# News this Week

Science  05 Nov 2004:
Vol. 306, Issue 5698, pp. 952
1. PLANETARY SCIENCE

# Titan Remains Mysterious With a Hint of the Familiar

1. Richard A. Kerr

Planetary scientists knew that Saturn's huge, haze-enshrouded moon Titan—by scientists' reckoning the solar system's “largest expanse of unexplored terrain”—would prove to have a most unearthly makeup. From theory and observation, they had recognized or inferred a cold-hardened bedrock of ice, liquefied-methane clouds, a gasoline-like drizzle, and a coating of pervasive organic goo. Still, as the Cassini spacecraft now orbiting Saturn homed in last week for its first close pass by Titan, scientists expected to glimpse reasonably familiar features, including continents, seas, clouds, and rain. Guess again.

After staring at the images returned by Cassini for a few bleary-eyed hours, “we're not quite sure what we're looking at,” said camera team leader Carolyn Porco of the Space Science Institute in Boulder, Colorado. “There isn't much we're absolutely confident about right now.” Two days' more reflection didn't help much. Team members did become confident that Titan has a youthful surface reshaped by surprisingly active geological processes, a couple of which look similar to other icy moons of the outer solar system. “Things are starting to make a minute amount of sense,” said planetary geologist Laurence Soderblom of the Cassini team and the U.S. Geological Survey in Flagstaff, Arizona. Yet Cassini scientists have not found the most anticipated feature of Titan: seas, or at least lakes, of liquid hydrocarbons.

From the two flybys of the Voyager spacecraft in 1981 and later Earth-based observations, Titan looked as if it could be a fairly simple world. The organic chemical factory in its dense atmosphere, where solar radiation interacts with methane, would account for the enveloping yellowish haze and scattered clouds, presumably composed of methane and ethane. At Titan's surface temperature of 94 degrees above absolute zero, the haze particles of organic crud would settle onto the water-ice surface of brighter “continents” or into the darker hydrocarbon “seas.” This light-dark patterning could be seen from Earth in haze- penetrating infrared telescopic images. Astronomers could only speculate, given chemists' inference of methane-ethane rain, that Titan's dark regions were seas, although radar astronomers reported that their radar beams often glinted off the surface like sunlight on a sea. Geologists weren't counting on finding anything more on the continents than the inevitable cratering by large impacts; Titan has no strong source of internal heat that could still be reshaping the surface, say by repaving it with icy “lava,” as happens on some other icy moons.

On first examining the Cassini data, team members found anything but the expected. “When you increase [imaging] resolution by a factor of 10,” said Porco, “you always find something new. It's just that we can't figure out what that is.” The camera found only the rare, fleeting cloud outside the south polar region, and the Visible Infrared Mapping Spectrometer (VIMS) could detect no methane in the polar clouds. Surprisingly, neither Cassini's imaging radar nor its camera could find any obvious impact craters. “Maybe there's something burying them,” said interdisciplinary science team member Jonathan Lunine of the University of Arizona (UA), Tucson.

That something could have been an eon's worth of haze particles settling to the surface. Radar team leader Charles Elachi, director of the Jet Propulsion Laboratory in Pasadena, California, reported that along the strip of surface probed by the radar's altimeter, the surface is so flat that its height varied by only 100 meters over many hundreds of kilometers. And the radar, operating in its radiometer mode, found a heat-emission signature consistent with a coating of organic material rather than bare ice. One area, at least, “is really covered in organics,” said radar team member Ralph Lorenz of UA.

Hundreds of meters of organic crud might explain the flatness of the region the altimeter surveyed, but where the camera and radar returned images of areas other than those surveyed by the altimeter, something more geologically active is suggested. “We have images of very complex geological processes,” says Lunine. “There's enormous diversity [of features] in just one area of the radar data.” There are what may be ridges squeezed up from a plain, says Soderblom, and bright “pieces of string” squiggling across the landscape that are familiar from Neptune's big moon Triton. The strings may be steep scarps eaten into the edges of thin layers of icy “volcanic” flows, he says. In both radar and VIMS images, there are strong hints of so-called grooved terrain that the Voyagers imaged over large parts of Jupiter's Ganymede and other outer-solar-system moons. And Elachi reported that some areas of Titan that are dark in radar images, at least, give a “kind of impression of lakes of liquid.”

Planetary geologist James Head of Brown University in Providence, Rhode Island, agrees that Titan's apparently grooved terrain reminds one of Ganymede's, but he notes that there is still no agreement about how those structures form on Ganymede. And the Cassini camera has yet to see sunlight glinting off the surface of Titan, as it must if much of the moon is covered by liquid. Indeed, most of the flyby analysis so far may be flat wrong, says Soderblom, who has long experience with Voyager flybys of icy moons. “This is certainly among the strangest worlds of the solar system,” he says.

Cassini team members agree that such strangeness will no doubt demand patient integration of observations from the spacecraft's dozen instruments during the 44 Titan flybys planned for the next 4 years. And those data will be anchored by the observations to be made by the Huygens probe as it drifts to the Titan surface on 14 January after release from Cassini in December. There it may add to the glamour of the mysterious Titan: What could be stranger than an alien craft tossed on a wine-dark sea of liquefied natural gas?

2. INFECTIOUS DISEASES

# Ducks May Magnify Threat of Avian Flu Virus

1. Dennis Normile

The H5N1 avian influenza virus that has devastated poultry flocks in Asia and raised the specter of a global human pandemic seems to be becoming more dangerous. New lab experiments suggest that the virus can replicate copiously in the guts of domestic ducks without making them sick. What's more, this year's strain survives longer in the environment than last year's, and ducks shed larger quantities of it. Together, the findings indicate that controlling the virus could be even more challenging than previously thought.

“This requires a rethinking of the entire strategy to control H5N1 in poultry,” says Klaus Stöhr, coordinator of the World Health Organization's (WHO's) Global Influenza Program. “We have to ask if it will be possible to eliminate the virus among chickens if there is a reservoir nearby which doesn't show it is harboring the virus.”

The experiments, carried out at the St. Jude Children's Research Hospital in Memphis, Tennessee, a WHO collaborating center for animal influenza viruses, have not yet been published, but WHO went public with the findings because of their potential impact on animal and public health efforts.

Although disconcerting, the findings are not completely unexpected. K. S. Li of Shantou University Medical College in China and colleagues at the University of Hong Kong and other institutions reported in Nature on 8 July that asymptomatic domestic ducks sampled at live animal markets in southern China were shedding H5N1, suggesting that ducks could be a key factor in the transmission of the virus.

In the new lab study, researchers infected ducks with several strains of the H5N1 virus collected from chickens and humans earlier this year and compared the results to those of a separate study that used strains collected in 2003. Ducks infected with the 2004 strains shed more virus for longer time periods than did ducks infected with earlier strains. And most of the ducks showed no signs of illness—a surprise for a virus that is so pathogenic in chickens it typically kills them within 24 hours of infection.

Yi Guan, a microbiologist at the University of Hong Kong and a member of the group that published the July Nature paper, notes that past strains of H5N1 did kill ducks. “It is natural that the virulence of the virus is gradually decreasing in aquatic birds,” he says, because viruses that cause rapid death have less chance to reproduce. But he says it is unusual and worrying that the virus is compatible in one bird species yet lethal in another when the two are often raised together. The 2004 virus also seems to have become more stable, surviving in the environment for 6 days at a temperature of 37°C, compared to 2 days for older strains.

Stöhr says additional studies are needed to determine how widespread the virus is in domestic duck flocks in Asia, if the virus is moving between ducks and migratory birds, and what risks infected ducks pose to humans. Although there have been no proven cases of viral transmission from ducks to humans, WHO now says it must be considered when tracking the route of human infections.

The United Nations Food and Agriculture Organization already suggests that chickens and other poultry species be raised separately. But enforcing this recommendation is difficult. In many Asian countries, free-ranging ducks and chickens mingle and frequently share the same water supplies. So far, says Stöhr, only Thailand is trying to get farmers to keep ducks and chickens apart.

