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Biology subjects

Woodruff, T. M.

Publications and source records attributed to Woodruff, T. M..

At least 19 recordsLinked to original sources

Consumption of processed foods impairs memory function through dietary advanced glycation end-products

Consumption of processed foods is associated with dementia, obesity, and other negative health outcomes. Sustained heat treatment, a common food processing approach to enhance flavor, induces the chemical Maillard reaction that promotes the formation of dietary advanced glycation end-products (AGEs). The neurocognitive impacts of consuming dietary AGEs are poorly understood. Here we modeled an AGE-rich diet through heat treatment fed to rats during adolescence, a critical period of neural development, to mechanistically evaluate the long-term impact of early life dietary AGEs on behavioral and neural processes. Consuming the AGE-rich diet impaired hippocampal-dependent memory function and altered the gut microbiome without inducing obesity or nonspecific behavioral deficits. AGE-induced memory deficits were coupled with impaired hippocampal glutamatergic synaptic neurotransmission and altered expression in the synapse-pruning complement system. Hippocampal synaptic deficits likely result from direct AGE-complement interactions, as our extended studies reveal competitive antagonist action of AGEs on complement receptors. Memory impairments were prevented by administration of the AGE-inhibitor, alagebrium, and by supplementation with an AGE-inhibiting bacterial taxon, Lactococcus lactis, which was depleted in the heat-treated diet. These findings reveal a functional connection between early life dietary AGEs, the microbiome, and memory impairments, thus illuminating mechanisms through which food processing negatively impacts neurocognition.

neuroscience↗

Targeting C5aR1 Reveals Protective Macrophage Maturation States in Intestinal Injury

Inflammatory intestinal injury, which may occur following cytotoxic cancer treatments such as radiotherapy, compromises homeostasis when intestinal stem cells undergo apoptosis and regeneration programmes are not effectively engaged. Recent evidence indicates the importance of macrophages in regulating stem regeneration following injury, but how to therapeutically promote specific macrophage subtypes which may tip the balance away from apoptosis and towards regeneration is still unclear. Here we show that targeting complement C5a receptor 1 (C5aR1) reduces radiation-induced intestinal injury by promoting macrophage phenotypes which reduce intestinal stem cell apoptosis. Specifically, C5aR1 signalling attenuates a macrophage maturation response characterised by increased IL10 and CX3CR1 expression. Our data indicate that increased IL10 signalling, and macrophage maturation induced following C5aR1 targeting are important for regulating crypt cell loss via radiotherapy-induced apoptosis. Interestingly, IL10-signalling is also critical for the improved tumour radiation response observed following C5aR1 targeting.

immunology↗

Paraquat Licences α-Synuclein to Trigger NLRP3-Dependent Pyroptosis in Microglia

Parkinsons disease (PD) is characterized by dopaminergic neurodegeneration and the pathological accumulation of -synuclein (Syn) aggregates. While its etiology is multifactorial, environmental toxicants such as paraquat, a widely used herbicide banned in over 70 countries but still permitted in some regions, including Australia, are strongly implicated. We previously showed that Syn activates the NLRP3 inflammasome in microglia, inducing interleukin-1{beta} (IL-1{beta}) release without triggering pyroptosis. Here, we examine how paraquat modulates microglial NLRP3 activation by Syn. Using primary microglia from wild-type (WT) and Nlrp3 knockout (Nlrp3-/-) mice, alongside human monocyte-derived microglia, we demonstrate that paraquat induces dose-dependent NLRP3 activation in LPS-primed WT microglia via NADPH oxidase-mediated reactive oxygen species (ROS) generation. This results in ASC speck formation and IL-1{beta} release, which is absent in Nlrp3-/- microglia and blocked by the selective NLRP3 inhibitor MCC950. While Syn alone fails to induce pyroptosis, co-treatment with paraquat resulted in caspase-1-dependent lactate dehydrogenase (LDH) release and gasdermin-D cleavage, hallmarks of pyroptotic cell death. This response was also suppressed by NLRP3 inhibition. Our findings reveal that paraquat acts as a critical second hit to license Syn-induced inflammasome-dependent pyroptosis in microglia, highlighting a mechanistic link between environmental toxin exposure and innate immune activation in PD pathogenesis. Targeting this axis with brain-penetrant NLRP3 inhibitors may offer an alternative therapeutic avenue for synucleinopathies associated with environmental toxins.

