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Archive for the ‘SARS -Cov2’ Category

covid19: corticosteroids, male susceptibility, evaluating health, remdesivir, coagulation factors

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from The Lancet, ‘the four horsemen of a viral apocalpse’

 

Canto: So short-course use of some steroids was being advocated in the medcram update 88, though without thorough RCT evidence. 

Jacinta: Well, data was presented from the Oxford RCT on those on oxygen or on ventilators showing a statistically significant reduction of mortality from short-course (up to 10 days) low dosage of dexamethasone, a freely-available steroid medication. The study involved some 2000 patients, but only those severely afflicted were helped by the medication. 

Canto: An interesting aside to the data is that in the study males outnumbered females by almost 2 to 1, and that accords with the overall ratio of male to female covid19 patients Dr Seheult is finding, which rather shocked me. Why would more males be coming down with the disease? Presumably that’s not the infection rate, but the rate at which they need to be hospitalised. 

Jacinta: Yes, you’re right, according to this Australian site (unfortunately undated):

Reports continue to emerge that men are significantly more vulnerable to COVID-19 than women. The commonly held perception that more men smoke and this makes them more susceptible along with other lifestyle factors does not tell the whole picture. White House COVID-19 Task Force director Dr Deborah Birx highlighted a “concerning trend” that men in all age brackets were becoming seriously ill from the virus at a higher rate than women, including younger males.

They’re suggesting more research needs to be done on this gender difference, for health issues in general. Some are claiming that estrogen makes a difference. In any case I think cardiovascular problems are more common in males – but maybe not so much in younger males. 

Canto: So update 89 is fairly short, and deals with US data about cases and deaths, most of it out of date now, and more on corticosteroids and the dangers of unsupervised use. Update 90 introduces us to a tool I’ve never heard of called ‘Discern’. Very useful for we autodidacts in helping us, for example, to enlighten our doctors as to our condition. Discern is a tool for evaluating internet health info, such as medcram’s updates on youtube, or anything else on youtube. The instrument asks you to evaluate the material according to 16 different criteria. Interestingly, this tool has been tested on covid19 material by a study out of Poland done in March. The results weren’t so good, especially for news channels. 

Jacinta: Yes, physicians’ information did best – but of course we don’t go to news channels for health information, and we’d advise against anyone else doing so. The study evaluated the Discern tool itself and found it excellent, then used the tool to evaluate health information, specifically on youtube. Of course know that there’s ‘viral misinformation’ from various news outlets that gets posted on youtube. And good to see that the medcram updates were some of the most highly rated using the Discern tool. 

Canto: So we’re now into reporting from early July with update 91. It starts by looking at a ‘covid risk calculator’ in which you can type in your age, gender, BMI, underlying conditions, waist circumference, and other data which you might need a full medical checkup to find out about (and that’s overdue for me), including, for example, %FMD, a measure I’ve never heard of, but which has to do with endothelial function. 

Jacinta: FMD stands for fibromuscular dysplasia. The Johns Hopkins medicine site describes it as a rare blood vessel disease in which the cells of some arteries become more stiff and fibrous and less flexible. This leads to weakness and damage. Not sure how it relates to covid19 but surely any pre-existing blood vessel damage is a danger for those contracting the virus. 

Canto: Right, so it’s unlikely anyone will know offhand their percentage of FMD. I don’t even know my HDL and LDL levels, never mind my HbA1c or lipids. I’d love to be able to take measures of all these myself, without visiting a doctor.

Jacinta: Typical male control freak. So all of this is to measure your risk of covid19 hospitalisation, ICU admission or mortality. Fun times. So next the update looks at Gilead, the makers of the antiviral remdesivir, who donated all their supplies of the drug to the USA in early May. But of course they kept manufacturing the drug and have to recoup the money they spent researching, developing and trialling it etc. The Wall Street Journal reports that a typical course of the drug will cost over $3000 per patient. Interestingly the Trump administration is wanting the drug to stay in the USA as much as possible, rather than be available overseas, and is spending money to that effect. 

Canto: Hmm. Is that protectionism? 

Jacinta: Yes I suppose. It’s not surprising that a country wants to look after its own first, especially via a product produced within its own borders. But I suspect this government would’t be interested in helping any other country – unless there was a quid pro quo. And there’s another antiviral, favipiravir, currently being trialled in Japan and the USA (I mean as of early July), and a vaccine, developed in China, is being used on the Chinese military in what seems a rather rushed and somewhat secretive fashion – we don’t know if they got the soldiers’ permission on this seemingly untried vaccine. At least at the phase 3 level.

Canto: Very CCP. 

Jacinta: So onto update 92, and we revisit the electron transport chain, with four successive electron transfers converting molecular oxygen into water. Problems within this chain can produce reactive oxygen species (ROS) such as superoxide, hydrogen peroxide and hydroxy radicals, which are destructive in excess. We also look, yet again, at covid19’s impact on angiotensin and particularly the production of superoxide, which in turn causes endothelial dysfunction, increased von Willebrand factor activity, which leads to thrombosis. People were presenting as ‘happy hypoxics’, looking and feeling fine but with very low oxygen levels, and autopsies revealed ‘microthrombi in the interalveolar septa’ of victims’ lungs. All this leading to a paper published in The Lancet which looked at factors in this process of coagulation and thrombosis:

We assessed markers of endothelial cell and platelet activation, including VWF antigen, soluble thrombomodulin [a marker of endothelial cell activation], soluble P-selectin [a marker of endothelial cell and platelet activation], and soluble CD40 ligand [a marker of platelet and T-cell activation], as well as coagulation factors, endogenous anticoagulants, and fibrinolytic enzymes.

So this was about getting to the bottom of the increased clotting. And the results were hardly surprising, but the final discussion section is worth quoting at length, as it seems to capture much that we know about covid19’s effects (at least short-term effects) at the moment: 

We therefore propose that COVID-19-associated coagulopathy is an endotheliopathy that results in augmented VWF release, platelet activation, and hypercoagulability, leading to the clinical prothrombotic manifestations of COVID-19-associated coagulopathy, which can include venous, arterial, and microvascular thrombosis. The factors responsible for this endotheliopathy and platelet activation are uncertain but could include direct viral infection of endothelial cells, collateral damage to the tissue as a result of immune infiltration and activation, complement activation, or any number of inflammatory cytokines believed to play a role in COVID-19 disease.

