Developmental fate and cellular maturity encoded in human regulatory DNA landscapes.

Cellular-state information between generations of developing cells may be propagated via regulatory regions. We report consistent patterns of gain and loss of DNase I-hypersensitive sites (DHSs) as cells progress from embryonic stem cells (ESCs) to terminal fates. DHS patterns alone convey rich information about cell fate and lineage relationships distinct from information conveyed by gene expression. Developing cells share a proportion of their DHS landscapes with ESCs; that proportion decreases continuously in each cell type as differentiation progresses, providing a quantitative benchmark of developmental maturity. Developmentally stable DHSs densely encode binding sites for transcription factors involved in autoregulatory feedback circuits. In contrast to normal cells, cancer cells extensively reactivate silenced ESC DHSs and those from developmental programs external to the cell lineage from which the malignancy derives. Our results point to changes in regulatory DNA landscapes as quantitative indicators of cell-fate transitions, lineage relationships, and dysfunction.

Feasibility study of micronutrient status and body mass index of newly diagnosed pediatric oncology patients: Research commentary.

We conducted a feasibility study to evaluate micronutrients and body mass index (BMI). Fat soluble vitamins A, D, E and trace elements copper (Cu), selenium (Se), and zinc (Zn) levels were evaluated. Weight, height, BMI, and Z-scores were recorded. Side effects or specific adverse events were documented. No patient had a Z-score for height, weight, or BMI of less than 2 SD or greater than 2 SD. Ninety percent of patients had one or more micronutrient levels below normal. These results suggest that micronutrient abnormalities are common despite no obvious evidence of malnutrition. Side effects of chemotherapy may be exacerbated by micronutrient depletion.

Contemporary Mechanics of Conductive Hearing Loss.

The middle ear plays a critical role for the conversion of acoustic energy to mechanical vibrations that subsequently enter the cochlea. It is middle ear impedance matching through ossicular coupling that has enabled land-dwelling vertebrates to hear soft airborne sounds. Conductive hearing loss may result from damage to the delicate middle ear structures following infection, trauma or rapid pressure changes. An understanding of the mechanics of the middle ear significantly improves the oto-surgeon's ability to effectively diagnose conductive hearing loss, localize the responsible lesion and then effectively correct the conduction abnormality. This article reviews some of the basic knowledge of middle ear mechanics for sound transmission, highlights recent advances in developing new techniques to assist in diagnosis of middle ear disease, and finally sheds light on future research aimed at improving the diagnosis and management of middle ear pathology.

Regulatory Role of Anti-Sigma Factor RsbW in Clostridioides difficile Stress Response, Persistence, and Infection.

The anaerobic pathogen Clostridioides difficile, which is a primary cause of antibiotic-associated diarrhea, faces a variety of stresses in the environment and in the mammalian gut. To cope with these stresses, alternative sigma factor B (σB) is employed to modulate gene transcription, and σB is regulated by an anti-sigma factor, RsbW. To understand the role of RsbW in C. difficile physiology, a rsbW mutant (ΔrsbW), in which σB is assumed to be "always on," was generated. ΔrsbW did not show fitness defects in the absence of stress but tolerated acidic environments and detoxified reactive oxygen and nitrogen species better compared to the parental strain. ΔrsbW was defective in spore and biofilm formation, but it displayed increased adhesion to human gut epithelia and was less virulent in a Galleria mellonella infection model. A transcriptomic analysis to understand the unique phenotype of ΔrsbW showed changes in expression of genes associated with stress responses, virulence, sporulation, phage, and several σB-controlled regulators, including the pleiotropic regulator sinRR'. While these profiles were distinct to ΔrsbW, changes in some σB-controlled stress-associated genes were similar to those reported in the absence of σB. Further analysis of ΔrsbW showed unexpected lower intracellular levels of σB, suggesting an additional post-translational control mechanism for σB in the absence of stress. Our study provides insight into the regulatory role of RsbW and the complexity of regulatory networks mediating stress responses in C. difficile. IMPORTANCE Pathogens like Clostridioides difficile face a range of stresses in the environment and within the host. Alternative transcriptional factors like sigma factor B (σB) enable the bacterium to respond quickly to different stresses. Anti-sigma factors like RsbW control sigma factors and therefore the activation of genes via these pathways. Some of these transcriptional control systems provide C. difficile with the ability to tolerate and detoxify harmful compounds. Here, we investigate the role of RsbW in C. difficile physiology. We demonstrate distinctive phenotypes for a rsbW mutant in growth, persistence, and virulence and suggest alternate σB control mechanisms in C. difficile. Understanding C. difficile responses to external stress is key to designing better strategies to combat this highly resilient bacterial pathogen.

