COVID-19 Causes Genetic Changes that Create Long-Haul COVID-19 Symptoms, Researchers Find

Gene changes caused by SARS-CoV-2 spike proteins provide a potential answer for what causes long-haul COVID-19, Texas researchers find 

As COVID-19 vaccines become widely available and cases of COVID-19 in the United States begin to drop, the medical community is beginning to focus more on the long-term effects of COVID-19. Sometimes called “long-haul COVID-19,” the varied and long-term effects that a SARS-CoV-2 infection can create are just starting to be understood (See Long-Haul COVID-19 Emerges as a Concern, Potentially Increasing Need for More COVID-19 Antibody Testing).

New research out of Texas Tech University provides a possible explanation for why these symptoms occur. Led by Sharilyn Almodovar, PhD, at the Texas Tech University Health Sciences Center, researchers found that exposing airway cells to the spike protein of the SARS-CoV-2 virus caused genetic changes.

Potential Impact of Exposure to SARS-CoV-2 Spike Protein Alone

“We found that exposure to the SARS-CoV-2 spike protein alone was enough to change baseline gene expression in airway cells,” said Nicholas Evans, a master’s student at the Texas Tech University Health Sciences Center and one of the researchers involved in the study. “This suggests that symptoms seen in patients may initially result from the spike protein interacting with the cells directly.”

This finding that changes in gene expression occur with exposure to the spike protein of SARS-CoV-2 provides a possible explanation of what causes the mysterious, unexplainable symptoms of long-haul COVID-19 that vary from patient to patient. Changes in gene expression can have different effects on different patients, depending on their genetic makeup and their exposure to the virus.

“Our work helps to elucidate changes occurring in patients on the genetic level, which could eventually provide insight into which treatments would work best for specific patients,” says Evans.

Research May Eventually Lead to Clues About Unexplained Illnesses

While the scientific community’s understanding of long-haul COVID-19 is still quite nascent, Almodovar’s team’s findings are one of the first findings in studying long-haul COVID-19 that provide a good explanation for what could potentially be the cause of these symptoms. This new research will undoubtedly lead to further research examining more in-depth which genes are affected and to what extent this impacts different individuals. These findings may also have implications for other previously unexplained illnesses, such as the long-term effects of Lyme disease.

One interesting result of the these findings is that they may explain why some people with long-haul COVID-19 symptoms have relief of their symptoms after getting vaccinated against COVID-19. Authorized COVID-19 vaccines are designed to use human cells to manufacture spike proteins and stimulate immunity. The finding that the spike protein of SARS-CoV-2 may cause long-haul COVID-19 symptoms could explain how vaccines that artificially create a form of the spike protein could cause these symptoms to change.

COVID-19 Vaccine Questions, Further Studies

Another question that this finding raises is if COVID-19 vaccines, which artificially create SARS-CoV-2 spike proteins, could also stimulate changes in gene expression, causing symptoms that mimic long-haul COVID-19 symptoms.

While Almodovar’s team’s research is only the beginning of study into the possibility of gene expression changes driving long-haul COVID-19 symptoms, it may become a foundational concept in this area of research.

Understanding the implications and effects of long-haul COVID-19 will be important for clinical laboratories that provide COVID-19 antibody testing. As the medical community’s understanding of long-haul COVID-19 increases, it may not only increase the demand for serology tests but may also create a demand for other related tests, particularly immunologically-related tests.

Clinical laboratories will benefit from keeping abreast of long-haul COVID-19 related research and being aware of developments that affect how testing will support clinical treatments and outcomes.

Long-Haulers: Spike Protein Present on Covid And in Vaccines Dangerously Modifies Genes Even After Exposure – Texas Tech Uni Study

study by the Texas Tech University published in Biochemistry and Molecular Biology section of the FASEB journal, has found that spike proteins which Covid-19 uses to penetrate cells and is also supplied in abundance by vaccines to stimulate the body’s immune response, modify the body’s genes.

The study coordinators, Nicholas Evans et al. found that even after exposure, these proteins stimulate continued gene expression and inflammatory events that may be behind the long haul Covid and post vaccine syndromes.

Quoting from a review of the study as we published earlier:

“Results from a new cell study at Texas Tech University Health Sciences Center, US, suggest that the SARS-CoV-2 Spike (S) protein can bring about long-term gene expression changes. The findings could help explain why some COVID-19 patients experience symptoms such as shortness of breath and dizziness long after clearing the infection, a condition known as long COVID.

“We found that exposure to the SARS-CoV-2 S protein alone was enough to change baseline gene expression in airway cells,” said Nicholas Evans, one of the researchers. “This suggests that symptoms seen in patients may initially result from the S protein interacting with the cells directly.””

The study is titled “Lung Time No See”: SARS-Cov-2 Spike Protein Changes Genetic Expression in Human Primary Bronchial Epithelial Cells After Recovery” and is published in the Biochemistry and Molecular Biology section of the FASEB journal.

Long Haul Covid and Long Haul Post-Vaccine Syndromes

Long haul covid or Long-COVID or COVID long-haulers according to a new review can present with as many as 55 long term symptoms. The most common of which are “fatigue (58%), headache (44%), attention disorder (27%), hair loss (25%), and dyspnea (24%)…Diseases such as stroke and diabetes mellitus were also present.” Psychiatric problems like dementia and insomnia are also included. Smell and taste deficiency may persist as also cough and lung abnormalities. Autoimmune problems where the body fights itself is also part of this plethora of presentations. Weight loss, palpitations, renal failure, mood disorders, throat pain and sputum, myocarditis, arrhythmia, OCD, intermittent fever, digestive problems are some more.

These same symptoms as well as symptoms of acute covid infection are now reported months after being vaccinated according to Dr. Bruce Patterson, a pioneer in figuring out Covid and Long haul syndromes.