neuroscience↗

A systemic clock brake: Period1 stabilizes the circadian network under environmental stress

Precise alignment between internal circadian clocks and environmental light cycles is essential for physiological homeostasis and survival. However, the molecular mechanisms that preserve this synchrony across central and peripheral tissues remain poorly defined. Here, we uncover an unexpected role for the core clock gene Period1 (Per1) as a systemic modulator of circadian stability, regulating light-induced re-entrainment across the brain and body. In Per1-deficient mice, we show that loss of Per1 accelerates clock realignment, influencing transcriptomic, metabolic, hormonal, and behavioral indicators of circadian realignment across multiple organ systems, including the suprachiasmatic nucleus (SCN) and peripheral tissues such as the liver, adipose tissue, and adrenal glands. Notably, this accelerated adaptation confers protection against jetlag-induced sleep disturbances, weight gain, and metabolic imbalance, underscoring a systemic role for Per1 in maintaining circadian network stability. Mechanistically, unbiased spatial transcriptomics identified reduced expression of the arginine vasopressin (AVP), a key neuropeptide mediating SCN intercellular coupling, as the driver of circadian network instability. Weakened SCN synchrony permits enhanced flexibility of peripheral oscillator responses, expediting whole-body adaptation to shifted light-dark schedules. These findings position Per1 as a critical regulator of circadian robustness, a buffer against light over-responsiveness, identifying a potential molecular target for mitigating circadian misalignment in contexts such as jetlag, shift work, and metabolic disease. TeaserWhat if beating jetlag was as simple as switching off a gene? Researchers show that disabling Per1, a core circadian regulator, accelerates body clock realignment and protects against sleep and metabolic disruption--highlighting new therapeutic possibilities for jetlag and shift work.

neuroscience↗

Molecular fingerprints of a convergent mechanism orchestrating diverse ligand recognition and species-specific pharmacology at the complement anaphylatoxin receptors

Complement anaphylatoxin receptors (C3aR and C5aR1) are prototypical G protein-coupled receptors (GPCRs) playing crucial physiological roles in innate immunity by combating pathogenic infections and orchestrating inflammatory responses. They continue to be important therapeutic targets for multiple disorders including autoimmune diseases, acute and chronic inflammation, and allergy-related conditions. Recent structural coverage has provided important insights into their activation and signaling, however, confounding observations in the literature related to ligand efficacy and functional responses, especially in different model systems, present a major challenge for drug discovery efforts. Here, we systematically and comprehensively profile a broad set of natural and synthetic ligands at C3aR and C5aR1 and discover a previously unanticipated level of functional specialization in terms of species-specific pharmacology and receptor activation. Taking a lead from this, we determine seventeen cryo-EM structures of different ligand-receptor-G-protein complexes and uncover distinct orientation of agonists between the human and mouse receptors despite an overlapping positioning in the orthosteric binding pocket. Combined with extensive mutagenesis and functional assays, these structural snapshots allow us to decode and validate a convergent molecular mechanism involving a "Five-Point-Switch" in these receptors that orchestrates the recognition and efficacy of diverse agonists. We also identify species-specific differences at the level of phosphorylation patterns encoded in the carboxyl-terminus of these receptors and directly visualize their impact on {beta}arr binding and activation using cryo-EM structures. Interestingly, we observe that {beta}arrs engage with the mouse C5aR1 using a variation of previously discovered P-X-P-P phosphorylation motif via a "Sliding-Mechanism" and also exhibit distinct oligomeric state for the human vs. mouse receptors. Taken together, this study elucidates functional specialization at the complement anaphylatoxin receptors and underlying molecular mechanisms, offering a previously lacking framework with direct and immediate implications for the development of novel therapeutics.

