Mental Stress-Induced Change in Plasma Stromal Cell-Derived Factor-1 and Adverse Cardiovascular Outcomes: A Cohort Study.

Background: Acute psychological stress can provoke mental stress-induced myocardial ischemia (MSIMI) in coronary artery disease (CAD). Stromal cell-derived factor 1 (SDF1) is released in response to hypoxia, and higher levels of SDF1 are associated with adverse outcomes. We examined whether an increase in SDF1 level in response to mental stress predicts adverse outcomes in CAD patients. Methods: A total of 554 patients with stable CAD (mean age 63 years; 76% male; 26% Black) underwent standardized mental stress testing. Plasma SDF1 levels were measured at rest and 90 minutes after mental stress, and MSIMI was evaluated by 99mTc-sestamibi perfusion imaging. Participants were followed for 5 years for the primary endpoint of composite of death and myocardial infarction (MI) and the secondary endpoint of composite of death, MI, and heart failure hospitalization. Cox hazard models were used to assess the association between SDF1 change and incident adverse events. Results: Mean (standard deviation) SDF1 change with mental stress was +56.0 (230) pg/mL. During follow-up, a rise of 1 standard deviation in SDF1 with mental stress was associated with a 32% higher risk for the primary endpoint of death and MI (95% confidence interval, 6%-64%), independent of the resting SDF1 level, demographic and clinical risk factors, and presence of ischemia. A rise of 1 standard deviation in SDF1 was associated with a 33% (95% confidence interval, 11%-59%) increase in the risk for the secondary endpoint, independent of the resting SDF1 level, demographic, and clinical risk factors and presence of ischemia. Conclusions: An increase in SDF1 level in response to mental stress is associated with a higher risk of adverse events in stable CAD, independent of MSIMI.

Contexte: Un stress psychologique aigu peut provoquer une ischémie myocardique induite par le stress mental chez les patients atteints d'une coronaropathie. Le facteur dérivé des cellules stromales de type 1 (SDF1) est libéré en réponse à une hypoxie, et des taux accrus de SDF1 sont associés à des résultats défavorables. Nous avons examiné si une élévation des taux de SDF1 en réponse à un stress mental permettait de prévoir la survenue de résultats défavorables chez des patients atteints d'une coronaropathie. Méthodologie: Au total, 554 patients présentant une coronaropathie stable (âge moyen de 63 ans; 76 % d'hommes; 26 % de patients de race noire) ont subi une évaluation standardisée du stress mental. Les taux plasmatiques de SDF1 ont été mesurés au repos et 90 minutes après un stress mental, et l'ischémie myocardique induite par le stress mental a été évaluée par imagerie avec injection de Tc-99m sestamibi. Les participants ont fait l'objet d'un suivi pendant cinq ans afin de surveiller la survenue des événements constituant le paramètre d'évaluation principal composé (décès et infarctus du myocarde [IM]) et le paramètre d'évaluation secondaire composé (décès, IM et hospitalisation en raison d'une insuffisance cardiaque). Des modèles de Cox ont été utilisés pour évaluer le lien entre la modification des taux de SDF1 et les événements indésirables susceptibles de survenir. Résultats: La variation moyenne du taux de SDF1 (écart-type) associée au stress mental a été de +56,0 (230) pg/ml. Pendant le suivi, une augmentation de 1 écart-type du taux de SDF1 en raison du stress mental a été associée à un risque 32 % plus élevé de survenue de l'un des événements constituant le paramètre d'évaluation principal (décès et IM) [intervalle de confiance [IC] à 95 % : 6 % à 64 %], indépendamment du taux de SDF1 au repos, des caractéristiques démographiques, des facteurs de risque clinique et de la présence d'une ischémie. Une augmentation de 1 écart-type du taux de SDF1 a été associée à un risque 33 % plus élevé (IC à 95 % : 11 % à 59 %) de survenue de l'un des événements constituant le paramètre d'évaluation secondaire, indépendamment du taux de SDF1 au repos, des caractéristiques démographiques, des facteurs de risque clinique et de la présence d'une ischémie. Conclusions: Une augmentation du taux de SDF1 en réponse à un stress mental est associée à une augmentation du risque d'événements indésirables chez les patients atteints d'une coronaropathie stable, indépendamment de la présence d'une ischémie myocardique induite par le stress mental.