They suggest anti-platelet therapy and endothelial cell modification treatments as well as anticoagulation treatments, and they suggest some agents ‘which might have therapeutic potential’.

Canto: Potential? You’d think they’d be onto all this by now. 

Jacinta: Well there’s also potential for untried medications – at least untried in this context – to go terribly wrong. And it’s also likely that some hospitals are already onto using the safer forms of treatment. Dr Seheult speaks of the antioxidant N-acetylcysteine (NAC) in this context, as it has been shown to be a thrombolytic when used intravenously. There are studies pending on the effects of NAC in treating covid19 patients. 

Canto: Now, I’ve just been watching something on monoclonal antibodies as perhaps the most promising treatment yet, short of a vaccine. Can you explain….

Jacinta: Yes I’ll try, maybe next time.

References

Coronavirus Pandemic Update 88: Dexamethasone History & Mortality Benefit Data Released From UK

Coronavirus Pandemic Update 89: COVID 19 Infections Rising in Many States; Dexamethasone Cautions

Coronavirus Pandemic Update 90: Assess The Quality of COVID-19 Info With A Validated Research Tool

Coronavirus Pandemic Update 91: Remdesivir Pricing & Disparities in Drug Availability

Coronavirus Pandemic Update 92: Blood Clots & COVID-19 – New Research & Potential Role of NAC

amhf.org.au/covid_19

http://www.discern.org.uk

https://www.thelancet.com/journals/lanhae/article/PIIS2352-3026(20)30216-7/fulltext

 

more covid 19: vitamin D, helper T cells, testing

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I’m continuing with my gleanings from the Medcram Covid-19 updates presented by Dr Roger Seheult, though I’m not up to date with them, because they’re quite comprehensive and nuanced, and I want that detail more than anything. I’m also reading the book Outbreaks and epidemics: battling infection from measles to coronavirus, by Meera Senthilingam, which among other things, highlights the importance of preparedness, co-ordination and resourcing to deal with new and unexpected pathogens but also upsurges and cross-border spread of diseases we haven’t sufficiently dealt with in the past. As we hurtle at an unprecedented rate towards a number of vaccines against SARS-CoV2, for example, we may have to deal firmly, on a governmental level, with the anti-vaccination movement and its disinformation campaigns, but we also have to deal with grossly uneven levels of healthcare within and across nations. This current pandemic has been revelatory, for all but those on the front lines, of the variable impact such outbreaks have on the different levels of empowerment within societies. To take a stark example, Boris Johnson, the British Prime Minister, very likely owes his life to the fact that he is the British Prime Minister. Had he been a fifty-something person of colour living in Dagenham (or most anywhere outside of a UK city), his Covid-19 case would surely have turned out quite differently.

Update 74 is quite brief and mainly touches on vitamin D, the ‘sunlight’ vitamin, also obtained from foods such as fish, especially salmon and tuna, and egg yolks, and mushrooms raised using UV light – but mostly from the sun’s UV. Vitamin D enhances bone and muscle strength and function. A Lancet article is discussed, which correlates ‘vitamin D status’, presumably meaning bodily levels, with Covid-19 mortality. Some surprises in the data – vitamin D deficiency was common in ‘sunny’ Italy and Spain, but less of a problem in Nordic countries, perhaps due to a high vitamin D diet. Deficiencies were greater in poorer regions and in black communities, as of course were higher Covid-19 mortalities. in fact, ‘black people in England and Wales are 4 times more likely to die from Covid-19 than white people’ according to the UK’s Office for National Statistics.

The Lancet article referred to points out two aspects of vitamin D’s possible protection against Covid-19. First, it ‘supports production of antimicrobial peptides in the respiratory epithelium’, which sounds positive, and second it may help to reduce the inflammatory response to the virus because it’s known to interact with and promote the ACE-2 protein, which the virus suppresses. Other articles emphasise the benefits, with no attendant harm, of vitamin D supplements, particularly for the elderly. There have been no systemised, detailed trials as yet relating vitamin D levels to Covid-19 outcomes, but it seems like a no-brainer.

Update 75 continues the argument about SARS-Cov2 attacking the lining of the blood vessels, i.e. the endothelium, with the resultant effect on von Willibrand factor. This happens in the lungs as well as the vascular system, creating clots as well as the growth of new blood vessels as a type of immune response. This essentially marks it out from any kind of influenza. The New England Journal of Medicine has an article, published late May, looking exactly at these differences in the autopsies of Covid-19 victims – endothelialitis (inflammation of the endothelium) and angiogenesis (the formation of new blood vessels). They compared Covid-19 lungs with the lungs of ARDS (acute respiratory distress syndrome) victims, associated with influenza A (H1N1), and with uninfected lungs. They found ‘alveolar capillary microthrombi’ – often difficult to detect with scans – in the Covid-19 lungs at nine times the level of the influenza lungs, and new vessel growth at almost three times that of the influenza lungs. Clearly the new vessel growth is caused by the blockages, and the need to circulate around them. Microscopic analysis shows lymphocytes infiltrating the lungs, adding to inflammation, stiffness and tissue damage. The clotting prevents oxygen being picked up from the alveolar space, leading to low oxygen saturation of the blood. Scanning electron micrographs of the lung endothelium revealed viral particles in the extracellular space, suggesting strongly that the virus itself, and not simply the immune response to it (perivascular inflammation) is causing damage. Dr Seheult brings up NAC again here, as a possible disruptor of the cascade of events, especially in the suppression of superoxide and in the cleaving of disulphide bonds in VWF.

