Novel intertypic recombination of Echovirus 11 in the Enterovirus species B.

Enteroviruses (EVs), single-stranded, positive-sense RNA viruses, can be classified into four species (A-D), which have previously been linked to a diverse range of disease manifestations and infections affecting the central nervous system. In the Enterovirus species B (EV-B), Echovirus type 11 (E11) has been observed to occasionally circulate in Taiwan, which was responsible for an epidemic of enterovirus infections in 2018. Here, 48 clinical specimens isolated in 2003, 2004, 2009, and 2018 were collected for the high-throughput sequencing. Notably, we identified 2018 Taiwanese strains having potential recombinations in the 3D gene, as well as one 2003 strain having a double recombination with E6 and Coxsackievirus B5 in the P2 and P3 regions, respectively. Additionally, one amino acid signature mutated from the Histidine (H) in throat swab specimens to the Tyrosine (Y) in cerebral spinal fluid specimens was detected at position 1496 (or 57) of the genomic coordinate (or 3A gene) to further demonstrate intra-host evolution in different organs. In conclusion, this study identifies potential intertypic recombination events and an intra-host signature mutation in E11 strains, isolated during a 2018 neurological disease outbreak in Taiwan, contributing to our understanding of its evolution and pathogenesis.

Prognostic factors and risk-stratification model of recurrent or metastatic head and neck squamous cell carcinoma treated with cetuximab containing regimen.

BACKGROUND: In recent years, the addition of cetuximab to chemotherapy has improved treatment outcomes for patients with recurrent/metastatic head and neck squamous cell carcinoma (R/M HNSCC). In this study, we present the real-world survival data of R/M HNSCC patients who received cetuximab-containing regimens from thirteen medical centers in Taiwan, as well as a three-level risk stratification model for this patient population. METHODS: This study enrolled R/M HNSCC patients from thirteen medical centers in Taiwan who received cetuximab-containing regimens from January 1, 2017 to June 6, 2022. The cases were divided into a training cohort and a validation cohort based on the start of treatment. Overall survival (OS) was evaluated in both cohorts and exploratory analysis was performed to identify associated adverse clinical and laboratory factors. The results of the exploratory analysis were used to construct a three-level risk stratification prediction model for OS. RESULTS: A total of 1434 patients with R/M HNSCC were enrolled in this study and received cetuximab-containing regimens. The overall population had a median OS of 8.57 months (95% CI: 8.07 - 9.08). Multivariate analysis of the training cohort identified poor ECOG performance status, heavy alcohol consumption, and prior adjuvant CCRT or lack of prior RT as adverse prognostic factors. Comparison of laboratory data between patients with OS≦6 and OS > 6 also revealed unfavorable factors, including increased white blood cell count, decreased hemoglobin level, increased platelet count, increased absolute neutrophil count, decreased absolute lymphocyte count, and increased neutrophil-to-lymphocyte ratio. Using forward prediction, a three-level risk stratification prediction model was constructed using the variables of ECOG performance status, alcohol consumption, skin metastasis, modality of radiation therapy, hemoglobin level, and neutrophil-to-lymphocyte ratio. The median OS in the low-risk, intermediate-risk, and high-risk groups were 12.02 months (95% CI 10.44-13.61), 7.5 months (95% CI 7.33-8.17), and 4.01 months (95% CI 3.94-4.08), respectively, with a log-rank test p-value < 0.001. CONCLUSION: This study presents a three-level risk stratification model with strong prediction ability for OS in R/M HNSCC patients who received cetuximab-containing regimens. The results are based on real-world data and may provide valuable information for clinicians in treatment planning and future drug development.

Synchronous Head and Neck Squamous Cell Carcinoma and Hematologic Malignancies.

Synchronous occurrence of solid tumors and hematological malignancy is a rare condition. There is no standard management or therapy for this complicated situation. The authors systematically reviewed articles searched using online databases. The patients are predominantly male, and the average age is similar to each malignancy. According to these studies, most patients are treated in order of aggressiveness. However, when a patient has multiple primary malignancies, all of which are aggressive, the treatment decision is challenging. Due to the limited number of cases, it is hard to have a firm conclusion about an optimal treatment policy. It appears that poor outcomes are mostly related to less adequate therapy. Because the condition is complicated in such cases and there is no standardized treatment, patients should be treated on an individual basis with treatments customized to a given patient's particular circumstances. In addition, multidisciplinary communication and cooperation are crucial in the management of these patients. More in-depth research is still needed to reach a more conclusive treatment strategy and predictable outcomes.

