Biology subjects

Clayton, S. A.

Publications and source records attributed to Clayton, S. A..

5 recordsLinked to original sources

bioRxiv2025

Dexamethasone inhibits the LPS-induced activation of hypoxia-inducible factor 1 alpha (HIF-1α) in macrophages independently of the DUSP1-p38-TTP-HIF-1α regulatory axis.

The oxygen-sensitive transcription factor HIF-1 (hypoxia-inducible factor 1) is a master regulator of cellular and organism-level adaptations to decreasing oxygen availability. Under conditions of oxygen sufficiency HIF-1 is constitutively produced and degraded. In response to hypoxia, the degradation pathway is inhibited and HIF-1 protein accumulates via mechanisms that are now well understood. Additional signals combine with the oxygen-sensitive protein degradation pathway to modulate the expression and/or activity of HIF-1. In the context of the immune system, HIF-1 can be activated under normoxic conditions by a variety of stimuli, including the pro-inflammatory agonist lipopolysaccharide. Here we use a variety of genetic and pharmacological approaches to reveal that lipopolysaccharide-induced HIF-1 accumulation in primary macrophages is dependent on mitogen-activated protein kinase p38. This response is not dependent on modulation of HIF-1 protein stability: instead, it involves the phosphorylation and inactivation of tristetraprolin, an mRNA destabilizing protein that targets Hif1a mRNA for degradation. We previously reported that the glucocorticoid dexamethasone inhibits lipopolysaccharide-induced HIF-1 accumulation and metabolic reprograming in primary macrophages. Here we tested and disproved the hypothesis that dexamethasone prevents HIF-1 accumulation by inducing expression of dual specificity phosphatase 1 and thereby inhibiting p38 function. Hence two novel mechanisms critically regulate HIF-1 activation in lipopolysaccharide-treated macrophages: a p38-dependent mechanism that operates at the post-transcriptional level to control Hif1a mRNA expression, and a glucocorticoid-sensitive mechanism that operates at the post-translational level to control HIF-1 protein stability. Combined targeting of these two mechanisms may exert therapeutic effects in contexts where HIF-1 contributes to immune-mediated inflammatory pathology.

immunology↗

bioRxiv2024

Preclinical Multi-Omic Assessment of Pioglitazone in Skeletal Muscles of Mice Implanted with Human HER2/neu Overexpressing Breast Cancer Xenografts

Breast cancer (BC) is the most prevalent cancer worldwide and is accompanied by fatigue during both active disease and remission in the majority of cases. Our lab has measured fatigue in isolated muscles from treatment-naive BC patient-derived orthotopic xenograft (BC-PDOX) mice. Here, we conducted a preclinical trial of pioglitazone in BC-PDOX mice to determine its efficacy in ameliorating BC-induced muscle fatigue, as well as its effects on transcriptomic, metabolomic, and lipidomic profiles in skeletal muscle. MethodsThe pioglitazone and vehicle groups were treated orally for 4 weeks upon reaching a tumor volume of 600 mm3. Whole-animal indirect calorimetry was used to evaluate systemic metabolic states. The transcriptome was profiled using short-read bulk RNA sequencing (RNA-seq). Liquid chromatography-tandem mass spectrometry (LC-MS/MS) was used to profile the metabolome and lipidome. Fast and slow skeletal muscle function were evaluated using isolated ex vivo testing. ResultsPioglitazone was associated with a significant overall decrease in metabolic rate, with no changes in substrate utilization. RNA-seq supported the downstream effects of pioglitazone on target genes and displayed considerable upregulation of mitochondrial bioenergetic pathways. Skeletal muscle metabolomic and lipidomic profiles exhibited dysregulation in response to BC, which was partially restored in pioglitazone-treated mice compared to vehicle-treated BC-PDOX mice. Despite molecular support for pioglitazones efficacy, isolated muscle function was not affected by pioglitazone treatment. ConclusionsBC induces multi-omic dysregulation in skeletal muscle, which pioglitazone partially ameliorates. Future research should focus on profiling systemic metabolic dysfunction, identifying molecular biomarkers of fatigue, and testing alternative pioglitazone treatment regimens. Statement of Translational RelevanceBreast cancer-induced fatigue is a prevalent and debilitating symptom that affects a majority of patients, leading to early treatment discontinuation and poorer outcomes. Despite its significant impact on patient quality of life, there are currently no approved therapies for this condition. Our previous work in the clinically relevant breast cancer patient-derived orthotopic xenograft (BC-PDOX) mouse model suggests that disruptions in the PPAR{gamma} signaling pathway may contribute to the development of cancer-related fatigue. Using this model that recapitulates the fatigue phenotype observed in patients, we conducted a preclinical trial evaluating the FDA-approved PPAR{gamma} agonist, pioglitazone, as a treatment for fatigue. Our multi-omic analysis of skeletal muscle from BC-PDOX mice revealed that pioglitazone treatment partially restored dysregulated lipid profiles and mitochondrial bioenergetic transcriptomic alterations. These findings suggest that pioglitazone may have potential as a therapeutic option for managing cancer-related fatigue in breast cancer patients.

cancer biology↗

bioRxiv2023

The glucocorticoid dexamethasone inhibits HIF-1alpha stabilisation and metabolic reprogramming in lipopolysaccharide-stimulated primary macrophages.