An article in Science, which refers to the adaptive immune system, is next discussed. The adaptive immune system, as opposed to the innate immune system, is a system that creates a memory of a pathogen in order to develop an enhanced response, a system exploited by vaccines. This system includes T cells, of which there are three types, memory, cytotoxic and helper. These cells are apparently involved in lifelong immunity. Vaccine researchers are concerned to create antibodies as protection against the virus, but T cells are also important in this regard, and researchers have found that many infected patients, and non-infected people, do have T cells that attack the virus, probably because they have been infected with other coronaviruses that share proteins, such as the spike protein, with SARS-CoV2. Researchers in fact found that Covid-19 patients all harboured helper T cells that recognised the SARS-CoV2 spike protein, and other SARS-CoV2 proteins, again suggesting the possibility/probability of lifelong immunity. Many others harboured the same helper T cells, which may be protecting them against the worst Covid-19 symptoms, before the fact. This is possibly a very important, and highly explanatory finding. Or maybe not. T cells are long-lasting, so these findings are certainly positive.

Update 76 starts with antibodies, and it’s a bit difficult to follow. It looks at the CDC’s ‘interim guidelines for Covid-19 antibody testing’, and a CNN health article summerizes it thus:

The CDC explains why testing can be wrong so often. A lot has to do with how common the virus is in the population being tested. For example, in a population where the prevalence is 5%, a test with 90% sensitivity and 95% specificity will yield a positive predictive value of 49%. In other words, less than half of those testing positive will truly have antibodies’, the CDC said.

This is hard to follow, but 5% prevalence is fairly standard for this virus, at least at the outset. And so false positives are a problem. To be clear about testing – a person either has the disease or not. If you have it and you test positive, fine, that’s a true positive. If you have it and test negative, that’s a false-negative. If you don’t have it and you test positive, that’s a false-positive. If you don’t have it and test negative, that’s a true negative.

So we can look at percentages and maths, and I’m following Seheult strictly here. So imagine we’ve tested 2100 people in a particular region – that’s everyone in the region. At this stage the disease has a prevalence of 5%, so about 100 out of 2100 have the disease (strictly speaking that’s 4.76%). The test has a sensitivity of 90% and specificity of 95% as above. 90% sensitivity means that the number of true positives from the test will be 90% of the number of those who actually have been infected by the virus. That means 90 people. 95% specificity is about those not infected. So you divide the true negatives by those uninfected to arrive at the 95%. The true negatives will amount to 1900. So 10 people will be false positive and 100 false negatives. When specificity rises, false positives decrease. When sensitivity increases, false negatives decrease. So with high sensitivity a negative result is more conclusive, and with high specificity, a positive result is more conclusive.

Imagine then that the prevalence of the infection has risen to 52% in the same population of 2100. That gives us 1094 with the disease, 1006 without. With the same values for sensitivity and specificity of testing, you’ll have 985 true positives and 50 false positives, and 956 true negatives and 109 false negatives. What you need to know with these results is how things stand for patient x, the person you’re dealing with. This means you need to know the predictive values, positive (PPV) or negative (NPV). This requires some simple maths. Given a positive test result, what chance is there of x having the disease? Or vice versa for a negative result. This means that for the PPV you divide the true positives by the total number of positives, and the same process applies for NPV. Going back to the situation where the prevalence was 5% we get a PPV of 47% and a NPV of 99%. What this means is that when the prevalence is low, the negative predictive value is much higher than the positive predictive value. The implication is important. It’s just not clear at this stage whether you have antibodies against the virus. So you need to raise the specificity of the test, especially if the virus or pathogen has a low prevalence. But looking at the 52% prevalence case, and using the same simple maths we find that the PPV is up at 95% and the NPV goes down to 90%. Prevalence, then, is the main determinant of predictive values.

For testing, this means, just as the disease is becoming prevalent, that’s to say, as it’s just being detected, you need a test with a very high specificity (admittedly a big ask) and/or you need to test those with a high probability, based on current knowledge, of being infected, and those in contact with them.

References

Coronavirus Pandemic Update 74: Vitamin D & COVID 19; Academic Censorship

Coronavirus Pandemic Update 75: COVID-19 Lung Autopsies – New Data

Coronavirus Pandemic Update 76: Antibody Testing False Positives in COVID-19

https://en.wikipedia.org/wiki/T_helper_cell

Written by stewart henderson

August 9, 2020 at 12:34 pm

covid-19 stuff: NAC, glutathione, RT-PCR testing, re-positives

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So, more struggles with biochemistry. Update 70 talks again about N-acetylcysteine (NAC), but goes on to talk about glutathione, and whether glutathione itself might be a type of medication. So let’s get clear, or try to.

Glutathione is a naturally occurring and abundant thiol polypeptide in animal cells. A thiol has an SH (sulfanyl) group attached to a hydrocarbon chain, essentially. As we know, it’s an antioxidant which can be reduced by NAC, and they have structural relations. As Dr Seheult describes glutathione, it’s a combination of three amino acids, with cysteine at the centre. The other two are glycine and glutamate, and the cysteine and the glycine together effectively make up N-acetylcysteine – so NAC is described as a by-product or precursor of glutathione. A case report (regarded as the weakest level of scientific evidence) describes efficacious treatment of two patients with Covid-19-type symptoms using IV and oral glutathione. This and other studies and analyses seem to be begging for full-scale clinical trials to be carried out, but nothing as of mid-May. The treatments could be effective for hypoxemia in particular, due to the action on the disulphide bonds in VWF which are leading to platelet-rich thrombosis.

In his update 71 Seheult broaches the controversial topic of hydroxychloroquine, along with azithromycin and zinc. He suggests there’s evidence that hydroxychloroquine can act as a ‘zinc ionophore’, inducing zinc uptake into cells. Zinc inhibits the RNA-dependent RNA polymerase which SARS-CoV-2 utilises to reproduce. There has been a retrospective study suggesting that treatment with this combination may ‘result in a statistically significant reduction of mortality’, though maybe this hasn’t borne more careful analysis considering the cold water being poured on chloroquine as a treatment in recent months. It may be because it just doesn’t raise zinc levels sufficiently. The findings of the study do suggest the treatment has a statistically significant effect on reducing symptoms in hospitalised patients who are not in ICU – that is, they have relatively mild symptoms. No significant effect for ICU patients.

I should add here that now in August health authorities are warning against any unprescribed use of hydroxychloroquine as a prophylactic due to ineffectiveness and side-effects.

