Integrated In-Plane Nanofluidic Devices for Resistive-Pulse Sensing.

Single-entity (or digital) measurements enhance sensitivity (10- to 100-fold improvement) and uncover heterogeneity within a population (one event in 100 to 10,000). Many biological systems are significantly influenced by rare or infrequent events, and determining what species is present, in what quantity, and role of that species is critically important to unraveling many questions. To develop these measurement systems, resistive-pulse sensing is used as a label-free, single-particle detection technique and can be combined with a range of functional elements, e.g., mixers, reactors, filters, separators, and pores. Virtually, any two-dimensional layout of the micro- and nanofluidic conduits can be envisioned, designed, and fabricated in the plane of the device. Multiple nanopores in series lead to higher-precision measurements of particle size, shape, and charge, and reactions coupled directly with the particle-size measurements improve temporal response. Moreover, other detection techniques, e.g., fluorescence, are highly compatible with the in-plane format. These integrated in-plane nanofluidic devices expand the toolbox of what is possible with single-entity measurements.

Oxidative Dearomatization of Pyridines.

Dearomatization of pyridines is a well-established synthetic approach to access piperidines. Although remarkably powerful, existing dearomatization processes have been limited to the hydrogenation or addition of carbon-based nucleophiles to activated pyridiniums. Here, we show that arenophile-mediated dearomatizations can be applied to pyridines to directly introduce heteroatom functionalities without prior substrate activation. The arenophile platform in combination with olefin oxidation chemistry provides access to dihydropyridine cis-diols and epoxides. These previously elusive compounds are now readily accessible and can be used for the downstream preparation of diversely functionalized piperidines.

The Lymphoma Epidemiology of Outcomes cohort study: Design, baseline characteristics, and early outcomes.

To address the current and long-term unmet health needs of the growing population of non-Hodgkin lymphoma (NHL) patients, we established the Lymphoma Epidemiology of Outcomes (LEO) cohort study (NCT02736357; https://leocohort.org/). A total of 7735 newly diagnosed patients aged 18 years and older with NHL were prospectively enrolled from 7/1/2015 to 5/31/2020 at 8 academic centers in the United States. The median age at diagnosis was 62 years (range, 18-99). Participants came from 49 US states and included 538 Black/African-Americans (AA), 822 Hispanics (regardless of race), 3386 women, 716 age <40 years, and 1513 rural residents. At study baseline, we abstracted clinical, pathology, and treatment data; banked serum/plasma (N = 5883, 76.0%) and germline DNA (N = 5465, 70.7%); constructed tissue microarrays for four major NHL subtypes (N = 1189); and collected quality of life (N = 5281, 68.3%) and epidemiologic risk factor (N = 4489, 58.0%) data. Through August 2022, there were 1492 deaths. Compared to population-based SEER data (2015-2019), LEO participants had a similar distribution of gender, AA race, Hispanic ethnicity, and NHL subtype, while LEO was underrepresented for patients who were Asian and aged 80 years and above. Observed overall survival rates for LEO at 1 and 2 years were similar to population-based SEER rates for indolent B-cell (follicular and marginal zone) and T-cell lymphomas, but were 10%-15% higher than SEER rates for aggressive B-cell subtypes (diffuse large B-cell and mantle cell). The LEO cohort is a robust and comprehensive national resource to address the role of clinical, tumor, host genetic, epidemiologic, and other biologic factors in NHL prognosis and survivorship.

Integrating Battlefield Documentation into Virtual Reality Medical Simulation Training: Virtual Battlefield Assisted Trauma Distributed Observation Kit (BATDOK).

