The First International Mini-Symposium on Methionine Restriction and Lifespan

It has been 20 years since the Orentreich Foundation for the Advancement of Science, under the leadership Dr. Norman Orentreich, first reported that low methionine (Met) ingestion by rats extends lifespan (Orentreich et al., 1993). Since then, several studies have replicated the effects of dietary methionine restricted (MR) in delaying age-related diseases (Richie et al., 1994; Miller et al., 2005; Ables et al., 2012; Sanchez-Roman and Barja, 2013). We report the abstracts from the First International Mini-Symposium on Methionine Restriction and Lifespan held in Tarrytown, NY, September 2013. The goals were (1) to gather researchers with an interest in MR and lifespan, (2) to exchange knowledge, (3) to generate ideas for future investigations, and (4) to strengthen relationships within this community. The presentations highlighted the importance of research on cysteine, growth hormone (GH), and ATF4 in the paradigm of aging. In addition, the effects of dietary restriction or MR in the kidneys, liver, bones, and the adipose tissue were discussed. The symposium also emphasized the value of other species, e.g., the naked mole rat, Brandt's bat, and Drosophila, in aging research. Overall, the symposium consolidated scientists with similar research interests and provided opportunities to conduct future collaborative studies (Figure 3)

Update on Ppar and Nonalcoholic Fatty Liver Disease

Nonalcoholic fatty liver disease (NAFLD) is the most common initial presentation of obesity and insulin resistance. Uninterrupted progression of hepatic lipid accumulation often leads to fatty liver disease and eventually cirrhosis. Insulin resistance is one of the characteristics of type 2 diabetes. Several types of treatment have been employed against type 2 diabetes some of which ameliorate NAFLD. The frequent line of treatment to improve insulin sensitivity is the use of thiazolidinediones (TZD) which activate the nuclear receptor, peroxisome proliferator activated receptor gamma (Pparγ). Although TZDs are proven to be very effective in promoting insulin sensitivity, its actions on Pparγ have been complicated, specifically on NAFLD. According to studies in different models, Pparγ manifests both beneficial and undesirable effects on NAFLD. This paper will focus on the current knowledge of Pparγ and its effect on NAFLD

Physical activity of mice on dietary sulfur amino acid restriction is influenced by age of diet initiation and biological sex

Abstract Please use the attached pdf file for corrections. Sulfur amino acid restriction (SAAR)—the reduction of methionine and cysteine concentrations either in the diet or by genetic manipulation—promotes health span and extends lifespan, but its effects on physical activity remain unclear. We investigated whether age of diet initiation and biological sex could influence physical activity in mice fed either a control diet (CF, 0.86% methionine w/w) or SAAR (0.12% methionine w/w). Quadriceps femoris muscle mass is smaller in SAAR than in CF mice. Young mice fed a chronic SAAR diet at 8 weeks of age exhibited improved wire hang and running wheel activities compared to young CF mice, while aged mice showed comparable results. The effects of chronic SAAR on physical activity was mildly influenced by sex as observed in middle-aged male SAAR mice who showed minor improvements than CF males while middle-aged females displayed no discernible effects. Muscle mass is minimally affected by changes in markers of protein synthesis, autophagy and atrophy. Improvements to physical activity in young SAAR mice could be partially attributed to increased skeletal muscle mitochondrial activity. Furthermore, SAAR in C2C12 myotubes increased citrate synthase protein expression and enhanced succinyl dehydrogenase enzyme activity compared to CF myotubes. Overall, our data reveal that SAAR can improve mouse physical activity without compromising muscle proteostasis. This is partially due to enhanced mitochondrial activity, but the effects are influenced by age of diet initiation and sex

Methionine restriction alters bone morphology and affects osteoblast differentiation

Methionine restriction (MR) extends the lifespan of a wide variety of species, including rodents, drosophila, nematodes, and yeasts. MR has also been demonstrated to affect the overall growth of mice and rats. The objective of this study was to evaluate the effect of MR on bone structure in young and aged male and female C57BL/6J mice. This study indicated that MR affected the growth rates of males and young females, but not aged females. MR reduced volumetric bone mass density (vBMD) and bone mineral content (BMC), while bone microarchitecture parameters were decreased in males and young females, but not in aged females compared to control-fed (CF) mice. However, when adjusted for bodyweight, the effect of MR in reducing vBMD, BMC and microarchitecture measurements was either attenuated or reversed suggesting that the smaller bones in MR mice is appropriate for its body size. In addition, CF and MR mice had similar intrinsic strength properties as measured by nanoindentation. Plasma biomarkers suggested that the low bone mass in MR mice could be due to increased collagen degradation, which may be influenced by leptin, IGF-1, adiponectin and FGF21 hormone levels. Mouse preosteoblast cell line cultured under low sulfur amino acid growth media attenuated gene expression levels of Col1al, Runx2, Bglap, Alpl and Spp1 suggesting delayed collagen formation and bone differentiation. Collectively, our studies revealed that MR altered bone morphology which could be mediated by delays in osteoblast differentiation. Keywords: Methionine restriction, Aged mice, Micro-computed tomography, Nanoindentation, MC3T3-E1 subclone

Lean Phenotype Induced by Methionine Restriction in Mice is Associated with Proteomic Changes in Perivascular Adipose Tissue

