Showing papers by "University of Tennessee Health Science Center published in 2022"

A Draft Human Pangenome Reference

Abstract: Abstract The Human Pangenome Reference Consortium (HPRC) presents a first draft human pangenome reference. The pangenome contains 47 phased, diploid assemblies from a cohort of genetically diverse individuals. These assemblies cover more than 99% of the expected sequence and are more than 99% accurate at the structural and base-pair levels. Based on alignments of the assemblies, we generated a draft pangenome that captures known variants and haplotypes, reveals novel alleles at structurally complex loci, and adds 119 million base pairs of euchromatic polymorphic sequence and 1,529 gene duplications relative to the existing reference, GRCh38. Roughly 90 million of the additional base pairs derive from structural variation. Using our draft pangenome to analyze short-read data reduces errors when discovering small variants by 34% and boosts the detected structural variants per haplotype by 104% compared to GRCh38-based workflows, and by 34% compared to using previous diversity sets of genome assemblies.

The panoramic view of amyotrophic lateral sclerosis: A fatal intricate neurological disorder

Abstract: Amyotrophic lateral sclerosis (ALS) is a progressive and fatal neurological disease affecting both upper and lower motor neurons. In the United States alone, there are 16,000-20,000 established cases of ALS. The early disease diagnosis is challenging due to many overlapping pathophysiologies with other neurological diseases. The etiology of ALS is unknown; however, it is divided into two categories: familial ALS (fALS) which occurs due to gene mutations & contributes to 5-10% of ALS, and sporadic ALS (sALS) which is due to environmental factors & contributes to 90-95% of ALS. There is still no curative treatment for ALS: palliative care and symptomatic treatment are therefore essential components in the management of these patients. In this review, we provide a panoramic view of ALS, which includes epidemiology, risk factors, pathophysiologies, biomarkers, diagnosis, therapeutics (natural, synthetic, gene-based, pharmacological, stem cell, extracellular vesicles, and physical therapy), controversies (in the clinical trials of ALS), the scope of nanomedicine in ALS, and future perspectives.

Unbiased pangenome graphs

Abstract: Abstract Motivation Pangenome variation graphs model the mutual alignment of collections of DNA sequences. A set of pairwise alignments implies a variation graph, but there are no scalable methods to generate such a graph from these alignments. Existing related approaches depend on a single reference, a specific ordering of genomes, or a de Bruijn model based on a fixed k -mer length. A scalable, self-contained method to build pangenome graphs without such limitations would be a key step in pangenome construction and manipulation pipelines. Results We design the seqwish algorithm, which builds a variation graph from a set of sequences and alignments between them. We first transform the alignment set into an implicit interval tree. To build up the variation graph, we query this tree-based representation of the alignments to reduce transitive matches into single DNA segments in a sequence graph. By recording the mapping from input sequence to output graph, we can trace the original paths through this graph, yielding a pangenome variation graph. We present an implementation that operates in external memory, using disk-backed data structures and lock-free parallel methods to drive the core graph induction step. We demonstrate that our method scales to very large graph induction problems by applying it to build pangenome graphs for several species. Availability seqwish is published as free software under the MIT open source license. Source code and documentation are available at https://github.com/ekg/seqwish . seqwish can be installed via Bioconda https://bioconda.github.io/recipes/seqwish/README.html or GNU Guix https://github.com/ekg/guix-genomics/blob/master/seqwish.scm . Contact egarris5@uthsc.edu

Failure load of milled, 3D-printed, and conventional chairside-dispensed interim 3-unit fixed dental prostheses

Abstract: New techniques and materials for the laboratory fabrication of interim fixed dental prostheses have gained in popularity, yet how their failure strengths compare with conventional chairside materials is unclear.The purpose of this in vitro study was to compare the strength of computer-aided design and computer-aided manufacturing (CAD-CAM) milled polymethylmethacrylate (PMMA) or 3-dimensionally (3D) printed bis-acryl interim fixed dental prostheses with a traditional chairside-dispensed autopolymerizing bis-acryl prosthesis while taking into account the effect of loading rate and storage time.A dentiform mandibular second premolar and second molar with a first molar pontic were prepared and scanned. Three groups of 3-unit interim fixed dental prostheses were fabricated: milled PMMA, 3D-printed bis-acryl, and chairside-dispensed autopolymerizing bis-acryl. The interim prostheses were evaluated for fit with a silicone disclosing material and cemented onto 3D-printed resin dies. The specimens were stored in 100% humidity at 37 °C. After 1 or 30 days of storage, the cemented interim prostheses were loaded to failure in a universal testing machine at 1 or 10 mm/min (n=15/group). Failure loads were analyzed by 3-way analysis of variance and multiple comparisons (α=.05).Mean ±standard deviation failure loads ranged from 363 ±93 N (3D-printed bis-acryl, 30 days, 1 mm/min) to 729 ±113 N (milled PMMA, 24 hours, 1 mm/min). Loading rate did not significantly affect failure load of the interim prostheses (P=.306). After 30 days of storage in 100% humidity, the failure load of milled PMMA and 3D-printed bis-acryl interim prostheses decreased significantly, but the chairside autopolymerizing bis-acryl prostheses were not affected. After 30 days of storage, the failure loads of milled PMMA and chairside autopolymerizing bis-acryl were not significantly different.Regardless of loading rate, interim fixed dental prostheses from milled PMMA had the highest initial strength 1 day after storage. Thirty days of exposure to humidity, however, reduced the strength of the CAD-CAM-manufactured interim prostheses, whereas the traditional chairside prostheses retained their strength.

