electrodermal studies in Fragile X syndrome

Attention Deficit Hyperactivity Disorder (ADHD) is seen in the majority of children with Fragile X Syndrome (FraX). Previous work has documented an enhanced sweat response to stimuli in children with FraX compared to controls utilizing electrodermal response (EDR) measures. The present study assesses the EDRs both on and off stimulants in 19 children with ADHD and FraX compared to 17 age- and IQ-matched control patients with ADHD and developmental delays. Although the baseline EDRs were comparable between FraX patients and controls, the patients with FraX had a significant decrease in EDR amplitude and number of peaks when treated with stimulants compared to controls. This suggests that patients with FraX are more responsive to the enhancement of inhibitory systems that occur with stimulant use for ADHD. The use of a quantifiable measure, such as EDR, is recommended in future studies of treatment efficacy.     

Cerebral growth in Fragile X syndrome: review and comparison with Down syndrome

Neuroimaging studies have shown selective changes in brain size in Fragile X syndrome (FraX), which include reductions in the posterior cerebellar vermis, age-dependent increases in hippocampal volume, and enlarged caudate nucleus and thalamus. Contrasting with these limbic and subcortical anomalies, much less is known about the neocortex in FraX. The present study attempted to examine cerebral and lobar-level volumetric changes in young males with FraX (2-7 years), by comparing groups of subjects with full mutation (FM) and mosaicism (Mos) with both age-matched controls and subjects with developmental language delay (DLD) and Down syndrome (DS). For this purpose, we used high resolution (i.e, SPGR) MRI scans and semi-automated methods for segmenting (tissue class) and parcellating (i.e., Talairach) the brain. In agreement with previous studies, we found no changes in overall brain or cerebrum size in FraX. Nevertheless, boys with FM FraX had relative reductions in temporal lobe volume (primarily gray matter) and relative preservation/enlargement of parietal white matter volume. While temporal lobe reductions were not specific, since they were also observed in DLD and DS subjects, parietal preservation/enlargement was only seen in FraX. The relevance of these preliminary findings was emphasized by comparisons between FraX groups, which revealed more marked changes in FM FraX than in Mos FraX (i.e., gene dosage). While cross-sectional analyses revealed marked age-dependent decreases in DS, a group showing marked global and lobar volumetric reductions, there were no changes over time in FraX. These neuroimaging data are discussed in the context of FraX neurobiology and other developmental disorders.     

Early development in males with Fragile X syndrome: a review of the literature

This article reviews the current bibliographic knowledge on early neurobehavioral development and milestones in Fragile X syndrome (FraX), with emphasis on males affected by the condition. Three broad areas of early development were examined: (1) gross and fine motor, (2) speech and language, and (3) social. The result of the current review indicates very limited information on the developmental milestones in all three areas. The scarce literature on motor development shows that in FraX there is an early developmental delay. Research on speech and language demonstrates pervasive deficits in conversational skills and severe developmental delay, with increasing discrepancy between language level and chronological age in young males with FraX. Finally, deficits in social development in FraX include abnormal gaze, approach and avoidance conflict, and high incidence of autistic spectrum disorders.     

A developmental approach to understanding Fragile X syndrome in females

The psychological phenotype of females with fragile X syndrome (FraX) is discussed, focusing primarily on empirical findings over the past decade and on studies of probands with the full mutation (FM). A developmental approach is used to help characterize specific patterns of cognitive, neuropsychological, social, emotional, and behavioral functioning across the lifespan of females with FraX. Approximately half of females with the syndrome present with cognitive abilities that fall in the borderline to mentally retarded range, and the remaining females with average intellectual functioning may experience relative deficits in math achievement and problems with attention and executive functioning. Reports of socio-emotional functioning are somewhat inconsistent, due in part, perhaps, to methodological differences in study design. To date, much of what we understand about the psychological phenotype of FraX is based on cross-sectional studies of girls and women with the disorder. Symptoms associated with shyness, and social anxiety and avoidance have been reported in some school-age, adolescent, and adult females with FraX. Only recently have efforts begun to identify the developmental trajectory of FraX in infants and toddlers. There is a void of information specific to these developmental periods. Identifying key deficits in cognitive and socio-emotional functioning has important implications for early detection and intervention for girls with FraX. Directions for future research are discussed.     