The research foundation for COVID-19 vaccine development.

The development of effective vaccines in <1 year to combat the spread of coronavirus disease 19 (COVID-19) is an example of particularly rapid progress in biomedicine. However, this was only made possible by decades of investment in scientific research. Many important research commentaries and reviews have been provided to describe the various contributions and scientific breakthroughs that led to the development of COVID-19 vaccines. In this work, we sought to complement those efforts by adding a systematic and quantitative study of the research foundations that led to these vaccines. Here, we analyzed citations from COVID-19 vaccine research articles to determine which scientific areas of study contributed the most to this research. Our findings revealed that coronavirus research was cited most often, and by a large margin. However, significant contributions were also seen from a diverse set of fields such as cancer, diabetes, and HIV/AIDS. In addition, we examined the publication history of the most prolific authors of COVID-19 vaccine research to determine their research expertise prior to the pandemic. Interestingly, although COVID-19 vaccine research relied most heavily on previous coronavirus work, we find that the most prolific authors on these publications most often had expertise in other areas including influenza, cancer, and HIV/AIDS. Finally, we used machine learning to identify and group together publications based on their major topic areas. This allowed us to elucidate the differences in citations between research areas. These findings highlight and quantify the relevance of prior research from a variety of scientific fields to the rapid development of a COVID-19 vaccine. This study also illustrates the importance of funding and sustaining a diverse research enterprise to facilitate a rapid response to future pandemics.

Levodopa responsive freezing of gait is associated with reduced norepinephrine transporter binding in Parkinson's disease.

BACKGROUND: Freezing of gait (FOG) is a major cause of falling in Parkinson's disease (PD) and can be responsive or unresponsive to levodopa. Pathophysiology is poorly understood. OBJECTIVE: To examine the link between noradrenergic systems, the development of FOG in PD and its responsiveness to levodopa. METHODS: We examined norepinephrine transporter (NET) binding via brain positron emission tomography (PET) to evaluate changes in NET density associated with FOG using the high affinity selective NET antagonist radioligand [11C]MeNER (2S,3S)(2-[α-(2-methoxyphenoxy)benzyl]morpholine) in 52 parkinsonian patients. We used a rigorous levodopa challenge paradigm to characterize PD patients as non-freezing (NO-FOG, N = 16), levodopa responsive freezing (OFF-FOG, N = 10), and levodopa-unresponsive freezing (ONOFF-FOG, N = 21), and also included a non-PD FOG group, primary progressive freezing of gait (PP-FOG, N = 5). RESULTS: Linear mixed models identified significant reductions in whole brain NET binding in the OFF-FOG group compared to the NO-FOG group (-16.8%, P = 0.021) and regionally in the frontal lobe, left and right thalamus, temporal lobe, and locus coeruleus, with the strongest effect in right thalamus (P = 0.038). Additional regions examined in a post hoc secondary analysis including the left and right amygdalae confirmed the contrast between OFF-FOG and NO-FOG (P = 0.003). A linear regression analysis identified an association between reduced NET binding in the right thalamus and more severe New FOG Questionnaire (N-FOG-Q) score only in the OFF-FOG group (P = 0.022). CONCLUSION: This is the first study to examine brain noradrenergic innervation using NET-PET in PD patients with and without FOG. Based on the normal regional distribution of noradrenergic innervation and pathological studies in the thalamus of PD patients, the implications of our findings suggest that noradrenergic limbic pathways may play a key role in OFF-FOG in PD. This finding could have implications for clinical subtyping of FOG as well as development of therapies.

