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As the Bushfire Royal Commission investigates the deadly "Black Summer" and how it could have been prevented, research from the Climate the ARC Centre of Excellence for Climate Extremes (CLEX) and Climate Change Research Centre at the University of New South Wales shows how climate change may alter prescribed burning in the future across Australia.

Unexpectedly, it is not all bad news, but as the coming century progresses there are definitely changes ahead for our firefighters in Australia when it comes to preparing for a fire season, especially along the east coast of Australia.

Most prescribed burning along this densely populated coastal region takes place in March, April and May, when conditions are safest for hazard reduction efforts. In the future as climate change takes hold and conditions change, climate models show that the burning windows for these months decline over many regions. However, at the same time new burning windows open up from June to August and even into the early parts of September.

"Our research shows that, considering the whole year, the number of prescribed burning days along the east coast of Australia will remain the same and may even increase in some cases but the timing of the burning windows shifts," said lead author Dr Giovanni Di Virgilio from the Climate Change Research Centre at the University of New South Wales.

"This is because the multiple conditions that make for a good day for prescribed burning - like mild and still days - shift to later in winter. It highlights the vagaries of climate change."

To get their results the researchers looked at five variables - maximum temperature, relative humidity, wind speed, fuel moisture, and the McArthur forest fire danger index. Assuming emissions continue to rise at their current pace, they modelled how these would combine at a regional level to create prescribed burning windows in the period 2060-2079. They then compared these results to observations from 1990-2009 to find out how they differed.

Changes to these burning windows occurred over many parts of Australia but it was not uniform across space and time. Much of the Australian east coast and South Australia saw seasonal shifts in burning windows amounting to around a 50% reduction of burning windows in March to May but an increase from June to October.

Meanwhile much of Victoria and in particular the southern regions saw an increase in burning windows during April to May and in some parts of the State through September and October as well.

Only the east Queensland coast saw a general overall reduction in prescribed burn days from April through to October.

While this is mostly good news for those involved in the bushfire mitigation process, a complicating factor arising from previous CLEX research is that the conditions which create suitable prescribed burning windows often correspond with conditions that form inversion layers.

"Our past research showed climate change makes inversion layers in the winter months even more common and these can trap the smoke and fine particulate matter emitted from fires close to the ground. This has adverse impacts on many groups, such as asthmatics and those affected by high concentrations of air pollution,' said co-author from the ARC Centre of Excellence for Climate Extremes Prof Jason Evans.

"It is clear that climate change will influence future hazard reduction burning activities, shifting them to new times of the year. But it is fortunate that for the next century at least, the available days where we can perform these important activities before a fire season will, for the most part, remain similar or even increase."

A research team from Japan has developed a single-camera machine vision algorithm, making it possible for lightweight hovering indoor robots to guide themselves by identifying and interpreting reference points on a tiled floor. The technology opens the door to a new breed of functional, low-cost drones with potentially wide-ranging uses.

The team, led by Chinthaka Premachandra, an Associate Professor in the Department of Electronic Engineering at Shibaura Institute of Technology, published its findings in IEEE/CAA Journal of Automatica Sinica, a joint publication of the Institute of Electrical and Electronics Engineers (IEEE) and the Chinese Association of Automation (CAA).

Since GPS signals are too weak to penetrate most structures, indoor drones must rely on environmental queues, which are typically visual. A drone designed for indoor use is likely to be smaller and lighter than an outdoor drone, according to Premachandra. "We considered different hardware options, including laser rangefinders," he said. "But rangefinders are too heavy, and infrared and ultrasonic sensors suffer from low precision. So that led us to using a camera as the robot's visual sensor. If you think of the camera in your cell phone, that gives you an idea of just how small and light they can be."

Premachandra's research team aimed to design the guidance algorithm to be as simple as possible, allowing for a small, inexpensive microprocessor. The team used the Raspberry pi3, an open-source computing platform that weighs approximately 45 grams.