biochemistry↗

Interaction of Gut-Microbial Amyloids with Endogenous Amyloids can Drives Microglial Hyperactivity and Neuroinflammation in Alzheimers Disease

Gut bacteria have emerged as silent drivers in the pathology of Alzheimers disease (AD). They also make amyloids with structure analogue to pathological amyloids and have potential to cross-seed and propagate in a prion-like manner. AD is characterised by the accumulation of mature extracellular Amyloid-{beta} (A{beta}) plaques which are surrounded by inflammatory microglia. We report that exposure to interspecies microbial amyloids of FapC (fimbriae) and CsgA (curli) from opportunistic gut pathogens Pseudomonas aeruginosa and Escherichia coli hyperactivates microglia against A{beta} fibrils. Microbial amyloids and A{beta} fibrils converge in phagocytic compartments through subsequent internalization, not observed with A{beta} fibrils alone. This convergence promotes pro-inflammatory microglia with a defective proteome similar to those observed in AD brains. The resulting clusters develop a pro-inflammatory, indigestible interactome that is eventually regurgitated, inducing progressive degeneration in bystander neurons and ultimately leading to cognitive decline. Collectively, these findings provide compelling evidence that microbial amyloids can trigger progressive AD pathology through microglia-driven neuroinflammation.

cell biology↗

Complement C5a receptor 1 blockade reverses cognitive deficits following cranialradiation therapy for brain cancer

Cranial radiation therapy (RT) for brain cancers leads to an irreversible decline in cognitive function without an available remedy. Radiation-induced cognitive deficits (RICD) are a particularly pressing problem for the survivors of pediatric and low grade glioma (LGG) cancers who often live long post-RT lives. Radiation-induced elevated neuroinflammation and gliosis, triggered by the detrimental CNS complement cascade, lead to excessive synaptic and cognitive loss. Using intact and brain cancer-bearing mouse models, we now show that targeting anaphylatoxin complement C5a receptor (C5aR1) is neuroprotective against RICD. We used a genetic knockout, C5aR1 KO mouse, and a pharmacologic approach, employing the orally active, brain penetrant C5aR1 antagonist PMX205 to reverse RICD. Irradiated C5aR1 KO and WT mice receiving PMX205 showed significant neurocognitive improvements in object recognition memory and memory consolidation tasks. Inhibiting C5a/C5aR1 axis reduced microglial activation, astrogliosis, and synaptic loss in the irradiated brain. Importantly, C5aR1 blockage in two syngeneic, orthotopic glioblastoma-bearing mice protected against RICD without interfering with the therapeutic efficacy of RT to reduce tumor volume in vivo. PMX205 clinical trials with healthy individuals and amyotrophic lateral sclerosis (ALS) patients showed no toxicity, drug-related adverse events, or infections. Thus, C5aR1 inhibition is a translationally feasible approach to address RICD, an unmet medical need. SIGNIFICANCECranial radiotherapy for brain cancers activates CNS complement cascade, leading to cognitive decline. Ablation of the complement C5a/C5aR1 axis alleviates radiation-induced neuroinflammation, synaptic loss, and cognitive dysfunction, providing a novel tractable approach.

neuroscience↗

The effect of a dominant kinase-dead Csf1r mutation associated with adult-onset leukoencephalopathy on brain development and neuropathology.

