A better way to culture central nervous cells

A protein associated with neuron damage in Alzheimer's patients provides a superior scaffold for growing central nervous system cells in the lab. The findings could have clinical implications for producing neural implants and offers new insights on the complex link between the apoE4 apolipoprotein and Alzheimer's disease. Results appear in the journal Biomaterials.

PROVIDENCE, R.I. [Brown University] — A protein associated with neuron damage in people with Alzheimer’s disease is surprisingly useful in promoting neuron growth in the lab, according to a new study by engineering researchers at Brown University. The findings, in press at the journal Biomaterials, suggest a better method of growing neurons outside the body that might then be implanted to treat people with neurodegenerative diseases.

A more dependable scaffold for neural cell culture
Rat central nervous system cells cultured in the apoE4
protein (right) fare better, with more axons and dendrites
than cells cultured in laminin (left). Ironically, apoE4 is
associated with the neural deficits of Alzheimer's disease
in the body. Credit: Palmore Lab/Brown University

The research compared the effects of two proteins that can be used as an artificial scaffold for growing neurons (nerve cells) from the central nervous system. The study found that central nervous system neurons from rats cultured in apolipoprotein E-4 (apoE4) grew better than neurons cultured in laminin, which had been considered the gold standard for growing mammalian neurons in the lab.

“Most scientists assumed that laminin was the best protein for growing CNS (central nervous system),” said Kwang-Min Kim, a biomedical engineering graduate student at Brown University and lead author of the study, “but we demonstrated that apoE4 has substantially better performance for mammalian CNS neurons.”

Kim performed the research under the direction of Tayhas Palmore, professor of engineering and medical science and Kim’s Ph.D. adviser. Also involved in the project was Janice Vicenty, an undergraduate from the University of Puerto Rico, who was working in the Palmore lab as a summer research fellow through the Leadership Alliance.

One size doesn't fit all
Tayhas Palmore and Kwang-Min Kim showed that lamnin,
the preferred scaffold for peripheral nerve cells, is not the
best choice for culturing cells from the central nervous
system. The protein apoE4 works much better.
Credit: Mike Cohea/Brown University

The results are surprising partly because of the association of apoE4 with Alzheimer’s. Apolipoproteins are responsible for distributing and depositing cholesterols and other lipids in the brain. They come in three varieties: apoE2, apoE3 and apoE4. People with the gene that produces apoE4 are at higher risk for amyloid plaques and neurofibrillary tangles, the hallmarks of Alzheimer’s. But exactly how the protein itself contributes to Alzheimer’s is not known.

This study suggests that outside the body, where the protein can be separated from the cholesterols it normally carries, apoE4 is actually beneficial in promoting neuron growth.


Growing new neurons
In the body, neurons grow in what’s called an extracellular matrix (ECM), a protein-rich scaffold that provides cells with nutrients and a molecular structure in which to grow. To grow neurons in the lab, scientists try to mimic the ECM present in the body. Laminin is a common protein in the body’s ECM, and studies have shown that laminin aids the growth of neurons from the peripheral nervous system (nerve cells that grow outside the brain and spinal cord).

It was largely assumed, Kim said, that because laminin was good for growing peripheral nerve cells, it would also be good for growing central nerve cells. That turns out not to be the case.

Kim was inspired to test the effects of apoE4 by a previous study that found that a mixture of apoE4 and laminin promoted CNS cell growth better than laminin alone. “The previous work hadn’t tested the effects apoE4 by itself,” Kim said. “So we started working on a side-by-side comparison of apoE4 and laminin.”

Kim and his colleagues cultured rat hippocampal cells — a model for mammalian CNS neurons — in four different treatments: laminin, laminin and apoE4 mixed, apoE4 alone, and bare glass. They found that cells cultured in apoE4 alone performed substantially better than any other treatment. The apoE4 cells were more likely to adhere to the protein scaffold, which is necessary for proper growth. They also showed more robust growth of axons and dendrites, the wire-like appendages that enable neurons to send and receive nerve signals.

Laminin doesn’t seem to be of much benefit at all for culturing CNS cells, according to the study. Cells cultured on laminin alone did not perform any better than cells cultured on bare glass.

That was another big surprise, Kim said, because laminin is so widely used in all kinds of neuron cultures.

A second part of the research looked at the chemical pathways through which proteins may enhance neuron growth. Previous work had found two neuron receptors, the gateways through which neurons interact with the outside world, that play a role in how external proteins trigger cell growth. However, when Kim blocked these two receptors, known as integrin and HSPG, he found that apoE4 still enhanced neuron growth. That finding suggests that neurons use an as yet unknown pathway to interact with apoE4.

“This discovery opens up a new target for researchers who are interested in identifying receptors that are important for spurring neural growth,” Palmore said.