Systems genetics analysis defines importance of TMEM43/<i>LUMA</i> for cardiac- and metabolic-related pathways

Abstract: Broad cellular functions and diseases including muscular dystrophy, arrhythmogenic right ventricular cardiomyopathy (ARVC5) and cancer are associated with transmembrane protein43 (TMEM43/LUMA). The study aimed to investigate biological roles of TMEM43 through genetic regulation, gene pathways and gene networks, candidate interacting genes, and up- or downstream regulators. Cardiac transcriptomes from 40 strains of recombinant inbred BXD mice and two parental strains representing murine genetic reference population (GRP) were applied for genetic correlation, functional enrichment, and coexpression network analysis using systems genetics approach. The results were validated in a newly created knock-in Tmem43-S358L mutation mouse model (Tmem43S358L) that displayed signs of cardiac dysfunction, resembling ARVC5 phenotype seen in humans. We found high Tmem43 levels among BXDs with broad variability in expression. Expression of Tmem43 highly negatively correlated with heart mass and heart rate among BXDs, whereas levels of Tmem43 highly positively correlated with plasma high-density lipoproteins (HDL). Through finding differentially expressed genes (DEGs) between Tmem43S358L mutant and wild-type (Tmem43WT) lines, 18 pathways (out of 42 found in BXDs GRP) that are involved in ARVC, hypertrophic cardiomyopathy, dilated cardiomyopathy, nonalcoholic fatty liver disease, Alzheimer's disease, Parkinson's disease, and Huntington's disease were verified. We further constructed Tmem43-mediated gene network, in which Ctnna1, Adcy6, Gnas, Ndufs6, and Uqcrc2 were significantly altered in Tmem43S358L mice versus Tmem43WT controls. Our study defined the importance of Tmem43 for cardiac- and metabolism-related pathways, suggesting that cardiovascular disease-relevant risk factors may also increase risk of metabolic and neurodegenerative diseases via TMEM43-mediated pathways.

Longitudinal associations between insomnia symptoms and all-cause mortality among middle-aged and older adults: a population-based cohort study

Abstract: To date, there is no scientific consensus on whether insomnia symptoms increase mortality risk. We investigated longitudinal associations between time-varying insomnia symptoms (difficulty initiating sleep, difficulty maintaining sleep, early-morning awakening, and nonrestorative sleep) and all-cause mortality among middle-aged and older adults during 14 years of follow-up. Data were obtained from 2004 through 2018 survey waves of the Health and Retirement Study in the United States for a population-representative sample of 15 511 respondents who were ≥50 years old in 2004. Respondents were interviewed biennially and followed through the end of the 2018 survey wave for the outcome. Marginal structural discrete-time survival analyses were employed to account for time-varying confounding and selection bias. Of the 15 511 cohort respondents (mean [±SD] age at baseline, 63.7 [±10.2] years; 56.0% females), 5878 (31.9%) died during follow-up. At baseline (2004), 41.6% reported experiencing at least one insomnia symptom. Respondents who experienced one (HR = 1.11; 95% CI: 1.03-1.20), two (HR = 1.12; 95% CI: 1.01-1.23), three (HR = 1.15; 95% CI: 1.05-1.27), or four (HR = 1.32; 95% CI: 1.12-1.56) insomnia symptoms had on average a higher hazard of all-cause mortality, compared to those who were symptom-free. For each insomnia symptom, respondents who experienced difficulty initiating sleep (HR = 1.12; 95% CI: 1.02-1.22), early-morning awakening (HR = 1.09; 95% CI: 1.01-1.18), and nonrestorative sleep (HR = 1.17; 95% CI: 1.09-1.26), had a higher hazard of all-cause mortality compared to those not experiencing the symptom. The findings demonstrate significant associations between insomnia symptoms and all-cause mortality, both on a cumulative scale and independently, except for difficulty maintaining sleep. Further research should investigate the underlying mechanisms linking insomnia symptoms and mortality.

