Medical Info

A scientist from the Florida campus of The Scripps Research Institute has devised a new method of analyzing and quantifying changes in proteins that result from a common chemical process. The new findings could provide new insights into the effects of a highly destructive form of stress on proteins in various disease models, particularly cancer.

The study, published January 5, 2011, in the online Early View of the journal Angewandte Chemie, was designated by the journal as a "very important paper," a distinction bestowed on less than five percent of its publications.
"This new technique allows us to home in on which proteins are modified to a significant extent during periods of stress and how that changes during disease progression," said Kate Carroll, an associate professor in the Scripps Research Department of Chemistry who conducted the study with Young Ho Seo, a research fellow at The University of Michigan. "It gives us the chance to look more closely at targets for possible therapeutic intervention. From a practical standpoint, the technique is simple and will be accessible to biologists and chemists alike."
The new technique focuses on the process of cysteine S-hydroxylation, which plays a significant role in a number of events related to physiology in both health and disease, including the regulation of signaling proteins in various disease states.
The ability of the new technique to focus on signaling pathways, particularly in diseases such as cancer, is critical.
"Chronic disease states such as cancer can involve the modification of signaling proteins through S-hydroxylation, but other housekeeping proteins may also be targets," she said. "Key to distinguishing which of these proteins may be involved in pathogenesis is the ability to measure the amount of S-hydroxylation at specific sites within a protein. Now you'll be able to tell. This should help accelerate target identification in these disease-related signaling pathways and allow us to focus on proteins that are important to the process."
During periods of cellular stress, caused by factors such as exposure to UV radiation or many disease states, the level of highly reactive oxygen-containing molecules can increase, resulting in inappropriate modification of proteins and cell damage.
One oxidant, hydrogen peroxide, functions as a messenger that can activate cell proliferation through oxidation of cysteine residues in signaling proteins, producing sulfenic acid (i.e., S-hydroxylation); cysteine is an amino acid is synthesized in the body.
Extending the Gains of an Earlier Study

In a 2009 study, Carroll found that sulfenic acid served as an early warning biomarker of the reaction between hydrogen peroxide and cysteine. Carroll tagged the miniscule reaction target with a fluorescent dye antibody. With it, Carroll was able to read the levels of sulfenic acid levels in various cell lines, including breast cancer cells.
The new technique takes those findings several steps further by allowing scientists not only to quantify the modifications to various proteins, but also to monitor these changes at the level of individual cysteines within a single protein.
Carroll used a class of reagents called isotope-coded dimedone and iododimedone, which traps and tags sulfenic acids, allowing the cysteine sites and modified proteins to be easily identified. These probes, which are highly selective for sulfenic acid, allow the S-hydroxylation process to be monitored at the exact site of the modification.
The tagged proteins can be then be analyzed by mass spectrometry, a standard technology used to determine the precise make-up of proteins and other molecules.
"This technique should be widely accessible to the scientific community because it's so simple," Carroll said. "It should allow researchers to identify proteins with altered S-hydroxylation profiles whose function may lend insight into events in disease progression and have utility as potential markers for disease detection."

Soon, your family doctor will no longer have to send blood or cancer cell samples to the laboratory. A little chip will give her test results on the spot.



