Assessing brain involvement in Fabry disease with deep learning and the brain-age paradigm.

While neurological manifestations are core features of Fabry disease (FD), quantitative neuroimaging biomarkers allowing to measure brain involvement are lacking. We used deep learning and the brain-age paradigm to assess whether FD patients' brains appear older than normal and to validate brain-predicted age difference (brain-PAD) as a possible disease severity biomarker. MRI scans of FD patients and healthy controls (HCs) from a single Institution were, retrospectively, studied. The Fabry stabilization index (FASTEX) was recorded as a measure of disease severity. Using minimally preprocessed 3D T1-weighted brain scans of healthy subjects from eight publicly available sources (N = 2160; mean age = 33 years [range 4-86]), we trained a model predicting chronological age based on a DenseNet architecture and used it to generate brain-age predictions in the internal cohort. Within a linear modeling framework, brain-PAD was tested for age/sex-adjusted associations with diagnostic group (FD vs. HC), FASTEX score, and both global and voxel-level neuroimaging measures. We studied 52 FD patients (40.6 ± 12.6 years; 28F) and 58 HC (38.4 ± 13.4 years; 28F). The brain-age model achieved accurate out-of-sample performance (mean absolute error = 4.01 years, R2 = .90). FD patients had significantly higher brain-PAD than HC (estimated marginal means: 3.1 vs. -0.1, p = .01). Brain-PAD was associated with FASTEX score (B = 0.10, p = .02), brain parenchymal fraction (B = -153.50, p = .001), white matter hyperintensities load (B = 0.85, p = .01), and tissue volume reduction throughout the brain. We demonstrated that FD patients' brains appear older than normal. Brain-PAD correlates with FD-related multi-organ damage and is influenced by both global brain volume and white matter hyperintensities, offering a comprehensive biomarker of (neurological) disease severity.

Evaluation of automated staple sizes on gastric layer incorporation and intraluminal pressure for partial gastrectomy closure in an ex vivo canine model.

OBJECTIVE: To evaluate the performance of automated staple sizes on a cadaveric canine partial gastrectomy model. METHODS: Stomachs were transected through the gastric body axis and randomly allocated to two closure groups: Group B, thoracoabdominal (TA) stapler 3.5 mm staple cartridge (blue); Group G, TA stapler 4.8 mm staple cartridge (green). After construct completion, leak testing was performed for both groups and compared. Initial leakage pressure (ILP), maximal leakage pressure (MLP) and leakage location were recorded. Staple lines were evaluated by direct observation and fluoroscopy to assess sub-mucosal layer incorporation and staple conformation. Staple shape was classified as optimal or suboptimal. Significance was set at p less than 0.5. RESULTS: Following gastrectomy, the mean double gastric wall thickness was 7.82 ± 2.05 mm at the gastric body. Mean ILP was significantly lower in groups G (17.13 ± 1.19 mmHg) compared to group B (50.46 ± 6.03 mmHg, p = 0.0013). Similarly, mean MLP was significantly lower in group G (21.41 ± 1.39 mmHg) compared to group B (64.61 ± 10.21 mmHg, p < 0.0001). Although group G had higher percentage of B-shaped staple formation compared to group B, this was not significant (group G; 92.38%, group B; 54.56%; p = 0.054). CONCLUSION: The 3.5 mm TA staple cartridge (blue) achieved superior bursting pressures compared with the 4.8 mm TA staple cartridge (blue) for the closure of a canine partial gastrectomy model. Both staple sizes incorporated all gastric layers. No differences were noticed in optimal staple conformation between groups. In vivo investigation is warranted to evaluate the use of different staple sizes on gastric tissue perfusion, successful healing and post-operative stasis and dehiscence.

A quantified comparison of cortical atlases on the basis of trait morphometricity.

