Flowing toward potential CV therapeutic

It’s not a blockbuster cardiovascular drug – yet. But the pathway from bench to bedside is easy to see.

In a recent eLife paper, Hanjoong Jo’s lab characterizes a “flow-kine”: a protein produced by endothelial cells in response to healthy blood flow patterns. Unlike other atherosclerosis-linked factors previously identified by Jo’s team, this one – called KLK10 — is secreted. That means that the KLK10 protein could morph into a therapeutic.

We can compare KLK10 to PCSK9 inhibitors, which lower LDL cholesterol and have a proven ability to prevent cardiovascular events. KLK10 acts in a different way, not affecting cholesterol, but instead inhibiting inflammation in endothelial cells. KLK10 can protect against atherosclerosis in animal models, when delivered by injection.

“The most important clinical implication is that we were able to see that human atherosclerotic plaques have a low level of KLK10,” Jo says. “In a healthy heart, the expression level is OK.”

Jo sees similarities between KLK10 and myokines, exercise-induced proteins secreted by skeletal muscle cells. Looking ahead, his lab has begun experiments testing how exercise affects KLK10 and other protective factors.

Jo and his colleagues are in the Wallace H. Coulter Department of Biomedical Engineering at Georgia Tech and Emory. Using a workhorse model of disturbed blood flow in atherosclerosis, his team has steadily identified a stream of genes involved in the disease process. KLK10 is one of several down-regulated by disturbed blood flow.

Jo cites the transcription factor KLF2 as another good example of a protective protein identified by his team’s approach. KLF2 has a similar protective function, but it is expressed inside endothelial cells and stays inside the cell. KLK10 is secreted into the circulation, giving it more obvious therapeutic potential.

Posted on February 14, 2022 by Quinn Eastman in Heart Leave a comment

Dissecting atherosclerosis at the single cell level: tasting each piece of a fruit salad

More than a decade ago, Hanjoong Jo and colleagues developed an elegant animal model allowing the dissection of atherosclerosis. It was the first to definitively show that disturbed patterns of blood flow determine where atherosclerotic plaques will later appear.

In atherosclerosis, arterial walls thicken and harden because of a gradual build-up of lipids, cholesterol and white blood cells, which occurs over the course of years in humans. The Jo lab’s model involves restricting blood flow in the carotid artery of mice, which are fed a high-fat diet and also have mutations in a gene (ApoE) involved in processing fat and cholesterol. The physical intervention causes atherosclerosis to appear within a couple weeks. Inflammation in endothelial cells, which line blood vessels, is visible within 48 hours.

*The shear-sensitive gene LMO4 is turned on in the middle boxed region, but not the other two, because of disturbed flow in that area of the aorta*

Now Jo’s lab has combined the model with recently developed techniques that permit scientists to see molecular changes in single cells. The results were published Tuesday in Cell Reports.

Jo’s lab is in the Wallace H. Coulter Department of Biomedical Engineering at Emory and Georgia Tech.

Previously, when they saw inflammation in blood vessels, researchers could not distinguish between intrinsic changes in endothelial cells and immune or other cells infiltrating into the blood vessel lining.

A video made by Harvard scientists who developed the single cell techniques describes the difference like this. Looking at the molecules in cells with standard techniques is like making a fruit smoothie – everything is blended together. But single cell techniques allow them to taste and evaluate each piece of fruit individually.

Posted on December 15, 2020 by Quinn Eastman in Heart Leave a comment

Long-lasting blood vessel repair in animals via stem cells

Stem cell researchers at Emory University School of Medicine have made an advance toward having a long-lasting “repair caulk” for blood vessels. The research could form the basis of a treatment for peripheral artery disease, derived from a patient’s own cells. Their results were recently published in the journal Circulation.

A team led by Young-sup Yoon, MD, PhD developed a new method for generating endothelial cells, which make up the lining of blood vessels, from human induced pluripotent stem cells.. When endothelial cells are surrounded by a supportive gel and implanted into mice with damaged blood vessels, they become part of the animals’ blood vessels, surviving for more than 10 months.

“We tried several different gels before finding the best one,” Yoon says. “This is the part that is my dream come true: the endothelial cells are really contributing to endogenous vessels. When I’ve shown these results to people in the field, they say ‘Wow.'”

Previous attempts to achieve the same effect elsewhere had implanted cells lasting only a few days to weeks, although those studies mostly used adult stem cells, such as mesenchymal stem cells or endothelial progenitor cells, he says.

“When cells are implanted on their own, many of them die quickly, and the main therapeutic benefits are from growth factors they secrete,” he adds. “When these endothelial cells are delivered in a gel, they are protected. It takes several weeks for most of them to migrate to vessels and incorporate into them.” Read more

Posted on October 23, 2017 by Quinn Eastman in Heart Leave a comment

Blood vessels and cardiac muscle cells off the shelf

Tube-forming ability of purified CD31+ endothelial cells derived from induced pluripotent stem cells after VEGF treatment.