INTRODUCTION: Clinical documentation is an essential component of the provision of medical care, enabling continuity of information across provider and site handoffs. This is particularly important in the combat casualty care setting when a single casualty may be treated by four or more or five completely disparate teams across the roles of care. The Battlefield Assisted Trauma Distributed Observation Kit (BATDOK) is a digital battlefield clinical documentation system developed by the Air Force Research Laboratory to address this need. To support the deployment of this tool, we integrated BATDOK into a commercially available virtual reality (VR) medical simulation platform used by the U.S. Air Force and Defense Health Agency personnel in order to provide an immersive simulation training experience which included battlefield documentation. METHODS: A multidisciplinary team consisting of medical educators, VR simulation engineers, emergency physicians and pararescuemen, and BATDOK developers first developed a specification for a virtual BATDOK capability, including a detailed listing of learning objectives, critical interfaces and task plans, and sensor integrations. These specifications were then implemented into the commercially available Virtual Advancement of Learning for Operational Readiness VR Medical Simulation System and underwent developmental testing and evaluation during pararescueman training exercises at the Air Force Special Operations Command Special Operations Center for Medical Integration and Development. RESULTS AND CONCLUSIONS: The BATDOK capability was successfully implemented within the VR Medical Simulation System. The capability consisted of a virtual tablet with replicated interfaces and capabilities based on the developed specifications. These capabilities included integrated point-of-care ultrasound capability, multi-patient management, vitals sign monitoring with sensor pairing and continuous monitoring, mechanism of injury documentation (including injury pattern documentation), intervention logging (including tourniquets, dressing, airways, lines, tubes and drains, splints, fluids, and medications), and event logging. The capability was found to be operational and in alignment with learning objectives and user acceptance goals.

CRISPR/Cas9 mutagenesis of the Arabidopsis GROWTH-REGULATING FACTOR (GRF) gene family.

Genome editing in plants typically relies on T-DNA plasmids that are mobilized by Agrobacterium-mediated transformation to deliver the CRISPR/Cas machinery. Here, we introduce a series of CRISPR/Cas9 T-DNA vectors for minimal settings, such as teaching labs. Gene-specific targeting sequences can be inserted as annealed short oligonucleotides in a single straightforward cloning step. Fluorescent markers expressed in mature seeds enable reliable selection of transgenic or transgene-free individuals using a combination of inexpensive LED lamps and colored-glass alternative filters. Testing these tools on the Arabidopsis GROWTH-REGULATING FACTOR (GRF) genes, we were able to create a collection of predicted null mutations in all nine family members with little effort. We then explored the effects of simultaneously targeting two, four and eight GRF genes on the rate of induced mutations at each target locus. In our hands, multiplexing was associated with pronounced disparities: while mutation rates at some loci remained consistently high, mutation rates at other loci dropped dramatically with increasing number of single guide RNA species, thereby preventing a systematic mutagenesis of the family.

Characterization of Extracellular Vesicles by Resistive-Pulse Sensing on In-Plane Multipore Nanofluidic Devices.

Extracellular vesicles (EVs) are cell-derived, naturally produced, membrane-bound nanoscale particles that are linked to cell-cell communication and the propagation of diseases. Here, we report the design and testing of in-plane nanofluidic devices for resistive-pulse measurements of EVs derived from bovine milk and human breast cancer cells. The devices were fabricated in plane with three nanopores in series to determine the particle volume and diameter, two pore-to-pore regions to measure the electrophoretic mobility and zeta potential, and an in-line filter to prevent cellular debris and aggregates from entering the nanopore region. Devices were tested with and without the channels coated with a short-chain PEG silane to minimize electroosmotic flow and permit an accurate measurement of the electrophoretic mobility and zeta potential of the EVs. To enhance throughput of EVs, vacuum was applied to the waste reservoir to increase particle frequencies up to 1000 min-1. The nanopores had cross-sections 200 nm wide and 200 nm deep and easily resolved EV diameters from 60 to 160 nm. EVs from bovine milk and human breast cancer cells had similar particle size distributions, but their zeta potentials differed by 2-fold, -8 ± 1 and -4 ± 1 mV, respectively.

Recurrent petit mal seizures in Erdheim-Chester disease mimicking an intra-axial brain tumor: illustrative case.