3. SCIENCE POLICY

# Nanotech Forum Aims to Head Off Replay of Past Blunders

1. Robert F. Service

Stung by memories of the bruising battles over genetically modified organisms (GMOs), leaders from industry, academia, and environmental organizations met in Houston, Texas, last week to launch a new forum for hashing out concerns over nanotechnology, the nascent field of building materials up from the atomic scale. The rollout of the International Council on Nanotechnology (ICON) had its share of hiccups. Three environmental organizations balked at becoming founding members, charging that ICON is too industry-focused. But two of those groups still took part in the meeting as “guests” and say they will consider joining down the road, a development that most see as an improvement over the GMO fiasco.

“All of the parties have a significant amount of goodwill and want to talk. That's not a bad starting point,” says Pat Mooney, who heads the ETC Group, an environmental organization based in Ottawa, Canada, that spearheaded the attack against GMOs and has advocated a cautious approach to nanotechnology. “There is a lot of trust that needs to be built from all sides,” says Stephen Harper, who heads environmental, health, and safety policy for the computer chip giant Intel in Washington, D.C.

Researchers with Rice University's Center for Biological and Environmental Nanotechnology dreamed up ICON to begin building that trust, write reports to help regulatory agencies, and possibly even fund original research. But Mooney and other environmentalists say they were initially wary because ICON's funding comes from industrial members, and some of the proposed initial research projects seemed aimed at convincing the public that nanotechnology is safe rather than addressing basic concerns about the revolutionary technology.

At the Houston meeting, participants avoided potential flash points—such as backing safety studies or researching whether nanotechnology will benefit developing countries—to focus initially on a subject virtually everyone could agree on: determining how to describe various nanosized clumps of matter. Nomenclature is particularly tricky for nanomaterials because different sized nanoparticles of a material often have drastically different properties. Straw-shaped carbon nanotubes, for example, conduct electricity either as semiconductors or metals depending on the pitch at which the atoms wind around the straws. Such complexity bedevils agencies responsible for regulating the handling, manufacture, and release of nanomaterials.

Many nanotechnology experts laud ICON's early focus on language. “It will be the key to getting the regulations right. It's exactly what needs to be done,” says David Rejeski, director of the foresight and governance project at the Woodrow Wilson International Center for Scholars in Washington, D.C., another organization working to build bridges between disparate nanotechnology stakeholders. But with hundreds of products containing nanomaterials already on the market and the field expected to balloon to a $1 trillion industry in less than a decade, Rejeski says, much more needs to be done. Mooney and other environmentalists urge governments to act quickly to ensure that products don't pose health and environmental threats and to include countries around the globe in discussions about how this emerging industry will affect their economies and societies. Other nanotechnology forums are springing up to tackle those issues. In February, for example, the Rockefeller Foundation is co-sponsoring a meeting in Alexandria, Egypt, called the Global Dialogue on Nanotechnology and the Poor. Leaders from governments and other organizations will grapple with problems such as ensuring access to revolutionary technologies and promoting research that benefits poor people as well as rich—and hope the cautionary tale of GMOs gives this story a happier ending. 4. FRENCH PROTESTS # Researchers Back a 70-Page Agenda to Reform Agencies, Boost Careers 1. Barbara Casassus* 1. Barbara Casassus is a writer in Paris. PARIS—Never before have French scientists been as single-minded as they were at a 2-day gathering last week in the Alpine town of Grenoble, when more than 900 of them voted for a raft of proposals on everything from cash to careers in the name of overhauling fundamental research. With remarkable unanimity, they approved a wish list for François Fillon, minister for education and research, who has promised to take a reform bill to Parliament next spring. This effort builds on last year's showdown with the government in which more than 2000 lab directors and research team leaders quit their administrative duties to protest funding and staffing cutbacks (Science, 16 April, p. 368). After the government backed down, hundreds of scientists across the country organized into working groups to prepare a string of reports, which were then honed into last week's proposals. They call for more coherent government oversight and stronger support of scientific careers. The proposals include creating a single research and higher education ministry, an independent higher science council to advise the government on strategy, a new body to evaluate all researchers, a long-term jobs plan for researchers, and more crossover between agency and academic science. They ask that lecturers' teaching loads be halved and that universities be reformed in depth. Doctoral students, who now have no health or social security coverage, should be given proper pay and working conditions, and postdocs should be given associate researcher contracts of up to 3 years. But how much of the wish list will find its way into the parliamentary bill is another matter. “We don't know what the government will do with it, but we shall remain vigilant,” says Alain Trautmann, co-director of the cell biology department at the Cochin Institute and spokesperson for the protest movement. Although Fillon told the Grenoble conference he would also take account of some 20 other reform proposals, Trautmann says, “ours outstrip the others by a wide margin” in representing the community. All the major political parties are paying attention: Their leaders were on hand last week, echoing a commitment to raise research spending to the European Union goal of 3% of gross domestic product (GDP) by 2010. And researchers were warmed by Fillon's clear admission that the government made mistakes last year when his predecessor Luc Ferry and former research chief Claudie Haigneré were in office. Although the show of hands in favor of the reform proposals was almost unanimous, the preceding 2 days of debate were far from docile. They were peppered with applause, boos, and a rowdy protest from an antiscience lobby that was silenced when the demonstrators were hustled from the conference room. The high spot, some delegates said, was the summing-up by Edouard Brézin, French Academy of Sciences vice president and co-president of the Initiative and Proposal Council (CIP), which was formed earlier this year to produce a consensus for change among scientists. “It was a moment of great emotion,” says Trautmann. “We had to produce [a document] that contained neither overspecific recommendations nor a compromise that was so general we would have looked ridiculous,” said Brézin, who will take over as academy president from endocrinologist Etienne-Emile Baulieu next January. “We have succeeded in drawing up precise and realistic proposals and have not simply issued a union-type demand for more money, more posts.” The unions are happy with the result. The proposals “constitute a working framework for the scientific community,” says the leading research union, SNCS. But university presidents are far from happy. They regret “the absence of important elements” they had suggested, such as merging agency and university researcher status to replace the separate agency and university academic researcher categories, and turning the leading agencies into mere suppliers of cash. CIP is putting finishing touches to the 70-page document before presenting it to Fillon and Research Minister François d'Aubert on 9 November. Most changes are modest, but the wording may prompt further discussion. For example, the report will explicitly reject the idea of making the National Research Agency, which will be created next year, the lead provider of research funds. Instead, delegates agreed, government subsidies to agencies should continue to provide the bulk of the cash. Even all that would not be enough to put French research back on track, according to Baulieu. He told a parliamentary conference on Tuesday that the government must inject an extra €1 billion into research each year for 5 years, or a total of €15 billion, if it is to achieve the 3% of GDP target. The French reform game is clearly far from over. 5. HIGH-ENERGY ASTROPHYSICS # Hot on the Trail of Cosmic Rays 1. Daniel Clery CAMBRIDGE, U.K.—Researchers are closing in on finding the source of galactic cosmic rays, charged particles that whiz around the galaxy in huge numbers and constantly bombard Earth's upper atmosphere. Images published this week in Nature show the production of high-energy gamma rays around the remnant of a supernova, known as RX J1713.7-3946. The pictures, taken with a new gamma ray telescope called the High Energy Spectroscopic System (HESS) in Namibia (Science, 3 September, p. 1393), mark the first time researchers have produced a resolved image of a supernova remnant at such high energies. Astrophysicists believe that these gamma rays, with energies of about 1012 electron volts (TeV), are produced at the same time as cosmic rays and so mark the location of their source, but they haven't got proof yet. “This strong signal is a breakthrough,” says Karl Mannheim of the University of Würzburg in Germany. “But there are many open questions.” Cosmic rays travel at speeds produced in the most powerful particle accelerators. Theorists believe that when particles streaming out of a supernova remnant hit interstellar gases, protons and other light nuclei get boosted by the shock wave and produce a few TeV gamma rays as a byproduct. The problem is, electrons streaming from a supernova remnant can also generate TeV gamma rays, without cosmic rays being involved. The HESS team should be able to figure out whether protons or electrons are the culprits by studying the supernova at other wavelengths, such as radio waves, to figure out the density of matter around it. “We're just gearing up that,” says HESS spokesperson Werner Hofmann of the Max Planck Institute for Nuclear Physics in Heidelberg. 6. RUSSIAN ACADEMY OF SCIENCES # Putin: Reform Begins at Home 1. Andrey Allakhverdov, 2. Vladimir Pokrovsky* 1. Andrey Allakhverdov and Vladimir Pokrovsky are writers in Moscow. MOSCOW—President Vladimir Putin has begun a campaign to trim Russia's vast network of state-funded research institutions. Although there is general agreement that the current system is broken, no consensus exists on what is needed. Many scientists worry that Putin's proposed fix could do serious damage to the Russian Academy of Sciences (RAS), whereas others argue that additional structural changes are also necessary. Speaking here last week to the United President's Council on Science, Technology, and Education, Putin squelched rumors that the government wants to eliminate RAS (Science, 29 October, p. 795). “Nobody wants to ruin RAS,” he said. “We want to preserve the academy. The question is how to adapt it to present realities.” That could be hard work, he acknowledged. “We expect a thoughtful and effective modernization. We expect proposals on a drastic reorganization of work.” Putin's comments appear to give a green light to a plan, formally known as the Concept of the Management of State Research Organisations, that would privatize, merge, sell off, or close most of the academy's roughly 5000 research institutions while increasing support for the 100 to 200 institutes that remain (Science, 24 September, p. 1889). Science and education minister Andrey Fursenko has promised a$26,000 boost by 2008 in per capita spending—including salaries, equipment, and other costs of doing science—for the nearly 150,000 scientists expected to remain at RAS. At the same time, however, the proposal would put RAS on a shorter leash. The Russian cabinet would be given the authority to approve RAS's operating charter, for example, and to ratify the choice of an RAS president by academicians.