Neural correlates of stress and leucocyte telomere length in patients with coronary artery disease.

BACKGROUND: Accelerated biological aging, as indicated by telomere shortening, is associated with CAD pathogenesis. In a cross-sectional study, we investigated neural correlates of acute psychological stress and short telomeres in patients with CAD. METHODS: Individuals with CAD (N = 168) underwent a validated mental stress protocol including public speaking and mental arithmetic. Imaging of the brain with [O-15] water and high-resolution positron emission tomography (HR-PET) was performed during mental stress and control conditions. Blood flow during stressful tasks (average of speech and arithmetic) and control tasks were assessed. Telomere length in peripheral leucocytes was measured by quantitative polymerase chain reaction and expressed as Telomere/Single Copy Gene (T/S) ratio. Voxel-wise regression models were constructed to assess the association between brain areas and activity during rest and mental stress after adjustments for demographic factors and clinical characteristics. RESULTS: The mean (SD) age of the sample was 62 (8) years, and 69% were men. Increased activation with mental stress in the lingual gyrus, cerebellum and superior and inferior frontal gyri were associated with reduced telomere length; 1.6 higher voxel activation of these areas was associated with 0.1 T/S-units reduction in telomere length (P < 0.005). Additionally, during neutral counting and speaking tasks, brain activity in the precentral, middle and superior frontal and middle temporal gyri was inversely associated with telomere length. Results remained consistent after adjustment for demographic and clinical risk factors. CONCLUSION: Increased stress-induced activity in brain areas mediating the stress response was associated with shortened telomere length in CAD patients.

Research productivity and collaboration of the NIH-funded HIV vaccine trials network: A bibliometric analysis.

The HIV Vaccine Trials Network (HVTN) is the world's largest publicly funded, multi-disciplinary international collaboration facilitating the development of vaccines to prevent HIV/AIDS and has conducted the vast majority of HIV/AIDS clinical trials since its inception in 1999. Although scientific findings from the program have been published in scholarly journals, the impact of a large scientific research network such as the HVTN on the HIV/AIDS vaccine field has not been assessed. This paper describes and elucidates the productivity, influence, and collaboration among HVTN researchers over the last two decades. Our analyses indicate that the HVTN has funded a large number of HIV/AIDS vaccine safety and efficacy clinical trials through a strong global network of clinical sites. In addition, several metrics indicate HVTN researchers also published original research articles that are influential in the HIV vaccine field. Scientific research collaboration is critically important in a complex and multidisciplinary field such as HIV vaccine development as it allows improved sharing of knowledge and expertise as well as the pooling of resources and data. We found that collaboration in the HIV vaccine field increased during this time period and collaboration among HVTN authors increased even more. Combining these productivity, influence, and collaboration metrics with research outcomes can provide a comprehensive assessment of large complex programs such as the HVTN.

Patterns of Prescribing Antiretroviral Therapy Upon Discharge to Psychiatry Inpatients With HIV/AIDS at a Large Urban Hospital.

BACKGROUND: HIV infection is more prevalent among people with severe mental illness (SMI) than in the general population. People with SMI may lack access to recommended antiretroviral therapy (ART), and inpatient psychiatric admissions may be opportunities to ensure that individuals receive recommended treatment. OBJECTIVE: To evaluate ART prescription patterns on an inpatient psychiatry service. METHODS: In this retrospective, observational study, patient and admission characteristics and ART prescriptions were obtained for 248 HIV-positive inpatients between 2006 and 2012. Receipt of any ART, any recommended ART regimen, and ART with potentially harmful adverse events and drug interactions were examined. General estimating equation models were used to evaluate prescription patterns in relation to patient and admission characteristics. RESULTS: ART was prescribed at 39% of discharges and increased by 51% during the study. Prescription was more common in admissions with an AIDS diagnosis and age greater than 29 years and less common in admissions associated with a psychotic diagnosis and shorter inpatient stays. When ART was prescribed, regimens were consistent with guideline recommendations 91% of the time. Prescription of potentially harmful regimens was limited. CONCLUSION AND RELEVANCE: In an acute inpatient psychiatry setting in an urban HIV/AIDS epicenter, where psychotic disorders and brief and involuntary admissions were the norm, guideline-recommended ART regimens were prescribed at almost 60% of discharges by the end of the study. Future studies should explore interventions to increase ART for high-risk subpopulations with SMI, including younger individuals or those with brief inpatient psychiatry hospitalizations.

