Toll like receptors (TLR) and ME

Basic knowledge about TLR

The innate immune system is the first line of defence against invading microorganisms. The system consists of cells: neutrophils, monocytes/macrophages, dendritic cells and natural killer cells, and of soluble elements such as cytokines and components of the complement system. Detection of these microorganisms is mediated by pattern recognition receptors (PRR) expressed on the cells of the innate immune system. The most important components of PRR are Toll-like receptors (TLR). TLR recognize structures common to many pathogens, the so-called pathogen associated molecular patterns (PAMP). PAMP are part of endotoxins, peptidoglycans and other microbial molecules of clinical relevance in infectious diseases.  Danish Reference

There are 12 known TLRs – named TLR1-TLR12. Each TLR recognize a different structure. Some of them are (look in Wikipedia for a complete list):
TLR1/TLR2:   Lipopeptides from bacteria
TLR3:             Double-strandend RNA from viruses
TLR4:             Lipopolysaccharide (LPS) from gramnegative bacteria
TLR5:             Flagellin from bacteria
TLR6:             Multiple diacyl lipopeptides from Mycoplasma
TLR7/TLR8:  Small synthetic compounds, single-stranded RNA
TLR9:             Unmethylated CpG Oligodeoxynucleotide DNA

Agonists and antagonists respectively activate and de-activate receptors. TLR agonists have the ability to turn on the immune system, and TLR antagonists have the potential to be immunosuppressive and anti-inflammatory.

TLRs and ME

From the Whittemore Petersons website you can read:
The microbial pattern recognition receptors, Toll-like receptors (TLRs) 3 and 4, are activated by pathogen-derived dsRNA and LPS, respectively, to produce these proinflammatory cytokines. This system is tightly regulated; however, in patients with ME/CFS, it is constitutively active. The focus of this research project is to better understand the mechanism of this dysregulation.

Light et al also mention TLR4: After a sustained moderate exercise test, CFS patients showed greater increases than control subjects in gene expression for metabolite detecting receptors ASIC3, P2X4, and P2X5, for SNS receptors alpha-2A, beta-1, beta-2, and COMT and IS genes for IL10 and TLR4 lasting from 0.5 to 48 hours (P < .05). Reference

Maes et al have several times referred to LPS translocation as part of ME (and I presume that must activate TLR4):
The findings show that increased IgA responses to commensal bacteria in ME/CFS are associated with inflammation and CMI activation, which are associated with symptom severity. It is concluded that increased translocation of commensal bacteria may be responsible for the disease activity in some ME/CFS patients. Reference

Ampligen/Rintatolimod used for ME is a TLR3 agonist. Reference

Some ME patients have tried Naltrexone, which is a TLR 4 antagonist.

TLR and other diseases

Is there more to learn about ME and TLRs? Let us look for inspiration in other diseases:

Toll-like receptor signaling in neural plasticity and disease
TLR signaling in immune cells, glia and neurons can play roles in the pathogenesis of stroke, Alzheimer's disease (AD) and multiple sclerosis (MS). Recent findings suggest that TLR signaling also influences multiple dynamic processes in the developing and adult central nervous system including neurogenesis, axonal growth and structural plasticity. In addition, TLRs are implicated in the regulation of behaviors including learning, memory and anxiety. This review describes recently discovered and unexpected roles for TLRs in neuroplasticity, and the implications of these findings for future basic and translational research studies.

Toll-like receptors in rheumatic diseases: are we paying a high price for our defense against bugs?
In the last decade Toll-like receptor (TLR) research has led to new insights in the pathogenesis of many rheumatic diseases. In autoimmune diseases like systemic lupus erythematosus, rheumatoid arthritis and systemic sclerosis TLR signaling is likely to be involved in tolerance breakthrough and chronic inflammation via combined Fc gamma receptors and TLR recognition of immune complexes. Furthermore, inflammatory diseases like psoriatic arthritis and gout also show more and more evidence for TLR involvement. In this review we will discuss the involvement of TLR signaling in several rheumatic diseases and stress their similarities and differences based on recent findings.