BACKGROUND: Many different brain atlases exist that subdivide the human cortex into dozens or hundreds of regions-of-interest (ROIs). Inconsistency across studies using one or another cortical atlas may contribute to the replication crisis across the neurosciences. METHODS: Here, we provide a quantitative comparison between seven popular cortical atlases (Yeo, Desikan-Killiany, Destrieux, Jülich-Brain, Gordon, Glasser, Schaefer) and vertex-wise measures (thickness, surface area, and volume), to determine which parcellation retains the most information in the analysis of behavioural traits (incl. age, sex, body mass index, and cognitive ability) in the UK Biobank sample (N∼40,000). We use linear mixed models to compare whole-brain morphometricity; the proportion of trait variance accounted for when using a given atlas. RESULTS: Commonly-used atlases resulted in a considerable loss of information compared to vertex-wise representations of cortical structure. Morphometricity increased linearly as a function of the log-number of ROIs included in an atlas, indicating atlas-based analyses miss many true associations and yield limited prediction accuracy. Likelihood ratio tests revealed that low-dimensional atlases accounted for unique trait variance rather than variance common between atlases, suggesting that previous studies likely returned atlas-specific findings. Finally, we found that the commonly-used atlases yielded brain-behaviour associations on par with those obtained with random parcellations, where specific region boundaries were randomly generated. DISCUSSION: Our findings motivate future structural neuroimaging studies to favour vertex-wise cortical representations over coarser atlases, or to consider repeating analyses across multiple atlases, should the use of low-dimensional atlases be necessary. The insights uncovered here imply that cortical atlas choices likely contribute to the lack of reproducibility in ROI-based studies.

NADH/NAD+ binding and linked tetrameric assembly of the oncogenic transcription factors CtBP1 and CtBP2.

The activation of oncogenic C-terminal binding Protein (CtBP) transcriptional activity is coupled with NAD(H) binding and homo-oligomeric assembly, although the level of CtBP assembly and nucleotide binding affinity continues to be debated. Here, we apply biophysical techniques to address these fundamental issues for CtBP1 and CtBP2. Our ultracentrifugation results unambiguously demonstrate that CtBP assembles into tetramers in the presence of saturating NAD+ or NADH with tetramer to dimer dissociation constants about 100 nm. Isothermal titration calorimetry measurements of NAD(H) binding to CtBP show dissociation constants between 30 and 500 nm, depending on the nucleotide and paralog. Given cellular levels of NAD+ , CtBP is likely to be fully saturated with NAD under physiological concentrations suggesting that CtBP is unable to act as a sensor for NADH levels.

Cannabis use in patients with early psychosis is associated with alterations in putamen and thalamic shape.

Around half of patients with early psychosis have a history of cannabis use. We aimed to determine if there are neurobiological differences in these the subgroups of persons with psychosis with and without a history of cannabis use. We expected to see regional deflations in hippocampus as a neurotoxic effect and regional inflations in striatal regions implicated in addictive processes. Volumetric, T1w MRIs were acquired from people with a diagnosis psychosis with (PwP + C = 28) or without (PwP - C = 26) a history of cannabis use; and Controls with (C + C = 16) or without (C - C = 22) cannabis use. We undertook vertex-based shape analysis of the brainstem, amygdala, hippocampus, globus pallidus, nucleus accumbens, caudate, putamen, thalamus using FSL FIRST. Clusters were defined through Threshold Free Cluster Enhancement and Family Wise Error was set at p < .05. We adjusted analyses for age, sex, tobacco and alcohol use. The putamen (bilaterally) and the right thalamus showed regional enlargement in PwP + C versus PwP - C. There were no areas of regional deflation. There were no significant differences between C + C and C - C. Cannabis use in participants with psychosis is associated with morphological alterations in subcortical structures. Putamen and thalamic enlargement may be related to compulsivity in patients with a history of cannabis use.

The My Cancer Genome clinical trial data model and trial curation workflow.

OBJECTIVE: As clinical trials evolve in complexity, clinical trial data models that can capture relevant trial data in meaningful, structured annotations and computable forms are needed to support accrual. MATERIAL AND METHODS: We have developed a clinical trial information model, curation information system, and a standard operating procedure for consistent and accurate annotation of cancer clinical trials. Clinical trial documents are pulled into the curation system from publicly available sources. Using a web-based interface, a curator creates structured assertions related to disease-biomarker eligibility criteria, therapeutic context, and treatment cohorts by leveraging our data model features. These structured assertions are published on the My Cancer Genome (MCG) website. RESULTS: To date, over 5000 oncology trials have been manually curated. All trial assertion data are available for public view on the MCG website. Querying our structured knowledge base, we performed a landscape analysis to assess the top diseases, biomarker alterations, and drugs featured across all cancer trials. DISCUSSION: Beyond curating commonly captured elements, such as disease and biomarker eligibility criteria, we have expanded our model to support the curation of trial interventions and therapeutic context (ie, neoadjuvant, metastatic, etc.), and the respective biomarker-disease treatment cohorts. To the best of our knowledge, this is the first effort to capture these fields in a structured format. CONCLUSION: This paper makes a significant contribution to the field of biomedical informatics and knowledge dissemination for precision oncology via the MCG website. KEY WORDS: knowledge representation, My Cancer Genome, precision oncology, knowledge curation, cancer informatics, clinical trial data model.

