The physics of caffeine extraction now provides solutions for nerve stimulation

Researchers at Karolinska Institutet have developed a method for producing flexible plastic electrodes capable of releasing the neurotransmitter acetylcholine upon an electrical trigger. This technology has applications in the treatment of Alzheimers’s Disease or muscular disorders where today, hard  metal electrodes are used to stimulate nerves to restore muscle function.

Using flexible plastics instead of metal is an advantage because it will cause less damage to the surrounding brain tissue. By stimulating brain regions the physiological way, with neurotransmitters instead of electricity, the researchers claim that the effect will be much more specific and will allow treatments that were not previously possible.

Impregnating conducting plastics with sufficient amounts of neurotransmitters, so that release is only possible due to an electronic trigger is difficult under normal conditions. The multidisciplinary team therefore used the unique properties of a supercritical fluid to achieve the desired effect.

They took carbon dioxide gas and put it under pressure while at the same time, increasing the temperature. At a specific point under these conditions, the carbon dioxide will take on the properties of both a liquid and a gas. The researches found that acetylcholine with a co-solvent would dissolve in supercritical carbon dioxide and penetrate into the conducting plastic material.

“This approach has additional benefits.” said Professor Agneta Richter-Dahlfors, the lead investigator of the study. “Supercritical carbon dioxide is good sterilization method for sensitive materials, so when we impregnate the plastic with this method, we are also preparing it for use in the clinic, you get two for one.”

In the second part of the research article the researchers showed that they could then release the acetylcholine on demand by stimulating the conducting plastic with an electrical signal. This caused the polymers of the plastic to expand, allowing the trapped acetylcholine to diffuse out into the surroundings. This could be used for treating muscular disorders where the motor neurones are damaged.

“We are very excited about the results. The field of Organic Bioelectronics is expanding and every way we turn we find new and exciting applications in medicine. There are many important neurotransmitters which we can use with this method to stimulate nerve cells specifically,” said Dr. Susanne Löffler.

Source: Susanne Löffler, Silke Seyock, Rolf Nybom, Gunilla B. Jacobson, Agneta Richter-Dahlfors, Electrochemically triggered release of acetylcholine from scCO2 impregnated conductive polymer films evokes intracellular Ca2 + signaling in neurotypic SH-SY5Y cells, Journal of Controlled Release, Volume 243, 10 December 2016, Pages 283-290, ISSN 0168-3659, http://dx.doi.org/10.1016/j.jconrel.2016.10.020.

NIST Releases Roadmap for Polymer-Based Additive Manufacturing

Additive manufacturing (AM) is a high-priority technology growth area for U.S. manufacturers. Innovative AM processes that fabricate parts layer-by-layer directly from a 3-D digital model have great potential for producing high-value, complex, individually customized parts. Companies are beginning to use AM as a tool for reducing time to market, improving product quality, and reducing the cost to manufacture products. Metal-based AM parts are already in use in a number of applications, including automotive engines, aircraft assemblies, power tools, and manufacturing tools.

IR thermography measurement of materials extrusion additive manufacturing. Image credit: Jon Seppala

In support of the development of polymer-based additive manufacturing, the National Institute of Standards and Technology (NIST) has released the Measurement Science Roadmap for Polymer-Based Additive Manufacturing(link is external), a guide that identifies future desired capabilities, challenges, and priority RD topics in polymer-based AM. The report is the result of the “Roadmap Workshop on Measurement Science for Polymer-Based Additive Manufacturing,” held June 9-10, 2016 at the NIST campus in Gaithersburg, Maryland.  The workshop brought together nearly 100 AM experts from industry, government, national laboratories, and academia to identify measurement science challenges and associated RD needs for polymer-based AM systems. The workshop was hosted by NIST, and sponsored by the National Science Foundation, Division of Civil, Mechanical and Manufacturing Innovation(link is external) and NIST’s Material Measurement Laboratory.  Additive manufacturing is an important research priority for NIST  and a key component of MML’s Five-Year Strategic Plan.