Such drastic moves have drawn criticism from RAS trade unions, which have demanded Fursenko's resignation. They also prompted an angry letter to Putin from a number of Russian Nobel Prize winners, who fear that RAS will be swallowed up by the Russian bureaucracy. “We indeed are worried about this concept, which has scared everyone so much,” says 2003 physics Nobelist Vitaly Ginzburg. “The academy must in no circumstances become a state department; this is the main point.”

Shortly after the meeting, RAS reported changes to its leadership that are believed to reflect Putin's new priorities. Valery Kozlov, former RAS vice president of youth policy, was appointed vice president in charge of the RAS restructuring. Vice president Gennady Mesyats, a fierce opponent of the reform plan, saw his authority over financial issues handed to Alexandr Nekipelov, who was appointed vice president. Nekipelov refused comment on the changes, and Mesyats was unavailable.

In the next few months the Russian Duma (parliament) is expected to take up changes in the tax laws and other provisions that would be needed to implement the Concept. But Boris Saltykov, who pushed unsuccessfully for reform as Russia's first science minister in the mid-1990s, believes that even stronger medicine is needed to correct what ails RAS. “If you give money to an old Soviet quasi-structure like RAS, it will be used as ineffectively as in the past and will not have any substantial effect,” he says. “While it is necessary to retain some form of RAS, its very structure must be reformed. But no one even speaks about structural reform.”

7. EUROPEAN UNION

# New Research Commissioner Caught in Controversy's Wake

1. Gretchen Vogel

BERLIN—Janez Potocnik should be safely installed as the European Union's (E.U.'s) new research commissioner by now, but his future remains in question after the entire slate of new commissioners, the E.U.'s policymaking body, was pulled last week just hours before an approval vote by the European Parliament. The unprecedented move was forced on President-designate Jose Manuel Barroso following controversial comments by the nominee for justice. The would-be commissioners now face a shakeup; the final outcome is unlikely to be settled before early December.

The Parliament has only a yes or no vote on the entire 25-member commission, so the furor over Italian nominee Rocco Buttiglioni's comments on homosexuality and the role of women forced Barroso to withdraw his whole slate before an expected no vote on 27 October. The comments were enough to convince a majority of the Parliament that he was not fit to oversee the E.U.'s antidiscrimination laws.

Buttiglioni resigned on 30 October, but the aftershocks of his replacement could trigger further changes. Several other proposed commissioners got low marks from Parliament, and Barroso must now decide whether to move some commissioners to new positions in an effort to find a slate likely to win the Parliament's approval. There is a chance that Potocnik could be shifted in the resulting reorganization.

Many European scientists are hoping that doesn't happen. In contrast to Buttiglioni, Slovakia's Potocnik got high marks from the Committee on Industry, Research, and Energy that questioned him. During his 3-hour hearing on 1 October, he voiced support for the creation of the European Research Council (ERC) that would fund basic research (Science, 29 October, p. 796) and a doubling of the budget for the next Framework program, which funds E.U. science (Science, 25 June, p. 1885).

Potocnik has also gotten positive early marks from observers outside the E.U. bureaucracy. “I was very favorably impressed,” says Jose Mariano Gago, former Portuguese science minister and head of the Initiative for Science in Europe, a group lobbying for the development of the ERC, who met with Potocnik last month. “I think he's a man of great capacity. He is totally committed to advancing scientific development in Europe.” Europe's scientists will have to wait to see if he gets a chance to do so.

8. SCIENCE POLICY

# Japan to Host Global Forum on Societal Impacts

1. Dennis Normile

TOKYO—It may be a truism that scientific progress has widened the gap between the industrial and the developing world. But next week in Kyoto, a group of Nobelists, government officials, and business leaders will discuss how scientific advances might also be used to bridge that gap.

The occasion is the first Science and Technology in Society forum. The driving force behind the project is Koji Omi, a veteran Japanese legislator and former science minister who envisions the forum as an annual event. Rather than just assembling scientists and engineers, including nine Nobel laureates, Omi decided to reach out to media representatives, politicians, and captains of industry, too. In three preparatory meetings, a group of 40 founding members—including Sydney Brenner of the Salk Institute in La Jolla, California; Lu Yongxiang, president of the Chinese Academy of Sciences; David King, scientific adviser to the U.K. government; and Henry McKinnell, CEO of Pfizer Inc.—agreed to concentrate initially on five areas: energy and the environment, bioethics, information technologies and their social impact, capacity building, and human health and bioterrorism issues. “Our hope is to build a network of people who will think deeply about these problems,” says Omi.

Bruce Alberts, president of the U.S. National Academy of Sciences and also a founding member, says that bridging the divide between advanced and developing countries is an important subtheme of the forum, which will feature twinned sessions to explore particular problems through the perspective of both advanced countries and the developing world. For example, speakers from Japan, the United Kingdom, and Germany will lead a discussion on “The Promise of Clean Energy,” while officials from India, China, and Sudan will offer their views on “The Challenge of Meeting Energy Needs in Developing Countries.” Alberts views the forum as “an experiment,” with the test being whether participants use it as a springboard for tackling intractable global problems such as the digital divide.

That kind of exchange is also the fond wish of 1999 chemistry Nobelist Ahmed Zewail, a native of Egypt and a professor at the California Institute of Technology in Pasadena. Zewail, another founding member, hopes this “dialogue of cultures” will lead to more “support for science education and the technologies that can help developing countries.”