The precise cause of Long-COVID and Long-Post-Vaccine is being investigated but it may be due to organ damage or persistent autoimmune or inflammatory damage after the infection. Another recent study found Epstein Bar virus reactivated in 73% of long haulers and blamed this for the chronic fatigue, raynaud’s phenomenon and other related symptoms in long haulers.

“We found over 73 percent of COVID-19 patients who were experiencing long COVID symptoms were also positive for EBV reactivation.”

The long-lasting gene changes that result from spike proteins as discovered by Texas Tech University may well be the culprits in these prolonged syndromes.

Watch Dr. Patterson Discuss Long Haul Post-Vaccine Syndrome on Dr. Bean

Video culled from @ 20mins

Dr. Thomas E. Levy, MD, JD writes in OrthoMolecular that “depending on the cell types to which such spike proteins bind, a wide variety of diseases with autoimmune qualities can result.” He recommends treating both syndromes the same way with Vitamin C, Ivermectin, Quercetin and other agents.

“long-haul COVID syndrome likely represents a low-grade unresolved smoldering COVID infection with the same kind of spike protein persistence and clinical impact as is seen in many individuals after their COVID vaccinations (Mendelson et al., 2020; Aucott and Rebman, 2021; Raveendran, 2021).”

He further postulates an effect of the spike protein on the ACE2 receptor which has roles in essential pathways that protect blood vessels and other vital physiological processes.

“By itself, the disruption of ACE2 receptor function in so many areas of the body has resulted in an array of different side effects (Ashraf et al., 2021).” Dr Levy writes.

Salk researchers earlier in April found that the spike protein was dangerous to blood vessels by itself, their investigations “proving that the spike protein alone was enough to cause disease.”

Texas Tech University described the dangerous gene-modifying effects of the spike protein as being long-lived and leading to genetic inflammatory changes even after exposure.

“The researchers found that cultured human airway cells exposed to both low and high concentrations of purified S protein showed differences in gene expression that remained even after the cells recovered from the exposure. The top genes included ones related to inflammatory response.”

The study researcher, Nicholas Evans, et al. Concluded:

“Our preliminary results suggest that the SARS-CoV-2 spike protein is enough to change the baseline protein expression in primary HBECs. After recovery, genes related to immune response retained changes in gene expression, and these may indicate relevant long-term effects in asymptomatic patients. Additionally, the interplay between immune response and other pathways after SARS-CoV-2 spike protein exposure should be investigated in the future.”

NewsRescue experts opine: “It is plausible that candidates with lower immunities face more adverse reactions and are on greater risk of Long haul post-vaccine syndrome because it takes longer for their immune system to kick off and wash out the spike proteins produced in the body post vaccine. Studies should investigate any relationship between Long haul post-vaccine and immune compromise.

“Of the various vaccines, AstraZeneca distributed in Africa and the rest of the poorer third world is the worst for many reasons including the fact that it delivers the wild-type unmodified spike protein which has the capacity to transform to the ‘post-fusion’ state and enter the cell and as such has the potential to cause more gene modification or other side effects.”

Animal studies have found toxic effects of spike proteins.

“…those animal studies where Spike protein was produced by a pseudovirus, or the S1 subunit was administered directly. Both of these caused pathology in the animals all by itself, without coronavirus itself being present.”

Vaccine Technology and the Spike Protein

All current vaccines introduce the spike protein in the recipient. Oxford/AstraZeneca and Janssen (Johnson and Johnson) use ‘vectors’, J&J uses adenovirus type 26 (Ad26) as its vector, while AstraZeneca uses a chimpanzee adenovirus to introduce the spike protein. Moderna, Pfizer/BioNTech vaccines introduce mRNA which is a sort of pre-protein. This mRNA is then read by your cells and translated into S1 and modified spike proteins.

While Oxford/AZ introduces the same exact spike protein the virus uses, Moderna, Pfizer/BioNTech and J&J make the body produce a modified spike protein which has the addition of two amino acids (these are the building blocks of proteins), which modify the spike protein so it stays stuck in the ‘pre-fusion’ state and does not switch to the ‘post-fusion’ state which can penetrate the cell.

– Read more:

Certainly more studies shall continue to help humanity deal with these complex new long haul syndromes and to completely reverse the effects of the possible gene modification as reported.

Gene changes and long-haul COVID

Authors: Nancy D. Lamontagne April 30, 2021

Airway cells exposed to SARS-CoV-2 spike protein exhibited persisting changes in gene expression

Results from a new cell study suggest that the SARS-CoV-2 spike protein can bring about long-term gene expression changes. The findings could help explain why some COVID-19 patients—referred to as COVID long-haulers—experience symptoms such as shortness of breath and dizziness long after clearing the infection.

SARS-CoV-2, the virus that causes COVID-19, is covered in tiny spike proteins. During infection, the spike proteins bind with receptors on cells in our body, starting a process that allows the virus to release its genetic material into the inside of the healthy cell. COURTESY OF JULIE A. FORREST Research team members included undergraduate student Ethan Salazar, principal investigator Sharilyn Almodovar and master’s student Nicholas Evans.

“We found that exposure to the SARS-CoV-2 spike protein alone was enough to change baseline gene expression in airway cells,” said Nicholas Evans, a master’s student in the laboratory of Sharilyn Almodovar, PhD, at the Texas Tech University Health Sciences Center. “This suggests that symptoms seen in patients may initially result from the spike protein interacting with the cells directly.”

Evans will present the research at the American Society for Biochemistry and Molecular Biology annual meeting during the virtual Experimental Biology 2021 meeting, to be held April 27–30.

Culturing human airway cells requires specific conditions that allow cells to mature into the differentiated cells that would be found in the airway. The researchers optimized a previously developed culturing approach known as the air–liquid interface technique so that it would more closely simulate the physiological conditions found in the lung airway. This involved exposing cells to air and then giving them time to mature into airway cells.