Strength and stiffness of interim materials and interim fixed dental prostheses when tested at different loading rates

Abstract: How the loading rate might affect the mechanical properties of interim materials and interim fixed dental prostheses is unclear.The purpose of this in vitro study was to compare the material stiffness, material strength, and structural strength of interim 3-unit fixed dental prostheses fabricated from 3 interim materials when stressed at different loading rates.Bar-shaped specimens and anatomically correct interim 3-unit fixed dental prostheses with a modified-ridge lap pontic were fabricated from polyethyl methacrylate resin (Trim) and 2 bis-acrylic composite resins (TempSmart; Integrity) (n=10). Flexural modulus and strength of the bar specimens, representing material stiffness and strength, were determined with a 4-point bend test in a universal testing machine. The structural strength of the prosthesis was assessed from the failure load from a vertical force applied on the occlusal surface of the pontic. Three loading rates, 0.5, 5, or 10 mm/min, were evaluated. Results were statistically analyzed with 2-way analysis of variance and multiple comparisons (α=.05).Loading rate and material significantly affected flexural modulus, flexural strength, and structural strength (P<.05). Increasing loading rate significantly increased the flexural modulus of all materials (P<.05), but the effect of loading rate on the flexural strength of bis-acrylic composite resins was mostly insignificant. Polyethyl methacrylate specimens did not fracture when loaded at 0.5 or 5 mm/min, and the interim fixed dental prostheses made from polyethyl methacrylate did not fracture at the 0.5 mm/min loading rate. Dual-polymerizing bis-acrylic composite resin had significantly higher flexural modulus and strengths than autopolymerizing bis-acrylic composite resin.Polyethyl methacrylate resin had the lowest stiffness among the interim materials tested and did not fracture but excessively deformed at the low loading rate. Dual-polymerizing bis-acrylic composite resin consistently had higher stiffness and material strength and provided higher structural strength than the autopolymerizing bis-acrylic composite resin. Loading rate significantly affected the mechanical properties of polyethyl methacrylate resin (P<.05), but the effect was indistinct for the bis-acrylic materials.

Comprehensive Review on Medicinal Applications of Coumarin-Derived Imine–Metal Complexes

Abstract: Coumarins are fused six-membered oxygen-containing benzoheterocycles that join two synthetically useful rings: α-pyrone and benzene. A survey of the literature shows that coumarins and their metal complexes have received great interest from synthetic chemists, medicinal scientists, and pharmacists due to their wide spectrum of biological applications. For instance, coumarin and its derivatives have been used as precursors to prepare a large variety of medicinal agents. Likewise, coumarin-derived imine–metal complexes have been found to display a variety of therapeutic applications, such as antibacterial, antifungal, anticancer, antioxidant, anthelmintic, pesticidal, and nematocidal activities. This review highlights the current synthetic methodologies and known bioactivities of coumarin-derived imine–metal complexes that make this molecule a more attractive scaffold for the discovery of newer drugs.

Use of Rapid Assessment Procedures when analyzing qualitative data in pharmacy research.

Abstract: Qualitative research analytics and methodology are a useful part of many research projects. However, qualitative data analysis may be time intensive causing delays in results. This is especially problematic in time-sensitive projects where there an urgent need for results and a rapidly evolving situation being studied, such as during health crisis or early stages of project implementation. An emerging body of literature around the use of Rapid Assessment Procedures (RAP) suggests that this method of qualitative assessment provides more efficient coding and categorizing of data without comprising rigor. The objectives of this manuscript are to: 1) describe how RAP can be used in pragmatic healthcare research studies and 2) provide an example of when RAP was applied to a qualitative research study in the healthcare setting. RAP includes 5 core features: 1) use in combination with quantitative outcomes or process data (mixed methods approach), 2) quick timeline from start to finish (weeks to months), 3) population of interest participation in planning and implementing the research, 4) team approach to research process, and 5) iterative cycle of data collection and analysis. Use of RAP provides key stakeholders and decision makers the ability to generate solutions to problems faster than ever before without compromising rigor, a method needed now more than ever. The progression of healthcare and clinical management is moving at an unprecedented rate, and RAP allows researchers to stay ahead by providing quicker results for better outcomes.

