bluuebunny/crossref_metadata_2025 huggingface dataset:
Targeted-Sequencing Workflows for Comprehensive Drug Resistance Profiling of Mycobacterium tuberculosis Cultures Using Two Commercial Sequencing Platforms: Comparison of Analytical and Diagnostic Performance, Turnaround Time, and Cost
Authors: Tafess Ketema , Ng Timothy Ting Leung , Lao Hiu Yin , Leung Kenneth Siu Sing , Tam Kingsley King Gee , Rajwani Rahim , Tam Sarah Tsz Yan , Ho Lily Pui Ki , Chu Corey Mang Kiu , Gonzalez Dimitri , Sayada Chalom , Ma Oliver Chiu Kit , Nega Belete Haile , Ameni Gobena , Yam Wing Cheong , Siu Gilman Kit Hang
Published: April 2020

Background

The emergence of Mycobacterium tuberculosis with complex drug resistance profiles necessitates a rapid and comprehensive drug susceptibility test for guidance of patient treatment. We developed two targeted-sequencing workflows based on Illumina MiSeq and Nanopore MinION for the prediction of drug resistance in M. tuberculosis toward 12 antibiotics.

Methods

A total of 163 M. tuberculosis isolates collected from Hong Kong and Ethiopia were subjected to a multiplex PCR for simultaneous amplification of 19 drug resistance-associated genetic regions. The amplicons were then barcoded and sequenced in parallel on MiSeq and MinION in respective batch sizes of 24 and 12 samples. A web-based bioinformatics pipeline, BacterioChek-TB, was developed to translate the raw datasets into clinician-friendly reports.

Results

Both platforms successfully sequenced all samples with mean read depths of 1,127× and 1,649×, respectively. The variant calling by MiSeq and MinION could achieve 100% agreement if variants with an allele frequency of <40% reported by MinION were excluded. Both workflows achieved a mean clinical sensitivity of 94.8% and clinical specificity of 98.0% when compared with phenotypic drug susceptibility test (pDST). Turnaround times for the MiSeq and MinION workflows were 38 and 15 h, facilitating the delivery of treatment guidance at least 17–18 days earlier than pDST, respectively. The higher cost per sample on the MinION platform ($71.56) versus the MiSeq platform ($67.83) was attributed to differences in batching capabilities.

Conclusion

Our study demonstrates the interchangeability of MiSeq and MinION platforms for generation of accurate and actionable results for the treatment of tuberculosis.
Hypertension, the renin–angiotensin system, and the risk of lower respiratory tract infections and lung injury: implications for COVID-19
Authors: Kreutz Reinhold , Algharably Engi Abd El-Hady , Azizi Michel , Dobrowolski Piotr , Guzik Tomasz , Januszewicz Andrzej , Persu Alexandre , Prejbisz Aleksander , Riemer Thomas Günther , Wang Ji-Guang , Burnier Michel
Published: April 2020

Systemic arterial hypertension (referred to as hypertension herein) is a major risk factor of mortality worldwide, and its importance is further emphasized in the context of the novel severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) infection referred to as COVID-19. Patients with severe COVID-19 infections commonly are older and have a history of hypertension. Almost 75% of patients who have died in the pandemic in Italy had hypertension. This raised multiple questions regarding a more severe course of COVID-19 in relation to hypertension itself as well as its treatment with renin–angiotensin system (RAS) blockers, e.g. angiotensin-converting enzyme inhibitors (ACEIs) and angiotensin receptor blockers (ARBs). We provide a critical review on the relationship of hypertension, RAS, and risk of lung injury. We demonstrate lack of sound evidence that hypertension per se is an independent risk factor for COVID-19. Interestingly, ACEIs and ARBs may be associated with lower incidence and/or improved outcome in patients with lower respiratory tract infections. We also review in detail the molecular mechanisms linking the RAS to lung damage and the potential clinical impact of treatment with RAS blockers in patients with COVID-19 and a high cardiovascular and renal risk. This is related to the role of angiotensin-converting enzyme 2 (ACE2) for SARS-CoV-2 entry into cells, and expression of ACE2 in the lung, cardiovascular system, kidney, and other tissues. In summary, a critical review of available evidence does not support a deleterious effect of RAS blockers in COVID-19 infections. Therefore, there is currently no reason to discontinue RAS blockers in stable patients facing the COVID-19 pandemic.
Multiresidue Analysis of Pesticides in Moringa Pods by GC-MS/MS and LC-MS/MS
Authors: Dutta Anirban , Hingmire Sandip , Banerjee Kaushik
Published: April 2020

Background

Moringa pods are known for their nutritional and health benefits. The cultivation of this crop receives frequent pesticide applications. In the absence of risk assessment data, maximum residue limits of pesticides in this crop are considered at the default level (0.01 mg/kg). However, there exists scarcely any validated method for pesticide residue analysis in this matrix.

Objective

This study was undertaken to develop and validate a multiresidue method for the simultaneous analysis of multi-class pesticides in moringa pods by gas chromatography-tandem mass spectrometry (GC-MS/MS), and liquid chromatography-tandem mass spectrometry (LC-MS/MS).

Method

The homogenized sample (10 g) was extracted with acetonitrile (10 mL). The extract was cleaned by dispersive solid-phase extraction using a combination of 50 mg primary secondary amine, 5 mg graphitized carbon black, and 25 mg C18 sorbents, and was directly analyzed by LC-MS/MS. Another portion of the extract was reconstituted in ethyl acetate before GC-MS/MS analysis. The method was validated as per the SANTE/12682/2019 guidelines using GC-MS/MS (180 pesticides) and LC-MS/MS instruments (203 pesticides).

