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Photocatalyst-based ViraMap platform maps SARS-CoV-2 spike protein interactions on the cell surface – News-Medical.Net



In a recent study posted to the bioRxiv* preprint server, researchers performed the photocatalytic mapping of the severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) spike protein-host cell membrane interactions.

Study: High resolution photocatalytic mapping of SARS-CoV-2 Spike protein-host cell membrane interactions. Image Credit: Kjpargeter/Shutterstock

Understanding viral entrance and pathogenicity can be improved by identifying protein habitats at the virus-host cell interface. The virus responsible for the ongoing coronavirus disease 2019 (COVID-19) pandemic, SARS-CoV-2, uses the angiotensin-converting enzyme-2 (ACE2) protein as a primary receptor. However, the role of other cellular proteins in the entrance process is uncertain.

About the study

In the present study, the team developed a viral-host protein microenvironment mapping technology (ViraMap) using iridium photocatalysts (IrPC) conjugated to SARS-CoV-2 spike protein for visible-light-driven proximity labeling on host cells.

The team generated spike protein photocatalyst conjugates for targeted labeling on ACE2-expressing cells to analyze the interactions in the SARS-CoV-2 spike-host cell surface microenvironment using ViraMap. The SARS-CoV-2 spike protein trimer and its associated variants, including the SARS-CoV-1 spike and SARS-CoV-2 D614G spike, were chosen for conjugation with IrPC.

Further, the researchers performed targeted labeling on the cells by exposing them to spike-IrPC conjugates in the presence of a biotin-diazirine probe. This was followed by blue light irradiation and subsequent monitoring with flow cytometry and confocal imaging analysis. To selectively label host cell surface contacts and avoid cellular internalization of spike-IrPC conjugates, HEK293T+ACE2 cells were treated with spike-IrPC at 4 °C.

The team assessed cell surface binding in the presence of free ACE2 protein to establish that the spike-IrPC compound preserved its affinity for ACE2 (ACE2-Fc). For cell surface proximity labeling on HEK293T+ACE2 cells, three spike-IrPC variants such as SARS-CoV-1, SARS-CoV-2, and SARS-CoV-2 D614G spike proteins, were employed.

Furthermore, the team used a primary-secondary antibody combination comprising an anti-ACE2 primary antibody and an antibody-photocatalyst conjugate for targeted labeling. After extraction from the cell membrane fraction and streptavidin enrichment, the biotinylated host proteins from these targeted labeling studies were quantitatively characterized using a tandem mass tag (TMT)-based liquid chromatography–mass spectrometry (LC-MS)/MS quantification.


When compared to a non-binding antibody immunoglobulin G (IgG) control conjugated with IrPC, flow cytometry analysis revealed a clear change in the biotinylation signal for spike-IrPC variant conjugates-treated cells. Compared to the mouse-IgG-IrPC control, confocal imaging of spike-IrPC induced labeling revealed biotin localisation to the host cell surface environment. Furthermore, because the recombinant spike construct had a poly-His-tag, cellular biotinylation might be achieved through targeted labeling with an anti-His antibody IrPC conjugate.

IrPC-conjugated SARS-CoV-1, SARS-CoV-2, SARS-CoV-2 D614G spike proteins, as well as anti-ACE2 primary/secondary antibody systems resulted in statistically significant enrichment of distinct groups of cellular proteins compared to the IrPC isotype conjugate as a negative control. Using the SAINTexpress and mass spectrometry interaction statistics (MiST) scoring algorithms, the team identified 96 high-confidence enriched proteins across all spike-IrPC labeling studies.

Almost 23 of the 96 high-confidence enriched proteins were shared by all three spike variants, whereas the remaining two, 25, and 46 enriched proteins were unique to the SARS-CoV-1, SARS-CoV-2, and SARS-CoV-2 D614G spike-IrPC spikes, respectively. The gene ontology enrichment analysis of the 23 enriched proteins found a significant relationship with viral entrance, followed by immune cell differentiation and co-stimulation activities. Furthermore, 70% of the enriched proteins had a high abundance in most human tissues, including the lung, kidney, gastrointestinal tract, fat, cardiac muscle, soft tissues, reproductive tissues, and brain.

