Tobacco-derived 4R-cembranoid confers neuroprotection against LPS-induced hippocampal inflammation in mice (decreases levels of pro-inflammatory cytokines; improves memory function; activates STAT3, Akt1, and CREB phosphorylation; and upregulates the mRNA levels of ORM2, GDNF, and C3) independent of the α7 nicotinic receptor

January 24, 2024
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Rojas-Colón, L.A., Dash, P.K., Morales-Vías, F.A. et al. 4R-cembranoid confers neuroprotection against LPS-induced hippocampal inflammation in mice. J Neuroinflammation 18, 95 (2021). https://doi.org/10.1186/s12974-021-02136-9

LPS: Lipopolysaccharide

“4R protects the hippocampus against inflammation and memory impairments triggered by LPS by lowering TNF-α and IL-1β levels and activation of the Akt1 and CREB signaling pathways. Astrocyte proteins involved in neuronal survival also seem to be modulated by 4R. The effects of 4R in the hippocampus are independent of α7 nicotinic receptors. The diagram below represents a model of 4R action in the hippocampus during inflammation.“

figure a

Some other notes

Lipopolysaccharides (LPS) are large molecules consisting of a lipid and a polysaccharide that are bacterial toxins. They are composed of an O-antigen, an outer core, and an inner core all joined by covalent bonds, and are found in the bacterial capsule, the outermost membrane of cell envelope of Gram-negative bacteria, such as E. coli and Salmonella. Today, the term endotoxin is often used synonymously with LPS, although there are a few endotoxins (in the original sense of toxins that are inside the bacterial cell that are released when the cell disintegrates) that are not related to LPS, such as the so-called delta endotoxin proteins produced by Bacillus thuringiensis. Lipopolysaccharides can have substantial impacts on human health, primarily through interactions with the immune system. LPS is a potent activator of the immune system and pyrogen (agent that causes fever). In severe cases, LPS can play a role in causing septic shock. In lower levels and over a longer time period, there is evidence LPS may play an important and harmful role in autoimmunity, obesity, depression, and cellular senescence.

Lipopolysaccharides are frequent contaminants in plasmid DNA prepared from bacteria or proteins expressed from bacteria, and must be removed from the DNA or protein to avoid contaminating experiments and to avoid toxicity of products manufactured using industrial fermentation.

  • Wicks IP, Howell ML, Hancock T, Kohsaka H, Olee T, Carson DA (March 1995). “Bacterial lipopolysaccharide copurifies with plasmid DNA: implications for animal models and human gene therapy”. Human Gene Therapy. 6 (3): 317–323. doi:10.1089/hum.1995.6.3-317. PMID 7779915.

Ovalbumin is frequently contaminated with endotoxins. Ovalbumin is one of the extensively studied proteins in animal models and also an established model allergen for airway hyper-responsiveness (AHR). Commercially available ovalbumin that is contaminated with LPS can falsify research results, as it does not accurately reflect the effect of the protein antigen on animal physiology.

In pharmaceutical production, it is necessary to remove all traces of endotoxin from drug product containers, as even small amounts of endotoxin will cause illness in humans. A depyrogenation oven is used for this purpose. Temperatures in excess of 300 °C are required to fully break down LPS.

The standard assay for detecting presence of endotoxin is the Limulus Amebocyte Lysate (LAL) assay, utilizing blood from the Horseshoe crab (Limulus polyphemus).

Very low levels of LPS can cause coagulation of the limulus lysate due to a powerful amplification through an enzymatic cascade. However, due to the dwindling population of horseshoe crabs, and the fact that there are factors that interfere with the LAL assay, efforts have been made to develop alternative assays, with the most promising ones being ELISA tests using a recombinant version of a protein in the LAL assay, Factor C.

Lipopolysaccharide, is a significant factor that makes bacteria harmful, and it helps categorize them into different groups based on their structure and function. This makes LPS a useful marker for telling apart various Gram-negative bacteria. Swiftly identifying and understanding the types of pathogens involved is crucial for promptly managing and treating infections. Since LPS is the main trigger for the immune response in our cells, it acts as an early signal of an acute infection. Therefore, LPS testing is more specific and meaningful than many other serological tests. The current methods for testing LPS are quite sensitive, but many of them struggle to differentiate between different LPS groups. Additionally, the nature of LPS, which has both water-attracting and water-repelling properties (amphiphilic), makes it challenging to develop sensitive and user-friendly tests. The typical detection methods rely on identifying the lipid A part of LPS. However, this method has limitations because Lipid A is very similar among different bacterial species and serotypes. LPS testing techniques fall into six categories, and they often overlap: in vivo tests, in vitro tests, modified immunoassays, biological assays, and chemical assays. 

