RESUMEN

Tumoral calcinosis (TC) is a rare condition characterized by dystrophic calcinosis. TC in end stage kidney disease is associated with severe hyperparathyroidism. It is radiologically characterized by multilobulated cystic calcifications in periarticular regions without erosive arthropathy or osseus destruction. Secondary TC may necessitate medical or surgical parathyroidectomy for symptom control.

RESUMEN

We report bio-structural, bio-chemical and bio-physical evidence demonstrating how small molecules can bind to both wild-type and mutant IDH1, but only inhibit the enzymatic activity of the mutant isoform. Enabled through x-ray crystallography, we characterized a series of small molecule inhibitors that bound to mutant IDH1 differently than the marketed inhibitor Ivosidenib, for which we have determined the x-ray crystal structure. Across the industry several mutant IDH1 inhibitor chemotypes bind to this allosteric IDH1 pocket and selectively inhibit the mutant enzyme. Detailed characterization by a variety of biophysical techniques and NMR studies led us to propose how compounds binding in the allosteric IDH1 R132H pocket inhibit the production of 2-Hydroxy glutarate.

Asunto(s)

Glutaratos , Isocitrato Deshidrogenasa , Mutación , Isocitrato Deshidrogenasa/genética , Isocitrato Deshidrogenasa/metabolismo , Isocitrato Deshidrogenasa/antagonistas & inhibidores , Isocitrato Deshidrogenasa/química , Cristalografía por Rayos X , Humanos , Glutaratos/metabolismo , Glutaratos/química , Inhibidores Enzimáticos/farmacología , Inhibidores Enzimáticos/química , Espectroscopía de Resonancia Magnética , Regulación Alostérica , Modelos Moleculares

RESUMEN

BACKGROUND: Rituximab with cyclophosphamide, doxorubicin, vincristine, and prednisone (R-CHOP) is one of the first line treatments for diffuse large B-cell lymphoma (DLBCL). Rituximab comprises most of the treatment cost for this regimen; therefore, biosimilars, such as rituximab-abbs are crucial to provide affordable care. Although rituximab-abbs was studied primarily in follicular lymphoma, the Food and Drug Administration (FDA) approved this drug for all indications of the reference product on the basis of extrapolation. Effectiveness and safety data surrounding the use of rituximab-abbs in DLBCL is lacking. OBJECTIVE: To evaluate the effectiveness and safety of rituximab-abbs and reference product rituximab as R-CHOP treatment for patients with DLBCL. PATIENTS AND METHODS: This noninferiority (NI) study compared the 2-year overall survival (OS), overall response rate (ORR), and incidence of adverse events (AEs) between rituximab-abbs and its reference product (RP) in R-CHOP among adult patients with newly diagnosed DLBCL. The study inclusion period was from 1 January 2019 to 31 December 2020. Analyses were performed on the basis of a noninferiority lower limit of 10% for OS and ORR, and an upper limit of 10% for serious AEs. RESULTS: There were 240 patients who received RP rituximab, while 295 patients received rituximab-abbs. The cohort had a mean age of 63.7±12.2 years and 43% were female. The 2-year OS was 81.0% and 79.6% (NI p < 0.01) while the ORR was 80.0% and 69.6% (NI p < 0.01), among the rituximab-abbs and rituximab groups, respectively. The incidence of infusion reaction AEs (NI p < 0.01) and noninfusion reaction AEs (NI p < 0.01) also met noninferiority. CONCLUSIONS: We demonstrated that rituximab-abbs was noninferior to rituximab in both effectiveness and safety among patients receiving R-CHOP for DLBCL in this study. Long-term follow-up would be needed to confirm these results.