By identifying high priority goals and challenges in polymer-based AM, the report can serve as a roadmap for RD, standards development, and other future efforts.  It includes detailed analyses of the complexities surrounding material characterization, process modeling, in situ measurement, performance, and other cross-cutting challenges for polymer-based AM.  As such, the report can help guide public and private decision-makers interested in furthering the capabilities of polymer-based AM, and accelerating its more widespread use, and contribute to a robust national research agenda for polymer-based AM.

Source: NIST

Finding Diamonds in the Rough

During the kraft process used to convert wood into wood pulp, the structural material lignin is partially converted into molecules like stilbene. Stilbenes are also naturally occurring in plants and some bacteria, and may play a role in plant pathogen resistance.

The crystal structure of NOV1, a stilbene cleaving oxygenase, shows the features of this enzyme at atomic resolution. (A) This protein fold view highlights the placement of an iron (orange), dioxygen (red), and resveratrol, a representative substrate (blue) in the active site of the enzyme. (B) This surface slice representation shows the shape of the active site cavity and the arrangement of iron, dioxygen, and resveratrol. Image credit: Ryan McAndrew/JBEI and Berkeley Lab

Currently, the deconstruction of plant biomass into cellulose and lignin is an expensive process. Lignin accounts for about 30 percent of plant cell wall carbon, and increasing the efficiency of its conversion into chemicals or fuels could have a significant positive impact on the economics of processing lignocellulosic biomass. Enzymes capable of producing useful compounds from the breakdown of stilbenes and similar molecules could be employed for this. Collaborators from two of the Department of Energy Bioenergy Research Centers now have gained first-hand insight into how a stilbene cleaving oxygenase (SCO) carries out this unusual chemical reaction.

Researchers from the Joint BioEnergy Institute (JBEI) and the Great Lakes Bioenergy Research Center (GLBRC) report the atomic-level structure of NOV1, an SCO that breaks down a stilbene substrate into two smaller compounds. Paul Adams, vice president for technology at JBEI and the senior author of the study, explained, “In order to get a complete picture of how this enzyme works, we solved the structure of NOV1 in complex with a representative stilbene substrate (resveratrol), a representative product (vanillin), as well as in its unbound form.”

“When we studied the structures of NOV1, we saw a ternary complex of protein, oxygen, and either the substrate or product in the active site. This has not been seen previously in any crystal structures of related carotenoid cleavage oxygenases (CCO),” said co-first author Ryan McAndrew, project scientist at JBEI. “Despite the fact that it is similar to CCOs, this NOV1 structure shows several key differences indicative of their substrate preferences and how the enzyme carries out its reaction.”

This enzyme’s active site contains a coordinated iron atom that forms a stable complex when exposed to nitric oxide. This allowed for study by electron paramagnetic resonance (EPR) spectroscopy, which confirmed the configuration of the atoms in the crystal structure active site and provided information useful for elucidating the enzyme mechanism. GLBRC Deconstruction lead Brian Fox said, “Through our work, we were able to propose a mechanism for the reaction that requires dioxygen and the unique arrangement shown in the active site by the crystal structure. This gives new insight into how an SCO might be used to generate desirable bioproducts. As an added benefit, this work helps us understand related enzymes like carotenoid cleavage oxygenases, which produce vitamin A and retinal found in the eye.”

Cost reduction of the plant biomass breakdown and conversion of deconstruction byproducts such as lignin into chemicals are core missions of the bioenergy research centers. The result of this interdisciplinary collaborative study is another step toward finding ways to change a very abundant material like lignin into beneficial valuable bioproducts. “Ultimately, enzymes like NOV1 could produce value in the biological production of molecular fragments derived from lignin,” said Adams. “This would contribute to the sustainable operation of a biorefinery for the production of biofuels and other bioproducts.”

Source: LBL

Compound protects transplanted hearts from rejection

An experimental drug that blocks the activation of an immune cell component effectively prevented rejection of heart transplants in mice, according to new research from scientists at Weill Cornell Medicine and Brigham and Women’s Hospital.

The findings, published in Proceedings of the National Academy of Sciences, describe a compound developed by Weill Cornell Medicine investigators that inhibits cellular structures called immunoproteasomes while sparing closely related structures called constitutive proteasomes. Proteasomes help cells regulate their behavior by breaking down regulatory proteins. Constitutive proteasomes are found in all cells, while immunoproteasomes are expressed chiefly in cells of the immune system.