Molecular pathways of oligodendrocyte apoptosis revealed by mutations in the proteolipid protein gene

A decade after the genetic link was established between mutations in the proteolipid protein gene and two leukodystrophies, Pelizaeus-Merzbacher disease and spastic paraplegia, the molecular mechanisms underlying pathogenesis are beginning to come to light. Data from animal models of these diseases suggest that the absence of proteolipid protein gene products in the central nervous system confers a relatively mild phenotype while missense mutations in and duplications of this gene give rise to mild or severe forms of disease. Previously, we have interpreted the disease process in terms of the accumulation of the mutant proteins in the secretory pathway and, herein, we review the evidence in favor of such a cellular mechanism. Furthermore, on the basis of recent data we suggest that the unfolded protein response may be involved in the pathogenesis of Pelizaeus-Merzbacher disease and spastic paraplegia through a kinase signaling cascade that links the accumulation of mutant proteins in the endoplasmic reticulum of oligodendrocytes with changes in gene regulation, protein synthesis, and possibly apoptosis.     

Identification and characterization of molecular targets of natural products by mass spectrometry

Natural products, and their derivatives and mimics, have contributed to the development of important therapeutics to combat diseases such as infections and cancers over the past decades. The value of natural products to modern drug discovery is still considerable. However, its development is hampered by a lack of a mechanistic understanding of their molecular action, as opposed to the emerging molecule‐targeted therapeutics that are tailored to a specific protein target(s). Recent advances in the mass spectrometry‐based proteomic approaches have the potential to offer unprecedented insights into the molecular action of natural products. Chemical proteomics is established as an invaluable tool for the identification of protein targets of natural products. Small‐molecule affinity selection combined with mass spectrometry is a successful strategy to “fish” cellular targets from the entire proteome. Mass spectrometry‐based profiling of protein expression is also routinely employed to elucidate molecular pathways involved in the therapeutic and possible toxicological responses upon treatment with natural products. In addition, mass spectrometry is increasingly utilized to probe structural aspects of natural products–protein interactions. Limited proteolysis, photoaffinity labeling, and hydrogen/deuterium exchange in conjunction with mass spectrometry are sensitive and high‐throughput strategies that provide low‐resolution structural information of non‐covalent natural product–protein complexes. In this review, we provide an overview on the applications of mass spectrometry‐based techniques in the identification and characterization of natural product–protein interactions, and we describe how these applications might revolutionize natural product‐based drug discovery. © 2009 Wiley Periodicals, Inc., Mass Spec Rev 29:126–155, 2010     

A novel high-throughput scanning microscope for label-free detection of protein and small-molecule chemical microarrays

We describe a novel scanning optical microscope based on a polarization-modulated nulling ellipsometry. The new microscope employs a combination of scanning mirror and sample translation and thus enables high-throughput label-free detection of biomolecular microarrays with more than 10 000 protein or small-molecule targets. For illustration, we show the image of a 2760-spot protein microarray on a functionalized glass slide obtained with such a microscope. The new scanning microscope is also capable of determining, in parallel, the real-time binding kinetics of multiple molecular species under aqueous conditions.     

Network pharmacology and molecular docking identify mechanisms of medicinal plant-derived 1,2,3,4,6-penta-O-galloyl-beta-D-glucose treating gastric cancer

