Metformin Life-Span Study
After finding that metformin produces a gene-expression profile similar to that of long-lived CR mice, we decided to study the effects of long-term administration of metformin on mean and maximum life span and other biomarkers of aging in mice. An initial metformin life-span study was begun at the University of California, Riverside, in 2003, but design flaws in this study convinced us to set up another metformin life-span study in our northern California lab.
In the new Metformin study, we used a large number of mice fed a normal diet and supplemented with a dose of metformin comparable to that usually used in humans. A control group was fed a normal diet without metformin. This study, begun in early 2004, is expected to be completed around September 2007.
In 2004, we repeated high-density microarray studies of the livers of normal and CR mice with new Affymetrix chips that measure the expression of 34,000 genes in order to generate data on the entire mouse genome. We found many changes in gene expression in the CR mice that are different from those found in normal mice. Analysis of these results continues.
Grape Seed Extract with Resveratrol
In September 2004, a group of normally fed mice was supplemented with either synthetic resveratrol or grape seed extract with resveratrol obtained from Life Extension. Our objective was to compare gene expression in the livers of the mice in the grape extract group to gene expression in the livers of normally fed and CR mice.
Life Extension’s grape seed extract with resveratrol included two standardized extracts from the seeds and skin of whole red grapes (Vitis vinifera), which contain proanthocyanidins, catechin, epicatechin, gallic acid, and 10% resveratrol from whole red grapes and Polygonum cuspidatum root extract. It also included vitamin C and calcium from calcium ascorbate.
When we compared gene expression in the livers of mice fed grape extract with resveratrol to that found in CR animals, we found that substantial numbers of genes were expressed in a similar fashion. These results suggest that the molecular pathways that these genes regulate are activated in both groups, which could mean that the grape seed extract with resveratrol is an anti-aging therapy. Further detailed pathway analysis is under way and we will likely do a life-span study using Life Extension’s grape seed extract in the future.
Other Gene-Expression Studies
We also conducted gene-expression studies of livers from mice treated with two types of the herbal extract cat’s claw. One of the cat’s claw extracts includes oxindole alkaloids (BMcp-1), while the other is depleted of polysaccharides, tannins, and oxindole alkaloids (BMcp-2).
Tissues from both groups of mice treated with either BMcp-1 or BMcp-2 were harvested and processed. Specimens have been sent to Stanford University for further gene-chip processing. Image files containing the data from these groups have been received and data analysis is under way.
Studies at the Chinese Academy of Sciences
In collaboration with researchers at the Chinese Academy of Sciences in Beijing, Life Extension has funded studies of its grape seed extract with resveratrol, BMcp-1, BMcp-2, and BM-A1 (a plant extract), as well as a green tea and ginkgo biloba extract developed by Dr. Baolu Zhao, an authority in free radical biology.
One of the Chinese studies examining the grape seed extract was in Drosophila (fruit flies) that were bred to contain the human mutant alpha-synuclein gene. This gene is expressed in the fruit fly brain similarly to that in humans, so that the flies replicate the essential features of human Parkinson’s disease—age-dependent loss of dopaminergic neurons and motor impairment—that manifests (in the flies) as a progressive loss of climbing ability.
Groups of these fruit flies received four doses of the Life Extension grape seed extract in their food. Climbing ability was evaluated in each group and the data were analyzed statistically. The results showed a progressive decline in climbing ability with advancing age, and that this decline was reduced in flies supplemented with grape seed extract, an effect that was more noticeable in male Parkinson’s flies than in female Parkinson’s flies. Interestingly, the dose of grape seed extract that most inhibited the decline in climbing ability of the Parkinson’s flies was similar to that used in a life-span study in which flies given the grape seed extract lived 15% longer than those fed a normal diet. Flies receiving the grape seed extract with natural resveratrol showed greater life-span extension than those receiving synthetic resveratrol.
In another study, mitochondria from rat livers were exposed to damaging carcinogens, followed by treatment with either Life Extension grape seed extract or BMcp-2. The results showed that treatment with certain doses of grape seed extract or BMcp-2 protected the mitochondria from damage. Mitchondria are the cellular power plants that fuel the body’s life processes.
