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A healthy, urban-dwelling man presented with lassitude, jaundice without increased liver enzymes or obstructive features on imaging, brief acute kidney injury, leucocytosis with near-normal C reactive protein and markedly increased serum amylase and lipase. Leptospirosis was not considered for 10 days because of the low incidence of the disease in the country, absent animal contact and physicians’ low index of suspicion. Presentation without fever and without the commonly associated abdominal pain, myalgia, headache, thrombocytopaenia or elevated serum creatine kinase added to the diagnostic challenge. Once an infectious cause of acute pancreatitis was contemplated, leptospirosis was immediately sought and diagnosed by PCR of urine and microscopic agglutination test, and he fully recovered on ceftriaxone. Physicians in countries with a low incidence of leptospirosis should be more aware of the possibility of the disease even when several key features such as fever or pain are missing and the patient has a rare infectious acute pancreatitis. [ABSTRACT FROM AUTHOR]

It is histologically indistinguishable from splenic marginal-zone lymphoma and villous lymphocytes have immunologic and immunophenotypical characters of marginal zone lymphocytes.[2] Clinically, moderate lymphocytosis, villous lymphocytes on peripheral blood smear, marked splenomegaly, no lymphadenopathy, serum monoclonal band and a slow benign course are typical. Blood smear demonstrated abundant atypical lymphocytes with villous projections (Figure 1B). Imaging revealed massive splenomegaly (homogeneous) occupying most of the abdominal cavity (Figure 1A). [Extracted from the article]

Expression of the key anti-inflammatory cytokine IL-10 in lipopolysaccharide (LPS)-stimulated macrophages is mediated by a delayed autocrine/paracrine loop of type I interferons (IFN) to ensure timely attenuation of inflammation. We have previously shown that cAMP synergizes with early IL-10 expression by LPS, but is unable to amplify the late type I IFN-dependent activity. We now examined the mechanism of this synergistic transcription in mouse macrophages at the promoter level, and explored the crosstalk between type I IFN signaling and cAMP, using the β-adrenergic receptor agonist, isoproterenol, as a cAMP inducer. We show that silencing of the type I IFN receptor enables isoproterenol to synergize with LPS also at the late phase, implying that autocrine type I IFN activity hinders synergistic augmentation of LPS-stimulated IL-10 expression by cAMP at the late phase. Furthermore, IL-10 expression in LPS-stimulated macrophages is exclusively stimulated by either IFNα or isoproterenol. We identified a set of two proximate and inter-dependent cAMP response element (CRE) sites that cooperatively regulate early IL-10 transcription in response to isoproterenol-stimulated CREB and that further synergize with a constitutive Sp1 site. At the late phase, up-regulation of Sp1 activity by LPS-stimulated type I IFN is correlated with loss of function of the CRE sites, suggesting a mechanism for the loss of synergism when LPS-stimulated macrophages switch to type I IFN-dependent IL-10 expression. This report delineates the molecular mechanism of cAMP-accelerated IL-10 transcription in LPS-stimulated murine macrophages that can limit inflammation at its onset. [ABSTRACT FROM AUTHOR]

Integrated microfluidics is a sophisticated three-dimensional (multi layer) solution for high complexity serial or parallel processes. Fabrication of integrated microfluidic devices requires soft lithography and the stacking of thin-patterned PDMS layers. Precise layer alignment and bonding is crucial. There are no previously reported standards for alignment of the layers, which is mostly performed using uncontrolled processes with very low alignment success. As a result, integrated microfluidics is mostly used in academia rather than in the many potential industrial applications. We have designed and manufactured a semiautomatic Microfluidic Device Assembly System (μDAS) for full device production. μDAS comprises an electrooptic mechanical system consisting of four main parts: optical system, smart media holder (for PDMS), a micropositioning xyzϑ system and a macropositioning XY mechanism. The use of the μDAS yielded valuable information regarding PDMS as the material for device fabrication, revealed previously unidentified errors, and enabled optimization of a robust fabrication process. In addition, we have demonstrated the utilization of the μDAS technology for fabrication of a complex 3 layered device with over 12 000 micromechanical valves and an array of 64 × 64 DNA spots on a glass substrate with high yield and high accuracy. We increased fabrication yield from 25% to about 85% with an average layer alignment error of just ∼4 μm. It also increased our protein expression yields from 80% to over 90%, allowing us to investigate more proteins per experiment. The μDAS has great potential to become a valuable tool for both advancing integrated microfluidics in academia and producing and applying microfluidic devices in the industry. [ABSTRACT FROM AUTHOR]