Amino acid substitutions in the kinase domain of the human CSF1R protein are associated with autosomal dominant adult-onset leukoencephalopathy with axonal spheroids and pigmented glia (ALSP). To model the human disease, we created a disease-associated mutation (Glu631Lys; E631K) in the mouse Csf1r locus. Previous analysis demonstrated that heterozygous mutation (Csf1rE631K/+) had a dominant inhibitory effect on CSF1R signaling in vitro and in vivo but did not recapitulate the pathology of the human disease. We speculated that leukoencephalopathy in humans requires an environmental trigger and/or epistatic interaction with common neurodegenerative disease-associated alleles. Here we examine the impact of heterozygous Csf1r mutation on microglial phenotype, normal postnatal brain development, age-related changes in gene expression and on two distinct pathologies in which microgliosis is a prominent feature, prion disease and experimental autoimmune encephalitis (EAE). The heterozygous Csf1rE631K/+ mutation reduced microglial abundance and the expression of microglial-associated transcripts relative to wild-type controls at 12 weeks and 43 weeks of age but had no selective effect on homeostatic markers such as P2ry12. An epistatic interaction was demonstrated between Csf1rE631K/+ and Cxc3r1EGFP/+ genotypes leading to dysregulated microglial and neuronal gene expression in both hippocampus and striatum. Heterozygous Csf1rE631K mutation reduced the microgliosis associated with both diseases. There was no significant impact on disease severity or progression in prion disease. In EAE, induced expression of inflammation-associated transcripts in the hippocampus and striatum was suppressed in parallel with microglia-specific transcripts, but spinal cord demyelination was exacerbated. The results support a dominant-negative model of CSF1R-associated leukoencephalopathy and likely contributions of an environmental trigger and/or genetic background to neuropathology.

neuroscience↗

Modulation of pedunculopontine input to the basal ganglia relieves motor symptoms in Parkinsonian mice

Locomotion, the ability to move, is driven by central pattern generators in the spinal cord. However, its initiation, selection of gait and arrest are controlled by command centers in the brain stem of which the best understood is the mesencephalic locomotor region (MLR). Stimulation of the MLR can initiate or arrest movement, but its neural projections and the neural circuits that mediate these actions are poorly understood. Here we show that glutamatergic neurons in the pedunculopontine nucleus (PPN), a major part of the MLR, project to the substantial nigra pars reticulata (SNr) where they innervate neurons that send afferents to the ventro medial motor thalamus (VM). Stimulation of this input to the SNr in mice slows voluntary and driven movement by modifying gait, with higher frequencies arresting gait. In Parkinsons disease (PD), a neurodegenerative disorder, the cardinal symptoms are bradykinesia, tremor and rigidity. Using an mouse model of PD, we show that inhibition of PPN input to the SNr rescues bradykinesia. This is the first description of a circuit in the mammalian brain that regulates movement and gait selection. Targeting of this circuit may be a more effective modality for treating the motor symptoms of PD.

neuroscience↗

Senolytic therapy alleviates physiological human brain aging and COVID-19 neuropathology.

Aging is the primary risk factor for most neurodegenerative diseases, and recently coronavirus disease 2019 (COVID-19) has been associated with severe neurological manifestations that can eventually impact neurodegenerative conditions in the long-term. The progressive accumulation of senescent cells in vivo strongly contributes to brain aging and neurodegenerative co-morbidities but the impact of virus-induced senescence in the aetiology of neuropathologies is unknown. Here, we show that senescent cells accumulate in physiologically aged brain organoids of human origin and that senolytic treatment reduces inflammation and cellular senescence; for which we found that combined treatment with the senolytic drugs dasatinib and quercetin rejuvenates transcriptomic human brain aging clocks. We further interrogated brain frontal cortex regions in postmortem patients who succumbed to severe COVID-19 and observed increased accumulation of senescent cells as compared to age-matched control brains from non-COVID-affected individuals. Moreover, we show that exposure of human brain organoids to SARS-CoV-2 evoked cellular senescence, and that spatial transcriptomic sequencing of virus-induced senescent cells identified a unique SARS-CoV-2 variant-specific inflammatory signature that is different from endogenous naturally-emerging senescent cells. Importantly, following SARS-CoV-2 infection of human brain organoids, treatment with senolytics blocked viral retention and prevented the emergence of senescent corticothalamic and GABAergic neurons. Furthermore, we demonstrate in human ACE2 overexpressing mice that senolytic treatment ameliorates COVID-19 brain pathology following infection with SARS-CoV-2. In vivo treatment with senolytics improved SARS-CoV-2 clinical phenotype and survival, alleviated brain senescence and reactive astrogliosis, promoted survival of dopaminergic neurons, and reduced viral and senescence-associated secretory phenotype gene expression in the brain. Collectively, our findings demonstrate SARS-CoV-2 can trigger cellular senescence in the brain, and that senolytic therapy mitigates senescence-driven brain aging and multiple neuropathological sequelae caused by neurotropic viruses, including SARS-CoV-2.

