Intact microdissection of stellate ganglia in a Parkinson\u27s disease model reveals aggregation of mutant human α-synuclein in their cell bodies

\ua9 2025 The Author(s). Experimental Physiology published by John Wiley & Sons Ltd on behalf of The Physiological Society.Cardiac dysautonomia plays an important role in understanding Parkinson\u27s disease (PD), with recent studies highlighting the presence of α-synuclein in cardiac tissue. We hypothesise that sympathetic dysregulation observed in PD may involve pathological changes caused by α-synuclein in stellate ganglia (SG). This study aimed to investigate α-synucleinopathy in SG of the genetic PD murine animal model. Mice overexpressing Ala30Pro (A30P) mutant α-synuclein were used. We here demonstrate a technique for meticulously dissecting SG. The collected SG from the transgenic mice were immunolabelled with neuronal markers, A30P human mutant α-synuclein and anti-α-synuclein aggregates. A30P mutant α-synuclein protein was expressed in the sympathetic neuronal (tyrosine hydroxylase (TH)-positive) cell bodies. Approximately 27% of the TH-positive cell bodies expressed the A30P mutant α-synuclein protein. The mutant protein was densely localised at the cardiopulmonary pole of the SG. Additionally, we observed that the A30P mutant protein formed fibril aggregation in the SG. Our findings suggest that α-synucleinopathy in the PD animal model can affect the sympathetic autonomic nervous system, providing insight for further research into targeting α-synuclein pathology in the SG as a potential link between cardiac dysautonomia and PD

Age-dependent reduction in voltage-gated inward sodium current and<em> Scn8a</em> gene expression in murine stellate ganglia

\ua9 2025 The Author(s). Annals of the New York Academy of Sciences published by Wiley Periodicals LLC on behalf of The New York Academy of Sciences.Stellate ganglia (SG) provide sympathetic innervation to the heart and may predispose the myocardial conducting system to arrhythmias. However, little is known about age-related changes in the electrophysiology of murine SG. We investigated alterations in the electrophysiological properties of SG with aging. The loose patch clamp technique was adapted to SG tissue to investigate the voltage-gated ionic currents in its neuronal cells. We compared SG and ventricular cells from young (4 months) and aged (13 months) C57BL/6J mice to explore age-related alterations in their voltage-gated ionic currents (n &gt; 30 patches, eight mice in each group). We observed that the voltage-gated inward sodium current (peak INa(Max)) was significantly decreased with aging in the SG, but not in the ventricle. Additionally, Scn8a gene expression, which encodes the Nav 1.6 channel, was decreased with aging in the SG. Application of loose patch clamp electrophysiology thus suggests that ionic current alterations with age in murine SG could contribute to cardiac autonomic dysregulation in geriatric conditions

In situ exogenous alpha-synuclein aggregates inhibit murine ventricular voltage-gated inward sodium and outward potassium currents

BackgroundAlpha-synuclein is associated with neurodegeneration in Parkinson\u27s disease (PD). Recent studies have increasingly recognized incidences of cardiac complaints in PD patients. In particular, the occurrence of arrhythmias in PD patients may indicate potential electrophysiological alterations in the heart. Alpha-synuclein aggregates have been known to have disruptive effects on cell membranes. However, the effect of alpha-synuclein on the heart and sympathetic neuronal tissues remains unknown.ObjectiveThis study investigated the electrophysiological effects of alpha-synuclein aggregates in myocardium and cardiac sympathetic nervous system, potentially reflecting cardiac electrophysiological alteration in PD.MethodsWe measured the in situ sodium and potassium currents from murine ventricular myocardium and stellate ganglia using the loose patch clamp technique. The tissues were exposed to bioactive alpha-synuclein aggregates, and currents were measured under three different conditions: baseline, alpha-synuclein treatment, and wash out.ResultsThe experiments showed that alpha-synuclein aggregates altered the maximum cardiac sodium current (INa(Max)) (ANOVA, p &lt; 0.008) and affected its gating properties for channel activation (ANOVA F2,54 = 6.408, p = 0.003) and inactivation (F2, 67 = 6.32, p = 0.003). The alpha-synuclein aggregates also reduced the maximum outward potassium current (IK(Max)) during channel activation (F2, 77 = 6.02, p = 0.002). However, the alpha-synuclein aggregates did not affect the ionic currents in the stellate ganglia.ConclusionsOur results demonstrate that extracellular alpha-synuclein aggregates can inhibit ventricular but not stellate ganglion ionic currents, suggesting a differential sensitivity between the myocardium and the stellate ganglia, and indicating a cardiac-specific toxicity of alpha-synuclein on cardiac electrophysiology.Alpha-synuclein is the pathological hallmark of Parkinson\u27s disease (PD). There is an increased recognition of incidences of heart-related complaints in PD patients. In particular, arrhythmias have been associated with PD. This may indicate potential pathological changes in the heart. Alpha-synuclein has been known to be toxic to mammalian cells. However, its effect has not been tested in the heart and heart-connecting nerve tissues. Therefore, we investigated the effect of alpha-synuclein aggregates on the heart and the heart-connecting nervous tissues. The heart and stellate ganglia conveying nerves to the heart were exposed to bioactive alpha-synuclein aggregates, and the ionic current in these tissues were measured. The results showed that alpha-synuclein aggregates reduced the size of the inward sodium current in the heart. However, the alpha-synuclein aggregates did not affect the ionic currents in the stellate ganglia. Our results revealed that alpha-synuclein aggregates can alter cardiac ionic currents, which imply that a cardiac-specific toxicity of alpha-synuclein aggregates on cardiac electrophysiology in PD