Background: 1,2,3,4,6-penta-O-galloyl-beta-D-glucose (PGG) is a natural polyphenolic compound derived from multiple medicinal plants with favorable anticancer activity. Methods: In this study, the mechanisms of PGG against gastric cancer were explored through network pharmacology and molecular docking. First, the targets of PGG were searched in the Herbal Ingredients’ Targets (HIT), Similarity Ensemble Approach (SEA), and Super-PRED databases. The potential targets related to gastric cancer were predicted from the Human Gene Database (GeneCards) and DisGeNET databases. The intersecting targets of PGG and gastric cancer were obtained by Venn diagram and then subjected to protein-protein interaction analysis to screen hub targets. Functional and pathway enrichment of hub targets were analyzed through Gene Ontology and Kyoto Encyclopedia of Genes and Genomes pathway databases. The differential expression and survival analysis of hub targets in gastric cancer were performed based on The Cancer Genome Atlas database. Finally, the affinity of PGG with hub targets was visualized by molecular docking. Results: Three hub targets were screened, including mitogen-activated protein kinase 14 (MAPK14), BCL2 like 1 (BCL2L1), and vascular endothelial growth factor A (VEGFA). MAPK14 had a higher expression, while BCL2L1 and VEGFA had lower expression in gastric cancer than in normal conditions. Enrichment analysis indicated enrichment of these hub targets in MAPK, neurotrophin, programmed death-ligand 1 (PD-L1) checkpoint, phosphatidylinositol 3-kinases/protein kinase B (PI3K-Akt), Ras, and hypoxia-inducible factor-1 (HIF-1) signaling pathways. Conclusion: Therefore, network pharmacology and molecular docking analyses revealed that PGG exerts a therapeutic efficacy on gastric cancer by multiple targets (MAPK14, BCL2L1, and VEGFA) and pathways (MAPK, PD-L1 checkpoint, PI3K-Akt, Ras, and HIF-1 pathways).     

Brief Note: Influence of 2-methoxyestradiol on MCF-7 cells: An improved differential interference contrasting technique and Bcl-2 and Bax protein expression levels

Proteins of the B-cell lymphoma 2 family are crucial for the regulation of apoptosis. B-cell lymphoma 2-associated X is a pro-apoptotic protein, while B-cell lymphoma 2 protein opposes apoptosis. The influence of 1 μM 2-methoxyestradiol was investigated on the expression levels of these two proteins in MCF-7 cells. 2-Methoxyestradiol exposure did not influence B-cell lymphoma 2 protein expression levels after 24 h of exposure. In contrast, B-cell lymphoma 2-associated X protein levels were significantly reduced. An improved differential interference contrasting technique revealed compromised cell density and the presence of a mitotic block in exposed cells. The study proposes that the influence of 2-methoxyestradiol on the expression of these proteins may be time- and cell type dependent and thus not evident during the mitotic block observed. Investigation of the regulation of the B-cell lymphoma 2 family will allow researchers to consider signaling pathways for diseases where apoptosis can potentially be controlled.     

Structure analysis of small proteins by electron microscopy: valinomycin, bacitracin and low molecular weight cell growth stimulators.

Dark field electron microscopy was combined with optical filtering to study at high resolution the structure of the cyclopeptide antibiotics, bacitracin and valinomycin, and two proteins of unknown structure, LMW-CSA N and B, low molecular weight granulocyte colony stimulating activity isolated from medium conditioned with normal or leukemic leukocytes. For bacitracin and valinomycin the images faithfully represented the known structural features at a resolution of 0.5 nm or better, depicting a two-ring structure for bacitracin, as well as the position of the potassium ion in valinomycin. Both proteins of unknown structrue had at least one cyclic peptide portion. LMW-CSA N had a size of 2.0 nm, LMW-CSA B of 2.4 nm. A potential site of the calcium ionophoric activity in the latter protein was found to be in the larger of the two ring portions constituting the molecule.     

Brain drug delivery, drug metabolism, and multidrug resistance at the choroid plexus

The choroid plexuses (CPs) have the capability to modulate drug delivery to the cerebrospinal fluid (CSF) and to participate in the overall cerebral biodisposition of drugs. The specific morphological properties of the choroidal epithelium and the existence of a CSF pathway for drug distribution to different targets in the central nervous system suggest that the CP-CSF route is more significant than previously thought for brain drug delivery. In contrast to its role in CSF penetration of drugs, CP is also involved in brain protection in that it has the capacity to clear the CSF from numerous potentially harmful CSF-borne exogenous and endogenous organic compounds into the blood. Furthermore, CP harbors a large panel of drug-metabolizing enzymes as well as transport proteins of the multidrug resistance phenotype, which modulate the cerebral bioavailability of drugs and toxins. The use of an in vitro model of the choroidal epithelium suitable for drug transport studies has allowed the demonstration of the choroidal epithelium acting as an effective metabolic blood-CSF barrier toward some xenobiotics, and that a vectorial, blood-facing efflux of conjugated metabolites occurs at the choroidal epithelium. This efflux involves a specific transporter with characteristics similar to those of the multidrug resistance associated protein (MRP) family members. Indeed, at least one member, MRP1, is largely expressed at the CP epithelium, and localizes at the basolateral membrane. These metabolic and transport features of the choroidal epithelium point out the CP as a major detoxification site within the brain.     

