Editing Spike as a superantigen


= Existing animal models =

=== COVID vaccine | Perimyocarditis ===

A Hong Kong group used the Pfizer vaccine on mice.  When the vaccine was injected intravenously (rather than via the intramuscular route), the mice developed perimyocarditis.  https://academic.oup.com/cid/article/74/11/1933/6353927

Female and male Balb/c (substrain OlaHsd) mice were used.

'''Dosage''':  COVID-19 mRNA vaccine (BNT162b2 lot number 1B004A, BioNTech, Germany) dose of 0.25 µg per gram of body weight was injected (about 5 µg in 50 µL per mouse; dose according to an immunogenicity study).  A Pfizer dose for a lightweight 40kg human would be 30 µg, or 0.00075 µg per gram of body weight.  The dosage used in the animal study was roughly 333X that of a human dose.

=== Gardasil HPV vaccine | HPV vaccination associated neuro-immunopathetic syndrome (HANS) ===

Satoko Aratani et al. at Tokyo Medical University.  HPV vaccine + pertussis toxin in mice.
* https://doi.org/10.1038/srep36943 - ''Murine hypothalamic destruction with vascular cell apoptosis subsequent to combined administration of human papilloma virus vaccine and pertussis toxin''

=== Gardasil HPV vaccine | ASIA syndrome ===

* https://autoimmunity-network.com/articles/Behavioral%20abnormalities%20in%20female%20mice%20following%20HPV%20vac%20%20%202017.pdf  Mice were given aluminium adjuvant only, Gardasil or Gardasil + pertussis toxin.  Forced swimming test 3 months after injection was the most sensitive test.
* https://autoimmunity-network.com/articles/Maria%20Biobody.pdf - Mice vaccinated with Gardasil and aluminium adjuvant only fared worse on a forced swim test.  Phospholipid supplementation can attenuate vaccine-induced depressive-like behavior.

=== COVID vaccine | Multiple organ failure ===

* A [https://igorchudov.substack.com/p/fully-vaccinated-gorilla-dies-of?s=r Substack article] speculates that a Western lowland gorilla died from COVID vaccination.

=== Hepatitis B vaccine | SLE / Lupus ===

* https://sci-hub.se/https://doi.org/10.1016/j.jaut.2014.06.006 - Mice were immunized with Hep-B vaccine, aluminium adjuvant, or the control injection (phosphate-buffered saline).  SLE-disease resulted for the vaccine and alum group.  Immunization with either HBVv or alum affected blood counts, neurocognitive functions and brain gliosis.  Various biomarkers were examined, though measurements had mixed results.

=== Multiple vaccinations | ASIA syndrome ===

ASIA syndrome - commercial sheep were given repeated vaccinations (thiomersal/thimerosal, aluminium salts, antigen) https://sci-hub.se/https://doi.org/10.1007/s12026-013-8404-0

=== SARS-CoV-2 spike protein | Acute COVID ===

Biancatelli, Solopov, et al. (in a group that included Paul Marik) tested SARS-CoV-2 spike protein on humanized mice and wild-type mice (https://doi.org/10.1152/ajplung.00223.2021).  The genetics of the Κ18-hACE2 mice were manipulated to overexpress human ACE2 receptors.  The humanized Κ18-hACE2 mice exhibited a more severe form of COVID-19 symptoms.

=== SARS-CoV-2 S1 and S2 spike protein | Inflammatory cytokines ===

https://doi.org/10.7554/eLife.68563 - Khan et al. injected wild-type and TLR2-deficient mice intraperitoneally.  IL-6, IL-1β, and TNFα were elevated following administration of S proteins in WT mice, whereas no such induction was observed in Tlr2−/− mice.

=== Aluminium adjuvant | Gulf War Illness / Motor deficits / Cognitive deficits ===

* https://link.springer.com/article/10.1385/NMM:9:1:83 - An animal model for examining the potential neurological impact of aluminum hydroxide, squalene, or aluminum hydroxide combined with squalene was developed.  Behavioral testing showed motor deficits in the aluminum treatment group that expressed as a progressive decrease in strength measured by the wire-mesh hang test (final deficit at 24 wk; about 50%). Significant cognitive deficits in water-maze learning were observed in the combined aluminum and squalene group (4.3 errors per trial) compared with the controls (0.2 errors per trial) after 20 wk.

=== Other models ===

Other models such as squalene, aluminium adjuvant, other vaccines, etc. etc. likely exist.