neuroscience↗

Innate immune receptor C5aR1 regulates cancer cell fate and can be targeted to improve radiotherapy in tumours with immunosuppressive microenvironments

An immunosuppressive microenvironment causes poor tumour T-cell infiltration and is associated with reduced patient overall survival in colorectal cancer. How to improve treatment responses in these tumours is still a challenge. Using an integrated screening approach to identify cancer-specific vulnerabilities, we identify complement receptor C5aR1 as a druggable target which when inhibited improves radiotherapy even in tumours displaying immunosuppressive features and poor CD8+ T-cell infiltration. While C5aR1 is well-known for its role in the immune compartment, we find that C5aR1 is also robustly expressed on malignant epithelial cells, highlighting potential tumour-cell specific functions. C5aR1 targeting results in increased NF-{kappa}B-dependent apoptosis specifically in tumours and not normal tissues; indicating that in malignant cells, C5aR1 primarily regulates cell fate. Collectively, these data reveal that increased complement gene expression is part of the stress response mounted by irradiated tumours and that targeting C5aR1 can improve radiotherapy even in tumours displaying immunosuppressive features.

cancer biology↗

The maternal microbiome regulates infant respiratory disease susceptibility via intestinal Flt3L expression and plasmacytoid dendritic cell hematopoiesis

Severe lower respiratory infection (sLRI) are a major cause of infant morbidity and mortality, and predispose to later chronic respiratory diseases such as asthma. Poor maternal diet during pregnancy is a risk factor for sLRI in the offspring. Here we demonstrate in mice that a maternal low-fibre diet (LFD) disrupts plasmacytoid and conventional dendritic cell (DC) hematopoiesis in the offspring, predisposing to sLRI and subsequent asthma. The LFD alters the composition of the maternal milk microbiome and assembling infant gut microbiome, ablating the induction of a developmental wave of the non-redundant DC growth factor Flt3L by neonatal intestinal epithelial cells. Therapy with a propionate-producing bacteria isolated from the milk of high-fibre diet-fed mothers, or supplementation with propionate, confers protection against sLRI by restoring gut Flt3L expression and pDC hematopoiesis. Our findings identify a microbiome-dependent Flt3L axis in the gut that regulates pDC hematopoiesis in early life and confers disease resistance. O_FIG O_LINKSMALLFIG WIDTH=200 HEIGHT=51 SRC="FIGDIR/small/522516v1_ufig1.gif" ALT="Figure 1"> View larger version (15K): org.highwire.dtl.DTLVardef@1a39990org.highwire.dtl.DTLVardef@1b6fe67org.highwire.dtl.DTLVardef@f8440dorg.highwire.dtl.DTLVardef@13bc67e_HPS_FORMAT_FIGEXP M_FIG C_FIG

immunology↗

SARS-CoV-2 drives NLRP3 inflammasome activation in human microglia through spike-ACE2 receptor interaction

Coronavirus disease-2019 (COVID-19) is primarily a respiratory disease, however, an increasing number of reports indicate that SARS-CoV-2 infection can also cause severe neurological manifestations, including precipitating cases of probable Parkinsons disease. As microglial NLRP3 inflammasome activation is a major driver of neurodegeneration, here we interrogated whether SARS-CoV-2 can promote microglial NLRP3 inflammasome activation utilising a model of human monocyte-derived microglia. We identified that SARS-CoV-2 isolates can bind and enter microglia, triggering inflammasome activation in the absence of viral replication. Mechanistically, microglial NLRP3 could be both primed and activated with SARS-CoV-2 spike glycoprotein in a NF-{kappa}B and ACE2-dependent manner. Notably, virus- and spike protein-mediated inflammasome activation in microglia was significantly enhanced in the presence of -synuclein fibrils, which was entirely ablated by NLRP3-inhibition. These results support a possible mechanism of microglia activation by SARS-CoV-2, which could explain the increased vulnerability to developing neurological symptoms akin to Parkinsons disease in certain COVID-19 infected individuals, and a potential therapeutic avenue for intervention. SIGNIFICANCE STATEMENTSevere acute respiratory syndrome coronavirus 2 (SARS-CoV-2) principally affects the lungs, however there is evidence that the virus can also reach the brain and lead to chronic neurological symptoms. In this study, we examined the interaction SARS-CoV-2 with brain immune cells, by using an ex-vivo model of human monocyte-derived microglia. We identified robust activation of the innate immune sensor complex, NLRP3 inflammasome, in cells exposed to SARS-CoV-2. This was dependent on spike protein-ACE2 receptor interaction and was potentiated in the presence of -synuclein. We therefore identify a possible mechanism for SARS-CoV-2 and increased vulnerability to developing neurological dysfunction. These findings support a potential therapeutic avenue for treatment of SARS-CoV-2 driven neurological manifestations, through use of NLRP3 inflammasome or ACE2 inhibitors.