The 11 to 14 November meeting is by invitation only. The Japanese government put up about a quarter of the $4 million needed for the meeting, with Japanese businesses contributing the rest. 9. PLANT BIOLOGY # NO-Making Enzyme No More: Cell, PNAS Papers Retracted 1. John Travis May 2003 was an up-and-down month for Daniel Klessig. On 7 May, he resigned as president of the Boyce Thompson Institute (BTI) for Plant Research in Ithaca, New York, for health reasons. But 9 days later, his lab published a paper in Cell that promised to be a major advance. His team had identified a long-sought enzyme that enables plants to make nitric oxide, especially in response to infections. Last Friday, however, Klessig and some of his co-authors retracted the Cell paper and a subsequent one published in the Proceedings of the National Academy of Sciences (PNAS) that was dependent on the earlier research. “This is the worst experience I've had in my life, and that includes open-heart surgery,” says Klessig. According to Klessig, an “absolutely critical” experiment described in the Cell paper cannot be reproduced by members of his lab, and he now considers the experiment's data to be “shaky.” Meena Chandok, the first author on both papers, did not approve the retractions. BTI is investigating the matter. Klessig acknowledges that Chandok left his laboratory in April: “I can't say a lot more than what's in the retraction.” (Science was unable to reach Chandok.) The initial Cell paper was considered a breakthrough because plant biologists have recently come to recognize that nitric oxide may have as many roles as a signaling molecule in plants as it does in animals, where it regulates immunity, nerve cell communication, and much more. Yet investigators had been unable to identify a plant version of a nitric oxide synthase (NOS), the enzyme that creates the gas in animals. But in the Cell report, Chandok, Klessig, and their colleagues described a tobacco plant protein that produced nitric oxide in response to infection. “The paper was very important, as it was the first discovery of a nitric oxide synthase enzyme … in plants; it provided a mechanism for making NO in response to pathogen infection, and it indicated that plants have a novel mechanism for producing NO compared with animals,” says plant biologist Nigel Crawford of the University of California, San Diego. “This is a major setback for the field.” The tobacco protein identified by the Ithaca team bore little resemblance to animal NOS enzymes, but a follow-up paper by the group in the 25 May 2004 PNAS seemed to clinch the case, notes Crawford. They reported that suppressing the enzyme's activity made tomatoes more susceptible to a bacterium. In the Cell retraction, Klessig's team says that the data showing that the recombinant form of the tobacco protein has NO-making properties cannot be reproduced, and they call the original data in the Cell paper “unreliable.” Although Klessig's lab continues to study the protein and hasn't ruled out that it may be an NOS, he advises others not to pursue the matter for now. “It's important that the rest of the scientific community not base their research on this unreliable data that we are no longer confident in,” he says. In any case, the hunt for a pathogen- responsive NOS in plants may be over. Last October in Science, Crawford's team identified a gene in the mustard plant Arabidopsis thaliana that encodes a protein, dubbed AtNOS1, with a strong resemblance to a NOS found in snails, and they showed that it produces the gas in response to hormonal signals. Now, in a paper in the 2 November PNAS, a group headed by Jörg Durner of the GSF-National Research Center for Environment and Health in Neuherberg, Germany, reports that mustard plants exposed to the bacterial cell surface components activate AtNOS1 as a way to make NO and induce defensive genes. Mutating the gene left the plants more vulnerable to bacterial infection. “It's a quite convincing story,” says Klessig. 10. INFECTIOUS DISEASES # Ethicists to Guide Rationing of Flu Vaccine 1. Jennifer Couzin Despite emergency efforts by the Centers for Disease Control and Prevention (CDC), medical experts have not been able to solve a pressing numbers problem: how to distribute 61 million or so existing doses of flu vaccine among the 95 million Americans in high-risk categories. So, with only half the expected supply of vaccine available and flu season imminent, CDC has turned to another class of experts—ethicists. The agency, facing calls for guidance from state and local health officials as they struggle to ration the vaccine, has formed its first-ever ethics panel to help guide flu vaccine distribution. The five-member group held its first two meetings last week, and CDC expects to issue recommendations later this month. Although this panel is a stopgap measure, CDC officials are planning to establish a more permanent ethics committee, which will consider a range of issues, including vaccine distribution. The flu panel can't act soon enough. Over 40 million doses of flu vaccine have already been shipped by their maker, Aventis; CDC doesn't know how many have been administered. Last week, the Department of Health and Human Services announced that it had located 5 million more doses and was hoping to purchase them, if they meet safety criteria. Meanwhile, state health officials are struggling with difficult questions. Should an elderly individual, who is in danger of dying from the flu but has already lived much of his or her life, receive vaccine before a 2-year-old, who is likelier than most people to get sick but will probably survive? (Last year, 36,000 people died of flu in the United States; 150 were young children.) If a nursing home has only enough vaccine for either its staff or its patients, who should receive it? The crisis erupted on 5 October after British authorities banned distribution of vaccine made at Chiron's U.K. plant because of contamination. Within 4 hours of hearing the news, frantic CDC officials convened a meeting of their Advisory Committee on Immunization Practices (ACIP) to revise recommendations on who should get flu shots. Before the shortage, CDC put the number of recommended recipients at 185 million. Based on various studies assessing a person's risk of contracting and dying from flu, ACIP members managed to shrink it to 95 million. For example, the committee agreed that only very young children needed to be vaccinated and that healthy 50- to 64-year-olds didn't. Available data, however, didn't permit ACIP to reduce the number further. “ACIP and CDC will go as far as the science will allow us to go,” says Lance Rodewald, director of the immunization services division in CDC's national immunization program. “There was a sense that we needed some help.” Five ethicists have agreed to provide it: Daniel Callahan of the Hastings Center in Garrison, New York; John Arras of the University of Virginia in Charlottesville; Kathleen Kinlaw of Emory University in Atlanta, Georgia; Thomas Beauchamp of Georgetown University in Washington, D.C.; and Robert J. Levine of Yale University in New Haven, Connecticut. The panel is first focusing on the relative responsibilities of CDC and local health departments in making recommendations about rationing. The group may consider questions such as the life expectancy of different populations with and without the vaccine, says Levine. It may also examine how to prevent shortages of other vaccines in the future. Eight of 11 childhood vaccines, for example, have been in short supply in recent years. “What the CDC is doing now is something that we need,” says Paul Offit, chief of infectious diseases at Children's Hospital of Philadelphia. Although vaccinating health care workers—as most states are doing—can help contain influenza's spread, “what are the ethics of immunizing a healthy adult who works in the hospital and is unlikely to die?” Offit asks. The issues are tough enough when they concern routine flu immunization. The CDC panel will also likely consider how vaccine should be rationed if a deadly new flu pandemic emerges, possibly from the avian flu strains now circulating in Asia. There, CDC won't be alone: On 11 November, the World Health Organization will hold a summit with country officials and 16 vaccine companies to determine how to boost vaccine and antiviral supplies now. 11. TOXICOLOGY # EPA Criticized for Study of Child Pesticide Exposure 1. Erik Stokstad* 1. With reporting by Jocelyn Kaiser. Last month researchers began recruiting in Florida for a$7 million study of how children age 3 and younger are exposed to indoor pesticides and several other household chemicals. Data from the Children's Environmental Exposure Research Study (CHEERS) will help the U.S. Environmental Protection Agency (EPA) determine safe levels for infants and toddlers. But the study is under attack because it is partially funded by a chemical trade association, and because parents will not be informed of the potential danger to their children from continued use of pesticides in the home. “The nuances of this study raise a series of questions,” says environmental health researcher Bruce Lanphear of the University of Cincinnati, Ohio.

Although many studies have looked at occupational exposure to pesticides and other chemicals, little is known about how pesticides get into children's bodies. CHEERS hopes to fill that gap by following 60 children for 2 years. Parents will track what their children eat and how active they are, while also keeping track of the pesticides used in the house. Field teams will visit every 6 months to sample floors and other surfaces and to collect urine to analyze for metabolites of the pesticides. The study is being conducted in and around Jacksonville in northeastern Florida. Each participating family will be paid $970 and will get to keep the camcorder they used to record their child's movements. EPA originally planned to study only pesticides, but last month it received a$2 million contribution from the American Chemistry Council to add several other kinds of common household chemicals, including flame retardants, perfluorinated chemicals, and ingredients in plastic products. The council, which represents 136 chemical companies, says it wants any government regulation to be based on facts about exposure.

That arrangement “erodes EPA's independence of research to regulate chemicals,” says Richard Wiles of the Environmental Working Group (EWG), a nonprofit advocacy organization based in Washington, D.C. EWG also accused the agency of, in effect, paying parents to expose their children to pesticides.

EPA denies those charges. “People would not do this just for the money,” says Linda Sheldon, acting director of EPA's Human Exposure and Atmospheric Sciences Division. Although household use of pesticides is one criterion for enrollment, parents are not required to continue applying pesticides during the duration of the study, she adds.

The study has been approved by EPA's Institutional Review Board (IRB), which examined subject protection and other issues. Green lights were also given by IRBs for EPA's partners—the Centers for Disease Control and Prevention and the Duval County Health Department—and one of its contractors.