Today, a blood sample whose protein content, genes and so on are to be read needs to be submitted to a series of complex processes, such as centrifugation, heat treatment, mixing with enzymes and concentration of disease markers. This means that samples are sent to central laboratories for analysis, and weeks may pass before the results are returned.
The same thing happens when women are checked for cervical cancer by taking a cell scrape from the cervix. The samples are then sent off and studied under the microscope. Diagnostic error rates can be high when abnormal cell appearance is determined by even experienced eyes.
Automated
The EU's MicroActive project has developed an integrated system based on microtechnology and biotechnology, that will enable a number of conditions to be diagnosed automatically in the doctor's own office.
The new "health chip" looks like a credit card and contains a complete laboratory. The EU project has used cells taken to diagnose cervical cancer as a case study, but in principle the chip can check out a number of different diseases caused by bacteria or viruses, as well as various types of cancer.
SINTEF has coordinated the project, whose other members include universities, hospitals and research institutes from Germany and Ireland. The Norwegian NorChip company had the idea for the chip, and has carried out full-scale tests during the project.
Advanced "credit card"
The chip is engraved with a number of very narrow channels that contain chemicals and enzymes in the correct proportions for each individual analysis. When the patient's sample has been drawn into the channels, these reagants are mixed.
"The health chip can analyse your blood or cells for eight different diseases," say Liv Furuberg and Michal Mielnik of SINTEF. "What these diseases have in common is that they are identified by means of special biomarkers that are found in the blood sample. These "labels" may be proteins that either ought or ought not to be there, DNA fragments or enzymes.
"This little chip is capable of carrying out the same processes as a large laboratory, and not only does it perform them faster, but the results are also far more accurate. The doctor simply inserts the card into a little machine, adds a few drop of the sample taken from the patient via a tube in the cardholder, and out come the results."
Scientists at SINTEF's MiNaLaB have developed a number of techniques for interpreting the results when the biomarkers have been found. For example, they can read them off in a spectrophotometer, an optical instrument in which the RNA molecules in different markers emit specific fluorescent signals.
"SINTEF's lab-on-a-chip projects have shown that it is possible to perform rapid, straightforward diagnostic analyses with the aid of microchips, and we are now working on several different types of chip, including a protein analysis chip for acute inflammations," says Liv Furuberg.
Mass production
NorChip has just started a new two-year EU project that aims to industrialise the diagnostic chip to the mass-production stage while the company will also evaluate market potential and industrial partners.
Chief scientist Frank Karlsen in NorChip says that the ways in which the chip can be used can be extended to enable patients themselves to take samples at home, and he expects that such special sampling systems will be ready for testing within a few years.

Researchers from the Royal College of Surgeons in Ireland (RCSI) have identified a biomarker in patient blood which can predict the severity of breast cancer in patients and may enable clinicians to track patients throughout the course of their treatment.

  Around 75 percent of breast cancers are fuelled by the hormone estrogen which encourages them to grow. Although this type of breast cancer can be treated with drugs such as tamoxifen, many people develop resistance and see their breast cancer return.
The study published in Cancer Research reports on the identification of biomarkers which may help distinguish breast cancer patients with a greater risk of disease recurrence. This may lead to the development of personalised treatment programmes for individual patients.
The research, funded by Science Foundation Ireland (SFI) and Breast Cancer Ireland (BCI) investigated the protein changes in breast cancer that is resistant to treatment and found a higher level of a protein called HOXC11 and the presence of a secreted molecule S100-beta that is detected via a simple blood test.
Lead researcher, Dr Leonie Young, Leader of Surgical Research in RCSI said "We hope this research will provide vital information about drug resistance to ensure that people with breast cancer are getting treatments that will benefit them."
Further work on this project is being funded by the UK based charity Breast Cancer Campaign and will provide vital information so that a new blood test can be designed to predict which breast cancers will become drug resistant to current therapies ensuring that only those who will benefit from a particular treatment actually receive it.
Breast cancer is the second most common cancer in Ireland after non-melanomatous skin cancer. Breast cancer accounts for 28% of all cancers in women in Ireland, with an average of 1726 new diagnosis each year. Though management has improved over the years, breast cancer continues to be responsible for an average of 644 Irish female deaths each year. It is the leading cause of cancer related deaths in women in Ireland, with mortality 33% higher in Ireland than in the USA.

A substance found in breast milk can kill cancer cells, reveal studies carried out by researchers at Lund University and the University of Gothenburg, Sweden.



Although the special substance, known as HAMLET (Human Alpha-lactalbumin Made LEthal to Tumour cells), was discovered in breast milk several years ago, it is only now that it has been possible to test it on humans. Patients with cancer of the bladder who were treated with the substance excreted dead cancer cells in their urine after each treatment, which has given rise to hopes that it can be developed into medication for cancer care in the future.

Discovered by chance

HAMLET was discovered by chance when researchers were studying the antibacterial properties of breast milk. Further studies showed that HAMLET comprises a protein and a fatty acid that are both found naturally in breast milk. So far, however, it has not been proven that the HAMLET complex is spontaneously formed in the milk. It is speculated, however, that HAMLET can form in the acidic environment of the babies´ stomachs. Laboratory experiments have shown that HAMLET kills 40 different types of cancer, and the researchers are now going on to study its effect on skin cancer, tumours in the mucous membranes and brain tumours. Importantly, HAMLET kills only cancer cells and does not affect healthy cells.

