Now You See It, Now You Don't: Point-of-Care Ultrasound Identification of Left Ventricular Thrombus-in-Transit.

BACKGROUND: Left-sided intracardiac thrombi are most commonly seen in conditions with decreased cardiac flow, such as myocardial infarction or atrial fibrillation. They can be propagated into the systemic circulation, leading to a cerebrovascular accident. Identification of thrombus-in-transit via point-of-care ultrasound (POCUS) has the potential to change patient management given its association with high patient morbidity and mortality. CASE REPORT: An intubated 60-year-old man was transferred to our emergency department for management of altered mental status and seizure-like activity. The patient was markedly hypotensive on arrival, and cardiac POCUS was performed to identify potential causes of hypotension. A left ventricular thrombus-in-transit was identified. The thrombus was notably absent on a repeat POCUS examination < 10 min later, which led to concern for thrombus propagation. Furthermore, the patient's vasopressor requirements had significantly increased in that time period. Subsequent emergent neuroimaging revealed a large ischemic stroke in the left internal carotid and middle cerebral artery distribution. The patient was, unfortunately, deemed to not be a candidate for either thrombectomy or thrombolysis and ultimately expired in the hospital. Why Should an Emergency Physician Be Aware of This? Serial POCUS examinations identified the propagation of this patient's thrombus-in-transit, leading the physician to change the initial presumptive diagnosis and treatment course, and pursue further imaging and workup for ischemic stroke. Identification of a thrombus-in-transit is a clue to potentially underlying critical pathology and should be followed with serial POCUS examinations to assess for treatment efficacy and thrombus propagation.

Applications of Advanced Natural Language Processing for Clinical Pharmacology.

Natural language processing (NLP) is a branch of artificial intelligence, which combines computational linguistics, machine learning, and deep learning models to process human language. Although there is a surge in NLP usage across various industries in recent years, NLP has not been widely evaluated and utilized to support drug development. To demonstrate how advanced NLP can expedite the extraction and analyses of information to help address clinical pharmacology questions, inform clinical trial designs, and support drug development, three use cases are described in this article: (1) dose optimization strategy in oncology, (2) common covariates on pharmacokinetic (PK) parameters in oncology, and (3) physiologically-based PK (PBPK) analyses for regulatory review and product label. The NLP workflow includes (1) preparation of source files, (2) NLP model building, and (3) automation of data extraction. The Clinical Pharmacology and Biopharmaceutics Summary Basis of Approval (SBA) documents, US package inserts (USPI), and approval letters from the US Food and Drug Administration (FDA) were used as our source data. As demonstrated in the three example use cases, advanced NLP can expedite the extraction and analyses of large amounts of information from regulatory review documents to help address important clinical pharmacology questions. Although this has not been adopted widely, integrating advanced NLP into the clinical pharmacology workflow can increase efficiency in extracting impactful information to advance drug development.

Blueprint for the development of resuscitation and emergency critical care fellowships.

The volume of critically ill patients presenting to the emergency department (ED) is increasing rapidly. Continued growth will likely further stress an already strained U.S. health care system. Numerous studies have demonstrated an association with worsened outcomes for critically ill patients boarding in the ED. To address the increasing volume and complexity of critically ill patients presenting to EDs nationwide, resuscitation and emergency critical care (RECC) fellowships were developed. RECC programs teach a general approach to the management of the undifferentiated critically ill patient, advanced management of critically ill patients by disease presentation, and ongoing supportive care of the critically ill patient boarding in the ED. The result is critical care training beyond that of a typical emergency medicine (EM) residency with a focus on the unique features and challenges of caring for critically ill patients in the ED not normally found in critical care fellowships. Graduates from RECC fellowships are well suited to practicing in any ED practice model and may be especially well prepared for EDs that distinguish acuity between zones (e.g., resuscitative care units, ED-based intensive care units). In addition to further developing clinical acumen, RECC fellowships provide graduates with a niche in EM education, research, and administration. In this article, we describe the philosophical principles and practical components necessary for the creation of future RECC fellowships.

XB130 deficiency enhances carcinogen-induced skin tumorigenesis.

