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Understanding the natural history of coronary artery atherosclerosis is necessary to determine prognosis and prescribe effective therapies. Traditional management of coronary artery disease has focused on the treatment of flow-limiting anatomical obstructions that lead to ischaemia. In most scenarios, revascularization of these atherosclerotic plaques has not substantially improved freedom from death or myocardial infarction, questioning the utility of contemporary revascularization strategies to improve prognosis. Advances in non-invasive and invasive imaging techniques have helped to identify the characteristics of obstructive and non-obstructive plaques that are precursors for plaque progression and future acute coronary syndromes as well as cardiac death. These 'vulnerable plaques' develop as a consequence of systemic inflammation and are prone to inducing thrombosis. Vulnerable plaques most commonly have a large plaque burden with a well-formed necrotic core and thin fibrous cap and are metabolically active. Perivascular adipose tissue might, in some patients, be used as a surrogate for coronary inflammation and predict future risk of adverse cardiac events. Vulnerable plaques can be identified in their quiescent state, offering the potential for therapeutic passivation. In this Review, we describe the biological and compositional features of vulnerable plaques, the non-invasive and invasive diagnostic modalities to characterize vulnerable plaques, the prognostic utility of identifying vulnerable plaques, and the future studies needed to explore the value of intensified pharmacological and focal treatments of vulnerable plaques.

SummaryMammalian outer radial glia (oRG) emerge as cortical progenitor cells that directly support the development of an enlarged outer subventricular zone (oSVZ) and, in turn, the expansion of the neocortex. Thein vitrogeneration of oRG is essential to model and investigate the underlying mechanisms of human neocortical development and expansion. By activating the STAT3 pathway using LIF, which is not produced in guided cortical organoids, we developed a cerebral organoid differentiation method from human pluripotent stem cells (hPSCs) that recapitulates the expansion of a progenitor pool into the oSVZ. The structured oSVZ is composed of progenitor cells expressing specific oRG markers such asGFAP, LIFR, HOPX, which closely matches human oRGin vivo. In this microenvironment, cortical neurons showed faster maturation with enhanced metabolic and functional activity. Incorporation of hPSC-derived brain vascular LIF- producing pericytes in cerebral organoids mimicked the effects of LIF treatment. These data indicate that the cellular complexity of the cortical microenvironment, including cell-types of the brain vasculature, favors the appearance of oRG and provides a platform to routinely study oRG in hPSC-derived brain organoids.

Objectives Nursing homes (NHs) are important health care and residential environments for the growing number of frail older adults. The COVID-19 pandemic highlighted the vulnerability of NHs as they became COVID-19 hotspots. This study examines the associations of NH design with COVID-19 cases, deaths, and transmissibility and provides relevant design recommendations. Design A cross-sectional, nationwide study was conducted after combining multiple national data sets about NHs. Setting and Participants A total of 7785 NHs were included in the study, which represent 50.8% of all Medicare and/or Medicaid NH providers in the United States. Methods Zero-inflated negative binomial models were used to predict the total number of COVID-19 resident cases and deaths, separately. The basic reproduction number (R0) was calculated for each NH to reflect the transmissibility of COVID-19 among residents within the facility, and a linear regression model was estimated to predict log(R0 – 1). Predictors of these models included community factors and NHs’ resident characteristics, management and rating factors, and physical environmental features. Results Increased share of private rooms, larger living area per bed, and presence of a ventilator-dependent unit are significantly associated with reductions in COVID-19 cases, deaths, and transmissibility among residents. Increased number of certified beds in the NH is associated with reduced resident cases and deaths, after setting the number of actual residents as the exposure variable and controlling for staff cases. It also correlates with reduced transmissibility among residents when other risk factors, including staff cases, are controlled. Conclusions and Implications Architectural design attributes have significant impacts on COVID-19 transmissions in NHs. Considering the vulnerability of NH residents in congregated living environments, NHs will continue to be high-risk settings for infection outbreaks. To improve safety and resilience of NHs against future health disasters, facility guidelines and regulations should consider the need to increase private rooms and living areas.