The researchers found that cultured human airway cells exposed to both low and high concentrations of purified spike protein showed differences in gene expression that remained even after the cells recovered from the exposure. The top genes included ones related to inflammatory response.

“Our work helps to elucidate changes occurring in patients on the genetic level, which could eventually provide insight into which treatments would work best for specific patients,” said Evans.

The researchers also compared their cultured human airway cells to studies from others where cells were collected from patients with COVID-19 infection. They were able to confirm that the optimized cell culture approach reflected what occurs in patients, making it useful for future translational studies. They plan to use the new approach to better understand how long the genetic changes last and the potential long-term consequences of these changes in relation to long-haul COVID-19 cases.

Antigen Presentation of mRNA-Based and Virus-Vectored SARS-CoV-2 Vaccines

Authors: Ger T. Rijkers,1,2,* Nynke Weterings,1 Andres Obregon-Henao,3 Michaëla Lepolder,1 Taru S. Dutt,3 Frans J. van Overveld,1 and Marcela Henao-Tamayo3


Infection with Severe Acute Respiratory Syndrome Coronavirus 2 (SARS-CoV-2) causes Coronavirus Disease 2019 (COVID-19), which has reached pandemic proportions. A number of effective vaccines have been produced, including mRNA vaccines and viral vector vaccines, which are now being implemented on a large scale in order to control the pandemic. The mRNA vaccines are composed of viral Spike S1 protein encoding mRNA incorporated in a lipid nanoparticle and stabilized by polyethylene glycol (PEG). The mRNA vaccines are novel in many respects, including cellular uptake and the intracellular routing, processing, and secretion of the viral protein. Viral vector vaccines have incorporated DNA sequences, encoding the SARS-CoV-2 Spike protein into (attenuated) adenoviruses. The antigen presentation routes in MHC class I and class II, in relation to the induction of virus-neutralizing antibodies and cytotoxic T-lymphocytes, will be reviewed. In rare cases, mRNA vaccines induce unwanted immune mediated side effects. The mRNA-based vaccines may lead to an anaphylactic reaction. This reaction may be triggered by PEG. The intracellular routing of PEG and potential presentation in the context of CD1 will be discussed. Adenovirus vector-based vaccines have been associated with thrombocytopenic thrombosis events. The anti-platelet factor 4 antibodies found in these patients could be generated due to conformational changes of relevant epitopes presented to the immune system.

1. Introduction

The high morbidity and mortality rate of coronavirus disease of 2019 (COVID-19) has triggered the rapid development of vaccines against its causative agent, Severe Acute Respiratory Syndrome Coronavirus 2 (SARS-CoV-2). Vaccines are the most effective way to eliminate and control the virus [1,2]. Most of the vaccines developed for COVID-19 have shown very high levels of protection. Within one year after the outbreak of the pandemic and identification of the genomic structure of SARS-CoV-2, a number of highly effective vaccines were approved and used globally, as over 2.5 billion vaccine doses have been administered [3] (dated 25 June 2021; World Health Organization). The two major categories of SARS-CoV-2 vaccines are mRNA-based vaccines and viral vector vaccines, both targeting the Spike protein of the virus [4]. Worldwide, the most used mRNA vaccines are those of Pfizer/BioNTech (BNT162b2, brand name Comirnaty) and of Moderna (mRNA-1273, brand name COVID-19 Vaccine Moderna). The most-used adenovirus vector vaccines are the ones of Oxford/AstraZeneca (ChAdOx1 nCoV-19, brand names Vaxzevria and Covishield) and Jansen/Johnson and Johnson (Ad26.COV2.S, brand name Janssen COVID-19 Vaccine), as well as the Sputnik-V and CanSino vaccines.

Both mRNA vaccines for SARS-CoV-2 as well as viral vector based vaccines have turned out to be highly effective for protection against mild and severe COVID-19 cases. After vaccination, high titers of IgG and IgA antibodies against the Spike protein are generated which, in vitro, show a virus neutralizing capacity, and cytotoxic T cells are activated [5,6,7].

The aim of this review is to delineate the molecular pathways, outside and inside of the cell, which ultimately lead to the presentation of Spike peptides to the immune system. Both the classical antigen presentation routes via MHC class I to CD8+ T cells and via MHC class II to CD4+ T cells, as well as the antigen-presenting routes for presentation to non-conventional T cells, will be reviewed and discussed.

While SARS-CoV-2 vaccines are protecting from the severe illness and deaths due to COVID-19, after large-scale implementation, rare immune-mediated side effects became apparent. In particular, anaphylactic reactions and various thrombotic or abnormal bleeding have raised concern [8,9]. These side effects may be due to abnormal handling or presentation of the vaccine or vaccine additives to the immune system, of which the potential scenarios will be discussed.

For A Detailed Explanation:

SARS-CoV-2 Spike Protein Impairs Endothelial Function via Downregulation of ACE 2

Authors: Yuyang Lei,1,2,*Jiao Zhang,1,2,5,*Cara R. Schiavon,8,9Ming He,5Lili Chen,2Hui Shen,5,10Yichi Zhang,5Qian Yin,2Yoshitake Cho,5Leonardo Andrade,8Gerald S. Shadel,9Mark Hepokoski,6Ting Lei,3Hongliang Wang,4Jin Zhang,7Jason X.-J. Yuan,6Atul Malhotra,6Uri Manor,8,†Shengpeng Wang,2,†Zu-Yi Yuan,1,† and John Y-J. Shyy5,†

SARS-CoV-2 (severe acute respiratory syndrome coronavirus 2) infection relies on the binding of S protein (Spike glycoprotein) to ACE (angiotensin-converting enzyme) 2 in the host cells. Vascular endothelium can be infected by SARS-CoV-2,1 which triggers mitochondrial reactive oxygen species production and glycolytic shift.2 Paradoxically, ACE2 is protective in the cardiovascular system, and SARS-CoV-1 S protein promotes lung injury by decreasing the level of ACE2 in the infected lungs.3 In the current study, we show that S protein alone can damage vascular endothelial cells (ECs) by downregulating ACE2 and consequently inhibiting mitochondrial function.