Asunto(s)

Protocolos de Quimioterapia Combinada Antineoplásica , Ciclofosfamida , Doxorrubicina , Linfoma de Células B Grandes Difuso , Prednisona , Rituximab , Vincristina , Humanos , Rituximab/uso terapéutico , Rituximab/efectos adversos , Rituximab/administración & dosificación , Linfoma de Células B Grandes Difuso/tratamiento farmacológico , Femenino , Persona de Mediana Edad , Masculino , Protocolos de Quimioterapia Combinada Antineoplásica/uso terapéutico , Protocolos de Quimioterapia Combinada Antineoplásica/efectos adversos , Doxorrubicina/uso terapéutico , Doxorrubicina/efectos adversos , Doxorrubicina/administración & dosificación , Ciclofosfamida/uso terapéutico , Ciclofosfamida/efectos adversos , Ciclofosfamida/administración & dosificación , Anciano , Prednisona/uso terapéutico , Prednisona/administración & dosificación , Prednisona/efectos adversos , Vincristina/uso terapéutico , Vincristina/efectos adversos , Vincristina/administración & dosificación , Adulto , Biosimilares Farmacéuticos/uso terapéutico , Biosimilares Farmacéuticos/efectos adversos , Biosimilares Farmacéuticos/administración & dosificación , Resultado del Tratamiento , Anciano de 80 o más Años

Asunto(s)

Cisplatino , Neoplasias Pulmonares , Humanos , Cisplatino/uso terapéutico , Neoplasias Pulmonares/tratamiento farmacológico , Vinblastina , Costos y Análisis de Costo , Análisis Costo-Beneficio , Protocolos de Quimioterapia Combinada Antineoplásica

RESUMEN

OBJECTIVE: To assess if the dental follicle volume of palatally impacted canines (PICs) affects the relative root position of the adjacent lateral incisors (LIs) and first premolars (FPs). MATERIALS AND METHODS: A retrospective cross-sectional study of 49 patients with unilaterally PICs with dental follicles who had CBCT imaging previously taken. Four orthodontic centers in different countries provided the sample. A mean difference of 5° between the angular measurements (mesiodistal tip, buccolingual inclination, or mesiodistal rotation) of the LI and FP adjacent to the PIC and the controls was considered clinically relevant. A value of 0.05 was set for significance level and a power of 80%. The minimum sample size was determined to be 26 patients. These patients were further assigned to an LI sample (n = 49) and an FP sample (n = 23), dependent on the direct contact of the dental follicle to that adjacent tooth. A manual segmentation technique was used to obtain the volumetric measurements of the dental follicle. Angular measurements of adjacent teeth were then compared to the contralateral nonimpacted side, which acted as the control. A multivariant regression analysis was performed using IBM SPSS software, and statistical significance was set at α = 0.05. RESULTS: Adequate intra-rater reliability was accomplished. The multivariant regression analysis implied that there is no difference in the mean change in the tip, torque, and rotation of the LI and FP between the impacted and control sides when dental follicle volumes are considered (P = .509 for the LI sample and P = .804 for the FP sample). LIMITATIONS: CBCT imaging of dental follicle border delimitations, partial volume effect, and scattering are limitations. This is a convenience sample where the FP subsample is small. CONCLUSION: The dental follicle volume of the PICs does not seem to influence the relative position of the adjacent LI and FP mesiodistal tip, buccolingual inclination, and mesiodistal rotation. Early intervention could have been suggested to avoid certain malocclusion traits if significant displacements were demonstrated.

Asunto(s)

Saco Dental , Diente Impactado , Humanos , Saco Dental/diagnóstico por imagen , Estudios Retrospectivos , Reproducibilidad de los Resultados , Estudios Transversales , Diente Canino/diagnóstico por imagen , Diente Impactado/diagnóstico por imagen , Tomografía Computarizada de Haz Cónico/métodos , Maxilar