Update 72 began by looking at the sensitivity and specificity of antibody tests available, presumably in the USA. A study examined ‘four new commercially available serological assays [i.e blood serum tests]’, from three German and one US company, and it was found that they all ‘have a sufficient sensitivity and specificity for identifying individuals with past SARS-CoV2 infection’. Of course, the principal issue with the testing is the time it takes to receive results, but maybe that’ll be addressed anon.

Apparently (news to me in very safe – so far – South Australia where hardly anyone I know has had to be tested) there’s a difference between sensitivity and specificity, illustrated by the ‘spin’ and ‘snout’ mnemonic. For a highly specific test if you test positive you’re very sure to be in trouble, and for a highly sensitive test if you test negative you’re sure to be out of danger.

Dr Seheult next describes a retrospective study which looks at glycosylated haemoglobin (HbA1c) as a Covid-19 risk factor. A person’s HbA1c levels (how much glucose is attached to their haemoglobin) are a measure of diabetes. A1c (blood sugar level) is measured in percentages, with 5-6% being normal. The study found that ‘high HbA1c levels is associated with inflammation, hypercoagulability and low SaO2 [oxygen saturation] in Covid-19 patients, and the mortality rate (27.7%) is higher in patients with diabetes’. So HbA1c levels need to be looked at as a priority.

The update next looks at dentistry during the pandemic, for which there’s been little guidance, at least from the CDC. Apparently, during the AIDS crisis, dentists were viewed as modes of transmission, partly due to a NYT article on the subject. In any case, fewer people are now seeing their dentists for obvious reasons, which could lead to an oral health crisis. A number of diseases, including coronary disease, are linked to periodontal problems, so this can exacerbate the pandemic – and dental health, in Australia as in the USA, is not treated with the same gravitas as other forms of health.

Update 73 starts with a look at testing, particularly the reverse transcriptase polymerase chain reaction (RT-PCR) test. So the coronavirus has these spike proteins protruding from a bilipid membrane, with the RNA wrapped inside bound together by disulphide bonds and the like, I think. The protein shell around the virus is called the nucleocapsid. Of course the RNA’s code is specific to SARS-CoV2, so a test needs to look at a segment of the viral RNA and identify it with sufficient – essentially total – specificity. RNA is made up of the four base pairs adenine (A), uracil (U), guanine (G) and cytosine (C), with A pairing always with U and G with C. With that I’m going to switch to Scientific American for more detail.

A test starts with a sterile swab from the back of the nasal passage, aka a sample. Sample collection needs to be done properly, or it could lead to a false-negative result. If there’s viral RNA present, it’s extracted and used to produce a complementary strand of DNA – that’s where the reverse transcriptase enzyme comes in, reversing the usual transcription process from DNA to RNA. This material is then amplified – thousands of copies are made – to ensure a measurable result. The different available test kits generally vary in the segment of genetic material chosen.

I’m hearing that there are serious delays, in the USA at least, in delivering test results. This is extraordinary as, according to the Scientific American article, which is dated late March,

the FDA recently began granting emergency use authorization (EUA) to rapid diagnostic PCR tests that manufacturers say can deliver results in less than an hour. The authorization allows medical devices that have not yet been approved by the agency to be used during public health emergencies. 

What’s happening? According to very recent article from Quartz magazine, the problem is that there are too many kinds of tests. The EUA system was utilised, partly because of the urgency, partly because of the disastrous problem caused by the use of faulty reagents by the CDC back in February. Now there are about 150 tests that have been given EUA approval. Testing delays at first resulted mainly from lack of general lab equipment and PPR for the testers, but increasingly there are problems due to different types of tests, the variability of the tests, knowing which test to use, having the right equipment for each test, the prioritising of certain groups, such as front-line health workers, over others, confirmation of test results by other labs, and of course the overload in demand. We’re talking about the USA here, of course, and it just seems another case of lack of centralised control and uniformity in a state with a failed federal government.

Returning to update 73, Seheult describes a situation in which a SARS-Cov2-infected individual’s immune system has broken down the virus into ineffectual strands of RNA, proteins and other particles. It’s possible that a RT-PCR test could pick up on an RNA fragment, and produce a positive test result in these apparently recovered patients, and in fact this has often occurred. This is called a re-positive. The update describes a study by the South Korean CDC which provides valuable evidence on these re-positive cases. Some 280 re-positive subjects were studied, and about half of them displayed Covid-19 symptoms (on average 14 days after ‘recovery’). Presumably this re-positive finding was after they’d tested negative, i.e they’d first tested positive, then negative, then later positive again, though this isn’t clear. In any case, they checked a percentage of the subjects for antibodies and the result was almost entirely positive. They checked a larger sample for viral particles and found ‘not a single whole viral particle’, according to Dr Seheult, by which I presume he means anything that was replicable or active. They also looked at close contacts of the subjects, in large numbers, and all of them tested negative. So the finding was that these re-positives were, it seemed, the results of ultra-sensitive testing that was picking up viral RNA fragments that were in effect innocuous. This would seem to be a lesson for developing the right types of test. Hopefully a lesson learned.

References

Coronavirus Pandemic Update 70: Glutathione Deficiency, Oxidative Stress, and COVID 19

Coronavirus Pandemic Update 71: New Data on Adding Zinc to Hydroxychloroquine + Azithromycin

Coronavirus Pandemic Update 72: Dentists; Diabetes; Sensitivity of COVID-19 Antibody Tests

Coronavirus Pandemic Update 73: Relapse, Reinfections, & Re-Positives – The Likely Explanation

https://www.scientificamerican.com/article/heres-how-coronavirus-tests-work-and-who-offers-them/

https://qz.com/1886940/why-covid-19-test-results-take-so-long/

Written by stewart henderson

August 5, 2020 at 2:34 pm

more Covid-19 gleanings from MedCram updates 67-69

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polymorphonuclear leukocytes (white blood cells)

I’m continuing my self-education re everything Covid-19 thanks to Dr Seheult’s updates and other useful sites. Update 67 carries on from where we left off, summarising again how SARS-CoV2 induces endothelial dysfunction, before focusing on thrombosis. So we repeat again that a key molecule in normal endothelial function and in the working of AT-1,7 is nitric oxide (NO). Endothelial function (and, to be clear, the endothelium lines the vasculature, which means the body’s blood vessels) is also dependent on the various other enzymes mentioned in the last post, e.g. superoxide dismutase (SOD), and glutathione peroxidase (GPx).