BACKGROUND: Erdheim-Chester disease (ECD) is a rare non-Langerhans cell histiocytosis characterized histologically by foamy histiocytes and Touton giant cells in a background of fibrosis. Bone pain with long bone osteosclerosis is highly specific for ECD. Central nervous system involvement is rare, although dural, hypothalamic, cerebellar, brainstem, and sellar region involvement has been described. OBSERVATIONS: A 59-year-old man with a history of ureteral obstruction, medically managed petit mal seizures, and a left temporal lesion followed with serial magnetic resonance imaging (MRI) presented with worsening seizure control. Repeat MRI identified bilateral amygdala region lesions. Gradual growth of the left temporal lesion over 1 year with increasing seizure frequency prompted resection. A non-Langerhans cell histiocytosis with a BRAF V600E mutation was identified on pathology. Imaging findings demonstrated retroperitoneal fibrosis and long bone osteosclerosis with increased fluorodeoxyglucose uptake that, together with the neuropathologic findings, were diagnostic of ECD. LESSONS: This case of biopsy-proven ECD is unique in that the singular symptom was seizures well controlled with medical management in the presence of similarly located bilateral anterior mesial temporal lobe lesions. Although ECD is rare intracranially, its variable imaging presentation, including the potential to mimic seizure-associated medial temporal lobe tumors, emphasizes the need for a wide differential diagnosis.

Role of omega-3 endocannabinoids in the modulation of T-cell activity in a multiple sclerosis experimental autoimmune encephalomyelitis (EAE) model.

Epidemiological studies show that omega-3 fatty acid consumption is associated with improved conditions in neurodegenerative diseases such as multiple sclerosis (MS). However, the mechanism of this association is not well understood. Emerging evidence suggests that parent molecules such as docosahexaenoic acid are converted into downstream metabolites that are capable of directly modulating immune responses. In vitro, we found that docosahexaenoyl ethanolamide (DHEA), another dietary component and its epoxide metabolite, reduced the polarization of naïve T-cells toward proinflammatory Th1 and Th17 phenotypes. Furthermore, we identified that DHEA and related endocannabinoids are changing during the disease progression in mice undergoing relapse-remitting experimental autoimmune encephalomyelitis (RR-EAE). In addition, daily administration of DHEA to mice delayed the onset of disease, the rate of relapse, and the severity of clinical scores at relapse in RR-EAE, an animal model of MS. Collectively, these data indicate that DHEA and their downstream metabolites reduce the disease severity in the RR-EAE model of MS and can be potential dietary adjuvants to existing MS therapeutics.

Amaryllidaceae Alkaloids Decrease the Proliferation, Invasion, and Secretion of Clinically Relevant Cytokines by Cultured Human Colon Cancer Cells.

Alkaloids isolated from members of the Amaryllidaceae plant family are promising anticancer agents. The purpose of the current study was to determine if the isocarbostyrils narciclasine, pancratistatin, lycorane, lycorine, crinane, and haemanthamine inhibit phenomena related to cancer progression in vitro. To achieve this, we examined the proliferation, adhesion, and invasion of cultured human colon cancer cells via MTT assay and Matrigel-coated Boyden chambers. In addition, Luminex assays were used to quantify the secretion of matrix metalloproteinases (MMP) and cytokines associated with poor clinical outcomes. We found that all alkaloids decreased cell proliferation regardless of TP53 status, with narciclasine exhibiting the greatest potency. The effects on cell proliferation also appear to be specific to cancer cells. Narciclasine, lycorine, and haemanthamine decrease both adhesion and invasion but with various potencies depending on the cell line. In addition, narciclasine, lycorine, and haemanthamine decreased the secretion of MMP-1, -2, and -7, as well as the secretion of the cytokines pentraxin 3 and vascular endothelial growth factor. In conclusion, the present study shows that Amaryllidaceae alkaloids decrease phenomena and cytokines associated with colorectal cancer progression, supporting future investigations regarding their potential as multifaceted drug candidates.

Dearomative Ring Expansion of Polycyclic Arenes.

Benzocycloheptenes constitute a common structural motif embedded in many natural products and biologically active compounds. Herein, we report their concise preparation from non-activated polycyclic arenes using a two-step sequence involving dearomative [4+2]-cycloaddition with arenophile in combination with palladium-catalyzed cyclopropanation, followed by cycloreversion-initiated ring expansion. The described strategy provides a working alternative to the Buchner reaction, which is limited to monocyclic arenes. Overall, this methylene-insertion molecular editing approach enables rapid and direct conversion of simple (hetero)arenes into a range of substituted (aza)benzocycloheptatrienes, which can undergo a myriad of downstream functionalizations.