To appropriately acclimate to environmental stresses, plants have to rapidly activate a specific transcriptional program. Yet, the identity and function of many of the transcriptional regulators that mediate early responses to abiotic stress stimuli is still unknown. In this work we employed the promoter of the multi-stress-responsive zinc-finger protein Zat12 in yeast one-hybrid ( Y1H) screens to identify early abiotic stress-responsive transcriptional regulators. Analysis of Zat12 promoter fragments fused to luciferase underlined an approximately 200 bp fragment responsive to NaCl and to reactive oxygen species ( ROS). Using these segments and others as baits against Y1H control or stress Arabidopsis prey libraries, we identified 15 potential Zat12 transcriptional regulators. Among the prominent proteins identified were known transcription factors including bZIP29 and ANAC91 as well as unknown function proteins such as a homolog of the human USB1, a U6 small nuclear RNA ( snRNA) processing protein, and dormancy/auxin-associated family protein 2 ( DRM2). Altered expression of Zat12 during high light stress in the knockout mutants further indicated the involvement of these proteins in the regulation of Zat12. Using a state of the art microfluidic approach we showed that AtUSB1 and DRM2 can specifically bind dsDNA and were able to identify the preferred DNA-binding motif of all four proteins. Overall, the proteins identified in this work provide an important start point for charting the earliest signaling network of Zat12 and of other genes required for acclimation to abiotic stresses. [ABSTRACT FROM AUTHOR]

Objective Clinical follow-up of aortic dimensions is performed interchangeably by multi-detector computed tomography (MDCT) and by cardiac echocardiography (ECHO). This study assesses the relationship between measurements of the aortic diameter by MDCT and ECHO at various predetermined locations using several methods. Methods The aortic diameter was measured at 6 locations between the aortic annulus and the aortic arch in 49 patients who underwent both MDCT and ECHO. Measurements were performed by three methods: internal-to-internal edge (INT), external-to-internal edge (MIX), and external-to-external edge (EXT). Measurements by MDCT and ECHO were made by an experienced radiologist and cardiologist, respectively, both blinded to results and images from the other modality. Results The average aortic diameter at all locations was significantly different between the MDCT and ECHO by all three methods (INT: 30.0 ± 5.8 mm vs. 27.8 ± 5.9 mm; MIX: 31.5 ± 5.8 mm vs. 30.8 ± 5.8 mm; EXT: 32.9 ± 6.6 mm vs. 33.8 ± 6.5 mm, p < 0.002 for all). While mean absolute differences between INT and EXT methods were similar (3.5 ± 3.1 mm and 3.4 ± 2.7 mm, respectively), the absolute difference using the MIX method was significantly smaller (3.1 ± 2.8 mm; p < 0.001 for INT vs. MIX; p < 0.05 for EXT vs. MIX). Conclusions There is considerable variability between MDCT and ECHO measurements of the ascending aorta. Measuring the aortic diameter by the MIX provides the closest measurements and is advised for long-term follow-up. [ABSTRACT FROM AUTHOR]