Multimodality neuroimaging brain-age in UK biobank: relationship to biomedical, lifestyle, and cognitive factors.

The brain-age paradigm is proving increasingly useful for exploring aging-related disease and can predict important future health outcomes. Most brain-age research uses structural neuroimaging to index brain volume. However, aging affects multiple aspects of brain structure and function, which can be examined using multimodality neuroimaging. Using UK Biobank, brain-age was modeled in n = 2205 healthy people with T1-weighted MRI, T2-FLAIR, T2∗, diffusion-MRI, task fMRI, and resting-state fMRI. In a held-out healthy validation set (n = 520), chronological age was accurately predicted (r = 0.78, mean absolute error = 3.55 years) using LASSO regression, higher than using any modality separately. Thirty-four neuroimaging phenotypes were deemed informative by the regression (after bootstrapping); predominantly gray-matter volume and white-matter microstructure measures. When applied to new individuals from UK Biobank (n = 14,701), significant associations with multimodality brain-predicted age difference (brain-PAD) were found for stroke history, diabetes diagnosis, smoking, alcohol intake and some, but not all, cognitive measures (corrected p < 0.05). Multimodality neuroimaging can improve brain-age prediction, and derived brain-PAD values are sensitive to biomedical and lifestyle factors that negatively impact brain and cognitive health.

Regulation of Protein Kinase R by Epstein-Barr Virus EBER1 RNA.

Protein kinase R (PKR) is a key antiviral component of the innate immune pathway and is activated by viral double-stranded RNAs (dsRNAs). Adenovirus-associated RNA 1 (VAI) is an abundant, noncoding viral RNA that functions as a decoy by binding PKR but not inducing activation, thereby inhibiting the antiviral response. In VAI, coaxial stacking produces an extended helix that mediates high-affinity PKR binding but is too short to result in activation. Like adenovirus, Epstein-Barr virus produces high concentrations of a noncoding RNA, EBER1. Here, we compare interactions of PKR with VAI and EBER1 and present a structural model of EBER1. Both RNAs function as inhibitors of dsRNA-mediated PKR activation. However, EBER1 weakly activates PKR whereas VAI does not. PKR binds EBER1 more weakly than VAI. Assays at physiological ion concentrations indicate that both RNAs can accommodate two PKR monomers and induce PKR dimerization. A structural model of EBER1 was obtained using constraints derived from chemical structure probing and small-angle X-ray scattering experiments. The central stem of EBER1 coaxially stacks with stem loop 4 and stem loop 1 to form an extended RNA duplex of ∼32 bp that binds PKR and promotes activation. Our observations that EBER1 binds PKR much more weakly than VAI and exhibits weak PKR activation suggest that EBER1 is less well suited to function as an RNA decoy.

Oligomerization of RIG-I and MDA5 2CARD domains.