Out of 23, a total of 10 high-confidence enriched proteins could be divided into two groups based on whether or not they showed co-enrichment with the anti-ACE2 primary/secondary antibody system. The first class of cell surface receptors included ACE2, neuropilin 1 (NRP1), prostaglandin F2 receptor negative (PTGFRN), and neurogenic locus notch homolog protein 2 (NOTCH-2). Evidence of PTGFRN protein and messenger ribonucleic acid (mRNA) expression in a wide range of tissue cells, particularly cardiomyocytes and lung fibroblasts, implied that these cell types are highly susceptible to SARS-CoV-2 via various entry points. Galectins were the second set of host proteins found in all three spike-IrPC labeling assays.

The results of the targeted labeling experiments point to putative cell surface co-receptors involved in viral entry and antiviral immunity. As a result, functional experiments were performed to assess which of the identified host proteins are involved in SARS-CoV-2 entrance into cells. For functional validation, the study focused on the ten overlapping proteins shared by all three spike-IrPC variants. The clustered regularly interspaced short palindromic repeats /Cas9-mediated knockdown (KD) only reduced ACE2 abundance by 30%. Although there was a 40% reduction in protein expression relative to controls, there was no significant change in pseudoparticle entrance following NRP1 KD.

Overall, the study findings showed that the ViraMap technology successfully facilitated the assessment of known as well as unknown virus-host protein interactions that occur on the outer cell membrane of the host cell using cell surface recognition targeting modalities.

*Important notice

bioRxiv 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:

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Using artificial sweeteners may raise the risk of heart disease, study shows – Prestige Online Malaysia




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Artificial sweeteners may increase risk of heart disease, new study shows




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‘Debilitating’ heart palpitations could be sign of Long Covid – do you have the condition? – Express



Heart palpitations can be a sign of several different problems – both mental and physical. Often, they’re caused by stress and anxiety. But over the course of the last few years, Long Covid has reportedly caused palpitations. A recent study has explored why this might happen.

Long Covid is when people suffer ongoing symptoms of Covid, 12 weeks after infection.

Some people with the condition have struggled with heart palpitations, chest pain, dizziness, or feeling faint.

Researchers, observing their patients, have concluded that these symptoms could be caused by problems with the autonomic nervous system – the part of your nervous system that monitors automatic activities such as heart rate, breathing, and blood pressure.

Doctors and nurses at Hammersmith Hospital and Imperial College London believe that the “debilitating” palpitations and other symptoms were caused by “orthostatic intolerance syndrome”.

READ MORE: Princess Beatrice’s ongoing difficulty with ‘muddled’ thoughts swirling in her head

Orthostatic intolerance syndromes are when moving from a sitting or lying position to an upright position causes a low blood pressure in your arteries.

The British Heart Foundation explains: “When a healthy person stands up, some of the blood in the body will flow downwards with the pull of gravity.

“The body responds to prevent blood pressure falling – blood vessels narrow and there is a slight increase in heart rate.

“But in people with orthostatic problems, these automatic changes don’t happen.


“Moving to an upright position causes a drop in the blood supply to the heart and brain and a fast heart rate as the body tries to compensate.”

For people that struggle with these issues with moving to an upright position, the study by Hammersmith Hospital and Imperial College London made recommendations.

It suggested: “Non-upright exercise such as cycling on a recumbent exercise bike and swimming are encouraged.”

It added: “The patient should be advised on rising cautiously from a lying or seated position and avoiding exacerbating factors such as prolonged standing, warm environments, and dehydration.”

READ MORE: Princess Beatrice’s ongoing difficulty with ‘muddled’ thoughts swirling in her head

Shingles, memory loss, tinnitus, itchy skin, and tremors were among the more abnormal symptoms experienced.

