A community of research leaders in biomedical engineering at Tufts has helped to expand the (ever-growing) list of medical uses of silk
We’ve all brushed against them, whether we’re moving through a summer cabin that’s been shut for a season or reaching for a light cord in some dark, musty basement—those pesky cobwebs that cling ever so lightly to our faces and hands and seem impossible to pick off our shirtsleeves.
But spider silk is a much more complex and wonderful material than you might suppose. An early fascination with the nature of the stuff opened a door for David Kaplan that he is still walking through, in a sense, helping to transform medical research as he goes.
“If you asked me 20 years ago, would I still be working on silk in 2010, I’d have thought you were crazy,” says Kaplan, professor and chair of biomedical engineering at Tufts University, shaking his head. “But silk continues to amaze me. I’ve been fortunate in the way that it keeps leading to new and exciting things.”
Back in 1988, Kaplan became intrigued by a scientific paper on the unusual mechanical properties of dragline spider silk. Consider the evidence. Here was a barely-there filament that emerged from a simple biological process and was up to five times stronger than steel, that was stable and durable and that could be easily shaped into a variety of shapes and forms.
“I started to wonder, what is this material and how did it arise from a simple biological process?” Kaplan remembers. He spent the next decade or so delving into the mystery.
Kaplan found that silk consists of a long chain of repeating sequences of a small number of amino acids. Tight hydrogen bonds in these amino acids strengthen the chain, making it three times tougher than Kevlar, the synthetic fabric used to make bulletproof vests. Silk’s organizing pattern is as elemental as a one-two punch and unbreakably strong.
Viewed in evolutionary terms, spider silk is the culmination of millions of years of refinement of web design granted the arachnids of our world. The silk could scarcely be improved upon for its task of snaring and holding prey. (For a number of reasons, including the ability to cultivate the product on a large scale, purified silk from silkworms, slightly different in its chemical composition, is more commonly used than spider silk in research.)
A nice bonus feature to add to the list of silk’s alluring traits, one that gave it immediate potential applicability, stood out from the rest. Silk was biocompatible, meaning that it could be put into the human body without arousing an immune response. In contrast with a host of synthetic substances commonly used to create medical implants, silk is just one step removed from human tissue, Kaplan explains. That means silk meshes well with living things.
What are the possibilities for silk in medicine? Since 2000, Kaplan has been exploring this question, enlisting others in his passion as he goes, like someone who just heard some unbelievably great news and can’t wait to share it. The man wears a number of faces, all of them agreeable and welcoming. He is at once the world’s leading ambassador for the biomedical marvels of silk and the avuncular neighbor who loans you his mower with a shrug and smile when yours is on the fritz.
Kaplan is easygoing (“He’s incredibly busy, but will always make time to help you,” says Kelly Sanford, his longtime executive assistant), but also a man on a mission, and he lives in the tension there.
Day by day, he’s enlarging the ring of true believers through his exuberance. A few years ago, he walked down the hall from his office at the Center for Science and Engineering on Tufts’ Medford/Somerville campus and showed his colleague, Fiorenzo Omenetto, a specialist in optics, just how flat and sheer a silk membrane could be.
“I had never seen anything like that,” says Omenetto, who began envisioning how easily the silk might hold biosensors and electrical micro-arrays; now the two men are close collaborators on developing such devices for human implantation. In an article in Science last summer, “New Opportunities for an Ancient Material,” Omenetto and Kaplan outlined a few of these envisioned uses.
“When the resonance is good, the flow is high,” says Omenetto, in a remark that typifies Kaplan’s involvement with others as well as the ever-widening potential for the wispy material he has taken to heart.
Completely Won Over
Michael Rosenblatt is a believer. A nationally known bone biologist and the former dean of Tufts Medical School, Rosenblatt has been concerned for some time with the process by which breast cancer cells move through the human body to the skeleton and extend their damage.