Their study prototype had a single downward facing camera with intentionally low resolution - only 80 by 80 pixels. "Our robot only needed to distinguish its direction of motion and identify corners. From there, our algorithm allows it to extrapolate its position in the room, helping it avoid contacting the walls," Premachandra said.

The team's program worked by taking each 80x80 image through a series of simple processing steps ending in a black and white grid, making it easier to quickly identify motion along the X and Y planes.

The guidance method is limited because it relies on a room with a tile floor and predictable patterns. Premachandra said next steps in research into lightweight, autonomous-hovering, indoor robots might include adapting the technology for infrared cameras so they could function in the dark, as well as adding a second camera so the robot could visually determine both its X,Y position and its altitude in the room.

"There are many potential applications," Premachandra said. "Hovering indoor robots may be useful in warehouses, distribution centers, and industrial applications to remotely monitor safety."

Professor Kimoon Kim's research group at POSTECH has developed a highly pure and efficient technique for purifying antiviral and anti-cancer protein therapeutics using molecular affinity interaction.

With the COVID-19 showing no signs of slowing down around the world, the development of a vaccine seems to be the sole solution to end the pandemic. However, even if a vaccine is developed, the task of manufacturing well-refined drug molecules, which treat a number of other comorbid conditions, remains elusive. Besides COVID-19, there are growing concerns for developing a source treatment protocol to deal with new infectious diseases that may find their way into our lives.

A joint research team made up of Professor Kimoon Kim (POSTECH University Professor of Department of Chemistry at POSTECH, Director of Center for Self-assembly and Complexity (CSC) and of Institute for Basic Science (IBS)) and Dr. Kyeng Min Park (Research fellow and Group leader at CSC and IBS) have together developed a source technology using the affinity interaction of cucurbiturils to purify recombinant therapeutic proteins used as antiviral drugs or as anti-cancer agents with high efficiency and high purity. The research findings were published online in Nature Biomedical Engineering (DOI: 10.1038/s41551-020-0589-7) on July 20.

Recombinant DNA technology uses biotechnological techniques to insert genes from one organism into another and activate them to change its genetic trait. Using this method, a vaccine can be developed to exterminate pathogenic microorganisms or weaken their toxicity by expressing all or part of the proteins. For the mass-production of the hormones, antibodies or vaccines made from this process, the proteins must be purified.

Until now, therapeutic protein purification technology has used protein-based affinity columns. However, they face financial and technological difficulties as the materials are costly, their storage or reuse is not efficient and their suitability and efficiency depends on the properties of each therapeutic protein.

The researchers succeeded in purifying various types of therapeutic proteins expressed in cells by harnessing the molecular affinity interaction using a synthetic host molecule cucurbit[7]uril (or CB[7] in short), which was first discovered by the same research group. The molecular affinity principle, a key element in the development of the purification technique, is dependent on the high-affinity and controllable host-guest interaction between CB[7] and guests such as adamantane.

The new purification technology has succeeded in purifying a monoclonal antibody drug, Herceptin (breast cancer treatment), as well as much smaller Interferon alpha (leukemia treatment) with high efficiency and high purity.

In particular, by applying small and stable synthetic molecules, the team succeeded in securing the manufacturability, sterilization, and recyclability of purified materials in a stable manner as well as increasing the purity and productivity of purified protein therapeutics. In addition, introducing adamantine (or AdA in short) to therapeutic proteins through genetic regulation and enzyme treatment can purify them regardless of their size or type.

This technique can be applied to most recombinant therapeutic proteins, including antibodies or fusion proteins that effectively prevent or treat fatal diseases such as viral infections or cancer, and is highly efficient and reusable. Furthermore, it is applicable to a wide variety of therapeutic proteins used in the development of vaccines or treatments which will speed up their production.

The research was conducted with the support from the Institute of Basic Science.

Depression in mothers during and after pregnancy increased the odds of depression in offspring during adolescence and adulthood by 70%, according to a new study by researchers at The University of Texas Health Science Center at Houston (UTHealth).