Some studies have suggested that long Covid is an autoimmune disease, similar to Parkinson’s disease. An autoimmune disease is when the body’s immune system attacks itself.

The body cannot tell the difference between your own cells and foreign cells so causes the body to attack healthy cells.

According to one small study from 2021, 44 percent of long Covid patients involved had high levels of a type of antibody connected with other autoimmune diseases and lupus.

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Genomic testing and medico-legal risk | InSight+ – MJA Insight



INTEGRATING genomics into mainstream health care has many benefits, but also presents challenges. One challenge is ensuring that doctors and patients can safely and sustainably benefit from genomic medicine without increased medico-legal risk.

While the number of legal cases in Australia is still relatively small, this moment in time represents an opportunity to develop tools, training and support so doctors and their patients can safely and sustainably benefit from the advancing capabilities without increased medico-legal risk.

Over the past 2 years, we have been exploring the practical and medico-legal risks associated with genomic medicine, particularly in the context of antenatal care. Through our work, we identified that informed consent to test and workforce and funding issues are significant barriers to more effective use of genomics in health care. What we are seeing and how these medico-legal risks can be addressed through tools, training and support are explored below.

Informed consent to test

At the centre of consultations involving genomic medicine are individuals and families grappling with complex and often life-altering decisions. Many people need support both to understand the issues involved and to deal with the consequences of their decisions.

The consent process for people considering genomic testing presents specific challenges compared with other testing and treatments.

Advancing technologies and the increasing availability of expanded screening and testing mean that the practitioner and the pregnant person need to discuss and understand more complex information and face value-laden choices even before deciding whether to go ahead with a test.

Concepts such as “chance” of a condition developing (and the severity of that condition) or “variants of uncertain significance” are somewhat unique to the field of genomics and can be difficult for doctors to explain and for patients to understand.

To put this issue into the antenatal context, guidelines from the Royal Australian and New Zealand College of Obstetricians and Gynaecologists and the Royal Australian College of General Practitioners (RACGP) state that information on carrier screening for the most frequent autosomal or X-linked genetic conditions should be offered to all women planning a pregnancy or in the first trimester of pregnancy. The Colleges also state that all women should be provided with information and have timely access to screening tests for fetal chromosome and genetic conditions.

Guidelines play an important role in outlining the standard of care expected of doctors. Therefore, it is incumbent on practitioners working in this field to be equipped with the knowledge, expertise and tools to have comprehensive and comprehensible consent discussions with their patients about genomic testing.

An additional challenge for the consent process is that technology often develops at a much quicker pace than laws and guidelines. The emergence of expanded screening for autosomal and X-linked conditions and for fetal chromosome conditions can present challenges for doctors and their patients. “Expanded” screening can test for over a thousand conditions, many of which would have very little impact on a person. Explaining the potential impact of a variant of uncertain significance in terms of the chance of a condition developing in a person is inherently difficult.

The well established principles of consent require a doctor to give the person enough information for them to make an informed decision about the test or treatment being offered. This is a patient-centred test, requiring a doctor to discuss not only what a reasonable person in the patient’s position would want to know but also what the particular patient wants to know. The doctor is also ethically and legally obliged to recognise whether the patient has understood this information.

It is also important to ensure that a patient’s choice to have genomic testing is voluntary. An article published in the MJA in 2021 noted that patients may feel foolish to reject a test that is on offer and that a “normalisation” of testing has begun.

This was echoed in a recent Avant webinar, where medical and legal experts discussed issues of genetic testing and pregnancy. There was sentiment from the doctors who attended the webinar that the concept of voluntariness in genetic testing in pregnancy may be eroded by an “expectation” or social “normalisation” that women will accept each test offered. One participant said:

“… practitioners need to be careful to convey respect to women who choose against genetic screening in pregnancy. There is a risk that repeated discussion and repeated offers of testing sends a message that the woman has made an incorrect decision.”

We support the development of standard information and forms to be used as part of the consent process for genetic testing in pregnancy. This would help patients to understand what is involved and assist in their decision making, as well as alleviate some of the medico-legal risk for doctors.