Application to neural prosthetics
Unlike other cells in the body, nerve cells tend not to regenerate after being damaged by disease or trauma. So researchers hope that they can eventually implant lab-grown cells in the body to treat trauma or neurodegenerative diseases like Alzheimer’s.

“People are looking at all these different proteins to see if we can make a material — a scaffold — that to a neuron, looks and feels like their natural environment,” said Palmore. “The finding that apoE4 is a better protein to add to neural scaffolds is a good breakthrough because most people have been using laminin for the central nervous system models, which turns out to be less than optimal.”

The research was supported by the National Science Foundation (HRD-0548311) and the National Institutes of Health.

- by Kevin Stacey

How ion bombardment reshapes metal surfaces

Ion bombardment of metal surfaces is an important, but poorly understood, nanomanufacturing technique. New research using sophisticated supercomputer simulations has shown what goes on in trillionths of a second. The advance could lead to better ways to predict the phenomenon and more uses of the technique to make new nanoscale products.

PROVIDENCE, R.I. [Brown University] — To modify a metal surface at the scale of atoms and molecules — for instance to refine the wiring in computer chips or the reflective silver in optical components — manufacturers shower it with ions. While the process may seem high-tech and precise, the technique has been limited by the lack of understanding of the underlying physics. In a new study, Brown University engineers modeled noble gas ion bombardments with unprecedented richness, providing long-sought insights into how it works.

Three new mechanisms at the nanoscale
A computer-model image of an island of metal atoms
formed after bombardment by noble gas ions. Atoms
disturbed by the bombardment cluster together under
the surface and then glide back up in a matter of 2.1
trillionths of a second, or picoseconds (ps).
Credit: Kim Lab/Brown University

“Surface patterns and stresses caused by ion beam bombardments have been extensively studied experimentally but could not be predicted accurately so far,” said Kyung-Suk Kim, professor of engineering at Brown and co-author of the study published May 23 in the Proceedings of the Royal Society A. “The new discovery is expected to provide predictive design capability for controlling the surface patterns and stresses in nanotechnology products.”

The improved understanding could open the door to new technologies, Kim said, such as new approaches to make flexible electronics, biocompatible surfaces for medical devices, and more damage-tolerant and radiation-resistant surfaces. The research applies to so-called “FCC” metals such as copper, silver, gold, nickel, and aluminum. Those metals are crystals made up of cubic arrangements of atoms with one at each corner and one in each cube-face center.

Scientists have been trying to explain the complicated process for decades, and more recently they have begun to try modeling it on computers. Kim said the analysis of the Brown team, including lead author and postdoctoral scholar Sang-Pil Kim, was more sophisticated than previous attempts that focused on a single bombardment event and only isolated point defects within the metal substrate.

“In this work, for the first time, we investigate collective behavior of those defects during ion bombardments in terms of ion-substrate combinations,” Kyung-Suk Kim said.

The new model revealed how ion bombardments can set three main mechanisms into motion in a matter of trillionths of a second. The researchers dubbed the mechanisms “dual layer formation,” “subway-glide mode growth,” and “adatom island eruption.” They are a consequence of how the incoming ions melt the metal and then how it resolidifies with the ions occasionally trapped inside.

When ions hit the metal surface, they penetrate it, knocking away nearby atoms like billiard balls in a process that is akin, at the atomic level, to melting. But rather than merely rolling away, the atoms are more like magnetic billiard balls in that they come back together, or resolidify, albeit in a different order.

Some atoms have been shifted out of place. There are some vacancies in the crystal nearer to the surface, and the atoms there pull together across the empty space, that creates a layer with more tension. Beneath that is a layer with more atoms that have been knocked into it. That crowding of atoms creates compression. Hence there are now two layers with different levels of compression and tension.This “dual layer formation” is the precursor to the “subway-glide mode growth” and “adatom island eruption”.

A hallmark of materials that have been bombarded with ions is that they sometimes produce a pattern of material that seems to have popped up out of the original surface. Previously, Kyung-Suk Kim said, scientists thought displaced atoms would individually just bob back up to the surface like fish killed in an underwater explosion. But what the team’s models show is that these molecular islands are formed by whole clusters of displaced atoms that bond together and appear to glide back up to the surface.

“The process is analogous to people getting on a subway train at suburban stations, and they all come out together to the surface once the train arrives at a downtown station during the morning rush hour,” Kyung-Suk Kim said.

The mechanisms, while offering a new explanation for the effects of ion bombardment, are just the beginning of this research.

 “As a next step, I will develop prediction models for nanopattern evolution during ion bombardment which can guide the nanomanufacturing processes,” Sang-Pil Kim said. “This research will also be expanded to other applications such as soft- or hard-materials under extreme conditions.”