The Cellular Characterization of SARS-CoV-2 Spike Protein in Virus-Infected Cells Using the Receptor Binding Domain Binding Specific Human Monoclonal Antibodies.

A human monoclonal antibody panel (PD4, PD5, PD7, SC23, and SC29) was isolated from the B cells of convalescent patients and used to examine the S protein in SARS-CoV-2-infected cells. While all five antibodies bound conformational-specific epitopes within SARS-CoV-2 spike (S) protein, only PD5, PD7, and SC23 were able to bind to the receptor binding domain (RBD). Immunofluorescence microscopy was used to examine the S protein RBD in cells infected with the Singapore isolates SARS-CoV-2/0334 and SARS-CoV-2/1302. The RBD-binders exhibited a distinct cytoplasmic staining pattern that was primarily localized within the Golgi complex and was distinct from the diffuse cytoplasmic staining pattern exhibited by the non-RBD-binders (PD4 and SC29). These data indicated that the S protein adopted a conformation in the Golgi complex that enabled the RBD recognition by the RBD-binders. The RBD-binders also recognized the uncleaved S protein, indicating that S protein cleavage was not required for RBD recognition. Electron microscopy indicated high levels of cell-associated virus particles, and multiple cycle virus infection using RBD-binder staining provided evidence for direct cell-to-cell transmission for both isolates. Although similar levels of RBD-binder staining were demonstrated for each isolate, SARS-CoV-2/1302 exhibited slower rates of cell-to-cell transmission. These data suggest that a conformational change in the S protein occurs during its transit through the Golgi complex that enables RBD recognition by the RBD-binders and suggests that these antibodies can be used to monitor S protein RBD formation during the early stages of infection. IMPORTANCE The SARS-CoV-2 spike (S) protein receptor binding domain (RBD) mediates the attachment of SARS-CoV-2 to the host cell. This interaction plays an essential role in initiating virus infection, and the S protein RBD is therefore a focus of therapeutic and vaccine interventions. However, new virus variants have emerged with altered biological properties in the RBD that can potentially negate these interventions. Therefore, an improved understanding of the biological properties of the RBD in virus-infected cells may offer future therapeutic strategies to mitigate SARS- CoV-2 infection. We used physiologically relevant antibodies that were isolated from the B cells of convalescent COVID-19 patients to monitor the RBD in cells infected with SARS-CoV-2 clinical isolates. These immunological reagents specifically recognize the correctly folded RBD and were used to monitor the appearance of the RBD in SARS-CoV-2-infected cells and identified the site where the RBD first appears.

Speech and Language outcomes in Auditory Neuropathy Spectrum Disorder (ANSD) children managed with amplification.

OBJECTIVE: To evaluate speech and language outcomes in children with Auditory Neuropathy Spectrum Disorder (ANSD) without significant comorbidities who received hearing rehabilitation in the form of hearing aids and/or cochlear implantation. METHODS: Retrospective chart review of pediatric ANSD patients at a large academic tertiary care institution from 2010 to 2019. Patients were included if they received a diagnosis of bilateral ANSD, had minimal to no comorbidities, and had speech and language testing (SLT) on at least two occasions. RESULTS: 51 patients were reviewed and 7 met inclusion criteria. Average age at ANSD diagnosis was 1 year and 11 months, and average age of hearing aid fitting was 3 years and 3 months. Hearing loss ranged from mild to profound, with four of the children wearing behind (BTE) hearing aids and three eventually receiving cochlear implants. Four of five patients who received hearing aids prior to their first speech and language evaluation demonstrated a delay at their initial evaluation, and all five patients continued to demonstrate a delay at their most recent evaluation, despite appropriate audiologic management and speech and language therapy. There were two patients who were unaided at the time of their initial and latest evaluations; one patient showed a delay at both timepoints, and one patient showed no speech delay at either timepoint. CONCLUSIONS: Pediatric ANSD patients, who are otherwise typically developing and received hearing rehabilitation and speech and language therapy, continue to show a speech and language delay (SLD). This outcome underscores the importance of close monitoring of speech and language development, providing early amplification and/or cochlear implantation, and promoting additional education and psychosocial support.

