Careful-Breathing Can Be Hazardous

The pessimists are right; even breathing can be hazardous to your health. Because as mammals humans inhale oxygen, they and other mammals open themselves up to oxidative stress, which can interfere with a cell's ability to perform normal functions. Diana Bigelow, associate professor of biochemistry and biophysics in KU's molecular biosciences deparment, is researching how increased oxidative stress, specifically as a result of the aging process, destroys a cell's ability to regulate calcium levels.

To understand just what that means for the body, Bigelow says it helps to understand calcium signaling. In a resting muscle cell, the cytoplasm normally maintains low concentrations of calcium. Calcium is stored in a cellular storehouse called the endoplasmic reticulum. When the brain tells a musclecell to contract, calcium provides the trigger for contraction. Proteins called calcium channels release calcium from the endoplasmic reticulum into the cytoplasm where calcium binds to the contractile fibers and causes them to change shape, or contract. For a muscle cell to return to a resting state (relaxation), proteins on the surface of the endoplasmic reticulum called calcium pumps reset the calcium gradient for the next contraction. So in a healthy muscle cell, calcium cycles in and out of the cytoplasm with a rapid frequency.

Bigelow's primary interest is twofold: First, she wants to understand how these protein pumps and channels work and secondly, how they are impaired when the body undergoes oxidative stress, primarily that experienced during the aging process.

"Oxygen is a very reactive element," Bigelow says. "And because we live on it and have to process it, this makes us highly susceptible to oxidative stress." The problem begins, she explains, when inhaled oxygen is inevitably converted into reactive types of chemicals called reactive oxygen species (ROS). Though cells are designed to capture most ROS, about 3 percent does escape into the cell and may damage cellular proteins with conccurrent loss of cellular function. In a healthy cell, cellular function is continually restored by repair of degradation of damaged proteins. When cellular conditions are such that damaged proteins are not repaired, this is called oxidative stress. Oxidative stress is largely responsible for the damage to muscle proteins associated with the aging process and with numerous illnesses such as Alzheimer's and Lou Gehrig's disease, emphysema, heart attacks, stroke and cancer.

When Bigelow and her researchers examine aging muscle cells, they see the same cycles of calcium release and removal as in healthy cells, but the whole process slows down considerably because of oxidative damage to both calcium pumps and channels. This damage contributes to muscle weakness associated with age.

Bigelow says if she and her researchers can better understand how healthy muscle cells handle ROS, this understanding could pave the way for the design of therapeutic strategies to mitigate oxidative damage in older or diseased cells.

To answer some of these fundamental questions, she and her research group have begun to analyze chemical changes that trigger structural and functional changes in these cellular proteins during oxidative stress. They then try to monitor how the whole cell responds to the oxidatively damaged proteins.

Bigelow says that when a protein molecule becomes damaged, it may or may not be crucial to cellular operation. She and her researchers are now trying to determine which cellular proteins are crucial to the cell's survival. One target for study is the process of nytrosyration of tyrosine, which Bigelow says forms in substantial amounts on the calcium pump in aging muscle cells, perturbing the pump's function.

Even more exciting to Bigelow is her lab's discovery of an activity in muscle cells that actually repairs this chemical modification. She says, "This repair process involves a denitrase, and our idea is that it removes the oxidatively damaged nitro group off the tyrosine." Tyrosine is a normal amino acid on a protein that is converted into nitrotyrosine by ROS. At this point, many of the details surrounding the repair process have yet to be discovered. "We're left with several significant research questions now," Bigelow says. "For example, when does this repair enzyme 'kick-in'? Does it only repair severely damaged cells under conditions of oxidative stress, or does it continually repair damage as it occurs? Does it mediate partial or total repair of the cell? What type of structural changes to damaged proteins are necessary for repair?"

Bigelow and her researchers also know that calcium pumps and channels are continually, albeit slowly, degraded and replaced with newly synthesized proteins to maintain cellular function in the muscle. What she would like to discover is the role these degradation-resynthesis pathways play in ridding the cell of proteins damaged by ROS. Bigelow would also like to know where, in an aging muscle cell, do the the defects that lead to the accumulation of nitrotyrosine-modified calcium pumps originate? She is also exploring whether defects exist in repair or degradation pathways, or whether the repair-degradation processes in some aging cells can no longer keep up with the amount of damage.

"Over the past six years, we've seen an increase in the number of people looking at calcium signaling in all kinds of cells, but no one has really looked at calcium signaling in the muscle cell as it relates to the aging process," Bigelow says. "My research group is part of a large, multi-investigator Program Project funded by the National Institute on Aging that allows us to pool answers with other researchers studying aspects of aging, oxidation, and calcium regulation. This makes our work both easier and more exciting."

Bigelow has been able to take advantage of the presence of considerable expertise already present at KU in both methods for analyzing chemical modifications to proteins and for the study of calcium signaling in cells.

Most importantly, the increased research into this area of calcium signaling could clear a path for a healthier life for all of us as we age.