“Once the cancer spreads to bone, the disease enters an incurable phase,” he notes. “That’s when we see pain, fractures, high calcium levels in the blood and ultimately death from bone metastases.”
Rosenblatt attended a Kaplan lecture a few years ago and came away persuaded that he might have found a better method for conducting his research. To simulate the process of cancer colonizing bone, Rosenblatt had been employing human breast cancer cells and human bone samples recovered from hip operations.
This was the normal way of doing things. One inescapable logistical problem was the variability in the bone samples. A donor might be male or female, young or old, and suffering from any of a range of medical ailments. “The person supplying the bone might have had osteoporosis,” Rosenblatt suggests.
In contrast to this largely random array of samples, a silk “scaffold” could supply a platform, “like steel bars in a skyscraper,” in Rosenblatt’s words, letting him grow bone from human stem cells in a pure, reproducible form. “Human bone is hard to work with. It’s pretty tough stuff. But you can control tissue-engineered bone. You can manipulate it easily,” says Rosenblatt, now chief medical officer at Merck, the pharmaceutical giant.
“This, in turn, opens up potential new targets for cancer treatments. Right now, most research in my area is targeted at the cancer cell. However, a better target might be something in the bone that’s enabling the cancer to colonize there. What is it that the breast cancer cells are sticking to?” With help from silk, Rosenblatt is exploring that promising new angle.
John Castellot, a professor and vascular specialist at the medical school, has his own raft of reasons to be excited about the tough, flexible, biologically inert material. Let’s start with silk’s seeming invisibility. “Silk doesn’t contain stretches of amino acids that are recognized by the immune system,” he explains. “Therefore it’s not attacked by the immune system. This is very uncommon. Silk is a single-protein fibroin. Cells on patrol in the body don’t recognize the silk as being non-self.”
Another nice feature? “Silk can be chemically manipulated,” he continues. “You can make it dissolve quickly or slowly in the body. This means that you can impregnate the silk with a drug and then control the release of that drug.” Again, as with Rosenblatt, the big emphasis here is on the word “control.”
Have a Heart
Castellot’s research focus is coronary artery disease. During bypass surgery or balloon angioplasty to clear out clogged arteries, certain damage is done to the blood vessel, he explains. Smooth muscle cells rapidly proliferate in response to the damage, and through their sheer proliferation act to close down the artery. Endothelial cells lining the artery walls fight back to keep the artery open and functioning well. “It’s a race between endothelial cells and smooth muscle cells,” says Castellot.
As often as 25 percent of the time, the smooth muscle cells win. To depict the struggle, Castellot displays a representative cross-section of a human artery in sequence: first immediately after the surgery, then two days later and finally 14 days later. It is like watching a straw whose sides are steadily collapsing.
The surgery itself disrupts the patient’s endothelial cells. At two days post-surgery, a smattering of endothelial cells is still visible, but the smooth muscle cells are already revving up their proliferation engines. After two weeks, anyone can see that the muscle cells have largely won control of the artery and re-blocked it in a process called restenosis. It happens that fast.
Normally a stent will be inserted to help prop open the artery. Often the stent is drug-coated to help eliminate those smooth muscle cells choking off the artery’s normal flow. But as Castellot explains, the problem with drug-coated stents, typically made of Mylar or metal, is that the only drugs that will adhere to them are as crude as shotgun blasts in their effect. “They don’t discriminate,” he says of the commonly used drugs. “They kill muscle cells but also endothelial cells.”
A half-million coronary bypass surgeries are performed each year in the U.S., and about 20 or 25 percent of them fail, in part because of the losing battle being waged between cell types in the patients’ arteries. By any measure, the loss of hard-won arterial flow has been an intractable problem in the field. Silk promises to change the historic terms of the fight, Castellot contends.
