Biology subjects

Thomas, J. M.

Publications and source records attributed to Thomas, J. M..

11 recordsLinked to original sources

bioRxiv2026

Eicosapentaenoic Acid Enhances Angiogenesis and Reperfusion After Ischemia: Comparative Effects of Omega-3 Fatty Acids in a Murine Hind Limb Model

BackgroundOmega-3 polyunsaturated fatty acids are known to confer benefits in the prevention of cardiovascular diseases. Among these, eicosapentaenoic acid (EPA) appears to be more effective than docosahexaenoic acid (DHA) in ischemic vascular conditions. However, the specific roles of EPA and DHA in limb angiogenesis and post-ischemic reperfusion remain unclear. Moreover, omega-3 fatty acids remain understudied for peripheral artery disease (PAD) intervention. The aim was to compare the effect of high dose omega-3 fatty acids, EPA and DHA on ischemic tissue reperfusion, angiogenesis and vascular remodelling in mice. MethodsHind limb ischemia (HLI) was performed in mice and reperfusion was measured using laser speckle contrast imaging over two weeks post-HLI. Mice were treated daily with oral high-dose EPA or DHA (600 mg/kg/day) or vehicle (olive oil). Gastrocnemius muscle tissue was collected for analysis of mRNA and protein markers of angiogenesis. ResultsFollowing HLI, blood flow was restored more rapidly in mice treated with EPA compared with vehicle. DHA treatment did not enhance reperfusion. Histological assessment revealed significant muscle fibre regeneration after HLI, which was further improved by EPA. CD31+ neo vessel density was also increased in the EPA group. Collectively, these findings indicate that EPA promotes angiogenesis after peripheral vascular ischemia, whereas DHA does not. The beneficial effects of EPA are associated with upregulation of hypoxia inducible factor . ConclusionsHigh-dose EPA accelerated post-ischemic reperfusion, while DHA was ineffective. These results highlight EPA as a potential therapeutic strategy for improving limb perfusion and vascular repair in patients with PAD. Research PerspectiveO_ST_ABSWhat Is New?C_ST_ABSO_LIOmega-3 fatty acid, eicosapentaenoic acid (EPA) accelerates post-ischaemic reperfusion following hind limb ischemia injury in mice. C_LIO_LIOmega 3 fatty acid, docosahexaenoic acid (DHA), does not show the same improvement in reperfusion after hind limb ischemia C_LIO_LIEPA can promote vasculogenesis and stimulate muscle fibre regeneration. C_LI What question should be addressed next?O_LIHigh dose purified EPA, in the form of icosapent ethyl reduces mortality from coronary artery disease. Peripheral artery disease (PAD) is a common co-morbidity yet high quality interventional trials for icosapent ethyl for PAD are lacking. Therapeutic angiogenesis has the potential to improve PAD symptoms and disease progression but there are no efficacious candidates. Icosapent ethyl should be trialled to determine whether functional outcomes are improved in PAD. C_LI

physiology↗

bioRxiv2022

Dynamics of repurposing synthetic and natural compounds against WNT/β-Catenin signaling in glioma- An in vivo approach

BackgroundGlioblastomas arise from multistep tumorigenesis of the glial cells and are associated with poor prognosis. Despite the current state-of-art treatment, tumor recurrence is inevitable. Thus, there exists a desperate need for effective therapeutic alternatives to improve glioblastoma outcome. Among the innovations blooming up, drug repurposing could provide a profound premises for glioblastoma treatment enhancement. While considering this strategy, the efficacy of the repurposed drugs as monotherapies were not up to par; hence, the focus has now shifted to investigate the multi-drug combinations to target glioblastomas. In line with this concept, we investigated the efficacy of a quadruple-combinatorial treatment comprising temozolomide (the benchmark drug) along with chloroquine (a synthetic drug), naringenin (a flavonoid) and phloroglucinol (a marine derivative) in an orthotopic glioma-induced xenograft model. MethodsAnti-proliferative effect of the drugs was assessed by immunostaining. The expression profiles of WNT/{beta}-catenin and apoptotic markers were evaluated by qRT-PCR, immunoblotting and ELISA. Patterns of mitochondrial depolarization was determined by flow cytometry. TUNEL assay was performed to affirm apoptosis induction. In vivo drug detection study was carried out by ESI-Q-TOF MS analysis. ResultsThe quadruple-drug treatment had significantly hampered GB proliferation and had induced apoptosis by modulating the WNT/{beta}-catenin signalling. Flow cytometric analysis revealed that the induction of apoptosis was associated with mitochondrial depolarization. Further the quadruple-drug cocktail, had breached the blood brain barrier and was detected in the brain tissue and plasma samples from various experimental groups. ConclusionThe quadruple-drug combination served as a promising adjuvant therapy to combat glioma lethality in vivo and can be probed for translation from bench to bedside.

