The surprisingly strong effect could have implications for drug design and delivery.

The humble membranes that enclose our cells have a surprising superpower: They can push away nano-sized molecules that happen to approach them. A team including scientists at the National Institute of Standards and Technology (NIST) has figured out why, by using artificial membranes that mimic the behavior of natural ones. Their discovery could make a difference in how we design the many drug treatments that target our cells.

Cell membranes generate powerful electric field gradients that are largely responsible for repelling nano-sized particles like proteins from the surface of the cell — a repulsion that notably affects uncharged nanoparticles. In this schematic drawing, a negatively charged membrane (at top, in red) attracts small, positively charged molecules (purple circles), which crowd the membrane and push away a far larger, neutral nanoparticle (pink). Credit: N. Hanacek/NIST

The team’s findings, which appear in the Journal of the American Chemical Society, confirm that the powerful electrical fields that cell membranes generate are largely responsible for repelling nanoscale particles from the surface of the cell. This repulsion notably affects neutral, uncharged nanoparticles, in part because the smaller, charged molecules the electric field attracts crowd the membrane and push away the larger particles. Since many drug treatments are built around proteins and other nanoscale particles that target the membrane, the repulsion could play a role in the treatments’ effectiveness.

The findings provide the first direct evidence that the electric fields are responsible for the repulsion. According to NIST’s David Hoogerheide, the effect deserves greater attention from the scientific community.

“This repulsion, along with the related crowding that the smaller molecules exert, is likely to play a significant role in how molecules with a weak charge interact with biological membranes and other charged surfaces,” said Hoogerheide, a physicist at the NIST Center for Neutron Research (NCNR) and one of the paper’s authors. “This has implications for drug design and delivery, and for the behavior of particles in crowded environments at the nanometer scale.”

Membranes form boundaries in nearly all kinds of cells. Not only does a cell have an outer membrane that contains and protects the interior, but often there are other membranes inside, forming parts of organelles such as mitochondria and the Golgi apparatus. Understanding membranes is important to medical science, not least because proteins lodged in the cell membrane are frequent drug targets. Some membrane proteins are like gates that regulate what gets into and out of the cell.

The region near these membranes can be a busy place. Thousands of types of different molecules crowd each other and the cell membrane — and as anyone who has tried to push through a crowd knows, it can be tough going. Smaller molecules such as salts move with relative ease because they can fit into tighter spots, but larger molecules, such as proteins, are limited in their movements.

This sort of molecular crowding has become a very active scientific research topic, Hoogerheide said, because it plays a real-world role in how the cell functions. How a cell behaves depends on the delicate interplay of the ingredients in this cellular “soup.” Now, it appears that the cell membrane might have an effect too, sorting molecules near itself by size and charge.

“How does crowding affect the cell and its behavior?” he said. “How, for example, do molecules in this soup get sorted inside the cell, making some of them available for biological functions, but not others? The effect of the membrane could make a difference.”

While researchers commonly use electric fields to move and separate molecules — a technique called dielectrophoresis — scientists have paid scant attention to this effect at the nanoscale because it takes extremely powerful fields to move nanoparticles. But powerful fields are just what an electrically charged membrane generates.

“The electric field right near a membrane in a salty solution like our bodies produce can be astoundingly strong,” Hoogerheide said. “Its strength falls off rapidly with distance, creating large field gradients that we figured might repel nearby particles. So we used neutron beams to look into it.”

Neutrons can distinguish between different isotopes of hydrogen, and the team designed experiments that explored a membrane’s effect on nearby molecules of PEG, a polymer that forms chargeless nano-sized particles. Hydrogen is a major constituent of PEG, and by immersing the membrane and PEG into a solution of heavy water — which is made with deuterium in place of ordinary water’s hydrogen atoms — the team could measure how closely the PEG particles approached the membrane. They used a technique known as neutron reflectometry at the NCNR as well as instruments at Oak Ridge National Laboratory.

Together with molecular dynamics simulations, the experiments revealed the first-ever evidence that the membranes’ powerful field gradients were the culprit behind the repulsion: The PEG molecules were more strongly repelled from charged surfaces than from neutral surfaces.

While the findings do not reveal any fundamentally new physics, Hoogerheide said, they do show well-known physics in an unexpected place, and that should encourage scientists to take notice — and explore it further.

“We need to add this to our understanding of how things interact at the nanoscale,” he said. “We’ve demonstrated the strength and significance of this interaction. Now we need to investigate how it affects these crowded environments where so much biology happens.”