Results

The method provided a satisfactory analysis of the targeted pesticides with good calibration linearity (r2>0.99), high precision (RSD < 20%), and accuracy (recoveries, 70 to 120%). The reconstitution of the acetonitrile extract in ethyl acetate significantly reduced the matrix effects on GC-MS/MS analysis. The use of matrix-matched standards could correct all recoveries.

Conclusions

The method offered a large-scale analysis of multi-class pesticides with high accuracy, and precision at 10 ng/g, and higher levels. The method performance complied with the regulatory requirements, and thus, can be implemented in routine testing purposes.

Highlights

The study reports a validated method for large-scale multiresidue analysis of pesticides in moringa matrix for the first time. The method provided a high throughput analysis of multi-class pesticides with satisfactory selectivity, sensitivity, accuracy, and precision.
The effectiveness of proton beam therapy for liver metastatic recurrence in gastric cancer patients
Authors: Yamaguchi Hisashi , Honda Michitaka , Hamada Koichi , Kobayashi Hiroshi , Todate Yukitoshi , Seto Ichiro , Takayama Kanako , Suzuki Motohisa , Wada Hitoshi , Kikuchi Yasuhiro , Murakami Masao
Published: April 2020

Objective

The purpose of this cross-sectional study was to evaluate the efficacy and safety of proton beam therapy for liver metastatic recurrence in gastric cancer patients.

Methods

Consecutive patients who underwent proton beam therapy from 2010 to 2015 were isolated from our institutional database. Patients with extrahepatic metastatic lesions were excluded. Seven patients were enrolled. The median diameter of target lesions was 31 mm (13–68 mm). The most frequent dosage was 72.6 Gy equivalent in 22 fractions. The effectiveness was assessed based on the local control, overall survival and progression-free survival rates. The local control, overall survival and progression-free survival rates were calculated using the Kaplan–Meier method. Adverse events were described according to the patients’ medical records.

Results

The median follow-up period was 41.7 months (20.7–66.3 months). The 3-year local control, overall survival and progression-free survival rates were 85.7, 68.6 and 43%, respectively. All patients completed proton beam therapy without interruption. No grade ≥3 adverse events were observed.

Conclusions

Proton beam therapy might be a treatment option for patients with liver metastasis of gastric cancer.
Sovereign debt crises and cross-country assistance
Authors: Grisse Christian , Natvik Gisle J.
Published: April 2020

We provide a parsimonious framework to study the interplay between cross-country assistance and expectations-driven sovereign debt crises. Our framework extends the traditional single-country model of how multiple perfect-foresight equilibria are possible when a sovereign attempts to service public debt. The extension is that a self-interested ‘safe’ country may choose to assist a ‘risky’ country which is prone to default. Investors internalize the potential for assistance when lending to fragile countries. If the safe country cannot commit to fixed cross-country transfers or rule them out completely, assistance improves equilibrium outcomes only if the risky country is fundamentally insolvent in the sense that it cannot repay existing debt at the risk-free interest rate. If a default requires pessimistic expectations, an incentive-compatible (IC) assistance policy has adverse side effects.
Automatic sleep staging using heart rate variability, body movements, and recurrent neural networks in a sleep disordered population
Authors: Fonseca Pedro , van Gilst Merel M , Radha Mustafa , Ross Marco , Moreau Arnaud , Cerny Andreas , Anderer Peter , Long Xi , van Dijk Johannes P , Overeem Sebastiaan
Published: April 2020

Study Objectives

To validate a previously developed sleep staging algorithm using heart rate variability (HRV) and body movements in an independent broad cohort of unselected sleep disordered patients.

Methods

We applied a previously designed algorithm for automatic sleep staging using long short-term memory recurrent neural networks to model sleep architecture. The classifier uses 132 HRV features computed from electrocardiography and activity counts from accelerometry. We retrained our algorithm using two public datasets containing both healthy sleepers and sleep disordered patients. We then tested the performance of the algorithm on an independent hold-out validation set of sleep recordings from a wide range of sleep disorders collected in a tertiary sleep medicine center.

Results

The classifier achieved substantial agreement on four-class sleep staging (wake/N1–N2/N3/rapid eye movement [REM]), with an average κ of 0.60 and accuracy of 75.9%. The performance of the sleep staging algorithm was significantly higher in insomnia patients (κ = 0.62, accuracy = 77.3%). Only in REM parasomnias, the performance was significantly lower (κ = 0.47, accuracy = 70.5%). For two-class wake/sleep classification, the classifier achieved a κ of 0.65, with a sensitivity (to wake) of 72.9% and specificity of 94.0%.

Conclusions

This study shows that the combination of HRV, body movements, and a state-of-the-art deep neural network can reach substantial agreement in automatic sleep staging compared with polysomnography, even in patients suffering from a multitude of sleep disorders. The physiological signals required can be obtained in various ways, including non-obtrusive wrist-worn sensors, opening up new avenues for clinical diagnostics.
Plates by TORTEBAT, 1760 (Paris): an Abstract of Anatomy, for painting and carving
Authors: Le Floch‐Prigent Patrice Pierre , Nguyen Juliette , Gillot Jean‐Bernard
Published: April 2020

This edition occured in 1760 and was produced in Paris by the printer J‐B Crépy from 10 (on 12) original copper plates of a first edition published in 1668 which were kept by Charles‐Antoine Jombert. The exact title was “Abrégé d'anatomie, accomodé aux arts de peinture et de sculpture” = Abstract of anatomy, adaptated to the arts of painting and carving. It opened on 4 figures of the skeleton, then 7 figures of myology. They were inspired by Vesalius. The judgment of Cushing was: “by common consent the most beautiful reproduction of Vesalius' illustrations ever to be made”. Six of the plates presented a printed text on the reverse side. This work is probably a collaboration between Roger de Piles (1635–1709) and the engraver François Tortebat (1621–1690). It is discussed if this work is the one of Roger de Piles alone, as a note in his “Cours de peinture par principes” (Paris, 1708, p. 153) assessed: “In the past, I have written under a borrowed name (i.e. Abrégé d'Anatomie by Tortebat) an abstract of anatomy adaptated to the arts of painting and and carving”.