cancer biology↗

bioRxiv2021

Probing small ribosomal subunit RNA helix 45 acetylation across eukaryotic evolution

NAT10 is an essential enzyme that catalyzes the formation of N4-acetylcytidine (ac4C) in eukaryotic transfer RNA (tRNA) and 18S ribosomal RNA (rRNA). Recent studies in human cells suggested that rRNA acetylation is dependent on SNORD13, a non-canonical box C/D small nucleolar RNA (SNORD) predicted to base-pair with 18S rRNA via two antisense elements. However, the selectivity of SNORD13-dependent cytidine acetylation and its relationship to NAT10s essential function in pre-rRNA processing remain to be defined. Here, we used CRISPR-Cas9 genome editing to formally demonstrate that SNORD13 is required for acetylation of a single cytidine residue of human and zebrafish 18S rRNA. In-depth characterization revealed that SNORD13-dependent ac4C is dispensable for yeast or human cell growth, ribosome biogenesis, translation, and the development of multicellular metazoan model organisms. This loss of function analysis inspired a cross-evolutionary survey of the eukaryotic rRNA acetylation machinery that led to the characterization of many novel SNORD13 genes in phylogenetically-distant metazoans and more deeply rooted photosynthetic organisms. This includes an atypical SNORD13-like RNA in D. melanogaster which appears to guide ac4C to 18S rRNA helix 45 despite lacking one of the two rRNA antisense elements. Finally, we discover that C. elegans 18S rRNA is not acetylated despite the presence of an essential NAT10 homolog. Altogether, our findings shed light on the molecular mechanisms underlying SNORD13-mediated rRNA acetylation across the eukaryotic tree of life and raise new questions regarding the biological function and evolutionary persistence of this highly conserved rRNA base modification.

molecular biology↗

bioRxiv2021

A novel gene-by-environment quantitative trait locus on mouse chromosome 15 underlies susceptibility to acute ozone-induced lung injury

Respiratory toxicity caused by the common urban air pollutant ozone (O3) varies considerably within the human population and across inbred mouse strains, suggestive of gene-environment interactions (GxE). Though previous studies genetic mapping studies using classical inbred strains have identified several and quantitative trait locus (QTL) and candidate genes underlying responses to O3 exposure, precise mechanisms of susceptibility remain incompletely described. We sought to expand our understanding of the genetic architecture of O3 responsiveness using the Collaborative Cross (CC) recombinant inbred mouse panel, which contains more genetic diversity than previous inbred strain panels. We evaluated hallmark O3-induced respiratory phenotypes in 56 CC strains after exposure to filtered air or 2 ppm O3, and performed focused genetic analysis of variation in lung injury as measured by the total bronchoalveolar lavage protein concentration. Because animals were exposed in sex- and batch-matched pairs, we defined a protein response phenotype as the difference in lavage protein between the O3- and FA-exposed animal within a pair. The protein response phenotype was heritable, and QTL mapping revealed two novel loci on Chromosomes 10 (peak: 26.2 Mb; 80% CI: 24.6-43.6 Mb) and 15 (peak: 47.1 Mb; 80% CI: 40.2-54.9 Mb), the latter surpassing the 95% significance threshold. At the Chr. 15 locus, C57BL/6J and CAST/EiJ founder haplotypes were associated with higher protein responses compared to all other CC founder strain haplotypes. Using additional statistical analysis and high-density SNP data, we delimited the Chr. 15 QTL to a [~]2 Mb region containing 21 genes (10 protein coding). Using a weight of evidence approach that incorporated candidate variant analysis, functional annotations, and publicly available lung gene expression data, we nominated three candidate genes (Oxr1, Rspo2, and Angpt1). In summary, we have shown that O3-induced lung injury is modulated by genetic variation and demonstrated the value of the CC for uncovering and dissecting gene-environment interactions.