Reactivity-dependent profiling of RNA 5-methylcytidine dioxygenases.

Epitranscriptomic RNA modifications can regulate fundamental biological processes, but we lack approaches to map modification sites and probe writer enzymes. Here we present a chemoproteomic strategy to characterize RNA 5-methylcytidine (m5C) dioxygenase enzymes in their native context based upon metabolic labeling and activity-based crosslinking with 5-ethynylcytidine (5-EC). We profile m5C dioxygenases in human cells including ALKBH1 and TET2 and show that ALKBH1 is the major hm5C- and f5C-forming enzyme in RNA. Further, we map ALKBH1 modification sites transcriptome-wide using 5-EC-iCLIP and ARP-based sequencing to identify ALKBH1-dependent m5C oxidation in a variety of tRNAs and mRNAs and analyze ALKBH1 substrate specificity in vitro. We also apply targeted pyridine borane-mediated sequencing to measure f5C sites on select tRNA. Finally, we show that f5C at the wobble position of tRNA-Leu-CAA plays a role in decoding Leu codons under stress. Our work provides powerful chemical approaches for studying RNA m5C dioxygenases and mapping oxidative m5C modifications and reveals the existence of novel epitranscriptomic pathways for regulating RNA function.

Nutritional and physico-chemical implications of avocado meal as a novel dietary fiber source in an extruded canine diet.

This study assessed the effects of a diet containing avocado meal (AMD), an underutilized by-product avocado oil processing, on apparent total tract digestibility (ATTD) and fecal fermentative end-products when compared with beet pulp (BPD) and cellulose (CD) diets targeting 15% total dietary fiber (TDF). The concentration of persin, a natural fungicidal toxin present in avocado, was also determined on several parts of the fruit and avocado meal. Nine intact female beagles (4.9 ± 0.6 yr and 11.98 ± 1.76 kg) were randomly grouped in a 3 × 3 replicated Latin square design. Periods were 14 d long, with 10 d of adaptation followed by 4 d of total fecal and urine collection for apparent total tract digestibility (ATTD) calculations. Fresh fecals were analyzed for fermentative end-products. The BPD (87.0 g/d) caused higher (P < 0.05) fecal output (as-is basis) than AMD (62.3 g/d) and CD (58.0 g/d). Fecal score for the BPD (3.1) was greater (P < 0.05) than for AMD (2.8) or CD (2.6). Acid-hydrolyzed fat ATTD was lower (P < 0.05) for the BPD (94.1%) than for the AMD (95.5%) and CD (95.7%). Crude protein ATTD was greater (P < 0.05) for the CD (88.5%) than the AMD (82.2%) or BPD (83.7%). Dogs fed AMD (49.9%) or BPD (51.0%) exhibited greater (P < 0.05) TDF ATTD than CD. The fermentative profile for the AMD (233.4, 70.9, 8.8, and 12.0 µmole/g DM, respectively) was similar (P > 0.05) to the CD (132.9, 61.7, 7.5, and 9.5 µmole/g DM, respectively) profile, with lower (P < 0.05) concentrations of acetate and propionate and higher (P < 0.05) concentrations of isovalerate and indoles compared to the BPD. Dogs fed AMD (47.0 µmole/g DM) or BPD (54.2 µmole/g DM) exhibited similar (P > 0.05) fecal butyrate concentrations greater (P < 0.05) than for CD (24.7 µmole/g DM). Given these results, avocado meal appears to be an adequate dietary fiber source when compared with traditional fiber sources used in canine diets. No health adverse effects were observed in dogs fed extruded diet containing as much as 18% of avocado meal (as-is basis).

Computational and experimental optimization of 135Xe production in calibration sources.