The Effect of Developmental Delay and Autism Spectrum Disorder on External Auditory Canal Foreign Body Extraction.

OBJECTIVES: To determine the effect of developmental delay (DD) and autism spectrum disorder (ASD) on pediatric external auditory canal foreign body (EAC FB) retrieval outcomes. METHODS: A retrospective chart review of children presenting with EAC FB at a tertiary children's hospital was performed between January 2018 and December 2019. Charts were reviewed for demographics, presence of otalgia, complications, number of EAC FB episodes, indications for operating room removal, DD, and ASD status. RESULTS: A total of 1467 patients underwent EAC FB removal. One hundred thirty-seven children (9.3%) had DD, and, of those with DD, 63 (46%) had ASD. Children with DD were 1.76 years older compared with children with non-DD (NDD) ( P < 0.0001) at the time of presentation, whereas children with ASD were 1.45 years older than children with NDD ( P = 0.0023). Children with DD and ASD were more likely to require removal of FB in the operating room (OR) compared with the NDD group (36.5% vs 16.7%, P = 0.0001). This was not true for children with DD without ASD. Patients with DD reported significantly less otalgia when compared with NDD patients (26.3% vs 37.4%, P = 0.0097). A similar trend, although not statistically significant, was observed when comparing children with ASD with NDD patients. The NDD patients (1.1) had fewer EAC FB episodes than patients with DD (1.6, P < 0.0001) and ASD (1.8, P < 0.0016). Hazard ratios for multiple episodes of FB were 4.5 (95% confidence interval, 2.9-6.8) for DD, and 5.6 for ASD (95% confidence interval, 3.2-9.9). The complication rate for all groups was low. CONCLUSIONS: Due to the different ways that children with DD and ASD present compared with NDD children, physicians should be vigilant when evaluating symptoms and conducting physical examinations for EAC FB in those patients. A lower threshold for referral to otolaryngologists may result in more favorable outcomes.

Toripalimab Plus Chemotherapy for Recurrent or Metastatic Nasopharyngeal Carcinoma: The JUPITER-02 Randomized Clinical Trial.

Importance: There are currently no therapies approved by the US Food and Drug Administration for nasopharyngeal carcinoma (NPC). Gemcitabine-cisplatin is the current standard of care for the first-line treatment of recurrent or metastatic NPC (RM-NPC). Objective: To determine whether toripalimab in combination with gemcitabine-cisplatin will significantly improve progression-free survival and overall survival as first-line treatment for RM-NPC, compared with gemcitabine-cisplatin alone. Design, Setting, and Participants: JUPITER-02 is an international, multicenter, randomized, double-blind phase 3 study conducted in NPC-endemic regions, including mainland China, Taiwan, and Singapore. From November 10, 2018, to October 20, 2019, 289 patients with RM-NPC with no prior systemic chemotherapy in the RM setting were enrolled from 35 participating centers. Interventions: Patients were randomized (1:1) to receive toripalimab (240 mg [n = 146]) or placebo (n = 143) in combination with gemcitabine-cisplatin for up to 6 cycles, followed by maintenance with toripalimab or placebo until disease progression, intolerable toxicity, or completion of 2 years of treatment. Main Outcome: Progression-free survival as assessed by a blinded independent central review. Secondary end points included objective response rate, overall survival, progression-free survival assessed by investigator, duration of response, and safety. Results: Among the 289 patients enrolled (median age, 46 [IQR, 38-53 years; 17% female), at the final progression-free survival analysis, toripalimab treatment had a significantly longer progression-free survival than placebo (median, 21.4 vs 8.2 months; HR, 0.52 [95% CI, 0.37-0.73]). With a median survival follow-up of 36.0 months, a significant improvement in overall survival was identified with toripalimab over placebo (hazard ratio [HR], 0.63 [95% CI, 0.45-0.89]; 2-sided P = .008). The median overall survival was not reached in the toripalimab group, while it was 33.7 months in the placebo group. A consistent effect on overall survival, favoring toripalimab, was found in subgroups with high and low PD-L1 (programmed death-ligand 1) expression. The incidence of all adverse events, grade 3 or greater adverse events, and fatal adverse events were similar between the 2 groups. However, adverse events leading to discontinuation of toripalimab or placebo (11.6% vs 4.9%), immune-related adverse events (54.1% vs 21.7%), and grade 3 or greater immune-related adverse events (9.6% vs 1.4%) were more frequent in the toripalimab group. Conclusions and Relevance: The addition of toripalimab to chemotherapy as first-line treatment for RM-NPC provided statistically significant and clinically meaningful progression-free survival and overall survival benefits compared with chemotherapy alone, with a manageable safety profile. These findings support the use of toripalimab plus gemcitabine-cisplatin as the new standard of care for this patient population. Trial Registration: ClinicalTrials.gov Identifier: NCT03581786.

