Mould, Sustained Stress, and PTSD Converge on the RAGE–TLR4 Axis: A Unifying Model of Neuroimmune Dysregulation

Dr Graham Exelby May 2025

Abstract 

Chronic mould exposure, sustained psychological stress, and PTSD have emerged as key environmental drivers of persistent immune activation, neuroinflammation, and autonomic dysregulation. This paper explores how these factors converge mechanistically on a shared axis involving Toll-like receptors (particularly TLR2 and TLR4), the Receptor for Advanced Glycation End Products (RAGE), and downstream NF-κB signaling. These pathways initiate a feed-forward loop of mast cell degranulation, microglial activation, astrocyte dysfunction, and mitochondrial injury. The resulting disruption of glymphatic clearance, glutamate excitotoxicity, and blood–brain barrier integrity forms a pathophysiological bridge between mould toxicity, trauma-driven sensitization, and post-viral syndromes such as Long COVID.

These pathways trigger NF-κB-mediated cytokine release, mast cell degranulation, microglial priming, and astrocyte dysfunction, culminating in a self-perpetuating loop of glymphatic impairment, excitotoxicity, and barrier breakdown. RAGE-TLR4 crosstalk, mitochondrial DAMPs, and glutamate-mediated sensitization are highlighted as key intersections in the pathophysiology of POTS, fibromyalgia, and Long COVID.

This central hypothesis, that  chronic mould exposure, sustained stress, and PTSD converge on a shared neuroimmune axis involving TLR4, RAGE, and NF-κB-is strongly supported by current immunological, neuroscientific, and translational research.   The roles of mast cells, microglia, astrocytes, mitochondrial dysfunction, and chemokines like CCL2 are all well-evidenced in the literature.  The paper acknowledges the evolving nature of the research in this field culminating in a feed-forward loop of glymphatic impairment, excitotoxicity, and barrier breakdown. Recent evidence of reversible PEM through lymphatic decompression underscores the relevance of extracellular toxin clearance as a modifiable therapeutic target in this model.

Introduction 

In the complex pathophysiology that underlies POTS, fibromyalgia, and their comorbidities, persistent mould exposure, sustained stress, and PTSD stand out in many patients. These environmental and psychological insults trigger immune dysregulation through activation of NF-κB, with cascading downstream effects. Such stressors may also worsen complications such as Long COVID, as viral persistence and inflammation intersect with disrupted neurotransmitter dynamics.

Emerging evidence positions RAGE as a central amplifier in this pathophysiological triad. Chronic activation of RAGE by endogenous and exogenous ligands—including AGEs, mycotoxins, mitochondrial DNA, and S100 proteins—amplifies NF-κB-driven inflammation, primes glial cells, and activates mast cells. We propose that this neuroimmune axis, underpinned by environmental biotoxins and traumatic stress, sustains the inflammation seen in these chronic syndromes.

The overlap of PTSD, chronic stress, and mould exposure is increasingly observed, particularly in flood-affected areas where individuals may face both environmental and psychological insults. This creates a perfect storm of immune dysregulation through parallel TLR2/NF-κB and RAGE pathway activation.

TLR4 signalling is a central player in this process, serving as a link between environmental stressors and the immune response, especially in the combined PTSD/ mould exposure.  Dysfunctional TLR4 signalling, resulting from chronic activation, may fail to resolve inflammation, leading to a sustained inflammatory state that can potentially break immune tolerance and contribute to autoimmune processes.  

RAGE perpetuates NF-κB activation via a positive feedback loop, leading to chronic glial activation. Crosstalk with TLR4 synergistically activates the NLRP3 inflammasome, resulting in IL-1β/IL-18 release and pyroptosis.   Pyroptosis is a type of programmed cell death that's characterized by its inflammatory nature. It's often triggered by infections and other stimuli, and involves cell swelling, plasma membrane rupture, and the release of pro-inflammatory cytokines. This cell death process is crucial for clearing pathogens and activating the immune system, but it can also contribute to disease.

TLR and RAGE Crosstalk in Blood-Brain Barrier Dysfunction 

The blood-brain barrier (BBB), comprising endothelial cells, pericytes, and astrocyte end-feet, protects the CNS from systemic immune stimuli.  This barrier is essential in maintaining CNS homeostasis and preventing infections from reaching the vulnerable neural tissue. Under normal conditions, the BBB tightly regulates the passage of molecules and immune cells. However, increasing evidence shows that chronic systemic inflammation—such as that triggered by mould-related biotoxins, mycotoxins, and pathogen-associated molecular patterns (PAMPs)—can compromise BBB integrity.

Activation of the TLR4–NF-κB axis by environmental toxins induces endothelial cell dysfunction, loss of tight junction proteins (e.g., claudins and occludins), and perivascular inflammation. Furthermore, the Receptor for Advanced Glycation End Products (RAGE), which is upregulated in response to oxidative stress and hypoxia, facilitates the translocation of inflammatory ligands such as HMGB1, S100 proteins, and mycotoxin-induced DAMPs across the BBB, sustaining a neuroinflammatory loop.

However, in the context of chronic mould exposure, this response can become dysregulated, leading to sustained microglial activation, astrocyte reactivity, and progressive neuroinflammation. This contributes to symptoms observed in mould-affected individuals, including cognitive dysfunction, fatigue, central sensitization, and altered autonomic regulation—hallmarks of neuroimmune sensitization and brainstem vulnerability.

Once breached, BBB permeability allows for neuroinflammatory ligands to access CNS parenchyma, triggering microglial activation. Microglia, as primary CNS immune sentinels, react rapidly by producing cytokines and reactive oxygen species, but when chronically activated by sustained insults, their inflammatory state persists.

Figure 1. TLR Signalling in innate immune cells

Source: Duan T, Du Y, Xing C, Wang HY, Wang RF. Toll-Like Receptor Signaling and Its Role in Cell-Mediated Immunity. Front Immunol. 2022 (1)

Figure 2. TLR4 and RAGE Signalling Integration: This diagram illustrates the convergence of TLR4 and RAGE pathways upon stimulation by ligands such as LPS and HMGB1, leading to downstream activation of NF-κB and MAPK pathways.

Source: Constructed by ChatGPT

Toll-Like Receptors

 To understand what is happening we start with the immune response to COVID.   The pathophysiology of COVID-19 is characterized by systemic inflammation, hypoxia resulting from respiratory failure, and neuroinflammation (either due to viral neurotropism, the ability of the virus to invade and live in neural tissue, or in response to the cytokine storm), affecting the brain. (Phulwani et al. 2008 (2))  

In humans there are 10 types of body threat receptors, or Toll-Like Receptors (TLRs) that respond to a variety of PAMPs (pathogen-associated molecular patterns associated with bacteria and viruses).   TLRs are crucial components in the initiation of the innate immune system, triggering the downstream production of pro-inflammatory cytokines, interferons (IFNs) and other mediators. 

 TLRs recognize invading pathogens by sensing PAMP and activate the regulation of innate immunity and cytokines. TLR activation leads to the production of proinflammatory cytokines and Interferon IFN.(Mantovani et al 2023 (3))  TLRs 1,2,4,5,6,10 are plasma protein TLRs, while TLR3 and 7 and TLR9 are on endosomes (intracellular sorting organelles). TLR2/6 and TLR4 are located on the cell membrane.(Figure 3) 

TLR4 signalling in COVID is activated by the Spike protein (S).  This can lead to a pro-thrombotic and pro-inflammatory state contributing to severe complications eg myocardial infarction and acute lung injury.(Aboudounya & Heads. 2021(4))(van der Donk et al. 2023 (5))

Figure 3. Downstream Signalling Pathways of TLRs

Source: Mantovani S, Oliviero B, Varchetta S, Renieri A, Mondelli MU. TLRs: Innate Immune Sentries against SARS-CoV-2 Infection. Int J Mol Sci. 2023 (3)

Toll-like receptors TLR5, TLR4 and the heterodimers of TLR2-TLR1 or TLR2-TLR6 prefer to recognize the membrane components of pathogens on the cell surface.  TLR3, TLR7-TLR8 and TLR9 localize to the endosome where they recognize nucleic acids from both the host and pathogens.  TLR4 localizes at the plasma membrane but it is endocytosed into endosomes upon activation.    The TLR signalling switches from MyD88 to TRIF once TLR4 moves to the endosomes. (van der Donk et al. 2023 (5))

