Reviewing the reviews

Here is the next in what will likely be a long series of semi-regular review roundups — links, without extensive further comment, to the reviews I found most intriguing over the past few weeks months (I went on hiatus during the winter holidays). For the previous foray into the secondary literature, see here.

Alzheimer’s:

• Threats to the Mind. Aging, Amyloid, and Hypertension, Iadecola et al.

Apoptosis & cancer:

• The genetics of the p53 pathway, apoptosis and cancer therapy, Vazquez et al.

Calorie restriction:

• Endocrine alterations in response to calorie restriction in humans, Redman & Ravussin

Diabetes:

• Diabetes and the risk of multi-system aging phenotypes: a systematic review and meta-analysis, Lu et al.

Klotho:

• The Klotho gene family, the endocrine fibroblast growth factors, Kurosu and Kuro-o

Sirtuins:

• How does SIRT1 affect metabolism, senescence and cancer?, Brooks & Gu

Stem cells:

• Stem Cells, Their Niches and the Systemic Environment: An Aging Network, Drummond-Barbossa

Telomeres:

• …and progeria: Telomere dysfunction and dyskeratosis congenita, Walne & Dokal
• …and structure: Telomere-associated proteins: cross-talk between telomere maintenance and telomere-lengthening mechanisms, De Boeck et al.

 

ER stress in Huntington’s disease

Expression of pathogenic polyQ-repeat-containing huntingtin cripples a major endoplasmic reticulum quality assurance and garbage disposal pathway, ER-associated protein degradation (ERAD). The mutant protein appears to sequester essential components of the ERAD pathway. From Duennwald & Lindquist:

Impaired ERAD and ER stress are early and specific events in polyglutamine toxicity

Protein misfolding, whether caused by aging, environmental factors, or genetic mutations, is a common basis for neurodegenerative diseases. The misfolding of proteins with abnormally long polyglutamine (polyQ) expansions causes several neurodegenerative disorders, such as Huntington’s disease (HD). Although many cellular pathways have been documented to be impaired in HD, the primary triggers of polyQ toxicity remain elusive. We report that yeast cells and neuron-like PC12 cells expressing polyQ-expanded huntingtin (htt) fragments display a surprisingly specific, immediate, and drastic defect in endoplasmic reticulum (ER)-associated degradation (ERAD). We further decipher the mechanistic basis for this defect in ERAD: the entrapment of the essential ERAD proteins Npl4, Ufd1, and p97 by polyQ-expanded htt fragments. In both yeast and mammalian neuron-like cells, overexpression of Npl4 and Ufd1 ameliorates polyQ toxicity. Our results establish that impaired ER protein homeostasis is a broad and highly conserved contributor to polyQ toxicity in yeast, in PC12 cells, and, importantly, in striatal cells expressing full-length polyQ-expanded huntingtin.

 

2008 Hillblom meeting: Morning session 2

(continued from our coverage of yesterday’s sessions: A B)

This session was devoted to presentations by postdoctoral fellows and recipients of faculty startup grants — talks tended to be more data-intensive than the network-grant talks yesterday morning. Here are some of the highlights:

• The cortex experiences significant synaptic loss in AD; Josh Trachtenberg (UCLA) has developed a method for imaging this loss as well as its repair in vivo in a mouse model of Alzheimer’s disease. This technology will be important in future studies of AD under the emerging paradigm of AD as a disease of neuronal connectivity (as opposed to cytotoxicity); more about this in yesterday’s coverage.
• Speaking of connectivity in AD: Beth Stevens (Harvard Medical School) has asked whether Alzheimer’s is caused by re-activation of a developmental mechanism of synapse elimination. She is studying the effect of astrocytes (non-neuronal cells in the brain, each of which can ensheath and contact up to 100,000 synapses) on developing synapses, and observed that the complement protein C1q — involved in opsonization and clearance of foreign bodies in the blood — is expressed at synapses during the time period in early deveopment when “pruning” of superfluous connections is taking place. (In C1q knockout mice, pruning doesn’t occur efficiently.) Stevens hypothesizes that this process may be reactivated in AD (as well as other diseases that involve synapse loss), resulting in the targeting and destruction of (desirable) synapses.
• I spoke during this session, partly about a paper about senescence-associated protein secretion that our lab has coming out in December and partly about my own studies of the relationship between micro-RNAs and cellular senescence. Oddly for an aging conference, my talk was one of only two about cancer and the only one about senescence.
• Sun Hur (UCSF) spoke about the relationship between RNA modification and the human progeroid syndrome dyskeratosis congenita. The dyskerin protein is required for covalent modifications to a number of RNAs, including the telomerase RNA (aka TERC). Hur has crystallized several protein-RNA complexes, and is using their crystallographic structures to learn about the functional importance of RNA modification in the cell.
• Ken Nakamura (UCSF) is studying Parkinson’s disease in a refreshingly original way: he has developed ways to monitor alpha-synuclein multimers in live cells, using fusion with fluorescent proteins. The pathological protein aggregates end up associating with membranes, including mitochondria — which then fragment, potentially contributing to cytotoxicity.