The systematic review was recently published in JAMA Network Open. It is the first study to examine the effects of maternal depression on children age 12 and older.

The authors note that 10-20% of mothers experience perinatal depression, which refers to the time during pregnancy (antenatal) or within the first year after birth (postnatal). Perinatal depression has been associated with reduced growth rates, malnutrition, and an increased risk of childhood health problems and obesity.

"There are a lot of studies that look at how perinatal depression affects a child's growth or emotional well-being, but we wanted to look at how it affects offspring later in life," said Vaishali Tirumalaraju, MBBS, a resident with the Louis A. Faillace, MD, Department of Psychiatry and Behavioral Sciences at McGovern Medical School at UTHealth and one of the study's first authors.

Researchers examined all published studies on the topic, zeroing in on six major cohorts following long-term symptoms in more than 15,000 children age 12 or older.

"We found that the offspring of mothers who had perinatal depression, especially antenatal depression, had a greater chance of having depression when they grow up," said Sudhakar Selvaraj, MD, PhD, an assistant professor of psychiatry with McGovern Medical School, the director of the Depression Research Program, and the study's senior author. "This is important because it shows perinatal depression not only negatively affects the mother, but also has a lasting negative impact on the child."

This research comes during a time of increased stress and strain on mental health, as the COVID-19 pandemic poses challenges for new and expecting mothers.

"Pregnancies during these times are particularly stressful given the social situation and physical distancing guidelines in place. There are a lot of moms without support who have to isolate, and many are handling extra stressors with limited resources," Tirumalaraju said.

"Stress can cause changes in eating habits, less desire to exercise, and overall less healthy behaviors that can lead to depression," said Selvaraj, a psychiatrist with UT Physicians, the clinical practice of McGovern Medical School. "Due to stigma, depression is hard to track because often mothers don't want to report that they are sad or having trouble mentally. Our study shows the potential long-term impact of depression in offspring and therefore the need for proper screening so that moms who could benefit from counseling and/or medication can be provided the resources they need."

Selvaraj is part of the UTHealth Women's Mental Health Program, which provides access to specialists in psychology and psychiatry in certain UT Physicians Women's Centers, simplifying the process for women to seek help for perinatal depression and other mental health conditions.

Study authors say more research into the factors involved in depression risk transmission and assessments of how to reduce this risk could help develop future strategies to mitigate depressive disorders in pregnancy.

HOUSTON - (July 27, 2020) - Income is inextricably linked to access to education in America and it has been for a century, according to a new study from researchers at Stanford University and Rice University.

"A century of educational inequality in the United States," published July 27 in Proceedings of the National Academy of Sciences, examines the link between education and income dating back to the early the 20th century. The research draws upon a dozen nationally representative datasets on college enrollment and completion between 1908 and 1995 as well as tax data from more recent years. It is one of the first studies to examine this link over such an extended period of time.

Researchers Michelle Jackson from Stanford and Brian Holzman from Rice's Houston Education Research Consortium, part of the Kinder Institute for Urban Research and School of Social Sciences, found that income and educational inequality moved in lockstep with one another throughout the 20th century. The authors said previous studies of this topic, which haven't examined data going so far back in time, did not reveal such a strong link.

Their paper detailed how inequality in college enrollment and completion rose in the 1930s and 1940s amid rising income inequality; was low for Americans born in the late 1950s and 1960s, when income inequality was low; and rose again for Americans born in the late 1980s, when income inequality peaked. This U-turn indicates the nation is experiencing levels of collegiate inequality not seen for generations, the authors wrote.

"Long story short, the findings reveal that longstanding worries about income inequality and its relationship to college opportunity are warranted," Holzman said.

One notable exception was during the Vietnam War. For young people at risk of serving in the war, collegiate inequality was high while income inequality was low. During this period, inequality in college enrollment and completion was significantly higher among men than women, suggesting a bona fide "Vietnam War effect," according to the paper.