Clostridioides difficile infection: traversing host-pathogen interactions in the gut.

C. difficile is the primary cause for nosocomial infective diarrhoea. For a successful infection, C. difficile must navigate between resident gut bacteria and the harsh host environment. The perturbation of the intestinal microbiota by broad-spectrum antibiotics alters the composition and the geography of the gut microbiota, deterring colonization resistance, and enabling C. difficile to colonize. This review will discuss how C. difficile interacts with and exploits the microbiota and the host epithelium to infect and persist. We provide an overview of C. difficile virulence factors and their interactions with the gut to aid adhesion, cause epithelial damage and mediate persistence. Finally, we document the host responses to C. difficile, describing the immune cells and host pathways that are associated and triggered during C. difficile infection.

Spectral clustering of single-cell multi-omics data on multilayer graphs.

MOTIVATION: Single-cell sequencing technologies that simultaneously generate multimodal cellular profiles present opportunities for improved understanding of cell heterogeneity in tissues. How the multimodal information can be integrated to obtain a common cell type identification, however, poses a computational challenge. Multilayer graphs provide a natural representation of multi-omic single-cell sequencing datasets, and finding cell clusters may be understood as a multilayer graph partition problem. RESULTS: We introduce two spectral algorithms on multilayer graphs, spectral clustering on multilayer graphs and the weighted locally linear (WLL) method, to cluster cells in multi-omic single-cell sequencing datasets. We connect these algorithms through a unifying mathematical framework that represents each layer using a Hamiltonian operator and a mixture of its eigenstates to integrate the multiple graph layers, demonstrating in the process that the WLL method is a rigorous multilayer spectral graph theoretic reformulation of the popular Seurat weighted nearest neighbor (WNN) algorithm. Implementing our algorithms and applying them to a CITE-seq dataset of cord blood mononuclear cells yields results similar to the Seurat WNN analysis. Our work thus extends spectral methods to multimodal single-cell data analysis. AVAILABILITY AND IMPLEMENTATION: The code used in this study can be found at https://github.com/jssong-lab/sc-spectrum. All public data used in the article are accurately cited and described in Materials and Methods and in Supplementary Information. SUPPLEMENTARY INFORMATION: Supplementary data are available at Bioinformatics online.

Methods for the calibration of bone conduction transducers at frequencies from 5 to 20 kHz.

Techniques for standardizing the output of bone conduction transducers over the 5-20 kHz range are presented. The techniques include definitions of the standard artificial mastoid (AM) impedance and force sensitivity in that high-frequency range using an impedance head coupled to a vibration source. The AM impedance is shown to vary with the contact area of the vibration source. The AM force sensitivity does not vary with the contact area but does exhibit sharp frequency dependences over the 14-20 kHz range. The sharp frequency dependence complicates the use of the AM force sensor as a stand-alone calibration device at these high frequencies. An alternative calibration scheme that uses an accelerometer interposed between the vibrator and the AM impedance is described. Comparisons of the two schemes demonstrate that the accelerometer method produces more consistent results. Comparisons of the force and acceleration output of one bone conduction vibrator at high frequencies suggest those outputs depend on the driven load. The loads used in the two calibration schemes are compared to what is known of the impedance of the skin-covered head.

Multiple angle digital holography for the shape measurement of the unpainted tympanic membrane.

The shape of the tympanic membrane (TM) plays an important role in sound transmission through the ear for hearing. Previously we developed a high-speed holographic system employing a tunable wavelength laser for rapid TM shape measurement. However, the tunable laser illumination was not sufficient to measure the shape of the unpainted TM due to the semi-transparency of the TM and short exposure time of the camera. This paper presents a new multiple angle illumination technique that allows us to use a higher power single wavelength laser to perform shape measurements on the unpainted TM. Accuracy of the new method is demonstrated by a measure of a step gauge provided by the National Institute of Standards and Technology. We successfully applied the new shape measurement method on a fresh postmortem human TM without any paint.