Sheldon says EPA approached the American Chemistry Council (ACC) for the additional funding because it has a program of research grants. ACC did not pick the additional chemicals, she says, and it will have no role in the study nor any say over publication. ACC's Carol Henry seconds that: “We have no control over the study.” Although EWG noted that an advisory committee will contain industry scientists, EPA picked them and will rely on them solely for technical advice, mainly about analytical techniques, says EPA's Roy Fortmann, a project leader.

Some researchers say the study has ethical shortcomings, especially because EPA doesn't plan to warn parents explicitly about the potential risks of pesticides to children. “It's unconscionable,” says Leo Trasande of Mount Sinai School of Medicine in New York, who works on biomonitoring and children's health. “EPA's policy is that if you apply pesticides according to the labels, then they should be safe for indoor use,” Sheldon says. Of course, it's the CHEERS study that will provide EPA with the data needed to determine ultimately what's safe for children.

12. PALEONTOLOGY

# Dinosaurs Under the Knife

With a wealth of good specimens now at their disposal, paleontologists are probing elegantly preserved fossil bone tissue for clues to how long-extinct animals grew and lived

With high-domed skulls built like battering rams, dinosaurs called pachycephalosaurs look for all the world as if they must have butted heads. Paleontologists imagined the males sparring for mates as bighorn sheep do, and the idea was bolstered by radiating bony structures that apparently strengthened the head against impacts. But did they actually knock noggins?

To find out, Mark Goodwin of the University of California, Berkeley, and John Horner of Montana State University in Bozeman did something that would give most museum curators the heebie-jeebies: They sawed open the skulls to examine the fossilized bone tissue. The answer was trapped within the domes, Goodwin says, and histology—the study of tissues—was the only way to get it.

Preserved in the bone, as in many fossils, was a beautiful record of the original tissue, down to the level of individual cells. That's beyond the resolution of computed tomography scanners. By studying pachycephalosaurs of various ages, Horner and Goodwin determined that the radial structures were ephemeral features associated with fast-growing bones of juveniles. There was no sign of stress to the skull bones, they reported in the spring issue of Paleobiology. “I didn't see any evidence that they head-butted,” Goodwin says. However, he and Horner did find bundles of so-called Sharpey's fibers, which anchor ligaments and also thick pads of keratin to bone. Horner and Goodwin speculate that this may have secured a crest to the top of the head, perhaps for display.

More and more paleontologists are putting their fossils under the knife—the rock saw, actually—to gain new insights into their biology. “The microstructure includes a tremendous amount of information,” says Armand de Ricqlès of the Université Paris VII. After removing a slice of bone, they glue it to a glass slide and then grind it until it is transparent. Studying this “thin section” of bone tissue with microscopes can explain the origin of strange structures, such as the thick heads of pachycephalosaurs and the plates of stegosaurs, and help test hypotheses about their function. “I get quite excited about the potential of using bone microstructure to flesh ancient animals out and make them more real,” says Anusuya Chinsamy-Turan of the University of Cape Town, South Africa.

Paleohistology is already shedding light on the question of how sauropods and tyrannosaurs attained their gigantic sizes and other evolutionary patterns. It can tell adult animals from juveniles, and it provides the only way to determine how old extinct animals were when they died and how quickly they grew—key questions for studying their population biology and ecology. “We're on the cusp of being able to learn a lot about the biology of these animals—things we thought we'd never be able to tease out of the bones,” says Lawrence Witmer of Ohio University College of Osteopathic Medicine in Athens. This week at the Society of Vertebrate Paleontology (SVP) annual meeting in Denver, Colorado, paleontologists unveiled a bumper crop of histological studies, from a possible determination of the sex of a Tyrannosaurus rex to identification of an island of dwarf sauropods. “This field is about to explode,” says paleobiologist Gregory Erickson of Florida State University, Tallahassee.

## Diverse tissues

Paleohistology has a long history. For 150 years, paleontologists have used the technique to classify ancient fishes. But not until the early 20th century did they begin to compare the microstructure of various fossil land animals. Studies by Rodolfo Amprino of the University of Turin, Italy, led to an observation in 1947 that is now called “Amprino's rule”: The rate of an animal's growth strongly influences the type of tissue deposited in its bones. A rapidly growing bone has many blood vessels. Its characteristic “fibrolamellar” texture is marked by quickly deposited fibers and holes that are then filled by bony structures called primary osteons. In contrast, a bone growing more slowly has a texture called lamellar-zonal with fewer blood vessels and a finely layered appearance.

Dinosaurs typically have bone tissue that more closely resembles the quickly growing bones of large birds and mammals than the slow, lamellar-zonal tissue of reptiles. In 1969 and in later papers, de Ricqlès suggested that the dinosaur bone tissue might indicate fast, continuous growth and an active metabolism like that of mammals and birds. This idea played an important role in the “renaissance” that changed the perception of dinosaurs from sluggish reptiles to active, possibly warm-blooded animals. Tissue type turned out not to be a simple indicator of metabolism, but it does indicate the general pace of growth.

Bone tissue offers another way to understand the growth of ancient animals. Bones sometimes lay down dark lines, called lines of arrested growth (LAGs), which represent periods when growth slowed or stopped for a while. LAGs are common in amphibians and reptiles, but modern birds typically lack them because they complete their growth in less than a year. In 1981, Robin Reid of Queen's University of Belfast reported that dinosaurs showed the lines, too. By counting them like tree rings, paleontologists can infer how many years a bone has grown, and by extension how long the dinosaur lived. This technique of skeletochronology is now widely used by biologists studying modern reptiles and amphibians thanks to Jacques Castanet and others in de Ricqlèes's laboratory in Paris, which has trained many paleohistologists.

Interpreting fossils can be tricky. For one thing, an animal's body continually dissolves primary bone—to extract calcium or to repair microfractures—and then deposits secondary bone, erasing the bone's early history. To account for missing LAGs, researchers must make assumptions about their spacing and about bone deposition rates—no simple task, because in living animals, deposition rates vary widely from species to species and even between bones in the same individual. Temperature and diet affect bone growth, too.

One solution is to look at many specimens of various ages, so that juvenile bone fills in the missing picture for adults. “As long as I have enough individuals and a diversity of bones, I can reconstruct what was going on,” says Kristi Curry Rogers of the Science Museum of Minnesota in St. Paul. By counting LAGs, researchers can assemble a series with individuals of various ages. Then, using techniques for estimating an animal's mass from the size of its bones, they plot how various types of dinosaurs typically grew over time. Most growth series are partial, but the hadrosaur Maiasaura is known from embryo to adult.

When growth curves were published in the 1990s, they revealed startling facts about dinosaurs. In a 1999 Journal of Vertebrate Paleontology paper, for example, Curry Rogers showed that the giant sauropod Apatosaurus reached full size—25 meters long—in just 8 to 11 years, not the decades that had long been assumed. “People would have laughed!” says Kevin Padian of the University of California, Berkeley. The quick growth rate complicates a long-standing puzzle: How did sauropods, with their relatively small mouths and simple teeth, manage to get so big, particularly during the Jurassic, when only cycads and other plants of meager nutrition were growing?

Researchers have also established a general pattern for dinosaurs, compared to other groups. In a pair of 2001 Nature papers, two groups—Padian's team and Erickson and colleagues—used different techniques to plot growth curves for several dinosaur species. Both concluded that dinosaurs grew faster than reptiles. The larger dinosaurs packed on weight at a pace comparable with that of mammals, but none grew as blindingly fast as modern birds do. Growth curves also show that, like birds and mammals, dinosaurs grew fast when young, then slowed down and stopped growing as adults. By contrast, nondinosaurian reptiles such as crocodiles grow more slowly.

Growth curves have been used to investigate how dinosaurs evolved various patterns of growth. In August, Erickson and several co-authors reported in Nature how T. rex evolved to its formidable size, relative to other tyrannosaurids (Science, 13 August, p. 930). Rather than extend its growth phase, T. rex accelerated its adolescent growth spurt—packing on up to 2 kilograms a day. Sauropods show similar changes, according to a paper by Martin Sander of the University of Bonn, Germany, and colleagues, in press at Organisms, Diversity & Evolution. “That's not information you can get from gross anatomy,” notes Allison Tumarkin-Deratzian of Vassar College in Poughkeepsie, New York. “The only way you can pin down accelerated rates of growth versus extended period of growth in the fossil record is by looking at histology.”