XB130 is an adaptor protein that functions as a mediator of multiple tyrosine kinases important for regulating cell proliferation, survival, migration and invasion. Formerly predicted as an oncogene, alterations of its expression are documented in various human cancers. However, the exact role of XB130 in tumorigenesis is unknown. To address its function in skin tumorigenesis, a two-stage dimethylbenzanthracene (DMBA)/12-O-tetradecanoylphorbol 13-acetate (TPA) study was performed on XB130 knockout (KO), heterozygous (HZ) and wild-type (WT) littermate mice. DMBA/TPA-treated XB130 KO and HZ males developed a significantly higher number of epidermal tumors that were notably larger in size than did WT mice. Interestingly, DMBA/TPA-treated female mice did not show any difference in tumor multiplicity regardless of the genotypes. The skin tumor lesions of XB130 KO males were more progressed with an increased frequency of keratoacanthoma. Deficiency of XB130 dramatically increased epidermal tumor cell proliferation. The responses to DMBA and TPA stimuli were also individually investigated to elucidate the mechanistic role of XB130 at different stages of tumorigenesis. DMBA-treated male XB130 KO mice showed compensatory p53-mediated stress response. TPA-treated XB130 KO males demonstrated more skin ulceration with more severe edema, enhanced cell proliferation, accumulation of infiltrating neutrophils and increased production of pro-inflammatory cytokine genes compared with WT mice. Enhanced activities of nuclear factor-kappa B pathway, increased protein expression of metalloproteinase-9 and ERK1/2 phosphorylation were found in these KO mice. These findings demonstrate that XB130 acts as a tumor suppressor in carcinogen-induced skin tumorigenesis that may be mediated through inhibiting inflammation.

Ischemia-reperfusion induces death receptor-independent necroptosis via calpain-STAT3 activation in a lung transplant setting.

Ischemia-reperfusion (I/R)-induced lung injury undermines lung transplantation (LTx) outcomes by predisposing lung grafts to primary graft dysfunction (PGD). Necrosis is a feature of I/R lung injury. However, regulated necrosis (RN) with specific signaling pathways has not been explored in an LTx setting. In this study, we investigated the role of RN in I/R-induced lung injury. To study I/R-induced cell death, we simulated an LTx procedure using our cell culture model with human lung epithelial (BEAS-2B) cells. After 18 h of cold ischemic time (CIT) followed by reperfusion, caspase-independent cell death, mitochondrial reactive oxygen species production, and mitochondrial membrane permeability were significantly increased. N-acetyl-Leu-Leu-norleucinal (ALLN) (calpain inhibitor) or necrostatin-1 (Nec-1) [receptor interacting serine/threonine kinase 1 (RIPK1) inhibitor] reduced these changes. ALLN altered RIPK1/RIPK3 expression and mixed lineage kinase domain-like (MLKL) phosphorylation, whereas Nec-1 did not change calpain/calpastatin expression. Furthermore, signal transducer and activator of transcription 3 (STAT3) was demonstrated to be downstream of calpain and regulate RIPK3 expression and MLKL phosphorylation during I/R. This calpain-STAT3-RIPK axis induces endoplasmic reticulum stress and mitochondrial calcium dysregulation. LTx patients' samples demonstrate that RIPK1, MLKL, and STAT3 mRNA expression increased from CIT to reperfusion. Moreover, the expressions of the key proteins are higher in PGD samples than in non-PGD samples. Cell death associated with prolonged lung preservation is mediated by the calpain-STAT3-RIPK axis. Inhibition of RIPK and/or calpain pathways could be an effective therapy in LTx.

Relationship Between the Subcostal and Right Lateral Ultrasound Views of Inferior Vena Cava Collapse: Implications for Clinical Use of Ultrasonography.