We administered a pseudovirus expressing S protein (Pseu-Spike) to Syrian hamsters intratracheally. Lung damage was apparent in animals receiving Pseu-Spike, revealed by thickening of the alveolar septa and increased infiltration of mononuclear cells (Figure [A]). AMPK (AMP-activated protein kinase) phosphorylates ACE2 Ser-680, MDM2 (murine double minute 2) ubiquitinates ACE2 Lys-788, and crosstalk between AMPK and MDM2 determines the ACE2 level.4 In the damaged lungs, levels of pAMPK (phospho-AMPK), pACE2 (phospho-ACE2), and ACE2 decreased but those of MDM2 increased (Figure [B], i). Furthermore, complementary increased and decreased phosphorylation of eNOS (endothelial NO synthase) Thr-494 and Ser-1176 indicated impaired eNOS activity. These changes of pACE2, ACE2, MDM2 expression, and AMPK activity in endothelium were recapitulated by in vitro experiments using pulmonary arterial ECs infected with Pseu-Spike which was rescued by treatment with N-acetyl-L-cysteine, a reactive oxygen species inhibitor (Figure [B], ii).Open in a separate windowFigure.

SARS-CoV-2 (Severe acute respiratory syndrome coronavirus 2) Spike protein exacerbates endothelial cell (EC) function via ACE (angiotensin-converting enzyme) 2 downregulation and mitochondrial impairment. A, Representative H&E histopathology of lung specimens from 8- to 12 wk-old male Syrian hamsters 5-day post administration of pseudovirus overexpressing Spike protein (Pseu-Spike) or mock virus in control group (n=3 mice per group, 1×108 PFU). Thickened alveolar septa (red arrowhead) and mononuclear cell (red arrow). Scale bar=20 μm. B, Pseu-Spike (n=4) or mock virus (n=4)–infected hamster lungs were subjected to Western blot analysis for pAMPK (phospho-AMPK) T172, AMPK, pACE2 (phospho angiotensin-converting enzyme) S680, ACE 2, MDM2, peNOS S1176, peNOS T494, eNOS (endothelial NO synthase), and β-actin (B, i). Human pulmonary arterial EC (PAECs) were infected with Pseu-Spike or mock virus for 24 h with or without N-acetyl-L-cysteine (NAC; 5 mmol/L) pretreatment for 2 h. The protein extracts were analyzed by Western blot using antibodies against proteins as indicated (n=4; B, ii). C, Representative confocal images of mitochondrial morphology of ECs treated with human recombinant S1 protein or IgG (4 μg/mL) for 24 h (C, i) or infected with human adenovirus ACE2 S680D (ACE2-D) or ACE2 S680L (ACE2-L; 10 MOI) for 48 h (C, ii). Mitochondria were visualized using TOM20 antibody (n=4, 50 cells counted for each replicate). Scale bar=2.5 μm. Tubular: the majority of mitochondria in ECs was >10 μm in length; Intermediate: the mitochondria were <≈10 μm; Fragment: the majority of mitochondria were spherical (no clear length or width). D, Measurement of oxygen consumption rate (OCR, D, i and iii) and extracellular acidification rate (ECAR, D, ii and iv) in ECs infected with ACE2-D vs ACE2-L (10 MOI) for 48 h (n=3) or treated with IgG vs S1 protein (4 μg/mL) for 24 h (n=3). E, Real-time quantitative polymerase chain reaction analysis of the indicated mRNA levels in lung ECs from ACE2-D (n=4) and ACE2-L (n=4) knock-in mice. Eight-week-old ACE2-D and ACE2-L male mice with C57BL/6 background were used. F, Dose-response curves of acetylcholine (ACh, left)- and sodium nitroprusside (SNP, right)–mediated relaxation on the tension of phenylephrine (1 μmol/L) precontracted intrapulmonary artery stripes from Pseu-Spike-(ACh n=8, SNP n=5) or mock (ACh n=6, SNP n=5) virus–infected Syrian hamsters (1×108 PFU; F, i) and ACE2-D (n=6) or ACE2-L (n=5) mice (F, ii). The animal experiments were approved by the ethical committee of Xi’an Jiaotong University. 2-DG indicates 2-Deoxy-D-glucose; ACE2-D, a phospho-mimetic ACE2 with increased stability; ACE2-L, a dephospho-mimetic ACE2 with decreased stability; AMPK, AMP-activated protein kinase; AA/R, antimycin A&Rotenone; ENO2, enolase 2; FCCP, carbonyl cyanide-p-(trifluoromethoxy)phenylhydrazone; H&E, Hematoxylin and Eosin; HK2, hexokinase 2; HO1, heme oxygenase-1; MDM2, murine double minute 2; MOI, multiplicity of infection; NRF1, nuclear respiratory factor 1; peNOS, phospho-eNOS; PFKFB3, 6-phosphofructo-2-kinase/fructose-2,6-biphosphatase 3; Resp, respiration; and TFAM, transcription factor A, mitochondrial.

