INSULIN AND SARCOPENIA IN THE ELDERLY (CYCLE NO, 2)

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This subproject is one of many research subprojects utilizing the resources provided by a Center grant funded by NIH/NCRR. The subproject and investigator (PI) may have received primary funding from another NIH source, and thus could be represented in other CRISP entries. The institution listed is for the Center, which is not necessarily the institution for the investigator. Background: Muscle mass and function progressively decline with aging. This process has been termed sarcopenia, and is associated with increased risk of falls and vulnerability to injury, especially bone fracture, which increase the risk of functional dependence. Further, decreases in muscle mass and function can lead to a reduction in physical activity, which may have possible effects including weak bone, obesity, and borderline diabetes. Undernutrition and disuse have also been identified as potentially preventable contributing factors. Nutritional intervention is an appealing method of prevention and treatment of muscle loss with aging (sarcopenia) in the elderly due to its wide applicability and safety. However, attempts to improve muscle mass and strength in the elderly with nutritional supplements have failed to demonstrate a beneficial effect. Furthermore, nutritional supplementation added to resistance exercise did not result in an increase in muscle mass or strength compared to exercise alone. Hypothesis: Our general hypothesis is that a reduced response of muscle protein synthesis to insulin plays an important role in the loss of muscle mass that takes place with aging. We further hypothesize that this change is specific for proteins regardless whether the patient is diabetic or not. Our preliminary results suggest that there is a change in the response of muscle protein synthesis to insulin in the elderly, which can compromise the positive effect of amino acids on muscle protein gain. Our goal is to determine the mechanisms underlying the age-related insulin resistance of muscle proteins, which will allow us to define specific interventions to target this defect and provide the scientific basis for the prevention and treatment of sarcopenia. Specific Aims and Procedures (summary): (1) To determine if the inhibition of insulin-induced vasodilation reduces the response of muscle protein synthesis and net balance to insulin in young healthy subjects. (2) To determine if the muscle growth in response to insulin or a mixed meal improves in older subjects when blood flow and muscle perfusion are restored to youthful values with the use of a medication. (3) To determine in older subjects if a bout of aerobic exercise improves the response of blood flow, muscle perfusion, and muscle growth to insulin or a mixed meal. Experimental Design (summary): Volunteers will be recruited by an institutional review board-approved advertisement that will be posted at the University of Texas Medical Branch, local newspapers, and social centers, and by word of mouth. The volunteers will be informed about the risks and the benefits of the studies and will be asked to sign an approved consent form. The volunteers will arrive fasted at the GCRC in the morning. We will check them for any diseases that would not allow them to participate in the study. They will be enrolled only if the screening tests show that they do not have any health problems that may prevent them from participating in the study. If subjects are assigned to the exercise group they will walk or run on a treadmill, or cycle on a stationary bike for 15 minutes with increasing intensity (speed increments on the treadmill, resistance increments on a bike) while we are analyzing the activity of your heart by EKG. Randomization: Subjects will be assigned randomly to any of the following groups: for 18-35 years old the possible groups are the following: insulin alone, insulin with L-NMMA, L-NMMA alone; meal ; for 65 or older the possible groups will be the following: insulin alone, insulin with sodium nitroprusside; insulin with exercise; meal alone; meal with sodium nitroprusside; meal with exercise. Admission: Subjects will be admitted to the GCRC the afternoon before the study. The nurses will measure weight, height, blood pressure, and draw blood to check again blood count, clotting, and some other blood tests including electrolytes (salts) and liver function. Should these tests result abnormal subjects will not be allowed to participate to the study. Once the admission procedure is completed, a DEXA scan will be performed. Subjects assigned to an exercise group will complete an exercise session before dinner. They will not be allowed to eat or drink anything but water after dinner. Water or iced water will be allowed at any time during the night and during the study. We will start the experiment the next morning. The experiment consists of placing catheters, draw blood from those catheters, taking muscle biopsies, and infusing stable isotopes, indocyanine green, Definity microbubbles. Additionally subjects may receive insulin or a liquid mixed meal, and/or L-NMMA or sodium nitroprusside, according to the group assigned. DEXA scan. The DEXA scan will allow us to measure the amount of bone, muscle and fat in the body. Subjects will have to lie down on the DEXA table for a few minutes, while a technician will take pictures of the bones, muscles, and fat. Exercise session: Subjects assigned to an exercise group will walk fast on the treadmill for 45 min before dinner. Catheters. The next morning we will place one catheter (a small tube for drawing blood or putting fluids in the body) in a vein of each arm. We will also place one catheter in the femoral artery and one in the femoral vein of one leg (the vessels that bring in and drain the blood out