Perivascular adipose tissue (PVAT) surrounds blood vessels, is thermogenic, and secretes adipokines that can be vasoprotective. During aging and obesity, PVAT loses its thermogenic capacity, shows increased inflammation, and secretes proatherogenic cytokines. Our collaborative research has previously shown that dietary methionine restriction (MetR) in mice can decrease adiposity, improve glucose sensitivity, and induce PVAT to become more thermogenic. To identify initiating signals from PVAT that correspond to thermogenic conversion, we challenged male C57BL/6 mice with a high fat diet (HFD, 60 kcal% fat, 0.86% methionine) until they weighed \u3e45g (~12 weeks of age). Weight matched mice were then either maintained on this diet or switched to a high fat, methionine restricted diet (60 kcal% fat, 0.12% methionine). We collected PVAT after 3, 5, or 10 days, and analyzed tissues by mass spectrometry using Sequential Window Acquisition of all Theoretical Spectra (SWATH) to identify proteomic signatures. When compared to control, PVAT from methionine restricted conditions exhibited significant differences (p\u3c0.05) in 624, 313, and 502 proteins on days 3, 5, and 10 respectively. Overall, we observed a significant increase in proteins involved in mitochondrial network formation, such as GRP75, a heat-shock protein that our previous data indicate may be a potentially novel thermogenic marker in PVAT. Our study identifies unique protein signatures that are responsive to dietary methionine concentration and are associated with a lean phenotype, despite a HFD. These profiles can be used to identify pathways that may initiate the thermogenic phenotype in PVAT, which is predicted to suppress cardiovascular disease

Methionine-Restricted C57BL/6J Mice Are Resistant to Diet-Induced Obesity and Insulin Resistance but Have Low Bone Density

<div><p>Dietary methionine restriction (MR) extends lifespan, an effect associated with reduction of body weight gain, and improvement of insulin sensitivity in mice and rats as a result of metabolic adaptations in liver, adipose tissue and skeletal muscle. To test whether MR confers resistance to adiposity and insulin resistance, C57BL/6J mice were fed a high fat diet (HFD) containing either 0.86% methionine (control fed; CF) or 0.12% methionine (methionine-restricted; MR). MR mice on HFD had lower body weight gain despite increased food intake and absorption efficiency compared to their CF counterparts. MR mice on HFD were more glucose tolerant and insulin sensitive with reduced accumulation of hepatic triglycerides. In plasma, MR mice on HFD had higher levels of adiponectin and FGF21 while leptin and IGF-1 levels were reduced. Hepatic gene expression showed the downregulation of <em>Scd1</em> while <em>Pparg</em>, <em>Atgl</em>, <em>Cd36</em>, <em>Jak2</em> and <em>Fgf21</em> were upregulated in MR mice on HFD. Restriction of growth rate in MR mice on HFD was also associated with lower bone mass and increased plasma levels of the collagen degradation marker C-terminal telopeptide of type 1 collagen (CTX-1). It is concluded that MR mice on HFD are metabolically healthy compared to CF mice on HFD but have decreased bone mass. These effects could be associated with the observed increase in FGF21 levels.</p> </div

Hepatic gene expression of MR mice on HFD corresponded with improved insulin sensitivity.

<p> (<b>A</b>) Hepatic gene expression analysis by Taqman qPCR using β-actin as the housekeeping gene. All data are expressed as the mean ± SD (n = 7–8 mice per feeding group) and analyzed by Student’s unpaired <i>t</i>-test. *p<0.05, ***p<0.001.</p

Weight Loss and Concomitant Adipose Autophagy in Methionine-Restricted Obese Mice is Not Dependent on Adiponectin or FGF21.

OBJECTIVE: Identifying novel approaches to combat obesity is important to improve health span. It was hypothesized that methionine restriction (MR) will induce weight loss in obese mice by reducing adipose tissue mass caused by increased energy expenditure and reprogramming of adipose tissue homeostasis. The roles of adiponectin (ADIPOQ) and fibroblast growth factor 21 (FGF21) during weight loss in MR mice were also tested. METHODS: Diet-induced obese (DIO) male C57BL/6J (wild type), Adipoq-deficient (Adipoq knockout [KO]), Fgf21-KO, and Adipoq-Fgf21 double-KO mice were used. Following a switch to high-fat control (DIO-CF, 60% fat/0.86% methionine) or MR (DIO-MR, 60% fat/0.12% methionine) diet, physiological parameters were measured, and inguinal and perigonadal adipose tissues were examined. RESULTS: Obese mice subjected to MR showed loss of body weight and adiposity, increased energy expenditure, and improved glucose tolerance that were independent of the actions of ADIPOQ and FGF21. MR induced reduction of circulating lipids, glucose, insulin, leptin, and insulin like growth factor 1 and increased β-hydroxybutyrate, ADIPOQ, and FGF21 concentrations. In fat, MR upregulated protein levels of adipose triglyceride lipase, apoptosis-inducing factor, lysosomal-associated membrane proteins 1 and 2, autophagy-related protein 5, beclin-1, and light chain 3B I and II. CONCLUSIONS: MR reduction of adipose tissue mass in obese mice is associated with elevated lipolysis, apoptosis, and autophagy and occurs independently of the actions of ADIPOQ and FGF21

Diet composition of HFD-fed CF and MR mice.

<p>High fat diets were purchased from Research Diets, Inc., New Brunswick, NJ. Control-fed (CF) on HFD catalog number: A11051306 and methionine-restricted (MR) on HFD catalog number: A11051305. Numbers in parenthesis are levels of DL-methionine and L-glutamic acid in the CF diet.</p

Blood biochemistry of HFD-fed CF and MR mice.

<p>Eight weeks old C57BL/6J mice were weight-matched and fed control fed (CF) on HFD (n = 7–8) and methionine-restricted (MR) on HFD (n = 7–8) diets for 14 weeks. Data are expressed as means ± SD and compared using Student’s unpaired <i>t</i>-test.</p>*<p>p<0.05,</p>**<p>p<0.01,</p>***<p>p<0.001.</p