## Giants and dwarfs

As a rule, dinosaurs and other vertebrates evolved to get bigger through the ages. But the bones show that some bucked the trend. At this week's SVP meeting, Sander and Octavio Mateus of the Museum of Lourinha in Portugal and colleagues announced that small sauropods discovered in Germany are not juveniles but “the first unequivocal case of dwarfing for any dinosaur.” The 10 individuals range in size from 1.8 to 6.2 meters long—much smaller than the 23-meter-long brachiosaurids to which they are closely related—but the tightly spaced growth lines in their bone tissue clearly show that they were full grown. The growth curve, based on seven leg bones, suggests that the dwarfs may have been sexually mature at as young as 2 to 3 years of age.

The dwarfs lived about 150 million years ago, on an island about half the size of New Zealand. So they could provide new data about the relationship between land area and the maximum size of animals. Curry Rogers and colleagues are working on the histology of other possible “island dwarf” sauropods, titanosaurs from Argentina and Romania.

Birds are another group that reduced their body size relative to their dinosaurian ancestors, and histology is helping researchers figure out how that happened. By comparing their tissue with those of the most birdlike dinosaurs, Padian and others have argued that they shrank by shortening the amount of time they spend growing most rapidly. (Most birds reach full size within a few weeks.) “It's a very smart idea,” says Luis Chiappe of the Natural History Museum of Los Angeles County. So even though they grow at a faster rate than dinosaurs, they end up smaller—which would have been a key step toward evolving the ability to fly.

Following up on research begun by de Ricqlès in the 1970s, Chinsamy-Turan is also looking at the beginnings of another fast-growing group: the mammals. Research on their therapsid ancestors shows that some of these so-called mammallike reptiles were still growing like reptiles, while others show some distinct evidence of more mammalian growth patterns. Now she's looking at Mesozoic mammals from the Gobi desert. “I have begged and really pleaded” to get access to these rare specimens, she says, to compare them to modern mammals.

## Paleo-exotica

Sometimes zeroing in on ancient bones turns up exotic results. At the SVP meeting, Horner, and Mary Schweitzer and Jennifer Wittmeyer of North Carolina State University in Raleigh, described tissue, never reported from a dinosaur before, from the femur of a T. rex. The tissue has a random structure and is much richer in blood vessels than surrounding tissue. The researchers propose that it functioned like tissue that female birds use to store calcium for making eggshell. If so, it would be the first time paleontologists have determined gender—and reproductive status—from a dinosaur bone. Some skeptics, however, think the tissue structure might be the result of injury or disease.

Other novel tissues have been reported from flying reptiles. Pterosaur bones have plywoodlike tissue made of layers stacked so that bone fibers run at right angles in alternate layers. Such crisscrossing structures are common in fish scales, but Horner, Padian, and de Ricqlès were the first to describe them in a four-limbed vertebrate. In 2000 in the Zoological Journal of the Linnean Society, they speculated that the tissue is an adaptation for the biomechanical demands of flight.

Another unusual tissue has been found in squat, armored dinosaurs called ankylosaurs. Examining the bony plates called scutes, Sander and Torsten Scheyer, also of the University of Bonn, found bundles of structural fibers arranged parallel, perpendicularly, and obliquely to the scute surface—a light, strong design that would have resisted impacts from all directions, they speculate in next month's issue of the Journal of Vertebrate Paleontology. “It's a highly developed, composite material, like a bulletproof vest, that would prevent penetration of sharp objects,” Sander says.

Histology can also be used to test hypotheses about the function of bizarre structures that no longer exist in the world. Stegosaurus plates have long attracted attention, and a prevalent idea is that they were used to regulate body temperature. Horner, Padian, de Ricqlès, and Russell Main, now a graduate student at Harvard University, decided to test that idea. After making thin sections of stegosaur plates and the smaller scutes of related dinosaurs, the team discovered that Stegosaur plates had evolved simply by expanding the keel of scutes. “We saw nothing special about the stegosaur plates” that would be an adaptation for thermoregulation, Main says. Moreover, structures originally described as blood vessels probably weren't. The plates were probably used instead for species recognition, the group proposes in a paper in press at Paleobiology.

Indirectly, bone histology can even shed light on long-vanished animals' behavior. Curious about whether baby hadrosaurs would have stayed in the nest or struck out on their own after hatching, Horner's team looked at the bone tissue of Maiasaura embryos, as well as embryos of alligators and ratite birds. “We didn't have much evidence until we looked at the histology of the bones,” Horner says. Unlike the ossified bones of alligators and ostriches, the tissue at the end of the hadrosaur limb bones consisted of calcified cartilage, suggesting that hatchlings couldn't walk immediately. They reported these findings in the Journal of Vertebrate Paleontology in 2000.

Walking, running, jumping, flying: Bones actively respond to the stresses of these and other physical activities. On the one hand, this can complicate the interpretation of bone tissue when researchers are trying to establish growth rates. But because physical activity affects bone, it may also be possible to extract that history from bone tissue, for example by studying the orientation of the strutlike trabecular tissue inside bones, which is often oriented perpendicularly to the major axis of strain. “It's tricky and requires a certain amount of interpretation,” cautions John Hutchinson of the Royal Veterinary College in London. “There's still a lot of work that needs to be done in modern animals to see how strain impacts bone remodeling.”

A good amount of that work is going on. For example, Main is studying goats to determine how biomechanics affects their bone histology. He hopes to find signals that could enable fossils to reveal posture, among other details. Other researchers are seeking similar clues in alligators, crocodiles, and birds. “Modern animals are some of the great unsung heroes of dinosaur paleontology,” says Curry Rogers.

Better known heroes are playing a key role too, especially when they are abundant. Horner, for example, continues to mine a rich deposit of hadrosaurs, with individuals of all ages and sizes. “We're cutting hundreds and hundreds of slides,” says Horner, who has a technician working on histology full-time. Once his group and others nail down what's normal for bone tissues, they may be able to probe the many influences that affect bone, extracting information about sexual dimorphism, climate, gait, and much else. “We've just begun to scratch the surface,” says Chinsamy-Turan.

13. CANCER RESEARCH

# Inflammation and Cancer: The Link Grows Stronger

1. Jean Marx

Research into a long-suspected association between chronic inflammation and cancer reveals how the immune system may be abetting tumors

Hepatitis B virus infects hundreds of millions of people worldwide, causing jaundice, fatigue, liver damage, and joint pain. More ominously, investigators have indicted it in another role: as co-conspirator in a far- ranging case they've been building for years linking chronic inflammation and cancer. Researchers have long known that patients with persistent hepatitis B infections experience inflammation and scarring of liver tissue and an increased risk of liver cancer. Other sources of chronic inflammation, including the ulcer-causing bacterium Helicobacter pylori and an immune disorder known as ulcerative colitis, predispose patients to cancers of the stomach and colon.

Based on their experience with these diseases, researchers estimate that inflammation contributes to the development of at least 15% of all cancers. Much less clear, however, is exactly how it does its dirty work. The inflammation-cancer connection is especially puzzling in light of other work suggesting that in some circumstances the immune system, which sustains inflammation, has the opposite effect: inhibiting tumor development. A flurry of results published over the past few months may now be resolving the mystery.

The new work has implicated an inflammation-induced protein called NF-κB as a key player. It is an intermediary in promoting the cellular changes leading to the uncontrolled growth of cancer cells and also to later changes that help metastatic cells escape from the original tumor and spread to new sites in the body.

Other studies have pointed to a source of trouble in the inflammatory cells that creep into a growing tumor, suggesting that they produce numerous substances that can contribute to tumor growth and survival, including some that might trigger increased NF-κB activity. Research into the inflammation- cancer link “is a very exciting area that is developing very rapidly,” says Raymond DuBois of Vanderbilt University Medical Center in Nashville, Tennessee.

