Experimental Studies in Stereolithography Resolution

As we move towards micron-scale rapid manufacturing, it is critical to understand build resolution of Stereolithography technology. In order to determine the resolution limitations, positive and negative features on Stereolithography parts were built and analyzed. Results from several experiments were compared to an analytical model and important resolution issues are highlighted. Based on these experimental results, parameters that will maximize build resolution for a number of well-understood shapes are suggested in the paper. Build resolution experimental results, analysis, and measurement techniques are discussed. Conclusions are drawn related to feature shape as resolution limits are approached.We gratefully acknowledge the support from the RPMI member companies and the George W. Woodruff School of Mechanical Engineering at Georgia Tech. This work was partially funded by the National Science Foundation under Grant Number DMI-9988664.Mechanical Engineerin

Impairment of Coronary Arteriolar Endothelium-Dependent Dilation after Multi-Walled Carbon Nanotube Inhalation: A Time-Course Study

Engineered nanomaterials have been developed for widespread applications due to many highly unique and desirable characteristics. The purpose of this study was to assess pulmonary inflammation and subepicardial arteriolar reactivity in response to multi-walled carbon nanotube (MWCNT) inhalation and evaluate the time course of vascular alterations. Rats were exposed to MWCNT aerosols producing pulmonary deposition. Pulmonary inflammation via bronchoalveolar lavage and MWCNT translocation from the lungs to systemic organs was evident 24 h post-inhalation. Coronary arterioles were evaluated 24–168 h post-exposure to determine microvascular response to changes in transmural pressure, endothelium-dependent and -independent reactivity. Myogenic responsiveness, vascular smooth muscle reactivity to nitric oxide, and α-adrenergic responses all remained intact. However, a severe impact on endothelium-dependent dilation was observed within 24 h after MWCNT inhalation, a condition which improved, but did not fully return to control after 168 h. In conclusion, results indicate that MWCNT inhalation not only leads to pulmonary inflammation and cytotoxicity at low lung burdens, but also a low level of particle translocation to systemic organs. MWCNT inhalation also leads to impairments of endothelium-dependent dilation in the coronary microcirculation within 24 h, a condition which does not fully dissipate within 168 h. The innovations within the field of nanotechnology, while exciting and novel, can only reach their full potential if toxicity is first properly assessed

Tumor suppressor Tsc1 is a new Hsp90 co-chaperone that facilitates folding of kinase and non-kinase clients

The tumor suppressors Tsc1 and Tsc2 form the tuberous sclerosis complex (TSC), a regulator of mTOR activity. Tsc1 stabilizes Tsc2; however, the precise mechanism involved remains elusive. The molecular chaperone heat-shock protein 90 (Hsp90) is an essen- tial component of the cellular homeostatic machinery in eukary- otes. Here, we show that Tsc1 is a new co-chaperone for Hsp90 that inhibits its ATPase activity. The C-terminal domain of Tsc1 (998–1,164 aa) forms a homodimer and binds to both protomers of the Hsp90 middle domain. This ensures inhibition of both subunits of the Hsp90 dimer and prevents the activating co- chaperone Aha1 from binding the middle domain of Hsp90. Conversely, phosphorylation of Aha1-Y223 increases its affinity for Hsp90 and displaces Tsc1, thereby providing a mechanism for equilibrium between binding of these two co-chaperones to Hsp90. Our findings establish an active role for Tsc1 as a facilita- tor of Hsp90-mediated folding of kinase and non-kinase clients— including Tsc2—thereby preventing their ubiquitination and proteasomal degradation

Expedition 391 Preliminary Report : Walvis Ridge Hotspot: drilling Walvis Ridge, Southeast Atlantic Ocean, to test models of ridge hotspot interaction, isotopic zonation, and the hotspot reference frame