The researchers hope the paper will further demonstrate the systemic nature of the link between income and education and inform future work on increasing educational opportunities, particularly for disadvantaged people.

What The Article Says: The presence of myocardial injury in patients recently recovered from COVID-19 are evaluated in this observational study.

Authors: Eike Nagel, M.D., of University Hospital Frankfurt in Germany, is the corresponding author.

To access the embargoed study: Visit our For The Media website at this link https://media.jamanetwork.com/

(doi:10.1001/jamacardio.2020.3557)

Editor's Note: The article includes conflict of interest and funding/support disclosures. Please see the article for additional information, including other authors, author contributions and affiliations, conflicts of interest and financial disclosures, and funding and support.

What The Study Did: The presence of SARS-CoV-2 in myocardial tissue from autopsy cases is evaluated in this observational study.

Authors: Dirk Westermann, M.D., of the University Heart and Vascular Centre Hamburg in Germany, is the corresponding author.

To access the embargoed study: Visit our For The Media website at this link https://media.jamanetwork.com/

(doi:10.1001/jamacardio.2020.3551)

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What The Study Did: The characteristics and expected strength of evidence of COVID-19 studies registered on ClinicalTrials.gov are evaluated in this observational study.

Authors: Mintu Turakhia, M.D., M.A.S., of the Stanford University School of Medicine in Palo Alto, California, is the corresponding author.

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(doi:10.1001/jamainternmed.2020.2904)

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The latest round of offshore wind farms to be built in the UK could reduce household energy bills by producing electricity very cheaply.

Renewable energy projects, including onshore and offshore wind and solar farms, have so far been subsidised by government support schemes. This has led to some to complain that clean energy is pushing up bills.

However, the most recently approved offshore wind projects will most likely operate with 'negative subsidies' - paying money back to the government. The money will go towards reducing household energy bills as the offshore wind farms start producing power in the mid-2020s.

This is the conclusion of an analysis by an international team led by Imperial College London researchers published today in Nature Energy.

Lead researcher Dr Malte Jansen, from the Centre for Environmental Policy at Imperial, said: "Offshore wind power will soon be so cheap to produce that it will undercut fossil-fuelled power stations and may be the cheapest form of energy for the UK. Energy subsidies used to push up energy bills, but within a few years cheap renewable energy will see them brought down for the first time. This is an astonishing development."

Negative subsidies

The analysis for five countries in Europe, including the UK, focused on a series of government auctions for offshore wind farms between February 2015 and September 2019. Companies that want to build wind farms bid in the auctions by stating the price at which they will sell the energy they produce to the government.

These are known as 'contracts for difference' or CfDs. If a company's bid is higher than the wholesale electricity price on the UK market once the wind farm is up and running, then the company will receive a subsidy from the government to top up the price.

However, if the stated price is less than the wholesale price, then the company will pay the government back the difference. This payback is then passed through to consumer's energy bills, reducing the amount that homes and businesses will pay for electricity.

The UK's September 2019 auction made the headlines as winning companies said they could build new offshore wind farms for around £40 per megawatt hour (MWh) of power. This was a new record set by these wind farms with bids 30 percent lower than just two years earlier.

While this was an impressive reduction, researchers could only speculate whether this meant offshore wind had become subsidy free or even subsidy negative, because that depends on how future wholesale electricity prices evolve.

The team analysed likely future electricity price trends and found that contracted price is very likely to be below the UK wholesale price over the lifetime that these wind farms would produce electricity, from the mid-2020s onwards.

The team say that these wind farms are likely to be built and run with these costs, since financing is now accessible at lower costs for such projects, owing to trust in the now mature technology.

A cheap tool for decarbonisation

The researchers analysed similar offshore wind auctions held by governments of five European countries. They found that Germany and the Netherlands have seen some zero-subsidy offshore wind farms winning auctions, but that the UK projects are likely to be the world's first negative-subsidy offshore wind farms.