Recognizing the Importance of Fast Nonisothermal Crystallization for High-Performance Two-Dimensional Dion-Jacobson Perovskite Solar Cells with High Fill Factors: A Comprehensive Mechanistic Study.

Two-dimensional (2D) Dion-Jacobson (DJ) perovskite solar cells (PSCs), despite their advantage in versatility of n-layer variation, are subject to poor photovoltaic efficiency, particularly in the fill factor (FF), compared to their three-dimensional counterparts. To enhance the performance of DJ PSCs, the process of growing crystals and hence the corresponding morphology of DJ perovskites are of prime importance. Herein, we report the fast nonisothermal (NIT) crystallization protocol that is previously unrecognized for 2D perovskites to significantly improve the morphology, orientation, and charge transport of the DJ perovskite films. Comprehensive mechanistic studies reveal that the NIT effect leads to the secondary crystallization stage, forming network-like channels that play a vital role in the FF's leap-forward improvement and hence the DJ PSC's performance. As a whole, the NIT crystallized PSCs demonstrate a high power conversion efficiency and an FF of up to 19.87 and 86.16%, respectively. This research thus provides new perspectives to achieve highly efficient DJ PSCs.

Peri-Neural Invasion Is an Important Prognostic Factor of T2N0 Oral Cancer.

Background and objectives: Among patients with pathologically proven T2N0 oral squamous cell carcinoma (OSCC), a notable amount of patients still die from tumor recurrence although they have radical surgery for early stage cancers. In literature, the prognostic indicators of this specific disease entity were rarely reported. This study aims at analyzing the prognostic factors of T2N0 OSCC patients and discussing possible managements to improve the survival. MATERIALS AND METHODS: From January 2012 to December 2017, the data of 166 pathologically proven T2N0 oral cancer patients proved by radical surgery were retrospectively collected. The clinical and pathologic factors including age, gender, tumor differentiation grade, perineural invasion (PNI), angiolymphatic invasion (ALI), margin status, and adjuvant therapy were analyzed by univariate and multivariate analysis to determine their association with disease-specific survival (DSS), and disease-free survival (DFS), which were calculated by Kaplan-Meier method. RESULTS: After median follow up time of 43.5 months, overall 3-year rates of DSS and DFS were 86.1% and 80.1% respectively for our 166 patients. Univariate analysis showed that the 3-year DSS of 90.8% for PNI negative patients was significantly better than DSS of 57.0% for PNI positive patients (p = 0.0006). The 3-year DFS of 84.2% for PNI negative patients was also significantly better than DFS of 54.6% for PNI positive patients (p = 0.001). Further multivariate analysis revealed PNI was the only independent prognostic factor associated with both DSS (Hazard Ratio (HR) = 5.02; 95% Confidence Interval (CI) = 1.99-12.6; p = 0.001), and DFS (HR = 3.92; 95% CI = 1.65-9.32; p = 0.002). Nearly 10% (16) of the 166 patients had adverse pathologic feature of PNI only. In the 11 patients without adjuvant therapy, 5 patients died from OSCC. No patients had recurrence or mortality after they received adjuvant therapy with chemotherapy ± radiotherapy. CONCLUSION: PNI was an independent prognostic factor for T2N0 oral cancer patients. Adjuvant chemotherapy and radiotherapy may benefit the survival of this specific disease entity, but further investigations are needed to elucidate the optimal regimen.