So how does Covid-19 bring about oxidative stress and how does this effect thrombosis? Seheult discusses an article from April this year which addresses this. It describes a previously healthy elderly male admitted to hospital with fever and respiratory symptoms. After rapid deterioration he was sent to ICU, having developed ARDS, acute renal insufficiency and other health problems. Among various measures noted was a ‘massive elevation of von Willebrand factor (VWF), as well as ‘factor VIII of the coagulation cascade’. To quote from the article:

The increased VWF points toward massive endothelial stimulation and damage with release of VWF from Weibel-Palade bodies. Interestingly, endothelial cells express ACE-2, the receptor for SARS-CoV2, thus possibly mediating endothelial activation.

To explain some of these terms: Weibel-Palade bodies are found only in epithelial cells, and they contain VWF, which are released when required for haemostasis and coagulation. VWF is a stringy material of amino acid proteins which combine with platelets (aka thrombocytes) to coagulate the blood. When endothelial cells suffer serious damage, Weibel-Palade bodies inject large amounts of VWF into the bloodstream. Dr Seheult presents the abstract from a 2017 article on the topic:

The main function of VWF is to initiate platelet adhesion upon vascular injury. The hallmark of acute and chronic inflammation is the widespread activation of endothelial cells which provokes excessive VWF secretion from the endothelial cell storage pool. The level of VWF in blood not only reflects the state of endothelial activation early on in the pathogenesis, but also predicts disease outcome. Elevation in the blood level of VWF occurs either by pathologic increase in the rate of basal VWF secretion or by increased evoked VWF release from dysfunctional/activated endothelial cells. The increase in plasma VWF is predictive of prothrombotic complications and multi-organ system failure associated with reduced survival in the context of severe inflammatory response syndrome, type 2 diabetes mellitus, stroke and other inflammatory cardiovascular disease states.

The article points out that an over-production of VWF in highly elongated form is an indication of pathology. This is apparently being seen in serious Covid-19 patients. On the molecular level, the VWF is able to remodel itself from its usual globular conformation when it senses shear forces – note this definition from Science Direct: Shear stress is defined as the frictional force generated by blood flow in the endothelium, that is, the force that the blood flow exerts on the vessel wall, expressed in force-area unit (typically dynes/cm2). The VWF, under this stress, ‘turns into an extended chain format that forms ultra-large strings to which platelets bind to initiate clot formation at sites of vascular damage’. When the shear stress reaches a certain level, factor VIII is released. All of this can be essential for haemostasis, but too much of the multimeric, elongated form of VWF will lead to thrombosis, as appeared to be occurring in the patient described above.

So, as Seheult summarises, SARS-CoV2 binds to ACE-2 receptors and reduces ACE-2 production. This reduction has the effect of increasing AT-2 production and reducing AT-1,7. This results in an increase in superoxide production, oxidative stress and endothelial dysfunction. This in turns leads to an increase in VWF activity in the bloodstream, and local thrombosis. There is evidence from autopsies that thrombosis is a feature of Covid-19 mortality.

In his update 68 Dr Seheult looks at the predisposition of some ethnic groups (in the USA) to the more severe symptoms associated with Covid-19. He discussed a May CDC MMWR (morbidity and mortality weekly report) on 580 hospitalised Covid-19 patients which found that 45% were white, as far as they could ascertain, compared to 55% in that region’s community. 33% were black, compared to 18% in the community, and 8% were Hispanic compared to 14% in the community. A smallish sample, but suggestive. The CDC also reported on New York figures showing that Covid-19 death rates among black/African Americans and Hispanic/Latino persons were substantially higher than in the white population. Many possible reasons – work and living conditions, lower access to care – all generally related to relative poverty. There may also be other, purely physiological grounds for the disparity. A 16-year-old research article published in Circulation describes the results of placing nanosensors in isolated human umbilical vein endothelial cells (HUVECS) from blacks and whites (pardon the over-simplification, I’m only the messenger), as an attempt to measure endothelial oxidative stress. I can’t follow the details of the research, but what they found was that blacks expressed much more NADPH oxidase than whites (that’s bad). Nitric oxide, a reducer of oxidative stress, was produced in greater quantities in whites than in blacks, and the bad superoxides were produced in greater quantities in blacks. I won’t go further into the complex biochemistry, but I must say I find these apparent racial differences very surprising.

Update 68 also looks at increasing hospitalisations (at least in May) of young children due to Kawasaki disease, or something similar. The disease is characterised by inflammation of blood vessels. Symptoms include fever, high heart rate and possibly sepsis. There are a number of similarities to Covid-19, including ‘systemic vascular lesions’. Kawasaki disease is normally rare, and believed to be viral, or a response to a virus. A ten-year-old research paper on the disease hypothesises that the infection enters through the respiratory or gastro-intestinal systems, and so unsurprisingly there are similarities to the reaction to SARS-CoV2. Whether there’s a connection between Covid-19 and an uptick in Kawasaki disease has yet to be confirmed (but I’m behind the times on the research on this).

I’m moving now to update 69, and I’m going to follow Dr Seheult through the whole oxidative stress process again. It’s about reduction of oxygen – the adding of electrons. Adding an electron to oxygen, mediated by NADPH oxidase, produces superoxide. Add another electron and you get hydrogen peroxide. Another electron produces hydroxyl, and yet another produces water, moving from most oxidised to most reduced, and adding electrons also brings on protons. So at both ends of this chain you have neutral or positive molecules, but in between you have, I think ROS, reactive oxygen species, which are a problem. The body’s defence against these include the enzyme superoxide dismutase (SOD), which converts superoxide into hydrogen peroxide and also back into oxygen, and catalase which converts hydrogen peroxide into water and oxygen. Another important enzyme which protects against oxidative damage is glutathione peroxidase (GPx). It takes reduced glutathione (2GS-H, called a sulph-hydryl group) and uses it to reduce hydrogen peroxide into water, in the process oxidising the glutathione into a form of disulphide G-S-S-G. This oxidised form is in turn ‘regenerated back’ by taking the reduced form of NADP+ (NADPH) and converting it via glutathione reductase to NADP+.

