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Aging Not Required

BioTechniques

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3/28/2014

As we age, our bodies seem to slip into disrepair. Our hearts enlarge. Our muscles limp back after injury. Our memories fade. It makes sense: everything, including the human body, naturally deteriorates over time. But a set of new studies dispels the notion that aging is simple wear and tear.

“It’s apparent to anyone who looks at an aging organism that things are changing all over the body, and they’re changing with some synchronicity,” said Amy Wagers of the Harvard Stem Cell Institute. “That suggests that there might be some coordination in the process.”

If, instead of haphazard and inescapable erosion of function, aging is more systematic and preordained, might we be able to put an end to this pathway of decline?

“It’s really a new appreciation in the field that the process is not only molecularly accessible and can be slowed, but it can also be reversed,” said Wagers. Researchers, then, are redefining “aging” itself.

Young Blood

Back in 2005, Wagers and her colleagues Tom Rando and Irina Conboy published a paper looking at aging liver and skeletal muscle (1). She knew that specific changes occurred in these tissues as animals aged (such as the loss of Notch signaling in muscle and a decline in hepatic progenitor cell proliferation in liver), and was curious to know whether these detrimental alterations could be halted.

So she dusted off an old method called parabiosis, an experimental model in which two animals are surgically joined so that they share a blood system. This method first emerged in the 19th century, but in 2005, Wagers took it one step further. She joined the skin surface microcirculation of mice in so-called heterochronic parabiotic pairings—basically, an old mouse attached to a young mouse.

Wagers found that old mice exposed to young blood enjoyed restored Notch signaling, improved proliferation and regeneration of muscle satellite cells, and increased hepatocyte proliferation. After injury, their muscles healed much better than old mice not joined with young mice.

“The effect was so profound,” said Wagers. But no one knew why young blood had this effect.

By 2013, Wagers’ and Richard Lee’s lab, also at the Harvard Stem Cell Institute, had begun examining the heart (2). As the heart ages, it often enlarges in a process called cardiac hypertrophy. Using heterochronic parabiosis again, this time in a mouse model of cardiac aging, the team joined young and old mice for four weeks and found that cardiac hypertrophy was strikingly reduced.

This time, they searched for what, exactly, caused the improvement. The team used a broad proteomics analysis approach that applied aptamer-based technology (the SomaLogic proteomics discovery platform SOMAscan) to quantify proteins that differed in the blood of old versus young mice. They found 13 that differed, and only one, growth differentiation factor 11 (GDF11), a member of the activin/TGF-beta superfamily of growth and differentiation factors, was higher in the blood of aged mice joined to young mice. Furthermore, injection of recombinant GDF11 into older mice for 30 days didn’t just slow cardiac hypertrophy—it reversed it.

“It tells us something about the fundamental biology of aging that there is just one protein that is communicating with cells in systems that are very different in their normal physiology, metabolism, response to injury, daily function,” said Wagers. “And yet they’re all listening to the same signal. I think that’s a fascinating observation.”

Brain Gain

Now, researchers are finding that the effect of GDF11 is even more widespread. In a May paper in Science, Lida Katsimpardi, a postdoc in Lee Rubin’s lab at the Harvard Stem Cell Institute, examined neurogenic niches, which are essentially nurseries in the adult brain that birth new neurons from neural stem cells (3). A network of vessels supplies blood to these regions, and as we age, the vasculature deteriorates and blood flow diminishes. This is associated with fewer new neurons being made and reduced cognitive function. Rubin, Katsimpardi, and their team hypothesized that restoring blood flow, and presumably functionality, to this niche should remove some of the negative effects of aging on cognition.

Katsimpardi joined young and old mice in the heterochronic parabiosis model for five weeks. She looked at the subventricular zone (SVZ) where new neurons are formed and found that young blood increased neural stem cells in older mice, while older blood reduced neurogenesis in young animals. Cultured neural stem cells from older animals joined to young mice generated more neurons than those from old control mice.

To see how these increases in neural stem cells alter function, Katsimpardi followed the neuron migration trail to the olfactory bulb. She saw a 92% increase in olfactory neurogenesis in older mice connected to young animals than in older control mice joined with other older mice. She then separated some mice from their parabiotic partners and exposed them to various odorant concentrations, finding that older mice previously joined to younger mice spent more time exploring low odorant concentrations and avoiding high concentrations, than older control mice, which spent the same amount of time exploring regardless of the concentration. This suggests that exposure to youthful blood allows older mice to better distinguish between odorant concentrations.

To address their original question, Katsimpardi created 3D reconstructions of blood vessels showing that although aging triggers a decrease in the volume of blood vessels, being joined to a young mouse reverses this decrease and also increases blood vessel branching by 21%. Additionally, using MRI, they found that cerebral blood flow (known to decrease with aging) increased in older mice to levels seen in young mice.

“We think that because the blood vessels rejuvenate in this system, the neurogenic niche rejuvenates as well,” said Katsimpardi.