Factors affecting patency of arteriovenous fistula following first percutaneous transluminal angioplasty.

BACKGROUND: Percutaneous transluminal angioplasty (PTA) has generally replaced surgical procedures to treat arteriovenous fistula (AVF) dysfunction, but the predictors of post-intervention patency are highly variable. This study aimed to determine predictors of primary patency following PTA of dysfunctional AVF. MATERIALS AND METHODS: Retrospective analysis of first-time PTA of 307 AVF in 307 patients (171 males, mean age 64.3 ± 12.4 years). Demographic, clinical, anatomical and medication variables were reviewed and subjected to univariate and multivariate Cox regression analysis. RESULTS: The post-intervention primary patency rates at 6, 12, 24, and 36 months were 76.3%, 58.3%, 43.2%, and 38.2%, respectively. The higher aortic arch calcification (AAC) grade patients were older, had higher incidence of comorbidities and cardiomegaly, and younger AVF age, but their dialysis vintage term was shorter and diastolic blood pressure was lower, and the maximum diameter of balloon angioplasty was mostly ≤ 6 mm, and had lower phosphorus level and less calcium-containing phosphate binder use. In multivariate Cox proportional hazard analysis, the presence of higher AAC grade [hazard ratio (95% confidence interval): (1.46 (1.02-2.09); p = 0.037)] and stenosis at upper arm [1.76 (1.16-2.67); p = 0.008] were associated with shorter post-intervention primary patency. CONCLUSION: In conclusion, higher AAC grade and anatomic factor related to the location of AVF (upper arm) were the important predictors of AVF dysfunction after PTA. These results could assist in tailoring surveillance programs and performing appropriate interventions for risky AVF.

Culture-Based Virus Isolation To Evaluate Potential Infectivity of Clinical Specimens Tested for COVID-19.

Real-time reverse transcription-PCR (RT-PCR) is currently the most sensitive method to detect severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) that causes coronavirus disease 2019 (COVID-19). However, the correlation between detectable viral RNA and culturable virus in clinical specimens remains unclear. Here, we performed virus culture for 60 specimens that were confirmed to be positive for SARS-CoV-2 RNA by real-time RT-PCR. The virus could be successfully isolated from 12 throat and nine nasopharyngeal swabs and two sputum specimens. The lowest copy number required for virus isolation was determined to be 5.4, 6.0, and 5.7 log10 genome copies/ml sample for detecting the nsp12, E, and N genes, respectively. We further examined the correlation of genome copy number and virus isolation in different regions of the viral genome, demonstrating that culturable specimens are characterized by high copy numbers with a linear correlation observed between copy numbers of amplicons targeting structural and nonstructural regions. Overall, these results indicate that in addition to the copy number, the integrity of the viral genome should be considered when evaluating the infectivity of clinical SARS-CoV-2 specimens.

A case of rare isolated agnathia and literature review.

Agnathia is a rare congenital malformation with unknown etiology characterized by absence of the mandible, microstomia, and tongue aplasia, often found to have other anomalies including holoprosencephaly. The purpose of this paper was to describe the symptoms and imaging of a case of isolated agnathia and to conduct a comprehensive literature review of reported patients with isolated agnathia. Case reports of isolated agnathia are very rare, with most infants as stillborn. We report a child's management of isolated agnathia with microstomia and tongue aplasia. A literature review was performed with focus on diagnosis, airway, and feeding management of isolated agnathia. Polyhydramnios was a common pregnancy complication reported in 25 out of the 39 patients in the case study. Five infants were stillborn, while 23 died within the neonatal period. Of the deceased infants within the neonatal period, 19 died within minutes to hours while four died within days to weeks. There are nine patients with agnathia that survived past infancy. The results of this study suggest that isolated agnathia is a rare malformation which requires a multi-disciplinary approach for airway and feeding management.

Quantification of the severity of outflow stenosis of hemodialysis fistulas with a pulse- and thrill-based scoring system.