Prenatal exposures and exposomics of asthma

This review examines the causal investigation of preclinical development of childhood asthma using exposomic tools. We examine the current state of knowledge regarding early-life exposure to non-biogenic indoor air pollution and the developmental modulation of the immune system. We examine how metabolomics technologies could aid not only in the biomarker identification of a particular asthma phenotype, but also the mechanisms underlying the immunopathologic process. Within such a framework, we propose alternate components of exposomic investigation of asthma in which, the exposome represents a reiterative investigative process of targeted biomarker identification, validation through computational systems biology and physical sampling of environmental medi

Effects of Ferumoxides – Protamine Sulfate Labeling on Immunomodulatory Characteristics of Macrophage-like THP-1 Cells

Superparamagnetic Iron Oxide (SPIO) complexed with cationic transfection agent is used to label various mammalian cells. Labeled cells can then be utilized as an in vivo magnetic resonance imaging (MRI) probes. However, certain number of in vivo administered labeled cells may be cleared from tissues by the host's macrophages. For successful translation to routine clinical application of SPIO labeling method it is important that this mode of in vivo clearance of iron does not elicit any diverse immunological effects. The purpose of this study was to demonstrate that SPIO agent ferumoxides-protamine sulfate (FePro) incorporation into macrophages does not alter immunological properties of these cells with regard to differentiation, chemotaxis, and ability to respond to the activation stimuli and to modulate T cell response. We used THP-1 cell line as a model for studying macrophage cell type. THP-1 cells were magnetically labeled with FePro, differentiated with 100 nM of phorbol ester, 12-Myristate-13-acetate (TPA) and stimulated with 100 ng/ml of LPS. The results showed 1) FePro labeling had no effect on the changes in morphology and expression of cell surface proteins associated with TPA induced differentiation; 2) FePro labeled cells responded to LPS with slightly higher levels of NFκB pathway activation, as shown by immunobloting; TNF-α secretion and cell surface expression levels of CD54 and CD83 activation markers, under these conditions, were still comparable to the levels observed in non-labeled cells; 3) FePro labeling exhibited differential, chemokine dependent, effect on THP-1 chemotaxis with a decrease in cell directional migration to MCP-1; 4) FePro labeling did not affect the ability of THP-1 cells to down-regulate T cell expression of CD4 and CD8 and to induce T cell proliferation. Our study demonstrated that intracellular incorporation of FePro complexes does not alter overall immunological properties of THP-1 cells. The described experiments provide the model for studying the effects of in vivo clearance of iron particles via incorporation into the host's macrophages that may follow after in vivo application of any type of magnetically labeled mammalian cells. To better mimic the complex in vivo scenario, this model may be further exploited by introducing additional cellular and biological, immunologically relevant, components

Multi-Level Targeting of the Phosphatidylinositol-3-Kinase Pathway in Non-Small Cell Lung Cancer Cells