Exploration of the oxidative-inflammatory potential targets of <i>Coicis Semen</i> in osteoarthritis: Data mining and systematic pharmacology

Objective: On the basis of data mining, systematic pharmacology, molecular docking, and experiment validation, the oxidative-inflammatory molecular targets of Coicis Semen in the therapy of osteoarthritis (OA) were explored. Methods: The association rule analysis was effectively applied to highlight the correlation between Coicis Semen and oxidative inflammation indices. The random walk model was subsequently used to evaluate the clinical efficacy of Coicis Semen. Network pharmacology was used to predict network targets. The binding affinity of the active ingredient in Coicis Semen to the key target of OA was also successfully predicted. Results: Coicis Semen showed a significant reduction in oxidative-inflammatory indicators of OA. A total of 108 promising targets were predicted for the 24 bioactive compounds in Coicis Semen. Eight target genes were considered core target genes. The enrichment analysis predicts that Coicis Semen may activate the interleukin (IL)-17, mitogen-activated protein kinase (MAPK), and nuclear factor kappa B (NF-kappa B) signaling pathways. Molecular docking demonstrated that stigmasterol, 2-monoolein, sitosterol, and sitosterol alpha1 had free binding energies to oxidative and inflammatory targets (MAPK1, Estrogen Receptor 1 [ESR1], and Peroxisome Proliferator-Activated Receptor Alpha [PPARA]). Both clinical trials and in vitro cell experiments revealed that Coicis Semen could increase ESR1 and PPAR-α levels while decreasing MAPK1 levels. Conclusions: Coicis Semen has a remarkable anti-OA effect. Precisely, the major components of Coicis Semen, including stigmasterol, sitosterol alpha1, sitosterol, and 2-monoolein, specifically inhibit MAPK1, ESR1, and PPARA to reduce the inflammatory response and oxidative damage in OA.     

Non-radioactive in situ detection of mRNA in ES cell-derived cardiomyocytes and in the developing heart

Non-radioactive in situ hybridisation is an excellent method to visualise mRNA molecules within their topographical context. Recently we have reported a new non-radioactive in situ hybridisation procedure on tissue sections that is essentially based on the whole mount in situ hybridisation procedure. This method is superior in spatial resolution and sensitivity compared to the radioactive in situ hybridisation procedure. Generally, low levels of gene expression, such as found with the developmental onset of gene expression and in differentiating embryonic stem cells, are difficult to detect by in situ hybridisation. Here an application of the protocol is presented which is based on tyramide signal amplification, which enables the detection of very low abundant mRNAs. The significance of this method is two-fold: (1) the molecular phenotype of embryonic stem cell-derived cardiomyocytes can be examined at the cellular level with high sensitivity, and (2) the number of cells that express the gene of interest can be assessed.     

XPN-FMS: A CAD tool for FMS modeling, analysis, animation, and simulation using Petri nets and X window