This abstract is from the Experimental Biology 2019 Meeting. There is no full text article associated with this abstract published in The FASEB Journal.
Similar skull morphologies are found in seals and sea lions due to combat or to feeding style
Authors: Smith Michael Lawrence , Dewar Eric Walter
Published: April 2020

Many carnivorans exhibit sexual dimorphism in the skeleton, variously due to aggression, feeding styles, or other behaviors. Some species of seals and sea lions only use vocal displays between males for access to mates and territory, but other species engage in outright combat between individuals. Species with higher intraspecific aggression are characterized by skulls with more crest development for the emplacement of biting muscles than less‐aggressive species. Are these more prominent skull characteristics due entirely to levels of aggression?

To answer this, we observed the placement of 26 bony landmarks on 14 species of seals and sea lions (n = 45, including 29 adult males and 16 adult females) using a geometric morphometric approach. We categorized these species by reported level of intraspecific aggression and diet. We found that skull proportions did differ between males from more‐aggressive and less‐aggressive categories. Particularly, we found that the skulls of most less‐aggressive species were characterized by little dimorphism between sexes, a more rounded cranium, and a relatively narrow zygomatic width. On the other hand, within more‐aggressive species, notable dimorphism in skull morphology existed between males and females. Most males from highly‐aggressive species developed more prominent nuchal and sagittal crests and wider zygomatic arches while females retained a more rounded skull morphology similar to less‐aggressive species.

However, there were species that were categorized as being less‐aggressive but still possessed the more developed skull morphology of more‐aggressive species. For example, the Weddell seal and the leopard seal both developed wider zygomatic arches and taller cranial crests and they also lack notable intraspecific sexual dimorphism, but are not reported to engage in frequent combat. We propose that their skull morphologies are more likely due to the emplacement of larger muscles for feeding on larger prey.

This abstract is from the Experimental Biology 2019 Meeting. There is no full text article associated with this abstract published in The FASEB Journal.
A Novel Bipartite Trapezius Muscle Variant: Functional and Clinical Considerations
Authors: Schettler Steve P , Griffiths Todd Justin , Soares Airton , Miller Joseph M.A.
Published: April 2020

During dissection of the posterior triangle in a human cadaver (Caucasian female, 102 years of age), we observed a novel arrangement of the trapezius muscle on the subject's left side only, where it consisted of two separate muscle masses. The trapezius muscle on the subject's right side appeared normal.

Regarding the trapezius muscle on the subject's left side: The superior group of muscle fibers (sometimes referred to as the descending muscle fibers of trapezius) had normal attachments to the medial superior nuchal line of the occipital bone, ligamentum nuchae, and 7th cervical vertebra. However, the superior group of muscle fibers was completely separate from the remainder of the trapezius muscle and formed a discrete muscle mass that passed antero‐inferiorly, crossing the middle of the posterior triangle of the neck, and attached anomalously to the superior surface of the medial ½ of the clavicle. This muscle mass covered approximately 25% of the surface area of the posterior triangle. To our knowledge, this variant has not been previously described in scientific literature. The inferior part of the trapezius, consisting of middle and inferior trapezius fibers, had normal attachments to the spinous processes of the thoracic vertebrae, spine and acromion of the scapula.

Since the superior group of muscle fibers of the trapezius muscle are normally involved in scapular elevation and superior rotation, variations of the bony attachment of these fibers would inevitably affect the normal function and kinematics of the shoulder girdle and upper limb on the affected side of the body. In addition to functional considerations, this anatomical variation has implications for surgical procedures in the posterior triangle of the neck.

This abstract is from the Experimental Biology 2019 Meeting. There is no full text article associated with this abstract published in The FASEB Journal.
Interplay between the Yes‐Associated protein and the matricellular protein CCN1 Regulates the phenotypical plasticity of endothelial cells in developing blood vessels
Authors: Chaqour Brahim , Leee Sangmi , Ahad Afrouja , Moon Sohyun
Published: April 2020

CCN1 is a heparin and integrin‐binding matricellular protein which localizes pericellularly and acts primarily on cells that produce it. CCN1 is highly expressed at sites of active angiogenesis and tissue repair. Our studies in conditional knockout mouse models, demonstrate that endothelium‐specific deletion of the CCN1 gene results in severe vascular defects, most notably uncontrolled angiogenic sprouting and increased endothelial cell (EC) proliferation, resulting in the formation of a denser, widely lumenized vascular network. In this study, we investigated the regulatory mechanisms and signaling pathways involved in CCN1 gene expression and activity during angiogenesis. We found that the CCN1 gene is a target of the transcriptional coactivator Yes‐Associated protein (YAP), which acts in concert with other co‐activators such as myocardin‐related transcription factor (MRTF)‐A to activate the CCN1 promoter in angiogenic ECs. Loss of YAP function in mice is associated with reduced CCN1 promoter enrichment with YAP and decreased CCN1 gene expression. YAP is a key checkpoint of the Hippo pathway controlling cell growth, differentiation and cell‐cell junctions. It comprises a kinase cascade that inactivates by phosphorylation YAP leading to its cytoplasmic sequestration and growth inhibition. Loss of function of CCN1 in mice was associated with accumulation of active nonphosphorylated YAP in developing blood vessels. Consequently, growing ECs incorporate into preformed vessels and are no longer directed toward angiogenic sprouts, resulting in the formation of a thicker denser vasculature. In cultured ECs, a sustained expression of CCN1 through adenoviral gene transfer increases YAP phosphorylation and reduced its localization into the nucleus. CCN1‐integrin signaling through focal adhesion and the actin cytoskeleton provides the cells with a soft compliant matrix putting them in in low contractility, YAP repressive cytoskeletal states. This crosstalk between CCN1 and YAP regulates phenotypical plasticity of ECs and promotes correct sprouting and branching patterns of developing blood vessels.