genetics↗

bioRxiv2021

Integrative phenotypic and genomic analyses reveal strain-dependent responses to acute ozone exposure and their associations with airway macrophage transcriptional activity

Acute ozone (O3) exposure is associated with multiple adverse cardiorespiratory outcomes, the severity of which varies across human populations and rodent models from diverse genetic backgrounds. However, molecular determinants of response, including biomarkers that distinguish which individuals will develop more severe injury and inflammation (i.e., high responders), are poorly characterized. Here, we exposed adult, female and male mice from 6 strains, including 5 Collaborative Cross (CC) strains, to filtered air (FA) or 2 ppm O3 for 3 hours, and measured several inflammatory and injury parameters 21 hours later. Additionally, we collected airway macrophages and performed RNA-seq analysis to investigate influences of strain, treatment, and strain-by-treatment interactions on gene expression as well as transcriptional correlates of lung phenotypes. Animals exposed to O3 developed airway neutrophilia and lung injury, with varying degrees of severity. We identified many genes that were altered by O3 exposure across all strains, and examination of genes whose expression was influenced by strain-by-treatment interactions revealed prominent differences in response between the CC017/Unc and CC003/Unc strains, which were low- and high-responders, respectively (as measured by cellular inflammation and injury). Further investigation of this contrast indicated that baseline gene expression differences likely contribute to their divergent post-O3 exposure transcriptional responses. We also observed alterations in chromatin accessibility that differed by strain and with strain-by-treatment interactions, lending further plausibility that baseline differences can modulate post-exposure responses. Together, these results suggest that aspects of the respiratory response to O3 exposure may be mediated through altered airway macrophage transcriptional signatures, and further confirms the importance of gene-by-environment interactions in mediating differential responsiveness to environmental agents.

pharmacology and toxicology↗

bioRxiv2020

Evaluation of Zoetis GeneMax Advantage genomic predictions in commercial Bos taurus Angus cattle

The GeneMax (GMX) Advantage test, developed by Zoetis, uses approximately 50,000 single nucleotide polymorphisms (SNP) to predict the genomic potential of a commercial Angus heifer. Genetic predictions are provided for Calving Ease Maternal, Weaning Weight, Heifer Pregnancy, Milk, Mature Weight, Dry Matter Intake, Carcass Weight, Marbling, Yield, and three economic selection indices. Test results can inform selection and culling decisions made by commercial beef cattle producers. To measure the accuracy of the trait predictions, data from commercial Angus females and their progeny at the University of Missouri Thompson Research Center were utilized to analyze weaning weight, milk, marbling, fat, ribeye area, and carcass weight. Progeny phenotypic data were matched to the respective dam, then the cows genomic predictions were compared to the calfs age-adjusted phenotypes using correlation and linear model effect sizes. All tested GeneMax scores of the dam were significantly correlated with and predicted calf performance. Our predicted effect sizes, except for fat thickness, were similar to those reported by Zoetis. In conclusion, the GeneMax Advantage test accurately ranks animals based on their genetic merit and is an effective selection tool in commercial cowherds.

genomics↗

bioRxiv2020

Evaluation of the 7 & 7 Synch and 7-day CO-Synch + CIDR protocols for estrus synchronization of beef cows prior to fixed-time artificial insemination with conventional or sex-sorted semen