TLR2 senses the SARS-CoV-2 envelope protein (E), resulting in production of inflammatory cytokines and chemokines, contributing to the hyperinflammatory state and tissue damage seen in severe Covid.  The severity of the Covid infection is thought to be largely determined by the E Protein /TLR2 activation rather than the S protein.(Mantovani et al. 2023 (3))(Sariol & Perlman. 2021(7))

The endosomal TLR3 senses intracellular viral dsRNA.  Activated TLR regulates the production of proinflammatory factors through a series of signalling in the NF‐κB pathway and activates IRF3/7 to produce I IFN.   A DNA variant in TLR3 has also been identified as increasing susceptibility and mortality to acute COVID infections by decreasing TLR3 expression and impairing recognition of SARS-Co-V dsRNA. (Mantovani et al. 2023 (3))(Jiang et al 2022 (8))

TLR9 activation helps limit excessive inflammatory responses following traumatic experiences.  The failure of TLR9 to limit inflammation appears to play a significant role in post-traumatic stress disorder (PTSD) and associated anxiety as it serves as a crucial protective mechanism against post-traumatic inflammation.   Genetic polymorphisms in the TLR9 gene may predispose individuals to TLR9 dysfunction which could potentially affect its expression or function, altering the innate immune response and increasing susceptibility to post-traumatic inflammation and anxiety.(Zimmerman et al. 2012.(9)) 

TLR Signalling and Inflammation:  

 TLR4 is a key pattern recognition receptor that can be activated by both pathogen-associated molecular patterns (PAMPs) and damage-associated molecular patterns (DAMPs).  In the context of PTSD, chronic stress, and mould exposure:  TLR4 activation triggers inflammatory signalling cascades, including the NF-κB pathway, leading to the production of pro-inflammatory cytokines. (Souza-Junior et al. 2022 (10))

• Mould components, particularly mycotoxins, can act as PAMPs and activate TLR2, TLR4, and TLR6, triggering downstream MyD88 and TRIF pathways as well as RAGE (Kondashevskaya et al. 2024 (11))

• TLR2/4 signalling activates NF-κB and IRF3/7, upregulating IL-6, IL-1β, and TNF-α.

• The mycotoxin-induced TLR4 activation mimics the inflammatory amplification seen in severe viral infections, including COVID-19.

• Chronic stress can lead to the release of endogenous DAMPs that activate TLR4 and RAGE.( Souza-Junior et al. 2022 (10))

  
  

RAGE Activation

The Receptor for Advanced Glycation End Products (RAGE) is an underrecognized but critically important pattern recognition receptor (PRR) involved in sustaining sterile inflammation—that is, inflammation not caused by pathogens but by endogenous stress signals.

RAGE binds a range of ligands including HMGB1, S100 proteins, amyloid fibrils, oxidized lipids, and advanced glycation end products (AGEs)—many of which are elevated in tissue hypoxia, oxidative stress, or metabolic dysfunction. Once activated, RAGE initiates intracellular signalling cascades that culminate in NF-κB activation, amplifying production of CCL2, IL-6, and other inflammatory mediators.

In dysautonomia, RAGE is particularly relevant to vascular inflammation, endothelial barrier dysfunction, microglial activation, and central sensitization. Its expression is heightened in the brainstem and peripheral nerves under conditions of chronic hypoxia or mitochondrial impairment—both key features in POTS and post-COVID syndromes.

RAGE also localizes to cerebral endothelial cells, vagal afferents, and the coeliac and cardiac plexuses, placing it anatomically and mechanistically at the centre of autonomic disruption.

Importantly, RAGE activation is not self-limiting; it amplifies its own ligands through oxidative feedback, establishing a pathological positive feedback loop. This makes it a critical target in conditions driven by unresolved immune activation, with famotidine, nicotinamide, and TLR4-RAGE inhibitors offering potential avenues for intervention.

RAGE activation represents a parallel and synergistic pathway to TLR4, triggered by a variety of DAMPs such as S100 proteins, HMGB1, Se Amyloid A,AGEs, and oxidized phospholipids. These ligands are elevated in oxidative stress and tissue injury, including mould exposure and trauma.

RAGE amplifies NF-κB and MAPK signalling, sustains cytokine release, and primes the NLRP3 inflammasome. The RAGE-NF-κB axis is particularly active in astrocytes, endothelial cells, and microglia—disrupting glymphatic flow, increasing BBB permeability, and contributing to excitotoxicity and neuroinflammation.

RAGE / NF-κB activation via a positive feedback loop, leads to chronic glial activation. Crosstalk with TLR4 synergistically activates the NLRP3 inflammasome, resulting in IL-1β/IL-18 release and pyroptosis.   The complex nature of the immune response and mast cell activation in now an integral part of Long Covid pathogenesis.   The same microglial activation has been demonstrated in other conditions -ME/CFS, ADHD, migraine, Fibromyalgia syndrome  and Endometriosis. (Exelby 2025 (12))

NFkB activation

 Nuclear Factor kappa B (NFkB) is a protein complex that plays a crucial role in regulating the immune response, inflammation, and cell survival. The primary function of NFkB is to control gene expression in response to various signals, such as pro-inflammatory cytokines, bacterial or viral products, stress, and oxidative damage.  NF-κB has long been considered a prototypical proinflammatory signalling pathway, largely based on the activation of NF-κB by proinflammatory cytokines such as interleukin 1 (IL-1) and tumour necrosis factor α (TNFα), and the role of NF-κB in the expression of other proinflammatory genes. 

Anilkumar & Wright-Jin 2024 (13) describe that under normal situations NF-kB  is inactivated and sequestered to the cytoplasm either by natural inhibitors of  NF-kB or inherent structural inactivation.   Dysregulation of NF-kB  signalling has been implicated in various health conditions, including processes of synaptic plasticity and memory, autoimmune disorders, inflammatory diseases, cancer, and neurodegenerative diseases.  Acting as primary immune responders in the CNS, microglia upregulate  NFkB and cross-talk with other cells in the CNS can induce cell death, exacerbating the disease pathology. (Figure 3) 

NF-kB  plays a multi-faceted role in the CNS, with diverse roles depending on physiological conditions.   NF-kB  activation in neurons is essential for information processing and transmission with both neuroprotective and neurodegenerative effects, and a key modulator of neuron survival, and typically protective.    NF-kB  activation in microglia and astrocytes is primarily associated with inflammation secondary to pathology and typically has detrimental effects.( Anilkumar & Wright-Jin 2024 (13))

Activation of NF-kB in astrocytes is multi-faceted mirroring the astrocyte function in the CNS.  NF-kB is implicated in astrocyte-dependent clearing of synaptic glutamate, metabolic control and modulation of astrocyte structural plasticity.  In disease, the astrocytic NF-κB signalling is predominantly associated with increased inflammation, and can be associated with adverse outcomes.  ( Anilkumar & Wright-Jin 2024 (13))

Hypoxia-induced NF-κB signalling is a major mediator of microglial inflammation associated with upregulated TLR4 activation.   NF-κB signalling is implicated in both amyloid beta plaques and tau fibrils in late-onset Alzheimer’s disease, each mediated in part by microglial activation. ( Anilkumar & Wright-Jin 2024 (13))

The dysregulation of the inflammatory response in COVID-19 plays a very important role in disease progression. It has been observed that abnormal activity of NF-κB is directly associated with increased production of proinflammatory factors.( Gudowska-Sawczuk & Mroczko. 2022 (14))

The Central Role of CCL2 in Neuroimmune Sensitization and Hypoxia

Chemokines are small signalling molecules, part of the broader cytokine family, that play a crucial role in directing the movement of cells, particularly leukocytes, to areas of inflammation or other biological needs. They act by binding to specific receptors on the surface of cells, triggering signals that lead to cell migration and chemotaxis (movement in response to a chemical stimulus).  