Support or Funding Information

R01EY022091‐05A1

R01EY024998‐01A1

This abstract is from the Experimental Biology 2019 Meeting. There is no full text article associated with this abstract published in The FASEB Journal.
Loss of RGS Control at Gαo Reveals a Balance Between Nociceptin and Mu‐opioid Receptor Systems
Authors: Kandasamy Ram , Senese Nicolas , Chadderdon Aaron , Kohut Kevin , Jutkiewicz Emily , Ingram Susan , Traynor John
Published: April 2020

Regulator of G‐protein signaling (RGS) proteins bind to the active GTP‐bound Gα subunit of heterotrimeric G‐proteins to accelerate hydrolysis of GTP and limit signaling downstream of G‐protein coupled receptors (GPCRs). Studies have shown that mice expressing Gαo protein that is insensitive to modulation by RGS proteins (RGSi Gαo) are less hyperalgesic due to enhanced signaling at the mu‐opioid receptor (MOPR). In contrast, data suggest these mice have a hyperalgesic response to mechanical stimulation. The goal of the present study was to determine the mechanism behind this apparent contradiction. Baseline mechanical hypersensitivity in naive RGSi Gαo mice was reversed by pretreatment with systemic or central administration of the nociceptin/orphanin FQ (N/OFQ) receptor (NOPR) selective antagonist J‐113397, while the opioid antagonist naltrexone further intensified the hypersensitivity. Intraplantar injection of λ‐carrageenan produced mechanical hypersensitivity, which was reversed by J‐113397 in wild‐type mice and exacerbated by naltrexone in mutant mice. Whole brain homogenates from RGSi Gαo mice showed no change from wild‐type littermates in levels of NOPR as determined by [3H]N/OFQ saturation binding and no change in the affinity of [3H]N/OFQ for NOPR, while N/OFQ had unaltered potency to activate G‐protein as measured by [35S]GTPγS binding. N/OFQ‐induced inhibition of presynaptic GABA release in the periaqueductal gray was reduced in the mutant mice compared to their wild‐type controls. Together, these results indicate that increased signaling downstream of NOPR leads to hyperalgesia in the RGSi Gαo knock‐in mice, while increased signaling downstream of MOPR reverses NOPR‐mediated hyperalgesia. Similar opposing actions of the systems are present after inflammatory pain in wild‐type mice, signifying that in both cases the balance between MOPR and NOPR signaling is disturbed. These data highlight a delicate homeostatic balance between two receptor systems that is dependent on pain state.

Support or Funding Information

Funded by R01 DA035316 and T32 DA007268.

This abstract is from the Experimental Biology 2019 Meeting. There is no full text article associated with this abstract published in The FASEB Journal.
The Regulation of Replication Stress Response at ALT telomeres by Timeless‐1
Authors: Niedzwecki Brian , Kambhampati Ooha , Granat Lauren , Zhang Dong
Published: April 2020

All cancer cells need to maintain their telomeres in order to continue proliferating. In 85–90% of all human cancers, maintenance and extension of these telomeres is accomplished by telomerase, which adds a short repetitive sequence to the ends of chromosomes. 10–15% of cancers adopt another method, called the Alternative Lengthening of Telomeres (ALT) pathway. ALT pathway uses homologous recombination to maintain the telomere length of ALT cancers and is vital for the continued growth of ALT cancers. Previously, our lab reported that depletion of FANCM, a key member of the Fanconi Anemia family of genes, induced replication stress response primarily at the ALT telomeres (Pan, PNAS 2017). The objectives of our study were: (1) Elucidate the biological function of human Timeless (Tim1) protein, a very important DNA repair protein, in DNA replication stress response at ALT telomeres. (2) Determine whether depletion of Tim1 affects the viability of ALT cells. To accomplish these goals, we first used siRNA transfection to deplete either Tim1, or FANCM, or both in ALT cells. These cells were then co‐stained with antibodies recognizing TRF2 (a telomere marker) and either BLM (a DNA repair marker) or pRPA (a DNA damage checkpoint marker). The percentage of cells with TRF2 & BLM as well as TRF2 & pRPA foci were quantified. In addition, we also monitored the amount of micronuclei formed in those cells. Formation of micronuclei often represent the unrepaired DNA damages. Finally, cell viability assays of the siRNA transfected cells were done. Cells were allowed to grow for 10 days, fixed and stained with crystal violet. Our results show that:

siRNA depletion of FANCM and Tim1 in ALT cells had synergistic/additive foci formation for TRF2 & BLM, and TRF2 & pRPA.

siRNA depletion of FANCM and Tim1 in ALT cells had no significant effect on micronuclei formation alone, but increased formation when combined.

siRNA depletion of FANCM and Tim1 in ALT cells appears to decrease cell viability to the greatest extent when co‐depleted, as opposed to either one depleted alone.