LPS is a powerful toxin that, when in the body, triggers inflammation by binding to cell receptors. Excessive LPS in the blood can lead to endotoxemia, potentially causing a harmful condition called septic shock. This condition includes symptoms like rapid heart rate, quick breathing, temperature changes, and blood clotting issues, resulting in blood vessels widening and reduced blood volume, leading to cellular dysfunction. Recent research indicates that even small LPS exposure is associated with autoimmune diseases and allergies. High levels of LPS in the blood can lead to metabolic syndrome, increasing the risk of conditions like diabetes, heart disease, and liver problems. LPS also plays a crucial role in symptoms caused by infections from harmful bacteria, including severe conditions like Waterhouse-Friderichsen syndrome, meningococcemia, and meningitis. Certain bacteria can adapt their LPS to cause long-lasting infections in the respiratory and digestive systems. Recent studies have shown that LPS disrupts cell membrane lipids, affecting cholesterol and metabolism, potentially leading to high cholesterol, abnormal blood lipid levels, and non-alcoholic fatty liver disease. In some cases, LPS can interfere with toxin clearance, which may be linked to neurological issues.

See also

STAT3

Niclosamide seems to inhibit the STAT3 signalling pathway.

Nicotinamide (a type of Vitamin B3 – the most common and very worst type IMO) naturally inhibits STAT3.

However NAC (Acetylcysteine) inhibits STAT3 inhibitors.

That’s very interesting to me because according to the internet, NAC (which has been available as a supplement is currently being hijacked by all manner of cretins) is the antidote to paracetamol/tylenol poisoning and maybe aspartame poisoning (which is apparently more common than they care to admit particularly in the nonsmoker as the nonsmoker does not have a human metabolism or clearance system)

STAT3 has been shown to interact with:

Akt1

RAC(Rho family)-alpha serine/threonine-protein kinase is an enzyme that in humans is encoded by the AKT1 gene. This enzyme belongs to the AKT subfamily of serine/threonine kinases that contain SH2 (Src homology 2-like) protein domains. It is commonly referred to as PKB, or by both names as “Akt/PKB”.

The serine-threonine protein kinase AKT1 is catalytically inactive in serum-starved primary and immortalized fibroblasts. AKT1 and the related AKT2 are activated by platelet-derived growth factor. The activation is rapid and specific, and it is abrogated by mutations in the pleckstrin homology domain of AKT1. It was shown that the activation occurs through phosphatidylinositol 3-kinase. In the developing nervous system AKT is a critical mediator of growth factor-induced neuronal survival. Survival factors can suppress apoptosis in a transcription-independent manner by activating the serine/threonine kinase AKT1, which then phosphorylates and inactivates components of the apoptotic machinery. Mice lacking Akt1 display a 25% reduction in body mass, indicating that Akt1 is critical for transmitting growth-promoting signals, most likely via the IGF1 receptor. Mice lacking Akt1 are also resistant to cancer: They experience considerable delay in tumor growth initiated by the large T antigen or the Neu oncogene.

A single-nucleotide polymorphism in this gene causes Proteus syndrome. In 2011, a mutation in AKT1 was strongly associated with Proteus syndrome, the disease that probably affected the Elephant Man.

  • Lindhurst MJ, Sapp JC, Teer JK, Johnston JJ, Finn EM, Peters K, Turner J, Cannons JL, Bick D, Blakemore L, Blumhorst C, Brockmann K, Calder P, Cherman N, Deardorff MA, Everman DB, Golas G, Greenstein RM, Kato BM, Keppler-Noreuil KM, Kuznetsov SA, Miyamoto RT, Newman K, Ng D, O’Brien K, Rothenberg S, Schwartzentruber DJ, Singhal V, Tirabosco R, Upton J, Wientroub S, Zackai EH, Hoag K, Whitewood-Neal T, Robey PG, Schwartzberg PL, Darling TN, Tosi LL, Mullikin JC, Biesecker LG (2011). “A mosaic activating mutation in AKT1 associated with the Proteus syndrome”. N. Engl. J. Med. 365 (7): 611–9. doi:10.1056/NEJMoa1104017. PMC 3170413. PMID 21793738.
  • Cohen MM (2014). “Proteus syndrome review: molecular, clinical, and pathologic features”. Clin. Genet. 85 (2): 111–9. doi:10.1111/cge.12266. PMID 23992099. S2CID 204999819.
  • Lindhurst MJ, Sapp JC, Teer JK, Johnston JJ, Finn EM, Peters K, Turner J, Cannons JL, Bick D, Blakemore L, Blumhorst C, Brockmann K, Calder P, Cherman N, Deardorff MA, Everman DB, Golas G, Greenstein RM, Kato BM, Keppler-Noreuil KM, Kuznetsov SA, Miyamoto RT, Newman K, Ng D, O’Brien K, Rothenberg S, Schwartzentruber DJ, Singhal V, Tirabosco R, Upton J, Wientroub S, Zackai EH, Hoag K, Whitewood-Neal T, Robey PG, Schwartzberg PL, Darling TN, Tosi LL, Mullikin JC, Biesecker LG (27 July 2011). “A Mosaic Activating Mutation in Associated with the Proteus Syndrome”. New England Journal of Medicine. 365 (7): 611–619. doi:10.1056/NEJMoa1104017. PMC 3170413. PMID 21793738