These findings suggest that Life Extension’s grape seed extract with resveratrol may eventually play a role in preventing or treating Parkinson’s disease in humans. They also suggest that Life Extension’s resveratrol, which is extracted from whole red grapes and Polygonum cuspidatum roots, may be superior to synthetic resveratrol.
Is BM-A1 an Anti-Obesity Therapy?
BM-A1, a unique plant extract formula developed by Dr. Zhao of the Chinese Academy of Sciences, contains polyphenols, proanthocyanidins, and flavones derived from green tea and gingko biloba. BM-A1 shows promise for promoting cardiovascular and metabolic health, preventing cancer, and fighting obesity.
In one study, the Chinese scientists fed rats a high-fat diet, which caused them to gain weight, develop larger and heavier livers, and demonstrate significant elevations in serum and liver triglyceride levels. Treatment with BM-A1 reduced the weight of rats fed the high-fat diet as well as those fed a normal diet, reduced the size and weight of their livers, and lowered their triglyceride levels.
This study identified mechanisms of action involved in preventing obesity, which are linked to an important molecular pathway related to fat metabolism. This pathway involves a set of nuclear receptors known as peroxisome proliferator-activated receptors (PPARs). The scientists found that BM-A1 exerts its anti-obesity effects by modulating PPAR pathways. In doing so, BM-A1 inhibits the accumulation of lipids in adipose tissue, stimulates the utilization of triglycerides, and reduces lipid peroxidation caused by free radical activity.
A clinical trial was conducted to gauge BM-A1’s effects on lipid, glucose, and blood pressure levels in humans. The subjects were given BM-A1 and then followed for three months. At the end of this period, 71% showed major lowering of blood lipids and 29% showed moderate lowering; 59% showed major lowering of blood glucose, with 41% showing moderate lowering; and 72% showed major lowering of blood pressure, with 28% showing moderate lowering. A clinical trial examining BM-A1’s effects on obesity in humans needs to be conducted. Since BM-A1’s effects in rats are similar to those produced by CR and metformin in mice, it will be interesting to investigate gene-expression changes in mice given BM-A1.
With Life Extension-funded collaborative studies under way at prestigious institutions in the US and China, we expect to move forward with greater speed in identifying the genes and molecular pathways involved in aging and life extension. Using this knowledge, we hope to develop therapies to control aging and extend the healthy human life span.
Life-Extending Promise of Hypothermia
Terri Schiavo suffered an estimated 10 minutes of cardiac arrest that severely damaged her brain. Afterwards, her relatives fought over whether to keep her in a “vegetative” state on artificial life support (in this case, a feeding tube and pump) or “turn off the machines” and let her die. A court ruling finally settled the matter by deciding that her prognosis was so poor that she should be allowed to die. An autopsy eventually showed that her brain was about half the weight of a normal brain, meaning that about half of its neurons were gone.
Every day, people suffer heart attacks that either kill them or cause severe brain damage. This occurs because, at normal body temperature, after about five minutes without oxygenated blood being pumped through the brain, the brain’s neurons suffer lethal damage. Severe brain damage caused by oxygen deprivation may also be caused by other conditions, including strokes and closed head injuries.
Although neurons may suffer lethal damage in five to 10 minutes, they do not actually die and disappear until after many hours, or even days. This leaves a window of opportunity to save them. For over 50 years, scientists have known that cooling the brain (and the rest of the body) can have dramatic effects. One of these is to protect against the biochemical processes that kill neurons in brains with inadequate oxygen, a state caused by reduced or absent blood circulation, or ischemia. The success of hypothermia in brain protection is demonstrated in drowning cases, where victims have been revived after as long as 60 minutes of immersion in icy water, because their brains were cooled. This state of cooling is known in medicine as hypothermia.
Hypothermia slows down life processes. If applied properly after a person suffers from acute oxygen deprivation, hypothermia has the power to slow down the dying process in damaged neurons. This could save the lives of millions of people. Dogs have been resuscitated after as long as 15 minutes without blood flow at normal body temperature. This was done by cooling the animals with a bypass machine after their hearts were restarted. The same should be able to be done in humans. Patients like Terri Schiavo could be cooled during resuscitation, so that they could eventually live a normal life instead of suffering permanent brain damage.