=Superantigen theory notes=

https://sci-hub.se/https://doi.org/10.1016/0882-4010(87)90003-9 - In the TMEV virus, inbred mice can be more susceptible or more resistant to the virus.

=== HPV Gardasil vaccine ===

https://www.nature.com/articles/srep36943 - This Japanese group demonstrated an animal model for the HPV vaccine using pertussis toxin to skew the autoimmunity towards neurological phenotypes that include low responsiveness of the tail reflex and locomotive mobility.  They compared HPV + pertussis to MOG (myelin oligodendrocyte glycoprotein) antigen + pertussis toxin.

=== Kawasaki disease (similar to MIS-C) ===

https://sci-hub.se/10.1097/00001432-199506000-00005 - This 1995 links superantigens from S. Aureus and group A streptococci to staphlococcal and streptococcal toxic shock syndrome, Kawasaki syndrome, and the chronic diseases arthritis, guttate psoriasis, and atopic dermatitis.

=== SARS-CoV-2 spike protein activates TLR4 and acts as a superantigen (or not) ===

https://www.nature.com/articles/s41422-021-00495-9 - SARS COV 2 spike protein activates TLR4 and acts as a superantigen

https://www.mdpi.com/2073-4409/11/16/2526 - '''Unlike SEB, the SARS-CoV-2 spike does not exhibit an intrinsic superantigen-like activity (!!!)'''.

=== Human Toll Like Receptors (and their adapters) ===

The table below summarizes TLRs (Toll Like Receptors) and ligands that set off these receptors.

* BLP = Bacterial Lipoproteins
* PGN = Peptidoglycan
* LPS = Lipopolysaccharide 
* dsRNA = Double-stranded RNA
* ssRNA = Single-stranded RNA
* Taxol = Paclitaxel, a chemotherapy medication

[[File:Human TLRs and their adapters.png]]

From DOI:[https://sci-hub.se/https://doi.org/10.1007/s12016-013-8402-y 10.1007/s12016-013-8402-y]

[[File:TLR2-TLR4-ligands.png]]

=== TLR4 ===

'''https://pubmed.ncbi.nlm.nih.gov/12799214/''' - LPS is used in Experimental autoimmune thyroiditis (EAT) model.  '''LPS''' and Freund's adjuvant behave differently.

'''https://www.jimmunol.org/content/175/2/959''' - '''LPS''' induces relapse of EAE (Experimental Autoimmune Encephalomyelitis) in mice.

'''https://pubmed.ncbi.nlm.nih.gov/18203139/''' - !!! The TLR4 pathway seems to regulate the priming of Th17 cells and reduce the severity of EAE.

https://pubmed.ncbi.nlm.nih.gov/19211042/ - "TLR4 down regulates disease severity in EAE and Th17 cell responses"

https://www.jimmunol.org/content/183/1/298.long - Early Life Exposure to Lipopolysaccharide Suppresses Experimental Autoimmune Encephalomyelitis by Promoting Tolerogenic Dendritic Cells and Regulatory T Cells

https://www.sciencedirect.com/science/article/abs/pii/S0165572806003717 - "Treatment with the bacterial product lipopolysaccharide (LPS) prior to the induction of experimental autoimmune encephalomyelitis (EAE) consistently led to a delayed onset of disease but not to a reduction in disease severity."

https://pubmed.ncbi.nlm.nih.gov/21685327/ - This study had conflicting results.  TLR4 knockout mice had the same susceptibility to experimental autoimmune encephalomyelitis (EAE).

https://www.ncbi.nlm.nih.gov/pmc/articles/PMC1375233/ - TLR4 ligands include '''LPS''', Taxol

https://jitc.biomedcentral.com/articles/10.1186/s40425-019-0625-x - '''LPS''' is used to treat cancer as a TLR4 ligand / vaccine adjuvant.

http://www.worldcat.org/title/1051926526 - LPS and Freund's are used in the livestock industry for a commercial purpose (???measuring vaccine response???)

https://www.ncbi.nlm.nih.gov/pmc/articles/PMC3064511/ - Review of TLRs (Toll-like receptors) in Multiple Sclerosis / experimental autoimmune encephalomyelitis (EAE)

https://www.ncbi.nlm.nih.gov/pmc/articles/PMC2327198/ - Bacterial '''LPS''' acts as an adjuvant to induce autoimmune arthritis in mice.  LPS enhances MRL/lpr nephritis, experimental autoimmune uveitis, experimental autoimmune myocarditis and experimental autoimmune enterocolitis.