Paper: M. Aguilella-Arzo, D.P. Hoogerheide, M. Doucet, H. Wang and V.M. Aguilella. Charged biological membranes repel large neutral molecules by surface dielectrophoresis and counterion pressure. Journal of the American Chemical Society. Published online Jan. 16, 2024. DOI: 10.1021/jacs.3c12348

Source: NIST

Are firefighters at risk of increased exposure to cancer-causing chemicals in their protective clothing?

Last year, a study by the National Institute of Standards and Technology (NIST) showed that the textiles used in protective clothing worn by firefighters often contain per- and polyfluoroalkyl substances, or PFAS, a class of chemicals that has been linked to an increased risk of cancer and other health effects.

A firefighter’s protective clothing includes three layers made of different types of textiles. A pair of studies by NIST has found that these textiles often contain potentially cancer-causing chemicals called PFAS and that they can release more of those chemicals when subject to simulated wear and tear. Credit: B. Hayes/NIST

Now, a follow-up study from NIST shows that the textiles used in that protective clothing, called turnout gear, tend to release more PFAS when they are subject to wear and tear. Taken together, the two studies identified the PFAS compounds present in selected turnout gear textiles, how much of each was present, and whether simulated wear and tear increased the amount of PFAS that the textiles released.

“The firefighter community has raised concerns about PFAS in turnout gear, but before these studies, there was very little data that address those concerns,” said NIST chemist and study co-author Rick Davis. “Based on these studies we can confidently say that more than 20 types of PFAS might be present in firefighter gear and that the amount and type of PFAS vary depending on the type of textile used and the amount of stress it has been subjected to.”

The NIST studies do not assess the health risks that firefighters might face due to the presence of PFAS in turnout gear. However, they provide previously unavailable data that toxicologists, epidemiologists and other health experts can use to assess those risks.

NIST conducted these studies at the behest of Congress, which called on NIST to study PFAS in firefighter gear in the 2021 National Defense Authorization Act.

PFAS are used in many products because they can make things resistant to oil, water and stains. They are often present in clothing, furniture, food packaging and nonstick cookware, among other things. They play a particularly important role in turnout gear by helping firefighters to do their job without getting totally drenched.

Because PFAS don’t break down in the environment, they are often referred to as “forever chemicals.” The Centers for Disease Control and Prevention has found that most Americans have detectable amounts of PFAS in their blood. Other studies have indicated that the blood of firefighters may have higher than average levels

of at least one type of PFAS. Research also suggests that firefighters may have a higher risk for certain types of cancer than the general population does, though that is not necessarily due to PFAS specifically.

Turnout gear includes pants, coats, gloves, boots and helmets. This study focused on the textiles used in pants and jackets, which typically contain three layers of material: a thermal layer nearest the body, a moisture barrier and an outer shell. In the earlier study, the researchers purchased 21 textiles that are typically used in each of these layers. They then tested those textiles for 53 different PFAS compounds and measured how much of each was present.

In the more recent study, the researchers stressed those same textiles using four techniques: abrasion, heat, laundering and weathering. The weathering was simulated by exposing the textiles to ultraviolet (UV) radiation and high humidity.

The researchers then measured the PFAS present after the textiles were stressed. The results showed that abrasion can cause measured PFAS concentrations to increase across all textiles tested. In addition, weathering and heat caused measured PFAS concentrations to increase in the outer shell materials. Finally, laundering had little effect, and in some cases reduced PFAS concentrations, presumably because PFAS were washed away into the wastewater.

Overall, both before and after stressing, measured PFAS concentrations were highest in outer shell fabrics that had been treated with a water-repellent coating. PFAS concentrations were lowest in the thermal layer, which is the layer nearest the firefighter’s body.

The researchers measured PFAS concentrations by first extracting PFAS from the textiles using a solvent. Based on this method, it is unclear what caused PFAS concentrations to change during stressing. Those changes might have been caused by chemical transformations, but it is also possible that stressing loosened the PFAS from the textile fibers, allowing more of it to be extracted.

Now that the researchers have measured PFAS in the textiles that have been stressed under highly controlled laboratory conditions, they are considering studying real gear that has been used for years. That may paint a more realistic picture, though a potentially more complicated one, as used gear might become contaminated with toxic compounds picked up at fire scenes.

Firefighter gear has to meet certain standards, including minimum requirements for water repellency. This research might point to new ways to meet those standards while reducing the risks of PFAS exposure. For instance, the amount and types of PFAS in the textiles varied from one manufacturer to the next, suggesting that some combinations might result in a lower risk of exposure than others. Or manufacturers might find alternative ways to meet the standards without relying on potentially toxic chemicals.

“Using PFAS in turnout gear may or may not be an acceptable risk, given all the other hazards that firefighters already face,” said NIST chemist and co-author John Kucklick. “This data will help people weigh those costs and benefits.”

Source: NIST