Standard information used in consent discussions also helps to ensure information to patients is balanced, evidence-based and informative. This would lessen the burden on practitioners and means there is less room for allegations of bias in the information they were providing. It also has the potential to make clearer the blurred lines between clinical and research use of genomic test findings, which could, for example, add to the body of information that informs the significance of a rare variant that may have been identified.

It is therefore pleasing to see the National Model of Consent for Clinical Genomics Testing developed by the NSW Ministry of Health in accordance with Action 4A of the implementation plan for the National Health Genomics Policy Framework, on behalf of the Australian Health Minister’s Advisory Council Project Reference Group on Health Genomics. The Clinical Genomic Testing Consent Form developed to support the model can be adapted across jurisdictions and can be used in conjunction with patient information materials to assist patient understanding during the consent discussion. Consent material has also been developed by Australian Genomics.

Workforce and funding issues

Avant has also identified significant workforce pressures and gaps in funding for timely and sustainable care in genomic medicine.

We welcome the Medicare funding announced for genetic carrier screening for all couples, testing for carriers of cystic fibrosis, spinal muscular atrophy and fragile X syndrome from 2023. However, as genetic carrier screening becomes more accessible, there are some practical and medico-legal issues to consider.

With Medicare-funded testing available, the standards for carrier screening could change, with testing becoming the standard professional practice of doctors. More doctors will need to be equipped with the knowledge and communication skills to discuss carrier screening with their patients.

The complexity of these discussions should be reflected in appropriate Medicare item numbers for the consultations. The RACGP is calling for Medicare Benefits Schedule amendments to better support longer consultations for antenatal care that would support the use of time-based level C and D Medicare items for longer antenatal consultations. Such changes would better allow time for the detailed discussions that are required to take place around genetic screening. An alternative would be to introduce an additional cobilled item number specific to genomic testing and counselling which would recognise the complexity of these discussions with patients.

We believe Medicare-funded testing is likely to lead to more testing and, therefore, more people receiving positive results. Doctors we spoke with noted the importance of being able to engage with genetic specialists and counsellors. They reported that patients may need genomic expertise and mental health support. Currently, there can be lengthy delays and challenges accessing these practitioners. Time is of the essence in pregnancy, and workforce constraints could add pressure and stress to patients and doctors. Also, it is not clear that the newly announced funding for carrier screening includes costs associated with genetic counselling to discuss the results.

Research from the Deeble Institute has found that the workforce shortages in clinical genetics and genetic counselling are partly due to constraints on numbers of professionals being trained, which in turn are linked to limited workplace rotations available. While this may be the case with many specialties, particularly following the COVID-19 pandemic and workforce shortages due to burnout, the increased availability of and funding for genomic testing means that workforce shortages in clinical genetics and genetic counselling need to be addressed as a matter of urgency. This will help ensure that patients are given the information they need and alleviate doctors’ concerns about medico-legal risk.

The potential for genomic medicine is great, but the challenges it presents are also significant. Keeping up to date with rapidly advancing knowledge and ensuring patients have enough information so they are making informed decisions about testing are key concerns of doctors practising in this area.

Now is the time to develop tools, training and support so doctors and their patients can safely and sustainably benefit from the advancing capabilities without increased medico-legal risk.

Rocky Ruperto is a solicitor and Legal and Policy Adviser at Avant.

Dr Sally Parsons is a Medical Advisor and Claims Manager at Avant and Clinical Director and GP in Adelaide. Avant is Australia’s largest medical defence organisation.

Disclaimer: This article is intended to provide commentary and general information. It does not constitute legal or medical advice. You should seek legal or other professional advice before relying on any content, and practise proper clinical decision making with regard to the individual circumstances.

The statements or opinions expressed in this article reflect the views of the authors and do not necessarily represent the official policy of the AMA, the MJA or InSight+ unless so stated.

Subscribe to the free InSight+ weekly newsletter here. It is available to all readers, not just registered medical practitioners.

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