Clinical Factors Affecting Survival in Pediatric Vascular Tumors of the Head and Neck.

Clinical factors associated with the behavior and outcomes of nonbenign, head, and neck vascular tumors in children are not well described. Our aim is to provide descriptive information and identify prognostic factors associated with lower overall survival for children with these types of tumors. A retrospective cohort study was performed using the SEER database (years 1973-2015). Children aged 18 years and under with the diagnosis of a vascular tumor with locally aggressive or borderline and malignant behavior, classified by ICD-O-3, within the head and neck were included. Vascular tumors with benign behavior as classified by ISSVA were excluded. One hundred forty-eight children were identified. Mean age was 9.9 years (SD = 6.4). A gender predilection was noted with more males affected, female (37.8%) and male (62.2%), P = 0.0031. Majority of the children were white (79.4%) and angiosarcoma was the most common histologic subtype (68.2%). Children had a significantly better overall survival than adults with head and neck vascular tumors, P < 0.0001. Univariate and multivariate analysis did not reveal any factors with significant association to overall survival. Vascular tumors in the head and neck in children most commonly affect males and white children, with angiosarcoma as the most common histologic subtype. Children seem to have a more favorable overall survival compared to adults.

Weight-based tacrolimus trough concentrations post liver transplant.

BACKGROUND: Tacrolimus is one of the most widely used liver transplant medications. With the increasing number of obese patients requiring liver transplants, knowledge of the effect of body size affecting post-transplant outcomes, for example drug exposure is increasingly required. AIMS: (i) To investigate whether patient body size (i.e. total bodyweight) affects trough plasma concentrations of tacrolimus when a standard mg/kg dosing regimen is used; and (ii) to investigate whether obese patients have different numbers of plasma concentrations outside the therapeutic range compared to non-obese patients in the first months after liver transplant. METHODS: Using a transplant database, data tacrolimus concentrations were available for 69 patients. Tacrolimus was initially dosed at a standard 0.1 mg/kg/day after liver transplant, and adjusted to maintain a target trough concentration. Trough blood samples, phenotypic and outcome variables were analysed. RESULTS: Trough concentrations were similar between obese and non-obese patients (P > 0.05) at each sampling day. At day 7 post-transplant, 85.7% and 79.5% of the observed plasma concentrations were outside the recommended therapeutic range for obese and non-obese patients respectively, at day 30, 52.9% and 57.4%, and at 6 months, 18.7% and 27.5%. CONCLUSION: In the first week post-transplant, tacrolimus trough concentrations after standard mg/kg dosing post liver transplant appear to be corrected by total bodyweight. Obese patients have a similar number of trough plasma concentrations outside the therapeutic range compared to non-obese patients.

Outcomes of surgical treatment for hemangiomas.

OBJECTIVE: To quantify complications and rates of adverse events associated with surgical excision of hemangiomas in children using a national surgical database. DATA SOURCE: Pediatric American College of Surgeons (ACS) National Surgical Quality Improvement Program® (ACS NSQIP® -pediatric), years 2012-2016. METHODS: Subjects included children under 18 years with a postoperative diagnosis of International Classification of Diseases (ICD), 9th revision code: 228.00 and 228.01, or ICD 10 codes D18.00 and D18.01-hemangioma, any site and hemangioma, of skin and subcutaneous tissue. RESULTS: A total of 1180 patients were included. The median age was 3.2 years (interquartile range [IQR] 1.2 to 6.4 years old). No occurrences of deep incisional surgical site infection, nerve injury, mortality, sepsis, septic shock, or blood stream infections were noted. In multivariate analysis, age ≤ 3 years and advanced American Society of Anesthesiologists class were significant predictors of adverse events (P = 0.035 and 0.001, respectively). CONCLUSIONS: For children with infantile hemangioma who fail to respond or have an incomplete response to medical therapy, surgical resection may be a relatively safe treatment alternative. Children older than 3 years of age with hemangiomas had less surgical adverse events than younger children. Overall, however, the surgical complications rates were low and deemed relatively minor.

Limitations of present models of blast-induced sound power conduction through the external and middle ear.