The left image shows a section of a transplanted heart in a mouse that was given only CTLA4-Ig, a standard anti-rejection therapy. A large number of infiltrating immune cells are visible by their blue-staining nuclei, showing that the heart is undergoing rejection. Heart muscle cell nuclei are also stained blue. The right image shows a section of a transplanted heart in a mouse given CTLA4-Ig together with the immunoproteasome inhibitor DPLG3. The heart looks normal. There are almost no immune cells, only nuclei of heart muscle. Credit: Dr. Jamil Azzi/Brigham and Women’s Hospital

Currently approved proteasome inhibitors target both types of proteasomes equally. They are used in transplant medicine to block immune cell function and reduce the rejection of the transplanted organs. However, inhibition of proteasomes in all cells throughout the body can cause toxicity in both the transplanted organ and the host. In their study, the investigators demonstrated that their compound, a highly selective and reversible immunoproteasome inhibitor called DPLG3, only shuts down immunoproteasomes, leaving constitutive proteasomes unscathed. DPLG3 reduced the number of immune cells in the graft. The inhibitor also increased the expression of “exhaustion markers” on the remaining immune cells. Exhaustion markers are typical of immune cells that have dialed down their ability to attack foreign cells. These changes protect the organ from rejection, while sparing the body from the toxic side effects of nonselective proteasome inhibitors, such as damage to bone marrow and the nervous system.

“Transplant patients often have to continue toxic, broadly immunosuppressive agents for a long period of time, increasing the risks of infection, cancer and toxicity to the graft itself,” said co-senior author Dr. Carl Nathan, chairman of the Department of Microbiology and Immunology, the R.A. Rees Pritchett Professor of Microbiology, and a professor of microbiology and immunology and of medicine at Weill Cornell Medicine. “Selectively blocking immunoproteasomes – without affecting the constitutive proteasomes in other host cells and in the cells of the transplanted organ – could help patients with long-term acceptance of their grafts and improve the outlook for organ transplantation.”

Weill Cornell Medicine researchers, led by co-senior study author Gang Lin, associate professor of research in microbiology and immunology, designed DPLG3 to be highly selective for immunoproteasomes and to remain highly selective during prolonged exposure, in contrast to other experimental agents directed at immunoproteasomes. Nathan and Lin are co-inventors of DPLG3 and have filed a patent for it.

Then, Dr. Jamil Azzi and his team at Brigham and Women’s Hospital in Boston tested DPLG3 on mice that received heart transplants. Mice given DPLG3 daily for 10 days after surgery accepted the transplanted hearts for an average of 13 days, compared with seven days for rodents that did not receive the compound. The researchers then combined a seven-day regimen of DPLG3 with a single dose of the immunosuppressive agent CTLA4-Ig, finding that the treatment protected the transplant for an average of 84 days, compared with 38.5 for mice receiving only CTLA4-Ig. A single dose of CTLA4-Ig and a 14-day DPLG3 program maintained the heart transplant for as long as the mice were studied – more than 100 days, on average – without any other treatment after day 14.

Nathan said that it is likely, based on work not included in the paper, that these compounds might be helpful in very different conditions that involve excessive inflammatory or immune reactions. “We think there’s a lot of work to do to document whether this is the case and to improve the compounds in terms of basic pharmaceutical properties, but we think the possibilities are bright,” he said.

Source: Cornell University

Water-loving chemist also explores clusters of gold atoms

University of Nebraska-Lincoln chemist Xiao Cheng Zeng recently conducted a simple scientific test during lunch at a local Chinese restaurant. He poured a few ounces of water onto the table top to see if the surface was water-loving (hydrophilic) or water-repellant (hydrophobic).

A rendering shows a nanodroplet of water sitting on a folded protein that is flanked by aligned amino acids. The contact angle, as measured from the top of the droplet to the protein surface, reveals the amino acids’ repellence to water – a characteristic important to understanding the proper functioning of biomolecules.

“This one I would say pretty much loves the water,” said Zeng, the Chancellor’s University Professor of Chemistry, as the water spread to cover a section several inches in diameter.