RESUMEN

BACKGROUND: Using high awake blood pressure (BP; ≥130/80 mm Hg) on ambulatory BP monitoring (ABPM) as a reference, the purpose of this study was to determine the accuracy of high office BP (≥130/80 mm Hg) at an initial visit and high confirmatory office BP (≥130/80 mm Hg), and separately, high home BP (≥130/80 mm Hg) among participants with high office BP (≥130/80 mm Hg) at an initial office visit. METHODS AND RESULTS: The accuracy of office BP measurements using the oscillometric method for detecting high BP on ABPM was determined among 379 participants with complete office BP and ABPM data in the IDH (Improving the Detection of Hypertension) study. For detecting high BP on ABPM, the accuracy of high confirmatory office BP using the oscillometric method and, separately, high home BP was also determined among the subgroup of 122 participants with high office BP at an initial visit and complete home BP monitoring data. High office BP had moderate sensitivity (0.61 [95% CI, 0.53-0.68]) and high specificity (0.85 [95% CI, 0.80-0.90]) for high awake BP. High confirmatory office BP and high home BP had moderate sensitivity (0.69 [95% CI, 0.59-0.79] and 0.79 [95% CI, 0.71-0.87], respectively) and low and moderate specificity (0.44 [95% CI, 0.27-0.61] and 0.72 [95% CI, 0.56-0.88], respectively). CONCLUSIONS: Many individuals with high BP on ABPM do not have high office BP. Confirmatory office BP and home blood pressure monitoring also had limited ability to identify individuals with high BP on ABPM.

Asunto(s)

Monitoreo Ambulatorio de la Presión Arterial , Hipertensión , Adulto , Humanos , Monitoreo Ambulatorio de la Presión Arterial/métodos , Presión Sanguínea , Determinación de la Presión Sanguínea/métodos , Hipertensión/diagnóstico , Visita a Consultorio Médico

RESUMEN

AlGaN-based UV-A LEDs have wide applications in medical treatment and chemical sensing; however, their efficiencies are still far behind visible LEDs or even shorter wavelengths UV-C counterparts because of the large lattice mismatch between the low-Al-content active region and the AlN substrate. In this report, we investigated the composition and thickness of the quantum barrier in the active region in terms of LED performance. Due to the improved strain management and better carrier confinement, efficient UV-A LEDs (320 nm - 330 nm) with EQEs up to 6.8% were demonstrated, among the highest efficiencies at this wavelength range.

RESUMEN

BACKGROUND: Home blood pressure (BP) monitoring over a 7-day period is recommended to confirm the diagnosis of hypertension. METHODS: We determined upper and lower home BP thresholds with >90% positive predictive value and >90% negative predictive value using 1 to 6 days of monitoring to identify high home BP (systolic BP ≥130 mm Hg or diastolic BP ≥80 mm Hg) based on 7 days of home BP monitoring. The sample included 361 adults from the Improving the Detection of Hypertension Study who were not taking antihypertensive medication. We used two 7-day periods, at least 3 days apart, the first being a sampling period and the second a reference period. For each number of days in the sampling period, we determined the percentage of participants who had a high likelihood of having (>90% positive predictive value) or not having (>90% negative predictive value) high BP and would not need to continue home BP monitoring. Only the participants in an uncertain category (ie, positive predictive value ≤90% and negative predictive value ≤90%) after each day were carried forward to the next day of home BP monitoring. RESULTS: Of the 361 participants (mean [SD] age of 41.3 [13.2] years; 60.4% women), 38.0% had high home BP during the reference period. There were 63.7%, 17.1%, 10.5%, 3.3%, 3.6%, and 1.4% participants who would not need to continue after 1, 2, 3, 4, 5, and 6 days of monitoring. CONCLUSIONS: In most people, high home BP can be identified or excluded with a high degree of confidence with 3 days or less of monitoring.

Asunto(s)

Hipertensión , Hipotensión , Adulto , Humanos , Femenino , Adolescente , Masculino , Presión Sanguínea/fisiología , Monitoreo Ambulatorio de la Presión Arterial , Hipertensión/diagnóstico , Hipertensión/tratamiento farmacológico , Antihipertensivos/uso terapéutico , Factores de Riesgo