=== TLR2 ===

https://doi.org/10.7554/eLife.68563 - A study involving TLR2-deficient macrophages suggests that spike protein (but not membrane, envelope, or nucleocapsid) triggers inflammatory cytokines and chemokines via the TLR2 pathway.  Spike protein injected into normal and TLR2-deficient mice also demonstrated inflammation dependent on the TLR2 pathway.  They also found an inflammatory response in TLR4 deficient macrophages (!), suggesting that the TLR2 pathway is what leads to an inflammatory response when cells are exposed to extracellular spike protein.  The S2 subunit of the spike protein was more pro-inflammatory than S1.  SARS-CoV-2 S protein induced proinflammatory cytokines and chemokines in macrophages and monocytes regardless of whether or not the ACE2 pathway was inhibited with MLN-4760.

https://pubmed.ncbi.nlm.nih.gov/12538041/ - This study's title argues: "Freund adjuvant induces TLR2 but not TLR4 expression in the liver of mice".  Note that it is the solvent in Freund's adjuvant that can trigger TLR2 mRNA expression on its own without killed mycobacterium.

=== Complete Freund's adjuvant ===

https://pubmed.ncbi.nlm.nih.gov/2139617/ - a single injection of complete Freund's adjuvant (CFA) given at an early age (5 wk) prevented the appearance of diabetes and greatly increased the life span of NOD mice without additional therapy

https://pubmed.ncbi.nlm.nih.gov/8436836/ - The BCG vaccine (or a single injection of CFA) protected mice against diabetes.  However, immunosuppression via the chemo drug cyclophosphamide could induce disease in 'protected' mice.

=== EAE - Experimental Allergic Encephalomyelitis  / Experimental Autoimmune Encephalomyelitis ===

https://oxfordrecoverycenter.com/wp-content/uploads/2017/02/Ameliorating-Effects-of-Hyperbaric-oxygenation-on-Experimental-Allergic-Encephalomyelitis.pdf - HBOT was successful in pushing EAE (experimental allergic encephalomyelitis) into remission.  CFA (complete freund's adjuvant) with guinea pig spinal cord.

=== Myocarditis ===

https://onlinelibrary.wiley.com/doi/full/10.1002/iid3.155 - Autoimmune myocarditis model, CFA / TiterMax Gold Adjuvant + myocarditogenic peptides in male mice

https://pubmed.ncbi.nlm.nih.gov/31005592/ - Leonurine alleviates LPS-induced myocarditis through suppressing the NF-кB signaling pathway

== Animal models ==

* Myocarditis
* Arthritis
* Encephalomyelitis / Multiple sclerosis
* Enterocolitis
* Uveitis
* Nephritis
* Non-obese diabetic / Type 1 diabetes
* Depressive-like behavior (forced swim test) in HPV-vaccinated mice https://autoimmunity-network.com/articles/Maria%20Biobody.pdf
* ASIA syndrome - commercial sheep were given repeated vaccinations (thiomersal/thimerosal, aluminium salts, antigen)  https://sci-hub.se/https://doi.org/10.1007/s12026-013-8404-0
* Vasculitis?  https://sci-hub.se/https://doi.org/10.1177/0961203311429553
* Acute motor axonal neuropathy (AMAN, a GBS variant) https://sci-hub.se/https://doi.org/10.1016/j.neulet.2004.06.059
* Peripheral neuropathy  [https://d1wqtxts1xzle7.cloudfront.net/49961679/s0304-3940_2801_2902143-720161029-9547-1alngxq-libre.pdf?1477752273=&response-content-disposition=inline%3B+filename%3DInflammation_with_no_axonal_damage_of_th.pdf&Expires=1663267752&Signature=S3cgHwJiRc~t1FSPwYSLTfFG68FUfz5DozQ3Cr0xSdZKDctpKJ0TP8vVm~e2sXd-QrCY0XDTf3MyiKr2SV2~FsoeMlZGDdMgT-DjOQHEOEi0pcezBOmM-cEc1pjW9-TAUIdiq1WV9dXpAHqSBaItuXbhNeHC7H-kwc3SUTA~V3IYSMVFkjinautflshxljQaLVRqukzr~KzbH63l80iEmfoH9f8g-lCi3IBNSyWR9xUVYE6BM29Z-yauCvoTkXWJYFqmhxF0fnvOT7bpofSICVckT764fFbhIRMHFBeJs42CrqaDcOtyadVM5DtOuo8NYP8T62XCyiVxQPqZEQafPg__&Key-Pair-Id=APKAJLOHF5GGSLRBV4ZA https://doi.org/10.1016/S0304-3940(01)02143-7] - CFA applied to the saphenous nerves (with its spread limited by Oxycel) leads to mechanosensitivity.
* Allergic neuritis (EAN / experimental allergic neuritis) - [https://sci-hub.se/https://doi.org/10.1111/j.1529-8027.2000.22-44.x Sci hub] | DOI:[https://doi.org/10.1111/j.1529-8027.2000.22-44.x 10.1111/j.1529-8027.2000.22-44.x]  - Triple combination of CFA, pertussis toxin in the presence/absence of murine recombinant IL-12, and bovine peripheral nerve myelin (BPNM).