The tumor necrosis factor receptor superfamily (TNFRSF) and their ligands mediate lymphoid tissue development and homeostasis in addition to key aspects of innate and adaptive immune responses. T cells of the adaptive immune system express a number of TNFRSF members that are used to receive signals at different instructive stages and produce several tumor necrosis factor superfamily (TNFSF) members as effector molecules. There is also one example of a TNFRSF member serving as a ligand for negative regulatory checkpoint receptors. In most cases, the ligands in afferent and efferent phases are membrane proteins and thus the interaction with TNFRSF members must take place in immunological synapses and other modes of cell-cell interaction. A particular feature of the TNFRSF-mediated signaling is the prominent use of linear ubiquitin chains as scaffolds for signaling complexes that activate nuclear factor κ-B and Fos/Jun transcriptional regulators. This review will focus on the signaling mechanisms triggered by TNFRSF members in their role as costimulators of early and late phases of T cell instruction and the delivery mechanism of TNFSF members through the immunological synapses of helper and cytotoxic effector cells.

In this chapter, we present techniques, based on molecular-scale nanofabrication and selective self-assembly, for the presentation of biomolecules of interest (ligands, receptors, etc.) on a surface with precise spatial control and arbitrary geometry at the single-molecule level. Metallic nanodot arrays are created on glass coverslips and are then used as anchors for the immobilization of biological ligands via thiol linking chemistry. The nanodot size is controlled by both lithography and metallization. The reagent concentration in self-assembly can be adjusted to ensure single-molecule occupancy for a given dot size. The surrounding glass is backfilled by a protein-repellent layer to prevent nonspecific adsorption. Moreover, bifunctional surfaces are created, whereby a second ligand is presented on the background, which is frequently a requirement for simulating complex cellular functions involving more than one key ligand. This platform serves as a novel and powerful tool for molecular and cellular biology, e.g., to study the fundamental mechanisms of receptor-mediated signaling.

Activation of various cell surface receptors triggers the reorganization of downstream signaling molecules into micron- or submicron-sized clusters. However, the functional consequences of such clustering has been unclear. We biochemically reconstituted a 12-component signaling pathway on model membranes, beginning with T cell receptor (TCR) activation and ending with actin assembly. When TCR phoshophorylation was triggered, downstream signaling proteins spontaneously separated into liquid-like clusters that promoted signaling outputs both in vitro and in human Jurkat T cells. Reconstituted clusters were enriched in kinases but excluded phosphatases, and enhanced actin filament assembly by recruiting and organizing actin regulators. These results demonstrate that protein phase separation can create a distinct physical and biochemical compartment that facilitates signaling.

Regulatory T-cells (Tregs) prevent autoimmune and alloimmune reactions. Augmenting Treg function may enhance Treg therapies for these diseases. Treg-specific inhibition of protein kinase C-theta (PKC-θ) enhances Treg function. However, it is unclear whether PKC-θ inhibition can boost Treg function in systemic inflammatory conditions. In a mouse model of acute GVHD, we found that Tregs treated with the PKC-θ inhibitor AEB071 reduced GVHD mortality and severity significantly better than DMSO treated Tregs. Compared to DMSO, AEB071 treated Tregs significantly reduced conventional T-cells (Tcon) proliferation on D4 after transplant. Multi-photon microscopy showed that AEB071 treated Tregs significantly increased Tcon velocity and displacement compared to DMSO. Mechanistically, AEB071 augments expression of Neuropilin-1 (Nrp1) and Lymphocyte activation gene 3 (Lag3). Antibody blockade of Nrp1 and Lag3 in transwell suppression assays reduced the effect of AEB071 on Treg function. PKC-θ inhibition also reduces phosphorylation of mTORC2 targets FoxO3a and Akt phospho-site S473, but not mTORC1 targets S6, 4E-BP1 or Akt phospho-site T308. Compared to DMSO, AEB071 treatment significantly increased fatty acid uptake and oxygen consumption rate (OCR). Phosphoproteomic analysis identified a significant alteration in the interaction between PKC-θ and the intermediate filament vimentin after AEB071 treatment, which was confirmed by confocal. Vimentin siRNA treatment also significantly reduces PKC-θ/vimentin interaction, increases Treg function, Nrp1 expression and OCR. In summary, PKC-θ and vimentin modulate multiple aspects of Treg function, and altering these molecules may enhance the efficacy of Treg therapeutics.