Hotspot tracks (quasilinear chains of seamounts, ridges, and other volcanic structures) provide important records of plate motions, as well as mantle geodynamics, magma flux, and mantle source compositions. The Tristan-Gough-Walvis Ridge (TGW) hotspot track, extending from the active volcanic islands of Tristan da Cunha and Gough through a province of guyots and then along Walvis Ridge to the Etendeka flood basalt province, forms one of the most prominent and complex global hotspot tracks. The TGW hotspot track displays a tight linear age progression in which ages increase from the islands to the flood basalts (covering ~135 My). Unlike Pacific tracks, which are simple chains of seamounts that are often compared to chains of pearls, the TGW track is alternately a steep-sided narrow ridge, an oceanic plateau, subparallel linear ridges and chains of seamounts, and areas of what appear to be randomly dispersed seamounts. The track displays isotopic zonation over the last ~70 My. The zonation appears near the middle of the track just before it splits into two to three chains of ridge- and guyot-type seamounts. The older ridge is also overprinted with age-progressive late-stage volcanism, which was emplaced ~30–40 My after the initial eruptions and has a distinct isotopic composition. The plan for Expedition 391 was to drill at six sites, three along Walvis Ridge and three in the seamount (guyot) province, to gather igneous rocks to better understand the formation of track edifices, the temporal and geochemical evolution of the hotspot, and the variation in paleolatitudes at which the volcanic edifices formed. After a delay of 18 days to address a shipboard outbreak of the coronavirus disease 2019 (COVID-19) virus, Expedition 391 proceeded to drill at four of the proposed sites: three sites on the eastern Walvis Ridge around Valdivia Bank, an ocean plateau within the ridge, and one site on the lower flank of a guyot in the Center track, a ridge located between the Tristan subtrack (which extends from the end of Walvis Ridge to the island of Tristan da Cunha) and the Gough subtrack (which extends from Walvis Ridge to the island of Gough). One hole was drilled at Site U1575, located on a low portion of the northeastern Walvis Ridge north of Valdivia Bank. At this location, 209.9 m of sediments and 122.4 m of igneous basement were cored. The latter comprised 10 submarine lava units consisting of pillow, lobate, sheet, and massive lava flows, the thickest of which was ~21 m. Most lavas are tholeiitic, but some alkalic basalts were recovered. A portion of the igneous succession consists of low-Ti basalts, which are unusual because they appear in the Etendeka flood basalts but have not been previously found on Walvis Ridge. Two holes were drilled at Site U1576 on the west flank of Valdivia Bank. The first hole was terminated because a bit jammed shortly after penetrating igneous basement. Hole U1576A recovered a remarkable ~380 m thick sedimentary section consisting mostly of chalk covering a nearly complete sequence from Paleocene to Late Cretaceous (Campanian). These sediments display short and long cyclic color changes that imply astronomically forced and longer term paleoenvironmental changes. The igneous basement yielded 11 submarine lava units ranging from pillows to massive flows, which have compositions varying from tholeiitic basalt to basaltic andesite, the first occurrence of this composition recovered from the TGW track. These units are separated by seven sedimentary chalk units that range in thickness from 0.1 to 11.6 m, implying a long-term interplay of sedimentation and lava eruptions. Coring at Site U1577, on the extreme eastern flank of Valdivia Bank, penetrated a 154 m thick sedimentary section, the bottom ~108 m of which is Maastrichtian–Campanian (possibly Santonian) chalk with vitric tephra layers. Igneous basement coring progressed only 39.1 m below the sediment-basalt contact, recovering three massive submarine tholeiite basalt lava flows that are 4.1, 15.5, and >19.1 m thick, respectively. Paleomagnetic data from Sites U1577 and U1576 indicate that their volcanic basements formed just before the end of the Cretaceous Normal Superchron and during Chron 33r, shortly afterward, respectively. Biostratigraphic and paleomagnetic data suggest an east–west age progression across Valdivia Bank, becoming younger westward. Site U1578, located on a Center track guyot, provided a long and varied igneous section. After coring through 184.3 m of pelagic carbonate sediments mainly consisting of Eocene and Paleocene chalk, Hole U1578A cored 302.1 m of igneous basement. Basement lavas are largely pillows but are interspersed with sheet and massive flows. Lava compositions are mostly alkalic basalts with some hawaiite. Several intervals contain abundant olivine, and some of the pillow stacks consist of basalt with remarkably high Ti content. The igneous sequence is interrupted by 10 sedimentary interbeds consisting of chalk and volcaniclastics and ranging in thickness from 0.46 to 10.19 m. Paleomagnetic data display a change in basement magnetic polarity ~100 m above the base of the hole. Combining magnetic stratigraphy with biostratigraphic data, the igneous section is inferred to span >1 My. Abundant glass from pillow lava margins was recovered at Sites U1575, U1576, and U1578. Although the igneous penetration was only two-thirds of the planned amount, drilling during Expedition 391 obtained samples that clearly will lead to a deeper understanding of the evolution of the Tristan-Gough hotspot and its track. Relatively fresh basalts with good recovery will provide ample samples for geochemical, geochronologic, and paleomagnetic studies. Good recovery of Late Cretaceous and early Cenozoic chalk successions provides samples for paleoenvironmental study

A Transient Transgenic RNAi Strategy for Rapid Characterization of Gene Function during Embryonic Development