Chunhui Xu’s lab in the Department of Pediatrics recently published a paper in Stem Cell Reports on the differentiation of endothelial cells, which line and maintain blood vessels. Her lab is part of the Emory-Children’s-Georgia Tech Pediatric Research Alliance. The first author was postdoc Rajneesh Jha.

This line of investigation could eventually lead to artificial blood vessels, grown with patients’ own cells or “off the shelf,” or biological/pharmaceutical treatments that promote the regeneration of damaged blood vessels. These treatments could be applied to peripheral artery disease and/or coronary artery disease.

Xu’s paper concerns the protein LGR5, part of the Wnt signaling pathway. The authors report that inhibiting LGR5 steers differentiating pluripotent stem cells toward endothelial cells and away from cardiac muscle cells. The source iPSCs were a widely used IMR90 line.

Young-sup Yoon’s lab at Emory has also been developing methods for the generation of endothelial cells via “direct reprogramming.”

Posted on August 15, 2017 by Quinn Eastman in Heart Leave a comment

Direct reprogramming into endothelial cells

Direct reprogramming has become a trend in the regenerative medicine field. It means taking readily available cells, such as skin cells or blood cells, and converting them into cells that researchers want for therapeutic purposes, skipping the stem cell stage.

In a way, this approach follows in Nobel Prize winner Shinya Yamanaka’s footsteps, but it also tunnels under the mountain he climbed. Direct reprogramming has been achieved for target cell types such as neurons and insulin-producing beta cells.

Young-sup Yoon, MD, PhD

In Circulation Research, Emory stem cell biologist Young-sup Yoon, MD, PhD and colleagues recently reported converting human skin fibroblast cells into endothelial cells, which line and maintain the health of blood vessels.

Once reprogrammed, a patient’s own cells could potentially be used to treat conditions such as peripheral artery disease, or to form vascular grafts. Exactly how reprogrammed cells should be deployed clinically still needs to be worked out.

In cardiovascular disease, many clinical trials have been performed using bone marrow cells that were not reprogrammed. Emory readers may be familiar with studies conducted by Arshed Quyyumi, MD and colleagues, in which treatment was delivered after patients’ heart attacks. In those studies, sorted progenitor cells, some of which could become endothelial cells, were introduced into the heart. To provide the observed effects, the introduced cells were more likely supplying supportive growth factors.

In contrast, Yoon’s team is able to produce cells that already have endothelial character hammered into them. The authors have applied for a patent. The co-first authors were instructor Sang-Ho Lee, PhD and Changwon Park, PhD, assistant professor of pediatrics. Read more

Posted on February 24, 2017 by Quinn Eastman in Heart Leave a comment

Flow mediated dilation

On Friday, researchers from Emory Clinical Cardiovascular Research Institute demonstrated a test for how much blood vessels adjust to changes in blood flow. This test is known as “flow-mediated dilation” or FMD. A blood pressure measurement cuff is tightened on the arm for five minutes, restricting blood flow.

ECCRI investigator Salman Sher, MD demonstrates flow-mediated dilation

When the cuff is released, blood flow increases, but how much the arm’s main artery expands depends on the endothelium – the lining of the artery — and its ability to respond to nitric oxide, which is induced by the increased flow. Researchers monitor the artery’s expansion by ultrasound.

ECCRI co-director Arshed Quyyumi and his colleagues at Emory have extensive experience using the FMD test. Impaired endothelial function is an early stage in the process of atherosclerosis.

The FMD test is relatively non-invasive, in that no catheter probe is necessary. However, practitioners need practice and careful study design to ensure accuracy, ECCRI investigator Salman Sher explained. Posture, time of day and whether the patient has eaten can all affect the results.

Lab Land asked Sher (seated in the photo) whether the effect was similar to the common experience of sleeping on an arm and having it turn numb, followed by “pins and needles” when the pressure is relieved. This feeling actually comes from nerve compression. Read more

Posted on November 21, 2016 by Quinn Eastman in Heart Leave a comment

CV cell therapy: bridge between nurse and building block

In the field of cell therapy for cardiovascular diseases, researchers see two main ways that the cells can provide benefits:

*As building blocks â€“ actually replacing dead cells in damaged tissues

*As nurses — supplying growth factors and other supportive signals, but not becoming part of damaged tissues

Tension between these two roles arises partly from the source of the cells.

Many clinical trials have used bone marrow-derived cells, and the benefits here appear to come mostly from the â€œparacrine” nurse function.Â A more ambitious approach is to use progenitor-type cells, which may have to come from iPS cells or cardiac stem cells isolated via biopsy-like procedures. These cells may have a better chance of actually becoming part of the damaged tissueâ€™s muscles or blood vessels, but they are more difficult to obtain and engineer.

A related concern: available evidence suggests introduced cells â€“ no matter if they are primarily serving as nurses or building blocks — donâ€™t survive or even stay in their target tissue for long.