In addition to Kyung-Suk Kim and Sang-Pil Kim, other authors include Huck Beng Chew, Eric Chason and Vivek Shenoy.

The research was funded by the Korea Institute of Science and Technology, the U.S. National Science Foundation, and the U.S. Department of Energy. The work used the Extreme Science and Engineering Discovery Environment (XSEDE), which is supported by National Science Foundation grant number OCI-1053575.

Brown Professor Kyung-Suk Kim PhD’80 to Receive 2012 Engineering Science Medal from SES

Brown University School of Engineering Professor Kyung-Suk Kim PhD ’80 will receive the 2012 Engineering Science Medal from the Society of Engineering Science (SES). The prize is awarded in recognition of a singularly important contribution to engineering science. Professor Kim will receive his award during the 49th Annual Technical Meeting of the Society of Engineering Science to be held at Georgia Institute of Technology from October 9-12, 2012. The Society of Engineering Science has only awarded the Engineering Science Medal eight previous times since its inception in 1987.

“This is a tremendous and well-deserved honor for Professor Kim,” said Dean Larry Larson. “As both a Brown Engineering alumnus and professor we are extremely proud of his accomplishments and look forward to his continued contributions to the field.”

Professor Kim receives the prize for his singularly important contributions to experimental micro and nano-mechanics. These include his inventions of transverse displacement interferometer for high strain rate combined normal and shearing load, stress intensity tracer for time dependent fracture testing, Moiré interferometry for finite displacement measurement at the micro and nano-length scales, field projection methods to extract cohesive laws, residual stress measurements via chemical etching, high resolution TEM analysis to extract near atomic resolution constitutive laws and extension of the AFM range to measure the size scaling in contact and adhesion.

Professor Kim received his B.S. and M.S. degrees from Seoul National University of Korea in 1974 and 1976, respectively, and his Ph.D. from Brown University in 1980.  He worked on the faculty of the University of Illinois at Urbana-Champaign from 1980-1989 before returning to Brown as Professor of Engineering in 1989. He is currently the director of Nano and Micromechanics Laboratory in the Mechanics of Solids and Structures Group in the School of Engineering at Brown University.

About the Society of Engineering Science
Founded in 1963, the Society of Engineering Science (SES) was established to promote the free exchange of information on all aspects of engineering science and to provide a forum for discussion, education, and recognition of the talents of the engineering science community. Since its founding in 1963, the SES has established its reputation as the most vibrant and relevant technical society to promote the field of engineering science, where science and engineering meet. The annual technical meetings organized by SES bring leading engineers, scientists and mathematicians from around the world together to tackle some of the most challenging problems at the interface between engineering, sciences and mathematics.

Professor Kyung-Suk Kim melds engineering with history and humanities

“The West had William Tell and the East had Yang Man-Choon in Korea”

When Kyung-Suk Kim, a renowned Korean-American scientist and professor of mechanical engineering at Brown University, says this in his class Dynamics and Vibrations, a required course for engineering students, students are generally puzzled.

Yang was the legendary lord of Ansi Castle in Korea’s ancient dynasty of Goguryeo. He has been known to hit Emperor Taizong of the Chinese Tang Dynasty with an arrow in 645 A.D., when Tang invaded Goguryeo.

Kim`s students, however, pay attention to his lecture that combines history with physics and mechanical engineering if he says, “I will explain the principle of bow’s operations in a mechanical engineering point of view. The Korean bow is considered to be the best in the world from an engineering perspective, which you can confirm through experiments.”

Since 1989, Kim has taught mechanical engineering at Brown University, a prestigious Ivy League university in the U.S., with a laboratory text he wrote himself. More than 1,000 students have attended his lectures and 25 students have completed doctoral and postdoctoral studies under his advising over the years. Indeed, Kim has played the role of missionary for the promotion of Korea`s scientific excellence in its culture.

Speaking to the Dong-A Ilbo, a Korean news paper, over the phone Sunday, he said, “In the early 1990s, Brown University suggested me to develop a laboratory for engineering students that reflects some aspects of humanities and history. So I began working on developing such laboratory courses that bring in scientific excellence of Korean culture.”

Through experiments, Kim and his students have unveiled the secret of an ancient Korean bow that flies arrows up to nearly 1 kilometer, twice and three times the range of British and Japanese bows, though the bowstring is just 120 centimeters, shorter than Britain`s (180 centimeters) and Japan’s (2 meters). Kim showed that the Korean bow has a thrust of double pushes while launched, analogous to the thrust of a two-staged rocket.

Many had thought Korean bowstrings too short since Koreans have small frames. Kim, however, said the short bowstring creates great impellent power by the double-push mechanism and Korean bows bend to increase such power.

