Reports from Other Journals
Research Advances by Angela G. King
DNA Computing Targets West Nile Virus, Other Deadly Diseases, and Tic-Tac-Toe A team of scientists from Columbia University Medical Center in New York and the University of New Mexico, Albuquerque, say that they have developed a DNA-based computer that could lead to faster, more accurate tests for diagnosing West Nile Virus and bird flu. Representing the first “medium-scale integrated molecular circuit,” it is the most powerful computing device of its type to date. The new technology could be used in the future, perhaps in five to ten years, to develop instruments that can simultaneously diagnose and treat cancer, diabetes, or other diseases. Scientists have tried for years to build computers out of DNA (deoxyribonucleic acid), nature’s chemical blueprint for life. But getting nano-sized pieces of DNA to act as electrical circuits capable of problem-solving like their silicon counterparts has remained a major challenge. “This is a big step in DNA computing,” says Joanne Macdonald, a virologist who led the research team that developed MAYA-II (Molecular Array of YES and AND logic gates)—a “computer” whose circuits consist of DNA instead of silicon. She likens the significance of the advance to the development of the earliest silicon chips. “The study shows that large-scale DNA computers are possible.” “These DNA computers won’t compete with silicon computing in terms of speed, but their advantage is that they can be used in fluids, such as a sample of blood or in the body, and make decisions at the level of a single cell,” says Macdonald, whose work is funded by the National Science Foundation. She is currently using the technology to improve disease diagnostics for West Nile Virus by building a device to quickly and accurately distinguish between various viral strains, and she hopes to use similar techniques to detect new
strains of bird flu. In the future it is possible that DNA computers could be implanted in the body to first diagnose and then kill cancer cells or to monitor and treat diabetes by dispensing insulin when needed. In a series of laboratory demonstrations over a two-year period, Macdonald and her associates showcased the computer’s potential by engaging MAYA-II in a complete game of tic-tac-toe against human opponents. Shown in the foreground of Figure 1 is a cell-culture plate containing pieces of DNA that code for possible “moves”; a display screen (background) shows that the computer (red squares) has won the game against its human opponent (blue). Composed of more than 100 DNA circuits, MAYA-II is quadruple the size of its predecessor, MAYA-I, a similar DNA-based computer developed three years ago by the research team. With limited moves, the first MAYA could only play an incomplete game of tic-tac-toe. The experimental device looks nothing like today’s hightech gaming consoles. MAYA-II consists of nine cell-culture wells arranged in a pattern that resembles a tic-tac-toe grid. Each well contains a solution of DNA material that is coded with “red” or “green” fluorescent dye. The computer always makes the first move by activating the center well. Instead of using buttons or joysticks, a human player makes a “move” by adding a DNA sequence corresponding to a specific move to all eight remaining wells. The well chosen for the move by the human player responds by fluorescing green, indicating a match to the player’s DNA input. The move also triggers a strategic counter-move by the computer in one of the remaining wells, which fluoresces red. Each move takes about 30 minutes.
More Information 1. Macdonald, Joanne; Li, Yang; Sutovic, Marko; Lederman, Harvey; Pendri, Kiran; Lu, Wanhong; Andrews, Benjamin L.; Stefanovic, Darko; Stojanovic, Milan N. Medium Scale Integration of Molecular Logic Gates in an Automaton. Nano Lett. 2006, 6, 2598–2603. 2. Background information on this concept has been published in this Journal. See Ward, Michael D. Chemistry and Molecular Electronics: New Molecules as Wires, Switches, and Logic Gates. J. Chem. Educ. 2001, 78, 321–328. 3. Additional information on this research can be found online at http://cpmcnet.columbia.edu/news/in-vivo/Vol2_Iss15_sept29_03/ index.html (accessed Nov 2006).
Marijuana Component May Offer Hope for Alzheimer’s Disease Treatment
Figure 1. Prototype DNA computer MAYA-II. Photo courtesy of Columbia University Medical Center.
378
Journal of Chemical Education
•
Scientists are reporting discovery of a previously unknown molecular mechanism in which the active ingredient in marijuana may slow the progression of Alzheimer’s disease (AD). Kim D. Janda and colleagues at Scripps Research
Vol. 84 No. 3 March 2007
•
www.JCE.DivCHED.org
Structure: by A. King
Figure 2. The structure of tetrahydrocannabinol (THC), the active component of marijuana.