BACKGROUND: Hyper-pulsatility of hemodialysis arteriovenous fistula (AVF) is the basic physical examination finding when there is outflow stenosis. The arm elevation test can also be utilized to detect outflow stenosis. If there is no significant outflow stenosis, the AVF should collapse, at least partially, because of the effect of gravity when the AVF-bearing arm is elevated to a level above that of the heart. However, if there is significant outflow stenosis, the portion of the AVF downstream of the stenosis will collapse, while the portion upstream of the stenosis will remain distended (Clin J Am Soc Nephro 8:1220-7, 2013). In our daily practice, when performing the arm elevation test, we not only observe the collapsibility of the access outflow but also palpate the outflow to identify a background thrill that sometimes disappears with the arm at rest, only to reappear when the arm is elevated. If there is no thrill upon arm elevation, we assume that the outflow stenosis is severe and refer to this condition as "physical examination significant outflow stenosis" (PESOS). The aim of this study is to characterize PESOS using percentage stenosis and Doppler flow parameters. METHODS: We performed a case-control study using data collected prospectively between June 2019 and December 2019. A pulse- and thrill-based score system was developed to assess the severity of AVF outflow stenosis. We recorded the outflow scores and Doppler measurements performed in 84 patients with mature fistulas over a 6-month period. Angiograms were reviewed to determine the severity of outflow stenosis, which was assessed by calculation of percentage stenosis. RESULTS: Receiver operating characteristic analysis showed that a cutoff value of ≥74.44% stenosis discriminated PESOS from other AVF outflow scores, with an area under the curve of 0.9011. PESOS diagnosed cases with ≥75% outflow stenosis in an AVF, with a sensitivity of 80.39%, a specificity of 78.79%, a positive predictive value of 85.42%, and a negative predictive value of 72.22%. CONCLUSIONS: PESOS can be used to diagnose ≥75% outflow stenosis in an AVF, with or without a significant collateral vein, and its diagnostic accuracy is high. The use of PESOS as an indicator for treatment implies that physical examination may represent a useful surveillance tool.

Population dynamics at neuraminidase position 151 of influenza A (H1N1)pdm09 virus in clinical specimens.

Influenza A virus mutates rapidly, allowing it to escape natural and vaccine-induced immunity. Neuraminidase (NA) is a surface protein capable of cleaving the glycosidic linkages of neuraminic acids to release newly formed virions from infected cells. Genetic variants within a viral population can influence the emergence of pandemic viruses as well as drug susceptibility and vaccine effectiveness. In the present study, 55 clinical specimens from patients infected with the 2009 pandemic influenza A/H1N1 virus, abbreviated as A(H1N1)pdm09, during the 2015-2016 outbreak season in Taiwan were collected. Whole genomes were obtained through next-generation sequencing. Based on the published sequences from A(H1N1)pdm09 strains worldwide, a mixed population of two distinct variants at NA position 151 was revealed. We initially reasoned that such a mixed population may have emerged during cell culture. However, additional investigations confirmed that these mixed variants were detectable in the specimens of patients. To further investigate the role of the two NA-151 variants in a dynamic population, a reverse genetics system was employed to generate recombinant A(H1N1)pdm09 viruses. It was observed that the mixture of the two distinct variants was characterized by a higher replication rate compared to the recombinant viruses harbouring a single variant. Moreover, an NA inhibition assay revealed that a high frequency of the minor NA-151 variant in A(H1N1)pdm09 was associated with a reduced susceptibility to NA inhibitors. We conclude that two distinct NA-151 variants can be identified in patient specimens and that such variants may increase viral replication and NA activity.

Measuring the palpable pulsatility length as a physical examination test in defining the severity of inflow stenosis for hemodialysis fistulas.