Novel PET and Near Infrared Imaging Probes for the Specific Detection of Bacterial Infections Associated With Cardiac Devices.

OBJECTIVES: The aim of this study was to develop imaging agents to detect early stage infections in implantable cardiac devices. BACKGROUND: Bacteria ingest maltodextrins through the specific maltodextrin transporter. We developed probes conjugated with either a fluorescent dye (maltohexaose fluorescent dye probe [MDP]) or a F-18 (F18 fluoromaltohexaose) and determined their usefulness in a model of infections associated with implanted cardiac devices. METHODS: Stainless steel mock-ups of medical devices were implanted subcutaneously in rats. On post-operative day 4, animals were injected with either Staphylococcus aureus around the mock-ups to induce a relatively mild infection or oil of turpentine to induce noninfectious inflammation. Animals with a sterile implant were used as control subjects. On post-operative day 6, either the MDP or F18 fluoromaltohexaose was injected intravenously, and the animals were scanned with the appropriate imaging device. Additional positron emission tomography imaging studies were performed with F18-fluorodeoxyglucose as a comparison of the specificity of our probes (n = 5 to 9 per group). RESULTS: The accumulation of the MDP in the infected rats was significantly increased at 1 h after injection when compared with the control and noninfectious inflammation groups (intensity ratio 1.54 ± 0.07 vs. 1.26 ± 0.04 and 1.20 ± 0.05, respectively; p < 0.05) and persisted for more than 24 h. In positron emission tomography imaging, both F18 fluoromaltohexaose and F18 fluorodeoxyglucose significantly accumulated in the infected area 30 min after the injection (maximum standard uptake value ratio 4.43 ± 0.30 and 4.87 ± 0.28, respectively). In control rats, there was no accumulation of imaging probes near the device. In the noninfectious inflammation rats, no significant accumulation was observed with F18 fluoromaltohexaose, but F18 fluorodeoxyglucose accumulated in the mock-up area (maximum standard uptake value 2.53 ± 0.39 vs. 4.74 ± 0.46, respectively; p < 0.05). CONCLUSIONS: Our results indicate that maltohexaose-based imaging probes are potentially useful for the specific and sensitive diagnosis of infections associated with implantable cardiac devices.

HJURP antagonizes CENP-A mislocalization driven by the H3.3 chaperones HIRA and DAXX.

The centromere specific histone H3 variant CENP-A/CENH3 specifies where the kinetochore is formed in most eukaryotes. Despite tight regulation of CENP-A levels in normal cells, overexpression of CENP-A is a feature shared by various types of solid tumors and results in its mislocalization to non-centromeric DNA. How CENP-A is assembled ectopically and the consequences of this mislocalization remain topics of high interest. Here, we report that in human colon cancer cells, the H3.3 chaperones HIRA and DAXX promote ectopic CENP-A deposition. Moreover, the correct balance between levels of the centromeric chaperone HJURP and CENP-A is essential to preclude ectopic assembly by H3.3 chaperones. In addition, we find that ectopic localization can recruit kinetochore components, and correlates with mitotic defects and DNA damage in G1 phase. Finally, CENP-A occupancy at the 8q24 locus is also correlated with amplification and overexpression of the MYC gene within that locus. Overall, these data provide insights into the causes and consequences of histone variant mislocalization in human cancer cells.