Background Tuberculosis (TB) is the most common opportunistic infection in HIV-infected adults in developing countries. Isoniazid (INH) is recommended for treatment of latent TB infection, however non-adherence is common. The purpose of this study was to apply in-house prepared isoniazid (INH) urine test strips in a clinical setting, and identify predictors of positive test results in an adherence questionnaire in HIV-infected adults taking INH for prevention of TB. Methods Cross-sectional study of adherence using a questionnaire and urine test strips for detection of INH metabolites at two hospitals in Pietermaritzburg, South Africa. Participants were aged at least 18 years, HIV positive, and receiving INH for prevention of tuberculosis disease. Univariate and multivariate analyses are used to identify factors relevant to adherence. Results 301 consecutive patients were recruited. 28% of participants had negative urine tests. 32 (37.2%, 95% CI25.4, 45.0) of the 86 patients who received INH from peripheral pharmacies said the pharmacy had run out of INH at some time, compared with central hospital pharmacies (p = 0.0001). In univariate analysis, a negative test was associated with self-reported missed INH doses (p = 0.043). Each 12-hour increment since last reported dose increased the likelihood of a negative test by 34% (p = 0.0007). Belief in INH safety was associated with a positive test (p = 0.021). In multivariate analysis, patients who believed INH is important for prevention of TB disease were more likely to be negative (p = 0.0086). Conclusion Adequate drug availability at peripheral pharmacies remains an important intervention for TB prevention. Key questions may identify potentially non-adherent patients. In-house prepared urine tests strips are an effective and cheap method of objectively assessing INH adherence, and could be used an important tool in TB control programs.

The purpose of Infrastructure Operations Tools Access (IOTA) effort is to accomplish development of an advanced architecture for ISR information management. The architecture will be based on a software component framework for information dissemination. The component framework will be based on Web Services standards and include provisions for enterprise workflow management. The component framework will leverage commercial off-the-shelf (COTS) technology and government off-the-shelf (GOTS) applications to provide mechanisms for data discovery, dissemination and visualization that can be rapidly adapted to varying information requirements. In particular, Java 2 Enterprise Edition (J2EE), will be used to provide a platform-independent set of standards and tools for developing the component framework. Broadsword Gatekeeper, a GOTS application, will also be used to define middleware capabilities for data access and discovery.

In summary, a history of previous angina before a nonthrombolized AMI is a marker of increased risk of infarct extension, recurrent ischemic pain, and mitral regurgitation. However, a history of previous angina was associated with trends toward smaller infarct size in patients who had spontaneous thrombolysis.

The sensory cues that developing neurons use to control the pattering of neurite outgrowth are currently not well defined. CXCR4 and the insulin like growth factor receptor (IGF1r) have been implicated in the regulation of chemotaxis, neuronal migration, and axonal guidance. In addition to these chemokine receptors role in the guidance of neuronal development, an increasing body of evidence shows they are up-regulated in many late stage and metastatic cancers. Neuroblastoma, a cancer of neural crest origin, often expresses high levels of CXCR4 and IGF1r, and depending upon the level of differentiation, is capable of producing fully extended axons. The mediator of the cytoskeletal changes seen in axonal extension is primarily carried out by actin polymerization. In order to better understand the role of CXCR4 and IGF1r role in mediating these cytoskeletal changes we used western blots and immunocytochemistry / confocal microscopy to analyze the neuroblastoma cell line SHSY-5Y. We were able to show that indeed both receptors are involved in neuronal outgrowth. However, the individual effects of treatment with the receptors ligand resulted in different cellular morphologies. CXCR4 stimulation resulted in a more differentiated neuronal form, while IGF1r stimulation resulted in a very immature neuronal morphology with shorter broader processes. Finally, we were able to quantitate the effects of treatment to a wide array of phenotypic patterns that shows the connection between our observed morphological changes to show that these observations are real effects of treatment. Citation Format: {Authors}. {Abstract title} [abstract]. In: Proceedings of the 102nd Annual Meeting of the American Association for Cancer Research; 2011 Apr 2-6; Orlando, FL. Philadelphia (PA): AACR; Cancer Res 2011;71(8 Suppl):Abstract nr 1435. doi:10.1158/1538-7445.AM2011-1435