CCL2, also known as monocyte chemoattractant protein-1 (MCP-1), is a key chemokine that orchestrates immune cell trafficking, particularly the recruitment of monocytes, memory T cells, and dendritic cells to sites of tissue inflammation.   Although largely unfamiliar to most clinicians, CCL2 plays a pivotal role in neuroimmune dysregulation, particularly in conditions such as POTS, Long COVID, and ME/CFS where chronic low-grade inflammation persists without overt infection.

CCL2 expression is tightly regulated but becomes markedly upregulated in response to a broad range of innate immune stimuli—most notably through TLR4, RAGE (receptor for advanced glycation end products), IL-1β, TNF-α, and oxidative stress-induced NF-κB activation.

Once engaged, CCL2 establishes a feed-forward loop of inflammation and tissue infiltration, particularly in the central nervous system and autonomic ganglia, where it has been shown to activate microglia, enhance sympathetic tone, and sustain mast cell–T cell–glial crosstalk.

In dysautonomia, especially POTS, elevated CCL2 levels may reflect a persistent "sterile inflammation" phenotype, linking metabolic dysfunction, endothelial injury, and neuroimmune activation. Its overexpression also correlates with impaired baroreceptor sensitivity and brainstem gliosis in experimental models.

Importantly, CCL2 signalling is attenuated by agents such as famotidine (via modulation of histamine and IL-6), N-acetylcysteine, and upstream blockade of TLR4/RAGE pathways, offering emerging therapeutic relevance.

Genetic variations in the CCL2 gene are associated with an increased risk of developing a wide number of  diseases by activating signalling pathways, including JAK/STAT and NF-κB which can influence cell behaviour, survival, and gene expression across different cell types and tissues.

CCL2 has been identified as a mediator that can be released by activated mast cells, and elevated levels of CCL2 have been found in the serum of patients with Mast cell activation syndrome (MCAS).  CCL2 is a potent activator of mast cells, which are implicated in many POTS patients.  Mast cell degranulation releases histamine and other mediators, exacerbating vasodilation and vascular permeability, tachycardia and orthostatic symptoms.

Dysregulation of CCL2 expression has been implicated in the pathogenesis of various health conditions, including Rheumatoid Arthritis (15), IBS (16), fibromyalgia (17), chronic fatigue (18), chronic pain syndromes (15), POTS, connective tissue disease (19), ADHD (20), autism (21)(22). 

The role of CCL2 in the dysregulated pathways are described in Vittone & Exelby 2023 (23)

In POTS, hypoxic brainstem glia likely secrete CCL2 in response to perfusion failure, sustaining baroreflex instability and neuroinflammation. In fibromyalgia, CCL2 expression correlates with pain intensity and microglial activation, while in Long COVID, spike protein–induced TLR4/RAGE activation elevates CCL2 systemically and within the CNS. Moreover, gut barrier dysfunction in all three conditions allows endotoxins and alarmins to circulate and stimulate CCR2⁺ immune cells, further amplifying systemic inflammation and autoimmunity.

Both mould exposure and PTSD converge on the upregulation of CCL2 (MCP-1) through distinct but synergistic pathways involving TLR2/4 and RAGE signalling, NF-κB activation, and glial priming. In mould-related illness, fungal toxins such as ochratoxin A and β-glucans stimulate astrocytes, microglia, and endothelial cells via pattern recognition receptors, leading to robust CCL2 transcription. This chemokine promotes monocyte recruitment, glial activation, and blood–brain barrier permeability, perpetuating neuroinflammation and excitotoxicity. Concurrently, mycotoxin-induced mitochondrial dysfunction and redox stress further amplify this loop via NF-κB and cytokine production, impairing glutamate clearance and glymphatic function.

In PTSD, chronic HPA axis dysregulation, glucocorticoid resistance, and sympathetic overdrive similarly drive CCL2 expression in brain regions such as the amygdala, hippocampus, and brainstem. Persistent microglial priming and astrocyte–microglia cross-talk create a feed-forward loop of neuroinflammatory signalling, particularly under secondary stress or inflammatory triggers.

This shared CCL2-centric mechanism contributes to the central sensitization, cognitive dysfunction, and autonomic dysregulation characteristic of POTS, Long COVID, and functional neurological syndromes. The convergence of mould and trauma thus establishes a neuroimmune milieu that maintains chronic glial activation, impaired detoxification, and hypersensitized central circuits, predisposing patients to prolonged symptom flares and multisystem instability.

Microglia, Astrocytes, and Mast Cells: A Tripartite Axis 

 Mast cells, microglia, and astrocytes play important roles both independently and collectively in mediating responses to sustained mould exposure, stress and PTSD.  Mast cells regulate the functions of immune cells such as dendritic cells, monocytes/macrophages, granulocytes, T cells, B cells and Natural Killer (NK) cells.  

Microglia:

Microglia are a type of neuroglia (glial cell in the CNS that do not produce electrical signals), that account for about 10-15% of cells found within the brain.   Microglia are key cells in overall brain maintenance and constantly monitor neuronal functions.

Microglia scan the tissue and modify their morphology and functions if and when necessary.  They are crucial for the formation, shaping, and functioning of synapses, fundamental for brain development during pre- and post-natal periods. (Steardo et al 2023 (24))

Clough et al 2021 (25)) describe: “Microglia are the resident immune cells of the Central Nervous System (CNS). Microglia have the capacity to migrate, proliferate and phagocytize.  Under physiological conditions, microglia exist in their “resting” state, however on exposure to a pathogen, microglia transition into an activated state and quickly mobilize to the site of injury to initiate an innate immune response.”  As the resident macrophage cells, they act as the first and main form of active immune defence in the CNS.

Damage to the brain triggers a specific type of reactive response mounted by neuroglia cells, in particular by microglia, the most prominent immune cells in the CNS and which are the first to respond to threat.(Phulwani et al. 2008 (2))  Inflammatory microglial activation (IL-6 and TNFa) is the most common brain pathology found in patients who died of COVID-19: 42% are affected, and another 15% have microclots in brain tissue.(Bayat et al. 2022 (26))

This is complicated by astrocyte/ microglial “cross-talk” and neurotransmitter dysregulation.(Figure 4)   The SARS-Co-V spike protein activates microglia leading to pro-inflammatory effects and microglial-mediated synapse elimination.  This microglial activation and neuroinflammation can disrupt the BBB.    COVID also reduces the morphology and distribution of microglia and astrocytes in the hippocampus which has a major role in learning and memory.     Mast cells promote cross-talk between T cells and myeloid cells like microglia during neuroinflammation, and the complex interplay between the activated microglia, reactive astrocytes and mast cells is a key part of the neurological manifestations of the COVID-19 infection.( Theoharides & Kempuraj. 2023.(27))( Bayat et al. 2022 (26))

Figure 4. Mast Cell, Microglia and Astrocyte Cross-Talk

The various histamine receptors have been described on each cell in this diagram, which may lead to a decision on where management may be directed.

Source: Carthy, Elliott & Ellender, Tommas. (2021). Histamine, Neuroinflammation and Neurodevelopment: A Review. Frontiers in Neuroscience. (28)

Astrocytes and impact of mould and PTSD dysregulation

Astrocytes are the most abundant glial cells in the CNS.   They are pivotal in maintaining CNS homeostasis, including neurotransmitter regulation, particularly glutamate.   It is believed that astrocyte reactivity and subsequent glutamate dysregulation contributes to neurological symptoms eg cognitive impairment, fatigue and mood disorders in COVID, very similar to the dysfunction that occurred in the Gulf War Illness. If the brain is not directly damaged, resolution of systemic pathology usually results in restoration of the physiological homeostatic status of neuroglial cells. (Phulwani et al. 2008 (2)) 

Astrocytes provide support for neurons and regulate the blood-brain barrier:

• They become reactive under stress conditions

• Reactive astrocytes release inflammatory mediators

• They can impair blood-brain barrier integrity

• Astrocytes modulate synaptic function and neurotransmission

Blood flow in the brain is regulated by neurons and astrocytes.  Attwell et al. 2010 (29)) describe “It is now recognized that neurotransmitter-mediated signalling has a key role in regulating cerebral blood flow, that much of this control is mediated by astrocytes, that oxygen modulates blood flow regulation, and that blood flow may be controlled by capillaries as well as by arterioles.”   Astrocytes can promote the induction and progression of inflammatory states, which are significantly associated with the disease status or severity.(Zhang et al. 2022 (30))

Astrocytes play a pivotal role in regulating the brain’s glymphatic system, a waste clearance mechanism dependent on paravascular fluid flow. The endfeet of astrocytes—specialized extensions that envelop cerebral blood vessels—form the paravascular spaces through which cerebrospinal fluid (CSF) circulates. These endfeet are densely enriched in aquaporin-4 (AQP4) water channels, which are highly polarized to the endfoot membrane and are essential for the bidirectional exchange between CSF and interstitial fluid (ISF).