Asterless is a Polo-like kinase 4 substrate that both activates and inhibits kinase activity depending on its phosphorylation state.

Centriole assembly initiates when Polo-like kinase 4 (Plk4) interacts with a centriole "targeting-factor." In Drosophila, Asterless/Asl (Cep152 in humans) fulfills the targeting role. Interestingly, Asl also regulates Plk4 levels. The N-terminus of Asl (Asl-A; amino acids 1-374) binds Plk4 and promotes Plk4 self-destruction, although it is unclear how this is achieved. Moreover, Plk4 phosphorylates the Cep152 N-terminus, but the functional consequence is unknown. Here, we show that Plk4 phosphorylates Asl and mapped 13 phospho-residues in Asl-A. Nonphosphorylatable alanine (13A) and phosphomimetic (13PM) mutants did not alter Asl function, presumably because of the dominant role of the Asl C-terminus in Plk4 stabilization and centriolar targeting. To address how Asl-A phosphorylation specifically affects Plk4 regulation, we generated Asl-A fragment phospho-mutants and expressed them in cultured Drosophila cells. Asl-A-13A stimulated kinase activity by relieving Plk4 autoinhibition. In contrast, Asl-A-13PM inhibited Plk4 activity by a novel mechanism involving autophosphorylation of Plk4's kinase domain. Thus, Asl-A's phosphorylation state determines which of Asl-A's two opposing effects are exerted on Plk4. Initially, nonphosphorylated Asl binds Plk4 and stimulates its kinase activity, but after Asl is phosphorylated, a negative-feedback mechanism suppresses Plk4 activity. This dual regulatory effect by Asl-A may limit Plk4 to bursts of activity that modulate centriole duplication.

The Art of War: harnessing the epigenome against cancer.

Histone chaperones are indispensable regulators of chromatin structure and function. Recent work has shown that they are frequently mis-regulated in cancer, which can have profound consequences on tumor growth and survival. Here, we focus on chaperones for the essential H3 histone variants H3.3 and CENP-A, specifically HIRA, DAXX/ATRX, DEK, and HJURP. This review summarizes recent studies elucidating their roles in regulating chromatin and discusses how cancer-specific chromatin interactions can be exploited to target cancer cells.

Quantitative outcome of registration methods for correcting cardiac drift in cardiac PET/CT imaging.

Myocardial perfusion studies suffer from artifacts caused by misalignment of the transmission and emission data due to the influences of voluntary and involuntary patient motion. Regardless of 68Ge or respiratory-averaged CT based attenuation correction and good patient cooperation, approximately 21% of perfusion studies exhibit artifacts arising from misalignment that cannot be corrected by manipulating the attenuation acquisition protocol. This misalignment, termed cardiac drift, is caused by slow-moving abdominal cavity contents that reposition the heart in the thorax and appear as myocardial uptake overlying the left CT lung in fused PET/CT images. This study evaluates three postimaging registration techniques to correct PET/CT misalignment by altering the transmission map to match myo-cardial uptake. Simulated misalignment studies were performed with a cardiac torso phantom filled with [18F]FDG at 10:1 myocardium/background. An air-filled saline bag affixed to the medial left lung surface served as a distensible lung. An initial CT acquisition was followed by successive PET acquisitions consisting of small displacements of the cardiac insert into the left lung. Phantom transmission scans were aligned to the myocardial uptake in the emission scans by applying 1) full rigid-body translations and rotations, 2) rigid-body restricted to medial / lateral and superior / inferior translation, or 3) an emission-driven method that adds myocardial tissue to the transmission scan. These methods were also applied to 10 low-likelihood coronary artery disease (CAD) patients showing signs of cardiac drift. Full rigid-body registration showed significant over-correction (p < 0.004) of activity concentrations in the artifact areas of the phantom data due the relocation of highly attenuating structures (i.e., spine). Inaccurate regional activity distributions were also observed as streaks extending from the spine and these results were replicated in the patient population. There was no significant difference between the true phantom activity concentration after correction with the emission-driven method. Misalignment corrected with the rigid-body registration results in an increase in activity concentration but fails to accurately recover the true concentration. These data suggest that a nonlinear image registration approach such as an emission-driven method results in a more uniform activity distribution throughout the myocardium, and is more appropriate for addressing the cardiac drift misalignment problem.