Characterization of Epigenomic Alterations in HPV16+ Head and Neck Squamous Cell Carcinomas

Abstract: Epigenetic changes associated with human papillomavirus (HPV)-driven tumors have been described; however, HPV type-specific alterations are less well understood. We sought to compare HPV16-specific methylation changes with those in virus-unassociated head and neck squamous cell carcinomas (HNSCC).Within The Cancer Genome Atlas, 59 HPV16+ HNSCC, 238 nonviral HNSCC (no detectable HPV or other viruses), and 50 normal head and neck tissues were evaluated. Significant differentially methylated regions (DMR) were selected, and key associated genes were identified. Partial least squares models were generated to predict HPV16 status in additional independent samples.HPV infection in HNSCC is associated with type-specific methylomic profiles. Multiple significant DMRs were identified between HPV16+, nonviral, and normal samples. The most significant differentially methylated genes, SYCP2, MSX2, HLTF, PITX2, and GRAMD4, demonstrated HPV16-associated methylation patterns with corresponding alterations in gene expression. Phylogenetically related HPV types (alpha-9 species; HPV31, HPV33, and HPV35) demonstrated a similar methylation profile to that of HPV16 but differed from those seen in other types, such as HPV18 and 45 (alpha-7).HNSCC linked to HPV16 and types from the same alpha species are associated with a distinct methylation profile. This HPV16-associated methylation pattern is also detected in cervical cancer and testicular germ cell tumors. We present insights into both shared and unique methylation alterations associated with HPV16+ tumors and may have implications for understanding the clinical behavior of HPV-associated HNSCC.HPV type-specific methylomic changes may contribute to understanding biologic mechanisms underlying differences in clinical behavior among different HPV+ and HPV- HNSCC.

C. elegans PEZO-1 is a mechanosensitive ion channel involved in food sensation.

Abstract: PIEZO channels are force sensors essential for physiological processes, including baroreception and proprioception. The Caenorhabditis elegans genome encodes an orthologue gene of the Piezo family, pezo-1, which is expressed in several tissues, including the pharynx. This myogenic pump is an essential component of the C. elegans alimentary canal, whose contraction and relaxation are modulated by mechanical stimulation elicited by food content. Whether pezo-1 encodes a mechanosensitive ion channel and contributes to pharyngeal function remains unknown. Here, we leverage genome editing, genetics, microfluidics, and electropharyngeogram recording to establish that pezo-1 is expressed in the pharynx, including in a proprioceptive-like neuron, and regulates pharyngeal function. Knockout (KO) and gain-of-function (GOF) mutants reveal that pezo-1 is involved in fine-tuning pharyngeal pumping frequency, as well as sensing osmolarity and food mechanical properties. Using pressure-clamp experiments in primary C. elegans embryo cultures, we determine that pezo-1 KO cells do not display mechanosensitive currents, whereas cells expressing wild-type or GOF PEZO-1 exhibit mechanosensitivity. Moreover, infecting the Spodoptera frugiperda cell line with a baculovirus containing the G-isoform of pezo-1 (among the longest isoforms) demonstrates that pezo-1 encodes a mechanosensitive channel. Our findings reveal that pezo-1 is a mechanosensitive ion channel that regulates food sensation in worms.

Estrogen receptor alpha and NFATc1 bind to a bone mineral density-associated SNP to repress WNT5B in osteoblasts

Abstract: Genetic factors and estrogen deficiency contribute to the development of osteoporosis. The single-nucleotide polymorphism (SNP) rs2887571 is predicted from genome-wide association studies (GWASs) to associate with osteoporosis but has had an unknown mechanism. Analysis of osteoblasts from 110 different individuals who underwent joint replacement revealed that the genotype of rs2887571 correlates with WNT5B expression. Analysis of our ChIP-sequencing data revealed that SNP rs2887571 overlaps with an estrogen receptor alpha (ERα) binding site. Here we show that 17β-estradiol (E2) suppresses WNT5B expression and further demonstrate the mechanism of ERα binding at the enhancer containing rs2887571 to suppress WNT5B expression differentially in each genotype. ERα interacts with NFATc1, which is predicted to bind directly at rs2887571. CRISPR-Cas9 and ChIP-qPCR experiments confirm differential regulation of WNT5B between each allele. Homozygous GG has a higher binding affinity for ERα than homozygous AA and results in greater suppression of WNT5B expression. Functionally, WNT5B represses alkaline phosphatase expression and activity, decreasing osteoblast differentiation and mineralization. Furthermore, WNT5B increases interleukin-6 expression and suppresses E2-induced expression of alkaline phosphatase during osteoblast differentiation. We show that WNT5B suppresses the differentiation of osteoblasts via receptor tyrosine kinase-like orphan receptor 1/2 (ROR1/2), which activates DVL2/3/RAC1/CDC42/JNK/SIN3A signaling and inhibits β-catenin activity. Together, our data provide mechanistic insight into how ERα and NFATc1 regulate the non-coding SNP rs2887571, as well as the function of WNT5B on osteoblasts, which could provide alternative therapeutic targets for osteoporosis.