= Researchers =

* Chronic Lyme researchers:  Ying Zhang, Monica Embers, Anna Goc
* Can Li et al.  Hong Kong SAR researchers.  The University of Hong Kong  https://academic.oup.com/cid/article/74/11/1933/6353927
* Satoko Aratani et al. - Tokyo Medical University - HPV vaccine animal model
* Kaohsiung Medical University / Chiou et al. - HBOT study on EAE.
* Bhagirath Singh - Department of Microbiology and Immunology, University of Western Ontario, London, Ontario N6A 5C1, Canada - [https://www.researchgate.net/publication/14762146_Complete_Freund's_adjuvant-induced_T_cells_prevent_the_development_and_adoptive_transfer_of_diabetes_in_nonobese_diabetic_mice publication on NOD mice given CFA and BCG vaccine].  Funded by Canadian government grants.
* Bassi et al. - University of Padova, Italy - talks about ASIA syndrome - https://sci-hub.se/https://doi.org/10.1177/0961203311429553
* Petrik et al. - University of British Columbia - https://link.springer.com/article/10.1385/NMM:9:1:83
* Biancatelli, Solopov, et al. - Frank Reidy Research Center for Bioelectrics, Old Dominion University (USA) - https://doi.org/10.1152/ajplung.00223.2021
* Khan et al. - Animal and cell line model of spike protein activating TLR2, TLR4 pathways - https://doi.org/10.7554/eLife.68563

= Treatments  + Autoimmunity inducers =

=== THC ===

* https://sci-hub.se/10.1016/0165-5728(89)90075-1 - THC for multiple sclerosis / experimental autoimmune encephalomyelitis (EAE).

=== HBOT ===

* https://oxfordrecoverycenter.com/wp-content/uploads/2017/02/Ameliorating-Effects-of-Hyperbaric-oxygenation-on-Experimental-Allergic-Encephalomyelitis.pdf - HBOT was successful in pushing EAE (experimental allergic encephalomyelitis) into remission. CFA (complete freund's adjuvant) with guinea pig spinal cord.
* [https://oxfordrecoverycenter.com/wp-content/uploads/2017/02/OXYGEN-IMMUNOSUPPRESSION-MODIFICATION-OF-EXPERIMENTAL-ALLERGIC-ENCEPHALOMYELITIS-IN-RODENTS.pdf Warren et al. 1978] - The development of the delayed hypersensitivity reaction to myelin basic protein and to tuberculin is also suppressed by 02 therapy indicating that its effects upon autoimmune encephalomyelitis involves fundamental alterations of the cellular components of the immune response, some or all of which are reversible.
* https://www.mdpi.com/2227-9059/9/8/943 - Study from Taiwanese researchers.  "semi-therapeutic HBOT significantly alleviated the progression of EAE, at least, via the suppression of Th17 response, the downregulation of CD4 T helper cells expressing GM-CSF or TNF-α, and the boosting of immunomodulatory IL-4 or IL-10-expressed CD4 T cells in the CNS lesions. Conclusively, HBOT attenuated EAE through the modulation of T cell responses in an earlier stage"

=== Tetanus vaccine ===

* https://journals.sagepub.com/doi/10.1177/0961203311429816 - Tetanus vaccine was used to induce antiphospholipid syndrome (APS).  APS was induced with 2 different adjuvants (glycerol, aluminium hydroxide) and different adjuvant pretreatments (glycerol, CFA).