Targeting Toll-like receptors: emerging therapeutics for multiple sclerosis management
Toll-like receptors (TLR) are important innate immune proteins for the identification and clearance of invading pathogen. TLR signal through adaptor proteins, most commonly myeloid differentiation primary response gene 88 (MyD88). Inappropriate response of specific TLR has been implicated in certain autoimmune diseases, such as multiple sclerosis (MS). Activation of TLR2, TLR4, TLR7 and TLR9 plays a role in experimental allergic encephalomyelitis (EAE), a murine model of MS, while TLR3 activation protects from disease. Therefore, TLR-modulation could be an important adjuvant to current treatments. Here, we focus on TLR involved in EAE and MS pathogenesis highlighting specific components targeting TLR that might offer further therapeutic possibilities.


The Emerging Role of TLR and Innate Immunity in Cardiovascular Disease
Cardiovascular disease is a complex disorder involving multiple pathophysiological processes, several of which involve activation of toll-like receptors (TLRs) of the innate immune system. As sentinels of innate immunity TLRs are nonclonally germline-encoded molecular pattern recognition receptors that recognize exogenous as well as tissue-derived molecular dangers signals promoting inflammation. In addition to their expression in immune cells, TLRs are found in other tissues and cell types including cardiomyocytes, endothelial and vascular smooth muscle cells. TLRs are differentially regulated in various cell types by several cardiovascular risk factors such as hypercholesterolemia, hyperlipidemia, and hyperglycemia and may represent a key mechanism linking chronic inflammation, cardiovascular disease progression, and activation of the immune system. Modulation of TLR signaling by specific TLR agonists or antagonists, alone or in combination, may be a useful therapeutic approach to treat various cardiovascular inflammatory conditions such as atherosclerosis, peripheral arterial disease, secondary microvascular complications of diabetes, autoimmune disease, and ischemia reperfusion injury. In this paper we discuss recent developments and current evidence for the role of TLR in cardiovascular disease as well as the therapeutic potential of various compounds on inhibition of TLR-mediated inflammatory responses.

TLR cross-talk specifically regulates cytokine production by B cells from chronic inflammatory disease patients
Chronic systemic inflammation links periodontal disease and diabetes to increased incidence of serious comorbidities. Activation of TLRs, particularly TLR2 and TLR4, promotes chronic systemic inflammation. Human B cells have been generally thought to lack these TLRs. However, recent work showed that an increased percentage of circulating B cells from inflammatory disease patients express TLR2 and TLR4, and that TLR engagement on B cells resulted in unexpected changes in gene expression. New data show that B cells from inflammatory disease patients secrete multiple cytokines in response to different classes of TLR ligands. Furthermore, the B cell response to combinations of TLR ligands is cytokine- and ligand-specific. Some cytokines (IL-1beta and IL-10) are predominantly regulated by TLR4, but others (IL-8 and TNF-alpha) are predominantly regulated by TLR2, due in part to TLR-dictated changes in transcription factor/promoter association. TLR2 and TLR9 also regulate B cell TLR4 expression, demonstrating that TLR cross-talk controls B cell responses at multiple levels. Parallel examination of B cells from periodontal disease and diabetes patients suggested that outcomes of TLR cross-talk are influenced by disease pathology. We conclude that disease-associated alteration of B cell TLR responses specifically regulates cytokine production and may influence chronic inflammation.


EBV Latent Membrane Protein 2A Induces Autoreactive B Cell Activation and TLR Hypersensitivity
TLR signaling has been suggested to be important to the activation of autoreactive B cells in autoimmunity…
Systemic lupus erythematosus (SLE) is a chronic autoimmune disorder characterized by the production of autoantibodies and multiorgan inflammatory damage. Both genetic and environmental factors contribute to the etiology of SLE, but a clear understanding of how these factors contribute to disease is still lacking. Among the environmental risk factors, EBV is the most closely associated with SLE. SLE patients have higher titers of anti-EBV Abs than control populations, and EBV infection is more common among juvenile and adult SLE patients than among control populations. Some cases of SLE appear to directly result from acute EBV infection or reactivation of EBV. In addition, SLE patients have a 40-fold higher EBV viral load in peripheral blood leukocytes than control populations due to poor cytotoxic T cell (CTL) responses and a higher frequency of infected B cells….
Since EBV-encoded latent membrane protein 2A (LMP2A) interferes with normal B cell differentiation and function, we sought to determine its effect on B cell tolerance…..
LMP2A induces a heightened sensitivity to TLR ligand stimulation, resulting in increased proliferation or antibody-secreting cell differentiation or both. Thus, we propose a model whereby LMP2A induces hypersensitivity to TLR stimulation, leading to activation of anti-Sm B cells through the BCR/TLR pathway..
These data further implicate TLRs in the etiology of SLE and suggest a mechanistic link between EBV infection and SLE…
We have focused on an SLE-specific autoantigen, but since this mechanism could act on B cells of specific for autoantigens targeted in other autoimmune diseases, EBV could have a role in the etiology of multiple autoimmune diseases such as Sjögren’s syndrome, rheumatoid arthritis, multiple sclerosis, and autoimmune hepatitis.