Dr Iain Staffell, from the Centre for Environmental Policy at Imperial, said: "The price of offshore wind power has plummeted in only a matter of a decade, surprising many in the field. The UK auctions in September 2019 gave prices that were around one-third lower than those of the last round in 2017, and two-thirds lower than we saw in 2015.

"This amazing progress has been made possible by new technology, economies of scale and efficient supply chains around the North Sea, but also by a decade of concerted policymaking designed to reduce the risk for investing in offshore wind, which has made financing these huge billion-pound projects much cheaper.

"These new wind farms set the stage for the rapid expansion needed to meet the government's target of producing 30 percent of the UK's energy needs from offshore wind by 2030. Offshore wind will be pivotal in helping the UK, and more broadly the world, to reach net-zero carbon emissions with the added bonus of reducing consumers' energy bills."

Mega turbines and hydrogen fuels

One reason the price of offshore wind has fallen so rapidly is technology development, in particular the ability to build larger wind turbines further out at sea. Larger turbines can harness more wind energy and have access to more consistent wind speeds at higher altitudes.

The biggest wind turbines under construction have rotor diameters of 220 metres - twice the diameter of the London Eye. At the same time, wind farms are getting larger; the newest wind farm at Dogger Bank has the same installed capacity as Hinkley Point C and is expected to produce about two-thirds of its annual electricity.

The success of UK offshore windfarms, which are now primarily built in the Dogger Bank region of the North Sea, also means the UK has considerable skills and expertise than can be exported around the world.

The researchers also say this success means even more ambitious projects may now be attempted at offshore wind farms, such as producing hydrogen fuels using the wind power on site, out at sea. Hydrogen fuels could be another key technology in helping decarbonise the UK, by replacing petrol used in transportation and natural gas used for heating homes.

Researchers from Harvard University, the Centro Nacional de Análisis Genómico (CNAG) and the Centre for Genomic Regulation (CRG), describe the first technology able to visualize hundreds to potentially thousands of genomes at the same time under the microscope. The tech images genomes more cheaply, more quickly and increases range of visibility compared to currently available methods. The technique is described in Nature Methods.

Each human cell has two metres of genome condensed down into 10 microns within the cell nucleus. This blueprint of life folds to help genes make physical contact with other genes that may be located quite a distance away along the chromosome. This three-dimensional organisation is crucial for cell function, but its complexity and constant dynamism make it incredibly difficult to visualize. Imaging more than a handful of genes at the same time has been impossible, limiting researchers' ability to characterize how genomes function.

One of the most common ways of studying the genome is by using fluorescence in situ hybridization (FISH), which uses fluorescent probes to mark the presence or absence of specific DNA sequences on chromosomes. Scientists have made landmark discoveries such as how cells divide thanks to FISH, and still use it this day for medical applications or species identification, amongst other applications.

First developed in the 1980s, FISH is an ageing technology that can only visualise a handful of genes at the same time. Since then, new methods have been created that can image genomes at a high resolution, but map a very limited number of regions or chromosomes at a time.

Today authors of a a new study in Nature Methods describe OligoFISSEQ, a technology using new computational methods that overcomes these limitations. They used it to create three-dimensional maps of 66 genomic targets across six chromosomes in hundreds of cells, showcasing OligoFISSEQ's potential to visualize the entire genome at a molecular resolution out of reach until now.

"Up to this moment, seeing a large number of different genes at the same time under the microscope was impossible," says Marc A. Marti-Renom, co-lead author of the study and ICREA Professor at CNAG-CRG. "We combined existing sequencing technologies in a smart way so that we can see hundreds of genes by sequencing their targets under the microscope. Before OligoFISSEQ, reaching this number of genes at the same time was slow and expensive. It is like upgrading from a dial-up phoneline to fibre-optic internet and paying 40 times less for it."