RESUMEN

Importance: Oral chemotherapy is often dispensed to patients as a 1-month supply, with pill dose and package size predetermined by the drug manufacturer; thus, changing the patient dosage may waste the remaining initial drug supply. The cost of pills wasted due to dose modification and discontinuation is often unreported. Objective: To estimate the cost of pill wastage due to dose modification and discontinuation for oral anticancer drugs that were recently approved by the US Food and Drug Administration (FDA) or that are commonly prescribed. Design, Setting, and Participants: This retrospective cross-sectional economic evaluation initially identified 26 oral anticancer drugs newly approved between January 1, 2020, and August 31, 2022, from the FDA website and the top 50 best-selling pharmaceuticals in 2021 abstracted from the Drug Discovery Trends website managed by Drug Discovery and Development. The monthly costs of each agent were extracted from the Micromedex RED BOOK database. The FDA package insert, and in some cases PubMed, of each identified drug and indication was searched (matching on trial registration number) for information on registration trials. Information extracted for each drug included the name of the drug approved, drug target, cost of the drug, number of pills per bottle, available strengths, indication, name of the trial, number of patients exposed to treatment drug, number of dose level reductions, median duration of treatment, percentage of patients who received dose reduction, and percentage of dose discontinuation. All variables included in calculations were derived from the package insert or original trial publication. Main Outcomes and Measures: The cost of wastage for selected oral anticancer drugs due to dose reduction or discontinuation and the percentage of wastage in comparison with the total cost of treatment. Results: After removing duplicates, 22 oral anticancer medications were included in the study. Because some drugs had more than 1 indication, data from 35 clinical trials were analyzed. Eight of the medications (covering 9 indications) had pill strengths divisible at each dose-reduction level; thus the cost of reduction for these pills was assumed to be zero. Two medications did not allow for dose reduction. The median cost of wastage from dose reduction and discontinuation was $1750 (range, $43-$27 200), with a mean cost of $4290 (SD, $5720) per patient. The median percentage of wastage from the total cost of treatment was 1.04% (range, 0.04%-10.80%) with a mean of 1.78% (SD, 2.21%). Conclusions and Relevance: This economic evaluation found that due to both the high cost per pill and limited pill strength availability, the mean cost of wastage associated with dose reduction or discontinuation was $4290 per patient. These results suggest that to reduce the financial burden for patients with cancer, regulatory bodies should enforce availability of pill strengths that will limit pill wastage during dose modification or recommend that drug manufacturers issue credit for unused pills.

Asunto(s)

Antineoplásicos , Costos de los Medicamentos , Humanos , Preparaciones Farmacéuticas , Estudios Retrospectivos , Estudios Transversales , Antineoplásicos/uso terapéutico

RESUMEN

A rapid and sensitive approach for the detection of endopeptidases via a new analyte-triggered mutual emancipation of linker-immobilized enzymes (AMELIE) mechanism has been developed and demonstrated using a matrix metallopeptidase, a collagenase, as the model endopeptidase analyte. AMELIE involves an autocatalytic loop created by a pair of selected enzymes immobilized on solid substrates via linkers with specific sites that can be proteolyzed by one another. These bound enzymes are spatially separated so that they cannot act upon their corresponding substrates until the introduction of the target endopeptidase analyte that can also cleave one of the linkers. This triggers the self-sustained loop of enzymatic activities to emancipate all the immobilized enzymes. In this proof of concept, signal transduction was achieved by a colorimetric horseradish peroxidase-tetramethylbenzidine (HRP-TMB-H2O2) reaction with HRP that are also being immobilized by one of the linkers. The pair of immobilized enzymes were collagenase and alginate lyase, and they were immobilized by an alginate linker and a short peptide chain containing the amino acid sequence of Leu-Gly-Pro-Ala for collagenase. A detection limit of 2.5 pg collagenase mL-1 with a wide linear range up to 4 orders of magnitude was achieved. The AMELIE biosensor can detect extracellular collagenase in the supernatant of various bacteria cultures, with a sensitivity as low as 103 cfu mL-1 of E. coli. AMELIE can readily be adapted to provide the sensitive detection of other endopeptidases.

RESUMEN

Transcription factor programs mediating the immune response to coronavirus disease 2019 (COVID-19) are not fully understood. Capturing active transcription initiation from cis-regulatory elements such as enhancers and promoters by capped small RNA sequencing (csRNA-seq), in contrast to capturing steady-state transcripts by conventional RNA-seq, allows unbiased identification of the underlying transcription factor activity and regulatory pathways. Here, we profile transcription initiation in critically ill COVID-19 patients, identifying transcription factor motifs that correlate with clinical lung injury and disease severity. Unbiased clustering reveals distinct subsets of cis-regulatory elements that delineate the cell type, pathway-specific, and combinatorial transcription factor activity. We find evidence of critical roles of regulatory networks, showing that STAT/BCL6 and E2F/MYB regulatory programs from myeloid cell populations are activated in patients with poor disease outcomes and associated with COVID-19 susceptibility genetic variants. More broadly, we demonstrate how capturing acute, disease-mediated changes in transcription initiation can provide insight into the underlying molecular mechanisms and stratify patient disease severity.