BACKGROUND: Pulsatility is an important property of hemodialysis arteriovenous fistulas (AVF) and can be perceived by the fingers as a gradual decrease in strength downstream from the anastomosis along the main trunk of the fistula. The distance from the point at which the pulse becomes imperceptible to the anastomosis is termed the palpable pulsatility length (PPL); we considered this length may play a role in assessing the severity of inflow stenosis for hemodialysis fistulas. METHODS: This study was performed by retrospective analysis of routinely collected data. Physical examinations and fistula measurements were performed in a selected population of 76 hemodialysis patients with mature fistulas during half a year. Fistula measurements included the PPL before and after treatment and the distance between the anastomosis and the arterial cannulation site (aPump length). The aPump index (API) was calculated by dividing the PPL by the aPump length. Angiograms were reviewed to determine the location and severity of stenosis. PPL and API were used to detect the critical inflow stenosis, which indicates severe inflow stenosis of an AVF. RESULTS: Receiver operating characteristic analysis showed that the area under the curve was 0.895 for API and 0.878 for PPL. A cutoff value of API < 1.29 and PPL < 11.0 cm were selected to detect the critical inflow stenosis. The sensitivity was 96.0% versus 80.0% and specificity was 84.31% versus 84.31% for API and PPL, respectively. CONCLUSIONS: PPL and API are useful tools in defining the severity of pure inflow stenosis for mature AVFs in the hands of trained examiners with high sensitivity and specificity.

Disruption of the human COQ5-containing protein complex is associated with diminished coenzyme Q10 levels under two different conditions of mitochondrial energy deficiency.

BACKGROUND: The Coq protein complex assembled from several Coq proteins is critical for coenzyme Q6 (CoQ6) biosynthesis in yeast. Secondary CoQ10 deficiency is associated with mitochondrial DNA (mtDNA) mutations in patients. We previously demonstrated that carbonyl cyanide-p-trifluoromethoxyphenylhydrazone (FCCP) suppressed CoQ10 levels and COQ5 protein maturation in human 143B cells. METHODS: This study explored the putative COQ protein complex in human cells through two-dimensional blue native-polyacrylamide gel electrophoresis and Western blotting to investigate its status in 143B cells after FCCP treatment and in cybrids harboring the mtDNA mutation that caused myoclonic epilepsy with ragged-red fibers (MERRF) syndrome. Ubiquinol-10 and ubiquinone-10 levels were detected by high-performance liquid chromatography. Mitochondrial energy status, mRNA levels of various PDSS and COQ genes, and protein levels of COQ5 and COQ9 in cybrids were examined. RESULTS: A high-molecular-weight protein complex containing COQ5, but not COQ9, in the mitochondria was identified and its level was suppressed by FCCP and in cybrids with MERRF mutation. That was associated with decreased mitochondrial membrane potential and mitochondrial ATP production. Total CoQ10 levels were decreased under both conditions, but the ubiquinol-10:ubiquinone-10 ratio was increased in mutant cybrids. The expression of COQ5 was increased but COQ5 protein maturation was suppressed in the mutant cybrids. CONCLUSIONS: A novel COQ5-containing protein complex was discovered in human cells. Its destabilization was associated with reduced CoQ10 levels and mitochondrial energy deficiency in human cells treated with FCCP or exhibiting MERRF mutation. GENERAL SIGNIFICANCE: The findings elucidate a possible mechanism for mitochondrial dysfunction-induced CoQ10 deficiency in human cells.

Heritability and genetic basis of protein level variation in an outbred population.

The genetic basis of heritable traits has been studied for decades. Although recent mapping efforts have elucidated genetic determinants of transcript levels, mapping of protein abundance has lagged. Here, we analyze levels of 4084 GFP-tagged yeast proteins in the progeny of a cross between a laboratory and a wild strain using flow cytometry and high-content microscopy. The genotype of trans variants contributed little to protein level variation between individual cells but explained >50% of the variance in the population's average protein abundance for half of the GFP fusions tested. To map trans-acting factors responsible, we performed flow sorting and bulk segregant analysis of 25 proteins, finding a median of five protein quantitative trait loci (pQTLs) per GFP fusion. Further, we find that cis-acting variants predominate; the genotype of a gene and its surrounding region had a large effect on protein level six times more frequently than the rest of the genome combined. We present evidence for both shared and independent genetic control of transcript and protein abundance: More than half of the expression QTLs (eQTLs) contribute to changes in protein levels of regulated genes, but several pQTLs do not affect their cognate transcript levels. Allele replacements of genes known to underlie trans eQTL hotspots confirmed the correlation of effects on mRNA and protein levels. This study represents the first genome-scale measurement of genetic contribution to protein levels in single cells and populations, identifies more than a hundred trans pQTLs, and validates the propagation of effects associated with transcript variation to protein abundance.

