The Effect of a Western Diet on Hepatic Autophagy in Age Accelerated SAMP8 Mice

Non-alcoholic steatohepatitis (NASH) is characterized as a dysregulation of hepatic lipid metabolism and a chronic inflammatory state. It is hypothesized the link between lipid dysregulation and inflammation may be due in part to defective hepatic autophagy and reduced mitochondrial capacity to oxidize fatty acids. It remains to be determined; however, the effects of a Western diet on hepatic autophagy and mitochondrial function during aging. PURPOSE: The purpose of this study was to determine the effect of a high-fat high fructose diet (HFF) on markers of hepatic autophagy and mitochondrial function in an age accelerated mouse model. METHODS: Twenty week old, male and female, SAMP8 mice (n=49) were randomly assigned, matching for gender, to either a standard chow (SC) or HFF (45% fat, 24% fructose) diet for 32 weeks. Liver tissue was analyzed for mRNA expression of autophagic (BNIP3, Beclin 1, p62, and Atg7) and mitochondrial (PGC1α and COXIV) genes. Differences between gender and dietary groups were identified by a 2 x 2 ANOVA and statistical significance was set at p\u3c0.05. RESULTS: Following 32 weeks of feeding, male mice fed the HFF diet were significantly heavier than male mice in the SC group (31.6 g vs 26.5 g; p=0.001); however, no difference was observed between diet groups for female mice. The HFF diet resulted in higher autophagic activity as observed by Beclin 1 (+36%; p=0.001) and BNIP3 (+40%; P=0.003) expression. Despite the higher autophagic activity, p62 was higher (+31%; p\u3c0.001) in the HFF compared to the SC group, suggesting impaired autophagic flux. In addition, mitochondrial COXIV expression was elevated (+43%; P\u3c0.001) in the HFF group compared to the SC group suggesting increased β-oxidation. Overall, the expression of all autophagic and mitochondrial markers was higher in male compared to female mice; however, both sexes responded similarly to the HFF diet. CONCLUSION: Despite the higher expression of autophagic and mitochondrial genes, elevated expression of p62 suggests an impaired autophagic flux in age accelerated mice following a Western diet

The importance of comparative phylogeography in diagnosing introduced species: a lesson from the seal salamander, Desmognathus monticola

<p>Abstract</p> <p>Background</p> <p>In most regions of the world human influences on the distribution of flora and fauna predate complete biotic surveys. In some cases this challenges our ability to discriminate native from introduced species. This distinction is particularly critical for isolated populations, because relicts of native species may need to be conserved, whereas introduced species may require immediate eradication. Recently an isolated population of seal salamanders, <it>Desmognathus monticola</it>, was discovered on the Ozark Plateau, ~700 km west of its broad continuous distribution in the Appalachian Mountains of eastern North America. Using Nested Clade Analysis (NCA) we test whether the Ozark isolate results from population fragmentation (a natural relict) or long distance dispersal (a human-mediated introduction).</p> <p>Results</p> <p>Despite its broad distribution in the Appalachian Mountains, the primary haplotype diversity of <it>D. monticola </it>is restricted to less than 2.5% of the distribution in the extreme southern Appalachians, where genetic diversity is high for other co-distributed species. By intensively sampling this genetically diverse region we located haplotypes identical to the Ozark isolate. Nested Clade Analysis supports the hypothesis that the Ozark population was introduced, but it was necessary to include haplotypes that are less than or equal to 0.733% divergent from the Ozark population in order to arrive at this conclusion. These critical haplotypes only occur in < 1.2% of the native distribution and NCA excluding them suggest that the Ozark population is a natural relict.</p> <p>Conclusion</p> <p>Our analyses suggest that the isolated population of <it>D. monticola </it>from the Ozarks is not native to the region and may need to be extirpated rather than conserved, particularly because of its potential negative impacts on endemic Ozark stream salamander communities. Diagnosing a species as introduced may require locating nearly identical haplotypes in the known native distribution, which may be a major undertaking. Our study demonstrates the importance of considering comparative phylogeographic information for locating critical haplotypes when distinguishing native from introduced species.</p

Dietary Enrichment of Fish-Oils Attenuates Diet-Induced Obesity and Hepatic Steatosis

Non-alcoholic fatty liver disease (NAFLD) is characterized by the accumulation of excess hepatic fat, exceeding 5% of total liver mass. NAFLD is present in one-third of Americans and up to 90% in those who are obese. NAFLD develops largely in part to consumption of a Western diet, defined as 40-60% kcal from saturated fats; however, a diet rich in fish-oils may prevent and reverse the development of steatosis. PURPOSE: To determine the effects of fish oils on the development of NAFLD. METHODS: C57BL/6 (n=91) mice were randomly assigned to four dietary groups for 32-weeks: 10% lard (LFL), 10% fish-oil (LFFO), 41% lard (HFL), or 41% fish-oil (HFFO) diet. Significant differences (p\u3c0.05) between groups were identified by a one-way ANOVA. RESULTS: When compared to HFFO, mice in the HFL group saw an greater (Table 1) body mass and net glucose AUC by 13% (p\u3c0.001) and 24% (p=0.08), respectively. No significant difference was observed between LFL and LFFO for body mass, net glucose AUC or HOMA-IR. This is interesting given no significant difference was observed between groups for the mean weekly caloric intake. HFFO mice showed an 86% lower (p\u3c0.001) total hepatic lipid and 4.8-fold lower (p\u3c0.001) hepatic triglyceride concentration when compared to HFL. HFFO mice also saw a 32% lower (p\u3c0.001) total hepatic cholesterol when compared to HFL. There was no significant difference in total hepatic lipids between LFL and LFFO. CONCLUSION: Despite for no significant difference in caloric intake between high-fat diet groups, consumption of a high-fat diet rich in fish-oils prevented dietary induced obesity, insulin resistance and hepatic steatosis. These results suggest that a diet rich in fish-oils have preventative effects on the development of NAFLD