An experiment was designed to compare the recently developed 7 & 7 Synch and the 7-day CO-Synch + CIDR protocols for synchronization of estrus among beef cows prior to fixed-time artificial insemination (FTAI) with conventional or sex-sorted semen. Bos taurus cows (n = 1538) were blocked based on age and days postpartum (DPP) and randomly assigned to protocol and semen type. Cows treated with the 7-day CO-Synch + CIDR protocol (n = 769) received administration of gonadotropin-releasing hormone (GnRH; 100 g gonadorelin) and insertion of a 1.38 g intravaginal progesterone releasing insert (CIDR) on Day -10, and administration of prostaglandin F2 (PG; 500 g cloprostenol) coincident with CIDR removal on Day -3. Cows treated with 7 & 7 Synch (n = 769) received PG and insertion of CIDR on Day -17, GnRH on Day -10, and PG coincident with CIDR removal on Day -3. Estrus detection aids (Estrotect) were applied to all cows on Day -3, and activation status was recorded at FTAI. Cows received FTAI 66 h after CIDR removal with either conventional (20 x 106 cells per unit) or sex-sorted (4 x 106 cells per unit; SexedULTRA 4M) semen. A greater proportion of cows expressed estrus (P = 0.01) prior to FTAI following 7 & 7 Synch (82%; 629/769) compared with the 7-day CO-Synch + CIDR (64%; 492/769). Estrus expression was also affected by protocol x DPP (P = 0.0004), with 7 & 7 Synch resulting in a greater increase in the proportion of cows expressing estrus prior to FTAI among cows with greater DPP. Across protocols, body condition score (BCS) affected (P = 0.05) estrus expression, with greater proportion of cows expressing estrus prior to FTAI at greater BCS. Sex-sorted semen resulted in reduced (P < 0.0001) pregnancy rates to FTAI. Irrespective of semen type, greater (P = 0.001) pregnancy rates to FTAI were obtained among cows treated with 7 & 7 Synch (conventional semen: 72% [280/389]; sex-sorted semen: 52% [199/380]) compared with the 7-day CO-Synch + CIDR (conventional semen: 60% [231/383]; sex-sorted semen: 44% [171/386]). In summary, 7 & 7 Synch resulted in an increased proportion of cows expressing estrus prior to FTAI and an increased pregnancy rate to FTAI with conventional and sexed semen. With these results and ease of application, 7 & 7 Synch offers potential as a platform to improve success with fixed-time AI in beef cows.

physiology↗

bioRxiv2020

Baseline and Innate Immune Response Characterization of a Zfp30 Knockout Mouse Strain

Airway neutrophilia is correlated with disease severity in a number of chronic and acute pulmonary diseases, and dysregulation of neutrophil chemotaxis can lead to host tissue damage. The gene Zfp30 was previously identified as a candidate regulator of neutrophil recruitment to the lungs and secretion of CXCL1, a potent neutrophil chemokine, in a genome-wide mapping study using the Collaborative Cross. ZFP30 is a putative transcriptional repressor with a KRAB domain capable of inducing heterochromatin formation. Using a CRISPR-mediated knockout mouse model, we investigated the role that Zfp30 plays in recruitment of neutrophils to the lung using models of allergic airway disease and acute lung injury. We found that the Zfp30 null allele did not affect CXCL1 secretion or neutrophil recruitment to the lungs in response to various innate immune stimuli. Intriguingly, despite the lack of neutrophil phenotype, we found there was a significant reduction in the proportion of live Zfp30 homozygous mutant mice produced from heterozygous matings. This deviation from the expected mendelian inheritance (i.e. transmission ratio distortion) implicates Zfp30 in fertility or embryonic development. Overall, our results indicate that Zfp30 is an essential gene but does not influence neutrophilic inflammation in this particular knockout model.

genetics↗

bioRxiv2019

Monodelphis domestica as a fetal intra-cerebral inoculation model for Zika virus pathogenesis

Monodelphis domestica, also known as the laboratory opossum, is a marsupial native to South America. At birth, these animals are developmentally equivalent to human embryos at approximately 5 weeks of gestation which, when coupled with other characteristics including the size of the animals, the development of a robust immune system during juvenile development, and the relative ease of experimental manipulation, have made M. domestica a valuable model in many areas of biomedical research. However, their suitability as models for infectious diseases, especially diseases caused by viruses such as Zika virus (ZIKV), is currently unknown. Here, we describe the replicative effects of ZIKV using a fetal intra-cerebral model of inoculation. Using immunohistochemistry and in situ hybridization, we found that opossum embryos and fetuses are susceptible to infection by ZIKV administered intra-cerebrally, that the infection persists long term, and that the infection and viral replication consistently results in neural pathology and may occasionally result in global growth restriction. These results demonstrate the utility of M. domestica as a new animal model for investigating ZIKV infection in vivo. This new model will facilitate further inquiry into viral pathogenesis, particularly for those viruses that are neurotropic, that may require a host with the ability to support sustained viral infection, and/or that may require intra-cerebral inoculations of large numbers of embryos or fetuses.\n\nAUTHOR SUMMARYHere we show that the laboratory opossum (Monodelphis domestica) is a valuable new model for studying Zika virus pathogenesis. Newborns are at the developmental stage of 5-week human embryos. Zika virus inoculated on a single occasion into the brains of pups at the human developmental stages of 8-20 weeks post conception replicated in neuronal cells and persisted as a chronic infection until the experimental endpoint at 74-days post infection. In addition, we observed global growth restriction in one of 16 inoculated animals; global growth restriction has been observed in humans and other animal models infected with Zika virus. The results illustrate great potential for this new animal model for high throughput research on the neurological effects of Zika virus infection of embryos and fetuses.