The use of models to predict the effect of blast-like impulses on hearing function is an ongoing topic of investigation relevant to hearing protection and hearing-loss prevention in the modern military. The first steps in the hearing process are the collection of sound power from the environment and its conduction through the external and middle ear into the inner ear. Present efforts to quantify the conduction of high-intensity sound power through the auditory periphery depend heavily on modeling. This paper reviews and elaborates on several existing models of the conduction of high-level sound from the environment into the inner ear and discusses the shortcomings of these models. A case is made that any attempt to more accurately define the workings of the middle ear during high-level sound stimulation needs to be based on additional data, some of which has been recently gathered.

Sound pressure distribution within human ear canals: II. Reverse mechanical stimulation.

This work is part of a study of the interactions of ear canal (EC) sound with tympanic membrane (TM) surface displacements. In human temporal bones, the ossicles were stimulated mechanically "in reverse" to mimic otoacoustic emissions (OAEs), and the sound field within the ear canal was sampled with 0.5-2 mm spacing near the TM surface and at more distal locations within the EC, including along the longitudinal EC axis. Sound fields were measured with the EC open or occluded. The reverse-driven sound field near the TM had larger and more irregular spatial variations below 10 kHz than with forward sound stimulation, consistent with a significant contribution of nonuniform sound modes. These variations generally did not propagate more than ∼4 mm laterally from the TM. Longitudinal sound field variations with the EC open or blocked were consistent with standing-wave patterns in tubes with open or closed ends. Relative contributions of the nonuniform components to the total sound pressure near the TM were largest at EC natural frequencies where the longitudinal component was small. Transverse variations in EC sound pressure can be reduced by reducing longitudinal EC sound pressure variations, e.g., via reducing reflections from occluding earplugs.

Tympanic membrane surface motions in forward and reverse middle ear transmissions.

Characterization of Tympanic Membrane (TM) surface motions with forward and reverse stimulation is important to understanding how the TM transduces acoustical and mechanical energy in both directions. In this paper, stroboscopic opto-electronic holography is used to quantify motions of the entire TM surface induced by forward sound and reverse mechanical stimulation in human cadaveric ears from 0.25 to 18.4 kHz. The forward sound stimulus was coupled to an anatomically realistic artificial ear canal that allowed optical access to the entire TM surface, and the reverse mechanical stimulus was applied to the body of the incus by a piezo-electric stimulator. The results show clear differences in TM surface motions evoked by the two stimuli. In the forward case, TM motion is dominated by standing-wave-like modal motions that are consistent with a relatively uniform sound-pressure load over the entire TM surface. With reverse mechanical stimulation, the TM surface shows more traveling waves, consistent with a localized mechanical drive applied to the manubrium embedded in the TM. With both stimuli, the manubrium moves less than the rest of the TM, consistent with the TM acting like a compliant membrane rather than a stiff diaphragm, and also consistent with catenary behavior due to the TM's curved shape.

National estimations of airway foreign bodies in children in the United States, 2000 to 2009.

OBJECTIVE: Identify risk factors associated with airway foreign bodies in children in the United States and report observed trends over time. DATA SOURCE: KID database (2000-2009). METHODS: ICD-9-CM codes for airway foreign bodies were used to identify patients. Risk factors were used for univariate analysis and a multivariate model to identify any increased risk of mortality. These factors were then also trended over time. RESULTS: Children with airway foreign bodies demonstrate similar risk factors as previously reported, such as male gender, age less than five years and lack of private insurance. The weighted mortality rate for paediatric inpatients with airway foreign bodies was about 2.75%. Fortunately, the rate remained relatively unchanged from 2000 to 2009. Geographically, urban hospital settings appeared to be more affected. Increased risks of mortality were noted for older age, urban hospital setting and teaching hospital status. CONCLUSIONS: Our findings confirm previous findings and identified that the diagnosis of airway foreign bodies in children were associated with male gender, age <5 years, lack of private insurance and geographic location in an urban setting. Further investigation may be warranted to provide clarity on other factors found to have increased association with mortality for quality improvement.

National Characteristics of Lymphatic Malformations in Children: Inpatient Estimates and Trends in the United States, 2000 to 2009.