Our findings suggest: (1) ALT cancer cells utilize both FANCM and Tim1 to resolve the replication stress at ALT telomeres; (2) co‐depletion of FANCM and Tim1 is synthetically lethal in ALT cancers. (3) Targeting both FANCM and Tim1 is a potential efficacious strategy in treating ALT cancers.

This abstract is from the Experimental Biology 2019 Meeting. There is no full text article associated with this abstract published in The FASEB Journal.
FoxO1 regulates beige adipogenesis and adiposity via autophagy
Authors: Cheng Zhiyong , Tao Zhipeng , Liu Longhua , Zheng Louise
Published: April 2020

The transcription factor forkhead box O1 (FoxO1) plays an important role in autophagy and metabolic homeostasis. Previous studies from our lab and others have shown that FoxO1 is required for autophagy and white adipocyte differentiation. However, it remains largely unknown whether and how FoxO1 mediates beige adipogenesis, a cellular event that promotes energy expenditure through mitochondrial uncoupling and thermogenesis. To address this question, we modulated FoxO1 activity in primary white adipocytes in vitro with FoxO1 antagonist (AS1842856). AS1842856 increased mitochondrial uncoupling protein UCP1, concomitant with upregulation of mitochondrial respiration chain proteins. These changes were associated with suppressed autophagy as evaluated by LC3 and p62. Interestingly, treatment of white adipocytes with classic autophagy inhibitor bafilomycin A1 also increased UCP1 and mitochondrial respiration chain protein, suggesting that FoxO1 may regulate beiging of white adipocytes via autophagy. Indeed, knockout (KO) of FoxO1 in mouse adipose tissue dampens autophagy and promotes beiging phenotype (i.e., upregulation of UCP1 and other mitochondrial contents). In line with beige adipogenesis priming energy expenditure through thermogenesis, the FoxO1‐KO mice showed significantly higher body temperature and lower adiposity. Together, our data reveals a role for FoxO1 to mediate beiging of white adipocytes by regulating autophagy, and suggests that targeting adipocyte FoxO1 may lead to new strategies to treat or prevent obesity.

Support or Funding Information

USDA National Institute of Food and Agriculture Hatch Project 1007334 (Z.C.) and American Heart Association Grant 18TPA34230082 (Z.C.)

This abstract is from the Experimental Biology 2019 Meeting. There is no full text article associated with this abstract published in The FASEB Journal.
Podocyte‐parietal epithelial cell cross‐talk: a role for paracrine KLF4‐STAT3 signaling in parietal epithelial cell proliferation in proliferative glomerulopathies
Authors: Pace Jesse , Guo Yiqing , Estrada Chelsea , Mallipattu Sandeep
Published: April 2020

Podocyte injury contributing to parietal epithelial cell (PEC) proliferation is a dominant histologic feature in both rapidly progressive glomerulonephritis (RPGN) and subtypes of focal segmental glomerulosclerosis (FSGS). Activation of Signal Transducer and Activator of Transcription 3 (STAT3) signaling is critical to the progression of both. We recently showed that podocyte‐specific loss of Krüppel‐Like Factor 4 (KLF4), a zinc‐finger transcription factor, activates STAT3 signaling leading to mitotic catastrophe in podocytes and eventual FSGS, while triggering PEC proliferation.

To determine the mechanism by which podocyte‐specific knockdown of KLF4 leads to podocyte loss and triggers PEC proliferation with dysregulated STAT3 signaling, we generated cultured human podocytes with KLF4 knockdown (KLF4‐shRNA) and screened several pharmacological STAT3‐specific inhibitors to determine that S3I‐201 prevented podocyte loss to a higher degree than other STAT3 inhibitors or DMSO‐treated KLF4‐shRNA podocytes. Furthermore, we demonstrated that S3I‐201‐treated PodCre‐KLF4fl/fl mice exhibited less proteinuria and improved renal function as compared to DMSO‐treated PodCre‐KLF4fl/fl mice. We further validated the reno‐protective effects of S3I‐201 in HIV‐1 transgenic mice, a known model of podocyte‐specific Klf4 loss and STAT3 activation, by demonstrating reduced albuminuria, podocyte loss, FSGS lesions, and parietal epithelial cell activation, with improved renal function.

We previously demonstrated that supernatant harvested from KLF4‐shRNA podocytes induces PEC proliferation. To elucidate the mechanism by which loss of KLF4 specifically in podocytes triggers PEC activation and proliferation, we performed unbiased quantitative proteomics on the supernatant of KLF4‐shRNA podocytes as compared to EV‐shRNA podocytes. We subsequently cross‐matched proteins upregulated in supernatant of KLF4‐shRNA podocytes with differentially expressed transcripts from RNA‐sequencing of isolated glomeruli from PodCre‐KLF4fl/fl mice as compared to wildtype KLF4fl/fl mice. Enrichment analysis of these differentially expressed targets revealed activation of STAT‐dependent and independent pathways involved in inflammation, extracellular matrix organization, and proliferation. Interestingly, the highest expressed STAT3 downstream targets were Plasminogen Activator Inhibitor 1 (PAI‐1) and Galectin‐1 (LGALS1), which are key mediators of epithelial cell migration and proliferation.

Collectively, these findings demonstrate the role of dysregulated KLF4‐STAT3 signaling in proliferative glomerulopathies and identify potential paracrine mediators of podocyte‐PEC cross‐talk.