Transplanted cells were labeled with a red dye, while a perfused green dye shows the extent of functional blood vessels. Blue is DAPI, staining nuclear DNA. Yellow arrows indicate where red cells appear to contribute to green blood vessels. Courtesy of Sangho Lee.

Stem cell biologist Young-sup Yoon and colleagues recently published a paper in Biomaterials in which the authors use chitosan, a gel-like carbohydrate material obtainedÂ by processing crustacean shells, to aid in cell retention and survival. Ravi Bellamkonda’s lab at Georgia Tech contributed to theÂ paper.

More refinement of these approaches are necessary before clinical use, Â but it illustrates how engineered mixtures of progenitor cells and supportive materials are becoming increasingly sophisticated and complicated.

The chitosan gelÂ resemblesÂ the alginate material used to encapsulate cellsÂ by the Taylor lab.Â Yoon’s teamÂ wasÂ testing efficacy in a hindlimb ischemia model, in which a mouseâ€™s leg is deprived of blood. This situation is analogous to peripheral artery disease, and the readout of success is the ability of experimental treatments to regrow capillaries in the damaged leg.

The current paper builds a bridge between the nurse and building block approaches, because the researchers mix two complementary types of cells: an angiogenic one derived from bone marrow cells that expands existing blood vessels, and a vasculogenic one derived from embryonic stem cells that drives formation of new blood vessels.Â Note: embryonic stem cells were of mouse origin, not human. Read more

Posted on November 16, 2015 by Quinn Eastman in Heart Leave a comment

Deliver, but not to the liver

The potential of a gene-silencing technique called RNA interference has long enticedÂ biotechnology researchers. Itâ€™s used routinelyÂ in the laboratory to shut down specific genes in cells. Still, the challenge of delivery has held back RNA-based drugsÂ inÂ treating human disease.

RNA is unstable and cumbersome, and just getting it into the body without having it break down is difficult. One that hurdle is met, there is another: the vast majority of the drug is taken upÂ by the liver. Many current RNA-based approaches turnÂ this apparent bug into a strength, because they seek to treat liver diseases. See these articles in The Scientist and in Technology Review for more.

But what if you need to deliver RNA somewhere besides the liver?

Biomedical engineer Hanjoong Joâ€™s lab at Emory/Georgia Tech, working with Katherine Ferraraâ€™s group at UC Davis, has developed technologyÂ to broadenÂ the liver-dominantÂ properties of RNA-based drugs.

Hanjoong Jo, PhD

The results were recently published in ACS Nano. The researchers show they can selectively target an anti-microRNAÂ agent to inflamed blood vessels in mice while avoiding other tissues.

â€œWe have solved a major obstacle of using anti-miRNA as a therapeutic by being able to do a targeted delivery to only inflamed endothelial cells while all other tissues examined, including liver, lung, kidney, blood cells, spleen, etc showed no detectable side-effects,â€ Jo says. Read more

Posted on September 1, 2015 by Quinn Eastman in Heart Leave a comment

Ebola’s capriciousness in kids

Anita McElroy, a pediatric infectious disease specialist at Emory, and her colleagues at the CDC, led by Christina Spiropoulou, have been getting some attention for their biomarker research on Ebola virus infection. Sheri FinkÂ from the New York Times highlighted their work in a Nov. 9 report on the infection’s capriciousness. Genetics may also play a role in surviving Ebola infection, as recent animal research has suggested.

McElroy’s team’s findings attracted notice because their results suggest that Ebola virus disease may affect children differently and thus, children may benefit from different treatment regimens than those for adults. The authors write that early intervention to prevent injury to the lining of blood vessels — using statins, possibly — might be a therapeutic strategy in pediatric patients. Read more

Posted on November 11, 2014 by Quinn Eastman in Immunology Leave a comment

Targeting antioxidants to mitochondria

Why arenâ€™t antioxidants magic cure-alls?

Itâ€™s not a silly question, when one sees how oxidative stress and reactive oxygen species have been implicated in so many diseases, ranging from hypertension and atherosclerosis to neurodegenerative disorders. Yet large-scale clinical trials supplementing participantsâ€™ diets with antioxidants have showed little benefit.

Emory University School of Medicine scientists have arrived at an essential insight: the cell isnâ€™t a tiny bucket with all the constituent chemicals sloshing around. To modulate reactive oxygen species effectively, an antioxidant needs to be targeted to the right place in the cell.

Sergei Dikalov and colleagues in the Division of Cardiology have a paper in the July 9 issue ofÂ Circulation Research, describing how targeting antioxidant molecules to mitochondria dramatically increases their effectiveness in tamping down hypertension.

Mitochondria are usually described as miniature power plants, but in the cells that line blood vessels, they have the potential to act as amplifiers. The authors describe a â€œvicious cycleâ€ of feedback between the cellular enzyme NADPH oxidase, which produces the reactive form of oxygen called superoxide, and the mitochondria, which can also make superoxide as a byproduct of their energy-producing function.

Posted on July 14, 2010 by Quinn Eastman in Heart Leave a comment