 

2008 Hillblom meeting: Morning session 1B

(continued from our coverage of the earlier session)

I’m going to cover just one talk from this session, from Charlie Glabe, who gave two of the more exciting talks at the last two LLHF meetings (see my review of the 2006 and 2007 meetings).

Glabe’s group (including several other PIs joined by a LLHF network grant) has been developing anti-amyloid antibodies, some of which are conformation-specific but not necessarily sequence-specific; in other words, antibodies that recognize common features of amyloid aggregates formed by many different types of protein (e.g., Aß but also alpha-synuclein, IAPP, and other peptides involved in aggregation-based diseases). These reagents will be useful in research but also potentially as therapies against multiple age-related illnesses.

Since last year, the group has been attempting to determine the structure of amyloid oligomers. Problem: amyloids don’t crystallize, so the current strategy is to form co-crystals between anti-amyloid antibodies and prefibrillar oligomers — or, failing that, crystallize the antibody alone and make inferences about the amyloid structure (which should be the ‘negative space’ of the antibody Fab fragment — assuming, of course, that the antibody doesn’t have to undergo a dramatic structural rearrangement in order to bind). Another member of the collaboration has been trying to understand the folded and unfolded states of amyloidogenic proteins, using solution-based techniques (EPR, NMR) rather than crystallography.

Another new direction in this project: studying the effect of amyloid oligomers on membrane conductance. Amyloid oligomers, which are toxic to cells, have a significant effect on the electrical properties of lipid bilayers: specifically, they increase the rate of depolarization. Novel, and this will be especially relevant to the emerging idea that AD is a disease of neuronal connectivity (i.e., interfering with membrane conductance) as well as cell toxicity.

Not a whole lot of new stuff on the therapeutic angle this time around, but you can’t win the lottery every year.

 

Sex and the aging brain

Aging is one of the most complex biological processes we know of, and the human brain the most complex biological system. Unsurprisingly, that makes figuring out how aging affects our brains – affects those processes we really care about like learning, behaviour, and memory – enormously difficult.

A whole host of gross-level neuroanatomical changes take place as we get older, but it’s unclear to what extent these can explain the cognitive deficits that characterize normal aging and diseases of age like Alzheimer’s. For example, while some parts of the hippocampus (a brain structure crucial for the formation of new memories, and linked to dementia) lose neurons as we age, other parts only grow more and more synaptic connections.

To get the complete picture, we need more detail – we need to know what is going at the level of individual genes. In recent years, two groups (Lu et al. and Erraji-Benchekroun et al.) have published microarray expression studies of the aging human brain. However, in both studies there is no control for gender – female and male brains are all lumped into one group –  and this adds substantial noise to their results. Male and female brains are known to develop differently, and even to age differently at the neuroanatomical level – for instance, men experience more (and earlier-onset) brain atrophy and a greater increase in cerebrospinal fluid.

Last month, Berchtold et al. published a gene expression study of aging in four areas of the human brain, and for the first time looked at gender differences in brain aging:

Gene expression changes in the course of normal brain aging are sexually dimorphic
Gene expression profiles were assessed in the hippocampus, entorhinal cortex, superior-frontal gyrus, and postcentral gyrus across the lifespan of 55 cognitively intact individuals aged 20–99 years. Perspectives on global gene changes that are associated with brain aging emerged, revealing two overarching concepts. First, different regions of the forebrain exhibited substantially different gene profile changes with age. For example, comparing equally powered groups, 5,029 probe sets were significantly altered with age in the superior-frontal gyrus, compared with 1,110 in the entorhinal cortex. Prominent change occurred in the sixth to seventh decades across cortical regions, suggesting that this period is a critical transition point in brain aging, particularly in males. Second, clear gender differences in brain aging were evident, suggesting that the brain undergoes sexually dimorphic changes in gene expression not only in development but also in later life. Globally across all brain regions, males showed more gene change than females. Further, Gene Ontology analysis revealed that different categories of genes were predominantly affected in males vs. females. Notably, the male brain was characterized by global decreased catabolic and anabolic capacity with aging, with down-regulated genes heavily enriched in energy production and protein synthesis/transport categories. Increased immune activation was a prominent feature of aging in both sexes, with proportionally greater activation in the female brain. These data open opportunities to explore age-dependent changes in gene expression that set the balance between neurodegeneration and compensatory mechanisms in the brain and suggest that this balance is set differently in males and females, an intriguing idea.