We next studied the impact of S protein on mitochondrial function. Confocal images of ECs treated with S1 protein revealed increased mitochondrial fragmentation, indicating altered mitochondrial dynamics (Figure [C], i). To examine whether these mitochondrial changes were due, in part, to the decreased amount of ACE2, we overexpressed ACE2 S680D (ACE2-D, a phospho-mimetic ACE2 with increased stability) or S680L (ACE2-L, a dephospho-mimetic with decreased stability)4 in ECs. As shown in Figure [C], ii, ECs with ACE2-L had a higher number of fragmented mitochondria when compared to those with ACE2-D. Performing oxygen consumption rate and extracellular acidification rate assays, we found that ECs overexpressing ACE2-L had reduced basal mitochondrial respiration, ATP production, and maximal respiration compared to ECs overexpressing ACE2-D (Figure [D], i). Moreover, ACE2-L overexpression caused increased basal acidification rate, glucose-induced glycolysis, maximal glycolytic capacity, and glycolytic reserve (Figure [D], ii). Also, ECs incubated with S1 protein had attenuated mitochondrial function but increased glycolysis, when compared with control cells treated with IgG (Figure [D], iii and iv). We also compared the expressions of mitochondria- and glycolysis-related genes in lung ECs isolated from ACE2-D or ACE2-L knock-in mice.4 Shown in Figure [E], the mRNA levels of NRF1HO1, and TFAM (mitochondria biogenesis-related genes) were increased, whereas those of HK2PFKFB3, and ENO2 (glycolysis-related genes) were decreased in lung ECs in ACE2-D mice, as compared to those in ACE2-L mice.

SARS-CoV-2 infection induces EC inflammation, leading to endotheliitis.1,5 Because S protein decreased ACE2 level and impaired NO bioavailability, we examined whether S protein entry is indispensable for dysfunctional endothelium. As shown in Figure [F], i, the endothelium-dependent vasodilation induced by acetylcholine was impaired in pulmonary arteries isolated from Pseu-Spike-administered hamsters, whereas the endothelium-independent vasodilation induced by sodium nitroprusside was not affected. We also compared the acetylcholine- and sodium nitroprusside–induced vasodilation of pulmonary vessels from ACE2-D or ACE2-L mice. As anticipated, acetylcholine-induced vasodilation was hindered in pulmonary arteries isolated from ACE2-L mice in comparison to ACE2-D mice (Figure [F], ii). There was, however, little difference in sodium nitroprusside–induced vasodilation between ACE2-D and ACE-L animals.

Although the use of a noninfectious pseudovirus is a limitation to this study, our data reveals that S protein alone can damage endothelium, manifested by impaired mitochondrial function and eNOS activity but increased glycolysis. It appears that S protein in ECs increases redox stress which may lead to AMPK deactivation, MDM2 upregulation, and ultimately ACE2 destabilization.4 Although these findings need to be confirmed with the SARS-CoV-2 virus in the future study, it seems paradoxical that ACE2 reduction by S protein would decrease the virus infectivity, thereby protecting endothelium. However, a dysregulated renin-angiotensin system due to ACE2 reduction may exacerbate endothelial dysfunction, leading to endotheliitis. Collectively, our results suggest that the S protein-exerted EC damage overrides the decreased virus infectivity. This conclusion suggests that vaccination-generated antibody and/or exogenous antibody against S protein not only protects the host from SARS-CoV-2 infectivity but also inhibits S protein-imposed endothelial injury.

For More Information:

Something Really Strange Is Happening At Hospitals All Over America

Authors: Authored by Michael Snyder via, TURSDAY, NOV 04, 2021 – 05:11

In a year that has been filled with so many mysteries already, I have another very odd one to share with you.  Emergency rooms are filled to overflowing all over America, and nobody can seem to explain why this is happening.  Right now, the number of new COVID cases in the United States each day is less than half of what it was just a couple of months ago.  That is really good news, and many believe that this is a sign that the pandemic is fading.  Let us hope that is true.  With less people catching the virus, you would think that would mean that our emergency rooms should be emptying out, but the opposite is actually happening.  All across the country, emergency rooms are absolutely packed, and in many cases we are seeing seriously ill patients being cared for in the hallways because all of the ER rooms are already full.

Let me give you an example of what I am talking about.  The following comes from an article entitled “ERs Are Swamped With Seriously Ill Patients, Although Many Don’t Have Covid”

Inside the emergency department at Sparrow Hospital in Lansing, Michigan, staff members are struggling to care for patients showing up much sicker than they’ve ever seen.

Tiffani Dusang, the ER’s nursing director, practically vibrates with pent-up anxiety, looking at patients lying on a long line of stretchers pushed up against the beige walls of the hospital hallways. “It’s hard to watch,” she said in a warm Texas twang.

But there’s nothing she can do. The ER’s 72 rooms are already filled.

Can anyone explain why this is happening?

If the number of COVID cases was starting to spike again, it would make sense for emergency rooms to be overflowing.

But at this particular hospital in Michigan, we are being told that some of the main things that are being treated include “abdominal pain”, “respiratory problems”, “blood clots” and “heart conditions”

Months of treatment delays have exacerbated chronic conditions and worsened symptoms. Doctors and nurses say the severity of illness ranges widely and includes abdominal pain, respiratory problems, blood clots, heart conditions and suicide attempts, among other conditions.

That mention of “heart conditions” immediately got my attention, because I have been seeing this so much in the news recently.

For instance, a high school senior in Pennsylvania just dropped dead from “a sudden cardiac incident”

The high school soccer manager ‘greatly enjoyed’ his team’s championship victory Saturday. Later that evening, he was dead.

Now, late student Blake Barklage’s high school is mourning his untimely death. As 6ABC in Philly reports, the tragedy occurred at La Salle College High School in Montgomery County, Pa.

In a letter to parents, the school announced that the senior died after ‘a sudden cardiac incident’ Saturday night.

Elsewhere in the same state, an otherwise healthy 12-year-old boy just suddenly died because of an issue with his coronary artery…

As family and friends grieve, the cause of death is in for a 12-year-old taken way too soon while warming up for school basketball practice.

As TribLive in Pittsburgh reports, Jayson Kidd, 12, of Bridgeville, Pa., died of natural causes involving his coronary artery, according to the Allegheny County Medical Examiner’s Office.

Heart problems kill elderly people all the time, but it is odd that so many healthy young people have been having these problems.