Institute used laboratory experiments to show that delta-9tetrahydrocannabinol (THC) inhibits amyloid fibril formation and preserves brain levels of the key neurotransmitter acetylcholine. One neuropathological characteristic of AD is the formation of amyloid plaques in areas of the brain responsible for cognition and memory function. Molecules that prevent -amyloid peptide (A) aggregation, and thus plaque formation, are candidates for drug research. Since AD impairs the cholinergic neurotransmitter system, molecules that interact with it are also AD drug candidates. Existing medications for AD, including donepezil and tacrine, relieve AD symptoms by inhibiting acetylcholinesterase (AChE), which breaks down acetylcholine. THC does so by inhibiting a different site on acetylcholinesterase from the existing medications and at lower concentrations, Janda’s group reports. Their experiments show that THC also prevents formation of the amyloid plaques that are a hallmark of AD and its damage to the brain. In addition to affecting acetylcholine levels, AChE plays a role in Alzheimer’s disease by serving as a chemical chaperone, accelerating the formation of amyloid fibrils and forming stable complexes with Ab at the region denoted as the peripheral anionic binding site (PAS). If nonpolar THC binds to the PAS region of AChE, it could retard the formation of the fibrils. Janda’s team began by modeling THC binding to AChE in silico. The results predicted that the fused ring system of THC and the indole sidechain of a tryptophan residue of AChE would bind, with additional interactions between THC and AChE backbone carbonyls. Encouraged by the modeling results, scientists conducted a steady-state kinetic analysis that revealed that TCH competitively inhibits AChE, with a Ki = 10.2 M. A thioflavin T-based fluorometric assay was used to stain putative A fibrils to measure the ability of THC to prevent A aggregation. THC completely blocked the promotion of A aggregation by AChE. In this role it performed even better than propidium, one of the best known aggregation inhibitors. “Our results provide a mechanism whereby the THC molecule can directly impact Alzheimer’s disease pathology,” the team states. They also note that THC may provide a “drug lead”—a model for developing new and more effective medications with more targeted effects on AD. The researchers explain that such compounds “may provide an improved therapeutic for Alzheimer’s disease, augmenting acetylcholine levels by preventing neurotransmitter degradation and reeduwww.JCE.DivCHED.org
•
Figure 3. Predicted binding mode of THC (gray) to AChE (orange ribbon). The catalytic residues of AChE (green) and water molecules included in the docking calculations (light blue spheres) are shown. Reprinted with permission from Mol. Pharm. 2006, 3, 773– 777. Copyright 2006 American Chemical Society and American Institute of Chemical Engineers.
cating amyloid beta aggregation, thereby simultaneously treating both the symptoms and progression of Alzheimer’s disease.” Alzheimer’s disease is currently the leading cause of dementia among the elderly. While an estimated 4.5 million persons in the U.S. suffer from Alzheimer’s at this time, by 2050 the number is expected to balloon to 11–16 million. The average lifetime care cost for an Alzheimer’s patient is $174,000. Direct and indirect annual costs of caring for individuals with Alzheimer’s disease are at least $100 billion, according to estimates used by the Alzheimer’s Association and the National Institute on Aging.
More Information 1. Eubanks, Lisa M.; Rogers, Claude J.; Beuscher, Albert E., IV; Koob, George F.; Olson, Arthur J.; Dickerson, Tobin J.; Janda, Kim D. A Molecular Link between the Active Component of Marijuana and Alzheimer’s Disease Pathology. Mol. Pharm. 2006, 3, 773–777. 2. Professor Janda’s research is described at http:// www.scripps.edu/chem/janda/ (accessed Nov 2006). 3. Information on Alzheimer’s disease, research, and advocacy can be found at http://www.alz.org/ (accessed Nov 2006).
New Wound Dressing May Lead to Maggot Therapy—Without the Maggots Scientists in the United Kingdom have developed a new wound dressing that could bring the benefits of maggot therapy to patients without putting live Greenbottle fly (blowfly) larvae into non-healing wounds. The joint research
Vol. 84 No. 3 March 2007
•
Journal of Chemical Education
379
Chemical Education Today
Reports from Other Journals
Figure 4. (A) Photomicrographs illustrating the application and removal of maggots from a chronic wound during a typical biotherapy treatment. (B) A schematic representing the delivery of maggot extract onto model wounds in monolayer cell culture. The cradle held the hydrogel 1 mm away from the cells. Reprinted with permission from Biotechnol. Prog. 2006, 22, 1690–1696. Copyright 2006 American Chemical Society and American Institute of Chemical Engineers.