We propose a CAD tool, XPN-FMS, which is primarily based on a unique Petri net (PN) synthesis method, called the knitting technique, developed by the authors. Petri net theory has been applied to specification, validation, performance analysis, control code generation, and simulation for manufacturing systems. The analysis of flexible manufacturing systems (FMSs) based on PNs suffers from the complexity problem of reachability analysis (Peterson, 1981). CAD tools are urgently needed. There is no existing CAD tool for FMSs as comprehensive as XPN-FMS, in the sense that the latter integrates the functions of drawing, analysis, reduction (Chao and Wang, 1992; Murata and Koh, 1980), synthesis, property queries, and animation of FMS operations in one software package. Using the X window graphical interface and animation, XPN-FMS makes the modeling and analysis of an FMS visualizable and easy to understand and manipulate. It lets a user draw the factory layout of an FMS on the screen of a monitor using the supplied tools. A corresponding PN model can also be drawn on the monitor screen. XPN-FMS can animate and simulate the overall operating process of the FMS. It is useful for FMS specification, validation, and exploration of different design alternatives, status monitoring, and control. Using XPN-FMS with various inputs and comparing the resulting outputs, the user can determine how to improve efficiency, reduce cost, and pinpoint bottlenecks. For the PN models of FMSs that are decision free, we extend the theory and algorithm of a unique matrix-based method (Chao and Wang, 1993b) to search for subcritical loops (including types A and B) and to support scheduling and dealing with transition periods. XPN-FMS implements this extended method to find the minimum cycle time, critical loop, subcritical loops, next critical loop, and scheduling ranges to avoid the transient period for static scheduling. This is implemented in XPN-FMS for the input sequence control.This project is partially funded by NJIT's Separately Budgeted Research Program. Portions of this article were presented in Chao, Chen, Wang, and Zhou (1992),Proceedings of the 1992 IEEE International Conference on Systems, Man, and Cybernetics, Chicago, Illinois, October 1992. The former name of the first author, which has appeared in some of his earlier publications, was Yuh Yaw.     

卫星飞轮隔振与吸振联合减振系统设计

分析了抑制卫星飞轮振动的方法,提出了隔振为主、吸振为辅的联合减振方案。研究了会聚式隔振系统参数对其减振性能的影响;针对会聚式隔振系统存在的不足,提出了圆周分布式吸振方法和相应的吸振系统,分析表明会聚式隔振系统在4个方向的隔振效率达90%以上。根据会聚式隔振系统在X平动方向隔振效率较低的问题,建立了会聚式隔振和圆周分布式吸振的联合减振系统仿真模型。仿真结果表明,联合减振系统较单纯的隔振系统在X平动方向减振效率提高近50%,且不改变其他方向的减振性能。因此,联合减振设计方法合理可行,适用于卫星飞轮等主要振源的振动抑制,并为飞轮联合减振系统的工程化设计提供了理论支持。     

Role of necroptosis in spinal cord injury and its therapeutic implications

Spinal cord injury (SCI), a complex neurological disorder, triggers a series of devastating neuropathological events such as ischemia, oxidative stress, inflammatory events, neuronal apoptosis, and motor dysfunction. However, the classical necrosome, which consists of receptor-interacting protein (RIP)1, RIP3, and mixed-lineage kinase domain-like protein, is believed to control a novel type of programmed cell death called necroptosis, through tumour necrosis factor-alpha/tumour necrosis factor receptor-1 signalling or other stimuli. Several studies reported that necroptosis plays an important role in neural cell damage, release of intracellular pro-inflammatory factors, lysosomal dysfunction and endoplasmic reticulum stress. Recent research indicates that necroptosis is crucial to the pathophysiology of a number of neurological disorders and SCIs. In our review, we summarize the potential role of programmed cell death regulated by necroptosis in SCI based on its molecular and pathophysiological mechanisms. We also summarize the targets of several necroptosis pathways, which provide a more reliable reference for the treatment of SCI.     

How does FtsZ’s treadmilling help bacterial cells divide?

Most bacteria assemble a ring-like macromolecular machinery scaffolded by the essential cytoskeletal protein FtsZ for cell division. Studies have broadly explored how FtsZ could polymerize at the correct place and time. Recently, the FtsZ-ring was found to exhibit dynamic treadmilling along the circumference of the division site, driven by GTP hydrolysis. This apparently directional motion of FtsZ seems to drive the movement of septal cell wall synthesis enzymes and to play an important role in modulating cell envelope constriction and septum morphogenesis. However, the relationship between FtsZ’s treadmilling dynamics and cell wall synthesis varies in different bacteria. More importantly, the biophysical and molecular mechanisms governing these dynamic processes are unclear. In this viewpoint, we will focus on some new and exciting studies surrounding this topic and discuss potential mechanisms that underlie how FtsZ’s treadmilling dynamics might regulate septal cell wall synthesis and cell division.