Was GDF11 responsible? In collaboration with Amy Wagers and Richard Lee, the team injected older mice daily with recombinant GDF11, and found that the blood vessel volume increased by 50% in treated mice and certain neural stem cells increased by 29% (but not as much as those triggered by heterochronic parabiosis).

Another study published in May in Nature Medicine (4) used heterochronic parabiosis to examine genomic changes in older animals exposed to young blood. Saul Villeda at the University of California San Francisco found transcriptional changes related to synaptic plasticity in the hippocampus of older mice exposed to young blood, an increase in dendritic spine density of mature neurons and improved synaptic plasticity in the hippocampus. Functionally, Villeda and his colleagues found that these older mice had better spatial learning and memory, as well as contextual fear conditioning. They didn’t find neurogenesis to be involved; thus the combination of these two brain studies may mean that both improved neurogenesis and synaptic plasticity play a role in the improvements seen in older mice exposed to young blood.

Pump You Up

Nestled within skeletal muscle fibers are specialized sets of mononuclear stem cells called satellite cells. Satellite cells replace damaged muscle fibers with new cells, keeping muscles working smoothly. As muscles age, they harbor fewer and fewer satellite cells, and the ones that remain don’t regenerate muscle cells as well. Wagers and her team wanted to see if young blood could rejuvenate old satellite cells. In a May Science paper (5), they sorted out the satellite cells in aged mice, using fluorescence-activated cell sorting, and found that DNA in aged cells was far more damaged than in young cells. Wagers joined young mice (2 months old) with aged mice (22 months old), and the results were profound: satellite cells from older mice joined to young mice were better able to form myogenic colonies, their genomic integrity was restored, and their DNA looked similar to that of young mice.

They also found less GDF11 in the muscles of the older mice, so they supplemented it with daily intraperitoneal injections of recombinant GDF11 (rGDF11). After four weeks, the treated muscles had more satellite cells and also more satellite cells with intact DNA compared to old control mice.

To test whether these new cells worked as well as young cells, the team injured some of the treated aged mice. They found that the mice who had received rGDF11 had more youthful-looking muscles: the mean size of the regenerating myofibers was 92% of those in young control mice. The rGDF11 also helped satellite cells regenerate: the treated aged animals harbored almost twice as many engrafted fibers as controls, and the fibers were also larger. Immunofluorescence showed that neuromuscular junctions in these animals were larger. Electron microscopy of uninjured, treated muscle showed large improvements in mitochondrial and myofibrillar morphology, reduced atypical and swollen mitochondria, and restored sarcomeric and interfibrillar mitochondrial patterning.

Wagers and her team then found that these changes do, indeed, lead to stronger mice. The aged mice treated with rGDF11 enjoyed an increased average exercise endurance (57 minutes versus 35 minutes), as well as improved clearance of systemic lactate and lower levels of glucose after 40 minutes of running, demonstrating improved mitochondrial function. The treated older mice also had a stronger grip.

The Fountain of Youth?

All this begs the question: “If GDF11 is so great, why does it go away?” asked Wagers. “We don’t know the answer, and the answer is critical; it’s a key to understanding why this whole process may be coordinated and why these effects may emerge.”

It could be that GDF11 declines because the cells that make it start making less or disappear altogether. Or perhaps cells that make GDF11 also make other proteins that become harmful with age, so those cells are tamped down.

“We are enthusiastic that it’s going to be a conserved response in mice and humans,” said Wagers. Wagers’ group is studying how the protein changes with age, especially in age-related disease, and whether different levels of the protein could predict health outcomes.

Perhaps someday GDF11 could be used therapeutically to reverse age-related disease, either by injection of the protein itself or as an orally-available small molecule that could trigger the body’s normal production of the protein. But the researchers caution that GDF11 won’t be a fountain of youth.

“First we should know more about the protein and how it works, and then the importance would be to treat people with diseases, such as neurodegenerative diseases,” said Lida Katsimpardi.

“The goal is not to extend lifespan,” said Wagers. “The goal is to extend the healthy years of life.”



References

1. Conboy IM et al. Rejuvenation of aged progenitor cells by exposure to a young systemic environment. Nature. 17 February 2005; 433:760-764.

2. Loffredo FS et al. Growth Differentiation Factor 11 is a circulating factor that reverses age-related cardiac hypertrophy. Cell. 9 May 2013; 153: 828-839.

3. Katsimpardi L et al. Vascular and neurogenic rejuvenation of the aging mouse brain by young systemic factors. Science. 9 May 2014; 344: 630-634.

4. Villeda SA et al. Young blood reverses age-related impairments in cognitive function and synaptic plasticity in mice. Nature Medicine. AOP.

5. Sinha M et al. Restoring systemic GDF11 levels reverses age-related dysfunction in mouse skeletal muscle. Science. 9 May 2014; 344: 649-652.


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