Single-Cell Classification Based on Population Nucleus Size Combining Microwave Impedance Spectroscopy and Machine Learning.

Many recent efforts in the diagnostic field address the accessibility of cancer diagnosis. Typical histological staining methods identify cancer cells visually by a larger nucleus with more condensed chromatin. Machine learning (ML) has been incorporated into image analysis for improving this process. Recently, impedance spectrometers have been shown to generate all-inclusive lab-on-a-chip platforms to detect nucleus abnormities. In this paper, a wideband electrical sensor and data analysis paradigm that can identify nuclear changes shows the realization of a single-cell microfluidic device to detect nuclei of altered sizes. To model cells of altered nucleus, Jurkat cells were treated to enlarge or shrink their nucleus followed by broadband sensing to obtain the S-parameters of single cells. The ability to deduce important frequencies associated with nucleus size is demonstrated and used to improve classification models in both binary and multiclass scenarios, despite a heterogeneous and overlapping cell population. The important frequency features match those predicted in a double-shell circuit model published in prior work, demonstrating a coherent new analytical technique for electrical data analysis. The electrical sensing platform assisted by ML with impressive accuracy of cell classification looks forward to a label-free and flexible approach to cancer diagnosis.

Broadband Electrical Sensing of a Live Biological Cell with In Situ Single-Connection Calibration.

Single-connection in situ calibration using biocompatible solutions is demonstrated in single-cell sensing from 0.5 to 9 GHz. The sensing is based on quickly trapping and releasing a live cell by dielectrophoresis on a coplanar transmission line with a little protrusion in one of its ground electrodes. The same transmission line is used as the calibration standard when covered by various solutions of known permittivities. The results show that the calibration technique may be precise enough to differentiate cells of different nucleus sizes, despite the measured difference being less than 0.01 dB in the deembedded scattering parameters. With better accuracy and throughput, the calibration technique may allow broadband electrical sensing of live cells in a high-throughput cytometer.

Multicenter phase 1 trial of intraventricular immunochemotherapy in recurrent CNS lymphoma.

UNLABELLED: Recurrent CNS lymphoma continues to be associated with poor outcomes in the rituximab era. Although IV rituximab mediates superior disease control of systemic non-Hodgkin lymphoma (NHL), it fails to completely eliminate the risk of meningeal recurrence, likely due to minimal CNS penetration. Given that rituximab acts synergistically with chemotherapy, we conducted the first phase 1 study of intraventricular immunochemotherapy in patients with recurrent CNS NHL. Fourteen patients received 10 mg or 25 mg intraventricular rituximab twice weekly for 4 weeks, with rituximab administered as monotherapy during the first treatment each week and rituximab administered in combination with methotrexate (MTX) during the second treatment each week. More than 150 doses were administered without serious toxicity. In a population with high-refractory CNS NHL, 75% of patients achieved complete cytologic responses and 43% achieved an overall complete response in CSF and/or brain parenchyma. Two patients achieved a first complete response of CNS NHL with intraventricular rituximab/MTX, including 1 with CNS lymphoma refractory to high-dose systemic and intrathecal MTX plus IV rituximab. We conclude that intraventricular rituximab in combination with MTX is feasible and highly active in the treatment of drug-resistant CNS NHL that is refractory or unresponsive to IV rituximab. KEY POINTS: Phase I study showed that intraventricular rituximab plus methotrexate is feasible and active in the treatment of refractory CNS lymphoma.

Biomechanical characteristics of the horizontal mattress stitch: implication for double-row and suture-bridge rotator cuff repair.