Chromatin Dynamics in Vivo: A Game of Musical Chairs.

Histones are a major component of chromatin, the nucleoprotein complex fundamental to regulating transcription, facilitating cell division, and maintaining genome integrity in almost all eukaryotes. In addition to canonical, replication-dependent histones, replication-independent histone variants exist in most eukaryotes. In recent years, steady progress has been made in understanding how histone variants assemble, their involvement in development, mitosis, transcription, and genome repair. In this review, we will focus on the localization of the major histone variants H3.3, CENP-A, H2A.Z, and macroH2A, as well as how these variants have evolved, their structural differences, and their functional significance in vivo.

Drosophila casein kinase I alpha regulates homolog pairing and genome organization by modulating condensin II subunit Cap-H2 levels.

The spatial organization of chromosomes within interphase nuclei is important for gene expression and epigenetic inheritance. Although the extent of physical interaction between chromosomes and their degree of compaction varies during development and between different cell-types, it is unclear how regulation of chromosome interactions and compaction relate to spatial organization of genomes. Drosophila is an excellent model system for studying chromosomal interactions including homolog pairing. Recent work has shown that condensin II governs both interphase chromosome compaction and homolog pairing and condensin II activity is controlled by the turnover of its regulatory subunit Cap-H2. Specifically, Cap-H2 is a target of the SCFSlimb E3 ubiquitin-ligase which down-regulates Cap-H2 in order to maintain homologous chromosome pairing, chromosome length and proper nuclear organization. Here, we identify Casein Kinase I alpha (CK1α) as an additional negative-regulator of Cap-H2. CK1α-depletion stabilizes Cap-H2 protein and results in an accumulation of Cap-H2 on chromosomes. Similar to Slimb mutation, CK1α depletion in cultured cells, larval salivary gland, and nurse cells results in several condensin II-dependent phenotypes including dispersal of centromeres, interphase chromosome compaction, and chromosome unpairing. Moreover, CK1α loss-of-function mutations dominantly suppress condensin II mutant phenotypes in vivo. Thus, CK1α facilitates Cap-H2 destruction and modulates nuclear organization by attenuating chromatin localized Cap-H2 protein.

Two Polo-like kinase 4 binding domains in Asterless perform distinct roles in regulating kinase stability.

Plk4 (Polo-like kinase 4) and its binding partner Asterless (Asl) are essential, conserved centriole assembly factors that induce centriole amplification when overexpressed. Previous studies found that Asl acts as a scaffolding protein; its N terminus binds Plk4's tandem Polo box cassette (PB1-PB2) and targets Plk4 to centrioles to initiate centriole duplication. However, how Asl overexpression drives centriole amplification is unknown. In this paper, we investigated the Asl-Plk4 interaction in Drosophila melanogaster cells. Surprisingly, the N-terminal region of Asl is not required for centriole duplication, but a previously unidentified Plk4-binding domain in the C terminus is required. Mechanistic analyses of the different Asl regions revealed that they act uniquely during the cell cycle: the Asl N terminus promotes Plk4 homodimerization and autophosphorylation during interphase, whereas the Asl C terminus stabilizes Plk4 during mitosis. Therefore, Asl affects Plk4 in multiple ways to regulate centriole duplication. Asl not only targets Plk4 to centrioles but also modulates Plk4 stability and activity, explaining the ability of overexpressed Asl to drive centriole amplification.

The use of cultured Drosophila cells for studying the microtubule cytoskeleton.