AQP4 facilitates the influx of CSF into the brain along arterial paravascular spaces, its mixing with interstitial solutes, and the efflux of waste-laden ISF along venous paravascular routes. This coordinated flow supports the clearance of metabolic by-products, neurotoxins (e.g., amyloid-β, tau, lactate), and inflammatory mediators, while also buffering intracranial pressure by modulating the dynamic balance between CSF and venous compartments.

Additionally, mast cell–induced collagenase release and fascia remodelling in the upper cervical region may mechanically impair venous and lymphatic drainage, especially at C1 and the thoracic inlet. This promotes accumulation of glutamate, lactate, and other mast cell–activating metabolites in the ECM, aggravating excitotoxic signalling and perpetuating PEM. MLD techniques that release fascial constriction at these choke points have shown rapid resolution of interstitial oedema and symptom flares, supporting a combined structural–biochemical hypothesis.

SARS-CoV-2 preferentially infects and replicates and propagates in astrocytes, particularly those adjacent to infected vasculature. In contrast, neurons and microglia are less likely to be directly infected. Importantly, while microglia and astrocytes are both reactivated, a direct dosage-sensitive effect of SARS-CoV-2 is only observed in reactive astrocytes.  Astrocytes are the primary targets of SARS-CoV-2 in the brain.    SARS-Co-V preferentially infects astrocytes over neurons resulting in astrocyte reactivation and neuronal death. (Bayat et al. 2022 (26))   Disruption of AQP4 localization or expression—due to astrocytic dysfunction—impairs glymphatic clearance, contributing to cerebral oedema, toxin accumulation, and intracranial hypertension. 

Astrocytic endfeet are also immune-metabolic sensors, susceptible to inflammatory stimuli. TLR2 and RAGE activation, commonly triggered by stress, infection, or mycotoxins, impairs AQP4 function and redistributes it away from the endfeet, thereby collapsing the structural integrity of the glymphatic system.

This is exacerbated in conditions such as Long COVID, where SARS-CoV-2 membrane proteins activate TLR2 on astrocytes, inducing pro-inflammatory cytokine release and mitochondrial dysfunction. In parallel, RAGE engagement by DAMPs (e.g., HMGB1, S100B) drives NF-κB–mediated inflammatory amplification, reducing glutamate clearance and contributing to excitotoxicity, neurovascular uncoupling, and raised intracranial pressure.

Thus, the loss of AQP4 polarization and endfoot integrity represents a critical nexus between immune activation, impaired fluid homeostasis, and neuroinflammatory persistence. This process underlies many of the symptoms observed in neuroimmune conditions such as POTS, fibromyalgia, and Long COVID, where impaired glymphatic flow contributes to brain fog, pressure headaches, fatigue, and autonomic dysregulation.

Both mould exposure and PTSD serve as upstream disruptors of this system. Mycotoxins such as ochratoxin A activate astrocytic TLR2 and RAGE, driving ROS generation, mitochondrial injury, and AQP4 mis-localization. Chronic glial activation and inflammatory pressure impair glymphatic flow, leading to accumulation of neurotoxic metabolites and increased intracranial pressure.

PTSD, through HPA axis dysregulation, glucocorticoid resistance, and central microglial priming, similarly disrupts AQP4 expression and astrocytic homeostasis. These insults converge to impair fluid dynamics, neurovascular coupling, and interstitial solute clearance—worsening symptoms and facilitating a state of sustained neuroimmune sensitization.

This shared pathway further reinforces the positioning of hypoxia, glial activation, and impaired drainage as central mechanisms in chronic neuroimmune disorders.

Mast cells

The mast cell is a potent immune cell known for its functions in host defence responses and diseases, such as asthma and allergies.  “Mast cells play a key role in homeostatic mechanisms and surveillance, recognizing and responding to different pathogens, and tissue injury.   An abundance of mast cells reside in connective tissue that borders with the external world (the skin as well as gastrointestinal, respiratory, and urogenital tracts.) (van der Donk et al 2023 (31))

Mast cells are located perivascularly close to nerve endings and ANS sites eg carotid bodies and the adrenals, allowing them to potentially regulate and be affected by autonomic function.  They can be triggered not only by allergens but also by triggers from the ANS, releasing neuro-sensitizing, pro-inflammatory and vasoactive mediators.

Mast cells regulate the functions of immune cells such as dendritic cells, monocytes/macrophages, granulocytes, T cells, B cells and Natural Killer (NK) cells.   They recruit immune cells to inflamed tissue by secreting chemokines and other mediators which locally increase vascular permeability.

Mast cells are activated by cytokines from TLR4.  They contribute to coronavirus-induced inflammation through mechanisms like degranulation and histamine release.  Mast cell mediators can disrupt connective tissue integrity.

Histamine and tryptase can degrade the extracellular matrix and disrupt the integrity of connective tissue.   Proteases eg  chymase can inhibit collagen synthesis by smooth muscle cells, weakening the connective tissue structure, and mast cell-derived mediators like TNFa can induce apoptosis of smooth muscle cells, further compromising the connective tissue.  Prostaglandins and leukotrienes contribute to inflammation and pain.(Viullapol et al 2015. (32))( Krystel-Whittemore et al. 2016 (33)) Clinic observations have demonstrated collagen changes occurring after COVID infections.

Mast cells are increasingly seen as important in the communication between peripheral nerve endings and cells of the immune system. Alim et al 2021 (34)  confirmed the binding of glutamate to glutamate receptors on the mast cell surface.    Further, glutamate had extensive effects on gene expression in the mast cells, including the upregulation of pro-inflammatory components such as IL-6 and CCL2.

Dong et al. 2017 (35) demonstrated that brain inflammation plays a critical role in the pathophysiology of brain diseases.  They demonstrated that in the brain, activation of mast cells triggers activation of microglia, increased cytokine and TLR4 expression, whereas stabilisation of mast cells with disodium cromoglycate (Cromolyn) inhibited the CNS inflammation that would otherwise result from activation of microglia.

Malone et al. 2021 (36) describe mast cell-derived histamine exerting its biological actions through four types of histamine  receptors (i.e., H1 receptor, H2 receptor, H3 receptor, and H4 receptor).   It also activates  acute immune-mediated reactions and enhances vascular smooth muscle contraction and the migration of other immune cells, antibodies, and mediators to the site of insult.  The release of histamine by perivascular mast cells may also affect adjacent lymphatic vessel function inducing immune cell trafficking through its lumen,which potentially contribute to acute inflammatory stimulus. 

Afrin, Weinstock & Molderings 2020.(37) describes “Fatigue and malaise are the most common complaints in MCAS.    Most patients remain functional, but some are severely impaired.   Low-grade temperature dysregulation is not uncommon, as are lymph node swelling, weight loss, unexplained weight gain, loss of appetite, fluctuating oedema, but it is the gain in adipose tissue that accounts for weight increase in most MCAS.   These patients may have bariatric surgery sometimes with complications of poor wound healing, and while there is initial weight loss, the other symptoms usually remain, and the weight gain slowly starts to return.  Mast cells are programmed to site themselves at environmental interfaces- lungs, gut, skin, bladder, nose and sinuses etc, so there can be a wide range of pathology in aberrant mast cell activation.”  