AKT (now also called AKT1) was originally identified as the oncogene in the transforming retrovirus, AKT8. The name Akt stands for Ak strain transforming. The origins of the Akt name date back to 1928,

AKT1 has been shown to interact with:

See also

  • AKT – the AKT family of proteins
  • AKT2 – the gene for the second member of the AKT family
  • AKT3 – the gene for the third member of the AKT family
  • Proteus syndrome

CREB phosphorylation

Not to be confused with Clean Renewable Energy Bonds.

CREB-TF (CREB, cAMP response element-binding protein) is a cellular transcription factor. It binds to certain DNA sequences called cAMP response elements (CRE), thereby increasing or decreasing the transcription of the genes. CREB was first described in 1987 as a cAMP-responsive transcription factor regulating the somatostatin gene. Genes whose transcription is regulated by CREB include: c-fos, BDNF, tyrosine hydroxylase, numerous neuropeptides (such as somatostatin, enkephalin, VGF, corticotropin-releasing hormone), and genes involved in the mammalian circadian clock (PER1, PER2). CREB is closely related in structure and function to CREM (cAMP response element modulator) and ATF-1 (activating transcription factor-1) proteins. CREB proteins are expressed in many animals, including humans. CREB has a well-documented role in neuronal plasticity and long-term memory formation in the brain and has been shown to be integral in the formation of spatial memory. CREB downregulation is implicated in the pathology of Alzheimer’s disease and increasing the expression of CREB is being considered as a possible therapeutic target for Alzheimer’s disease. CREB also has a role in photoentrainment in mammals.

See also

ORM2 (this doesn’t have a Wikipedia page. It is briefly mentioned on several others including ORM)

Orosomucoid (ORM) or alpha-1-acid glycoprotein (α1AGp, AGP or AAG) is an acute phase protein found in plasma. It is an alpha-globulinglycoprotein and is modulated by two polymorphic genes. It is synthesized primarily in hepatocytes and has a normal plasma concentration between 0.6–1.2 mg/mL (1–3% plasma protein). Plasma levels are affected by pregnancy, burns, certain drugs, and certain diseases, particularly HIV.

See also

Lipocalins mentions ORM2 as one of the human proteins that contain lipocalin domain

The lipocalins are a family of proteins which transport small hydrophobic molecules such as steroids, bilins, retinoids, and lipids, and most lipocalins are also able to bind to complexed iron (via siderophores or flavonoids) as well as heme. They share limited regions of sequence homology and a common tertiary structure architecture. This is an eight stranded antiparallel beta barrel with a repeated + 1 topology enclosing an internal ligand binding site. These proteins are found in gram negative bacteria, vertebrate cells, and invertebrate cells, and in plants. Lipocalins have been associated with many biological processes, among them immune response, pheromone transport, biological prostaglandin synthesis, retinoid binding, and cancer cell interactions.

The name “lipocalin” has been proposed for this protein family, but cytosolic fatty acid binding proteins are also included. The sequences of most members of the family, the core or kernel lipocalins, are characterised by three short conserved stretches of residues, while others, the outlier lipocalin group, share only one or two of these. Proteins known to belong to this family include alpha-1-microglobulin (protein HC); major urinary proteins; alpha-1-acid glycoprotein (orosomucoid); aphrodisin; apolipoprotein D; beta-lactoglobulin; complement component C8 gamma chain; crustacyanin; epididymal-retinoic acid binding protein (E-RABP); insectacyanin; odorant binding protein (OBP); human pregnancy-associated endometrial alpha-2 globulin (PAEP); probasin (PB), a prostatic protein; prostaglandin D synthase; purpurin; Von Ebner’s gland protein (VEGP); and lizard epididymal secretory protein IV (LESP IV).