The researchers also tested OligoFISSEQ by mapping all of the 46 regions along the length of the human X chromosome. The higher resolution of the technology revealed new patterns in the way the genome organizes itself, including that the length of the chromosome's arm is not correlated with its angle. The researchers hypothesise this may be indicative of cell type, cell state or cellular health or age.

"The resolution offered by OligoFISSEQ has huge potential to shed new light on patterns we could not see before", concludes Marti-Renom. "Because of the coverage it provides to study the genome, it is well suited for spotting what may seem like a minor, seemingly inconsequential change in one part of the nucleus that may have a 'butterfly effect' elsewhere. What may have previously have been thought of as random may turn out to be anything but. That is the power of this tool."

What The Study Did: Viral aerosol emissions from simulated individuals with asymptomatic to moderate COVID-19 are estimated in this mathematical modeling study.

Authors: Michael Riediker, of the Swiss Centre for Occupational and Environmental Health, in Winterthur, Switzerland, is the corresponding author.

To access the embargoed study:  Visit our For The Media website at this link https://media.jamanetwork.com/

(doi:10.1001/jamanetworkopen.2020.13807)

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About JAMA Network Open: JAMA Network Open is the new online-only open access general medical journal from the JAMA Network. On weekdays, the journal publishes peer-reviewed clinical research and commentary in more than 40 medical and health subject areas. Every article is free online from the day of publication.

What The Study Did: Rates of SARS-CoV-2 infection among asymptomatic health care workers and community residents in Texas are examined in this observational study.

Authors: Roberta L. Schwartz, Ph.D., of the Houston Methodist Academic Institute in Texas, is the corresponding author.

To access the embargoed study: Visit our For The Media website at this link https://media.jamanetwork.com/

(doi:10.1001/jamanetworkopen.2020.16451)

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About JAMA Network Open: JAMA Network Open is the new online-only open access general medical journal from the JAMA Network. On weekdays, the journal publishes peer-reviewed clinical research and commentary in more than 40 medical and health subject areas. Every article is free online from the day of publication.

Appendicitis is the most common cause for emergency abdominal surgery in childhood, affecting 80,000 children in the United States each year, but nonoperative treatment options are viable. A study performed by the Midwest Pediatric Surgery Consortium, led by Peter Minneci, MD, and Katherine Deans, MD, co-founders and directors of the Center for Surgical Outcomes Research at Nationwide Children's Hospital, and published online today in JAMA, found antibiotics alone successfully treated children with uncomplicated appendicitis and was associated with fewer disability days at one year.

Of 1,068 patients from 10 health centers enrolled in the study, 67.1% of those who elected to initially manage their care through antibiotics alone experienced no harmful side effects and did not later require an appendectomy by their one-year follow-up appointment. Patients in the non-operative group experienced an average of 6.6 disability days, compared to the 10.9 days in the surgery group. Non-operative management was also associated with fewer disability days for caregivers.

This research, funded by a Patient Centered Outcomes Research Institute (PCORI) grant, expands on an initial pilot study Drs. Minneci and Deans published in 2015, which first demonstrated the efficacy and safety of non-operative management of appendicitis in children by showing that children who were hospitalized for uncomplicated appendicitis--who experienced abdominal pain for no more than 48 hours, had a white blood cell count below 18,000 and underwent an ultrasound or CT scan to rule out rupture and to verify that their appendix was 1.1 centimeter thick or smaller with no evidence of an abscess or fecalith--and who elected initially to be treated with antibiotics could be successfully sent home without the use of traditional surgery.

"For surgery, patients need to go under general anesthesia, and there is 1-2% chance of a major complication and 5-10% chance of a minor complication," said Dr. Minneci, principal investigator of the studies with Dr. Deans. "And patients will definitely experience post-operative pain and disability. Treatment-related disability is important to kids, because it means missing activities in their lives that may directly affect their development and quality of life such as school, athletics and vacations."

Additionally, the study, which was designed to mimic clinical practice and used a decision aid to educate patients about the risks and benefits of each treatment option, found that both the patients who elected to undergo surgery and those who chose nonoperative care management with antibiotics alone had similar rates of complicated appendicitis, and reported similar health care satisfaction at 30 days and quality of life at 1 year.