Support or Funding Information

NIH/NIDDK, SBU URECA

This abstract is from the Experimental Biology 2019 Meeting. There is no full text article associated with this abstract published in The FASEB Journal.
High Fructose Consumption during Pregnancy and Lactation Leads to Maternal Renal Function Alteration
Authors: Gomes Guiomar Nascimento , Kimura Lichtenecker Debora C. , Argeri Rogerio
Published: April 2020

The consumption of foods that have fructose in its formulations has grown in the last decades. Nevertheless, increased consumption of this sugar has been related to the development of diseases such as obesity, diabetes and others. Many physiological changes occur during pregnancy and lactation and high consumption of fructose in these periods can interfere with maternal health. Thus, this study aims to evaluate the effects of increased fructose consumption on gestation and lactation on blood pressure and renal function.

Methods

Females Wistar rats were randomly assigned to groups: Control (C) and Fructose (F). The group C received food and drinking water ad libitum, and F received food and D‐fructose solution (20%) for drinking ad libitum. In F, the offer of D‐fructose solution begun 1week before mating and continued during pregnancy and lactation. Renal function was evaluated 21 days after offspring birth. Indirect measurement of systolic blood pressure (BPi), body weight, arterial blood pH, glycemia, ureia concentration in plasma, glomerular filtration rate ‐ GFR (creatinine clearance), excreted load of sodium (ENa+) and potassium (EK+) were measured. Ethic Committee on Animal Use of the Federal University of Sao Paulo approval number 757270117.

Results

Compared with C group, F presented higher values of BPi (129.7±1.4 vs 118.4±1.9 mmHg; p=0.0012), body weight (281.7±4.9 vs 232.7± 22.9 g; p=0.0245) and urinary flow (33.4±3.6 vs 14.6±2.1 mL; p=0.0035). F also had decreased GFR (3.8±0.4 vs 6.0± 0.5 mL/min/kg; p=0.0082), ENa+ (0.88±0.12 vs 1.42±0.25 mEq/24h; p=0.0350) and EK+ (2.24±0.38 vs 3.73±0.51 mEq/24h, p=0.0350). There were no significant differences between the groups regarding the arterial blood pH (7.40±0.01 vs 7.39±0.02; p=0.7599), glycemia (160.7± 3.5 vs 146.7±4.7 mg/dL; p=0.0565) and ureia concentration in plasma (36.7±1.2 vs 34.9±1.1 mg/dL; p=0.4645). Values presented as mean ± sem, Mann‐Whitney test, p ≤ 0.05; number of animals: C=6, F=7.

Conclusions

The results suggest that in rats, fructose overload during pregnancy and lactation can lead to maternal renal dysfunction (reduced GFR and sodium excretion), changes probably related to the increase in blood pressure observed in these animals.

Support or Funding Information

FAPESP, CNPq

This abstract is from the Experimental Biology 2019 Meeting. There is no full text article associated with this abstract published in The FASEB Journal.
Biosynthesis of Complex Plant‐Derived Natural Products
Authors: Smolke Christina
Published: April 2020

Plants are a rich source of unique molecules, including 25% of natural‐product‐derived drugs. However, the discovery, synthesis, and overall material supply chains for sourcing plant‐based medicines remain ad hoc, biased, and tedious. While microbial biosynthesis presents compelling alternatives to traditional approaches based on extraction from natural plant hosts, many challenges exist in the reconstruction of plant specialized metabolic pathways in microbial hosts. We have developed approaches to address the challenges that arise in the reconstruction of complex plant biosynthetic pathways in microbial hosts. We have utilized these strategies to develop yeast production platforms for an important class of plant alkaloids, which include the medicinal opioids and noscapinoids. The intersection of synthetic biology, genomics, and informatics will lead to transformative advances in how we make and discover essential medicines.

This abstract is from the Experimental Biology 2019 Meeting. There is no full text article associated with this abstract published in The FASEB Journal.
MicroRNA‐181b Regulates Ca2+ Influx by Targeting TRPC6 in PASMC from Patients with Idiopathic Pulmonary Arterial Hypertension
Authors: Babicheva Aleksandra , Makino Ayako , Yuan Jason X.‐J.
Published: April 2020

Introduction

We have previously showed that upregulated canonical transient receptor potential 6 (TRPC6) contributes to elevated cytosolic Ca2+ concentration ([Ca2+]cyt) in pulmonary artery smooth muscle cells (PASMC) isolated from patients with idiopathic pulmonary arterial hypertension (IPAH). An increase in [Ca2+]cyt is a major stimulus for pulmonary vasoconstriction and PASMC proliferation. Pulmonary vascular remodeling characterized by excessive PASMC proliferation, and sustained vasoconstriction are two important factors for elevated pulmonary vascular resistance (PVR) and pulmonary arterial pressure (PAP) in IPAH patients. However, the molecular mechanisms that regulate [Ca2+]cyt and PASMC proliferation still remain unknown. In this study, we investigate the role of microRNA (miRNA), small non‐coding RNA, which is able to regulate gene expression post‐transcriptionally by inhibiting translation and stability of mRNAs. We hypothesized that decreased miRNA‐181b (miR‐181b) resulted in the enhanced [Ca2+]cyt by upregulating TRPC6 in PASMC isolated from IPAH patients.

Methods

PASMC from healthy subjects and patients with IPAH were used for this study. miRNA expression profile was explored using the Human Cancer Pathway Finder miRNA PCR Array. In silico analysis was performed by using standard online software (Targetscan.org) to predict direct targets of miR‐181b. miR‐181b level was quantified by real‐time RT‐PCR in normal and IPAH PASMC. Normal PASMC were transfected with miR‐181b inhibitor while IPAH‐PASMC were overexpressed with miR‐181b. mRNA and protein expression of TRPC6 channels were measured by RT‐PCR and Western blot, respectively. [Ca2+]cyt was measured using fura‐2 AM, a membrane‐permeable Ca2+‐sensitive fluorescent indicator.