The excitement stems in part from the expectation that this emerging understanding could lead to improved cancer prevention and treatment. Epidemiological studies have already shown that people who regularly take NSAIDs—nonsteroidal anti-inflammatory drugs—have a lower risk of developing cancer than people who don't take the drugs. But the efficacy of NSAIDs is not ideal, and the first generation of these drugs, such as aspirin, can cause life-threatening stomach ulcers. Now, even the newer NSAIDs—the so-called COX-2 inhibitors, which were designed to avoid that side effect—may have problems: On 30 September, the pharmaceutical company Merck removed its blockbuster COX-2 inhibitor Vioxx from the market because it increased patients' risks of having heart attacks and strokes.

Researchers hope that if they learn how chronic inflammation leads to cancer, they will be able to design new drugs that counter its effects. The information may also aid the development of vaccines and other strategies to enhance immune attacks on tumors. But researchers will have to toe a fine line: The new work suggests that some approaches might enhance the growth of tumors rather than kill them. “It's a complicated field, but it's extremely important,” says Albert Baldwin of the University of North Carolina School of Medicine in Chapel Hill (UNC).

## Jekyll-and-Hyde macrophages

One line of evidence linking inflammation to cancer comes from the somewhat surprising finding that immune cells can foster tumor development. Lisa Coussens, Douglas Hanahan, and their colleagues at the University of California School of Medicine in San Francisco have found that this can happen in a model of skin cancer they devised in mice that ordinarily develop invasive skin carcinomas by age 1.

The tumors also show infiltration by immune cells, an inflammatory response the researchers initially viewed in a positive light as the body's attempt to destroy the tumor cells. But when they turned down the immune response by crossing their mice with another strain lacking a particular type of T immune cell, the resulting animals showed reduced tumor formation rather than the expected increase. “If we abate inflammation, we abate cancer development,” Coussens says.

Similarly, Jeffrey Pollard and his colleagues at Albert Einstein College of Medicine in New York City have evidence that infiltration with inflammatory cells called macrophages promotes both the development of breast cancers and their eventual spread to other sites in the body. The team showed this by knocking out the gene for colony-stimulating factor 1 (CSF-1), a so-called cytokine that attracts macrophages, in a mouse strain genetically engineered to develop mammary cancers.

Compared to controls, Pollard says, “the incidence and rate of early [tumor] growth in the resulting animals was not changed, … but progression to malignancy was slowed, and there was almost no metastasis.” Consistent with this, Pollard's team and others have also shown that CSF-1 levels are elevated in human breast, ovarian, and uterine cancers and that this elevation correlates with a poor prognosis.

Why the immune system can promote cancer development at times and in other cases help keep it in check remains unclear. But activating macrophages appears to be especially dangerous; many of the weapons these immune system scavengers release can promote cancer. The noxious substances include highly reactive forms of oxygen that can cause carcinogenic mutations. Cells are especially vulnerable to these assaults when they are dividing rapidly, as may be the case when tissues try to repair the damage caused by viral or bacterial infections. Macrophages also produce growth factors, enzymes that can help cancer cells escape from tumors and migrate through the body, and still other proteins that stimulate the formation of blood vessels needed for tumor growth.

## Inflammation hallmark implicated

Although the overall picture is far from complete, recent evidence has now firmly tied one protein, NF-κB, into the cancer-promoting action of inflammatory cells. Suspicion fell on the protein several years ago. It is highly active in both inflammatory cells, such as macrophages, and in other cells of inflamed tissues. Indeed, NF-κB activity is “almost a hallmark of inflammation,” says a researcher involved in this work, Yinon Ben-Neriah of Hebrew University-Hadassah Medical School in Jerusalem, Israel.

Researchers have also found that the protein is abnormally active in some cancers and that that portends a bad prognosis. This may be due to the fact that NF-κB activity leads to inhibition of the programmed cell death (apoptosis) that can eliminate defective cells, thus contributing both to cancer development and resistance to drug and radiation therapies.

But the case against NF-κB was largely circumstantial—until this summer, when Michael Karin and his colleagues at the University of California, San Diego, helped uncover a smoking gun. They found that the protein contributes to cancer development in two distinct ways. (The results appeared in the 6 August issue of Cell.)

For these experiments, the researchers turned to a mouse model of colitis-associated cancer developed about 8 years ago by a team led by Isao Okayasu of Kitasato University School of Medicine in Japan. This involves giving young mice a single injection of a cancer-triggering chemical and then dosing them repeatedly with a salt that irritates the intestinal lining, causing chronic inflammation of the colon. This treatment produces numerous colon tumors.

To test the involvement of NF-κB, the Karin team used two different strains of genetically altered mice; in one, the gene for an essential NF-κB activator enzyme called IKKβ had been knocked out only in the cells that give rise to macrophages, and in the second strain the same gene was knocked out in the epithelial cells lining their intestines. In the absence of the IKKβ gene, neither cell type could activate its NF-κB.

Loss of NF-κB function in macrophages reduced tumor incidence in the animals by about 50%, and the tumors that did form were smaller than those in controls. Further work traced this reduced tumor growth to loss of growth factors produced by inflammatory cells in response to NF-κB activation.

The picture was different in mice whose intestinal epithelial cells couldn't activate NF-κB. These animals “had an 80% reduction in tumor incidence,” Karin says. That difference was not due to reduced intestinal inflammation because it was more severe in the knockouts than in the controls. Instead, with NF-κB eliminated, apoptosis was no longer inhibited in intestinal cells, which presumably helped the knockout animals eliminate cells with cancer-promoting mutations. “Apoptosis is sort of like a safety mechanism,” Karin explains. “It makes sure tumor formation is low.”

Something similar appears to be going on in inflammation-associated liver cancer, as described by Ben-Neriah and his colleagues in Nature (published online on 25 August). These researchers started with a genetically altered strain of mice that develop severe liver inflammation—and cancer. Then they made a second genetic alteration, adding a gene for a natural NF-κB inhibitor called IκB that carried a regulatory sequence allowing the gene to be turned off at will by giving the animals an antibiotic.

When the IκB gene was on—and NF-κB was inactive—the livers of the mice showed early precancerous changes such as increased cell division and the formation of small growths called adenomas for 7 months. But compared to controls, very few of those adenomas—about 10%—progressed to full-fledged cancers.

When the researchers then turned the IκB gene off, Ben-Neriah says, “we started to see tumors as if there had been no [earlier] NF-κB switch-off.” This suggests that the protein is not required for the early changes that put cells on the path to cancerous transformation but is required for later progression to malignancy. Again, it apparently promotes liver cancer development by inhibiting apoptosis.

What's more, NF-κB may be needed for one of the most dangerous features of cancer cells: their ability to spread throughout the body and seed new tumors. Some of this work, described in the August Journal of Clinical Investigation, comes from a team led by Thomas Wirth of the University of Ulm, Germany. Working with cultured mammary epithelial cells that had been transformed with the cancer-causing Ras oncogene, the researchers showed that the NF-κB protein is needed for something called the epithelial-mesenchymal transition, in which normally sedentary epithelial cells undergo changes that allow them to migrate.

Consistent with those findings, the German team also showed that cells in which NF-κB had been inhibited by addition of an active IκB gene form far fewer lung metastases when injected into mice. “NF-κB may be involved both early and late” in cancer development, Wirth says. If so, it would be a good target for potential therapeutic drugs.

Karin's team provides further confirmation that inflammation-induced NF-κB activity is needed for growth of tumor metastases. To mimic metastatic growth, they injected colon cancer cells into mice and observed them forming metastases in the animals' lungs. When the researchers also injected a bacterial lipopolysaccharide (LPS) to stimulate an inflammatory response in the mice, the metastases “pretty much doubled in size,” Karin says. That response also required a protein called tumor necrosis factor-α (TNF-α that's made by macrophages. But when the researchers performed the same experiment with tumor cells bearing an NF-κB inhibitor, the tumors actually shrank following LPS injection due to increased apoptosis.

Although much evidence supports the idea that NF-κB promotes cancer by inhibiting apoptosis, it may contribute in numerous other ways as well. Some may be direct outgrowths of the protein's role in inflammation. The genes activated by NF-κB include the one that makes COX-2, an enzyme needed for the synthesis of a pro-inflammatory compound called PGE-2. This can bring in more immune cells to maintain the inflammation and further prod tumor growth. COX-2 also promotes blood vessel growth. Actions such as these may explain why COX-2 inhibitors have cancer-fighting effects.