BACKGROUND: We investigated the effects of bite-size horizontal mattress stitch (distance between the limbs passed through the tendon) on the biomechanical properties of the repaired tendon. METHODS: We anchored 20 bovine Achilles tendons to bone using no. 2 high-strength suture and 5-mm titanium suture anchors in a mattress-suture technique. Tendons were allocated randomly into two groups of ten each to receive stitches with a 4- or 10-mm bite. Specimens underwent cyclic loading from 5 to 30 N at 1 mm/s for 30 cycles, followed by tensile testing to failure. Gap formation, tendon strain, hysteresis, stiffness, yield load, ultimate load, energy to yield load, and energy to ultimate load were compared between groups using unpaired t tests. RESULTS: The 4-mm group had less (p < 0.05) gap formation and less (p < 0.05) longitudinal strain than did the 10-mm group. Ultimate load (293.6 vs. 148.9 N) and energy to ultimate load (2,563 vs. 1,472 N-mm) were greater (p < 0.001) for the 10-mm group than the 4-mm group. All tendons repaired with 4-mm suturing failed at the suture-tendon interface, with sutures pulling through the tendon, whereas the suture itself failed before the tendon did in seven of the ten specimens in the 10-mm group. CONCLUSIONS: Whereas a 4-mm bite fixed the tendon more tightly but at the cost of decreased ultimate strength, a 10-mm bite conveyed greater ultimate strength but with increased gap and strain. These results suggest that for the conventional double-row repair, small mattress stitches provide a tighter repair, whereas large stitches are beneficial to prevent sutures from pulling through the tendon after surgery. For suture-bridge rotator cuff repair, large stitches are beneficial because the repaired tendon has a higher strength, and the slightly mobile medial knot can be tightened by lateral fixation.

Mind the gap: the relevance of the genome reference to resolve rare and pathogenic inversions.

Both long-read genome sequencing (lrGS) and the recently published Telomere to Telomere (T2T) reference genome provide increased coverage and resolution across repetitive regions promising heightened structural variant detection and improved mapping. Inversions (INV), intrachromosomal segments which are rotated 180° and inserted back into the same chromosome, are a class of structural variants particularly challenging to detect due to their copy-number neutral state and association with repetitive regions. Inversions represent about 1/20 of all balanced structural chromosome aberrations and can lead to disease by gene disruption or altering regulatory regions of dosage sensitive genes in cis . Here we remapped the genome data from six individuals carrying unsolved cytogenetically detected inversions. An INV6 and INV10 were resolved using GRCh38 and T2T-CHM13. Finally, an INV9 required optical genome mapping, de novo assembly of lrGS data and T2T-CHM13. This inversion disrupted intron 25 of EHMT1, confirming a diagnosis of Kleefstra syndrome 1 (MIM#610253). These three inversions, only mappable in specific references, prompted us to investigate the presence and population frequencies of differential reference regions (DRRs) between T2T-CHM13, GRCh37, GRCh38, the chimpanzee and bonobo, and hundreds of megabases of DRRs were identified. Our results emphasize the significance of the chosen reference genome and the added benefits of lrGS and optical genome mapping in solving rearrangements in challenging regions of the genome. This is particularly important for inversions and may impact clinical diagnostics.

Inverted triplications formed by iterative template switches generate structural variant diversity at genomic disorder loci.

The duplication-triplication/inverted-duplication (DUP-TRP/INV-DUP) structure is a complex genomic rearrangement (CGR). Although it has been identified as an important pathogenic DNA mutation signature in genomic disorders and cancer genomes, its architecture remains unresolved. Here, we studied the genomic architecture of DUP-TRP/INV-DUP by investigating the DNA of 24 patients identified by array comparative genomic hybridization (aCGH) on whom we found evidence for the existence of 4 out of 4 predicted structural variant (SV) haplotypes. Using a combination of short-read genome sequencing (GS), long-read GS, optical genome mapping, and single-cell DNA template strand sequencing (strand-seq), the haplotype structure was resolved in 18 samples. The point of template switching in 4 samples was shown to be a segment of ∼2.2-5.5 kb of 100% nucleotide similarity within inverted repeat pairs. These data provide experimental evidence that inverted low-copy repeats act as recombinant substrates. This type of CGR can result in multiple conformers generating diverse SV haplotypes in susceptible dosage-sensitive loci.

Bunch of grapes - I saw the sign.

As early as the late 1800s, the appearance of hydatidiform moles have been likened to grapes. The bunch of grapes sign is an ultrasound finding of a heterogenous mass within the uterine cavity with multiple anechoic spaces. We saw the bunch of grapes sign on a 28-year-old on ultrasound.