Synthetic glucocorticoids are used to treat many chronic and acute inflammatory conditions. Frequent adverse effects of prolonged exposure to glucocorticoids include disturbances of glucose homeostasis, caused by changes of glucose traffic and metabolism in muscle, liver and adipose tissues. Macrophages are important targets for the anti-inflammatory actions of glucocorticoids. These cells rely on aerobic glycolysis to support various pro-inflammatory and antimicrobial functions. Employing a potent pro-inflammatory stimulus in two commonly-used model systems (mouse bone marrow-derived and human monocyte-derived macrophages), we showed that the synthetic glucocorticoid dexamethasone inhibited lipopolysaccharide-mediated activation of the hypoxia- inducible transcription factor HIF-1, a critical driver of glycolysis. In both cell types, dexamethasone-mediated inhibition of HIF-1 reduced the expression of the glucose transporter GLUT1, which imports glucose to fuel aerobic glycolysis. Aside from this conserved response, other metabolic effects of lipopolysaccharide and dexamethasone differed between human and mouse macrophages. These findings suggest that glucocorticoids exert anti-inflammatory effects by impairing HIF-1-dependent glucose uptake in activated macrophages. Furthermore, harmful and beneficial (anti-inflammatory) effects of glucocorticoids may have a shared mechanistic basis, depending on alteration of glucose utilisation. GRAPHICAL ABSTRACT O_FIG O_LINKSMALLFIG WIDTH=200 HEIGHT=128 SRC="FIGDIR/small/558626v1_ufig1.gif" ALT="Figure 1"> View larger version (24K): org.highwire.dtl.DTLVardef@4edbb5org.highwire.dtl.DTLVardef@13a5ab2org.highwire.dtl.DTLVardef@18776borg.highwire.dtl.DTLVardef@4816fa_HPS_FORMAT_FIGEXP M_FIG C_FIG

immunology↗

bioRxiv2023

Sexual Dimorphism of Skeletal Muscle in a Mouse Model of Breast Cancer: A Functional and Molecular Analysis

1.Breast cancer incidence in men is statistically rare; however, given the lack of screening in males, more advanced stages at initial diagnosis results in lower 5-year survival rates for men with breast cancer compared to women. A sexual dimorphism, with respect to the effect of tumor growth on cachexia incidence and severity, has also been reported across cancer types. The purpose of this study was to examine the sexual dimorphism of breast cancer as it pertains to skeletal muscle function and molecular composition. Using female and male transgenic PyMT mice, we tested the hypothesis that isometric contractile properties and molecular composition of skeletal muscle would be differentially affected by breast tumors. PyMT tumor-bearing mice of each sex, corresponding to maximal tumor burden, were compared to their respective controls. RNA-sequencing of skeletal muscle revealed different pathway alterations that were exclusive to each sex. Further, differentially expressed genes and pathways were substantially more abundant in female tumor mice, with only minimal dysregulation in male tumor mice, each compared to their respective controls. These differences in the transcriptome were mirrored in isometric contractile properties, with greater tumor-induced dysfunction in females than male mice, as well as muscle wasting. Collectively, these data support the concept of sexually dimorphic responses to cancer in skeletal muscle and suggest these responses may be associated with the clinical differences in breast cancer between the sexes. The identified sex-dependent pathways within muscle of male and female mice provide a framework to evaluate therapeutic strategies targeting tumor-associated skeletal muscle alterations. Statement of significanceThe PyMT mouse model of breast cancer, which recapitulates clinical characteristics, exhibits differences in molecular and functional responses of skeletal muscle that are sex-dependent.

cancer biology↗

bioRxiv2023

The glucocorticoid dexamethasone impairs the expression of anti-viral mediators in activated macrophages by inhibiting both expression and function of interferon beta.

Glucocorticoids potently inhibit expression of many inflammatory mediators, and have been very widely used to treat both acute and chronic inflammatory diseases for more than seventy years. However, they can have several unwanted effects, amongst which immunosuppression is one of the most common. Here we investigated effects of the synthetic glucocorticoid dexamethasone on the responses of primary mouse bone marrow-derived macrophages to the pro-inflammatory agonist lipopolysaccharide (LPS). At the mRNA level, dexamethasone inhibited the LPS-induced expression of more than 100 genes that are involved in cell-intrinsic defence against viral pathogens. Expression of most of the corresponding proteins was also reduced by dexamethasone. This antiviral disarmament occurred at two distinct levels. First, dexamethasone strongly and dose-dependently inhibited the expression of the type I interferon IFN{beta} by LPS-activated macrophages. IFN{beta} mediates an autocrine positive feedback loop in LPS-treated macrophages, promoting the expression of antiviral genes and other interferon-stimulated genes. Hence reduction of IFN{beta} expression contributes to impaired expression of antiviral genes. Dexamethasone also acted downstream of IFN{beta} to inhibit expression of a subset of interferon-regulated genes. We tested a number of hypotheses based on previous publications, but found that no single mechanism could account for more than a small fraction of the broad suppressive impact of dexamethasone on macrophage type I interferon signaling, underlining the complexity of this pathway. Preliminary experiments indicated that dexamethasone exerted similar inhibitory effects on primary human monocyte-derived or alveolar macrophages.

immunology↗