Introduction: We assessed expression of p85 and p110a PI3K subunits in non-small cell lung cancer (NSCLC) specimens and the association with mTOR expression, and studied effects of targeting the PI3K/AKT/mTOR pathway in NSCLC cell lines. Methods: Using Automated Quantitative Analysis we quantified expression of PI3K subunits in two cohorts of 190 and 168 NSCLC specimens and correlated it with mTOR expression. We studied effects of two PI3K inhibitors, LY294002 and NVP-BKM120, alone and in combination with rapamycin in 6 NSCLC cell lines. We assessed activity of a dual PI3K/mTOR inhibitor

The power of civilizational nationalism in Russian foreign policy making

The article draws upon theories of identity to understand Russian foreign policy towards Ukraine since 2000. The article argues that contemporary Russian foreign policy can be best understood as an articulation of ‘civilizational nationalism’ which relies on the myth of cultural superiority. The focus is on not only treating Russia as an imperial power, but on the cultural claims that this relies upon and its configuration within changing historical ideas of ‘Russianness’. Since the Orange Revolution, Russian presidents have accused Ukraine of following anti-Russian policies. This has been aided by a discourse of ‘civilizational nationalism’ where Ukraine is described as being part of a ‘Greater Russia’, rather than as a sovereign territory. This article analyses how imagined civilization and greatness of Russian culture is driving foreign policy making towards the Ukraine. Rather than an external territory, Ukraine is constructed as a ‘little brother’ which renders interventions legitimate

Mutation D816V Alters the Internal Structure and Dynamics of c-KIT Receptor Cytoplasmic Region: Implications for Dimerization and Activation Mechanisms

The type III receptor tyrosine kinase (RTK) KIT plays a crucial role in the transmission of cellular signals through phosphorylation events that are associated with a switching of the protein conformation between inactive and active states. D816V KIT mutation is associated with various pathologies including mastocytosis and cancers. D816V-mutated KIT is constitutively active, and resistant to treatment with the anti-cancer drug Imatinib. To elucidate the activating molecular mechanism of this mutation, we applied a multi-approach procedure combining molecular dynamics (MD) simulations, normal modes analysis (NMA) and binding site prediction. Multiple 50-ns MD simulations of wild-type KIT and its mutant D816V were recorded using the inactive auto-inhibited structure of the protein, characteristic of type III RTKs. Computed free energy differences enabled us to quantify the impact of D816V on protein stability in the inactive state. We evidenced a local structural alteration of the activation loop (A-loop) upon mutation, and a long-range structural re-organization of the juxta-membrane region (JMR) followed by a weakening of the interaction network with the kinase domain. A thorough normal mode analysis of several MD conformations led to a plausible molecular rationale to propose that JMR is able to depart its auto-inhibitory position more easily in the mutant than in wild-type KIT and is thus able to promote kinase mutant dimerization without the need for extra-cellular ligand binding. Pocket detection at the surface of NMA-displaced conformations finally revealed that detachment of JMR from the kinase domain in the mutant was sufficient to open an access to the catalytic and substrate binding sites

Is exposure to formaldehyde in air causally associated with leukemia?—A hypothesis-based weight-of-evidence analysis

Recent scientific debate has focused on the potential for inhaled formaldehyde to cause lymphohematopoietic cancers, particularly leukemias, in humans. The concern stems from certain epidemiology studies reporting an association, although particulars of endpoints and dosimetry are inconsistent across studies and several other studies show no such effects. Animal studies generally report neither hematotoxicity nor leukemia associated with formaldehyde inhalation, and hematotoxicity studies in humans are inconsistent. Formaldehyde's reactivity has been thought to preclude systemic exposure following inhalation, and its apparent inability to reach and affect the target tissues attacked by known leukemogens has, heretofore, led to skepticism regarding its potential to cause human lymphohematopoietic cancers. Recently, however, potential modes of action for formaldehyde leukemogenesis have been hypothesized, and it has been suggested that formaldehyde be identified as a known human leukemogen. In this article, we apply our hypothesis-based weight-of-evidence (HBWoE) approach to evaluate the large body of evidence regarding formaldehyde and leukemogenesis, attending to how human, animal, and mode-of-action results inform one another. We trace the logic of inference within and across all studies, and articulate how one could account for the suite of available observations under the various proposed hypotheses. Upon comparison of alternative proposals regarding what causal processes may have led to the array of observations as we see them, we conclude that the case fora causal association is weak and strains biological plausibility. Instead, apparent association between formaldehyde inhalation and leukemia in some human studies is better interpreted as due to chance or confounding