After completing graduate studies at Seoul National University, Kim went to the U.S. in 1976 for his PhD. He joined the Brown faculty in 1989 as a full professor. As the Director of the Nano and Micro Mechanics Laboratory at Brown, he received world attention last year with an article on the principle of precisely cutting carbon nano tubes using ultrasonic waves, written jointly with his collaborators at the Korean Institute of Science and Technology. The article was published in the Proceedings of the Royal Society, London.

- Courtesy of the Dong-A Ilbo (Korea)

Nanowrinkles, nanofolds yield strange hidden channels

Wrinkles and folds, common in nature, do something unusual at the nanoscale. Researchers at Brown University and in Korea have discovered that wrinkles on super-thin films have hidden long waves. The team also found that folds in the film produce nanochannels, like thousands of tiny subsurface pipes. The research could lead to advances in medicine,  electronics and energy. Results appear in Proceedings of the Royal Society A.

PROVIDENCE, R.I. [Brown University] — Wrinkles and folds are ubiquitous. They occur in furrowed brows, planetary topology, the surface of the human brain, even the bottom of a gecko’s foot. In many cases, they are nature’s ingenious way of packing more surface area into a limited space. Scientists, mimicking nature, have long sought to manipulate surfaces to create wrinkles and folds to make smaller, more flexible electronic devices, fluid-carrying nanochannels or even printable cell phones and computers.

A subsurface system of nanopipesResearchers at Brown University and in Korea used focusedion beams to extract a cross-section of compressed goldnanofilm. When tips of regular, neighboring folds touched,nanopipes were created beneath the surface.Credit: Kim Lab/Brown University

But to attain those technology-bending feats, scientists must fully understand the profile and performance of wrinkles and folds at the nanoscale, dimensions 1/50,000th the thickness of a human hair. In a series of observations and experiments, engineers at Brown University and in Korea have discovered unusual properties in wrinkles and folds at the nanoscale. The researchers report that wrinkles created on super-thin films have hidden long waves that lengthen even when the film is compressed. The team also discovered that when folds are formed in such films, closed nanochannels appear below the surface, like thousands of super-tiny pipes.

“Wrinkles are everywhere in science,” said Kyung-Suk Kim, professor of engineering at Brown and corresponding author of the paper published in the journal Proceedings of the Royal Society A. “But they hold certain secrets. With this study, we have found mathematically how the wrinkle spacings of a thin sheet are determined on a largely deformed soft substrate and how the wrinkles evolve into regular folds.”

Wrinkles are made when a thin stiff sheet is buckled on a soft foundation or in a soft surrounding. They are precursors of regular folds: When the sheet is compressed enough, the wrinkles are so closely spaced that they form folds. The folds are interesting to manufacturers, because they can fit a large surface area of a sheet in a finite space.

Kim and his team laid gold nanogranular film sheets ranging from 20 to 80 nanometers thick on a rubbery substrate commonly used in the microelectronics industry. The researchers compressed the film, creating wrinkles and examined their properties. As in previous studies, they saw primary wrinkles with short periodicities, the distance between individual wrinkles’ peaks or valleys. But Kim and his colleagues discovered a second type of wrinkle, with a much longer periodicity than the primary wrinkles — like a hidden long wave. As the researchers compressed the gold nanogranular film, the primary wrinkles’ periodicity decreased, as expected. But the periodicity between the hidden long waves, which the group labeled secondary wrinkles, lengthened.

“We thought that was strange,” Kim said.

It got even stranger when the group formed folds in the gold nanogranular sheets. On the surface, everything appeared normal. The folds were created as the peaks of neighboring wrinkles got so close that they touched. But the research team calculated that those folds, if elongated, did not match the length of the film before it had been compressed. A piece of the original film surface was not accounted for, “as if it had been buried,” Kim said.

Indeed, it had been, as nano-size closed channels. Previous researchers, using atomic force microscopy that scans the film’s surface, had been unable to see the buried channels. Kim's group turned to focused ion beams to extract a cross-section of the film. There, below the surface, were rows of closed channels, about 50 to a few 100 nanometers in diameter. “They were hidden,” Kim said. “We were the first ones to cut (the film) and see that there are channels underneath.”

The enclosed nano channels are important because they could be used to funnel liquids, from drugs on patches to treat diseases or infections, to clean water and energy harvesting, like a microscopic hydraulic pump.

Contributing authors include Jeong-Yun Sun and Kyu Hwan Oh from Seoul National University; Myoung-Woon Moon from the Korea Institute of Science and Technology; and Shuman Xia, a researcher at Brown and now at the Georgia Institute of Technology. The National Science Foundation, the Korea Institute of Science and Technology, the Ministry of Knowledge Economy of Korea, and the Ministry of Education, Science, and Technology of Korea supported the research.