RNA interference (RNAi) is a powerful strategy for studying the phenotypic consequences of reduced gene expression levels in model systems. To develop a method for the rapid characterization of the developmental consequences of gene dysregulation, we tested the use of RNAi for “transient transgenic” knockdown of mRNA in mouse embryos. These methods included lentiviral infection as well as transposition using the Sleeping Beauty (SB) and PiggyBac (PB) transposable element systems. This approach can be useful for phenotypic validation of putative mutant loci, as we demonstrate by confirming that knockdown of Prdm16 phenocopies the ENU-induced cleft palate (CP) mutant, csp1. This strategy is attractive as an alternative to gene targeting in embryonic stem cells, as it is simple and yields phenotypic information in a matter of weeks. Of the three methodologies tested, the PB transposon system produced high numbers of transgenic embryos with the expected phenotype, demonstrating its utility as a screening method

The trans-Golgi SNARE syntaxin 6 is recruited to the chlamydial inclusion membrane

Chlamydia trachomatis is an obligate intracellular pathogen that replicates within a parasitophorous vacuole termed an inclusion. The chlamydial inclusion is isolated from the endocytic pathway but fusogenic with Golgi-derived exocytic vesicles containing sphingomyelin and cholesterol. Sphingolipids are incorporated into the chlamydial cell wall and are considered essential for chlamydial development and viability. The mechanisms by which chlamydiae obtain eukaryotic lipids are poorly understood but require chlamydial protein synthesis and presumably modification of the inclusion membrane to initiate this interaction. A polarized cell model of chlamydial infection has demonstrated that chlamydiae preferentially intercept basolaterally directed, sphingomyelin-containing exocytic vesicles. Here we examine the localization and potential function of trans-Golgi and/or basolaterally associated soluble N-ethylmaleimide-sensitive factor attachment protein receptor (SNARE) proteins in chlamydia-infected cells. The trans-Golgi SNARE protein syntaxin 6 is recruited to the chlamydial inclusion in a manner that requires chlamydial protein synthesis and is conserved among all chlamydial species examined. The localization of syntaxin 6 to the chlamydial inclusion requires a tyrosine motif or plasma membrane retrieval signal (YGRL). Thus in addition to expression of at least two inclusion membrane proteins that contain SNARE-like motifs, chlamydiae also actively recruit eukaryotic SNARE-family proteins

Differential effects of glucagon-like peptide-1 receptor agonists on heart rate

Abstract While glucagon-like peptide-1 receptor agonists (GLP-1 RAs) are known to increase heart rate (HR), it is insufficiently recognized that the extent varies greatly between the various agonists and is affected by the assessment methods employed. Here we review published data from 24-h time-averaged HR monitoring in healthy individuals and subjects with type 2 diabetes mellitus (T2DM) treated with either short-acting GLP-1 RAs, lixisenatide or exenatide, or long-acting GLP-1 RAs, exenatide LAR, liraglutide, albiglutide, or dulaglutide (N\ua0=\ua01112; active-treatment arms). HR effects observed in two independent head-to-head trials of lixisenatide and liraglutide (N\ua0=\ua0202; active-treatment arms) are also reviewed. Short-acting GLP-1 RAs, exenatide and lixisenatide, are associated with a transient (1\u201312\ua0h) mean placebo- and baseline-adjusted 24-h HR increase of 1\u20133\ua0beats per minute (bpm). Conversely, long-acting GLP-1 RAs are associated with more pronounced increases in mean 24-h HR; the highest seen with liraglutide and albiglutide at 6\u201310\ua0bpm compared with dulaglutide and exenatide LAR at 3\u20134\ua0bpm. For both liraglutide and dulaglutide, HR increases were recorded during both the day and at night. In two head-to-head comparisons, a small, transient mean increase in HR from baseline was observed with lixisenatide; liraglutide induced a substantially greater increase that remained significantly elevated over 24\ua0h. The underlying mechanism for increased HR remains to be elucidated; however, it could be related to a direct effect at the sinus node and/or stimulation of the sympathetic nervous system, with this effect related to the duration of action of the respective GLP-1 RAs. In conclusion, this review indicates that the effects on HR differ within the class of GLP-1 RAs: short-acting GLP-1 RAs are associated with a modest and transient HR increase before returning to baseline levels, while some long-acting GLP-1 RAs are associated with a more pronounced and sustained increase during the day and night. Findings from recently completed trials indicate that a GLP-1 RA-induced increase in HR, regardless of magnitude, does not present an increased cardiovascular risk for subjects with T2DM, although a pronounced increase in HR may be associated with adverse clinical outcomes in those with advanced heart failure

Novel Methods for Analysing Bacterial Tracks Reveal Persistence in Rhodobacter sphaeroides