Asunto(s)

COVID-19 , Factores de Transcripción , Humanos , Factores de Transcripción/genética , Regulación de la Expresión Génica , Leucocitos/metabolismo , Unidades de Cuidados Intensivos

RESUMEN

OBJECTIVES: Suicide is a leading cause of maternal mortality. Suicidality during and around the time of pregnancy can have detrimental impacts on a child's development and outcomes. This paper examines prevalence, demographic characteristics, and timing of initial contact with first responders and health services for a cohort of women who experienced suicidality during and around the time of pregnancy. METHODS: Findings are drawn from the Partners in Prevention (PiP) study, a population-wide linked data set of suicide-related attendances by police or paramedics in Queensland, Australia. A sub-cohort of women was identified, who were between 6 months preconception and 2 years postpartum at the time of a suicide-related contact with police or paramedics (PiP-Maternal). Findings are compared to other girls and women who had a suicide-related contact with police or paramedics (PiP-Female). Prevalence, demographic characteristics, timing of contact with first responders and health services, re-presentations, and mortality are reported. RESULTS: The PiP-Maternal cohort comprised 3020 individuals and 3400 births. Women in the PiP-Maternal cohort were younger, more likely to be of Aboriginal and/or Torres Strait Islander descent and live outside of a major city than the PiP-Female cohort. There were high rates of out-of-hours calls to police and ambulance, and similar perceived seriousness of the call between women in the PiP-Maternal and PiP-Female cohorts. Women in the PiP-Maternal cohort were less likely to be admitted to an emergency department within 24 hours, even after matching on covariates. Prevalence of suicidality for women who were pregnant and up to 2 years postpartum was 1.32% (95% CI = [1.27, 1.37]). CONCLUSION: Vulnerabilities and high rates of contact with police or paramedics, coupled with lower levels of follow-up, highlight the critical need to improve service responses for women with mental health needs during these phases of life.

Asunto(s)

Socorristas , Suicidio , Embarazo , Niño , Femenino , Humanos , Madres , Prevalencia , Servicios de Salud

RESUMEN

Chronic exposure to Δ-9-tetrahydrocannabinol (THC) during adolescence is associated with long-lasting cognitive impairments and enhanced susceptibility to anxiety and mood disorders. Previous evidence has revealed functional and anatomical dissociations between the posterior vs. anterior portions of the hippocampal formation, which are classified as the dorsal and ventral regions in rodents, respectively. Notably, the dorsal hippocampus is critical for cognitive and contextual processing, whereas the ventral region is critical for affective and emotional processing. While adolescent THC exposure can induce significant morphological disturbances and glutamatergic signaling abnormalities in the hippocampus, it is not currently understood how the dorsal vs. ventral hippocampal regions are affected by THC during neurodevelopment. In the present study, we used an integrative combination of behavioral, molecular, and neural assays in a neurodevelopmental rodent model of adolescent THC exposure. We report that adolescent THC exposure induces long-lasting memory deficits and anxiety like-behaviors concomitant with a wide range of differential molecular and neuronal abnormalities in dorsal vs. ventral hippocampal regions. In addition, using matrix-assisted laser desorption/ionization imaging mass spectrometry (MALDI-IMS), we show for the first time that adolescent THC exposure induces significant and enduring dysregulation of GABA and glutamate levels in dorsal vs. ventral hippocampus. Finally, adolescent THC exposure induced dissociable dysregulations of hippocampal glutamatergic signaling, characterized by differential glutamatergic receptor expression markers, profound alterations in pyramidal neuronal activity and associated oscillatory patterns in dorsal vs. ventral hippocampal subregions.