Postpartum left circumflex to left atrial fistula: A rare presentation.

Coronary artery fistulas (CAFs) are rare and often asymptomatic, but severe complications can occur, resulting in heart failure and cardiac arrhythmia. They have been associated with iatrogenic or traumatic injuries as well as systemic inflammatory conditions. However, there have been very few documented cases of pregnancy associated CAFs. We observed a case of left circumflex to left atrium fistula in a 37-year-old female presenting with insidious onset of progressive dyspnea during pregnancy.

The Association Between Mood and Anxiety Disorders With Perioperative Outcomes Following Radical Cystectomy.

OBJECTIVE: To identify associations between preoperative psychiatric diagnoses and perioperative outcomes after RC. METHODS: The Florida Inpatient Data File was used to identify patients who underwent RC from 2013 to 2019. ICD-10 codes for a mood or anxiety disorder were identified and analyzed as a 3-level variable: neither, one of these, or both. Outcomes included inpatient mortality, non-home discharge, in-hospital complications, and length of stay. Mixed-effects logistic regression (accounting for clustering within hospitals) and negative binomial regression models were utilized. RESULTS: We identified 4396 RC patients, including 306 (7.0%) with a mood disorder and 389 (8.8%) with an anxiety disorder. After multivariable adjustment, there was no significant association between mood and/or anxiety disorders with mortality or the presence or number of in-hospital complications. However, a mood or anxiety disorder was significantly associated with increased odds of non-home discharge (OR 1.60, 95% CI 1.20-2.14) and longer length of stay (IRR 1.13, 95% CI 1.07-1.19); these associations were also increased among patients with both mood and anxiety disorder diagnoses (non-home discharge OR 2.66, 95% CI 1.61-4.38; length of stay IRR 1.12, 95% CI 1.01-1.24). CONCLUSION: Patients with mood and/or anxiety disorders undergoing RC had longer length of stay and increased odds of discharge to a non-home facility despite similar risks of perioperative complications. These data suggest an opportunity for perioperative intervention to address these disparities in postoperative outcomes. However, further work is needed to determine the underlying causes of these differences and to develop effective interventions.

Lactated Ringer's solution reduces systemic inflammation compared with saline in patients with acute pancreatitis.

BACKGROUND & AIMS: Aggressive fluid resuscitation is recommended for initial management of acute pancreatitis. We performed a randomized controlled trial to evaluate the impact of a goal-directed fluid resuscitation protocol on systemic inflammation in patients with acute pancreatitis. We then determined the impact of resuscitation with lactated Ringer's solution, compared with normal saline. METHODS: We performed a randomized controlled trial of 40 patients with acute pancreatitis at 3 New England hospitals from May 2009-February 2010. Patients received goal-directed fluid resuscitation with lactated Ringer's solution, goal-directed fluid resuscitation with normal saline, standard fluid resuscitation with lactated Ringer's solution, or standard fluid resuscitation with normal saline. Systemic inflammation was measured on the basis of levels of systemic inflammatory response syndrome (SIRS) and C-reactive protein (CRP) level after 24 hours. RESULTS: The volumes of fluid administered during a 24-hour period were similar among patients given goal-directed or standard fluid resuscitation (mean, 4300 vs 4600 mL, respectively; P = .87). Goal-directed resuscitation did not significantly reduce incidence of SIRS, compared with standard resuscitation (11.8% vs 13.0%, respectively; P = .85) or levels of CRP after 24 hours (87.1 vs 69.2 mg/dL, respectively; P = .75). By contrast, there was a significant reduction in SIRS after 24 hours among subjects resuscitated with lactated Ringer's solution, compared with normal saline (84% reduction vs 0%, respectively; P = .035); administration of lactated Ringer's solution also reduced levels of CRP, compared with normal saline (51.5 vs 104 mg/dL, respectively; P = .02). CONCLUSIONS: Patients with acute pancreatitis who were resuscitated with lactated Ringer's solution had reduced systemic inflammation compared with those who received saline.