Cultured Drosophila cell lines have been developed into a powerful tool for studying a wide variety of cellular processes. Their ability to be easily and cheaply cultured as well as their susceptibility to protein knockdown via double-stranded RNA-mediated interference (RNAi) has made them the model system of choice for many researchers in the fields of cell biology and functional genomics. Here we describe basic techniques for gene knockdown, transgene expression, preparation for fluorescence microscopy, and centrosome enrichment using cultured Drosophila cells with an emphasis on studying the microtubule cytoskeleton.

Accuracy Evaluation of a 3D Ultrasound-guided Biopsy System.

Early detection of prostate cancer is critical in maximizing the probability of successful treatment. Current systematic biopsy approach takes 12 or more randomly distributed core tissue samples within the prostate and can have a high potential, especially with early disease, for a false negative diagnosis. The purpose of this study is to determine the accuracy of a 3D ultrasound-guided biopsy system. Testing was conducted on prostate phantoms created from an agar mixture which had embedded markers. The phantoms were scanned and the 3D ultrasound system was used to direct the biopsy. Each phantom was analyzed with a CT scan to obtain needle deflection measurements. The deflection experienced throughout the biopsy process was dependent on the depth of the biopsy target. The results for markers at a depth of less than 20 mm, 20-30 mm, and greater than 30 mm were 3.3 mm, 4.7 mm, and 6.2 mm, respectively. This measurement encapsulates the entire biopsy process, from the scanning of the phantom to the firing of the biopsy needle. Increased depth of the biopsy target caused a greater deflection from the intended path in most cases which was due to an angular incidence of the biopsy needle. Although some deflection was present, this system exhibits a clear advantage in the targeted biopsy of prostate cancer and has the potential to reduce the number of false negative biopsies for large lesions.

The structure of the plk4 cryptic polo box reveals two tandem polo boxes required for centriole duplication.

Centrioles are key microtubule polarity determinants. Centriole duplication is tightly controlled to prevent cells from developing multipolar spindles, a situation that promotes chromosomal instability. A conserved component in the duplication pathway is Plk4, a polo kinase family member that localizes to centrioles in M/G1. To limit centriole duplication, Plk4 levels are controlled through trans-autophosphorylation that primes ubiquitination. In contrast to Plks 1-3, Plk4 possesses a unique central region called the "cryptic polo box." Here, we present the crystal structure of this region at 2.3 Å resolution. Surprisingly, the structure reveals two tandem homodimerized polo boxes, PB1-PB2, that form a unique winged architecture. The full PB1-PB2 cassette is required for binding the centriolar protein Asterless as well as robust centriole targeting. Thus, with its C-terminal polo box (PB3), Plk4 has a triple polo box architecture that facilitates oligomerization, targeting, and promotes trans-autophosphorylation, limiting centriole duplication to once per cell cycle.

Posterior bladder layering of excreted 18F-FDG on PET/CT.

PURPOSE: This study was carried out to determine the frequency, characteristics and possible causes and clinical significance of occasionally observed posterior layering of excreted 2-deoxy-2-[F]fluoro-D-glucose (F-FDG) in the bladder. PROCEDURES: A review of 567 consecutive patients referred for positron emission tomography/computerized tomography studies was performed. Urinary bladder size was measured in patients with and without layering. Urine samples from two patients with layering were imaged ex vivo. RESULTS: Twenty-four of 567 studies (4%) showed F-FDG posterior bladder layering. Mean volume+/-SD of the bladder was 175+/-161 ml in patients with layering and 93+/-83 ml in patients without layering. Urine samples collected from two randomly chosen patients with layering in vivo failed to show layering ex vivo. CONCLUSION: Posterior F-FDG layering occurs in 4% of positron emission tomography/computerized tomography cases and highly correlates with bladder volume. The mechanism is hypothesized to be because of slow F-FDG excretion in patients with a distended urinary bladder resulting in delayed mixing with urine, but needs to be further investigated with a more comprehensive study.