Toll-Like Receptor Agonists Synergistically Increase Proliferation and Activation of B Cells by Epstein-Barr Virus
Epstein-Barr virus (EBV) efficiently drives proliferation of human primary B cells in vitro, a process relevant for human diseases such as infectious mononucleosis and posttransplant lymphoproliferative disease. Human B-cell proliferation is also driven by ligands of Toll-like receptors (TLRs), notably viral or bacterial DNA containing unmethylated CpG dinucleotides, which triggers TLR9. Here we quantitatively investigated how TLR stimuli influence EBV-driven B-cell proliferation and expression of effector molecules. CpG DNA synergistically increased EBV-driven proliferation and transformation, T-cell costimulatory molecules, and early production of interleukin-6. CpG DNA alone activated only memory B cells, but CpG DNA enhanced EBVmediated transformation of both memory and naive B cells. Ligands for TLR2 or TLR7/8 or whole bacteria had a weaker but still superadditive effect on B-cell transformation. Additionally, CpG DNA facilitated the release of transforming virus by established EBV-infected lymphoblastoid cell lines. These results suggest that the proliferation of EBV-infected B cells and their capability to interact with immune effector cells may be directly influenced by components of bacteria or other microbes present at the site of infection.

Toll-like receptor 4 in CNS pathologies
Because microglial activation is widely controlled by pathogen recognition receptors (PRRs), TLR4 is implicated in the microglia-mediated neurotoxicity that occurs in many brain pathologies….
,,, If the TLR4 pathway is erroneously activated, or if a signal is amplified out of control, the cytokine response may have deleterious effects on the nervous system….
…Mice lacking functional TLR4 expression in CNS are exempt from long-term progressive neuron loss. This example illustrates the paradoxical nature of TLR4 signaling: it is necessary for defense, yet it invokes a powerful cascade that can be toxic. The stakes are high in the CNS, where subtle modifications can tip TLR4 signaling over the neurotoxic edge…
…Recent reports of TLR4 activation by endogenous ligands link TLR4 to autoimmunity as well as legitimate inflammation…
…Microglia, the immunocompetent cells of the CNS, are important contributors to chronic pain pathologies (Milligan and Watkins 2009). The TLR4 receptor is one avenue through which microglia can be activated and primed for the pain response. TLR4 influences the CNS pain response, invoking the production of pro-inflammatory cytokines and reactive oxygen species…
…LPS induces glial activation, and this activation can be ameliorated by naloxone administration (Wu et al. 2006). Based on the ability of naloxone to interfere with LPS signaling, the TLR4 pathway is implicated as a mediator of non-classical opioid responses…
…Consistent with the ability of naltrexone to reduce chronic pain symptoms, opioid
antagonists were found to inhibit TLR4 signaling and the production of pro-inflammatory substances. TLR4 inhibition was proportional to morphine analgesia effectiveness….
…Altogether, innate immunity and the responses coordinated by PRRs are extremely powerful modulators of the CNS environment…


Painful pathways induced by TLR stimulation of dorsal root ganglion neurons 
…TLR ligands upregulated the expression of a nociceptive receptor, transient receptor potential vanilloid type 1 (TRPV1), and enhanced calcium flux by TRPV1-expressing DRGNs. Using a tumor-induced temperature sensitivity model, we showed that in vivo administration of a TLR9 antagonist, known as a suppressive oligodeoxynucleotide, blocked tumor-induced temperature sensitivity. Taken together, these data indicate that stimulation of peripheral neurons by TLR ligands can induce nerve pain…

Emerging role of Toll-like receptors in the control of pain and itch
... In particular, primary sensory neurons, such as nociceptors, express TLRs (e.g., TLR4 and TLR7) to sense exogenous PAMPs and endogenous DAMPs released after tissue injury and cellular stress. These neuronal TLRs are new players in the processing of pain and itch by increasing the excitability of primary sensory neurons. Given the prevalence of chronic pain and itch and the suffering of affected people, insights into TLR signaling in the nervous system will open a new avenue for the management of clinical pain and itch...