So the point is that the accumulation of superoxide in people with diabetes, hypertension, coronory disease etc will be exacerbated by Covid-19. And going through that once more, Covid-19 blocks the ACE-2 receptor, causing an accumulation of AT-2 which stimulates superoxide production, and also a deficiency of AT-1,7, which, mediated by nitric oxide, inhibits superoxide production. The SARS-CoV2 virus also attracts PMNs (polymorphonuclear leukocytes – immune cells including neutrophils), which boost superoxide production, with attendant endothelial damage.

I’ll be continuing this series, and no doubt getting further behind, over the next few weeks.

References

Coronavirus pandemic update 67, presented by Dr Roger Seheult, as with all other updates

Coronavirus pandemic update 68

Coronavirus pandemic update 69 (first 5 minutes or so)

https://www.verywellhealth.com/polymorphonuclear-leukocyte-2252099

Written by stewart henderson

July 29, 2020 at 11:11 am

more on oxidative stress and covid-19

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So, much of this piece will rely on Dr Seheult’s coronavirus update 65. We have this constant set of reactions in the body that reduce oxygen – adding electrons – until we get to water molecules, producing reactive oxygen species (ROS) along the way. This is often described as the oxygen metabolism process. Reactive oxygen species come essentially in three types, superoxides, hydroxy radicals and hydrogen peroxide. The three forms of the enzyme SOD, superoxide dismutase, convert superoxide into oxygen and hydrogen peroxide (H2O2), and then the H2O2 is reduced to H2O by means of glutathione peroxidase (GPx). The GPx, which is broken down in the process is recharged by the enzyme glutathione reductase (GR), which is in turn recharged by other antioxidant products. Also the enzyme catalase, which requires iron, can break H2O2 down into O2 and H2O.

People with diabetes, hypertension and overweight issues, among other things, may have compromised antioxidant systems (too many ROS), linked to angiotensin-converting enzyme 2 (ACE-2) and angiotensin-2. In creating ROS, oxygen is reduced to superoxide by means of the enzyme NADPH oxidase. So, as part of the renin-angiotensin system, angiotensin-2 (AT-2) is converted to angiotensin 1,7 (AT-1,7) by means of angiotensin-converting enzyme 2 (ACE-2). This is important because AT-1,7 effectively blocks superoxide production, while AT-2 promotes it. The virus SARS-CoV2 binds with, and so inactivates, ACE-2, preventing the production of AT-1,7. This action also means that there will be more AT-2 available, and so more superoxides. SARS-CoV2 also, according to Seheult, causes inflammation by recruiting polymorphonuclear neutrophils (PMNs), which stimulate production of superoxides by means of NADPH oxidase. So this, in essence, is why Covid-19 is bringing about oxidative stress.

Seheult next goes on to look at the research evidence for the preceding. A review article from 2005 points out that evidence from animal studies and cell culture studies shows that NADPH oxidase-derived oxidative stress is increased in vascular cells by AT-2, among other ‘agonists’ (chemicals that bind to receptors, thereby producing a response). Another article from 2012 describes several enzyme systems that act to form ROS, including ‘mitochondrial electron leakage from the electron transport chain’ as described in my previous post on the subject, and in Seheult’s update 63. It points out that ROS levels can rise dramatically in older people suffering from oxidative stress due to heart issues such as ischemia-reperfusion (referring to problems with oxygenated blood supply to the heart or other organs). It also points out that it has been shown experimentally that AT-2 stimulates an increase in ROS. A more recent article pertaining to SARS-CoV2 looked at patients in Wuhan and found a substantial increase in neutrophils in the most severe cases. Neutrophils cause ROS to be generated by NADPH oxidase. So Dr Seheult is carefully building up evidence for the case. The last point to deal with is AT-1,7 effects. Seheult has found a 2008 article entitled ‘Angiotensin converting enzyme 2 confers endothelial protection and attenuates atherosclerosis’. Seheult quotes the last line from the abstract:

These data indicate that ACE-2, in an AT-1,7-dependent fashion, functions to improve endothelial homeostasis via a mechanism that may involve attenuation of NADPHox-induced reactive oxygen species production. ACE-2-based treatment approaches may be a novel approach to limit aberrant vascular responses and atherothrombosis.

Atherothrombosis involves disruption of atherosclerotic plaques, which can be an immediate cause of heart attacks. Another article from 2015 essentially confirms the findings, as indicated by its title, ‘ACE-2 and AT-1,7 protect endothelial cell function and prevent early atherosclerosis by inhibiting inflammatory response’. A more recent article, from January 2020, describes how AT-1,7 administration improves endothelial function in women who have suffered from preeclampsia (vasoconstriction, high blood pressure and organ damage due to pregnancy). To give more detail, women in the last stages of pregnancy often suffer vasoconstriction and high protein levels, which is believed to be related to AT-2 levels. Researchers administered local AT-1,7, which is ‘an endogenous inhibitor of... AT-2′, to see if this reduced vasodilation and other symptoms of preeclampsia. What they found was that ‘AT-1,7 increased endothelium-dependent vasodilation via nitric oxide synthase-mediated pathways and attenuated AT-2-mediated constriction in women who have had preeclampsia, suggesting that AT-1,7 may be a viable therapeutic target for improve d microvascular function in women who have had a preeclamptic pregnancy’.