Studying the integration of the substance

Researchers at the University of Gothenburg are focusing on how HAMLET can be taken up into tumour cells. The researchers, Roger Karlsson, Maja Puchades and Ingela Lanekoff, are attempting to gain an in-depth understanding of how the substance interacts with cell membranes, and their findings were recently published in the journal PLoS One.

Like a fence or barricade intended to stop unwanted intruders, the skin serves as a barrier protecting the body from the hundreds of allergens, irritants, pollutants and microbes people come in contact with every day. In patients with eczema, or atopic dermatitis, the most common inflammatory human skin disease, the skin barrier is leaky, allowing intruders -- pollen, mold, pet dander, dust mites and others -- to be sensed by the skin and subsequently wreak havoc on the immune system.



While the upper-most layer of the skin -- the stratum corneum -- has been pinned as the culprit in previous research, a new study published in the Journal of Allergy and Clinical Immunology found that a second skin barrier structure, consisting of cell-to-cell connections known as tight junctions, is also faulty in eczema patients and likely plays a role in the development of the disease. Tightening both leaky barriers may be an effective treatment strategy for eczema patients, who often have limited options to temper the disease.
"Over the past five years, disruption of the skin barrier has become a central hypothesis to explain the development of eczema," said Lisa Beck, M.D., lead study author and associate professor in the Department of Dermatology at the University of Rochester Medical Center. "Our findings challenge the belief that the top layer of the skin or stratum corneum is the sole barrier structure: It suggests that both the stratum corneum and tight junctions need to be defective to jumpstart the disease."
Currently, there are no treatments that target skin barrier dysfunction in eczema. To treat eczema, which causes dry, red, itchy skin, physicians typically prescribe anti-inflammatory drugs, like prednisone, and a variety of topical anti-inflammatory creams and ointments. But, modest benefit, negative side effects and cost concerns associated with these therapies leave patients and doctors eagerly awaiting new alternatives.
"We want to figure out what current eczema therapies do to both barrier structures and start thinking about new treatments to close the breaks that let irritants in and water out and subsequently drive the inflammation and dryness that is characteristic of the disease," noted Beck, who treats eczema patients in addition to conducting research on the condition.
To better understand the role of tight junctions in eczema, Beck and her team studied skin samples from eczema patients and healthy individuals. Using resistance and permeability tests, they discovered that tight junctions, which act like a gate controlling the passage of water and particles, were strong and tight in healthy skin samples, yet loose and porous in the skin of eczema patients.
On further investigation, they found that a particular tight junction protein, claudin-1, which determines the strength and permeability of tight junctions in skin, is significantly reduced in the skin of eczema patients, but not in healthy individuals or individuals with psoriasis, another common chronic skin disease. They demonstrated that reducing claudin-1 expression in skin cells from healthy donors made the tight junctions leaky and more permeable, a finding in line with results of other research groups.
"Since claudin-1 was only reduced in eczema patients, and not the other controls, it may prove to be a new susceptibility gene in this disease," said Anna De Benedetto, M.D., postdoctoral-fellow at the Medical Center and first author of the new study. "Our hypothesis is that reduced claudin-1 may enhance the reactivity to environmental antigens and lead to greater allergen sensitization and susceptibility in people with eczema."
If the team's hypothesis stands up in future research, increasing claudin-1 to combat eczema could be a new treatment approach worth exploring. The University of Rochester has applied for patent protection for increasing claudin-1 with drug compounds to treat eczema.
Barrier problems, and in particular tight junction defects, are recognized as a common feature in many other inflammatory diseases, such as inflammatory bowel disease and asthma, where the lining of the intestine and the airways is weakened, which is why Beck and her team decided to focus on the role of this barrier structure in eczema.
Eczema affects up to 17 percent of children and about six percent of adults in the United States -- close to 15 million Americans. While there are varying severities of eczema, all have an itch that can make it difficult to focus on daily activities and to sleep. People with eczema are often counseled to minimize their exposure to allergens, but that is a difficult task given the hundreds of allergens people are exposed to each day.
Beck, who began the research while at Johns Hopkins University, plans to build on these findings by investigating the immunologic consequences of tight junction disruption in the skin and whether there is a relationship between barrier disruption and subjects' intractable itch. In addition, as part of a contract with the National Institute of Allergy and Infectious Diseases (NIAID) at the National Institutes of Health, called the Atopic Dermatitis Research Network, Beck, in collaboration with Kathleen Barnes, Ph.D., at Johns Hopkins, will perform gene mapping of claudin-1 to try to identify mutations in patients with eczema.
The current research was funded by the Atopic Dermatitis and Vaccinia Network at NIAID, the National Eczema Association and the Mary Beryl Patch Turnbull Scholar Program. Along with Beck and De Benedetto, Andrei I. Ivanov, Ph.D., Steve N. Georas, M.D., Kunzhong Zhang, Ph.D., Sadasivan Vidyasagar, M.D., Ph.D., and Takeshi Yoshida, Ph.D., from the University of Rochester Medical Center participated in the research. Scientists from Johns Hopkins University School of Medicine, National Jewish Health, the University of California, San Diego, Children's Hospital Boston, Oregon Health & Science University, the University of Bonn (Germany), Technische Universität München (Germany) and Johns Hopkins Bloomberg School of Public Health contributed as well.