Systemic challenge with the TLR3 agonist poly I:C induces amplified IFNalpha/beta and IL-1beta responses in the diseased brain and exacerbates chronic neurodegeneration
The role of inflammation in the progression of neurodegenerative disease remains unclear. We have shown that systemic bacterial insults accelerate disease progression in animals and in patients with Alzheimer's disease. Disease exacerbation is associated with exaggerated CNS inflammatory responses to systemic inflammation mediated by microglia that become 'primed' by the underlying neurodegeneration. The impact of systemic viral insults on existing neurodegenerative disease has not been investigated. Polyinosinic:polycytidylic acid (poly I:C) is a toll-like receptor-3 (TLR3) agonist and induces type I interferons, thus mimicking inflammatory responses to systemic viral infection. In the current study we hypothesized that systemic challenge with poly I:C, during chronic neurodegenerative disease, would amplify CNS inflammation and exacerbate disease. Using the ME7 model of prion disease and systemic challenge with poly I:C (12 mg/kg i.p.) we have shown an amplified expression of IFN-alpha and beta and of the pro-inflammatory genes IL-1beta and IL-6. Similarly amplified expression of specific IFN-dependent genes confirmed that type I IFNs were secreted and active in the brain and this appeared to have anti-inflammatory consequences. However, prion-diseased animals were susceptible to heightened acute sickness behaviour and acute neurological impairments in response to poly I:C and this treatment also accelerated disease progression in diseased animals without effect in normal animals. Increased apoptosis coupled with double-stranded RNA-dependent protein kinase (PKR) and Fas transcription suggested activation of interferon-dependent, pro-apoptotic pathways in the brain of ME7+poly I:C animals. That systemic poly I:C accelerates neurodegeneration has implications for the control of systemic viral infection during chronic neurodegeneration and indicates that type I interferon responses in the brain merit further study.

Chronic ethanol increases systemic TLR3 agonist-induced neuroinflammation and neurodegeneration
Increasing evidence links systemic inflammation to neuroinflammation and neurodegeneration. We previously found that systemic endotoxin, a TLR4 agonist or TNFalpha, increased blood TNFalpha that entered the brain activating microglia and persistent neuroinflammation. Further, we found that models of ethanol binge drinking sensitized blood and brain proinflammatory responses. We hypothesized that blood cytokines contribute to the magnitude of neuroinflammation and that ethanol primes proinflammatory responses. Here, we investigate the effects of chronic ethanol on neuroinflammation and neurodegeneration triggered by toll-like receptor 3 (TLR3) agonist poly I:C.
Poly I:C increased blood and brain TNFalpha that peaked at three hours. Blood levels returned within one day, whereas brain levels remained elevated for at least three days. Escalating blood and brain proinflammatory responses were found with ethanol, poly I:C, and ethanol-poly I:C treatment. Ethanol pretreatment potentiated poly I:C-induced brain TNFalpha (345%), IL-1beta (331%), IL-6 (255%), and MCP-1(190%). Increased levels of brain cytokines coincided with increased microglial activation, NOX gp91phox, superoxide and markers of neurodegeneration (activated caspase-3 and Fluoro-Jade B). Ethanol potentiation of poly I:C was associated with ethanol-increased expression of TLR3 and endogenous agonist HMGB1 in the brain. Minocycline and naltrexone blocked microglial activation and neurodegeneration.
Ethanol potentiation of TLR3 agonist responses is consistent with priming microglia-monocytes and increased NOX, ROS, HMGB1-TLR3 and markers of neurodegeneration. These studies indicate that TLR3 agonists increase blood cytokines that contribute to neurodegeneration and that ethanol binge drinking potentiates these responses.

Apparently TLRs are involved in many conditions and diseases - so this is one topic more to keep an eye on.