Biologists from Austria and Singapore developed a technique that adds a new twist on the relationship between biology and art. In an article recently published online in The FASEB Journal (http://www.fasebj.org) and scheduled for the August 2008 print issue, these researchers describe how they were able to coat—or paint—viruses with proteins. This breakthrough should give a much-needed boost to the efficiency of some forms of gene therapy, help track and treat viral disease and evolution, improve the efficiency of vaccines, and ultimately allow health care professionals track the movement of viral infections within the body. Specifically, the new method should make it easier to track and treat infectious diseases such as HIV/AIDS, influenza, hepatitis C, and dengue fever. And because viruses can also be used to introduce biotechnology drugs and replacement genes, and act as vaccines, this research should lead to new treatments for cancer, cardiovascular, metabolic and inherited disorders.

“This technology should provide a new tool for the treatment of many diseases,” said Brian Salmons, one of scientists who co-authored the study. “Even if you are working with a virus that is unknown or poorly characterized, it is still possible to modify or paint it. This is very interesting for emerging diseases.”

In the article, Salmons and colleagues explain how they mixed purified proteins (glycosylphophatidylinositol anchor proteins) with lipid membranes to make it possible to bind these proteins to the outer “skin” (the lipid envelope) of viruses. Even with the new paint job, the viruses remained infectious. While the experiment only involved one type of protein and two types of viral vectors, Salmons says the technique could be expanded and used to apply “paint” made up of other proteins, dyes, and a variety of unique markers.

“Biology and art converge daily: people paint their nails, color their hair, and tattoo their skin,” said Gerald Weissmann, M.D., Editor-in-Chief of The FASEB Journal. “Now this convergence has entered a new dimension as painted viruses permit scientists to track, cure and prevent disease.”

University of Illinois researchers have developed a technique for imaging cells under an electron microscope that yields a sharper image of the structure of chromatin, the tightly wound bundle of genetic material and proteins that makes up the chromosomes. The findings appeared in Nature Methods.

Scientists have known for more than a century that proteins, such as histones, aid in packing DNA into the nucleus of a cell.  Human cells contain 2 to 3 meters of DNA, which must be kinked and coiled enough to fit into a region 1/10 the width of a human hair.

Despite the use of powerful, high-resolution imaging techniques such as electron microscopy, the mechanism by which this chromatin packing occurs remains a mystery.  The densely coiled chromatin fibers are very difficult to visualize, and little is known about how they condense during cell division, or unwind to allow gene expression.

In developing their method, the Illinois team tackled a key difficulty in imaging cells using electron microscopy.  Traditional studies “fix” the cells with potent chemicals (called fixatives) to preserve their structure for viewing under a microscope.  But standard fixation methods interfere with another step in the imaging process: the use of tagged antibodies to label key components of the cells.

These antibodies, which target and latch on to specific proteins in the cell, can be tagged with fluorescent labels for detection in light microscopy, or with metal particles (gold, in this case) for electron microscopy.

“If you fix the cells first, you have a dramatic drop in the efficiency of these immunochemical reactions,” said Igor Kireev, a visiting scientist in the department of cell and developmental biology and lead author of the paper.  Electron microscopy image Click photo to enlarge Image courtesy of Andrew Belmont and Igor Kireev The new technique exposes living cells to labeled antibodies, an approach that yields a much stronger signal for electron microscopy.

“And if your target is inside the condensed chromatin, the antibodies have no way to penetrate.”

Instead of fixing the cells before staining with antibodies, the researchers first exposed living animal cells to the labeled antibodies.  This allowed the antibodies to penetrate more deeply into the chromatin structure, and boosted the number of gold particles adhering to regions of interest.  The signal was enhanced by adding a silver solution that precipitated (solidified) upon contact with the gold.

“We are interested in chromatin structure, so our targets are mostly chromatin-bound proteins,” Kireev said.

The researchers had inserted several copies of a bacterial DNA, called the Lac operator, into the chromosomes.  A bacterial protein, the Lac repressor, recognizes and binds to the Lac operator in living cells.

The researchers combined a Lac repressor protein with another protein that fluoresces green under blue light.  This engineered protein adhered to the chromosomes in regions containing the Lac operator sequences.  Under blue light, these regions fluoresced.  A gold-tagged antibody targeted against green fluorescent protein (GFP) was then microinjected into the nucleus of a living cell, which added a metallic signal that could be boosted with silver.