The innate immune system is the first line of defense against invading pathogens. The retinoic acid-inducible gene I (RIG-I) like receptors (RLRs), RIG-I and melanoma differentiation-associated protein 5 (MDA5), are critical for host recognition of viral RNAs. These receptors contain a pair of N-terminal tandem caspase activation and recruitment domains (2CARD), an SF2 helicase core domain, and a C-terminal regulatory domain. Upon RLR activation, 2CARD associates with the CARD domain of MAVS, leading to the oligomerization of MAVS, downstream signaling and interferon induction. Unanchored K63-linked polyubiquitin chains (polyUb) interacts with the 2CARD domain, and in the case of RIG-I, induce tetramer formation. However, the nature of the MDA5 2CARD signaling complex is not known. We have used sedimentation velocity analytical ultracentrifugation to compare MDA5 2CARD and RIG-I 2CARD binding to polyUb and to characterize the assembly of MDA5 2CARD oligomers in the absence of polyUb. Multi-signal sedimentation velocity analysis indicates that Ub4 binds to RIG-I 2CARD with a 3:4 stoichiometry and cooperatively induces formation of an RIG-I 2CARD tetramer. In contrast, Ub4 and Ub7 interact with MDA5 2CARD weakly and form complexes with 1:1 and 2:1 stoichiometries but do not induce 2CARD oligomerization. In the absence of polyUb, MDA5 2CARD self-associates to forms large oligomers in a concentration-dependent manner. Thus, RIG-I and MDA5 2CARD assembly processes are distinct. MDA5 2CARD concentration-dependent self-association, rather than polyUb binding, drives oligomerization and MDA5 2CARD forms oligomers larger than tetramer. We propose a mechanism where MDA5 2CARD oligomers, rather than a stable tetramer, function to nucleate MAVS polymerization.

Opportunistic pulse checks in primary care to improve recognition of atrial fibrillation: a retrospective analysis of electronic patient records.

BACKGROUND: Atrial fibrillation (AF) is an important and modifiable risk factor for stroke. Earlier identification may reduce stroke-related morbidity and mortality. Trial evidence shows that opportunistic pulse regularity checks in individuals aged ≥65 years increases detection of AF. However, this is not currently recommended by the National Screening Programme or implemented by most clinical commissioning groups (CCGs). AIM: To evaluate the impact of a systematic programme to promote pulse regularity checks, the programme's uptake in general practice, and the prevalence of AF. DESIGN AND SETTING: Retrospective analysis of electronic primary care patient records in three east London CCGs (City and Hackney, Newham, and Tower Hamlets) over 10 years. METHOD: Rates of pulse regularity checks and prevalence of AF in individuals aged ≥65 years were compared from the pre-intervention period, 2007-2011, to the post-intervention period, 2012-2017. RESULTS: Across the three CCGs, rates of pulse regularity checks increased from a mean of 7.3% pre-intervention to 66.4% post-intervention, achieving 93.1% (n = 58 722) in the final year. Age-standardised prevalence of AF in individuals aged ≥65 years increased significantly from a pre-intervention mean of 61.4/1000 to a post-intervention mean of 64.5/1000. There was a significant increase in a post-intervention trend to a final-year mean of 67.3/1000: an improvement of 9.6% (5.9/1000) with 790 additional new cases identified. CONCLUSION: Organisational alignment, standardised data entry, peer-performance dashboards, and financial incentives rapidly and generally increased opportunistic screening with pulse regularity checks. This was associated with a significant increase in detection and prevalence of AF and is of public health importance.

No Evidence for Accelerated Aging-Related Brain Pathology in Treated Human Immunodeficiency Virus: Longitudinal Neuroimaging Results From the Comorbidity in Relation to AIDS (COBRA) Project.

Background: Despite successful antiretroviral therapy, people living with human immunodeficiency virus (PLWH) experience higher rates of age-related morbidity, including abnormal brain structure, brain function, and cognitive impairment. This has raised concerns that PLWH may experience accelerated aging-related brain pathology. Methods: We performed a multicenter longitudinal study of 134 virologically suppressed PLWH (median age, 56.0 years) and 79 demographically similar human immunodeficiency virus (HIV)-negative controls (median age, 57.2 years). To measure cognitive performance and brain pathology, we conducted detailed neuropsychological assessments and multimodality neuroimaging (T1-weighted, T2-weighted, diffusion magnetic resonance imaging [MRI], resting-state functional MRI, spectroscopy, arterial spin labeling) at baseline and at 2 years. Group differences in rates of change were assessed using linear mixed effects models. Results: One hundred twenty-three PLWH and 78 HIV-negative controls completed longitudinal assessments (median interval, 1.97 years). There were no differences between PLWH and HIV-negative controls in age, sex, years of education, smoking or alcohol use. At baseline, PLWH had poorer global cognitive performance (P < .01), lower gray matter volume (P = .04), higher white matter hyperintensity load (P = .02), abnormal white matter microstructure (P < .005), and greater brain-predicted age difference (P = .01). Longitudinally, there were no significant differences in rates of change in any neuroimaging measure between PLWH and HIV-negative controls (P > .1). Cognitive performance was longitudinally stable in both groups. Conclusions: We found no evidence that middle-aged PLWH, when receiving successful treatment, are at increased risk of accelerated aging-related brain changes or cognitive decline over 2 years.