pathology↗

bioRxiv2019

Transcriptional profiling of the murine airway response to acute ozone exposure

Exposure to ambient ozone (O3) pollution causes airway inflammation, epithelial injury, and decreased lung function. Long-term exposure is associated with increased mortality and exacerbations of respiratory conditions. While the adverse health effects of O3 exposure have been thoroughly described, less is known about the molecular processes that drive these outcomes. The aim of this study was to describe the cellular and molecular alterations observed in murine airways after exposure to either 1 or 2 ppm O3. After exposing adult, female C57BL/6J mice to filtered air, 1 or 2 ppm O3 for 3 hours, we assessed hallmark responses including airway inflammatory cell counts, epithelial permeability, cytokine secretion, and morphological alterations of the large airways. Further, we performed RNA-seq to profile gene expression in two critical tissues involved in O3 responses: conducting airways (CA) and airway macrophages (AM). We observed a concentration-dependent increase in airway inflammation and injury, and a large number of genes were differentially expressed in both target tissues at both concentrations of O3. Genes that were differentially expressed in CA were generally associated with barrier function, detoxification processes, and cellular proliferation. The differentially expressed genes in AM were associated with innate immune signaling, cytokine production, and extracellular matrix remodeling. Overall, our study has described transcriptional responses to acute O3 exposure, revealing both shared and unique gene expression patterns across multiple concentrations of O3 and in two important O3-responsive tissues. These profiles provide broad mechanistic insight into pulmonary O3 toxicity, and reveal a variety of targets for refined follow-up studies.

pharmacology and toxicology↗

bioRxiv2018

The bacillithiol pathway is required for biofilm formation in Staphylococcus aureus

Staphylococcus aureus is a major human pathogen that can cause infections that range from superficial skin and mucosal infections to life threatening disseminated infections. S. aureus can attach to medical devices and host tissues and form biofilms that allow the bacteria to evade the host immune system and provide protection from antimicrobial agents. To counter host-generated oxidative and nitrosative stress mechanisms that are part of the normal host responses to invading pathogens, S. aureus utilizes low molecular weight (LMW) thiols, such as bacillithiol (BSH). Additionally, S. aureus synthesizes its own nitric oxide (NO), which combined with its downstream metabolites may also protect the bacteria against specific host responses. We have previously shown that LMW thiols are required for biofilm formation in Mycobacterium smegmatis and Pseudomonas aeruginosa. Our data show that the bshC mutant, which is defective in the last step of the bacillithiol pathway and lacks BSH, is impaired in biofilm formation. We also identify a putative S-nitrosobacillithiol reductase (BSNOR), similar to a S-nitrosomycothiol reductase found in M. smegmatis, and show that the BSNOR mutant has reduced levels of BSH and decreased biofilm formation. Our studies also show that NO plays an important role in biofilm formation and that acidified sodium nitrite severely reduces biofilm thickness. These studies provide insight into the roles of oxidative and nitrosative stress mechanisms on biofilm formation and indicate that bacillithiol and nitric oxide are key players in normal biofilm formation in S. aureus.\n\nImportanceStaphylococcus aureus is the most frequent cause of biofilm-associated infections in hospital settings. The biofilm mode of growth allows the pathogen to escape the host immune response and is extremely difficult to combat, as biofilms are highly resistant to physical and chemical stressors. As outbreaks of antibiotic-resistant bacterial strains become more commonplace, it is essential to understand the pathways involved in biofilm formation in order to target this important virulence factor. Low molecular weight thiols enable S. aureus to combat oxidative and nitrosative stress mechanisms that are used by host cells to defend against infection. Our findings indicate that bacillithiol and nitric oxide, which are produced by S. aureus to combat these host generated stressors, are important for biofilm development, and that disruption of these pathways results in biofilm defects. In the long term, this work may lead to new solutions to eradicate S. aureus biofilms in the clinic.

microbiology↗