“Mast cell activation syndrome is known to permanently escalate its baseline level of dysfunction of the affected mast cells shortly after a major stressor, likely due to complex interactions between epigenetic abnormalities and the stressor’s induced cytokine storm- of additional mutations by the mutated stem cells from which the mutated /dysfunctional mast cells are derived.”( Afrin, Weinstock & Molderings 2020.(37))

The interplay between chronic stress, PTSD, mould exposure, and mast cell dysfunction creates a complex cycle that can significantly impact overall health and immune function. Addressing the root causes and supporting mast cell stabilization may be key to improving symptoms and immune regulation in affected individuals.

Mast cells are often the initiators of this neuroimmune storm. Activated by mould toxins, psychological stress, and PAMPs/DAMPs, they release tryptase, histamine, leukotrienes, and prostaglandins. These mediators not only increase vascular permeability but also activate nearby glial cells. Notably, glutamate receptors on mast cells facilitate pro-inflammatory gene expression.

Mast cells act as "first responders" in stress conditions, rapidly degranulating and releasing pre-formed mediators.    Chronic stress and PTSD can dysregulate mast cells by increased mast cell activation and degranulation due to elevated stress hormones like cortisol and noradrenalin, with enhanced production of pro-inflammatory mediators by mast cells, including histamine, cytokines, and chemokines, playing a crucial role in both peripheral inflammation and neuroinflammation in PTSD.

Chronic stress and PTSD can lead to dysregulation of mast cells in several ways:

• Increased mast cell activation and degranulation due to elevated stress hormones like cortisol and noradrenalin, with enhanced production of pro-inflammatory mediators by mast cells, including histamine, cytokines, and chemokines.

• Lowered activation threshold of mast cells, making them more sensitive to triggers.

• Increased numbers of mast cells in stress-responsive areas of the brain and body.

Mould can impact on mast cells through:

• Direct activation of mast cells by mould spores and mycotoxins with increased production of inflammatory mediators like histamine and leukotrienes.

• Sensitization of mast cells, leading to heightened reactivity to other triggers.

The combined effects on mast cells from chronic stress, PTSD, and mould exposure can lead to broader immune system dysfunction with dysregulation of T-cell responses and increased autoimmune reactivity, manifest as Mast Cell Activation Syndrome (MCAS), chronic fatigue, autoimmune disorder, multiple chemical sensitivities, GI tract dysfunction with compromised barrier function in the gut, lungs, and brain  and cognitive impairment:

Neuro-Metabolic Dysfunction from Mould and PTSD

Astrocyte–glutamate dysfunction has been implicated in Gulf War Illness, where exposed service personnel developed a range of neurocognitive and systemic symptoms, likely triggered by toxicant-induced neuroinflammation. Emerging data from Long COVID—notably the work of  Hotowitz,Guedj et al.2023 (38) suggests a convergent mechanism involving astroglial activation, glutamate excitotoxicity, and impaired neuro-energetic coupling.

Astrocytic dysfunction impairs glymphatic clearance, contributing to both fatigue and intracranial hypertension, by reducing the brain’s ability to eliminate metabolic waste and modulate cerebrospinal fluid dynamics.

Elevated extracellular glutamate and reduced GABA buffering, secondary to astrocytic EAAT2 downregulation and altered glutamate-glutamine cycling, may underlie excitatory–inhibitory imbalance seen across a range of neuropsychiatric and neurological conditions.

Both mould exposure and PTSD are potent upstream drivers of this astroglial dysfunction. Mycotoxins induce oxidative stress, inhibit mitochondrial enzymes (e.g., PDH, MDH2), and downregulate glutamate transporters (EAAT1/2), impairing glutamate clearance and promoting neuroexcitotoxicity. These toxins also activate astrocytic TLR2, TLR4, and RAGE receptors, shifting astrocytes to a proinflammatory, neurotoxic phenotype (A1), and promoting feed-forward neuroinflammation via IL-1β and CCL2 signalling.

Similarly, PTSD induces chronic astrocyte–microglial priming through sustained HPA axis dysregulation, catecholaminergic sensitization, and TLR4/NLRP3 inflammasome activation. This results in reduced GABAergic tone, elevated glutamate, and suppression of astrocyte-derived metabolic support (e.g., lactate, glutamine, and ethanolamine). Both mould and PTSD also compromise glymphatic flow through altered aquaporin-4 channel regulation, worsening interstitial fluid stasis and toxin accumulation.

Together, these mechanisms underlie the neuroinflammatory and excitatory dysregulation seen in conditions such as fibromyalgia, ADHD, autism spectrum disorder, migraine, visual snow, Alzheimer's disease, and Parkinson’s disease—each sharing a core signature of astrocytic impairment, excitotoxicity, and disrupted synaptic homeostasis.

Figure 5.  Proposed mechanism of ME/CFS linked to amino acid catabolism.

Category I: amino acids are converted to pyruvate, and therefore depend on PDH to be further oxidized. These are alanine (Ala), cysteine (Cys), glycine (Gly), serine (Ser), and threonine (Thr).

Category II: amino acids that enter the oxidation pathway as acetyl-CoA, which directly and independently of PDH fuels the TCA cycle for degradation to CO2. These are isoleucine (Ile), leucine

(Leu), lysine (Lys), phenylalanine (Phe), tryptophan (Trp), and tyrosine (Tyr).

Category III consists of amino acids that are converted to TCA cycle intermediates, thereby replenishing and supporting the metabolic capacity of the cycle- histidine (His), and proline (Pro)

The asterisks indicate the amino acids significantly reduced in ME/CFS patients.

 Source: Fluge et al., Metabolic profiling indicates impaired pyruvate dehydrogenase function in myalgic encephalopathy/chronic fatigue syndrome. JCI Insight. 2016 (39)

Clinic amino acid profiling in Long COVID and POTS (irrespective of activator), based on research by Fluge et al 2016,(39) has shown significant metabolic dysfunction in 3 main pathways, the PDH, lactate and malate/aspartate shuttles. 

PEM, ECM Sequestration, and Lymphatic Clearance: A Missing Link in Chronic Sensitization Syndromes

Clinical observation now supports that post-exertional malaise (PEM)—a hallmark symptom of ME/CFS, POTS, and Long COVID—may be mediated not purely by internal mitochondrial distress, but also by extracellular accumulation of hypoxic and inflammatory end-products, including lactate, glutamate, oxidized phospholipids, and bradykinin. These accumulate in the extracellular matrix (ECM) under conditions of impaired glymphatic and lymphatic flow, particularly in regions with venous congestion or fascial compression (e.g., C1, thoracic outlet).

Recent case series employing manual lymphatic drainage (MLD) have demonstrated immediate resolution of PEM and interscapular oedema, strongly implicating ECM stasis as a reversible contributor to symptom persistence. This suggests that the ECM may function as a transient “toxic reservoir”, wherein delayed clearance of neuroactive metabolites perpetuates mast cell activation, RAGE signalling, and glial sensitization.

Aspartate and GABA, which are consistently suppressed in urinary amino acid profiles of patients with PEM, may normalize following effective ECM decompression- this study is underway. These amino acids are involved in mitochondrial redox cycling (malate-aspartate shuttle), excitatory–inhibitory balance, and metabolic detoxification, underscoring the metabolic and immunological implications of mechanical lymphatic obstruction.

These findings shift the model of PEM from one of fixed mitochondrial injury to one of dynamic neuroimmune–mechanical dysfunction, wherein interstitial clearance is a key therapeutic target.

GABA Dysfunction

GABA is normally reduced in clinic testing of urinary amino acids,.   The GABAergic system, particularly GABA-A receptor function and GABA synthesis, is highly vulnerable to disruption by both mould exposure and PTSD, and the downstream effects of this dysfunction have significant implications for central sensitization, autonomic dysregulation, and cognitive impairment—all common in POTS, CFS, and Long COVID.

Aspartate-Malate Shuttle (AMS) Dysfunction

This AMS mechanism sits at the intersection of mitochondrial metabolism, glial–neuronal signalling, and neuroimmune injury. The AMS is critical for mitochondrial NADH import, maintaining redox balance and oxidative phosphorylation.   This facilitates ATP production, GABA/glutamate balance, and urea/ammonia detoxification via the malate–aspartate cycle. 