Structural Brain Abnormalities in Successfully Treated HIV Infection: Associations With Disease and Cerebrospinal Fluid Biomarkers.

Background: Brain structural abnormalities have been reported in persons living with human immunodeficiency virus (HIV; PLWH) who are receiving suppressive combination antiretroviral therapy (cART), but their pathophysiology remains unclear. Methods: We investigated factors associated with brain tissue volumes and white matter microstructure (fractional anisotropy) in 134 PLWH receiving suppressive cART and 79 comparable HIV-negative controls, aged ≥45 years, from the Comorbidity in Relation to AIDS cohort, using multimodal neuroimaging and cerebrospinal fluid biomarkers. Results: Compared with controls, PLWH had lower gray matter volumes (-13.7 mL; 95% confidence interval, -25.1 to -2.2) and fractional anisotropy (-0.0073; 95% confidence interval, -.012 to -.0024), with the largest differences observed in those with prior clinical AIDS. Hypertension and the soluble CD14 concentration in cerebrospinal fluid were associated with lower fractional anisotropy. These associations were independent of HIV serostatus (Pinteraction = .32 and Pinteraction = .59, respectively) and did not explain the greater abnormalities in brain structure in relation to HIV infection. Conclusions: The presence of lower gray matter volumes and more white matter microstructural abnormalities in well-treated PLWH partly reflect a combination of historical effects of AIDS, as well as the more general influence of systemic factors, such as hypertension and ongoing neuroinflammation. Additional mechanisms explaining the accentuation of brain structure abnormalities in treated HIV infection remain to be identified.

Increased brain-predicted aging in treated HIV disease.

OBJECTIVE: To establish whether HIV disease is associated with abnormal levels of age-related brain atrophy, by estimating apparent brain age using neuroimaging and exploring whether these estimates related to HIV status, age, cognitive performance, and HIV-related clinical parameters. METHODS: A large sample of virologically suppressed HIV-positive adults (n = 162, age 45-82 years) and highly comparable HIV-negative controls (n = 105) were recruited as part of the Comorbidity in Relation to AIDS (COBRA) collaboration. Using T1-weighted MRI scans, a machine-learning model of healthy brain aging was defined in an independent cohort (n = 2,001, aged 18-90 years). Neuroimaging data from HIV-positive and HIV-negative individuals were then used to estimate brain-predicted age; then brain-predicted age difference (brain-PAD = brain-predicted brain age - chronological age) scores were calculated. Neuropsychological and clinical assessments were also carried out. RESULTS: HIV-positive individuals had greater brain-PAD score (mean ± SD 2.15 ± 7.79 years) compared to HIV-negative individuals (-0.87 ± 8.40 years; b = 3.48, p < 0.01). Increased brain-PAD score was associated with decreased performance in multiple cognitive domains (information processing speed, executive function, memory) and general cognitive performance across all participants. Brain-PAD score was not associated with age, duration of HIV infection, or other HIV-related measures. CONCLUSION: Increased apparent brain aging, predicted using neuroimaging, was observed in HIV-positive adults, despite effective viral suppression. Furthermore, the magnitude of increased apparent brain aging related to cognitive deficits. However, predicted brain age difference did not correlate with chronological age or duration of HIV infection, suggesting that HIV disease may accentuate rather than accelerate brain aging.

Loss of Interneuron-Derived Collagen XIX Leads to a Reduction in Perineuronal Nets in the Mammalian Telencephalon.