Varicella Zoster Virus (VZV), the alphaherpesvirus that causes varicella upon primary infection and Herpes zoster (shingles) following reactivation in latently infected neurons, is known to be fusogenic. It forms polynuclear syncytia in culture, in varicella skin lesions and in infected fetal human ganglia xenografted to mice. After axonal infection using VZV expressing green fluorescent protein (GFP) in compartmentalized microfluidic cultures there is diffuse filling of axons with GFP as well as punctate fluorescence corresponding to capsids. Use of viruses with fluorescent fusions to VZV proteins reveals that both proteins encoded by VZV genes and those of the infecting cell are transferred in bulk from infecting non-neuronal cells to axons. Similar transfer of protein to axons was observed following cell associated HSV1 infection. Fluorescence recovery after photobleaching (FRAP) experiments provide evidence that this transfer is by diffusion of proteins from the infecting cells into axons. Time-lapse movies and immunocytochemical experiments in co-cultures demonstrate that non-neuronal cells fuse with neuronal somata and proteins from both cell types are present in the syncytia formed. The fusogenic nature of VZV therefore may enable not only conventional entry of virions and capsids into axonal endings in the skin by classical entry mechanisms, but also by cytoplasmic fusion that permits viral protein transfer to neurons in bulk. [ABSTRACT FROM AUTHOR]

Summary: A pathogen’s ability to engage host receptors is a critical determinant of its host range and interspecies transmissibility, key issues for understanding emerging diseases. However, the identification of host receptors, which are also attractive drug targets, remains a major challenge. Our structural bioinformatics studies reveal that both bacterial and viral pathogens have evolved to structurally mimic native host ligands (ligand mimicry), thus enabling engagement of their cognate host receptors. In contrast to the structural homology, amino acid sequence similarity between pathogen molecules and the mimicked host ligands was low. We illustrate the utility of this concept to identify pathogen receptors by delineating receptor tyrosine kinase Axl as a candidate receptor for the polyomavirus SV40. The SV40-Axl interaction was validated, and its participation in the infection process was verified. Our results suggest that ligand mimicry is widespread, and we present a quick tool to screen for pathogen-host receptor interactions. [Copyright &y& Elsevier]

To attain fertilization the spermatozoon binds to the egg zona pellucida (ZP) via sperm receptor(s) and undergoes an acrosome reaction (AR). Several sperm receptors have been described in the literature; however, the identity of this receptor is not yet certain. In this study, we suggest that theα7 nicotinic acetylcholine receptor (α7nAChR) might be a sperm receptor activated by ZP to induce epidermal growth factor receptor (EGFR)-mediated AR. We found that isolated ZP or α7 agonists induced the AR in sperm from WT but not α7-null spermatozoa, and the induced AR was inhibited by α7 or EGFR antagonists. Moreover, α7-null sperm showed very little binding to the egg, and microfluidic affinity in vitro assay clearly showed that α7nAChR, as well as EGFR, interacted with ZP3. Induction of EGFR activation and the AR by an α7 agonist was inhibited by a Src family kinase (SFK) inhibitor. In conclusion we suggest that activation ofα7 by ZP leads to SFK-dependent EGFR activation, Ca2+ influx, and the acrosome reaction. [ABSTRACT FROM AUTHOR]

A healthy young man presented to the emergency department with mild hemoptysis associated with cannabis abuse. He was on no medications and cocaine abuse was ruled out by both history and negative toxicology screens. There were neither signs of an infection, nor of a systemic or cardiac disease. Imaging studies were consistent with diffuse alveolar hemorrhage, a reported, albeit rare adverse drug reaction of tetrahydrocannabinol (Naranjo score 6). The patient improved spontaneously within a few days, hemoptysis stopped and repeat imaging was entirely normal. With the increase in cannabis abuse and enhanced cannabis potency worldwide clinicians may increasingly encounter even unusual cannabis-associated adverse drug reactions, including associated diffuse alveolar hemorrhage. [ABSTRACT FROM AUTHOR]