Microstructure of sputter-deposited strontium titanate films.

The origin of a marked difference in a dielectric constant, κ, observed between two types of strontium titanium oxide (STO) films sputter-deposited on platinum layers was investigated using a transmission electron microscopy method. The first type of STO films having a low κ value initially grew as an amorphous phase, followed by the formation of a randomly oriented polycrystalline phase. The second type with a high κ, on the other hand, not only grew as a crystalline phase throughout the entire film thickness, but also exhibited a strong [111] fiber texture. The observed difference in κ between these two types of STO films can thus be explained in terms of the degree of film crystallinity and texture.

Development and validation of a web-based assessment tool for the extended focused assessment with sonography in trauma examination.

OBJECTIVES: Extended focused assessment with sonography in trauma (EFAST) examinations are routinely performed by emergency physicians and general surgeons as an integral part of trauma care. Although guidelines for competency in the EFAST examination exist, tools to assess competency are lacking. Our goal was to develop and validate a Web-based competency assessment tool to evaluate providers who perform the EFAST examination. METHODS: A multiple-choice test regarding the performance and interpretation of the EFAST examination was developed following National Board of Medical Examiner guidelines. Five emergency physician experts with fellowship training in emergency ultrasound established face and content validity. The test was administered to emergency medicine residents and ultrasound fellows. Concurrent validity was evaluated by assessing the correlation of scores on our test with guidelines set by the American College of Emergency Physicians (ACEP) for emergency ultrasound. Scores were analyzed, and statistical analysis was performed. RESULTS: Sixty-three emergency medicine residents and 2 ultrasound fellows from 2 residency programs completed the assessment tool. Examinees who met ACEP guidelines scored significantly higher than those who had not: 70.4% (95% confidence interval, 67.3%-73.4%) versus 48.3% (43.2%-53.5%). Evaluation of scores showed a significant correlation between an increased level of training, participation in an ultrasound rotation, and the number of EFAST and total ultrasound examinations performed with higher test scores. However, overall test scores were lower than initially anticipated. CONCLUSIONS: Use of this assessment tool for interpretation of EFAST images showed face and content validity. Score trends showed a significant correlation with existing ACEP guidelines, ultrasound experience, and the training level. Scores continued to improve with experience beyond ACEP recommended guidelines.

Are live instructors replaceable? Computer vs. classroom lectures for EFAST training.

BACKGROUND: The EFAST (extended focused assessment with sonography for trauma) is part of the recommended curriculum for Emergency Medicine and Surgery residents. Computer-based lectures may represent a time-efficient alternative to traditional lectures. OBJECTIVES: Our hypothesis was that computer lectures in basic ultrasound and the EFAST are not inferior to classroom lectures in test score improvement for residents with or without prior training. METHODS: First-year Emergency Medicine and Surgery residents were enrolled and completed a pre-test. Subjects were then randomized into a classroom group, which attended traditional lectures, and a computer group, which listened to narrated lectures on computers. After the didactic training, all subjects completed a post-test. RESULTS: Forty-four subjects completed the study: 64% were General Surgery residents, 66% were male. Overall, mean test score improvements were higher in the classroom than in the computer group (28.0% vs. 18.4%). In 25 residents without prior training, mean improvements in the computer and classroom groups were 25% and 27%, respectively. The 95% confidence limit around the difference was 9%, falling within the a priori non-inferiority range of 10%, and consistent with non-inferiority of computer-based lectures. In 19 residents with prior training, mean test score improvements for the computer and classroom groups were 13% and 29%, respectively. The 95% confidence limit of 24% exceeded the non-inferiority range, consistent with inferiority of computer-based lectures. CONCLUSIONS: Computer-based lectures are not inferior to classroom lectures and may represent a worthwhile substitution in subjects without prior ultrasound education. Our data suggest that didactic ultrasound training through classroom lectures is more effective than computer-based lectures in individuals with prior training.

Trifocal Tibial Bone Transport Using a Magnetic Intramedullary Nail: A Case Report.