Preparation of Drosophila S2 cells for light microscopy.

The ideal experimental system would be cheap and easy to maintain, amenable to a variety of techniques, and would be supported by an extensive literature and genome sequence database. Cultured Drosophila S2 cells, the product of disassociated 20-24 hour old embryos, possess all these properties. Consequently, S2 cells are extremely well-suited for the analysis of cellular processes, including the discovery of the genes encoding the molecular components of the process or mechanism of interest. The features of S2 cells that are most responsible for their utility are the ease with which they are maintained, their exquisite sensitivity to double-stranded (ds)RNA-mediated interference (RNAi), and their tractability to fluorescence microscopy as either live or fixed cells. S2 cells can be grown in a variety of media, including a number of inexpensive, commercially-available, fully-defined, serum-free media. In addition, they grow optimally and quickly at 21-24 degrees C and can be cultured in a variety of containers. Unlike mammalian cells, S2 cells do not require a regulated atmosphere, but instead do well with normal air and can even be maintained in sealed flasks. Complementing the ease of RNAi in S2 cells is the ability to readily analyze experimentally-induced phenotypes by phase or fluorescence microscopy of fixed or live cells. S2 cells grow in culture as a single monolayer but do not display contact inhibition. Instead, cells tend to grow in colonies in dense cultures. At low density, S2 cultures grown on glass or tissue culture-treated plastic are round and loosely-attached. However, the cytology of S2 cells can be greatly improved by inducing them to flatten extensively by briefly culturing them on a surface coated with the lectin, concanavalin A (ConA). S2 cells can also be stably transfected with fluorescently-tagged markers to label structures or organelles of interest in live or fixed cells. Therefore, the usual scenario for the microscopic analysis of cells is this: first, S2 cells (which can possess transgenes to express tagged markers) are treated by RNAi to eliminate a target protein(s). RNAi treatment time can be adjusted to allow for differences in protein turn-over kinetics and to minimize cell trauma/death if the target protein is important for viability. Next, the treated cells are transferred to a dish containing a coverslip pre-coated with conA to induce cells to spread and tightly adhere to the glass. Finally, cells are imaged with the researcher's choice of microscopy modes. S2 cells are particularly good for studies requiring extended visualization of live cells since these cells stay healthy at room temperature and normal atmosphere.

Prompt-gamma compensation in Rb-82 myocardial perfusion 3D PET/CT.

OBJECTIVE: To compare the diagnostic accuracy of Rb-82 myocardial perfusion three-dimensional (3D) PET with and without prompt-gamma compensation (PGC). METHODS AND RESULTS: Retrospective, single center study of 76 patients who had rest and adenosine stress Rb-82 myocardial perfusion 3D PET. All studies were acquired using a Siemens Biograph-40 PET/CT scanner and were reconstructed with and without PGC. Fifty-seven patients (mean age 63 +/- 11 years, 26 men) had coronary angiography within 40 days of Rb-82 imaging. Nineteen patients (mean age 43 +/- 7 years, 10 men) had low likelihood of coronary artery disease (CAD). All PET images were scored by consensus of two blinded readers on a standard 5-point scale using a 17-segment left ventricular model. A normal PET test was defined as a summed stress score of less than four. Obstructive CAD at coronary angiography was used as the gold-standard and was defined as luminal stenoses > or =50% in one or more major coronary arteries. The prevalence of obstructive disease at coronary angiography was 68% (39/57). The mean summed stress score was 12 +/- 12 for PGC images and was 18 +/- 14 for non-PGC images. Sensitivity and specificity for obstructive CAD were 90% (95% CI 88-99) and 72% (95% CI 52-93) for PGC images and 95% (95% CI 88-100) and 22% (95% CI 3-41) for non-PGC images. CONCLUSION: PGC in Rb-82 3D PET improves the specificity for obstructive CAD at coronary angiography with no significant loss in sensitivity.