The pro-longevity enzyme SIRT6 regulates various metabolic pathways. Gene expression analyses in SIRT6 heterozygotic mice identify significant decreases in PPARα signaling, known to regulate multiple metabolic pathways. SIRT6 binds PPARα and its response element within promoter regions and activates gene transcription. Sirt6 +/− results in significantly reduced PPARα-induced β-oxidation and its metabolites and reduced alanine and lactate levels, while inducing pyruvate oxidation. Reciprocally, starved SIRT6 transgenic mice show increased pyruvate, acetylcarnitine, and glycerol levels and significantly induce β-oxidation genes in a PPARα-dependent manner. Furthermore, SIRT6 mediates PPARα inhibition of SREBP-dependent cholesterol and triglyceride synthesis. Mechanistically, SIRT6 binds PPARα coactivator NCOA2 and decreases liver NCOA2 K780 acetylation, which stimulates its activation of PPARα in a SIRT6-dependent manner. These coordinated SIRT6 activities lead to regulation of whole-body respiratory exchange ratio and liver fat content, revealing the interactions whereby SIRT6 synchronizes various metabolic pathways, and suggest a mechanism by which SIRT6 maintains healthy liver. • PPARα mediates various SIRT6-regulated metabolic pathways • PPARα binds to and is activated by SIRT6 to promote fatty acid beta oxidation • SIRT6 decreases NCOA2 acetylation and induces its co-activation of PPARα • Coordinated SIRT6-PPARα activities control energy production under limited nutrients How the pro-longevity enzyme SIRT6 coordinates between various metabolic pathways is still obscure. Here, Naiman et al. show that SIRT6 activates PPARα to promote fatty acid beta oxidation and inhibit pyruvate oxidation during fasting. This ultimately decides the energy source under nutrient-limited conditions, promoting fat usage over other energy sources. [ABSTRACT FROM AUTHOR]

Transcription of protein-coding genes is highly dependent on the RNA polymerase II core promoter. Core promoters, generally defined as the regions that direct transcription initiation, consist of functional core promoter motifs (such as the TATA-box, initiator [Inr], and downstream core promoter element [DPE]) that confer specific properties to the core promoter. The known basal transcription factors that support TATA-dependent transcription are insufficient for in vitro transcription of DPE-dependent promoters. In search of a transcription factor that supports DPE-dependent transcription, we used a biochemical complementation approach and identified the Drosophila TBP (TATA-box-binding protein)-related factor 2 (TRF2) as an enriched factor in the fractions that support DPE-dependent transcription. We demonstrate that the short TRF2 isoform preferentially activates DPE-dependent promoters. DNA microarray analysis reveals the enrichment of DPE promoters among short TRF2 up-regulated genes. Using primer extension analysis and reporter assays, we show the importance of the DPE in transcriptional regulation of TRF2 target genes. It was previously shown that, unlike TBP, TRF2 fails to bind DNA containing TATA-boxes. Using microfluidic affinity analysis, we discovered that short TRF2-bound DNA oligos are enriched for Inr and DPE motifs. Taken together, our findings highlight the role of short TRF2 as a preferential core promoter regulator. [ABSTRACT FROM AUTHOR]

An older adult patient was admitted with epigastric pain and vomiting and found to have an abdominal mass, increased cholestatic liver enzymes and markedly elevated serum lactate dehydrogenase (LDH). Imaging revealed extensive liver metastases of unknown primary but also an unusual splenic metastasis diagnosed by liver biopsy as malignant melanoma. The patient became lethargic and developed mental status changes associated with asterixis, abnormal EEG, and increased serum ammonia levels. All reversed with high-dose lactulose and had no alternative explanation other than an unusual hepatic encephalopathy secondary to portosystemic shunts bypassing the extensively metastatic liver., (© The Author(s) 2024. Published by Oxford University Press on behalf of the Association of Physicians. All rights reserved. For permissions, please email: journals.permissions@oup.com.)