Impaired aspartate function impacts on clearance of  hypoxic end products from mitochondria and is evident as post-exertional malaise (PEM,) aspartate/glutamate imbalance, BBB integrity, cognitive dysfunction, and functional neurological disorder (FND). 

Mould mycotoxins such as ochratoxin A:

• Inhibit NAD⁺ regeneration → suppress MDH2 and GOT2 activity.

• Downregulate AGC1/2 via redox-sensitive transcriptional repression

• Promote astrocyte reactivity, reducing aspartate export to neurons and increasing extracellular glutamate via EAAT dysfunction.

Chronic stress and PTSD downregulates SLC25A12, impeding aspartate–glutamate exchange. Elevated cortisol and noradrenaline shift metabolism toward glycolysis leading to an accumulation of cytosolic NADH and stalling of the shuttle.   IL-1β, TNF-α, and CRH disrupt mitochondrial enzymes and suppress GOT2 expression.

Aspartate deficiency leads to a failure to replenish TCA intermediates post-exertion causing post-exertional malaise (PEM).  It also reduces GABA synthesis (via failure to convert glutamate), amplifying excitotoxicity.

Glutamate Accumulation and Excitotoxicity- An impaired aspartate-malate shuttle stalls glutamate–aspartate exchange which leads to intracellular glutamate buildup.   Astrocytes become ineffective buffers of extracellular glutamate causing excitotoxicity.   GABA synthesis is reduced due to glutamate misrouting.   EAAT2 downregulation (in mould and stress) exacerbates this.

Excess glutamate disrupts tight junction proteins (occludin, claudins) via NMDA receptor-mediated calcium influx in endothelial cells disrupting the blood-brain barrier (BBB.) Glutamate also activates microglia and mast cells at the BBB causing a further breakdown via MMPs and ROS.   The results is paracellular leak, cytokine influx, DAMP/PAMP penetration.

AMS, Cognitive Dysfunction and Functional Neurological Disorders (FND)

A hypofunctional AMS leads to metabolic underdrive in high-demand circuits (e.g., prefrontal cortex, limbic structures).

Sustained glutamate toxicity leads to:

• Impaired working memory, executive function, verbal fluency.

• Cortical–subcortical dissociation, a proposed mechanism for FND.

• Inability to integrate sensory–motor signals under stress → motor/sensory conversion symptoms.

Lactate Shuttle Dysfunction

A dysfunctional lactate shuttle—particularly astrocyte–neuron lactate transport—is strongly implicated in cognitive impairment, and this mechanism appears highly relevant to conditions like CFS, POTS and Long COVID.   When this shuttle is impaired , neurons are deprived of a critical energy substrate during activation.  They suffer energy crisis, redox imbalance, and increased oxidative stress, reduced mitochondrial membrane potential and synaptic efficiency.

Both mould exposure and PTSD are potent disruptors of astrocyte–neuron metabolic coupling, and they converge via inflammatory, oxidative, and neuroimmune sensitization pathways that critically impair the lactate shuttle and cerebral energy homeostasis

Linking COVID, Mould, PTSD, and Chronic Stress through TLR4 and Mast Cell Pathophysiology

There is increasing recognition that COVID-19, mould exposure, PTSD, and chronic psychological stress share a common mechanistic thread: innate immune activation via Toll-like receptor 4 (TLR4) and mast cell dysregulation.

TLR4 serves as a central pattern recognition receptor that bridges environmental and psychological stressors with immune and neuroinflammatory responses. Whether activated by viral PAMPs (such as the SARS-CoV-2 spike protein), fungal DAMPs and mycotoxins, or stress-induced alarmins (e.g., HMGB1, heat shock proteins), chronic TLR4 signalling promotes a persistent pro-inflammatory state characterized by NF-κB activation, cytokine release, and glial sensitization.

When unresolved, this leads to immune exhaustion, loss of peripheral tolerance, and breakdown of self-recognition pathways—laying the groundwork for autoimmune activation.

In COVID-19, dysfunctional TLR4 activation contributes to the cytokine storm, endothelial damage, and hypercoagulability observed in severe cases. The SARS-CoV-2 spike protein has been shown to act as a potent TLR4 agonist, amplifying IL-6, TNF-α, and CCL2 production, even independent of viral replication.

Similarly, in PTSD and chronic stress, TLR4 is activated by endogenous DAMPs released during tissue damage or repeated HPA axis dysregulation, reinforcing a cycle of neuroinflammation, glucocorticoid resistance, and autonomic instability.

Mould exposure activates TLR4 via β-glucans, mannan, and mycotoxins, triggering mast cell degranulation, astrocyte activation, and the RAGE–TLR4–NF-κB axis, which sustains inflammation even after the original insult resolves.

Importantly, mast cells—strategically positioned across the blood–brain barrier, vasculature, and gut—respond to these diverse insults with rapid degranulation and mediator release, including histamine, leukotrienes, and tryptase, which in turn activate microglia and perpetuate glial–mast cell cross-talk (Figure 4).

The convergence of these insults on TLR4 and mast cell pathways results in shared symptomatology: cognitive dysfunction, fatigue, dysautonomia, post-exertional malaise, and in many cases, autoimmune comorbidity (e.g., Hashimoto’s thyroiditis, lupus, or Sjögren’s syndrome). Thus, TLR4 and mast cell dysregulation form a final common pathway through which diverse stressors—infectious, environmental, or psychological—can produce a similar neuroimmune phenotype, particularly in genetically or epigenetically primed individuals.

Chronic Stress, Immune Dysregulation, and Neuroinflammation

Chronic psychological stress exerts profound effects on the hypothalamic–pituitary–adrenal (HPA) axis, the autonomic nervous system, and immune signalling networks, shifting the body into a sustained sympathoadrenal–inflammatory state.  While acute stress may transiently enhance immune responsiveness, prolonged or repeated stress diminishes immune competence, induces cortisol resistance, and leads to sympathetic overdrive.

This altered internal milieu upregulates RAGE, disrupts TLR9-mediated anti-inflammatory control, and promotes feed-forward inflammatory cascades via TLR2, TLR4, and NF-κB activation. The resulting immune dysregulation facilitates the persistence of latent infections (e.g., EBV, HSV), disrupts mitochondrial redox balance, and primes both central and peripheral inflammatory loops.

Central to this pathology is the interplay between mast cells, microglia, and astrocytes. Mast cells, as rapid responders to internal and external stressors, degranulate within minutes, releasing histamine, tryptase, prostaglandins, and cytokines that recruit and activate glial cells.

Hendriksen et al 2015 (40) described that under chronic stress, microglia adopt a pro-inflammatory phenotype, producing IL-1β and TNF-α, while astrocytes lose regulatory control of glutamate homeostasis, contributing to excitotoxicity, BBB dysfunction, and impaired glymphatic clearance.

These interactions are further amplified by aging and repetitive microtrauma, setting the stage for central sensitization, cognitive impairment, and metabolic dysfunction. The glutamatergic system, in particular, becomes destabilized under astrocytic dysfunction, contributing to the neurocognitive and fatigue-related symptoms observed in stress-related syndromes- (Morey et al 2015 (41))

Chronic stress also modulates the expression and function of Toll-like receptors (TLRs), especially TLR4, which is closely linked to stress-induced neuroinflammation. TLR3, 7, and 9 are implicated in autoimmunity and viral reactivation, while TLR2 has been associated with EBV/HSV reactivation and bacterial pattern recognition.

Importantly, the interaction between stress and TLR signalling is bi-directional—stress modulates TLR expression, while TLR activation feeds back to dysregulate the HPA axis and increase sympathetic tone.

This closed-loop interaction contributes to the immune–neuroendocrine dysregulation seen in POTS, fibromyalgia, and chronic fatigue syndrome. Furthermore, mast cell–endothelial interactions increase vascular permeability, contributing to orthostatic intolerance, fluid shifts, and potential collagen dysfunction. This mechanistic overlap strongly supports the emerging model of chronic stress and PTSD as priming events in multi-system inflammatory sensitization, in which neuroimmune loops become progressively harder to extinguish.