The deepest hypothermia has been achieved in medical centers using a cardiopulmonary bypass system, with a heat exchanger to cool blood directly while it circulates outside the body. Such systems have lowered brain (and body) temperatures rapidly enough to permit doctors to operate successfully on patients in the absence of blood flow. For example, doctors need to induce hypothermia safely in order to perform operations on the vessels of the brain and heart, which cannot be performed while the heart is still beating and blood is flowing.
However, induced hypothermia has had limited use in medicine because it has proved difficult to cool patients rapidly in emergencies without inducing some degree of damage. In resuscitation, the use of hypothermia has been limited because it cannot be done rapidly and safely enough in the field to protect the patient completely until he or she can be moved to a well-equipped medical center. Even then, inducing even mild hypothermia takes hours. The vast majority of people who suffer heart attacks, stroke, and head injuries do so at home, work, or play, not when they are in a hospital. In the case of soldiers who suffer cardiac arrest from loss of blood after severe injury, safe and rapid cooling may currently be impossible.
Funding Induced Hypothermia Research
Life Extension has been funding induced hypothermia research at Critical Care Research (CCR), a laboratory in California. Directed by Steven Harris, MD, the CCR research program is developing an advanced technology to deliver safe and rapid cooling in hospitals, clinics, and ambulances. At CCR, scientists have identified several destructive processes that cause damage in oxygen-deprived brains, including an acute inflammatory response that responds to treatment with cold. Cooling the brain after brain injury has proven to be very similar to treating a burn or sports injury with the application of ice.
The technology that CCR invented and has been developing is called liquid ventilation lavage. This process involves the use of a fluorocarbon liquid cooled to near-freezing temperature in order to lavage (wash in cycles) the lungs of experimental anesthetized dogs. In this process, the lungs are not completely filled with fluid, and they work as both heat exchangers and gas exchangers. This process is able to produce very rapid cooling of the animal’s brain and body core, from 100° to 85° Fahrenheit (F) in less than 20 minutes. More than 100 dogs have been successfully revived using this process. Most of these animals have shown signs of minor lung damage for a couple of days after revival, but have recovered completely afterwards. Current liquid ventilation lavage research at CCR is aimed at eliminating all lung damage from dogs that undergo rapid cooling, and redesigning the components of the system into a portable unit capable of being operated by paramedics in an ambulance.
|Figure 7. Portable liquid ventilation system used by CCR.|
Developing Portable Liquid Ventilation
For several years, CCR worked with an engineering firm to place its lavage equipment into a prototype portable system. This required a redesign and computerization of the system’s first pump module, which originally weighed more than 100 lbs. The design eventually was reduced to a double-chamber piston system, which weighed less than 35 lbs. and was small enough to be carried in one hand with a strap. This system needed to be gas powered from a small pressure tank, and also required a vacuum source. Another CCR invention is portable heat exchangers made from pre-frozen “ice cartridges,” which work by routing fluorocarbon through fine tubes immersed in ice. With this system, the amount of ice needed for adequate cooling of a 45-lb. dog was reduced to only 8 lbs. CCR continues to conduct dog experiments with this system. (Fig. 7.)
In 2003, CCR engaged the services of a second engineering team to see whether it could help solve some problems that had developed with the first portable system. The new team proposed a redesign of the system that would use only electrically driven pumps. This allowed for direct electrical drive of both lavage liquid infusion and suction, and eliminated the need for both the vacuum and compressed air sources that drive the older system. A new prototype lavage machine has now been built as a breadboard device and has been tested successfully in animal experiments. A few problems remain. Future devices will address problems in the delicate timing of lavage cycles under software control. This approach is expected to result in a smaller, more portable system driven directly by AC (alternating current) power or batteries, which will do away with the need for a heavy vacuum or compressor system.
Critical Care Research now routinely cools dogs from 100° to 92° F by liquid lavage with almost no lung problems, resulting in animals that are up and walking the next day, usually “without even a wheeze,” according to Dr. Harris. He notes that since CCR published the first report of fluorocarbon lung lavage cooling in 2001, the lavage method for cooling and heating animals has been confirmed in other laboratories working with rabbits and pigs. With liquid lavage, there is not as much liquid to move compared with total liquid ventilation, and so there is less pressure and damage. Gas ventilation can be used simultaneously to take care of oxygen and carbon dioxide, so that the animal can be cooled without damage.