CASE: The patient sustained a pathologic fracture through chronic osteomyelitis of the tibia. Treatment involved resection of bone and soft tissue, leaving a 15-cm diaphyseal tibial defect. We designed a construct to allow for trifocal bone transport using a magnetic intramedullary lengthening nail connected to cables. This technique uses cables secured to the proximal tibia, the nail, and the transport segments. CONCLUSION: This case details the successful use of trifocal cable-assisted bone transport through a magnetic lengthening intramedullary nail. This technique can successfully be used to reconstruct diaphyseal bone defects that are longer than the stroke length of the magnetic nail.

Broadband electrical impedance as a novel characterization of oxidative stress in single L6 skeletal muscle cells.

Oxidative stress (OS) is one of the leading causes of cytotoxicity and is linked to many human physio-pathological conditions. In particular, myalgic encephalomyelitis/chronic fatigue syndrome (ME/CFS) induced by OS is debilitating to quality of life, while no clear biological markers have been identified for diagnostic measures. Recently, impedance measurements of peripheral blood cells of ME/CFS patients have been shown as a promising approach to diagnose the disease. Inspired by this study and aiming to interrogate muscle cells directly, we investigated if broadband measurements of single muscle cells could differentiate normal and oxidatively stressed cell populations. We first optimized a protocol through H2O2 treatment to introduce oxidative stress to cultured rat L6 skeletal muscle cells. The treated cells were further characterized through broadband impedance spectroscopy of single cells using a microfluidic lab-on-a-chip system. The resulting dielectric properties of cytoplasm permittivity and conductivity are electrically distinct from normally cultured cells. The reflection and transmission coefficients, ΔS11 and ΔS21, of the normal cells are tightly clustered and closely resemble those of the cell-free solution across the frequency range of 9 kHz to 9 GHz. On the other hand, dielectric properties of the oxidized cells have a wide distribution in the GHz range, deviating both in the positive and negative directions from the normally cultured cells. Simulation results guide our hypothesis that the dielectric differences could be linked to ion alterations, while calcium imaging directly supports the contribution of calcium flux to the observed deviation of S parameters. The unique electrical profile associated with oxidized cells in the GHz frequencies provide a framework for future development of technologies to diagnose oxidative-stress related diseases such as ME/CFS.

Early comprehensive testing for COVID-19 is essential to protect trauma centers.

BACKGROUND: The severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) pandemic presents a threat to health care systems worldwide. Trauma centers may be uniquely impacted, given the need for rapid invasive interventions in severely injured and the growing incidence of community infection. We discuss the impact that SARS-CoV-2 has had in our trauma center and our steps to limit the potential exposures. METHODS: We performed a retrospective evaluation of the trauma service, from March 16 to 30, following the appearance of SARS-CoV-2 in our state. We recorded the daily number of trauma patients diagnosed with SARS-CoV-2 infection, the presence of clinical symptoms or radiological signs of COVID-19, and the results of verbal symptom screen (for new admissions). The number of trauma activations, admissions, and census, as well as staff exposures and infections, was recorded daily. RESULTS: Over the 14-day evaluation period, we tested 85 trauma patients for SARS-CoV-2 infection, and 21 (25%) were found to be positive. Sixty percent of the patients in the trauma/burn intensive care unit were infected with SARS-CoV-2. Positive verbal screen results, presence of ground glass opacities on admission chest CT, and presence of clinical symptoms were not significantly different in patients with or without SARS-CoV-2 infection (p > 0.05). Many infected patients were without clinical symptoms (9/21, 43%) or radiological signs on admission (18/21, 86%) of COVID-19. CONCLUSION: Forty-five percent of trauma patients are asymptomatic at the time of SARS-CoV-2 diagnosis. Respiratory symptoms, as well as verbal screening (recent fevers, shortness of breath, cough, international travel, and close contact with known SARS-CoV-2 carriers), are inaccurate in the trauma population. These findings demonstrate the need for comprehensive rapid testing of all trauma patients upon presentation to the trauma bay. LEVEL OF EVIDENCE: Diagnostic tests or criteria, level III, Therapeutic/care management, level IV.