neuroscience↗

SARS-CoV-2 triggers complement activation through interactions with heparan sulfate

The complement system has been heavily implicated in severe COVID-19 with clinical studies revealing widespread gene induction, deposition, and activation. However, the mechanism by which complement is activated in this disease remains incompletely understood. Herein we examined the relationship between SARS-CoV-2 and complement by inoculating the virus in lepirudin-anticoagulated human blood. This caused progressive C5a production after 30 minutes and 24 hours, which was blocked entirely by inhibitors for factor B, C3, C5, and heparan sulfate. However, this phenomenon could not be replicated in cell-free plasma, highlighting the requirement for cell surface deposition of complement and interactions with heparan sulfate. Additional functional analysis revealed that complement-dependent granulocyte and monocyte activation was delayed. Indeed, C5aR1 internalisation and CD11b upregulation on these cells only occurred after 24 hours. Thus, SARS-CoV-2 is a non-canonical complement activator that triggers the alternative pathway through interactions with heparan sulfate.

immunology↗

Intrinsic bias at non-canonical, β-arrestin-coupled seven transmembrane receptors

G protein-coupled receptors (GPCRs) are typically characterized by their seven transmembrane (7TM) architecture, and interaction with two universal signal-transducers namely, the heterotrimeric G-proteins and {beta}-arrestins ({beta}arrs). Synthetic ligands and receptor mutants have been designed to elicit transducer-coupling preferences and distinct downstream signaling outcomes for many GPCRs. This raises the question if some naturally-occurring 7TMRs may selectively engage one of these two signal-transducers, even in response to their endogenous agonists. Although there are scattered hints in the literature that some 7TMRs lack G-protein coupling but interact with {beta}arrs, an in-depth understanding of their transducer-coupling preference, GRK-engagement, downstream signaling and structural mechanism remains elusive. Here, we use an array of cellular, biochemical and structural approaches to comprehensively characterize two non-canonical 7TMRs namely, the human decoy D6 receptor (D6R) and the human complement C5a receptor (C5aR2), in parallel with their canonical GPCR counterparts, CCR2 and C5aR1, respectively. We discover that D6R and C5aR2 couple exclusively to {beta}arrs, exhibit distinct GRK-preference, and activate non-canonical downstream signaling partners. We also observe that {beta}arrs, in complex with these receptors, adopt distinct conformations compared to their canonical GPCR counterparts despite being activated by a common natural agonist. Our study therefore establishes D6R and C5aR2 as bona-fide arrestin-coupled receptors (ACRs), and provides important insights into their regulation by GRKs and downstream signaling with direct implications for biased agonism.