Results

miRNA PCR microarray showed that 5 out of 89 miRNAs were significantly downregulated in IPAH‐PASMC compared to normal PASMC. In silico analysis revealed that out of 5 candidates miR‐181b has the most potential to directly target TRPC6. Downregulated miR‐181b is correlated with increased TRPC6 expression and [Ca2+]cyt in PASMC from IPAH patients compared to PASMC isolated from healthy subjects. Inhibition of miR‐181b in normal PASMC significantly enhanced [Ca2+]cyt and upregulates TRPC6 expression while overexpression of miR‐181b in IPAH‐PASMC inhibits [Ca2+]cyt and decreases TRPC6 level.

Conclusion

Increased TRPC6 and elevated Ca2+ influx mediated by downregulated miR‐181b could contributes to excessive PASMC and sustained vasoconstriction in patients with IPAH.

Support or Funding Information

This work was supported, in part, by grants from the National Heart, Lung, and Blood Institute of the National Institutes of Health (HL125208, HL135807, and HL126609).

This abstract is from the Experimental Biology 2019 Meeting. There is no full text article associated with this abstract published in The FASEB Journal.
Stability of Early Auditory Evoked Potential Components Over Extended Test-Retest Intervals in Young Adults
Authors: Bieber Rebecca E. , Fernandez Katharine , Zalewski Chris , Cheng Hui , Brewer Carmen C.
Published: April 2020

Objectives:

Synaptic damage from noise exposures can occur even in the absence of changes in hearing sensitivity in animal models. There is an unmet clinical need for measurements sensitive to such damage to the human auditory system that can augment the pure-tone audiogram. Early components (i.e., <10 msec) of the auditory evoked potential (AEP) may be useful noninvasive indicators of synaptic integrity. Wave I is a measure of synchronous neural activity at the level of the synapse between cochlear inner hair cells and the auditory nerve and may be of particular clinical utility. This amplitude measure has historically been classified as too variable in humans to be used for clinical waveform interpretation, though several recent reliability studies have challenged this view. The focus of the present study is to examine across-session stability of early AEP amplitude measures.

Design:

In this study, amplitudes of early components (wave I, wave V, summating potential [SP]) of the AEP were measured in a cohort of 38 young adults aged 19 to 33 years (21 female). Stability of these amplitude measures was examined in a subset of 12 young adults (8 female), at time intervals ranging from 15 hr to 328 days between tests. Eligibility criteria included normal pure-tone hearing sensitivity, normal tympanometry, and intact acoustic reflexes. Participants were tested at up to four time points. Each evaluation included pure-tone thresholds, tympanometry, speech-in-noise testing, distortion-product otoacoustic emissions (DPOAE), and early AEPs. AEPs were collected in response to click and tone burst stimuli, with both ear canal and mastoid electrode montages.

Results:

No clinical changes in pure-tone hearing were found between baseline and follow-up visits. Intraclass correlation coefficients (ICCs) indicated good to excellent reliability for wave I and wave V peak-to-trough amplitudes within individuals across time, with greatest reliability (0.92, 95% confidence interval [0.81 to 0.96]) and largest amplitudes for wave I when measured from the ear canal in response to a click stimulus. Other measures such as amplitude ratios of waves V/I and the SP and action potential (AP) showed lower ICC values when measured from the ear canal, with SP/AP ratio demonstrating the lowest reliability.

Conclusions:

The results of this study suggest that, when recorded under certain conditions, wave I amplitude can be a stable measure in humans. These findings are consistent with previous work and may inform the development of clinical protocols that utilize wave I amplitude to infer inner ear integrity.
Gut microbiota dysbiosis in preeclampsia patients in the second and third trimesters
Authors: Wang Jing , Shi Zhong-Hua , Yang Jing , Wei Yuan , Wang Xiao-Ye , Zhao Yang-Yu
Published: April 2020

Background

Preeclampsia (PE) is a serious complication that affects maternal and perinatal outcomes. However, the mechanisms have not been fully explained. This study was designed to analyze longitudinal gut microbiota alterations in pregnant women with and without PE in the second (T2) and third trimesters (T3).

Methods

In this nested case-control study, which was conducted at Nanjing Maternity and Child Health Care Hospital, fecal samples from 25 PE patients (25 fecal samples obtained in T2 and 15 fecal samples obtained in T3) and 25 matched healthy controls (25 fecal samples obtained in T2 and 22 fecal samples obtained in T3) were collected, and the microbiota were analyzed using 16S rRNA gene sequencing. The diversity and composition of the microbiota of PE cases and controls were compared.