Over the weekend, Barcelona striker Sergio Aguero suddenly collapsed on the pitch during a match.

He was later diagnosed with “a cardiac arrhythmia”

Sergio “Kun” Aguero, a striker for the Barcelona soccer team, has been diagnosed with a cardiac arrhythmia after collapsing during Saturday’s match against Alaves.

The 33-year-old Argentinian was examined by medical staff at the stadium before being taken to a nearby hospital where he is still waiting to undergo further examination.

Just two days later, a match in Norway was brought to a screeching halt after a player experienced “cardiac arrest” right in the middle of a match…

A football match in Norway’s second division was halted on Monday after Icelandic midfielder Emil Pálsson suffered a cardiac arrest during play.

The 28-year-old Sogndal player suffered the attack as the game against Stjordals-Blink entered the 12th minute, his club said in a statement.

I have been seeing so many stories like this.

So why are so many young people suddenly having such serious problems with their hearts?

Can anyone out there explain this to me?

Link between fever, diarrhea, severe COVID-19, and persistent anti-SARS-CoV-2 antibodies

Authors: By Dr. Liji Thomas, MD Jan 7 2021

Ever since the coronavirus disease 2019 (COVID-19) pandemic began, there have been many attempts to understand the nature and duration of immunity against the causative agent, the severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2).

A new preprint research paper appearing on the medRxiv* server describes a link between the persistence of neutralizing antibodies against the virus, disease severity, and specific COVID-19 symptoms.

Permanent immunity is essential if the pandemic is to end. In the earlier SARS epidemic, antibodies were found to last for three or more years after infection in most patients. With the current virus, it may last for six or more months at least, as appears from some reports. Other researchers have concluded that immunity wanes rapidly over the same period, with some patients who were tested positive for antibodies becoming seronegative later on. This discrepancy may be traceable to variation in testing methods, sample sizes and testing time points, as well as disease severity.

Study details

The current study looked at a population of over a hundred convalescent COVID-19 patients, testing most of them for antibodies at five weeks and three months from symptom resolution.

The researchers used a multiplex assay that measured the Immunoglobulin G (IgG) levels against four SARS-CoV-2 antigens, one from SARS-CoV, and four from circulating seasonal coronaviruses. In addition, they carried out an inhibition assay against SARS-CoV-2 spike receptor-binding domain (RBD)-angiotensin-converting enzyme 2 (ACE2) binding and a neutralization assay against the virus. The antibody titers were then plotted against various clinical features and demographic factors.

Antibody titers higher in COVID-19 convalescents

The researchers found that severe disease is correlated with advanced age and the male sex. Patients with underlying vascular disease were more likely to be hospitalized with COVID-19, but those with asthma were relatively spared.

Convalescent COVID-19 patients had higher IgG levels against all four SARS-CoV-2 antigens, relative to controls, and in 98% of cases, at least one of the tests was likely to show higher binding compared to controls. IgGs targeting the viral spike and RBD were likely to be much more discriminatory between SARS-CoV-2 patients and controls. Interestingly, anti-SARS-CoV IgG, as well as anti-seasonal betacoronavirus antibodies, were likely to be higher in these patients.

Anti-spike and anti-nucleocapsid IgG levels, as well as neutralizing antibody titers, were higher in convalescent hospitalized COVID-19 patients than in convalescent non-hospitalized patients, and the titers were positively associated with disease severity.Antibodies against SARS-CoV-2 persist three months after COVID-19 symptom resolution. Sera from COVID-19 convalescent subjects (n=79) collected 5 weeks (w) and 3 months (m) after symptom resolution were subjected to multiplex assay to detect IgG that binds to SARS-CoV-2 S, NTD, RBD and N antigens (A), to RBD-ACE2 binding inhibition assay (B), and to SARS-CoV-2 neutralization assay (C). Dots, lines, and asterisks in red represent non-hospitalized (n=67) and in blue represent hospitalized (n=12) subjects with lines connecting the two time points for individual subjects (*p<0.05 and **p<0.01 by paired t test).Antibodies against SARS-CoV-2 persist three months after COVID-19 symptom resolution. Sera from COVID-19 convalescent subjects (n=79) collected 5 weeks (w) and 3 months (m) after symptom resolution were subjected to multiplex assay to detect IgG that binds to SARS-CoV-2 S, NTD, RBD and N antigens (A), to RBD-ACE2 binding inhibition assay (B), and to SARS-CoV-2 neutralization assay (C). Dots, lines, and asterisks in red represent non-hospitalized (n=67) and in blue represent hospitalized (n=12) subjects with lines connecting the two time points for individual subjects (*p<0.05 and **p<0.01 by paired t test).

Clinical correlates of higher antibody titer

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When antibody titers in non-hospitalized subjects were compared with clinical and demographic variables, they found that older males with a higher body mass index (BMI) and a Charlson Comorbidity Index score >2 were likely to have higher antibody titers. COVID-19 symptoms that correlated with higher antibody levels in these patients comprise fever, diarrhea, abdominal pain and loss of appetite. Chest tightening, headache and sore throat were associated with less severe symptoms.

The link between the specific symptoms listed above with higher antibody titers could indicate that they mark a robust systemic inflammatory response, which in turn is necessary for a strong antibody response. Diarrhea may mark severe disease, but it is strange that in this case, it was not more frequent in the hospitalized cohort. Alternatively, diarrhea may have strengthened the immune antibody response via the exposure of the virus to more immune cells via the damaged enteric mucosa. More study is required to clarify this finding.

Potential substitute for neutralizing assay

The binding assay showed that the convalescent serum at five weeks inhibited RBD-ACE2 binding much more powerfully than control serum. Neutralizing activity was also higher in these sera, but in 15% of cases, convalescent patients showed comparable neutralizing antibody titers to those in control sera. On the whole, however, there was a positive association between neutralizing antibody titer, anti-SARS-CoV-2 IgG titers, and inhibition of ACE2 binding.