project of Stephen Britland and Annie Smith from Bradford University, David Pritchard of Nottingham University, and colleagues from the biotechnology company AGT Sciences Ltd developed and conducted preliminary testing of the new hydrogel dressing that is based on maggot therapy but does not involve live maggots. Non-parasitic larvae, such as those of the Greenbottle fly, have historically been used to clean and debride wounds, particularly wounds that will not heal. While performing this role, the larvae also secrete antibacterial mixtures that prevent microorganisms from infecting the wound. The researchers noted resurgence in medical use of larval biotherapy — intentionally introducing blowfly maggots into non-healing wounds to clean away dead tissue. Medical use of the technique led to observations suggesting that maggots’ excretions and secretions (ESs) also may encourage regeneration of tissue and wound healing. Researchers, led by Stephen Britland, used computer software to measure the size of model wounds in cell cultures with and without ESs added to the culture medium. They found that the optimal dose of 50 g/mL dramatically decreased the time needed for the wound to close. Quantitative image analysis revealed that the active substances within the maggot secretions accelerated the closure of model wounds in cell culture. Previous zymography and protease inhibitor studies showed that this effect was due at least in part to the enzymes within the secretions, most likely the serine protease chymotrypsin. The mechanism of action could be
380
Journal of Chemical Education
•
Figure 5. Effect of maggot extract (ES) released from a prototype hydrogel wound dressing on the rate of closure of model wounds in cell cultures. Data shown are means ⫾ SEM of surface area of the wound remaining after 12 h of treatment with hydrogel sheets impregnated with 1 mg/mL (ES) or controls containing vehicle buffer only. Reprinted with permission from Biotechnol. Prog. 2006, 22, 1690–1696. Copyright 2006 American Chemical Society and American Institute of Chemical Engineers.
through protease activated receptors (PARs) expressed by these cells, or alternatively through protease-dependant substratum degradation mimicking the provisional matrix of healing wounds. Proteolytic products of extracellular matrix molecules, some of which have analogs in serum, are known to be motogenic for cells. That the proteases were in fact stimulating cell motility was manifest by upregulation of signal transduction pathways dependent upon phosphotyrosine activity revealed by immunocytochemistry. That the stimulatory effect of the maggot secretions did not influence cell division (mitogenesis), a very important factor in tissue regeneration, was confirmed by the MTT cell viability assay for cell proliferation. However there are drawbacks and barriers to using live larva to introduce the ESs to a wound. They remain in this stage of their life cycle for only 2–3 days and cannot be stored for long before application. Historically, attempts to administer ESs, known at the time as “maggot active principle” were ineffective due to a standard form of administration. Realizing that the ESs would have to be delivered in a controlled fashion, Britland’s group developed a hydrogel dressing, which slowly releases maggot ESs. The scientists cast a poly(vinyl alcohol-co-vinyl acetate) polymer hydrogel, and cut the hydrogel into 5 mm disks. (Hydrogels are colloidal gels in which water is the dispersion medium.) The disks were dehydrated, and then rehydrated in a solution 1 mg/mL ES in phosphate buffer or untreated buffer as a control. This con-
Vol. 84 No. 3 March 2007
•
www.JCE.DivCHED.org
Chemical Education Today
centration of ES produced disks that released into cell cultures 50 g/mL ES, the previously established optimal dose. For studies, the disks were suspended with well inserts above cell cultures. Colorimetric protease assay showed a time-dependent release of ES in the disc supernatant, and treated cell cultures showed a significant increase in wound closure. “The present prototype hydrogel wound dressing could potentially be deployed as a device to deliver insect-derived active products to skin wounds in vivo to encourage tissue regeneration,” says Britland. The prevalence of chronic (intractable) wounds is astonishingly high, and the quality of life implications for the patients potentially treatable by a commercially developed ES-based hydrogel merits further exploration. “It has been reported that in the UK at any one time 1% of the population will have a hard-to-heal wound costing the nation (through its National Health Service) several billion pounds per annum to treat. Extrapolating these figures to the rest of the developed world speaks for itself.”
www.JCE.DivCHED.org
•
More Information 1. Smith, Annie G.; Powis, Rachel A.; Pritchard, David I.; Britland, Stephen T. Greenbottle (Lucilia sericata) Larval Secretions Delivered from a Prototype Hydrogel Wound Dressing Accelerate the Closure of Model Wounds. Biotechnol. Prog. 2006, 22, 1690– 1696. 2. Hydrogels also have use in cataract surgery. See J. Chem. Educ. 2005, 82, 346. 3. An undergraduate lab isolating PVA is available in this Journal. See Chen, Yueh-Huey; Yaung, Jing-Fun. A Polymer in Everyday Life: The Isolation of Poly(vinyl alcohol) from Aqueous PVA Glues. An Undergraduate Chemistry Experiment. J. Chem. Educ. 2006, 83, 1534–1536.
Angela G. King is Senior Lecturer in Chemistry at Wake Forest University, P.O. Box 7486, Winston-Salem, NC 27109;
[email protected].
Vol. 84 No. 3 March 2007
•
Journal of Chemical Education
381
