an autodidact meets a dilettante…

‘Rise above yourself and grasp the world’ Archimedes – attribution

Archive for the ‘comorbidities’ Category

covid19: autopsy analyses, biomarkers, von Willebrand factor

leave a comment »

von Willibrand factor, a multimeric blood protein which plays a central role in blood clotting

Canto: So we’re working hard to get through what has been reported on medcram update 95, even though it’s taking us further behind the times in terms of what’s happening in the fight against this virus – there’s been some controversy on convalescent plasma recently for example – because it’s important to get the most out of every report before going onto the next one.

Jacinta: Yes, which means we need to work harder and faster. So in this study of a number of fatal cases of covid19 they found ‘no endothelial abnormalities on microscopic review, in alignment with previous studies’, which suggests that evidence of endothelial damage just doesn’t seem to be there, but they couldn’t rule out pro-coagulant endothelial dysfunction in the absence of ‘histopathological evidence of cell activation or erosion’, and they referred to another autopsy study with specialised equipment which ‘demonstrated ultrastructural endothelial damage’. So it seems they’re struggling with causes.

Canto: What they call the precise aetiology of the disease. 

Jacinta: Yes that’s what we’re after. So they do mention elevated troponin in covid19, which appears to be found regularly. Troponins are ‘a group of proteins found in skeletal and cardiac muscle fibres that regulate muscular contraction’. As the update tells us, troponin tests measure cardiac-specific troponin in the blood as a sign of heart injury. This Australian site tells us more:

For patients who are hospitalised with COVID-19, mild elevation of troponin is common (19.7%) and frequently correlates with disease severity, acting as a marker for cardiac injury. The cause of troponin elevation in serious infection is multifactorial.

In the study under discussion, they consider that the elevated troponin has to do with ‘thrombosis of the microvasculature and cardiac veins’. This cardiac vein finding is apparently important – they found, they believe for the first time, that thrombosis of a cardiac vein can cause myocardial infarction. They also write about renal findings in their subjects, to ‘shed light on the pathogenesis of acute kidney injury in covid19’. They found virions in proximal tubular cells. A virion is essentially a full, active molecule of a virus (there’s still some disagreement about these definitions, it seems). The proximal tubules are components of nephrons, the most important functional units of kidneys. They found acute tubular necrosis and other damage, and noted that this was common to other covid19 autopsy findings, perhaps unsurprisingly as these tubular cells present ACE2, the receptor for the virus. Dr Seheult then goes on to another study from Switzerland. This study looked at 639 critically ill covid10 patients, to determine which factors were most associated with survival or otherwise. So in general they found that this group suffered a ‘moderate’ mortality rate of 24%. To understand the findings will require quite a bit of medico-immunological knowledge, but here goes: they found that ‘PCT and IL-6 levels remained similar in ICU survivors and non-survivors throughout the ICU stay’. PCT is procalcitonin. According to Medscape:

Procalcitonin (PCT) is a biomarker that exhibits greater specificity than other proinflammatory markers (eg, cytokines) in identifying sepsis and can be used in the diagnosis of bacterial infections. Procalcitonin is also produced by the neuroendocrine cells of the lung and intestine and is released as an acute-phase reactant in response to inflammatory stimuli, especially those of bacterial origin. This raised procalcitonin level during inflammation is associated with bacterial endotoxin and inflammatory cytokines.

IL-6 is interleukin-6. An opinion article in Frontiers in Microbiology  entitled ‘The Role of Interleukin-6 During Viral Infections’ describes IL-6:

IL-6 is a pleiotropic cytokine produced in response to tissue damage and infections…  Multiple cell types including fibroblasts, keratinocytes, mesangial cells, vascular endothelial cells, mast cells, macrophages, dendritic cells, and T and B cells are associated with the production of this cytokine….

Pleiotropic cytokines – a cytokine is a type of small protein – affect the activity of multiple cell types. The complex pleiotropic nature of IL-6 unsurprisingly implicates it in both pro-inflammatory and anti-inflammatory effects. So, PCT and Il-6 levels remained similar for these study subjects, but ‘CRP, creatinine, troponin, D-dimer, lactate, neutrophil count, P/F diverged within the first seven days.’  Okay, C-reactive protein (CRP) is produced in the liver, from which it enters the bloodstream, and its levels ‘start to increase very soon after any inflammation or infection affects the body’, according to Australia’s healthdirect website. Creatinine is a waste product found in everyone’s bloodstream, and it’s produced by muscle metabolism. It’s generally filtered out by the kidneys. Too much blood creatinine may be a sign of kidney dysfunction. D-Dimer, the fibrin degradation product, always contains ‘two D fragments of the fibrin product joined by a cross-link’. I won’t try to explain much further at present. Neutrophils, remember, are infection-fighting white blood cells, and P/F ratio, aka PaO2/FiO2 ratio, is, briefly, an assessment of lung function. So with that, and some more, the study looked at levels of different markers most associated with mortality. To quote from the study: 

In contrast to risk factors in hospitalised patients reported in other studies, the main mortality predictors in these critically ill patients were markers of oxygenation deficit, renal and microvascular dysfunction, and coagulatory activation. Elevated risk of bloodstream infections underscores the need to exercise caution with off-label therapies. 