Perineuronal nets (PNNs) are lattice-like supramolecular assemblies of extracellular glycoproteins that surround subsets of neuronal cell bodies in the mammalian telencephalon. PNNs emerge at the end of the critical period of brain development, limit neuronal plasticity in the adult brain, and are lost in a variety of complex brain disorders diseases, including schizophrenia. The link between PNNs and schizophrenia led us to question whether neuronally expressed extracellular matrix (ECM) molecules associated with schizophrenia contribute to the assembly of these specialized supramolecular ECM assemblies. We focused on collagen XIX-a minor, nonfibrillar collagen expressed by subsets of telencephalic interneurons. Genetic alterations in the region encoding collagen XIX have been associated with familial schizophrenia, and loss of this collagen in mice results in altered inhibitory synapses, seizures, and the acquisition of schizophrenia-related behaviors. Here, we demonstrate that loss of collagen XIX also results in a reduction of telencephalic PNNs. Loss of PNNs was accompanied with reduced levels of aggrecan (Acan), a major component of PNNs. Despite reduced levels of PNN constituents in collagen XIX-deficient mice ( col19a1-/-), we failed to detect reduced expression of genes encoding these ECM molecules. Instead, we discovered a widespread upregulation of extracellular proteases capable of cleaving Acan and other PNN constituents in col19a1-/- brains. Taken together, these results suggest a mechanism by which the loss of collagen XIX speeds PNN degradation and they identify a novel mechanism by which the loss of collagen XIX may contribute to complex brain disorders.

New Murine Model of Early Onset Autoimmune Thyroid Disease/Hypothyroidism and Autoimmune Exocrinopathy of the Salivary Gland.

Sixty to seventy percent of IFN-γ(-/-) NOD.H-2h4 mice given sodium iodide (NaI)-supplemented water develop a slow onset autoimmune thyroid disease, characterized by thyrocyte epithelial cell (TEC) hyperplasia and proliferation (H/P). TEC H/P develops much earlier in CD28(-/-) mice and nearly 100% (both sexes) have severe TEC H/P at 4 mo of age. Without NaI supplementation, 50% of 5- to 6-mo-old CD28(-/-)IFN-γ(-/-) mice develop severe TEC H/P, and 2-3 wk of NaI is sufficient for optimal development of severe TEC H/P. Mice with severe TEC H/P are hypothyroid, and normalization of serum thyroxine levels does not reduce TEC H/P. Activated CD4(+) T cells are sufficient to transfer TEC H/P to SCID recipients. Thyroids of mice with TEC H/P have infiltrating T cells and expanded numbers of proliferating thyrocytes that highly express CD40. CD40 facilitates, but is not required for, development of severe TEC H/P, as CD40(-/-)IFN-γ(-/-)CD28(-/-) mice develop severe TEC H/P. Accelerated development of TEC H/P in IFN-γ(-/-)CD28(-/-) mice is a result of reduced regulatory T cell (Treg) numbers, as CD28(-/-) mice have significantly fewer Tregs, and transfer of CD28(+) Tregs inhibits TEC H/P. Essentially all female IFN-γ(-/-)CD28(-/-) NOD.H-2h4 mice have substantial lymphocytic infiltration of salivary glands and reduced salivary flow by 6 mo of age, thereby providing an excellent new model of autoimmune exocrinopathy of the salivary gland. This is one of very few models where autoimmune thyroid disease and hypothyroidism develop in most mice by 4 mo of age. This model will be useful for studying the effects of hypothyroidism on multiple organ systems.

Solvent Properties of Water in Aqueous Solutions of Elastin-Like Polypeptide.

The phase-transition temperatures of an elastin-like polypeptide (ELP) with the (GVGVP)40 sequence and solvent dipolarity/polarizability, hydrogen-bond donor acidity, and hydrogen-bond acceptor basicity in its aqueous solutions were quantified in the absence and presence of different salts (Na2SO4, NaCl, NaClO4, and NaSCN) and various osmolytes (sucrose, sorbitol, trehalose, and trimethylamine N-oxide (TMAO)). All osmolytes decreased the ELP phase-transition temperature, whereas NaCl and Na2SO4 decreased, and NaSCN and NaClO4 increased it. The determined phase-transition temperatures may be described as a linear combination of the solvent's dipolarity/polarizability and hydrogen-bond donor acidity. The linear relationship established for the phase-transition temperature in the presence of salts differs quantitatively from that in the presence of osmolytes, in agreement with different (direct and indirect) mechanisms of the influence of salts and osmolytes on the ELP phase-transition temperature.

Multifunctional nanoparticles for use in theranostic applications.