BACKGROUND: The collapsibility index of the inferior vena cava is traditionally visualized from the subcostal region in the sagittal plane, referred to here as cIVCSS. Alternatively, the collapsibility index of the inferior vena cava can be visualized from the right midaxillary line in the coronal plane, referred to here as cIVCRC. It is unclear whether values of cIVCRC are comparable with values of cIVCSS because the inferior vena cava collapses asymmetrically into an elliptical form, quantified as the flat ratio of the inferior vena cava (F-IVC). This study aimed (1) to establish if cIVCRC is concordant or discordant to cIVCSS, and (2) to describe how this concordance or discordance is related to F-IVC. METHODS: This single-center cross-sectional study enrolled 110 spontaneously breathing patients. Values of cIVCRC were compared with cIVCSS. Performance of cIVCRC ≥ 42% in predicting fluid responsiveness, defined as cIVCSS ≥ 42%, was assessed. F-IVC was also correlated to the difference between cIVCSS and cIVCRC. RESULTS: cIVCRC ≥ 42% was 61.5% sensitive (95% CI, 31.58%-86.14%) and 67.1% specific (95% CI, 55.81%-77.06%) for predicting cIVCSS ≥ 42%. cIVCRC underestimated cIVCSS. The degree of discordance between cIVCRC and cIVCSS was proportional to the value of F-IVC. CONCLUSIONS: cIVCRC and cIVCSS measures are discordant, where cIVCRC underestimates cIVCSS. The degree of discordance is directly proportional to the value of F-IVC. Therefore, we recommend that cIVCRC ≥ 42% be used to rule in, but not to rule out, fluid responsivity. Wherever possible, F-IVC should be assessed to understand the clinical relevance of cIVCRC.

Intraosseous infusion is unreliable for adenosine delivery in the treatment of supraventricular tachycardia.

Supraventricular tachycardia (SVT) is a common tachyarrhythmia in the pediatric population that can necessitate immediate treatment. Adenosine has been well studied as a mainstay treatment, but the methods of adenosine administration have not been very well delineated. The intraosseous technique has presented itself as a possible method of administration. We describe 2 cases in which adenosine was administered through bone marrow infusion to convert SVT without success. The cases we describe show that intraosseous is not a reliable method of administering adenosine to stop SVT. Both patients presented with SVT refractory to vagal maneuvers and difficult intravenous placement. Intraosseous access was achieved, but administration of adenosine at increasing doses was unable to successfully convert the arrhythmia.

Regulatory networks define phenotypic classes of human stem cell lines.

Stem cells are defined as self-renewing cell populations that can differentiate into multiple distinct cell types. However, hundreds of different human cell lines from embryonic, fetal and adult sources have been called stem cells, even though they range from pluripotent cells-typified by embryonic stem cells, which are capable of virtually unlimited proliferation and differentiation-to adult stem cell lines, which can generate a far more limited repertoire of differentiated cell types. The rapid increase in reports of new sources of stem cells and their anticipated value to regenerative medicine has highlighted the need for a general, reproducible method for classification of these cells. We report here the creation and analysis of a database of global gene expression profiles (which we call the 'stem cell matrix') that enables the classification of cultured human stem cells in the context of a wide variety of pluripotent, multipotent and differentiated cell types. Using an unsupervised clustering method to categorize a collection of approximately 150 cell samples, we discovered that pluripotent stem cell lines group together, whereas other cell types, including brain-derived neural stem cell lines, are very diverse. Using further bioinformatic analysis we uncovered a protein-protein network (PluriNet) that is shared by the pluripotent cells (embryonic stem cells, embryonal carcinomas and induced pluripotent cells). Analysis of published data showed that the PluriNet seems to be a common characteristic of pluripotent cells, including mouse embryonic stem and induced pluripotent cells and human oocytes. Our results offer a new strategy for classifying stem cells and support the idea that pluripotency and self-renewal are under tight control by specific molecular networks.

Comprehensive microRNA profiling reveals a unique human embryonic stem cell signature dominated by a single seed sequence.

Embryonic stem cells are unique among cultured cells in their ability to self-renew and differentiate into a wide diversity of cell types, suggesting that a specific molecular control network underlies these features. Human embryonic stem cells (hESCs) are known to have distinct mRNA expression, global DNA methylation, and chromatin profiles, but the involvement of high-level regulators, such as microRNAs (miRNA), in the hESC-specific molecular network is poorly understood. We report that global miRNA expression profiling of hESCs and a variety of stem cell and differentiated cell types using a novel microarray platform revealed a unique set of miRNAs differentially regulated in hESCs, including numerous miRNAs not previously linked to hESCs. These hESC-associated miRNAs were more likely to be located in large genomic clusters, and less likely to be located in introns of coding genes. hESCs had higher expression of oncogenic miRNAs and lower expression of tumor suppressor miRNAs than the other cell types. Many miRNAs upregulated in hESCs share a common consensus seed sequence, suggesting that there is cooperative regulation of a critical set of target miRNAs. We propose that miRNAs are coordinately controlled in hESCs, and are key regulators of pluripotence and differentiation. Disclosure of potential conflicts of interest is found at the end of this article.