Some NF-κB actions may be independent of inflammation, however. Although NF-κB activity is not necessary for cancerous transformation by Ras, Baldwin says, it does foster cancer growth, similar to what Karin's team found in their experiments. But when Ras activates NF-κB, the consequences may be different than when TNF-α or other inflammatory factors do. In work published in the 15 October issue of Cancer Cell, Baldwin and his colleagues analyzed the genes turned on by NF-κB following its activation by Ras. The genes affected included several that make growth-promoting proteins, but for reasons not yet understood, none of the inflammation-promoting genes typically activated in response to TNF-α. Although this fosters cancer growth, NF-κB activity in response to Ras apparently activates a different set of genes than when TNF-α is the activator.

However NF-κB works, it's looking more and more like a good target for anticancer drugs. “There are definitely ways to take advantage of this,” Karin predicts. The pharmaceutical industry is currently working to develop NF-κB inhibitors, and even some low-tech compounds such as the active ingredients in green tea and red wine, which are thought to have anticancer properties, are turning out to act on the protein. “Almost every cancer preventive is an NF-κB inhibitor,” Baldwin says.

14. INVASIVE SPECIES

# Expanding Trade With China Creates Ecological Backlash

1. Dennis Normile

Scientists in the United States and China are scrambling to cope with an unintended consequence of increasing economic ties—a two-way flow of unwelcome plants and animals

To the unknowing eye, the reeds growing along the Yangtze River near Shanghai—a burst of green in the summer, with tips that turn golden brown in the fall—belong on picture postcards. But ecologists know them as a biological experiment run amok. The salt marsh grass Spartina alterniflora, a native of eastern North America introduced in 1979 to check erosion, has now spread across southeastern China, choking estuaries, crowding out native grasses, and reducing feed and habitat for fish and migratory birds.

Across the Pacific, the tiny holes in the bark of maples, willows, and elms in New York, New Jersey, and Illinois come from the Asian longhorned beetle. Anoplophora glabripennis is an unwelcome hitchhiker from China that most likely arrived in the United States a decade ago aboard wooden shipping crates. Unless checked, the beetle threatens to bore its way through billions of dollars' worth of valuable timber, shade, and maple syrup trees.

Different continents, different species, different routes of introduction—but a common problem. Since China opened its doors to the West in the early 1980s, its burgeoning trade with the United States (see graph) has meant more marine organisms in ballast water and insects in packing crates accidentally transported across the Pacific in both directions. The influx has been supplemented by intentional introductions by commercial U.S. nurseries and horticultural collectors looking for exotic Chinese specimens. Many of these species flourish thanks to similar habitats and climates in the two countries. The result, says Li Bo, a plant ecologist at Shanghai's Fudan University, is that “American species can easily get established in China, and Chinese species can easily get established in America.”

Many introduced species never expand beyond a beachhead, and most horticultural species are well behaved. But for reasons that are still being debated, some introductions lead to ecological disaster. This summer Chinese and U.S. scientists held two meetings* in hopes of sharing information and developing strategies that might help avert dire consequences from this two-way traffic. “There is a chance we could stop this wave of invasive species exchange between China and the United States” through stricter inspections and stiffer regulations, believes Peter Alpert, a plant ecologist at the University of Massachusetts, Amherst. But he says “there are as yet no effective policies to control species exchanges.”

## Proceed with caution

Controlling invasives is already a major headache for both countries. David Pimentel, an entomologist and systems ecologist at Cornell University in Ithaca, New York, concluded in 2000 that invasive species cost the United States more than $137 billion per year. A similar study presented this summer at the Beijing symposium by Xu Haigen of the Nanjing Institute of Environmental Sciences, working with colleagues there and at the Nanjing Forestry University, concluded that invasives had caused$2.4 billion in damages to eight major Chinese industries alone. Many researchers on both sides say these numbers are probably low and certain to rise.

The first bilateral efforts to control invasive species in Asia and North America occurred in the 1980s, when U.S. scientists visited China in search of natural predators for species imported as horticultural ornamentals in the late 19th and early 20th centuries. Many of these, such as Chinese privet, are now widespread in North America. Those initial surveys led to the creation of a Sino-American Biological Control Laboratory in Beijing in 1989 to identify and exchange insects that eat invasive plant species.

One apparent success story is a leaf beetle (Diorhabda elongata) native to Xianjiang Province. It attacks tamarisks (Tamarix ramosissima), an invasive Eurasian species that has displaced native plants along rivers and in arid regions of the American west. The beetle was released 7 years ago, and it “looks like it will be a terrific success” in reducing the spread of tamarisk, says Robert Pemberton, an entomologist at the U.S. Department of Agriculture's (USDA's) Agricultural Research Service in Fort Lauderdale, Florida. So too does the 1980 introduction into China of two beetles, originally from South America, to control the spread of water hyacinth.

Introducing insects to eat invasive plants is a laborious and risky process. The insects must be carefully screened to ensure that they don't attack native plant species, as happened with the weevil Rhinocyllus conicus, brought into Canada and the United States from Europe in the late 1960s to battle exotic thistles. It turned out to have a taste for native thistles as well. Ding Jianqing, an entomology postdoc at Cornell University who previously headed a biocontrol lab at the Chinese Academy of Agricultural Sciences, says that concerns about unintended consequences have limited the number of introductions in the past 15 years to just four predatory insects from the United States to China, and only three in the opposite direction.

In situations in which eradication is difficult or impossible, scientists would settle for preventing further spread of the invasive plant. But they are hoping that early intervention may actually stem the infestation of the Asian longhorned beetle. USDA researchers have been cooperating with their Chinese counterparts to develop better methods to detect tree infestation early as well as to identify possible lures to attract and kill the beetles. But this is still a work in progress.

Similarly, Duane Chapman, a fisheries biologist at the U.S. Geological Survey's Columbia Environmental Research Center in Missouri, is struggling to reduce damage from bighead and silver carp imported from Taiwan in the 1970s for aquaculture farms in Arkansas. The carp escaped and are now crowding out native species. Chapman and Chinese counterparts are summarizing and translating Chinese studies of the carp in their native habitat, information which may lead to more effective control strategies.

## A closer look

Despite good working relationships between individual scientists, U.S. and Chinese researchers say that they won't make a serious dent in controlling invasive species until their governments make it a higher priority. The United States spent \$1 billion this year on efforts such as inspections of cargo and baggage for accidental introductions and vetting deliberately imported species. A 2002 National Academy of Sciences (NAS) study says that USDA's Animal and Plant Health Inspection Service (APHIS) intercepts more than 53,000 arthropods, pathogens, and noxious plants each year. But that's only a tiny fraction of what needs to be done, say scientists. Only 2% of cargo shipments are checked, they note. And although packing crates infested with the Asian longhorned beetle have been stopped at ports in 17 states, according to USDA, enough slipped through to start an invasion.

There's also growing concern about deliberately introduced horticultural specimens. “We're going to have to take a much more aggressive position on evaluating whether a species can become a threat to any aspect of the U.S. biota prior to its release into the marketplace,” says Richard Mack, an ecologist at Washington State University in Pullman. Mack chaired the panel that produced the 2002 NAS report Predicting Invasions of Nonindigenous Plants and Plant Pests. Its recommendations are being discussed by scientists and federal officials, he says, “but I don't see any strong movement yet” to implement them.

Fudan's Li says China has only recently begun to conduct inspections and implement quarantine requirements. Last year the government published a “black list” of species not to be brought into the country, but it simply covered known problems, such as Spartina alterniflora. This month, however, the Chinese Academy of Agricultural Sciences, with the support of the ministries of Science and Technology and of Agriculture, is holding a workshop to develop a national invasive species strategy that would cover prevention, early detection, and on-the-ground management. That strategy should give scientists on both sides of the Pacific more ammunition to battle an increasing onslaught of invasive species.

• *Beijing International Symposium on Biological Invasions, 8–15 June 2004. Biological Invasions: Species Exchanges between Eastern Asia and North America, Portland, Oregon, 2 August 2004, held in conjunction with the Ecological Society of America meeting.