Theranostics is a promising field that combines therapeutics and diagnostics into single multifunctional formulations. This field is driven by advancements in nanoparticle systems capable of providing the necessary functionalities. By utilizing these powerful nanomedicines, the concept of personalized medicine can be realized by tailoring treatment strategies to the individual. This review gives a brief overview of the components of a theranostic system and the challenges that designing truly multifunctional nanoparticles present. Considerations when choosing a class of nanoparticle include the size, shape, charge, and surface chemistry, while classes of nanoparticles discussed are polymers, liposomes, dendrimers, and polymeric micelles. Targeting to disease states can be achieved either through passive or active targeting which uses specific ligands to target receptors that are overexpressed in tumors and common targeting elements are presented. To image the interactions with disease states, contrast agents are included in the nanoparticle formulation. Imaging options include optical imaging techniques, computed tomography, nuclear based, and magnetic resonance imaging. The interplay between all of these components needs to be carefully considered when designing a theranostic system.

Structural analysis of adenovirus VAI RNA defines the mechanism of inhibition of PKR.

Protein kinase R (PKR) is activated by dsRNA produced during virus replication and plays a major role in the innate immunity response to virus infection. In response, viruses have evolved multiple strategies to evade PKR. Adenovirus virus-associated RNA-I (VAI) is a short, noncoding transcript that functions as an RNA decoy to sequester PKR in an inactive state. VAI consists of an apical stem-loop, a highly structured central domain, and a terminal stem. Chemical probing and mutagenesis demonstrate that the central domain is stabilized by a pseudoknot. A structural model of VAI was obtained from constraints derived from chemical probing and small angle x-ray scattering (SAXS) measurements. VAI adopts a flat, extended conformation with the apical and terminal stems emanating from a protuberance in the center. This model reveals how the apical stem and central domain assemble to produce an extended duplex that is precisely tuned to bind a single PKR monomer with high affinity, thereby inhibiting activation of PKR by viral dsRNA.

Analysis of SecA dimerization in solution.

The Sec pathway mediates translocation of protein across the inner membrane of bacteria. SecA is a motor protein that drives translocation of preprotein through the SecYEG channel. SecA reversibly dimerizes under physiological conditions, but different dimer interfaces have been observed in SecA crystal structures. Here, we have used biophysical approaches to address the nature of the SecA dimer that exists in solution. We have taken advantage of the extreme salt sensitivity of SecA dimerization to compare the rates of hydrogen-deuterium exchange of the monomer and dimer and have analyzed the effects of single-alanine substitutions on dimerization affinity. Our results support the antiparallel dimer arrangement observed in one of the crystal structures of Bacillus subtilis SecA. Additional residues lying within the preprotein binding domain and the C-terminus are also protected from exchange upon dimerization, indicating linkage to a conformational transition of the preprotein binding domain from an open to a closed state. In agreement with this interpretation, normal mode analysis demonstrates that the SecA dimer interface influences the global dynamics of SecA such that dimerization stabilizes the closed conformation.

Domain interactions in adenovirus VAI RNA mediate high-affinity PKR binding.

Protein kinase R (PKR) is a component of the innate immunity antiviral pathway. PKR is activated upon binding to double-stranded RNA (dsRNA) to undergo dimerization and autophosphorylation. Adenovirus-associated RNA I (VAI) is a short, non-coding transcript whose major function is to inhibit the activity of PKR. VAI contains three domains: an apical stem-loop, a highly structured central domain, and a terminal stem. Previous studies have localized PKR binding to the apical stem and to the central domain. However, the molecular mechanism for inhibition of PKR is not known. We have characterized the stoichiometry and affinity of PKR binding to VAI and several domain constructs using analytical ultracentrifugation and correlated VAI binding and PKR inhibition. Although PKR binding to simple dsRNAs is not regulated by divalent ion, analysis of the interaction of the isolated dsRNA binding domain with VAI reveals that the binding affinity is enhanced by divalent ion. Dissection of VAI into its constituent domains indicates that none of the isolated domains retains the PKR binding affinity or inhibitory potency of the full-length RNA. PKR is capable of binding the isolated terminal stem, but deletion of this domain from VAI does not affect PKR binding or inhibition. These results indicate that both the apical stem and the central domain are required to form a high-affinity PKR binding site. Our data support a model whereby VAI functions as a PKR inhibitor because it binds a monomer tightly but does not facilitate dimerization.