Pooled-matrix protein interaction screens using Barcode Fusion Genetics.

High-throughput binary protein interaction mapping is continuing to extend our understanding of cellular function and disease mechanisms. However, we remain one or two orders of magnitude away from a complete interaction map for humans and other major model organisms. Completion will require screening at substantially larger scales with many complementary assays, requiring further efficiency gains in proteome-scale interaction mapping. Here, we report Barcode Fusion Genetics-Yeast Two-Hybrid (BFG-Y2H), by which a full matrix of protein pairs can be screened in a single multiplexed strain pool. BFG-Y2H uses Cre recombination to fuse DNA barcodes from distinct plasmids, generating chimeric protein-pair barcodes that can be quantified via next-generation sequencing. We applied BFG-Y2H to four different matrices ranging in scale from ~25 K to 2.5 M protein pairs. The results show that BFG-Y2H increases the efficiency of protein matrix screening, with quality that is on par with state-of-the-art Y2H methods.

Size-Tunable Low Molecular Weight Pectin-Based Electrospun Nanofibers Blended with Low Content of Poly(ethylene oxide).

Pectin, a natural plant polysaccharide, holds great potential for biomedicine. Developing low molecular weight (Mw) pectin-based nanofibers is desirable for biomedical applications in which fast degradation and elimination of polymer from the body are required. Here, we report the first work on fabricating low Mw pectin-based nanofibers through electrospinning, among which the content of carrier polymer, poly(ethylene oxide) (PEO), can be minimized to 10%. Surfactant (Triton X-100), high polymer concentration and cosolvent were essential to electrospin bead-free nanofibers at low PEO content. The size of pectin nanofibers was dependent on polymer concentration and cosolvent. The presence of cosolvent inhibited the crystallization of PEO, but enhanced the crystallization of pectin. Meanwhile, glycerol as cosolvent could lead to phase separation of polymers. This work provides a new prospective for the fabrication of low Mw pectin nanofibers suitable for in vivo applications with the demand of fast degradation.

Genome-Wide Analysis of DNA Methylation in Hyperoxia-Exposed Newborn Rat Lung.

PURPOSE: Oxygen therapy is often required to treat newborn infants with respiratory disorders. Prolonged exposure of neonatal rats to hyperoxia reduced alveolar septation, increased terminal air space size, and increased lung fibrosis; these conditions are very similar to those of human bronchopulmonary dysplasia. Epigenetic regulation of gene expression plays a crucial role in bronchopulmonary dysplasia development. METHOD: We reared Sprague-Dawley rat pups in either room air (RA, n = 24) or an atmosphere containing 85% O2 (n = 26) from Postnatal Days 1 to 14. Methylated DNA immunoprecipitation (MeDIP) was used to analyze genome-wide DNA methylation in lung tissues of neonatal rats. Hyperoxia-exposed rats exhibited larger air spaces and thinner septa than RA-exposed rats did on Postnatal Day 14. The rats exposed to hyperoxia exhibited significantly higher mean linear intercepts than did the rats exposed to RA. We applied MeDIP next-generation sequencing for profiling changes in DNA methylation in the rat lungs exposed to hyperoxia and RA. We performed bioinformatics and pathway analyses on the raw sequencing data to identify differentially methylated candidate genes. RESULTS: Our in vivo model revealed that neonatal hyperoxia exposure arrested alveolarization on Postnatal Day 14. We found that the ErbB, actin cytoskeleton, and focal adhesion signaling pathways are epigenetically modulated by exposure to hyperoxia. We demonstrated that hyperoxia exposure contribute in delaying lung development through an epigenetic mechanism by disrupting the expression of genes in lungs that might be involved in alveolarization. CONCLUSIONS: These data indicate that aberrant DNA methylation and deregulation of the actin cytoskeleton and focal adhesion pathways of lung tissues may be involved in the pathophysiology of hyperoxia-induced arrested alveolarization.