<scp>STAT3</scp> suppresses the <scp>AMPKα</scp> / <scp>ULK1</scp> ‐dependent induction of autophagy in glioblastoma cells

Abstract: Despite advances in molecular characterization, glioblastoma (GBM) remains the most common and lethal brain tumour with high mortality rates in both paediatric and adult patients. The signal transducer and activator of transcription 3 (STAT3) is an important oncogenic driver of GBM. Although STAT3 reportedly plays a role in autophagy of some cells, its role in cancer cell autophagy remains unclear. In this study, we found Serine-727 and Tyrosine-705 phosphorylation of STAT3 was constitutive in GBM cell lines. Tyrosine phosphorylation of STAT3 in GBM cells suppresses autophagy, whereas knockout (KO) of STAT3 increases ULK1 gene expression, increases TSC2-AMPKα-ULK1 signalling, and increases lysosomal Cathepsin D processing, leading to the stimulation of autophagy. Rescue of STAT3-KO cells by the enforced expression of wild-type (WT) STAT3 reverses these pathways and inhibits autophagy. Conversely, expression of Y705F- and S727A-STAT3 phosphorylation deficient mutants in STAT3-KO cells did not suppress autophagy. Inhibition of ULK1 activity (by treatment with MRT68921) or its expression (by siRNA knockdown) in STAT3-KO cells inhibits autophagy and sensitizes cells to apoptosis. Taken together, our findings suggest that serine and tyrosine phosphorylation of STAT3 play critical roles in STAT3-dependent autophagy in GBM, and thus are potential targets to treat GBM.

When disadvantage collides: The concentrated effects of energy insecurity and internet burdens in the United States

Abstract: Energy and internet insecurity are exacerbated by the compounding of multiple forms of social-economic disadvantage during extreme events. This study demonstrates the effects of concentrated disadvantage on internet and energy burdens and utility hardships in the United States during the COVID-19 pandemic in 2021. Based on 1991 online respondents, we found that internet and energy burdens are higher in Florida than in California, but utility hardship is greater in California. Women, renters, low-income households, and people of color have higher internet and energy burdens than their counterparts. Unique to this study, people with higher energy medical needs are more likely to suffer from energy and internet insecurity than people without such needs. Low-income women, low-income homeowners, and homeowners of color with more energy medical needs have- higher energy burdens than their counterparts. Low-income men, people of color, and Black/Latino residents with higher levels of energy medical needs, and renters with disabilities and homeowners with medical needs affected by heating and cooling experienced higher levels of utility hardship than their counterparts. These findings suggest that energy insecurity is not just determined by income but by other social and health factors. The findings provide policy implications.

Targeting Genetic Modifiers of HBG Gene Expression in Sickle Cell Disease: The miRNA Option

Abstract: Sickle cell disease (SCD) is one of the most common inherited hemoglobinopathy disorders that affects millions of people worldwide. Reactivation of HBG (HBG1, HBG2) gene expression and induction of fetal hemoglobin (HbF) is an important therapeutic strategy for ameliorating the clinical symptoms and severity of SCD. Hydroxyurea is the only US FDA-approved drug with proven efficacy to induce HbF in SCD patients, yet serious complications have been associated with its use. Over the last three decades, numerous additional pharmacological agents that reactivate HBG transcription in vitro have been investigated, but few have proceeded to FDA approval, with the exception of arginine butyrate and decitabine; however, neither drug met the requirements for routine clinical use due to difficulties with oral delivery and inability to achieve therapeutic levels. Thus, novel approaches that produce sufficient efficacy, specificity, and sustainable HbF induction with low adverse effects are desirable. More recently, microRNAs (miRNAs) have gained attention for their diagnostic and therapeutic potential to treat various diseases ranging from cancer to Alzheimer’s disease via targeting oncogenes and their gene products. Thus, it is plausible that miRNAs that target HBG regulatory genes may be useful for inducing HbF as a treatment for SCD. Our laboratory and others have documented the association of miRNAs with HBG activation or suppression via silencing transcriptional repressors and activators, respectively, of HBG expression. Herein, we review progress made in understanding molecular mechanisms of miRNA-mediated HBG regulation and discuss the extent to which molecular targets of HBG might be suitable prospects for development of SCD clinical therapy. Lastly, we discuss challenges with the application of miRNA delivery in vivo and provide potential strategies for overcoming barriers in the future.