Persistent immunity at three months

This study also shows that SARS-CoV-2 antibodies persist in these patients at even three months after symptoms subside, with persistent IgG titers against the SARS-CoV-2 spike, RBD, nucleocapsid and N-terminal domain antigens. Binding and neutralization assays remained highly inhibitory throughout this period. The same was true of antibodies against the other coronaviruses tested as well, an effect that has been seen with other viruses and could be the result of cross-reactive anti-SARS-CoV-2 antibodies. Alternatively, it could be due to the activation of memory B cells formed in response to infection by the seasonal beta-coronaviruses.


IgG titers, particularly against S and RBD, and RBD-ACE2 binding inhibition better differentiate between COVID-19 convalescent and naive individuals than the neutralizing assay,” the researchers concluded.

These could be combined into a single diagnostic test, they suggest, with extreme sensitivity and specificity. The correlation with neutralizing antibody titers could indicate that the neutralizing assay, which is more expensive, sophisticated and expensive, as well as more dangerous for the investigators, could be replaced by the other antibody tests without loss of value.

In short, the study shows that specific antibodies persist for three months at least following recovery; antibody titers correlate with COVID-19-related fever, loss of appetite, abdominal pain and diarrhea; and are also higher in older males with more severe disease, a higher BMI and CCI above 2. Further research would help understand the lowest protective titer that prevents reinfection, and the duration of immunity.

*Important Notice

medRxiv publishes preliminary scientific reports that are not peer-reviewed and, therefore, should not be regarded as conclusive, guide clinical practice/health-related behavior, or treated as established information.Journal reference:

The origin of SARS-CoV-2 furin cleavage site remains a mystery

Authors: By Dr. Liji Thomas, MD Feb 17 2021

The ongoing pandemic of coronavirus disease 2019 (COVID-19), caused by the severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2), has largely defied attempts to contain its spread by non-pharmaceutical interventions (NPIs). With the massive loss of life and economic damage, the only way out, in the absence of specific antiviral therapeutics, has been the development of vaccines to achieve population immunity.

A new study on the Preprints server discusses the origin of the furin cleavage site on the SARS-CoV-2 spike protein, which is responsible for the virus’s relatively high infectivity compared to relatives in the betacoronavirus subgenus.

The furin cleavage site

The SARS-CoV-2 is a betacoronavirus, and is most closely related to the bat SARS-related coronavirus (SARSr-CoV) represented by the genome sequence RaTG13, which shares 96% identity with the former. This has made the bat virus the most probable precursor of the virus in current circulation.

The origin of this strain is linked to the emergence of the novel furin cleavage site in the viral spike glycoprotein. The furin is a serine protease widely expressed in human cells, that cleaves the SARS-CoV-2 spike at the interface of its two subunits. It is encoded by a gene on chromosome 15.

Furin acts on substrates with single or paired basic residues during the processing of proteins within cells. Such a polybasic furin cleavage site is found in various proteins from many viruses, including Betacoronavirus Embecoviruses, and the Merbecovirus. However, within the betacoronaviruses of the sarbecovirus lineage B, this type of site is unique to SARS-CoV-2.

The study used a bioinformatic approach using the genomic data available on the National Center for Biotechnological Information (NCBI) databases, to identify the origin of the furin cleavage site.

Same ancestor

They found three coronaviruses that were very similar to the SARS-CoV-2 at the genomic level. These are Pangolin-CoVs (2017, 2019), Bat-SARS-like (CoVZC45, CoVZXC21) and bat RatG13.

The three genomic fingerprints used to identify these matches include fingerprint 1, in the orf1a RNA polymerase gene, including the nsp2 and nsp3 genes; fingerprint 2, at the beginning of S gene, covering the part encoding the N-terminal domain and the receptor-binding domain (RBD) that mediates attachment to the host cell receptor, the angiotensin-converting enzyme 2 (ACE2).; and fingerprint 3, the orf8 gene.

These fingerprints are distinctive to the three closely related coronaviruses only at the RNA level, but the amino acid sequences in the translated proteins are similar to other sarbecoviruses.

The sharing of these genomic sequences indicates their common ancestry, supported by other short sequence features, with one deletion and three insertions. All three strains show the same deletion-insertion pattern at the same four different locations in the spike gene.

Spike gene recombination in a common ancestor

The analysis of the phylogeny of these three strains showed that the first to diverge was the pangolin coronavirus, with the RatG13 being the closest. However, when only the spike is analyzed, there is a high similarity between the pangolin CoV, RaTG13 and SARS-CoV-2.

This may indicate the occurrence of recombination events between the Pangolin-CoV (2017) and RatG13 ancestors. This was followed by the shift of the pangolin CoV to pangolin hosts.Phylogenetic tree of the closely related SARS-CoV-2 coronaviruses based on complete genomesPhylogenetic tree of the closely related SARS-CoV-2 coronaviruses based on complete genomes.

Unique codons encoding arginines in the furin cleavage site

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The furin cleavage site consists of four amino acids PRRA, which are encoded by 12 inserted nucleotides in the S gene. A characteristic feature of this site is an arginine doublet.

This insertion could have occurred by random insertion mutation, recombination or by laboratory insertion. The researchers say the possibility of random insertion is too low to explain the origin of this motif.

Surprisingly, the CGGCGG codons encoding the two arginines of the doublet in SARS-CoV-2 are not found in any of the furin sites in other viral proteins expressed by a wide range of viruses.

Even within the SARS-CoV-2, where arginine is encoded by six codons, only a minority of arginine residues are encoded by the CGG codon. Again, only two of the 42 arginines in the SARS-CoV-2 spike are encoded by this codon – and these are in the PRRA motif.