of the leg), at the top of the thigh, close to the groin fold. The femoral catheters will be placed using a sterile procedure to avoid infections, plus local anesthesia with lidocaine (numbing medicine) will be injected to eliminate pain. An ultrasound machine will be used to better visualize the blood vessels of the leg. Stable isotopes. After the catheters are placed, we will start to give stable isotopes through one of the arm catheters. This will go on until the end of the study, about 7 hours. In general, the stable isotopes of a material are very similar to the regular material, except that they are heavier, but not radioactive. An example of stable isotope is the "heavy water", which is just a stable isotope of water. Stable isotopes are natural compounds also present in very small quantities in the body. We will infuse an extra amount of stable isotopes of amino acids (the building blocks of proteins) and glucose (sugar) to increase the level of the naturally occurring isotopes by 2-5 %. The isotopes are tested in order to assure their sterility (absence of microorganisms, such as bacteria, that may give infections and fever). We will then be able, using highly sophisticated analytical instruments, to measure the small changes in the concentration of these substances in the blood and muscle. This will allow us to compute the rate of metabolism, exchange, or turnover of these substances in the body. Muscle biopsies. During the study we will take 3 muscles biopsies, 3 hours apart from each other, from the same leg in which we placed the catheters. This procedure involves the taking of a small piece of muscle from the leg. The skin is cleaned and made sterile, and the skin and tissue below are injected with local anesthetic (numbing medicine) to eliminate pain. We will make a small incision about the size of this dash "__" approximately at mid-thigh, through which a needle about the size of this letter "O" will be advanced into the muscle. A piece of the thigh muscle will then be removed with the needle, the skin closed with a steri-strip, and a light dressing applied. The last two biopsies will be taken from the same incision where the first one was taken. After the last biopsy the incision will be closed with one stitch or a special skin glue (Dermabond). Blood draws. The total amount of blood drawn during the study will be approximately 200 ml (approximately 6.5 ounces or 13 tablespoons). Indocyanine green. During the study, indocyanine green, which is a dye, will be given in the artery of the leg to measure the blood flow. Insulin. During the last three hours of the infusion study, for subjects assigned to an insulin group we will give insulin into the leg through the arterial catheter. This method will allow us to expose the leg muscles to an amount of insulin much higher than the rest of your body, so that the blood sugar and the blood amino acids (the building blocks of proteins) should not drop very much. The insulin will be diluted in sterile saline (diluted salted water) containing a small amount (less than 1 teaspoon) of plasma (a component of your blood). During the administration of insulin we will frequently draw very little amounts of blood (1 ml, approximately a few drops) to check that blood sugar remains normal. Should the blood sugar drops due to insulin we will give some extra sugar through one of the arm catheters to maintain the blood sugar normal. L-NMMA infusion. L-NMMA is a vasoconstrictor (reduces blood supply). Subjects assigned to the L-NMMA group, we will infuse it in the femoral artery during the insulin infusion to prevent the normal increase in blood supply that follows insulin infusion. We will continuously check the blood supply to the leg and adjust the L-NMMA infusion to avoid a reduction of blood supply to the leg below the initial level. Sodium nitroprusside. Sodium nitroprusside is vasodilator normally used to increase blood supply and reduce blood pressure. Subjects assigned to one of the sodium nitroprusside groups we will infuse it in the femoral artery of one leg in order to double the blood supply to that leg. Definity Microbubbles. This is a contrast agent approved by the FDA for measuring blood supply to the heart. We will use it to visualize the blood supply in the leg capillaries (very small blood vessels) with an ultrasound machine. While this contrast agent is referred to as microbubbles, there is no risk of blocking a vessel or introducing air into the circulation. The microbubbles are smaller than a red blood cell and are known to move through the body similar to red blood cells. Length of protocol. The total length of the study will be approximately 7 (seven) hours. However, once the stable isotopes and insulin are stopped and the catheters removed, subjects will have to lie down in the bed for additional 2 (two) hours, to make sure that the femoral vessels are not leaking, which could make a big bruise. A focused physical examination will be performed by a physician during and after the experiment to determine the presence of any abnormality in the affected leg. Physical activity after the study. To avoid problems at the site of insertion of the femoral catheters we recommend that subjects do not engage in strenuous physical activity (running, weight lifting, etc.) during the 24 hours following the study. Significance (summary): Our project is designed to clarify part of the complex mechanisms that induce the loss of muscle mass in aging. Specifically, we will focus on the changes that occur with aging in the response of muscle perfusion and muscle protein metabolism to insulin. Overall, we believe that this proposal will allow us to better delineate the changes in muscle metabolism with aging, thus providing a solid scientific basis for future intervention to prevent and treat muscle loss with aging (sarcopenia).