Human proteins that contain lipocalin domain include:

See also

Anticalin proteins are artificial proteins that are able to bind to antigens, either to proteins or to small molecules. They are not structurally related to antibodies, which makes them a type of antibody mimetic. Instead, they are derived from human lipocalins which are a family of naturally binding proteins. Anticalin proteins are being used in lieu of monoclonal antibodies, but are about eight times smaller with a size of about 180 amino acids and a mass of about 20 kDa. The Anticalin technology is exclusively commercialized by Pieris Pharmaceuticals in Freising, Germany. Anticalin is a registered trademark of Pieris.[citation needed] Anticalin proteins have better tissue penetration than antibodies and are stable at temperatures up to 70 °C. Unlike antibodies, they can be produced in bacterial cells like E. coli in large amounts. While antibodies can only be directed at macromolecules such as proteins and at small molecules (haptens) only if bound to macromolecules, Anticalin proteins are able to selectively bind to small molecules as well.[citation needed] They were mainly developed at the Technical University of Munich and are currently used as research tools. Diagnostic and therapeutic applications, including the use for targeted drug delivery, are being aimed at. The underlying technology was nominated for the German Future Prize in 2004. Characteristic for Anticalin proteins is their barrel structure formed by eight antiparallel β-strands pairwise connected by loops and an attached α-helix. The main structure of Anticalin proteins is identical to wild type lipocalins. Conformational deviations are primarily located in the four loops reaching in the ligand binding site. Mutagenesis of amino acids at the binding site allows for changing the affinity and selectivity.[citation needed]

Engineered monoclonal antibodies and antibody mimetics

Categories: 

GDNF

Glial cell line-derived neurotrophic factor (GDNF) is a protein that, in humans, is encoded by the GDNFgene. GDNF is a small protein that potently promotes the survival of many types of neurons. It signals through GFRα receptors, particularly GFRα1. It is also responsible for the determination of spermatogonia into primary spermatocytes, i.e. it is received by RET proto-oncogene (RET) and by forming gradient with SCF it divides the spermatogonia into two cells. As the result there is retention of spermatogonia and formation of spermatocyte. GDNF was discovered in 1991, and is the first member of the GDNF family of ligands (GFL) found. GDNF is highly distributed throughout both the peripheral and central nervous system. It can be secreted by astrocytes, oligodendrocytes, Schwann cells, motor neurons, and skeletal muscle during the development and growth of neurons and other peripheral cells. The GDNF gene encodes a highly conserved neurotrophic factor. The recombinant form of this protein was shown to promote the survival and differentiation of dopaminergic neurons in culture, and was able to prevent apoptosis of motor neurons induced by axotomy. 

C3

Complement component 3, often simply called C3, is a protein of the immune system that is found primarily in the blood. It plays a central role in the complement system of vertebrate animals and contributes to innate immunity. In humans it is encoded on chromosome 19 by a gene called C3. Deficiencies and defects of C3 result in the affected person being immunocompromised and particularly vulnerable to bacterial infections. In humans, C3 is predominantly synthesised by liver hepatocytes and to some degree by epidermis keratinocytes. Factor H is the primary regulator of C3. Deficiency of Factor H may lead to uncontrolled C3 activity through the alternative pathway of the complement system.

Deficiency of C3 results in the affected person being immunocompromised. Specifically, they are vulnerable to bacterial pathogens, including repeat infections by the same organism, but are not susceptible to viruses. This vulnerability also occurs in an individual deficient in C1, C2, C4, or any of their required components or associated proteins, and the clinical effects are very similar regardless of the specific deficiency. This is because all of these must work with C3 for the complement system to function. Affected people are particularly vulnerable to infections with Gram-negative organisms such as pathogenic E. coli or Salmonella enterica. Additionally, C3 and other complement deficiencies are associated with frequent and severe respiratory infections, as well as other infections that invade and penetrate tissue layers. Some data shows that acquired C3 deficiency, including when this is intentionally done for medical immunosuppression purposes, may not significantly impact a person’s immune function long-term. However, by contrast, congenital C3 deficiency is known to cause chronic illness. Additionally, several forms of C3 deficiency contribute to the development of systemic lupus erythematosus and other autoimmune diseases.

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