Here we present a new method of irradiating 134Xe capsules to produce 135Xe gas standards which maximize the ratio of 135Xe to 133Xe production due to (n,g) and (n,2n) reactions, respectively. We performed "Spectral tuning" of the University of Utah TRIGA Reactor (UUTR) neutron spectrum to increase the length of time that 135Xe dominates undesirable 135Xe in the sample, so that the capsules - used for calibration and quality control testing of Xe gas detection equipment in support of the Comprehensive Test Ban Treaty (CTBT) - will remain viable for longer periods post-irradiation. Moreover, optimized methods of computation and analysis were developed yielding improved computational efficiency over standard Monte Carlo approaches. These methods provided valuable insight into the final design and manufacture of new, ex-core Teflon irradiation chambers tested in the UUTR. The methods of computation and analysis, as well as the physical irradiation chamber designs, were derived such that they could be readily applied to any reactor for spectral tuning of a specific reactor's flux profile. Results of the physical experiments employing the optimized irradiation chamber designs demonstrated sample viability time improvements of over 60% when compared to conventional, un-optimized methods of gas sample generation. Thus, use of these methods enhance both the CTBT-related calibrations and performance testing, and the continued stability of the CTBT monitoring network overall.

COVID-19 and Lung Cavitation: A Clue to Pathogenesis?

Description Lung cavitation as a complication of COVID-19 is rare. A 56-year-old male presented with lung cavitation, small volume hemoptysis, and violaceous discoloration of the right great toe, 5 weeks after diagnosis with COVID-19 pneumonia. The digital changes were consistent with previously described microvascular changes called "COVID toe." CT angiography of the chest was negative for pulmonary embolism but showed a 2.5 x 3.1 x 2.2 cm cavitation within the right lung. Extensive evaluation for commonly implicated infectious and autoimmune causes was negative. We concluded that the cavitary lung lesions were likely a complication of COVID-19 pneumonia and may implicate microangiopathy as an important component of pathogenesis. This case highlights a rare complication of COVID-19 of which clinicians should be aware.

Inadequate Metadiaphyseal Fill of a Modern Taper-Wedge Stem Increases Subsidence and Risk of Aseptic Loosening: Technique and Distal Canal Fill Matter!

BACKGROUND: Adequate interference fit and mechanical stability through optimal surgical technique are essential to prevent subsidence and loosening in cementless total hip arthroplasty. The purpose of this study is to determine the effect of surgical technique on radiographic subsidence and subsequent stability of a modern taper-wedge cementless stem. METHODS: A retrospective review of 250 consecutive cementless primary total hip arthroplasties performed by 2 surgeons was completed. Surgeon A vigorously broached, maximizing the mediolateral stem dimension and confirmed final broach stability with a torsional test, whereas Surgeon B did not. All patients received identical taper-wedge stems. Preoperative bone morphology (canal flare index), postoperative subsidence, and canal fill were radiographically assessed. RESULTS: Canal flare index was not different between groups (P = .747). There was significantly less subsidence at 1 month for Surgeon A (0.3 vs 1.3 mm, P < .001). Additional subsidence at 1 year occurred in only 0.8% of Surgeon A (1/119) compared to 51.6% of Surgeon B stems (33/64, P < .001). Surgeon technique and canal fill measured at 60 mm below the lesser trochanter were the only variables predictive for subsidence, where Surgeon A and B had a mean canal fill of 95% and 86%, respectively. Surgeon B had 2 cases of aseptic loosening (2%) at 2 and 3 years postoperatively. CONCLUSION: These observations support that maximizing mediolateral canal fill and avoiding under-sizing the femoral implant with meticulous broaching technique minimizes subsidence and optimizes stability of modern cementless taper-wedge stems. Failure to optimize canal fill with appropriate broaching and surgical technique may predispose femoral components to failure from aseptic loosening.

Development of a Scalable and Sublimation-Free Route to MTAD.

The cyclic azodicarbonyl 4-methyl-1,2,4-triazoline-3,5-dione (MTAD) is a versatile and powerful reagent used mainly in cycloaddition chemistry. Though known for more than 50 years, its unsafe preparation, as well as purification by sublimation, hampered its widespread applicability on a larger scale. Herein we report a scalable and safe route to MTAD, which avoids the generation of methyl isocyanate. Moreover, we demonstrate that sublimation can be circumvented by the application of judicious oxidation conditions, followed by simple filtration. Overall, up to 25 g of MTAD was prepared in a single batch from commercial starting materials in three steps, with recrystallization serving as the only purification in the sequence. When employed in dearomative methodologies, the MTAD obtained by this protocol displayed synthetic efficiency equivalent to that of MTAD purified by sublimation.