"Whiplash"-type injuries are commonly encountered and often cause neck pain, neck stiffness, and headaches. However, these injuries can have rare and poorly recognized complications, such as the development of a prevertebral hematoma leading to acute respiratory failure in the emergency department, followed by severe, life-threatening dysphagia and recurrent aspirations. In the patient described herein, a whiplash injury was accompanied by vocal cord paralysis and dysphonia (vagus nerve), dysgeusia (glossopharyngeal nerve, vagus nerve), and upper esophageal spasm (cricopharyngeal muscle, vagus nerve). It is unlikely that this was a complication of cervical fusion surgery. Instead, a combined stretch-induced lower cranial nerve injury, possibly on the exit of these nerves through the jugular foramen, seems to be a likely, but underappreciated mechanism occurring in rare instances of whiplash injuries., Competing Interests: Conflicts of interest The authors have no conflicts of interest to declare., (© 2022 The Korean Society of Traumatology.)

Realization of bioinspired molecular machines that can perform many and diverse operations in response to external chemical commands is a major goal in nanotechnology, but current molecular machines respond to only a few sequential commands. Lack of effective methods for introduction and removal of command compounds and low efficiencies of the reactions involved are major reasons for the limited performance. We introduce here a user interface based on a microfluidics device and single-molecule fluorescence spectroscopy that allows efficient introduction and removal of chemical commands and enables detailed study of the reaction mechanisms involved in the operation of synthetic molecular machines. The microfluidics provided 64 consecutive DNA strand commands to a DNA-based motor system immobilized inside the microfluidics, driving a bipedal walker to perform 32 steps on a DNA origami track. The microfluidics enabled removal of redundant strands, resulting in a 6-fold increase in processivity relative to an identical motor operated without strand removal and significantly more operations than previously reported for user-controlled DNA nanomachines. In the motor operated without strand removal, redundant strands interfere with motor operation and reduce its performance. The microfluidics also enabled computer control of motor direction and speed. Furthermore, analysis of the reaction kinetics and motor performance in the absence of redundant strands, made possible by the microfluidics, enabled accurate modeling of the walker processivity. This enabled identification of dynamic boundaries and provided an explanation, based on the "trap state" mechanism, for why the motor did not perform an even larger number of steps. This understanding is very important for the development of future motors with significantly improved performance. Our universal interface enables two-way communication between user and molecular machine and, relying on concepts similar to that of solid-phase synthesis, removes limitations on the number of external stimuli. This interface, therefore, is an important step toward realization of reliable, processive, reproducible, and useful externally controlled DNA nanomachines.

Protein post-translational modifications mediate dynamic cellular processes with broad implications in human disease pathogenesis. There is a large demand for high-throughput technologies supporting post-translational modifications research, and both mass spectrometry and protein arrays have been successfully utilized for this purpose. Protein arrays override the major limitation of target protein abundance inherently associated with MS analysis. This technology, however, is typically restricted to pre-purified proteins spotted in a fixed composition on chips with limited life-time and functionality. In addition, the chips are expensive and designed for a single use, making complex experiments cost-prohibitive. Combining microfluidics with in situ protein expression from a cDNA microarray addressed these limitations. Based on this approach, we introduce a modular integrated microfluidic platform for multiple post-translational modifications analysis of freshly synthesized protein arrays (IMPA). The system's potency, specificity and flexibility are demonstrated for tyrosine phosphorylation and ubiquitination in quasicellular environments. Unlimited by design and protein composition, and relying on minute amounts of biological material and cost-effective technology, this unique approach is applicable for a broad range of basic, biomedical and biomarker research., (© 2015 by The American Society for Biochemistry and Molecular Biology, Inc.)

Viral-host interactions represent potential drug targets for novel antiviral strategies (Flisiak et al., Hepatology, 2008, 47, 817-26). Hence, it is important to establish an adequate platform for identifying and analyzing such interactions. In this review, we discuss bottlenecks in conventional protein-protein interaction methodologies and present the contribution of innovative microfluidic-based technologies towards a solution to these problems with respect to viral-host proteomics.