Notochord vacuoles absorb compressive bone growth during zebrafish spine formation.

The vertebral column or spine assembles around the notochord rod which contains a core made of large vacuolated cells. Each vacuolated cell possesses a single fluid-filled vacuole, and loss or fragmentation of these vacuoles in zebrafish leads to spine kinking. Here, we identified a mutation in the kinase gene dstyk that causes fragmentation of notochord vacuoles and a severe congenital scoliosis-like phenotype in zebrafish. Live imaging revealed that Dstyk regulates fusion of membranes with the vacuole. We find that localized disruption of notochord vacuoles causes vertebral malformation and curving of the spine axis at those sites. Accordingly, in dstyk mutants the spine curves increasingly over time as vertebral bone formation compresses the notochord asymmetrically, causing vertebral malformations and kinking of the axis. Together, our data show that notochord vacuoles function as a hydrostatic scaffold that guides symmetrical growth of vertebrae and spine formation.

Black Phosphorus High-Frequency Transistors with Local Contact Bias.

Having a sizable band gap and high carrier mobility, black phosphorus (BP) is a promising two-dimensional material for high-frequency electronic and optoelectronic devices. Further, for metal-oxide-semiconductor field-effect transistors (MOSFETs) operating at high frequencies, they must have a top gate of submicron length instead of the commonly used global back gate. However, without the global back gate to electrostatically induce doping in BP, top-gated submicron BP MOSFETs have not reached their full potential mainly due to large contact resistances. Here, we report top-gated submicron BP MOSFETs with local contact bias electrodes to induce doping in the contact region. This resulted in reduced contact resistance and, in turn, orders of magnitude improvement in current capacity (>500 µA/µm) and peak transconductance (>40 µS/µm), if compared with top-gated BP transistors without any back-gating scheme. In turn, these improvements resulted in a forward current gain cutoff frequency of 37 GHz and a maximum frequency of oscillation of 22 GHz at room temperature, the highest reported for BP MOSFETs up to date.

Correlation Between Optical Fluorescence and Microwave Transmission During Single-Cell Electroporation.

OBJECTIVE: Multimodal characterization of a mammalian cell by optical and microwave techniques simultaneously during electroporation. METHODS: Using a coplanar waveguide with a Jurkat cell trapped in the middle of its center conductor, continuous waves at 100 kHz of different amplitudes were applied for 20 s, while microwave transmission coefficients at 9 GHz were measured every 0.4 s. RESULTS: The onset of electroporation was indicated by abrupt changes in both fluorescence intensity and transmission coefficient. Additionally, in measurements that lasted 300 s, the transmission coefficient was found to recover to the pre-poration level, while the fluorescence intensity remained different. Since the cells were confirmed viable through post-poration staining, the recovery of the transmission coefficient suggested reversible electroporation. CONCLUSION: These experimental results showed that the transmission coefficient could serve as a label-free indicator of cell membrane permeability during and after electroporation. Furthermore, it could be used to expeditiously differentiate reversible electroporation from the irreversible one. SIGNIFICANCE: This study should aid fundamental analysis of cell physiology, as well as molecular delivery, in cell engineering and electrotherapy.

Rate of decline in oxygen saturation at various pulse oximetry values with prehospital rapid sequence intubation.

BACKGROUND: A high incidence of desaturations has been observed during prehospital rapid sequence intubation (RSI). The rate of decline in oxygen saturation (SpO2) at various pulse oximetry values has not been defined with emergency RSI. Objective. To define the rate of SpO2 decline at various pulse oximetry values and identify a threshold below which active BVM should be performed during prehospital RSI. METHODS: Traumatic brain injury (TBI) patients undergoing RSI by prehospital providers were included in this analysis. The time period from the highest to the lowest preintubation SpO2 value was selected for review. The mean rate of SpO2 decline was calculated for each SpO2 value and then used to define a theoretical SpO2 desaturation curve. The rate of desaturation to hypoxemia (SpO2