Canto: That last point seems important- it’s all about the blood. Or mostly..?

Jacinta: They presented a number of graphs which Dr Seheult interprets for us, but basically they are all likely to mark higher levels of microthrombi in the patients who died, and this seemed more clearly so in the D-dimer levels. High lactate levels are a sign of anaerobic metabolism, a problem with oxygenation. Ischemic heart disease was also measured, and this has to do with narrowing of the arteries. So blood oxygenation, or lack thereof, and coagulation, which can happen just about anywhere, seems to be happening early, leading to a wide range of symptoms, especially in patients with comorbidities, some of them previously undetected. 

Canto: So we’re moving on to update 96, which starts again with thrombosis due to endothelial damage causing increased production or release of von Willibrand factor (VWF).

Jacinta: Yes, and they’re apparently finding that different blood groups or types – and that’s a topic we could spend a lot of time on – affect the level and activity of VWF. As do other factors, according to Russian researcher Anna Aksenova:

The level and activity of VWF in the blood in people can be different. The lowest values are associated with von Willebrand disease. It is a hereditary blood disease that is characterized by spontaneous bleeding. Additionally, it differs markedly among healthy people. For example, it is higher among: African Americans than among Europeans; in men than in women; in adults than in children; and in the elderly than in middle-aged people. Also, academic papers have described the VWF and blood group relationship—its level is lower among people with blood group 0, and is higher among those with blood group A. The different amount and activity of VWF in people with different blood groups has a very interesting explanation: this protein is modified by oligosaccharide chains of antigenic determinants of the AB0 system (one of the blood group systems), and this affects its stability and activity.

She points out that ‘to date, the way in which the level of VWF is regulated in the blood has not yet been fully studied’, and then she describes some of what we do know, that it’s stored in special organelles (Weibel-Palade bodies) from where it’s secreted in multimeric form. She argues that, in order to determine the level of involvement of VWF in the progress of covid19, ‘large scale and comprehensive research’ needs to be carried out. Another article which is looking at emergency covid19 treatment has the title ‘targeting raised VWF levels and macrophage activation in severe covid19: consider low volume plasma exchange and low dose steroid’. It points out that VWF is such a large protein that it can only really be removed from the body through plasma exchange. This may be a way to reduce thrombosis in serious cases. Another interesting commentary piece is titled ‘microthrombotic complications of covid19 are likely due to embolism of circulating endothelial-derived ultralarge von Willebrand Factor (eULVWF) decorated-platelet strings’. 

Canto: An embolism being a blockage, caused by an embolus. That embolus could be a blood clot (a thrombus) or a fat globule or an air or gas bubble. 

Jacinta: Yes, and VWF can come in these long strings of platelets. In fact the platelets adhere to the strings. Anyway, that’ll do for now. We’ll go on about ivermectin and the Moderna vaccine trials next time. 

References

Coronavirus Pandemic Update 95: Widespread Clotting on Autopsy; New COVID-19 Prognostic Data

Coronavirus Pandemic Update 96: RNA Vaccine; Ivermectin; von Willebrand Factor and COVID-19

https://labtestsonline.org/tests/troponin#:~:text=Troponins%20are%20a%20group%20of,to%20help%20detect%20heart%20injury.

https://www1.racgp.org.au/ajgp/coronavirus/cardiovascular-conditions-and-covid-19#:~:text=Elevated%20biomarkers%3A%20Troponin%20and%20natriuretic,a%20marker%20for%20cardiac%20injury.&text=The%20cause%20of%20troponin%20elevation%20in%20serious%20infection%20is%20multifactorial.

https://www.medscape.com/answers/2096589-179642/what-is-procalcitonin-pct

https://www.frontiersin.org/articles/10.3389/fmicb.2019.01057/full

https://www.medicinenet.com/script/main/art.asp?articlekey=26197

https://www.healthdirect.gov.au/c-reactive-protein-CRP-test

https://medicalxpress.com/news/2020-07-complications-covid-von-willebrand-factor.html

Written by stewart henderson

September 6, 2020 at 1:44 pm

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

leave a comment »

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 on oxidative stress and covid-19

leave a comment »

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