All of this is interesting in itself, of course, and is a little crash course in how research is helping us to tweak our immune systems, but in relation to Covid-19 these finding are of importance due to the comorbidities and general characteristics of patients being hospitalised with Covid-19. Dr Seheult, in his update 65 video, shows that, contrary to what was initially thought, i.e that Covid-19 is primarily a virus affecting the lungs and respiratory system, it may be much more of a problem for those with hypertension, cardiovascular issues and obesity – all of which are related to oxidative stress, as are diabetes and many forms of cancer. They contribute to endothelial dysfunction, which inevitably leads to oxidative stress, and may lead to thrombosis. Seheult here refers to a lengthy 2018 review article, ‘nutrients and oxidative stress: friend or foe?’, which among other things makes useful dietary suggestions for the combatting of oxidative stress – whole grains, nuts, fruit and vegetables, fish and legumes.

It’s been known for some time that endothelial cell dysfunction (ECD) can lead to thrombosis, as it is a major function of these cells to prevent thrombosis. The abstract from a 2002 study finds that ECD ‘is associated with decreased synthesis and oxidative inactivation of nitric oxide (NO)’ and it lists four types of antioxidant enzymes ‘essential for eliminating ROS that can inactivate NO’. It seems that the promotion of these enzymes can be associated with diet as above and with the reduction of risk factors such as hypertension, hypercholesterolaemia (high blood cholesterol), hyperhomocysteinaemia (homocysteine is an amino acid which can contribute to arterial damage and blood clots, and the condition is often associated with lack of vitamin B-12 or folate), cigarette smoking and diabetes mellitus. NO is the key molecule in maintaining endothelial function through these enzymes.

Now I’m having a look at Dr Seheult’s update 66 on blood pressure medications known as ACE inhibitors or ARBs. He cites an editorial article for the New England Journal of Medicine, on ‘inhibitors of the renin-angiotensin-aldosterone system and Covid-19’. This is a triple hormone system responsible for blood pressure regulation and fluid balance. Now, to return to what was outlined before, angiotensin-2 (AT-2) is converted to AT-1,7 by an angiotensin-converting enzyme (ACE-2). The SARS-CoV2 virus binds to the ACE-2 receptor and inhibits the enzyme’s production. This is problematic because AT-2 stimulates superoxide production (that’s bad), while the antioxidant AT-1,7 blocks it, so reducing oxidative stress. SARS-CoV2 also stimulates the production of PMNs, as above, which activates oxidative stress. Another part of this picture is that AT-1 is converted to AT-2 by ACE. There are blood pressure lowering medications, such as benazepril and lisinopril, aka ACE inhibitors, which reduce the production of AT-2. There are also angiotensin receptor blockers (ARBs), which may up-regulate ACE-2 (it isn’t clear, apparently). ACE inhibitors may do the same. The question being asked is, assuming these medications produce more ACE-2, will this lead to more infections because SARS-CoV2 has more ACE-2 to work with? Clearly it would be important to know whether to maintain these medications or not, that’s to say, whether these medications are a risk factor for contracting the virus or recovering from it. The above-mentioned article discusses three studies from different parts of the world, each involving thousands of participants. They all found no risks associating ACE inhibitors and ARBs with a higher risk of infection, severity of illness or death from Covid-19. One of the studies found that ACE inhibitors and statins were associated with a decreased risk of mortality, but these are observational studies and further research would need to be done.

So the above is a rather technical piece, highly reliant on the experts. I write to inform myself, and I’ve certainly been informed by writing this one. Apologies for its laboriousness, but I’ll be continuing… Please consult the references yourself if there’s anything you don’t understand.

References

Coronavirus Pandemic Update 65: COVID-19 and Oxidative Stress (Prevention & Risk Factors)

Coronavirus Pandemic Update 66: ACE-Inhibitors and ARBs – Hypertension Medications with COVID-1

https://www.mayoclinic.org/diseases-conditions/high-blood-pressure/in-depth/ace-inhibitors/art-20047480

https://www.healthline.com/health/homocysteine-levels

https://www.mayoclinic.org/diseases-conditions/high-blood-pressure/in-depth/ace-inhibitors/art-20047480

https://www.ncbi.nlm.nih.gov/pmc/articles/PMC5831951/

Written by stewart henderson

July 16, 2020 at 4:21 pm

SARS-Cov2 and oxidative stress

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Dr Roger Seheult, just doing his job, workaholically

So I feel it’s time for me to get back to the epidemiology and immunology stuff that I know so little about, especially as it pertains to SARS-Cov2. Watching Dr Seheult’s Medcram updates again after a long hiatus, and catching up with them from the end of April, I note that he’s arguing – and I presume this is a mainstream view, as he clearly keeps an eye on the latest research – that the virus mostly does its damage in attacking the body’s endothelium, and that this in turn causes oxidative stress. The endothelium is a thin layer of cells, or a layer of thin cells, that form the inner lining of the blood and lymph vessels (one day I’ll find out what lymph actually is and does).

Oxidative stress is associated with an imbalance in the level of oxidants such as super-oxide anion and hydrogen peroxide, reduced forms of oxygen (with extra electrons). I don’t really understand this, so I’ll start from scratch. But just preliminary to that, the effects of oxidative stress are manifold. Here’s a summary from news-medical.net:

Oxidative stress leads to many pathophysiological conditions in the body. Some of these include neurodegenerative diseases such as Parkinson’s disease and Alzheimer’s disease, gene mutations and cancers, chronic fatigue syndrome, fragile X syndrome, heart and blood vessel disorders, atherosclerosis, heart failure, heart attack and inflammatory diseases.