Asunto(s)

Dronabinol , Hipocampo , Dronabinol/farmacología , Hipocampo/metabolismo , Transducción de Señal , Ácido Glutámico/metabolismo , Células Piramidales

RESUMEN

OBJECTIVES: The objective of this study is to describe the epidemiological features of each presentation with a primary dermatological diagnosis to a regional emergency department (ED). DESIGN: 1-year retrospective audit. SETTING: Regional Victorian hospital emergency department. PARTICIPANTS: Any presentation to this regional emergency department with a dermatological condition from 1 January 2020 to 31 December 2020. MAIN OUTCOME MEASURES: Dermatology presentations to the ED in 2020 and the prevalence of the associated primary diagnosis. RESULTS: In total, 4.7% (n = 1873) of ED presentations had a primary dermatological diagnosis. Of these, 1484 were ≥18 years of age and 389 were ≤17 years of age. Cellulitis (26.1%, n = 388) was the most common primary diagnosis among presentations ≥18 years. Non-specific rash was the most common diagnosis (23.6%, n = 92) in presentations ≤17 years. Indigenous Australians ≥18 years were more likely to be in a younger age group (p < 0.01), and dermatitis/eczema presentations ≥18 years (n = 10) were the largest diagnostic group referred to a dermatologist. A total of 134 (7.1%) patients ≥18 years travelled more than 50 km to the ED. There were no dermatological emergencies identified. CONCLUSIONS: A high proportion of presentations to this regional ED with a dermatological diagnosis could be well managed by a dermatologist or general practitioner (GP) as an outpatient. The findings of this study inform the need for future rural public dermatology services. Options include teledermatology, or a public weekly or fortnightly rapid review dermatology clinic with a visiting dermatologist, in the absence of a dermatologist onsite.

Asunto(s)

Servicio de Urgencia en Hospital , Médicos Generales , Humanos , Estudios Retrospectivos , Australia , Hospitales

RESUMEN

Background: Identification of circulating tumor DNA (ctDNA) following curative intent therapies is a surrogate for microscopic residual disease for patients with metastatic colorectal cancer (mCRC). Preclinically, in micrometastatic microsatellite stable (MSS) CRC, increased TGF-ß signaling results in exclusion of anti-tumor cytotoxic T cells from the tumor microenvironment. Bintrafusp alfa (BA) is a bifunctional fusion protein composed of the extracellular domain of the TGF-ßRII receptor ("TGF-ß trap") and anti-PD-L1 antibody. Methods: Patients with liver-limited, MSS mCRC and with detected ctDNA after complete resection of all known tumors and standard-of-care therapy were treated with 1200 mg of BA intravenously every 14 days for six doses. The primary endpoint was ctDNA clearance. Radiographic characteristics at recurrence were compared using independent t-tests to historical data from a similar cohort of patients with liver-limited mCRC who underwent observation. Results: Only 4 of 15 planned patients received BA before the study was stopped early for loss of equipoise. There was no grade ≥3 AE. None of the patients cleared ctDNA. All patients developed radiographic recurrence by the first planned restaging. Although not detectable at prior to treatment, TGFß3 was found in circulation in all patients at cycle 2 day 1. Compared to a historical cohort, patients administered BA developed more metastases (15 versus 2, p=0.005) and greater tumor volumes (9 cm vs 2 cm, p=0.05). Conclusions: Treatment with BA in patients with ctDNA-detected, liver-limited mCRC did not clear ctDNA and was associated with large-volume recurrence, highlighting the potential context-specific complexity of dual TGF-ß and PD-L1 inhibition.

Asunto(s)

Neoplasias del Colon , Neoplasias Colorrectales , Neoplasias Hepáticas , Neoplasias del Recto , Humanos , Neoplasias Colorrectales/tratamiento farmacológico , Neoplasias Hepáticas/tratamiento farmacológico , Factores Inmunológicos/uso terapéutico , Microambiente Tumoral