Before controlling for gender, Berchtold et al. examined the entire set of brain samples to see if there were any general trends across brain regions. Surprisingly, they found that the superior-frontal gyrus and the postcentral gyrus consistently showed the most aging-related changes. This is unexpected, because it’s the other two brain regions – the hippocampus and the entorhinal cortex – that are most associated with age-related brain diseases and cognitive decline.

Berchtold et al. then sorted brain samples by gender. They found that between young (20-59yrs) and old age (60-99yrs), the male brain undergoes three times as many changes in gene expression as the female brain. Also, men exhibit significantly higher levels of change in all areas save the hippocampus, where men and women experience roughly the same number of gene changes.

To get a more detailed picture, Berchtold et al. classified brains into four age groups: 20-39yrs, 40-59yrs, 60-79yrs and 80-99yrs. For men, the largest number of gene expression changes (about 5000) was observed between the age categories of 40-59yrs and 60-79yrs, and there were few changes in subsequent decades, i.e., the male brain seemed to stabilize. In contrast, the female brain showed substantially fewer changes (about 1000) between those age categories, and showed the most changes (about 3500) between later age categories 60-79yrs and 80-99yrs. The authors take this as evidence that the aging female brain never stabilizes in the way that the aging male brain does; it would be interesting to divide the final female age category into two finer categories (80-89yrs and 90-99yrs) to verify that stabilization never happens. They also point out that this trend is consistent with what we know about the incidence of dementia: dementia risk stabilizes for men around age 85, but increases for women from ages 77 to 95.

Because women and men have different life expectancies (women live on average 5-10 years longer), Berchtold et al. were concerned that the differences in aging between men and women might just be reflecting the difference in longevity – i.e., that women might show almost the same sequence of aging events as men, only drawn out over a longer scale. To test this idea, they compared lists of significantly differentially expressed genes from the most critical aging period for men and women. They found that more than 75% of gene expression changes for each sex were unique to that sex – i.e., most gene change differences can’t be ascribed to simple differences in longevity.

Finally, Berchtold et al. used Gene Ontology annotations to determine whether any functional categories of genes were significantly differentially regulated with age between young (20-59yrs) and old (60-99yrs) brains. In males (but not females), they found a general decreasing capacity for energy production with age (e.g., several relevant Gene Ontology categories were downregulated, including electron transport, oxidative phosphorylation, ATP metabolism, mitochondrial transport, etc.). In females (but not males), categories for neuronal morphogenesis and intracellular signalling were significantly downregulated. In both sexes, genes associated with synaptic transmission were downregulated, and genes associated with cell death and angiogenesis were upregulated. Interestingly, for both sexes many genes associated with inflammation and the immune system were upregulated (while a bit of inflammation can be neuroprotective, too much is undesirable, and a general feature of aging in many tissues).

…….So what do all these results mean? Can we immediately conclude that male brains change for the worse at a relatively young age – but then stabilize – and that female brains just keep on getting worse with increasing age? While evolutionary theory predicts that many of the observed gene expression changes are deleterious, others could be adaptive damage-control responses, beneficial, or even just plain neutral. Also, to interpret this data properly, we need to be able to disentangle cause from effect – some gene expression changes matter a lot more than others. Berchtold et al. have given us a wonderful resource to study how aging affects the brain at the gene level, but we still need to do a lot of work before we can connect this vast catalogue of gene expression values to the higher-level biological and cognitive phenomena that we are most interested in.

 

Turning back the CLK on neurodegeneration

Mutations in the CLK-1 gene, which is involved in the synthesis of coenzyme Q (ubiquinone), slow down aging in both worms and mice. The gene’s mechanism of action has been murky: Deficiency in the gene leads to a dietary dependence on ubiquinone and accumulation of a precursor molecule, DMQ, but neither the high levels of DMQ nor a shortage of Q is responsible for the physiological changes observed in CLK-1^-/- mutants — therefore, it’s possible that the lifespan function of CLK-1 protein is unrelated to its role in co-Q biosynthesis.