The reason is silk’s ability to hold beneficial proteins in place on its surface. Proteins are able to distinguish between muscle cells and endothelial cells, inhibiting the growth of the former while ignoring the latter. The adhesion between proteins and silk has to do with the natural surface characteristics of the fabric. Proteins are big molecules, at least 100 times larger than taxol or rapamycin, two commonly used stent coatings, Castellot says. A conventional stent simply couldn’t hold molecules that large in place without special coatings to keep them on; they would slide right off.
One goal under joint development in the Kaplan and Castellot labs is to perfect silk-coated stents impregnated with therapeutic drugs. A second tantalizing objective is to make small-bore blood vessels out of silk. These would be about the size of human coronary arteries—roughly the diameter of a toothpick—and offer cardiologists something new in the fight for coronary health.
A silken coronary artery implanted in the heart could easily be coated with molecules to prevent muscle cell proliferation and stimulate endothelial cells, the scientist suggests, thereby working to block restenosis. “You can’t stick cells onto Dacron or nylon,” Castellot notes. “They won’t stay put. One of the great things about silk is that it can be chemically modified, so that you can build it layer by layer and piece by piece.”
In the space of a few short years, Rosenblatt and Castellot have become true believers in the magic of silk—Rosenblatt for the easy way the material has simplified his research and given him control over its variables, and Castellot for the way this eerily neutral material can be inserted in the body and loaded to release chemicals where they can do the most good. But the impact of silk extends well beyond the ravages of bone cancer and clogged arteries.
Cultivating Human Cells in 3-D
Michael House, an assistant professor of obstetrics and gynecology, has found a use for silk as he examines the critical role played by abnormal cervical tissue in pre-term delivery. Pre-term births account for some 12 percent of all births in the U.S., with related costs amounting to $26 billion annually, according to the Institute of Medicine.
This is in addition to high mortality rates and a host of developmental issues associated with premature infants. Historically, the focus of most medical researchers has been on the uterus, House says, but “there’s a growing appreciation for the role of the cervix.”
The smooth-muscled uterus and the cervix, with its fibrous connective tissue, work together to permit a normal birth, the doctor reminds us. The cervix forms the bottom of the uterus, and the function of the two organs should be synchronized for a healthy delivery process. If cervical length shortens too much, House routinely tells his patient: “Ma’am, you’re at high risk for pre-term delivery.”
In his research, House has taken a step back to ask some basic questions. What is the cervix made of, and how does it change during pregnancy? “Our hypothesis,” House explains, “is that the cervix is shortening because the cervical tissue is weak and unable to stay closed.” That’s the starting point.
Studying cervical tissue can be problematic, though. The usual method is to subject cervical cells to testing in a Petri dish, but the limitations of this approach are well known. Repeated studies have shown that cells behave differently in three dimensions than they do in two, House points out. So how could that tactical problem be solved?
About three years ago, House bumped into the silk evangelist David Kaplan, who said to him, “Let’s make cervical tissue.” House’s reply: “Great. How do you do that?”
The answer was to isolate human cervical cells taken from women undergoing surgery and attach the cells to tissue scaffolding made of silk. The goal is to build an in vitro cervical model that closely resembles its real-life source.
“This hasn’t been attempted before,” says House. “What’s novel here is that we’re cultivating human cells in 3-D in order to match the body’s biological, mechanical environment. In other words, we’re trying to create a biomimetic environment for the cells. You can’t get this in a Petri dish.”
Previously, House had tried using collagen gel to build his cervical model. Silk was simpler and quicker. “It’s easy to use and it works,” he says, sounding like someone who’s not apt to be going back to the old ways.
A Better Building Material
Ronald Perrone is yet another example of a Tufts researcher who has collaborated with Kaplan and been won over by silk. He is a professor of medicine and a nephrologist at Tufts Medical Center. His clinical practice emphasizes polycystic kidney disease, or PKD, a hereditary disorder in which the kidneys are enlarged by fluid-filled cysts, resulting in a gradual breakdown of the kidney’s ability to filter blood.