Tracking bacteria using video microscopy is a powerful experimental approach to probe their motile behaviour. The trajectories obtained contain much information relating to the complex patterns of bacterial motility. However, methods for the quantitative analysis of such data are limited. Most swimming bacteria move in approximately straight lines, interspersed with random reorientation phases. It is therefore necessary to segment observed tracks into swimming and reorientation phases to extract useful statistics. We present novel robust analysis tools to discern these two phases in tracks. Our methods comprise a simple and effective protocol for removing spurious tracks from tracking datasets, followed by analysis based on a two-state hidden Markov model, taking advantage of the availability of mutant strains that exhibit swimming-only or reorientating-only motion to generate an empirical prior distribution. Using simulated tracks with varying levels of added noise, we validate our methods and compare them with an existing heuristic method. To our knowledge this is the first example of a systematic assessment of analysis methods in this field. The new methods are substantially more robust to noise and introduce less systematic bias than the heuristic method. We apply our methods to tracks obtained from the bacterial species Rhodobacter sphaeroides and Escherichia coli. Our results demonstrate that R. sphaeroides exhibits persistence over the course of a tumbling event, which is a novel result with important implications in the study of this and similar species

Self-Administered Intranasal Etripamil Using a Symptom-Prompted, Repeat-Dose Regimen for Atrioventricular-Nodal-Dependent Supraventricular Tachycardia (RAPID): A Multicentre, Randomised Trial

BACKGROUND: Etripamil is a fast-acting, intranasally administered calcium-channel blocker in development for on-demand therapy outside a health-care setting for paroxysmal supraventricular tachycardia. We aimed to evaluate the efficacy and safety of etripamil 70 mg nasal spray using a symptom-prompted, repeat-dose regimen for acute conversion of atrioventricular-nodal-dependent paroxysmal supraventricular tachycardia to sinus rhythm within 30 min. METHODS: RAPID was a multicentre, randomised, placebo-controlled, event-driven trial, conducted at 160 sites in North America and Europe as part 2 of the NODE-301 study. Eligible patients were aged at least 18 years and had a history of paroxysmal supraventricular tachycardia with sustained, symptomatic episodes (≥20 min) as documented by electrocardiogram. Patients were administered two test doses of intranasal etripamil (each 70 mg, 10 min apart) during sinus rhythm; those who tolerated the test doses were randomly assigned (1:1) using an interactive response technology system to receive either etripamil or placebo. Prompted by symptoms of paroxysmal supraventricular tachycardia, patients self-administered a first dose of intranasal 70 mg etripamil or placebo and, if symptoms persisted beyond 10 min, a repeat dose. Continuously recorded electrocardiographic data were adjudicated, by individuals masked to patient assignment, for the primary endpoint of time to conversion of paroxysmal supraventricular tachycardia to sinus rhythm for at least 30 s within 30 min after the first dose, which was measured in all patients who administered blinded study drug for a confirmed atrioventricular-nodal-dependent event. Safety outcomes were assessed in all patients who self-administered blinded study drug for an episode of perceived paroxysmal supraventricular tachycardia. This trial is registered at ClinicalTrials.gov, NCT03464019, and is complete. FINDINGS: Between Oct 13, 2020, and July 20, 2022, among 692 patients randomly assigned, 184 (99 from the etripamil group and 85 from the placebo group) self-administered study drug for atrioventricular-nodal-dependent paroxysmal supraventricular tachycardia, with diagnosis and timing confirmed. Kaplan-Meier estimates of conversion rates by 30 min were 64% (63/99) with etripamil and 31% (26/85) with placebo (hazard ratio 2·62; 95% CI 1·66-4·15; p INTERPRETATION: Using a symptom-prompted, self-administered, initial and optional-repeat-dosing regimen, intranasal etripamil was well tolerated, safe, and superior to placebo for the rapid conversion of atrioventricular-nodal-dependent paroxysmal supraventricular tachycardia to sinus rhythm. This approach could empower patients to treat paroxysmal supraventricular tachycardia themselves outside of a health-care setting, and has the potential to reduce the need for additional medical interventions, such as intravenous medications given in an acute-care setting. FUNDING: Milestone Pharmaceuticals

Phylogenomic analysis of the Chlamydomonas genome unmasks proteins potentially involved in photosynthetic function and regulation

Chlamydomonas reinhardtii, a unicellular green alga, has been exploited as a reference organism for identifying proteins and activities associated with the photosynthetic apparatus and the functioning of chloroplasts. Recently, the full genome sequence of Chlamydomonas was generated and a set of gene models, representing all genes on the genome, was developed. Using these gene models, and gene models developed for the genomes of other organisms, a phylogenomic, comparative analysis was performed to identify proteins encoded on the Chlamydomonas genome which were likely involved in chloroplast functions (or specifically associated with the green algal lineage); this set of proteins has been designated the GreenCut. Further analyses of those GreenCut proteins with uncharacterized functions and the generation of mutant strains aberrant for these proteins are beginning to unmask new layers of functionality/regulation that are integrated into the workings of the photosynthetic apparatus