For recombination to occur, there must be a donor, from another furin site and probably from another virus. In the absence of a known virus containing this arginine doublet encoded by the CGGCGG codons, the researchers discount the recombination theory as the mechanism underlying the emergence of PRRA in SARS-CoV-2.

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Pathophysiology of COVID-19:

Mechanisms Underlying Disease Severity and Progression

Authors: Mary Kathryn Bohn,1,2, Alexandra Hall,1 Lusia Sepiashvili,1,2, Benjamin Jung,1,2 Shannon, Steele,1 and Khosrow Adeli1,2,3

The global epidemiology of coronavirus disease 2019 (COVID-19) suggests a wide spectrum of clinical severity, ranging from asymptomatic to fatal. Although the clinical and laboratory characteristics of COVID-19 patients have been well characterized, the pathophysiological mechanisms underlying disease severity and progression remain unclear. This review highlights key mechanisms that have been proposed to contribute to COVID-19 progression from viral entry to multisystem organ failure, as well as the central role of the immune response in successful viral clearance or progression to death.


Coronavirus disease 2019 (COVID-19) is caused by a novel beta-coronavirus known as Severe Acute Respiratory Syndrome Coronavirus 2 (SARS-CoV-2). As of June 15, 2020, the number of global confirmed cases has surpassed 8 million, with over 400,000 reported mortalities. The unparalleled pathogenicity and global impact of this pandemic has rapidly engaged the scientific community in urgently needed research. Preliminary reports from the Chinese Center for Disease Control and Prevention have estimated that the large majority of confirmed SARS-CoV-2 cases are mild (81%), with ~14% progressing to severe pneumonia and 5% developing acute respiratory distress syndrome (ARDS), sepsis, and/or multisystem organ failure (MOF) (144). Although more data is urgently needed to elucidate the global epidemiology of COVID-19 (80), a wide spectrum of clinical severity is evident, with most patients able to mount a sufficient and appropriate immune response, ultimately leading to viral clearance and case resolution. However, a significant subset of patients present with severe clinical manifestations, requiring life-supporting treatment (51). The pathophysiological mechanisms behind key events in the progression from mild to severe disease remain unclear, warranting further investigation to inform therapeutic decisions. Here, we review the current literature and summarize key proposed mechanisms of COVID-19 pathophysiological progression (FIGURE 1). Key Pathophysiological Mechanisms: Our Current Understanding Viral Invasion The first step in COVID-19 pathogenesis is viral invasion via its target host cell receptors. SARSCoV-2 viral entry has been described in detail elsewhere (138). In brief, SARS-CoV-2 consists of four main structural glycoproteins: spike (S), membrane (M), envelope (E), and nucleocapsid (N). The M, E, and N proteins are critical for viral particle assembly and release, whereas the S protein is responsible for viral binding and entry into host cells (33, 76, 89, 143, 148). Similar to SARS-CoV, several researchers have identified human angiotensin converting enzyme 2 (ACE2) as an entry receptor for SARS-CoV-2 (75, 99, 148, 156). SARSCoV-2 is mostly transmissible through large respiratory droplets, directly infecting cells of the upper and lower respiratory tract, especially nasal ciliated and alveolar epithelial cells (161). In addition to the lungs, ACE2 is also expressed in various other human tissues, such as the small intestine, kidneys, heart, thyroid, testis, and adipose tissue, indicating the virus may directly infect cells of other organ systems when viremia is present (77). Interestingly, although the S proteins of SARS-CoV-2 and SARSCoV share 72% homology in amino acid sequences, SARS-CoV-2 has been reported to have a higher affinity for the ACE2 receptor (18, 21, 143). Following host cell binding, viral and cell membranes fuse, enabling the virus to enter into the cell (89). For many coronaviruses, including SARS-CoV, host cell binding alone is insufficient to facilitate membrane fusion, requiring S-protein priming or cleavage by host cell proteases or transmembrane serine proteases (9, 10, 90, 94, 108). Indeed, Hoffman and colleagues demonstrated that S-protein priming by transmembrane serine protease 2 (TMPRSS2), which may be substituted by cathepsin B/L, is required to facilitate SARS-CoV-2 entry into host cells (58). In addition, unlike other coronaviruses, SARS-CoV-2 has been reported to possess a furin-like cleavage site in the S-protein domain, located between the S1 and S2 subunits (31, 138). Furin-like proteases are ubiquitously expressed, albeit at low levels, indicating that S-protein priming at this cleavage site may contribute to the widened cell tropism and enhanced transmissibility of SARS-CoV-2 (123). However, whether furin-like protease-mediated cleavage is required for SARS-CoV-2 host entry has yet to be determined. Blocking or inhibiting these processing enzymes may serve as a potential antiviral target (130). Interestingly, SARS-CoV-2 has developed a unique S1/S2 cleavage site in its S protein, characterized by a four-amino acid insertion, which seems to be absent in all other coronaviruses (4). This molecular mimicry has been identified as an efficient evolutionary adaptation that some viruses have acquired for exploiting the host cellular machinery. Once the nucleocapsid is deposited into the cytoplasm of the host cell, the RNA genome is replicated and translated into structural and accessory proteins. Vesicles containing the newly formed viral particles are then transported to and fuse with the plasma membrane, releasing them to infect other host cells in the same fashion (33, 89, 105). Although much progress has been made in our understanding of the mechanisms underlying SARS-CoV-2 invasion, additional research is needed to delineate exactly how cleavage of the S proteins by TMPRSS2 confers viral particle entry as well as how S-protein cleavage by membrane proteases contributes to viral penetration.

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Non-structural protein 2a

NCBI ProteinQ80872
TaxonomyHuman coronavirus OC43

1Names and Identifiers


Non-structural protein 2a


32 kDa accessory protein

32 kDa non-structural protein


1.2Other Identifiers

1.2.1UniProt ID



>sp|Q80872|NS2A_CVHOC Non-structural protein 2a (Run BLAST)


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