biochemistry↗

Targeting C5aR1 Increases the Therapeutic Window of Radiotherapy

Engaging innate immune pathways is emerging as a productive way of achieving durable anti-tumor responses. However, systemic administration of these therapies can result in toxicity, deemed to be particularly problematic when combined with current standard-of-care cytotoxic treatments such as radiotherapy. Increasing the therapeutic window of radiotherapy may be achieved by using targeted therapies, however, few pre-clinical studies investigate both tumor and normal tissue responses in detail. Here we show that targeting innate immune receptor C5aR1 improves tumor radiation response while reducing radiation-induced normal tissue toxicity, thereby increasing the therapeutic window. Genetically or pharmacologically targeting C5aR1 increases both IL-10 expression in the small intestine and IL-10 secretion by tumor cells. Increased IL-10 attenuates RelA phosphorylation and increases apoptosis in tumor cells, leading to improved radiation responses in murine models. Of note, these radiosensitizing effects are tumor-specific since, in the gastrointestinal tract, targeting C5aR1 instead results in decreased crypt cell apoptosis reduced signs of histological damage and improved survival following total abdominal irradiation in mice. Furthermore, the potent and orally active C5aR1 inhibitor, PMX205, improves tumor radiation responses even in a context of reduced/absent CD8+ T cell infiltration. These data indicate that PMX205 can modulate cancer-cell intrinsic functions to potentiate anti-tumor radiation responses even in tumors displaying features of T-cell deficiency or exclusion. Finally, using a preclinical murine model allowing the simultaneous assessment of tumor and normal tissue radiation responses, we show that PMX205 treatment reduces histological and functional markers of small-bowel toxicity while affording a positive tumor response. Our data, therefore, suggest that targeting C5aR1 could be a promising approach for increasing the therapeutic window of radiotherapy.

cancer biology↗

The 'C3aR antagonist' SB290157 is a partial C5aR2 agonist

Innate immune complement activation generates the C3 and C5 protein cleavage products C3a and C5a, defined classically as anaphylatoxins. C3a activates C3a receptors (C3aR), while C5a activates two receptors (C5aR1 and C5aR2) to exert their immunomodulatory activities. The non-peptide compound, SB290157, was originally reported in 2001 as the first C3aR antagonist. In 2005, the first report on non-selective nature of SB290157 was published, where the compound exerted clear agonistic, not antagonistic, activity in variety of cells. Other studies also documented non-selective activities of this drug in vivo. These findings severely hamper data interpretation regarding C3aR when using this compound. Unfortunately, given the dearth of C3aR inhibitors, SB290157 still remains widely used to explore C3aR biology (>70 publications to date). Given these issues, in the present study we aimed to further explore SB290157s pharmacological selectivity by screening the drug against three human anaphylatoxin receptors, C3aR, C5aR1 and C5aR2, using transfected cells. We first confirmed that SB290157 acts as a potent agonist at human C3aR. We also identified that SB290157 exerts partial agonist activity at C5aR2 by mediating {beta}-arrestin recruitment at higher compound doses. Notably, SB290157s activity at C5aR2 was more potent and efficacious than the current lead C5aR2 agonist P32. Notwithstanding this, SB290157 showed inhibitory effect on C3a-mediated signalling in primary human macrophages. Our results therefore provide even more caution against using SB290157 as a research tool to explore C3aR function. Given the reported immunomodulatory and anti-inflammatory activities of C3aR and C5aR2 agonism, any function observed with SB290157 could be due to these off target activities.

pharmacology and toxicology↗

Glucose clearance and uptake is increased in the SOD1G93A mouse model of amyotrophic lateral sclerosis in an insulin-independent mechanism

Metabolic disturbances are associated with the progression of the neurodegenerative disorder, amyotrophic lateral sclerosis (ALS), however the molecular events that drive energy imbalances in ALS are not completely understood. In this study we aimed to elucidate deficits in energy homeostasis in the SOD1G93A mouse model of ALS. We identified that SOD1G93A mice at mid-symptomatic disease stage have increased oxygen consumption and faster exogenous glucose uptake, despite presenting with normal insulin tolerance. Fasting glucose homeostasis was also disturbed, along with increased liver glycogen stores, despite elevated circulating glucagon, suggesting that glucagon signalling is impaired. Metabolic gene expression profiling of livers indicated that glucose cannot be utilised efficiently in SOD1G93A mice. Overall, we demonstrate that glucose homeostasis and uptake are altered in SOD1G93A mice, which is linked to an increase in insulin-independent glucose uptake and a disturbance in glucagon sensitivity, suggesting glucagon secretion and signalling could be potential therapeutic targets for ALS.

physiology↗