Fish-oils Increase BAMBI Expression to Protect Against Fibrotic Activity in LPS Stimulated Hepatic Tissue

Non-alcoholic steatohepatitis (NASH), defined as excess hepatic lipid and chronic inflammation, provides an environment prone for the development of hepatic fibrosis. Recent evidence suggests that the antifibrotic protein BAMBI (BMP-Activin membrane bound inhibitor) is downregulated in the presence of inflammation, and may be central to the development of fibrosis. Diets rich in omega-3 (w-3) fatty acids are known to provide anti-inflammatory effects; however, the effects of w-3 fatty acids on hepatic fibrosis are not well-established. PURPOSE: To determine the effects of fish-oils on the hepatic fibrosis signaling cascade, following 32-weeks of high-fat feeding in a LPS-induced model of NASH. METHODS: Male C57BL/6 mice were randomly assigned to one of four diets for 32 weeks (n=9/group): low-fat lard based (LFL, 10% kcal fat), low-fat fish-oil based (LFFO, 10% kcal fat), high-fat lard based (HFL, 41% kcal fat), or high-fat fish-oil based (HFFO, 41% kcal fat). Following in situ LPS stimulation, liver mRNA expression of CD14, TLR4, MyD88, BAMBI, and TGF-β1 was quantified using quantitative RT-PCR. Differences between diets were identified using a one-way ANOVA with statistical significance set at p\u3c0.05. RESULTS: Following LPS stimulation, CD14 was increased 2.5 fold (p=0.020) in HFFO when compared to HFL. Despite the increase in CD14, TLR4 showed no difference between groups. In contrast, MyD88 was 2.8 fold greater (p\u3c0.001) in HFL compared to HFFO. In comparison to untreated tissue, BAMBI was 1.7 fold (p=0.017) higher in the HFFO LPS-stimulated tissue, which best explained the 1-fold (p=0.004) lower expression of TGF-β1 in HFFO when compared to HFL post-LPS stimulation. CONCLUSION: Despite the increase in extracellular LPS signaling receptor CD14, the consumption of fish-oils produced a protective intracellular response as observed by an increase in BAMBI and decrease in TGF-β1. These results suggest that a diet high in w-3 fatty acids may protect against the development of hepatic fibrosi

Diet and Sex Differences Induce Unique Alterations of Markers for Blood Brain Barrier Integrity in Age-Accelerated Mice

The role of diet on brain health has received significant attention, with the Western diet (WD) contributing to cerebrovascular alterations and neurodegenerative disease. The blood-brain barrier (BBB) may play a particularly important role as it forms the interface between the peripheral circulation and the central nervous system. The WD has been shown to negatively impact the BBB. Whether there are sex specific differences with diet on BBB integrity remains unclear. PURPOSE: To determine the effect of diet and sex on the mRNA expression of markers of BBB integrity in an age-accelerated mouse model. METHODS: Male and female Senescence Accelerated Mouse-Prone 8 (SAMP8) mice were randomly assigned to a standard diet (SD) or WD formula for a 32-week period, matched for sex, ending at 12-months of age (n=10-14/group). At 12-months of age, cortical brain tissue was evaluated for the expression of mRNA for targets associated with BBB integrity (Cldn-1, Cldn-3, Cldn-5, Cldn-12, F11r, Lsr, Msfd2a, Ocln, Tjp) using quantitative RT-PCR. A two-way ANOVA was used to identify whether mRNA expression of these targets differed with sex, diet, and their interaction. RESULTS: A significant (pCONCLUSION: Overall, female mice presented with higher expression of mRNA markers for BBB integrity, which may be a protective factor. Furthermore, mice fed the WD had lower mRNA expression of markers of BBB integrity suggesting that a Western diet may accelerate the pathogenesis of the disease state

Omega-3-Fatty Acids Hold Therapeutic Potential for the Prevention and Treatment of Diabetic Neuropathy