A soluble molecule secreted by human blastocysts modulates regulation of HOXA10 expression in an epithelial endometrial cell line.

OBJECTIVE: To determine the existence of a soluble signal, secreted from the human blastocyst embryo, that induces HOXA10 gene expression before cell-cell contact. DESIGN: To analyze, by semiquantitative reverse transcriptase polymerase chain reaction (RT-PCR), cell-free media that had contained human embryos cultured to the blastocyst stage for a soluble molecule that induces HOXA10 expression in an endometrial epithelial cell line (Ishikawa). SETTING: Assisted reproduction technology program of Yale University, New Haven, Connecticut. PATIENT(S): Patients undergoing intracytoplasmic sperm injection (ICSI) or in vitro fertilization (IVF) cycles. Treatment of Ishikawa cells with blastocyst-conditioned media. MAIN OUTCOME MEASURE(S): Determination of HOXA10 gene expression. RESULT(S): We demonstrate that cell-free media that had contained human embryos cultured to the blastocyst stage contain a soluble molecule that induces HOXA10 expression in an endometrial epithelial cell line (Ishikawa). We found that hCG does not induce HOXA10 in Ishikawa cells. CONCLUSION(S): Soluble molecules induce a well-characterized marker of endometrial receptivity in endometrial cells without blastocyst apposition. Additionally, HOXA10 induction can serve as a means of evaluating human embryos cultured for IVF and ET. High quality embryos may induce local endometrial receptivity before trophectoderm-endometrial contact.

CD4 T cell priming in dendritic cell-deficient mice.

Bone marrow (BM) chimeras (BMC) generated from mice carrying a null (-/-) mutation in the relB gene of the NF-kappaB family represent an ideal model for in vivo studies on the role of dendritic cells (DC) in the adaptive immune response. The spleen and lymph nodes (LN) of relB(-/-) BMC contain a small number of residual DC, mainly CD8alpha(+), that fail to up-regulate MHC class II and co-stimulatory molecules after stimulation in vitro. Moreover, residual spleen DC of relB(-/-) BMC have a 4-fold decrease in the ability to uptake and process soluble model antigen, ovalbumin (OVA), and failed to prime CD4 and CD8 T cells in vitro and in vivo. In addition, they also failed to present OVA peptide to OT-II transgenic T lymphocytes at a normal 1:10 (stimulator:responder) cell ratio. In spite of these multiple DC defects, relB(-/-) BMC immunized with plasmid DNA targeted to the spleen as the site of immune induction develop a specific CD4(+) T cell response comparable to that of relB competent mice. These data demonstrate that CD4( +) T cells can be primed in the absence of functional DC and suggest that relB may gauge the T cell response in vivo.

Interleukin-10 (IL-10) in experimental visceral leishmaniasis and IL-10 receptor blockade as immunotherapy.

Interleukin-10 (IL-10) is thought to promote intracellular infection, including human visceral leishmaniasis, by disabling Th1 cell-type responses and/or deactivating parasitized tissue macrophages. To develop a rationale for IL-10 inhibition as treatment in visceral infection, Th1 cytokine-driven responses were characterized in Leishmania donovani-infected BALB/c mice in which IL-10 was absent or overexpressed or its receptor (IL-10R) was blockaded. IL-10 knockout and normal mice treated prophylactically with anti-IL-10R demonstrated accelerated granuloma assembly and rapid parasite killing without untoward tissue inflammation; IL-12 and gamma interferon mRNA expression, inducible nitric oxide synthase reactivity, and responsiveness to antimony chemotherapy were also enhanced in knockout mice. In IL-10 transgenic mice, parasite replication was unrestrained, and except for antimony responsiveness, measured Th1 cell-dependent events were all initially impaired. Despite subsequent granuloma assembly, high-level infection persisted, and antimony-treated transgenic mice also relapsed. In normal mice with established infection, anti-IL-10R treatment was remarkably active, inducing near-cure by itself and synergism with antimony. IL-10's deactivating effects regulate outcome in experimental visceral leishmaniasis, and IL-10R blockade represents a potential immuno- and/or immunochemotherapeutic approach in this infection.