Drs. Minneci and Deans said future research could study how to disseminate these results so that more patients can be informed of the two options and the risks and benefits of each. The decision aid and treatment protocols developed for this study were developed to minimize risks and can be easily translated into pediatric clinical practice.

"Culture change and rethinking how we treat patients is always hard," said Dr. Deans. "Right now, some of the standards for success among surgeons are different than among patients and families. Surgeons' tend to be passionate about operations, and an appendectomy is a well-tested and trusted procedure. However, some patients want to avoid surgery at all costs, and the results of our studies reflect the effectiveness of offering a non-operative management to patients and their families in clinical practice. This allows us to move away from a one-size-fits-all model of appendicitis care and treat each child based on his or her values and preferences."

MADISON -- T cells are the immune soldiers at the frontlines of the battle with infiltrating pathogens that seek to cause disease. A new study published in Nature Biomedical Engineering describes a novel label-free imaging technique that can differentiate active T cells from those off duty.

The method could help assess T cell involvement in immunotherapies for cancer treatment or autoimmune diseases.

"T cells have a metabolic switch that regulates their activity," says Melissa Skala, principal investigator at the Morgridge Institute and associate professor of biomedical engineering at UW-Madison. In a healthy individual, most T cells are in a quiescent state--they're inactive, but ready and waiting for the signal to join in active combat against an invading virus or bacteria.

"We wanted to test if our imaging technology could tell the difference between the quiescent T cells and activated T cells," says Alexandra Walsh, formerly an assistant scientist at the Morgridge Institute who is now an assistant professor of biomedical engineering at Texas A&M University.

Most methods for characterizing T cells are antibody-based, such as flow cytometry or immunohistochemistry. These require staining with antibodies or contrast agents, a process that is destructive to the cells.

Alternatively, Walsh and Skala's method detects autofluorescence from molecules within the cell that naturally emit light when imaged by a microscope paired with an infrared laser. This label-free process is non-damaging and doesn't alter the behavior of the cell. The technique could be adapted to image cells in a plate or dish, tissue samples, or even in vivo imaging of a complete organism.

"It's super novel," Skala says. "Most people aren't using these techniques--you don't see a lot of autofluorescence studies in immunology."

To validate their approach, the researchers acquired blood samples from healthy donors, isolated the T cells, and measured autofluorescence of NAD(P)H and FAD, two molecules that are involved in cellular metabolism.

"We kept some of the T cells in their quiescent state, and then we added antibodies to a group to activate them," says Walsh.

Images of the quiescent cells versus the activated cells revealed differences in metabolic function, most notably through a change in NAD(P)H autofluorescence in the activated T cell populations. They also observed that active T cells were slightly larger in size than quiescent cells.

The activation protocol and imaging capabilities will be useful for manufacturing the CAR-T cells used in immunotherapies, says Skala. These re-engineered T cells are often co-cultured with other cells, like cancer cells, to test their reactivity.

However, using additional harsh reagents or antibody-labels to further characterize the T cell is a bottleneck for CAR-T cell manufacturers. The autofluorescent approach provides an attractive way to perform those experiments by imaging the same cells across multiple timepoints in a way that's non-damaging.

"We showed that you can resolve temporal changes with our imaging technique," says Walsh. "We were able to see changes in the imaging endpoints within minutes after adding the activating antibodies."

Walsh adds that it would be difficult to see these dynamic changes using flow cytometry, since the time required for staining and incubation make it difficult to capture multiple timepoints.

The Skala lab plans to continue this line of investigation to better understand how a cancer patient's T cells might respond as the tumor grows or as they're treated with immunotherapies.

"These technologies could tell us something about tumors or about T cell manufacturing that we didn't know," adds Skala, "because previously we didn't have the methods to monitor T cell behavior over time.