RESUMEN

Designing studies of immunotherapy is limited due to a lack of pre-clinical models that reliably predict effective immunotherapy responses. To address this gap, we developed humanized mouse models of colorectal cancer (CRC) incorporating patient-derived xenografts (PDX) with human peripheral blood mononuclear cells (PBMC). Humanized mice with CRC PDXs were generated via engraftment of autologous (isolated from the same patients as the PDXs) or allogeneic (isolated from healthy donors) PBMCs. Human T cells were detected in mouse blood, tissues, and infiltrated the implanted PDXs. The inclusion of anti-PD-1 therapy revealed that tumor responses in autologous but not allogeneic models were more comparable to that of patients. An overall non-specific graft-vs-tumor effect occurred in allogeneic models and negatively correlated with that seen in patients. In contrast, autologous humanized mice more accurately correlated with treatment outcomes by engaging pre-existing tumor specific T-cell populations. As autologous T cells appear to be the major drivers of tumor response thus, autologous humanized mice may serve as models at predicting treatment outcomes in pre-clinical settings for therapies reliant on pre-existing tumor specific T-cell populations.

RESUMEN

Increased plasma mitochondrial DNA concentrations are associated with poor outcomes in multiple critical illnesses, including COVID-19. However, current methods of cell-free mitochondrial DNA quantification in plasma are time-consuming and lack reproducibility. Here, we used next-generation sequencing to characterize the size and genome location of circulating mitochondrial DNA in critically ill subjects with COVID-19 to develop a facile and optimal method of quantification by droplet digital PCR. Sequencing revealed a large percentage of small mitochondrial DNA fragments in plasma with wide variability in coverage by genome location. We identified probes for the mitochondrial DNA genes, cytochrome B and NADH dehydrogenase 1, in regions of relatively high coverage that target small sequences potentially missed by other methods. Serial assessments of absolute mitochondrial DNA concentrations were then determined in plasma from 20 critically ill subjects with COVID-19 without a DNA isolation step. Mitochondrial DNA concentrations on the day of enrollment were increased significantly in patients with moderate or severe acute respiratory distress syndrome (ARDS) compared with those with no or mild ARDS. Comparisons of mitochondrial DNA concentrations over time between patients with no/mild ARDS who survived, patients with moderate/severe ARDS who survived, and nonsurvivors showed the highest concentrations in patients with more severe disease. Absolute mitochondrial DNA quantification by droplet digital PCR is time-efficient and reproducible; thus, we provide a valuable tool and rationale for future studies evaluating mitochondrial DNA as a real-time biomarker to guide clinical decision-making in critically ill subjects with COVID-19.

Asunto(s)

COVID-19 , Síndrome de Dificultad Respiratoria , COVID-19/diagnóstico , COVID-19/genética , Enfermedad Crítica , ADN Mitocondrial/genética , Humanos , Unidades de Cuidados Intensivos , Reacción en Cadena de la Polimerasa , Reproducibilidad de los Resultados , Síndrome de Dificultad Respiratoria/diagnóstico , Síndrome de Dificultad Respiratoria/genética

RESUMEN

Multiple microRNA (miRNA) logical assays have attracted wide attention recently, which can be applied to mimic and reveal cellular events at the molecular level. However, it remains challenging to develop labeling- and amplification-free approaches to perform logical functions with low levels of miRNA molecules. Herein, we proposed a strategy for miRNA logic operations using plasmid DNA-derived nanowires produced from a facile biosynthesis method. First, let-7d was chosen as the model target of the plasmid DNA-derived nanowire strategy, which showed good selectivity and a response sensitivity of as low as the femtomolar level. The operations of the miRNA logic gates proved the programmability of the constructed plasmid DNA-derived nanowire system for two inputs (let-7d and miR-21). Finally, three pairs of DNA nanowires were combined together to demonstrate the availability of this strategy in parallel multiple miRNAs assays. In this strategy, readout signals can be directly obtained from agarose gel without extra chemical labeling or amplification procedures. Considering the excellent performance of the logic gates with low levels of inputs, our plasmid DNA-derived nanowire strategy could provide a facile method to promote simultaneous multiple miRNA assays for the benefit of diagnosis and could be applied for the assembly of complex DNA nanostructures.

Asunto(s)

MicroARNs , Nanocables , Computadores Moleculares , ADN/química , ADN/genética , MicroARNs/genética , Plásmidos/genética

Asunto(s)

COVID-19 , Crup , Infecciones del Sistema Respiratorio , Niño , Crup/diagnóstico , Crup/epidemiología , Crup/etiología , Humanos , Lactante