“All this combined gives us a much better signal, a much stronger signal, with the very best structural preservation,” Kireev said.

The fluorescing protein helped the researchers find the regions of interest in the cells.  These areas were then “immunogold” labeled and targeted for electron microscopy.

In the resulting micrographs the researchers saw enhanced staining of the chromosomes.

“We can now apply this same live-cell labeling method to study at high resolution many different GFP-tagged proteins in the cell cytoplasm or nucleus,” said Andrew Belmont, a professor of cell and developmental biology and senior author of the paper.

“In trying to understand chromosomes, people have largely been limited to low resolution visualization of specific chromosomal proteins using light microscopy,” Belmost said.  “This meant everyone has had to do a lot of guessing of how things are put together, leading in many cases to vague, cartoon models of what are likely to be complicated chromosomal structures carrying out DNA functions such as replication and transcription.”

“Now we hope we can simply look and see the real structure using the more than 10-fold higher resolution of electron microscopy,” Belmont said.  “We are really excited to see what we will find using our new method”

New cellular therapies benefits came to light as a result of powerful PET and SPECT imaging in a recent study reported in the April issue of the Journal of Nuclear Medicine. Researchers in Germany were able to observe the repair action of circulating progenitor cells (CPCs), immature blood-derived cells capable of developing into adult stem cells, as they successfully preserved healthy heart tissue and corrected blood flow imbalance within the heart.

Twenty-six patients took part in the randomized, placebo-controlled and double-blinded study. Following the recanalization of blocked coronary arteries (the surgical reopening or formation of new paths for blood flow), one group received an infusion of progenitor cells. FDG PET and 99mTc-tetrofosmine-SPECT were then used to image relative changes in myocardial perfusion (blood flow through the middle and thickest part of the heart) and glucose metabolism.

The results were compared with a control group that had undergone recanalization but did not receive CPCs. In the CPC group, normalization of glucose metabolism and coronary blood flow was seen in nearly 50 percent of the repaired artery segments.

“PET and SPECT are the only techniques capable of validating the metabolic changes we needed to observe in the heart once we had administered the progenitor cells,” said Kai Kendziorra, M.D., a specialist in Nuclear Medicine at the University of Leipzig in Leipzig, Germany. “The results shown by these imaging modalities provide the evidence needed to expand the use of CPC treatment.”

Earlier research has shown that when a patient’s progenitor cells are activated by growth factors, the result is increased cell division, which is vital to the tissue repair process. In this study, progenitor cells developed from circulating blood were also found to be capable of repairing dysfunctional—yet viable—myocardial tissue, a condition referred to as “hibernating myocardium.”

Kendziorra said he believes that in addition to assisting in monitoring and guiding treatment of heart patients, PET scans may also be helpful in selecting those who would profit the most from CPC administration.

“Early detection of hibernating myocardial tissue via noninvasive imaging modalities such as PET and SPECT will help us to assess a patient’s myocardial metabolism and blood flow,” he said. “Subsequent early coronary recanalization and CPC administration may lead to treatment-specific normalization and reduce the risk of cardiac events over longer periods.”

“For decades, nuclear medicine imaging has contributed functional assessment to the anatomical definition of the presence or absence of disease,” said Alexander J. McEwan, M.D., president of SNM. “Today molecular imaging is on the way to revolutionizing patient care—by integrating information about location, structure, function and biology—leading to a package of non-invasive imaging tools with enormous potential for improving patient care and outcomes.”

Co-authors of “Effect of Progenitor Cells on Myocardial Perfusion and Metabolism in Patients After Recanalizatoin of a Chronically Occluded Coronary Artery” include Henryk Barthel, Osama Sabri and Regine Kluge, Department of Nuclear Medicine; Sandra Erbs and Gerhard Schuler, Heart Center Leipzig GmbH; and Frank Emmrich, Institute of Clinical Immunology and Transfusion Medicine, all with the University of Leipzig, Leipzig, Germany; and Rainer Hambrecht, Department of Nuclear Medicine, University of Leipzig, Leipzig, Germany and Heart Center Bremen, Bremen, Germany.