Diabetic neuropathy is a debilitating complication of diabetes, affecting over 50% of diabetic patients. Overweight humans display manifestations of diabetic neuropathy before developing overt diabetes and mice fed a high fat diet exhibit signs of neuropathy including mechanical hindpaw hypersensitivity and neuronal inflammation, suggesting fat diet-induced inflammation may play a role in the development of neuropathy. Omega-3 (n-3) fatty acids have anti-inflammatory properties and may hold therapeutic potential as a preventative treatment for prediabetic and diabetic patients at risk for neuropathy. PURPOSE: Investigate the impact of diet composition on signs of neuropathy. We hypothesized that a diet rich in n-3 fatty acids would attenuate hindpaw hypersensitivity during prolonged feeding of a high fat diet. METHODS: C57BL/6 mice were randomized into four diet groups (n = 12/group) for 32 weeks: 10% low fat-fish oil (LFFO), 41% high fat-fish oil (HFFO), 10% low fat-lard (LFL), or 41% high fat-lard (HFL). Neuropathy was characterized at baseline and every other week thereafter using the von Frey behavioral test for hindpaw mechanical sensitivity. A glucose tolerance test was performed at end study, and total area under the curve (AUC) was calculated using the trapezoidal method. RESULTS: At end study, body weight was greater in HFL compared to all other groups. Body weight was also greater in HFFO compared to LFFO. Fasting glucose and glucose AUC were higher in HFL compared to LFFO and HFFO. Following the same pattern as body weight, fasting glucose was higher in HFFO compared to LFFO. Although percent paw withdrawal was greater in HFL compared to HFFO and LFFO, there were no significant differences for LF vs. HF for fish oil or lard. CONCLUSION: A HFL diet induced signs of neuropathy including hindpaw hypersensitivity, whereas a fish oil diet was protective against hindpaw hypersensitivity. Moreover, omega-3-fatty acids may hold therapeutic potential for neuropathy prevention in nondiabetic and diabetic patients

The Gly2019Ser mutation in LRRK2 is not fully penetrant in familial Parkinson's disease: the GenePD study

<p>Abstract</p> <p>Background</p> <p>We report age-dependent penetrance estimates for leucine-rich repeat kinase 2 (<it>LRRK2</it>)-related Parkinson's disease (PD) in a large sample of familial PD. The most frequently seen <it>LRRK2 </it>mutation, Gly2019Ser (G2019S), is associated with approximately 5 to 6% of familial PD cases and 1 to 2% of idiopathic cases, making it the most common known genetic cause of PD. Studies of the penetrance of <it>LRRK2 </it>mutations have produced a wide range of estimates, possibly due to differences in study design and recruitment, including in particular differences between samples of familial PD versus sporadic PD.</p> <p>Methods</p> <p>A sample, including 903 affected and 58 unaffected members from 509 families ascertained for having two or more PD-affected members, 126 randomly ascertained PD patients and 197 controls, was screened for five different <it>LRRK2 </it>mutations. Penetrance was estimated in families of <it>LRRK2 </it>carriers with consideration of the inherent bias towards increased penetrance in a familial sample.</p> <p>Results</p> <p>Thirty-one out of 509 families with multiple cases of PD (6.1%) were found to have 58 <it>LRRK2 </it>mutation carriers (6.4%). Twenty-nine of the 31 families had G2019S mutations while two had R1441C mutations. No mutations were identified among controls or unaffected relatives of PD cases. Nine PD-affected relatives of G2019S carriers did not carry the <it>LRRK2 </it>mutation themselves. At the maximum observed age range of 90 to 94 years, the unbiased estimated penetrance was 67% for G2019S families, compared with a baseline PD risk of 17% seen in the non-<it>LRRK2</it>-related PD families.</p> <p>Conclusion</p> <p>Lifetime penetrance of <it>LRRK2 </it>estimated in the unascertained relatives of multiplex PD families is greater than that reported in studies of sporadically ascertained <it>LRRK2 </it>cases, suggesting that inherited susceptibility factors may modify the penetrance of <it>LRRK2 </it>mutations. In addition, the presence of nine PD phenocopies in the <it>LRRK2 </it>families suggests that these susceptibility factors may also increase the risk of non-<it>LRRK2</it>-related PD. No differences in penetrance were found between men and women, suggesting that the factors that influence penetrance for <it>LRRK2 </it>carriers are independent of the factors which increase PD prevalence in men.</p

ADDITIVE MANUFACTURING OF SILICON CARBIDE (SIC) CERAMIC ROCKET NOZZLES

Rocket motor nozzles are typically made of materials such as graphite and fiber-reinforced phenolics or epoxies with low erosion rates, relatively low manufacturing costs, and/or reduced weight. Additive manufacturing (AM) methods enable unique control of geometry and composition and can improve the weight and thermal performance of rocket nozzles. A newly developed AM process called vibration assisted printing (VAP) can process a unique combination of ceramic-forming polymer binders with a large amount of solid particles at very high solid loadings and viscosities. This research investigated the formulation, pre-processing and post-processing of these materials, determined their final microstructure and phase composition, and compared them to some of the materials utilized in rockets today. Some of the parameters investigated included mixture ratio, curing temperature, and sintering temperature of silicon carbide (SiC)-ceramic forming polymer particles. These mixtures were used for 3D printing of SiC-ceramic matrix/SiC composite parts. The results of the study will help improve the performance of modern-day rockets and other applications within the aerospace industry.Outstanding ThesisLieutenant, United States NavyApproved for public release. distribution is unlimite