Oxidative stress has been long known to fuel disease, but how exactly it damages various organs has been challenging to sort out. Now scientists from Johns Hopkins say research in mice reveals why oxidation comes to be so corrosive to heart muscle.
A report on the results, published online May 4 in The Journal of Clinical Investigation, shows that oxidation inside the cardiac cells precipitates heart failure by disrupting the work of a heart-shielding protein called PKG, known to act as a natural "brake" against biological stressors like chronically elevated blood pressure, inflammation and pressure overload. The team's experiments show that oxidation alters a key part in PKG's structure, interfering with this natural braking system and diminishing its ability to blunt stress on the heart muscle.
Known triggers of oxidative stress in the heart include heart attacks, choked off flow of oxygen to the heart muscle known as ischemia, smoking and even environmental pollution. Because the new findings unravel some of the mechanisms of damage caused by oxidative stress, the researchers say these insights could spark the development of therapies to halt or slow down the development of heart failure, a condition estimated to affect 23 million people worldwide.
Marked by the gradual weakening, enlargement or stiffening of the cardiac muscle, heart failure culminates in the organ's loss of blood-pumping ability.
"Our results suggest that precision-targeted therapies that prevent oxidation in key proteins of the cardiac cell could boost self-protective mechanisms and stave off the development or stop the progression of heart failure," says senior investigator David Kass, M.D., professor of medicine at the Johns Hopkins University School of Medicine and its Heart and Vascular Institute.
In an initial set of observations, the researchers demonstrated that PKG is indeed oxidized in the heart cells of mice and men. Next, to test how oxidation affects the heart muscle, researchers surgically altered the hearts of mice in a way that forced them to pump harder -- a technique that induces heart failure. Mice with normal forms of PKG developed full-blown disease, marked by weakened and enlarged heart muscle. However, animals engineered to have oxidant-resistant forms of PKG had much milder disease, a critical clue that oxidation is a key catalyst in heart disease, the researchers say.
Chemically, oxidization occurs when a molecule loses an electron, or gets "oxidized," though despite its name, the reaction does not require a gain or loss of oxygen. Everyday examples of oxidation include metal rust or brown spots on an apple. Biologically speaking, some oxidation is normal. A byproduct of cellular function, it can be an important defense mechanism to thwart infection and kill harmful viruses and bacteria. Under normal conditions, the process is kept in check by cellular antioxidants that mop up excess oxidation. However, when certain cell proteins become highly oxidized, the result is often cell death, tissue damage and, eventually, overt disease.
In the case of PKG, Kass says, oxidation alters its shape by forming a chemical bond -- akin to inserting a "strut"-- in a key segment of the protein that functions as a zipper that opens and closes to bind to certain molecules. The team's experiments revealed that oxidation interferes with the zipper's normal function, preventing PKG from binding to key substances. PKG's ability to interact with other proteins is critical, Kass says, because these substances help ferry PKG around the cell, allowing it to eventually move to the most advantageous, strategic spot from where it can best shield cardiac cells from stress.
A set of experiments underscored the link between location and performance -- where PKG was turned out to be critical to how well it did its job, the researchers found. After briefly exposing cardiac cells to the potent oxidizer hydrogen peroxide, PKG made its way from the interior of the cell to the cell membrane where it bonded with and blunted the effects of a misbehaving protein, TRPC6, known to stimulate heart muscle scarring and enlargement. However, prolonged exposure to hydrogen peroxide coaxed PKG to leave the membrane and return to the cell interior, where it could no longer perform its gatekeeping duties. The heart cells of mice engineered to resist oxidation, however, kept PKG safely secluded in the membrane even after prolonged oxidation exposure, the study showed. When researchers analyzed cardiac cells from mice with surgically induced heart failure, they noted the same effect: Normal PKG was diffused inside the cell, while oxidation-resistant PKG remained in the outer membrane.
"Location does matter," says study lead author Taishi Nakamura, M.D., Ph.D., a cardiology fellow at the Johns Hopkins University School of Medicine. "Like a sentry who abandons his post in the watchtower, once back inside the cell, PKG can no longer spot and disarm harmful proteins on the periphery. It fails to protect the 'castle.'"
Results of the latest study, the research team says, highlight the need to develop "smart" treatments that keep excess oxidation in check. Understanding which proteins get oxidized and fuel disease is the key to designing targeted treatments that extinguish small flare-ups without completely shutting off normal oxidation.
"Figuratively speaking, oxidation is the cellular equivalent to having a fireplace inside the home," Kass says. "Poor insulation and lack of regular maintenance could cause it to burn the house down, but to prevent a fire, you don't get rid of the fireplace. You tend to it carefully to quash problems before they arise."
Previous work by Kass' team demonstrated that glitches in two separate signaling pathways also interfere with PKG and cause heart muscle damage. These earlier results, he says, together with findings from the latest study, reinforce the idea that PKG is an all-important heart protector whose good work can be sabotaged on multiple fronts by various breakdowns in the cellular apparatus of the heart.
The work was supported by the National Institutes of Health under grants HL-119012, HL-093432, HL-114910, HHSN268201000032C, and HL-107153; the Fondation Leducq Transatlantic Networks of Excellence; Abraham and Virginia Weiss and Michael and Janet Huff Endowments; the National Heart, Lung and Blood Institute grant R01-GM090161 and NHLBI 32-HL-07227k; the American Heart Association; the Japan Heart Foundation; the Sarnoff Foundation; and the British Heart Foundation under grant RG/12/12/29872.
Other investigators involved in the study included Mark Ranek, Dong Lee, Virginia Shalkey Hahn of Johns Hopkins; Philip Eaton of King's College in London; and Choel Kim of the Baylor College of Medicine.
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