Walking may slow cognitive decline in adults with mild cognitive impairment (MCI) and Alzheimer's disease, as well as in healthy adults, according to a study presented November 29 at the annual meeting of the Radiological Society of North America (RSNA).



"We found that walking five miles per week protects the brain structure over 10 years in people with Alzheimer's and MCI, especially in areas of the brain's key memory and learning centers," said Cyrus Raji, Ph.D., from the Department of Radiology at the University of Pittsburgh in Pennsylvania. "We also found that these people had a slower decline in memory loss over five years."
Alzheimer's disease is an irreversible, progressive brain disease that slowly destroys memory and cognitive skills. According to the National Institute on Aging, between 2.4 million and 5.1 million Americans have Alzheimer's disease. Based on current population trends, that number is expected to increase significantly over the next decade.
In cases of MCI, a person has cognitive or memory problems exceeding typical age-related memory loss, but not yet as severe as those found in Alzheimer's disease. About half of the people with MCI progress to Alzheimer's disease.
"Because a cure for Alzheimer's is not yet a reality, we hope to find ways of alleviating disease progression or symptoms in people who are already cognitively impaired," Dr. Raji said.
For the ongoing 20-year study, Dr. Raji and colleagues analyzed the relationship between physical activity and brain structure in 426 people, including 299 healthy adults (mean age 78), and 127 cognitively impaired adults (mean age 81), including 83 adults with MCI and 44 adults with Alzheimer's dementia.
Patients were recruited from the Cardiovascular Health Study. The researchers monitored how far each of the patients walked in a week. After 10 years, all patients underwent 3-D MRI exams to identify changes in brain volume.
"Volume is a vital sign for the brain," Dr. Raji said. "When it decreases, that means brain cells are dying. But when it remains higher, brain health is being maintained."
In addition, patients were given the mini-mental state exam (MMSE) to track cognitive decline over five years. Physical activity levels were correlated with MRI and MMSE results. The analysis adjusted for age, gender, body fat composition, head size, education and other factors.
The findings showed across the board that greater amounts of physical activity were associated with greater brain volume. Cognitively impaired people needed to walk at least 58 city blocks, or approximately five miles, per week to maintain brain volume and slow cognitive decline. The healthy adults needed to walk at least 72 city blocks, or six miles, per week to maintain brain volume and significantly reduce their risk for cognitive decline.
Over five years, MMSE scores decreased by an average of five points in cognitively impaired patients who did not engage in a sufficient level of physical activity, compared with a decrease of only one point in patients who met the physical activity requirement.
"Alzheimer's is a devastating illness, and unfortunately, walking is not a cure," Dr. Raji said. "But walking can improve your brain's resistance to the disease and reduce memory loss over time."
Coauthors are Kirk Erickson, Ph.D., Oscar Lopez, M.D., James Becker, Ph.D., Caterina Rosano, M.D., Anne Newman, M.D., M.P.H., H. Michael Gach, Ph.D., Paul Thompson, Ph.D., April Ho, B.S., and Lewis Kuller, M.D.