It’s known that SARS-Cov2 enters via the lungs, and does damage there, but it’s now thought that most of the damage is done in the endothelium. To understand this, Dr Seheult is going to teach me some ‘basic’ stuff about metabolism, oxidation, energy production and such. So, we start with mitochondria, the energy-producing organelles inside our cells, which have their own DNA passed down the female line. Looking into a mitochondrion, we have the matrix inside, and around it, between the inner and outer membranes, is the inter-membrane space (IMS). Our food, broken down into its essential components, carbs, fats and proteins, is absorbed into the matrix, and somehow turned into ‘two-carbon units’ called acetyl coenzyme A. This is metabolism, apparently. These molecules go through a famous process called the Krebs cycle, of which I know nothing except that it’s about more metabolism… Although now I know that it produces electrons, tied up in two important molecules, NADH and FADH2. These electrons ‘love to be given up’, a way of saying they ‘want’ to be reduced. The molecule that gives up electrons is said to be oxidised, the receiving molecule is reduced. So think of a molecule being reduced as the opposite of losing, rather counter-intuitively. The oxidised molecule is the one that loses electrons. All this is about energy production within the matrix, and the aim is to end up with a molecule I’ve heard and forgotten much about, adenosine triphosphate (ATP). This molecule is the energy molecule, apparently, and the energy is produced by ‘knocking off’ one of the phosphates, according to Dr Seheult, leaving, apparently, adenosine diphosphate (ADP) plus ‘energy’ (clearly, this part needs a little more detail). So going from the diphosphate form to the triphosphate requires energy, going the other way releases energy – none of which really explains why ATP is the body’s energy source. Anyway…

Returning to the carbs, fats and proteins, they go through these mitochondrial processes to produce electrons which want to reduce stuff. So NADH goes to the membrane which separates the IMS from the matrix of the mitochondrion, where proteins can be found that are willing to accept electrons, i.e. to be reduced. The electrons are brought in ‘at the very top of the scale’ (?) and lose some of their reducing ability, so they go down to a lower state of reduction, and protons are pumped into the IMS. (I’m sure this is all true but making sense of it is another matter. It certainly makes me think of proton pump inhibitors, drugs that reduce gastric reflux, but that would be the subject of another set of posts). Then ‘it goes to another species’ by which I think Seheult means another protein, judging from the video, but what he means by ‘it’ I’ve no idea. The NADH? The wave/body of electrons? Anyway, things keep going down to a lower level, becoming more oxidised, and more and more protons are pumped out. So there comes to be a very high concentration of protons (H+) in the IMS, creating a very low PH (high acidity). Meanwhile, the electron transport chain has gone down so many levels that it can only reduce oxygen itself, which by accepting electrons turns finally into water. It’s apparently essential to have sufficient oxygen to keep this cycle going, and to keep the protons pumping, because the protons in the IMS want to move to a place of lower concentration, in the matrix. In doing this, they pass through a channel, which involves, somehow, a coupling of ADP to ATP. Without enough oxygen, this process is stymied, ATP can’t be supplied, leading to insufficient energy and cell death.

So, I think I understand this, as far as it goes. Now, if you over-eat, with lots of high-calorie fats and carbs entering the cells, you’ll likely end up with a surplus of electrons, tied up in NADH and FADH2, which can cause problems. This is where super-oxides come in.

Oxygen is the final electron acceptor in the electron transport chain, and when you add an electron to this final acceptor you get a super-oxide, an oxygen molecule with an additional electron, aka a radical. These are very reactive and dangerous. They can cause DNA damage and serious inflammation, and the body uses them to kill bacteria. If you add another electron, you get H2O2, hydrogen peroxide, and another one again produces a hydroxy radical, OH. Another electron gives water, so it’s these intermediate molecules that are called ‘dangerous species’. Cells such as neutrophils (a type of white blood cell) make these, via an enzyme called NADPH oxidase, as part of their defence against antigens, but an accumulation of these radicals is problematic and needs to be dealt with.

from Dr Seheult’s presentation, showing the production of reactive oxygen species (ROS) – super-oxide, hydrogen peroxide and hydroxy radicals

One enzyme the body uses to bring down these accumulating radicals is super-oxide dismutase (SOD), which takes two super-oxides and converts them into O2 and H2O2. SOD comes in three types, related to where they reside – in the mitochondria, the cytosol and the extracellular matrix. These enzymes are powered by zinc, copper and, in the mitochondria, manganese. So what happens to the extra hydrogen peroxide created? An enzyme called glutathione peroxidase (GPx) reduces H2O2 to water by giving it two electrons. Where do these electrons come from? According to Seheult, and this is presumably ‘basic’ microbiology, the antioxidant glutathione has two forms, oxidised and reduced. The reduced form is 2GS-H, with a hydrogen bonded to the sulphur group. The oxidised form is G-S-S-G, a disulphide bond replacing the hydrogen. With the reduced form, GPx donates its extra two electrons to H2O2, reducing it to water. The glutathione system is recharged by reducing it back with NADPH, which has two electrons which are converted to NADP+ (?) Glutathione reductase is the key enzyme in that process. It might take me a few lifetimes to get my head around just this much.

Meanwhile there’s another system… Catalase, an iron-boosted enzyme, can convert two molecules of H2O2 into O2 and H2O. This occurs in organelles called peroxisomes. The major point to remember in all this is that super-oxides are harmful species that can cause oxidative stress, and the major solutions come in the form of SOD and GPx. In fact the general name for these harmful molecules – super-oxides, hydrogen peroxide, and hydroxy radicals – is reactive oxygen species (ROS).

So we have to relate all this to the effects of SARS-Cov2, which enters the body through the ACE-2 (angiotensin-converting enzyme-2) receptor. According to a 2008 research paper, ACE-2, the receptor for which is blocked by SARS-Cov2, ‘confers endothelial protection and attenuates atherosclerosis’. Quoting from the paper, we find a section called ‘ACE-2 modulates ANG II(angiotensin 2)-induced ROS production in endothelial cells’. The researchers’ essential finding was that ‘ACE-2 functions to improve endothelial homeostasis’, and it seems this function is being disrupted by SARS-Cov2. As Dr Seheult puts it, SARS-Cov2 inhibits the inhibitor, that is it inhibits ACE-2, which normally acts to regulate angiotensin 1,7 (not explained in this particular video), thus allowing NADPH oxidase to keep producing super-oxides, with the resultant oxidative stress. As Seheult concludes here, subjects with compromised systems caused by diabetes, cardiovascular disease or obesity, affecting the production or effectiveness of SOD and GPx, might be relying on ACE-2 and angiotensin 1,7 to maintain some semblance of health. Are these the subjects that are succumbing most to the virus? That’s to be explored in future videos, and future posts here.

Reference

Coronavirus Pandemic Update 63: Is COVID-19 a Disease of the Endothelium (Blood Vessels and Clots)? (video by Dr Roger Seheult – clearly a hero in this time)


Written by stewart henderson

July 5, 2020 at 11:46 pm