Wang et al. have characterized the wide-spectrum anti-neurodegeneration drug cliquinol, and discovered that it downregulates the activity of the CLK-1 enzyme. The data suggests that heavy metal cations are important for the protein’s longevity-regulation function:

The development of neurodegenerative diseases such as Alzheimer’s, Parkinson’s, and Huntington’s disease is strongly age-dependent. Discovering drugs that act on the high rate of aging in older individuals could be a means of combating these diseases. Reduction of the activity of the mitochondrial enzyme CLK-1 (also known as COQ7) slows down aging in C. elegans and in mice. Clioquinol is a metal chelator that has beneficial effects in several cellular and animal models of neurodegenerative diseases as well as on Alzheimer’s disease patients. Here we show that clioquinol inhibits the activity of mammalian CLK-1 in cultured cells, an inhibition that can be blocked by iron or cobalt cations, suggesting that chelation is involved in the mechanism of action of clioquinol on CLK-1. We also show that treatment of nematodes and mice with CQ mimics a variety of phenotypes produced by mutational reduction of CLK-1 activity in these organisms. These results suggest that the surprising action of clioquinol on several age-dependent neurodegenerative diseases with distinct etiologies might result from a slowing down of the aging process through action of the drug on CLK-1. Our findings support the hypothesis that pharmacologically targeting aging-associated proteins could help relieve age-dependent diseases.

Note the underlined section (emphasis mine) — this reminds me of a speculation I offered up a couple of years ago: Could coenzyme Q supplementation, which has a variety of health benefits in mammals, act by end-product inhibition of the ubiquinone synthesis pathway? In other words, high levels of the pathway’s ultimate product could be shutting down the both the enzymatic and lifespan-regulatory function of CLK-1, resulting in a phenocopy of the CLK-1 loss-of-function mutation. Here, the authors argue that direct enzymatic inhibition of the pathway might be preventing neurodegeneration primarily by slowing down the aging process.

Of course, it’s also possible that the effect on CLK-1 is an epiphenomenon and that heavy metal cations cause neurodegeneration directly, so that chelating them is a generally good thing totally unrelated to the drug’s effect on ubiquinone synthesis. (If that were true, then we would predict that clioquinone would further prevent the delayed neurodegeneration that eventually occurs in CLK-1 mutants as well.)

 

SIRT1 in metabolism, signaling and disease

Racing toward its ultimate goal of being involved in every aspect of biology, the mammalian sirtuin SIRT1 has been the subject of a number of recent papers, each dealing with a different aspect of the protein’s role. (Abstracts are excerpted; ellipses, emphases, and interpolated commentary are mine.)

In energy metabolism and liver cirrhosis: Sirt1 is involved in energy metabolism: The role of chronic ethanol feeding and resveratrol, Oliva et al.:

These results support the concept that ethanol induces the Sirt1/PGC1α pathway of gene regulation and both naringin and resveratrol prevent the activation of this pathway by ethanol. However, resveratrol did not reduce the liver pathology caused by chronic ethanol feeding [In other words, it’s probably not a good idea to get your resveratrol by drinking 1000 bottles of red wine a day.]

In diseases of protein aggregation: The role of calorie restriction and SIRT1 in prion-mediated neurodegeneration, Chen et al. [a collaboration between the Lindquist and Guarente labs]:

We tested the role of SIRT1 in mediating the effects of CR in a mouse model of prion disease. … We report that the onset of prion disease is delayed by CR and in the SIRT1 KO mice fed ad libitum. CR exerts no further effect on the SIRT1 KO strain, suggesting the effects of CR and SIRT1 deletion are mechanistically coupled. In conjunction, SIRT1 is downregulated in certain brain regions of CR mice. … Surprisingly, CR greatly shortens the duration of clinical symptoms of prion disease and ultimately shortens lifespan of prion-inoculated mice in a manner that is independent of SIRT1. [i.e., CR isn’t actually therapeutically beneficial since the mice die young.]

In inflammation, inflammaging, and HIV/AIDS: SIRT1 longevity factor suppresses NFκB -driven immune responses: regulation of aging via NFκB acetylation?, Salminen et al. (review):

HIV-1 Tat protein binds to SIRT1 protein, a well-known longevity factor, and inhibits the SIRT1-mediated deacetylation of the p65 component of the NFκB complex. As a consequence, the transactivation efficiency of the NFκB factor was greatly potentiated, leading to the activation of immune system and later to the decline of adaptive immunity. … Longevity factors, such as SIRT1 and its activators, might regulate the efficiency of the NFκB signaling, the major outcome of which is inflamm-aging via proinflammatory responses.

In Notch regulation of stem cell aging: Sirt1, Notch and stem cell “age asymmetry”, Mantel et al. (review):

The protein-deacetylase, SIRT1, has received much attention because of its roles in oxygen metabolism, cellular stress response, aging, and has been investigated in various species and cell types including embryonic stem cells. However, there is a dearth of information on SIRT1 in adult stem cells, which have a pivotal role in adult aging processes. Here, we discuss the potential relationships between SIRT1 and the surface receptor protein, Notch, with stem cell self-renewal, asymmetric cell division, signaling, and stem cell aging.