Comparative Analysis of LASIK Flap Diameter and its Centration Using Two Different Femtosecond Lasers.

The aim of this study was to compare the diameter, accuracy, variability, and centration with respect to the limbus of corneal flaps created by two femtosecond lasers, the VisuMax, and Wavelight FS200, for laser in situ keratomileusis (LASIK) and how these flaps affect visual outcomes. This is a retrospective chart review of flap morphology created during LASIK Surgery. Overall, 168 eyes underwent flap creation using the WaveLight FS200 laser, and on 189 eyes, the VisuMax laser was used. Of these total number, flap morphology was analyzed in a random sample of 158 eyes; 80 with the Visumax laser and 78 with the WaveLight FS200 laser. Intraoperative photos of the flaps taken by the Wavelight Allegretto EX500 were analyzed. Flap diameters and centration were measured using Adobe Acrobat Pro. All patients had visual acuity measurements including uncorrected distance visual acuity (UDVA), corrected distance visual acuity (CDVA), spherical equivalent refraction (SE) and refractive astigmatism recorded three months postoperatively. Greater than 90% of patients in both groups achieved a UDVA of 20/20 postoperatively. The mean difference between targeted and achieved flap diameter was 0.50 +/- 0.15 mm in the VisuMax group and 0.35 +/- 0.15 millimeters (mm) in the FS200 group (P<0.01). The flap diameters of the VisuMax group were more precise with a variance of 0.024 mm compared to a variance of 0.038 mm in the FS200 group (P<0.05). VisuMax flaps were more nasally displaced (log(NA/TA) = -0.21 +/- 0.10 mm) compared to the FS200 flaps (log(NA/TA) = 0.03 +/- 0.10 mm), (P< 0.01). We concluded that both the VisuMax and FS200 created flaps larger than the preoperative targeted diameter. VisuMax created corneal flaps that had a greater degree of deviation from the targeted diameter when compared to flaps from the FS200. However, there was less variance in the VisuMax flap diameter. In addition, VisuMax flaps were more nasally displaced. There were no statistically significant differences in visual outcomes when comparing the two femtosecond lasers.

Astigmatic correction with implantation of a light adjustable vs monofocal lens: a single site analysis of a randomized controlled trial.

AIM: To evaluate the light adjustable lens (LAL) vs a standard monofocal lens in achieving target astigmatic refraction and improving postoperative uncorrected distance visual acuity (UDVA). METHODS: This randomized controlled clinical trial included 40 patients with pre-existing astigmatism and visually significant cataract. Twenty-eight patients received the LAL and 12 control patients received a monofocal intraocular lens (IOL) after cataract extraction at a single institution. The patients with the LAL underwent adjustment by ultraviolet (UV) light postoperatively plus subsequent lock-in procedures and all patients returned to clinic for follow up of study parameters at 6, 9, and 12mo. Manifest refraction, distance visual acuity, and adverse events were recorded at each visit. RESULTS: The mean cylinder before adjustment in eyes with the LAL was -0.89±0.58 D (-2.00 to 0.00 D) and -0.34±0.34 D (-1.25 to 0.00 D) after lock-in (P=1.68x10-8). The mean cylinder in patients with the monofocal lens was -1.00±0.32 D (-1.50 to -0.50 D) at 17-21d postoperatively, which was statistically different from the LAL cylinder post lock-in (P=1.43x10-6). UDVA in the LAL group was 20/20 or better in 79% of patients post lock-in with good stability over 12mo compared with 33% of the control patients with UDVA of 20/20 or better. CONCLUSION: These results demonstrate that the LAL is more effective in achieving target refractions and improving postoperative UDVA in patients with pre-existing corneal astigmatism than a standard monofocal lens.

A paradigm shift in eye banking: how new models are challenging the status quo.

This article presents a fact-based, direct approach in order to evaluate the strengths and short comings of both local eye banks and larger companies that have penetrated the market of eye donation. By taking a non-biased approach to the topic, the goal of this article is to create a platform, which furthers the discussion. We focus on SightLife and CorneaGen as well as local eye banks and how their models differ in terms of tissue procurement, distribution, and surgeon partnership.