Results

No significant differences in diversity were found between the PE and control groups (P > 0.05). In the control group, from T2 to T3, the relative abundances of Proteobacteria (median [Q1, Q3]: 2.25% [1.24%, 3.30%] vs. 0.64% [0.20%, 1.20%], Z = −3.880, P < 0.05), and Tenericutes (median [Q1, Q3]: 0.12% [0.03%, 3.10%] vs. 0.03% [0.02%, 0.17%], Z = −2.369, P < 0.05) decreased significantly. In the PE group, the relative abundance of Bacteroidetes in T2 was lower than in T3 (median [Q1, Q3]: 18.16% [12.99%, 30.46%] vs. 31.09% [19.89%, 46.06%], Z = −2.417, P < 0.05). In T2, the relative abundances of mircrobiota showed no significant differences between the PE group and the control group. However, in T3, the relative abundance of Firmicutes was significantly lower in the PE group than in the control group (mean ± standard deviation: 60.62% ± 15.17% vs. 75.57% ± 11.53%, t = −3.405, P < 0.05). The relative abundances of Bacteroidetes, Proteobacteria, and Enterobacteriaceae were significantly higher in the PE group than in the control group (median [Q1, Q3]: 31.09% [19.89%, 46.06%] vs. 18.24% [12.90%, 32.04%], Z = −2.537, P < 0.05; 1.52% [1.05%, 2.61%] vs. 0.64% [0.20%, 1.20%], Z = −3.310, P < 0.05; 0.75% [0.20%, 1.00%] vs. 0.01% [0.004%, 0.023%], Z = −4.152, P < 0.05). Linear discriminant analysis combined effect size measurements analysis showed that the relative abundances of the phylum Bacteroidetes, class Bacteroidia and order Bacteroidales were increased in the PE group, while those of the phylum Firmicutes, the class Clostridia, the order Clostridiales, and the genus unidentified Lachnospiraceae were decreased in the PE group; and these differences were identified as taxonomic biomarkers of PE in T3.

Conclusion

From T2 to T3, there was an obvious alteration in the gut microbiota. The gut microbiota of PE patients in T3 was significantly different from that of the control group.
Comparison of fresh frozen plasma vs. coagulation factor concentrates for reconstitution of blood
Authors: Gratz Johannes , Ponschab Martin , Iapichino Giacomo E. , Schlimp Christoph J. , Cadamuro Janne , Grottke Oliver , Zipperle Johannes , Oberladstätter Daniel , Gabriel Christian , Ziegler Bernhard , Schöchl Herbert
Published: April 2020

BACKGROUND

Many trauma centres have adopted the administration of fixed ratios of packed red blood cells (PRBCs), platelet concentrates and fresh frozen plasma (FFP) for bleeding patients. However, the haemostatic efficacy of this concept is not well proven.

OBJECTIVE

Our objective was to characterise the haemostatic profile of different ratios (2 : 1 : 1, 1 : 1 : 1 and 1 : 1 : 2) of PRBCs, platelet concentrates and FFP in comparison with coagulation factor concentrates (fibrinogen and/or prothrombin complex concentrate).

DESIGN

An in vitro study.

SETTING

Research laboratories of the department of transfusion medicine, Linz, Austria.

MATERIALS

Whole blood donations from a total of 20 male volunteers.

INTERVENTION

Reconstitution of blood at different ratios of PRBCs, platelet concentrates and FFP or coagulation factor concentrates.

MAIN OUTCOME MEASURES

Cell count, conventional and thromboelastometric coagulation parameters, single coagulation factor activities as well as endogenous thrombin potential.

RESULTS

Fibrinogen levels and haematocrit were lower in the FFP group at any ratio compared with the concentrate-based groups (P < 0.0001). Reconstitution of blood with FFP at different ratios resulted in haematocrit or fibrinogen levels that were borderline with regard to recommended substitution triggers (haematocrit 41 ± 2% and fibrinogen 1.5 ± 0.3 g l−1 at the 2 : 1 : 1 ratio vs. 21 ± 1% and 2.1 ± 0.4 g l−1 respectively at the 1 : 1 : 2 ratio). Compared with FFP at any ratio, maximum clot firmness showed higher values in the groups using fibrinogen concentrate (P < 0.0001), whereas endogenous thrombin potential revealed higher values in the groups using prothrombin complex concentrate (P < 0.0001).

CONCLUSION

Use of coagulation factor concentrates for the reconstitution of blood allows for delivery of a higher haematocrit and a higher fibrinogen content compared with FFP. However, prothrombin complex concentrate might result in an unnecessary excess of thrombin generation. Clinical studies are warranted to further investigate these in vitro findings.
The role of organizational learning and resilience for change in building quality improvement capacity in primary care
Authors: Schuttner Linnaea , Coleman Katie , Ralston James , Parchman Michael
Published: April 2020

Background

The extent that organizational learning and resilience for the change process, that is, adaptive reserve (AR), is a component of building practice capacity for continuous quality improvement (QI) is unknown.

Purpose

The aim of the study was to examine the association of AR and development of QI capacity.

Methodology

One hundred forty-two primary care practices were evaluated at baseline and 12 months in a randomized trial to improve care quality. Practice AR was measured by staff survey along with a validated QI capacity assessment (QICA). We assessed the association of baseline QICA with baseline AR and both baseline and change in AR with change in QICA from 0 to 12 months. Effect modification by presence of QI infrastructure in parent organizations and trial arm was examined.

Results

Mean QICA increased from 6.5 to 8.1 (p < .001), and mean AR increased from 71.8 to 73.9 points (p < .001). At baseline, there was a significant association between AR and QICA scores: The QICA averaged 0.34 points higher (95% CI [0.04, 0.64], p = .03) per 10-point difference in AR. There was a significant association between baseline AR and 12-month QICA—which averaged 0.30 points higher (95% CI [0.02, 0.57], p = .04) per 10 points in baseline AR. There was no association between changes in AR and the QICA from 0 to 12 months and no effect modification by trial arm or external QI infrastructure.

Conclusions

Baseline AR was positively associated with both baseline and follow-up QI capacity, but there was no association between change in AR and change in the QICA, suggesting AR may be a precondition to growth in QI capacity.

Practice Implications

Findings suggest that developing AR may be a valuable step prior to undertaking QI-oriented growth, with implications for sequencing of development strategies, including added gain in QI capacity development from building AR prior to engaging in transformation efforts.