PTSD, Immune Sensitization, and Neuroinflammatory Convergence

Post-traumatic stress disorder (PTSD) is increasingly recognized not only as a psychological disorder but also as a neuroimmune–metabolic condition with delayed, progressive, and systemic manifestations. Many individuals who initially cope with trauma later develop symptoms months or even years afterward—delayed-onset PTSD. 

Subclinical dysregulation often precedes overt symptoms, driven by progressive sensitization to environmental stressors, including physical trauma, infection, mould exposure, or dietary inflammatory triggers. This delayed evolution challenges older paradigms of PTSD as an immediate response to trauma and aligns with models involving accumulated allostatic load, in which chronic stress and repeated sympathetic activation disrupt HPA axis regulation, immune surveillance, and metabolic homeostasis.(Cohen et al. 2011 (42))

Neuroimmune mechanisms underlie much of PTSD’s chronicity. Trauma primes mast cells, microglia, and astrocytes, initiating a feed-forward inflammatory loop that involves TLR activation (notably TLR2, TLR4, and TLR9), NF-κB transcriptional signalling, and cytokine release (e.g., IL-1β, TNF-α, IL-6).

Mast cells, as peripheral and central “first responders,” rapidly degranulate in response to perceived threats, releasing histamine, tryptase, prostaglandins, and neuropeptides. This in turn activates adjacent microglia and astrocytes, resulting in sustained neuroinflammation, BBB disruption, and glutamate/GABA imbalance.

Astrocytic dysfunction—particularly impaired glutamate clearance via EAAT2 and altered lactate shuttling—contributes to excitotoxicity, cognitive decline, and fatigue. These glial–mast cell interactions are central to the pathophysiology of PTSD and intersect with key symptom domains observed in fibromyalgia, chronic fatigue syndrome, irritable bowel syndrome, and functional neurological disorders.

Moreover, PTSD exhibits profound metabolic consequences. Chronic HPA dysregulation and sympathetic hyperactivation contribute to hypertension, hyperlipidaemia, obesity, and coronary artery disease, with overlapping pathophysiological links to mast cell activation and immune–glucocorticoid interactions. Lowered cortisol availability impairs NF-κB suppression, exacerbating systemic inflammation. TLR9, while protective in limiting neuroinflammatory overshoot, is often dysfunctional in PTSD, leading to unrestrained IL-1β and volumetric changes in stress-sensitive brain regions like the hippocampus and amygdala.

These same circuits are implicated in fear reconsolidation, a process tightly linked to NF-κB activation in the basolateral amygdala. This supports a model wherein repeated trauma exposure, immune sensitization, and glial priming culminate in central neuroimmune dysregulation and multisystem involvement.  Sensitization is a critical process in the onset of pain syndromes and PTSD.  The central role of the amygdalae in the kindling in PTSD is similar to the phenomena of windup of C fibre evoked pain in fibromyalgia, IBS and chronic fatigue.   Furthermore this has been associated with modified autonomic function (dysautonomia).

The recognition of PTSD as a shared axis disorder—overlapping with fibromyalgia, POTS, and chronic fatigue—underscores the need for integrative treatment models. Therapeutic approaches targeting mast cell–glial interactions, TLR signalling, or NF-κB modulation may hold promise, but require deeper translational investigation. Ultimately, PTSD represents not just a psychiatric condition but a systemic sensitization syndrome, where past trauma creates a vulnerable physiological terrain prone to flare with new triggers and cumulative environmental load.(Zimmerman et al. 2012.(9))(Katrinli et al 2022 (43))

Mould-Induced Immune and Metabolic Dysregulation

Mast cells, microglia, and astrocytes play crucial roles in mediating central nervous system responses to sustained mould exposure. These cell types operate both independently and in a tightly integrated network, where chronic immune activation by fungal antigens and mycotoxins leads to persistent neuroinflammation, oxidative stress, and metabolic disruption. Prolonged mould exposure has been shown to result in:

• Persistent neuroinflammation and glial priming (Harding et al 2019) (46))(Kraft et al. 2021)(47))

• Blood–brain barrier (BBB) permeability and paracellular leak

• Neuronal injury and excitotoxicity

• Cognitive and behavioural impairments, including memory loss, mood alterations, and executive dysfunction (Harcha et al 2021 (48))(Ehsanifar et al 2023 (49))

Experimental models show that mould exposure drives hippocampal immune activation and inhibits adult neurogenesis, resulting in impaired memory consolidation, learning, and emotional regulation. Additionally, mould exposure modulates pain perception and anxiety-like behaviour, consistent with central sensitization syndromes.

Mould and Mast Cell Activation

Mould has a dual capacity to both directly activate and sensitize mast cells, potentiating broad neuroimmune dysregulation. This occurs through:

• Direct activation of mast cells by mycotoxins and fungal β-glucans, leading to the release of histamine, leukotrienes, prostaglandins, and tryptase

• Lowering of activation thresholds, whereby chronic low-level mould exposure leads to mast cell sensitization, increasing reactivity to unrelated triggers.

This mast cell activation intersects with glial signalling, as histamine and cytokine release further activate astrocytes and microglia, feeding into a chronic neuroinflammatory loop. Mast cell–glial–vascular crosstalk contributes to BBB disruption, central sensitization, and limbic hyperexcitability, hallmarks of complex multisystem illnesses like POTS, fibromyalgia, and Long COVID.

Mould and TLR, RAGE, and NF-κB Pathway Activation

Mould is a potent trigger of innate immune activation via pattern recognition receptors (PRRs), particularly Toll-like receptors (TLR2, TLR4, and TLR6) and Receptor for Advanced Glycation End Products (RAGE). These receptors recognize pathogen-associated molecular patterns (PAMPs) and damage-associated molecular patterns (DAMPs) present in fungal spores, mycotoxins, and cell wall components (e.g., β-glucans, mannan). This activates a canonical MyD88–NF-κB signalling cascade, culminating in the transcription of pro-inflammatory mediators such as:

• IL-6

• TNF-α

• CCL2 (MCP-1)

• IL-1β

These mediators not only sustain peripheral inflammation but also recruit immune cells into the CNS, amplifying glial activation and altering synaptic plasticity and metabolic tone.

Mitochondrial Dysfunction, Oxidative Stress, and the Cell Danger Response

Mycotoxins such as ochratoxin A and trichothecenes directly impair mitochondrial function, inhibiting complexes I and III, depleting NAD⁺ and ATP, and generating reactive oxygen species (ROS). These stressors induce:

1. Release of mitochondrial DAMPs (e.g., mtDNA, cardiolipin) which amplify innate immune signalling.

2. Astrocyte dysfunction, compromising lactate production, glutamate clearance, and aquaporin-4-regulated glymphatic flow.

3. Activation of the Cell Danger Response (CDR)—a hypometabolic protective state—resulting in suppressed mitochondrial respiration, sustained glycolysis, and altered membrane signalling.

These disturbances create a self-sustaining inflammatory and metabolic loop, which underpins chronic fatigue, post-exertional malaise (PEM), neurocognitive dysfunction, and autonomic dysregulation in mould-sensitive individuals.

Autoimmune Activation Pathways in Stress, Mould, and PTSD

Chronic exposure to environmental and psychological stressors—including mould-derived mycotoxins, traumatic experiences, and prolonged psychosocial stress—can converge on shared innate immune pathways that contribute to the loss of immune tolerance and activation of autoimmunity, particularly in susceptible individuals.

The central axis in this convergence is persistent activation of Toll-like receptor 4 (TLR4), which serves as a sentinel receptor for both exogenous PAMPs (e.g., β-glucans, lipopolysaccharide, viral proteins) and endogenous DAMPs (e.g., HMGB1, mitochondrial DNA, oxidized phospholipids).

1. Sustained TLR4 activation by stress and mould exposure promotes chronic inflammation:

   • In mould-sensitive individuals, exposure to fungal components such as β-glucans and mycotoxins activates TLR4 on mast cells, astrocytes, and microglia, initiating an NF-κB-driven pro-inflammatory program.

   • In PTSD and chronic stress, cellular stress signals (e.g., ATP, heat shock proteins, ROS) act as DAMPs to activate the same receptors.