Far more complicated, however, are the findings of his recent paper on the hydrophobicity of proteins. Zeng and his co-authors recently published their computational data on the molecular hydrophobicity of amino acid chains in the Proceedings of the National Academy of Sciences.

Hydrophobic interactions help drive protein folding, a necessary step for the molecules to perform their biological functions.

“Misfolded proteins could cause diseases such as mad cow disease,” said Zeng, whose research is supported by the National Science Foundation.

The work of Zeng and his associates could help researchers decipher the role of hydrophobic interactions in protein folding, but measurements are critical. In the macro-environment of a kitchen countertop or a frying pan, engineers can optically measure the contact angle of a water droplet. The contact angle is the angular distance from the top of the surface to the top of the droplet.

“If the angle is big, then that means water doesn’t like to wet the surface,” Zeng explained. “If the angle is zero degrees, the water pretty much wets the entire surface.”

But the optical approach that engineers use to measure the contact angles on large, flat, wet surfaces doesn’t work well for proteins, which are microscopic and three-dimensional. Protein chemists measure hydrophobicity instead by studying the complex interactions between proteins and water. Zeng, however, sought to devise a measurement of protein hydrophobicity that he could share with engineers.

To do this, Zeng and colleague Joseph Francisco, the Elmer H. and Ruby M. Cordes Chair in Chemistry and dean of the College of Arts and Sciences, collaborated with Chinese physicists to create a network of proteins on a flat, artificial surface in a computer simulation.

“We stretch them and put them together side by side and then align them into a network,” Zeng explained. In the final stage, they add a virtual droplet of water to the mix to see what happens.

A single protein could contain millions of amino acid groups. In the past, however, chemists could measure only one such group per protein. But the Zeng-Francisco team’s work at Nebraska’s Holland Computing Center knows no such limitation.

“We have applied this computational measurement to all 20 types of amino acids in nature,” Zeng said, resulting in a new amino acid hydrophobicity scale.

Going for gold

Zeng’s theoretical and computational research spans energy science, materials science and atmospheric chemistry. A second paper published online Dec. 2 in Nature Communications displays his research versatility. In that paper, Zeng and three co-authors propose a grand unified model for understanding the structural richness of the 71 liganded gold clusters now known.

Ligands are ions or molecules that permit the clusters to form different species in solution. A better understanding of the atomic structure of the clusters could accelerate the development of potential applications.

“Liganded gold clusters have attracted intensive interest over the past 10 years owing to their broad and practical implications in catalysis, electrochemistry, quantum electronics and biomedicine,” Zeng and his co-authors wrote. The team performed supporting molecular computation at supercomputing centers in China and at the Nebraska Cluster for Computational Chemistry.

Nobel laureate Roger Kornberg of Stanford University first determined the structure of a big liganded gold cluster using X-ray crystallography in 2007. His work ignited new interest in the field.

Zeng’s latest paper represents a milestone in liganded gold cluster research, Francisco said.

“This is a major contribution,” said Francisco, one of Zeng’s frequent collaborators.

Zeng and his associates borrowed an idea from the world of high-energy physics for their classification scheme. In high-energy physics, quarks, the smallest known constituent of matter, come in six types or flavors: up, down, strange, charm, top and bottom. Now the field of liganded gold clusters has three flavors of its own: bottom, middle and top. These flavors represent valence states, which categorize the clusters’ ability to chemically unite with one another.

When gold clusters are small, they are like children who exhibit a different look and behavior as they age from 2 or 3 years old to 4 or 5, Zeng said. Similarly, a liganded gold cluster consisting of four atoms has a different structure and properties than a cluster consisting of five.

This structure-property relationship is important only for clusters ranging in size from two atoms up to about 1,000. At that point, the clusters are like children who have matured into adulthood and behave like regular gold, Zeng said.

Of the 71 known liganded gold clusters, scientists have crystallized the structures of 54 in the laboratory. Researchers have computationally predicted the remaining 17.

“We have 54 but maybe there are 100 or 200 to go,” Zeng said. Zeng predicts that his team’s new grand unified model will be able to accommodate them all.

Source: University of Nebraska-Lincoln