With ever increasing demands to manage finite resources for health care utilization, we performed an investigation to identify inpatient clinical characteristics and trends in children with lymphatic malformations using the Kids' Inpatient Database, years 2000 to 2009, to help identify populations best suited for resource deployment. Subjects included children 18 years and below with International Classification of Diseases (ICD), ninth revision code: 228.1-lymphangioma, any site. In the United States, between 2000 and 2009, inpatient pediatric patients with lymphatic malformations most commonly affected children aged 3 years and younger, urban hospital locations, and the South and West regions. There was no significant change in age of children with lymphatic malformations or the distribution of their age from year to year, P=0.948 and 0.4223, respectively. No significant evidence for seasonal variation or effect on inpatient admission was identified, P=0.7071. A great majority of admissions (>96%) were in urban locations across each year. There was also no significant change in breakdown of admissions by geographic location, P=0.7133. Further investigation may help to elucidate how to improve access to multidisciplinary vascular anomalies teams to optimize care for these children with unique and complex lymphatic malformations.

Minimally Invasive Approach for Resection of Masseteric Vascular Malformations.

OBJECTIVE: Vascular malformations (VMs) in the head and neck region often cause esthetic as well as functional problems for patients. Intramuscular VMs (IVM), such as those in the masseter, can cause severe facial asymmetry and typically are excised transcutaneously to facilitate wide exposure and safe dissection from the facial nerve. This requires extensive dissection, prolonged healing, and can lead to suboptimal facial scarring. METHODS: We describe the technique of resecting large IVMs of the masseter muscle in 3 patients using an entirely intraoral approach with continuous nerve monitoring and without visible facial scarring or secondary deformity. Preoperative injection of sclerotherapy was performed to reduce intra-operative bleeding and optimize resection. RESULTS: Successful excision was performed without complication in 3 patients to date. Total average operating room time was 120 minutes (range 95-145 minutes). Estimated blood loss was 213 mL (range 180-240 mL). The patients were discharged home either post-operative day (POD) 1 or 2, with 1 returning to work POD 4. Facial nerve function was normal postoperatively and no hematomas developed. Subjective masticatory function was equivalent to preoperative levels in all patients. CONCLUSIONS: Intraoral excision of VMs of the masseter muscle can be safely performed without added risk or complication. Continuous facial nerve monitoring allows minimally invasive approaches to be considered with less risk of iatrogenic facial nerve injury. We purport that this is a safe and effective method with substantially better esthetic outcomes compared with traditional transcutaneous approaches.

Estimating absolute methylation levels at single-CpG resolution from methylation enrichment and restriction enzyme sequencing methods.

Recent advancements in sequencing-based DNA methylation profiling methods provide an unprecedented opportunity to map complete DNA methylomes. These include whole-genome bisulfite sequencing (WGBS, MethylC-seq, or BS-seq), reduced-representation bisulfite sequencing (RRBS), and enrichment-based methods such as MeDIP-seq, MBD-seq, and MRE-seq. These methods yield largely comparable results but differ significantly in extent of genomic CpG coverage, resolution, quantitative accuracy, and cost, at least while using current algorithms to interrogate the data. None of these existing methods provides single-CpG resolution, comprehensive genome-wide coverage, and cost feasibility for a typical laboratory. We introduce methylCRF, a novel conditional random fields-based algorithm that integrates methylated DNA immunoprecipitation (MeDIP-seq) and methylation-sensitive restriction enzyme (MRE-seq) sequencing data to predict DNA methylation levels at single-CpG resolution. Our method is a combined computational and experimental strategy to produce DNA methylomes of all 28 million CpGs in the human genome for a fraction (<10%) of the cost of whole-genome bisulfite sequencing methods. methylCRF was benchmarked for accuracy against Infinium arrays, RRBS, WGBS sequencing, and locus-specific bisulfite sequencing performed on the same human embryonic stem cell line. methylCRF transformation of MeDIP-seq/MRE-seq was equivalent to a biological replicate of WGBS in quantification, coverage, and resolution. We used conventional bisulfite conversion, PCR, cloning, and sequencing to validate loci where our predictions do not agree with whole-genome bisulfite data, and in 11 out of 12 cases, methylCRF predictions of methylation level agree better with validated results than does whole-genome bisulfite sequencing. Therefore, methylCRF transformation of MeDIP-seq/MRE-seq data provides an accurate, inexpensive, and widely accessible strategy to create full DNA methylomes.