=== Gardasil HPV vaccine, aluminium adjuvant (and phospholipid supplementation) ===

* https://autoimmunity-network.com/articles/Behavioral%20abnormalities%20in%20female%20mice%20following%20HPV%20vac%20%20%202017.pdf  Mice were given aluminium adjuvant only, Gardasil or Gardasil + pertussis toxin.  Forced swimming test 3 months after injection was the most sensitive test.
* https://autoimmunity-network.com/articles/Maria%20Biobody.pdf - Mice vaccinated with Gardasil and aluminium adjuvant only fared worse on a forced swim test.  Phospholipid supplementation can attenuate vaccine-induced depressive-like behavior.

=== Hepatitis B vaccine? ===

* https://sci-hub.se/https://doi.org/10.1016/j.jaut.2014.06.006 - Mice were immunized with Hep-B vaccine, aluminium adjuvant, or the control injection (phosphate-buffered saline).  SLE-disease resulted for the vaccine and alum group.  Immunization with either HBVv or alum affected blood counts, neurocognitive functions and brain gliosis.  Various biomarkers were examined, though measurements had mixed results.

=== Repeated vaccination ===

* https://sci-hub.se/https://doi.org/10.1007/s12026-013-8404-0 - An experimental flock of sheep were given 14 inoculations containing thiomersal, aluminium salt, and ovine pathogens.  Lower weight in the vaccinated group was observed, becoming statistically significant on day 177.

=== Infections that worsen/improve autoimmunity ===

* https://journals.plos.org/plosone/article?id=10.1371/journal.pone.0017849#pone.0017849-Brocke1 - Plasmodium chabaudi (an eukaryote parasite) infection modifies the clinical course of EAE

=== Multiple sclerosis drugs, interferons ===

* [https://www.ncbi.nlm.nih.gov/pmc/articles/PMC3981554/ A review article] on the EAE model for Multiple Sclerosis discusses how all approved MS drugs have some effectiveness in the EAE model.  Several therapies such as glatiramer acetate (copaxone) and natalizumab (Tysabri) were tested first in the mouse model of EAE as part of their successful drug development cycle.

== Metrics / biomarkers of disease severity ==

https://sci-hub.se/https://doi.org/10.1016/j.jaut.2014.06.006
* Forced swim test / immobility time
* Number of rears (standing on rear limbs)
* Stairs climbed
* Antibodies, e.g. ds-DNA
* Autoantibodies
* Weight, organ weight
* Urine protein concentration (SLE model)
* Blood counts
* Novel object recognition (NOR) test 
* Y maze test
* Staircase - number of climbed stairs, rearing events during stair climb

https://journals.sagepub.com/doi/10.1177/0961203311429553 - 3X injection of CFA
* Anti-dsDNA
* Urine / proteinuria

AMAN (GBS variant) model - https://sci-hub.se/https://doi.org/10.1016/j.neulet.2004.06.059
* Abnormal posture
* Neck weakness
* Slip of forelimbs
* Slip of hindlimbs
* Dragging forelimbs
* Dragging hindlimbs
* Unable to walk
* Slow righting reflex
* No righting reflex
* Ptosis
* Tremor of limbs or trunk induced by pulling tail
* Spontaneous tremor of limbs or trunk
* Animal appeared systemically unwell

= Priority research questions =

* Are existing therapies effective?
* What repurposed drugs are effective for treating long haul?
* Is there some way to avoid long-lasting worsening from the best treatments?  Is there some test that can predict serious adverse reactions to treatment?
* Are drug combinations useful for treating long haul?

* Do humanized ACE2 receptors matter in the animal model?

=== Research questions that matter less than they first appear ===

* Are there treatments that are effective in animal models but not in humans?  →  The practical workaround is to first test repurposed drugs with excellent safety profiles.  If these drugs fail in humans, the downside is very low.

=== Less important research questions ===

* Do all the spike protein vaccines cause vaccine injury in animals?
* Does the animal model exhibit stereotypical symptoms such as fatigue and post exertional malaise?
* Why are some people non-responders to treatment?
* Does immunosuppression cause long-term harm?
* Do COVID vaccines cause stillbirths?
* Is the same animal model also viable for Long COVID research?  (The advantage of vaccine-induced 'Long COVID' is that no biosafety level 3 lab would be required, saving money.  However, obtaining mRNA vaccines may be difficult.)
* Do chronic illness patients react badly to other vaccinations (e.g. flu), contrast dye, etc.?