While this new technique offers many advantages over traditional methods, there are still limitations. For one, autofluorescence imaging isn't very sensitive.

"We aren't relying on really specific labels, we're relying on the metabolism of the cells," Skala says. "That's only going to get you so far in differentiating the cell types."

Additionally, the technique requires experienced people to perform the microscopic imaging and analyze the data, says Walsh.

The Skala lab is working on developing a prototype to take the imaging capability of their large-scale microscope and translate it into a "box-sized" system.

"You won't have to be a specialized optical engineer to use it," Skala says. "That's the direction we're trying to go. We're trying to make it more accessible."

Mariel Mohns, mmohns@morgridge.org

Journal

Nature Biomedical Engineering

DOI

10.1038/s41551-020-0592-z

Two papers in Genome Research by the FANTOM Consortium have provided new insights into the core regulatory networks governing cell types in different vertebrate species, and the role of RNA as regulators of cell function and identity.

The FANTOM Consortium was established at RIKEN two decades ago to go beyond genomics and examine RNA--known as the transcriptome. Understanding the transcriptome is crucial for further advances in biology because although the cells in our bodies share the same genomic DNA, their diversity is attributed to their RNA make up, with more than 400 types defined and many more thought to exist. Thus, understanding how RNA is expressed is a key for grasping how each cell type establishes its distinctive function, morphology, and behavior by activating specific transcriptional programs. Both studies published today were based on the CAGE technology that was developed at RIKEN to profile the transcriptome using next-generation sequencers.

The first study (Alam et al.) compares transcriptome data from matching primary cell types in human, mouse, rat, dog, and chicken. While the group found that the transcriptome measured by CAGE for the same cell type differed markedly between species, they identified a core regulatory network defining each cell type that is common between species. In general, the genes encoding products involved in RNA biology in the cell nucleus were found to be activated consistently in the same cell type regardless of the species. According to Michiel de Hoon, the corresponding author of the paper, "We identified genes acting within the nucleus whose usage was conserved for 100's of millions of years of evolution. On the other hand, genes that primarily act in communication between cells had diverged and were being used differently in different species, implying that the distinctive phenotype of each species is to a great extent due to the specific way that cells in an organism communicate with each other."

The second study (Ramilowski J., Yip CW., et al.), part of FANTOM 6--the latest edition of the project--looked at human long non-coding RNAs, which outnumber protein-coding genes in mammals but whose function is still poorly understood. The researchers selectively targeted nearly 300 long non-coding RNAs for suppression in human fibroblast cells using an automated robotics system (Figure 1), and combined live cell imaging with CAGE to observe how cells respond at both the cellular and the molecular level. Jay Shin, one of the corresponding authors of this study, emphasized that "it was critical to automate our efforts as much as possible to reduce biases in our experimental design, and to quickly identify and correct any that remained." Based on the analysis, over 25 percent of long non-coding RNAs were found to affect cell growth and morphology, as well as cell migration, which is important in cancer. Surprisingly, targeting different isoforms (variants) of the same long non-coding RNA led to profoundly different cellular and molecular phenotypes, giving rise to the enticing conjecture that each long non-coding RNA isoform produced by a cell might have its own specific regulatory function.

According to Jordan Ramilowski, one of the first authors of the study, "Deep CAGE profiling of the molecular state of the cells after suppression of each long non-coding RNA allowed us to perform a functional analysis of long non-coding RNAs at an unprecedented level, and provides a valuable resource for a detailed investigating and understanding of the RNA biology and its potential application to enhancing human health."

Piero Carninci commented that "although this is still a pilot project, the results show involvement of lncRNAs in a broad variety of cellular processes and functions, which makes the case for extension of these studies to a broader number of cells and lncRNAs. We are excited to see that these RNAs, often considered 'junk' when discovered some 15 years ago, are often proven to be functional. We also believe that that the nomenclature should shift from 'non-coding' to terminology that better reflects their role, such as 'regulatory RNAs' or 'structural RNAs'."