   • This results in a persistent cytokine environment, rich in IL-1β, IL-6, TNF-α, and CCL2, which impairs the resolution phase of inflammation and disrupts regulatory T cell (Treg) function.

2. Loss of immune tolerance through molecular mimicry, bystander activation, and impaired antigen clearance:

   • Chronic glial activation and BBB dysfunction increase CNS antigen exposure to peripheral immune cells.

   • Repeated mast cell–microglial activation creates an inflammatory milieu that favours MHC-II upregulation and epitope spreading, increasing the likelihood of autoreactive T and B cell recruitment.

   • Dysregulated antigen presentation under inflammatory stress promotes neoantigen formation, particularly via oxidatively modified self-proteins, which may trigger autoantibody production.

3. Mast cell–glial feedback amplifies neuroimmune dysfunction in autoimmune-prone tissues:

   • Mast cells, via histamine, tryptase, and IL-6 release, activate microglia and astrocytes, which in turn produce CCL2 and glutamate.

   • This feedback loop promotes central sensitization and contributes to the breakdown of immune privilege in the CNS.

   • In conditions such as Hashimoto’s thyroiditis, lupus, Sjögren’s syndrome, and MS-like syndromes, this mechanism may underpin early symptoms of fatigue, brain fog, or neuropathic pain, even prior to classic seroconversion.

4. Interaction with HPA axis dysfunction and viral reactivation:

   • Chronic stress impairs cortisol-mediated immunoregulation, reducing the suppression of Th17 and Th1 cytokines.

   • This enhances susceptibility to autoimmune flares and enables viral reactivation (e.g., EBV, HSV), which can act as a cofactor in bystander activation of autoreactive lymphocytes.

   • TLR4 and TLR2 also facilitate viral reactivation, contributing to cytokine escalation and further breaking of immune tolerance.

This autoimmune priming mechanism helps to explain why individuals with combined mould exposure, trauma histories, and persistent stress are disproportionately represented among patients with dysautonomia, fibromyalgia, autoimmune thyroiditis, and non-specific autoimmune serologies (e.g., ANA+, SSA+, low complement). The TLR4–mast cell–glial axis, when chronically activated, creates a permissive environment for autoimmunity, particularly in genetically predisposed individuals with impaired TLR9 or IL-10 regulatory circuits.

Neuroimmune Sensitization in Chronic Conditions

These chronic conditions—POTS, fibromyalgia, and Long COVID—are increasingly understood as manifestations of a shared neuroimmune sensitization state, driven by converging insults to vascular, glial, and immune homeostasis. A unifying principle across all is the presence of regional hypoxia, particularly affecting the brainstem, limbic system, and peripheral autonomic network.

In POTS, impaired venous return and cerebral hypoperfusion initiate brainstem glial activation, mast cell degranulation, and baroreflex instability. In fibromyalgia, hypoperfusion and microvascular dysfunction contribute to astrocyte–glutamate dysregulation and C-fibre wind-up, while in Long COVID, endotheliitis, microthrombi, and viral persistence amplify hypoxic signalling across the neurovascular unit.

A key amplifier in this shared pathway is CCL2 (monocyte chemoattractant protein-1), which is robustly induced by hypoxia, oxidative stress, and activation of TLR4 and RAGE—common features of all three conditions. CCL2 promotes recruitment of CCR2⁺ monocytes, mast cells, and microglial precursors into the CNS, particularly within hypoxic and inflamed regions such as the brainstem and meninges. This chemokine not only facilitates perivascular immune infiltration but sustains glial reactivity, mast cell–microglia cross-talk, and barrier dysfunction, thereby driving chronic neuroinflammation and sensitization. Elevated CCL2 also contributes to immune dysregulation and autoimmune propagation, particularly in individuals with impaired regulatory T cell activity or ongoing viral reactivation.

Hypoxia acts as both a trigger and a perpetuator of this cascade—upregulating HIF-1α, NF-κB, and downstream inflammatory mediators, while simultaneously impairing mitochondrial metabolism, lactate clearance, and redox cycling. This results in bioenergetic failure, post-exertional malaise, and exacerbation of neurovascular and autonomic instability. CCL2 is therefore positioned not as a secondary byproduct, but as a central node in the feed-forward loop linking vascular insufficiency, immune cell recruitment, glial sensitization, and systemic autoimmunity.

By placing hypoxia and CCL2-driven immune amplification at the heart of these conditions, we can better understand their overlapping symptoms, fluctuating severity, and shared therapeutic targets—pointing toward interventions that modulate perfusion, innate immune signalling, and neuroglial homeostasis.

PTSD, Mould, Chronic Stress, and Gut–Brain Axis Dysfunction

PTSD, chronic psychological stress, and mould exposure compromise the intestinal barrier, facilitating bacterial translocation and converge on a shared pathophysiological axis that disrupts the gut–brain barrier, initiates TLR4-mediated systemic inflammation, and destabilizes immune tolerance.(11)(18)

Mycotoxins, particularly trichothecenes and ochratoxin A, compromise intestinal epithelial tight junctions, resulting in increased intestinal permeability or "leaky gut." This allows bacterial translocation and the entry of lipopolysaccharide (LPS) and other endotoxins into systemic circulation, triggering TLR4 activation on monocytes, dendritic cells, and mast cells. The resulting inflammatory cascade primes the adaptive immune system, lowers tolerance thresholds, and facilitates autoantibody production, contributing to systemic and neuroimmune sensitization.

This pathway mirrors that seen in COVID-19, where gut epithelial disruption and microbial dysbiosis promote increased mucosal CD4⁺/CD8⁺ T cell activation, neutrophil infiltration, and a decline in regulatory T cell (Treg) populations. These changes collectively drive a hyperinflammatory milieu that potentiates autoimmunity, particularly in genetically predisposed individuals. (Eleftheriotis et al 2023 (50))(Sun et al 2022 (51))

In parallel, chronic stress and PTSD alter vagal tone and sympathetic output, impairing mucosal immune surveillance and increasing intestinal permeability via glucocorticoid resistance and sympathetic overdrive. The gut microbiota itself is disrupted, with loss of commensals and overgrowth of opportunistic pathobionts, leading to dysbiosis, diminished production of short-chain fatty acids, and reduced capacity to neutralize or bind mycotoxins.

Compounding this, both stress and mould exposure promote the upregulation of RAGE under conditions of oxidative stress. RAGE ligation by HMGB1, AGEs, and serum amyloid A (SAA), all elevated in post-infectious and toxic-inflammatory states, reinforces NF-κB activation through a positive feedback loop. RAGE–TLR4 crosstalk leads to NLRP3 inflammasome activation, with subsequent release of IL-1β and IL-18 and induction of pyroptotic cell death in enterocytes and glia.

This feed-forward inflammatory state extends from the gut mucosa to the central nervous system via both humoral and vagal pathways, contributing to central sensitization, neuroinflammation, and autonomic dysfunction. The result is a destabilized gut–brain axis, in which immune, microbial, and neural signals perpetuate chronic symptoms across the gastrointestinal, immune, and central nervous systems.

Conclusion:

The shared immunopathology underpinning mould-related illness, PTSD, and Long COVID centres on chronic activation of the TLR4–RAGE–NF-κB axis, amplified by mast cell–glial crosstalk and regional hypoxia. These conditions converge on a final common pathway of neuroimmune sensitization, marked by glymphatic failure, excitotoxicity, and sustained inflammation.

Central to this model is the role of CCL2, which amplifies immune cell recruitment and reinforces barrier breakdown and autoimmunity. Understanding these shared mechanisms provides a unified framework for diagnosis and treatment—emphasizing the need to modulate neuroimmune interfaces, restore glial function, and target inflammatory signalling at its origin.

These insights reinforce the need to expand the current neuroimmune model to encompass mechanical lymphatic obstruction and ECM bioaccumulation as co-drivers of immune dysregulation. By restoring lymphatic flow and targeting ECM-bound inflammatory metabolites, clinicians may disrupt the otherwise intractable cycle of glial priming, mast cell reactivity, and autonomic destabilization. This paradigm shift positions lymphatic decompression as both a diagnostic probe and a therapeutic tool in multisystem chronic illness.

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