Nationwide, there are about 600,000 cases of people with PKD. “I see hundreds of patients with this condition,” says Perrone, who is also medical director of the kidney transplantation program at Tufts. “My goal is to create effective treatments.”
After their initial meeting at a lecture that Kaplan gave on the Boston campus, the medical engineering guru invited Perrone to his lab to show him how a kidney membrane might be simulated using silk tissue scaffolding and human cells. The approach was similar to House’s building the cervix model. Silk evinced all the old virtues for Perrone that had charmed the other scientists: the stuff was biologically neutral and could hold human cells in place; its surface could be tightly controlled; it was an easy material to mold into any form you wanted; and it was relatively cheap and could be deployed on a large scale.
That last feature stood out for Perrone. “Kidney disease in humans is rather slow,” he notes. “In animal models, the usual method for studying the process, the disease takes five months to manifest. That means that for those five months, we’ve got to be taking care of animals every day, feeding and caring for them. That gets to be expensive and labor-intensive. But with silk we can set things up for a high-throughput capacity.” And how is the PKD research going? “So far, so good,” says Perrone.
Implants That Melt into Place
The door to the medical uses of silk has barely swung open, yet the enthusiasm for the cause is growing steadily. Juan Enriquez, founding director of the Life Sciences Project at Harvard Business School, a leading authority on the economics of biotechnology, is an ardent fan of the vision advanced by Kaplan and Omenetto.
“Silk has led to the rise and fall of civilizations, and now here are two interesting, quirky, smart scientists leading the re-engineering of this familiar material in a huge range of ways that nobody’s thought of,” he told a reporter recently.
Many such approaches are already in development.
Doped silk implants, for example, promise to revolutionize drug delivery, especially for chronic conditions. Eleanor Pritchard, an undergraduate student in biomedical engineering who worked recently in Kaplan’s lab, explains that the dime-sized implants are made by blending drugs into a liquid silk solution, stabilizing the drugs and then hardening the mixture to form a film. When the film is implanted under a patient’s skin, the silk biodegrades on a controlled schedule, gradually releasing the drug.
“Instead of taking daily growth hormone shots, or dosages of anti-seizure medication,” Pritchard, E11, points out, “patients can get these implants and receive a constant, stable dosage of whatever it is they need.”
Using silk embedded with nano-electronics and coated with a variety of substances, Omenetto is helping to create an entirely new field of medical signaling devices. Together with John Rogers, a professor of materials science and engineering at the University of Illinois, he and Kaplan created silk sensors that can be placed under the skin to monitor glucose levels in the blood. A next stage of development could include microscopic antennas able to transmit data from inside the body, perhaps by text message, to a receiver in the patient’s hand or doctor’s office.
Brain implants that can “melt into place” are another promising avenue. Kaplan and Omenetto have worked with Rogers and Brian Litt, a neurologist at the University of Pennsylvania, to fashion a bit of stretchable silk that holds metal electrodes 500 microns thick, much thinner than a sheet of paper. The silk is inserted into the brain cavity. When flushed with saline, the silk dissolves, leaving the electrodes draped in place. (The research was published in Nature Materials last spring.)
This approach offers multiple advantages. In contrast to the rigid silicon grid normally used for brain implants, silk is able to wrap itself around the folds of the brain, hugging its contours, as one observer noted, “the same way a silk dress clings to the hips.”
The intrusion is temporary and benign. Not only is potential damage to brain tissue reduced by using the softer, thinner material, but the greater surface adhesion afforded by silk means that more signals are picked up by the implant.
Walter Koroshetz of the National Institute of Neurological Disorders and Stroke, part of the National Institutes of Health, confirms the advantage. “These implants have the potential to maximize the contact between electrodes and brain tissue, while minimizing damage to the brain,” he says. “They could provide a platform for a range of devices with applications in epilepsy, spinal cord injuries and other neurological disorders.” In one such version, electrodes in contact with the brain could detect seizures as they start and deliver a series of pulses to shut the seizure down.