Palmitic acid triggers inflammatory responses in N42 cultured hypothalamic cells partially via ceramide synthesis but not via TLR4.

A high-fat diet induces hypothalamic inflammation in rodents which, in turn, contributes to the development of obesity by eliciting both insulin and leptin resistance. However, the mechanism by which long-chain saturated fatty acids trigger inflammation is still contentious. To elucidate this mechanism, the effect of fatty acids on the expression of the pro-inflammatory cytokines IL-6 and TNFα was investigated in the mHypoE-N42 hypothalamic cell line (N42). N42 cells were treated with lauric acid (LA) and palmitic acid (PA). PA challenge was carried out in the presence of either a TLR4 inhibitor, a ceramide synthesis inhibitor (L-cycloserine), oleic acid (OA) or eicosapentaenoic acid (EPA). Intracellular ceramide accumulation was quantified using LC-ESI-MS/MS. PA but not LA upregulated IL-6 and TNFα. L-cycloserine, OA and EPA all counteracted PA-induced intracellular ceramide accumulation leading to a downregulation of IL-6 and TNFα. However, a TLR4 inhibitor failed to inhibit PA-induced upregulation of pro-inflammatory cytokines.In conclusion, PA induced the expression of IL-6 and TNFα in N42 neuronal cells independently of TLR4 but, partially, via ceramide synthesis with OA and EPA being anti-inflammatory by decreasing PA-induced intracellular ceramide build-up. Thus, ceramide accumulation represents one on the mechanisms by which PA induces inflammation in neurons.

Early and reversible changes to the hippocampal proteome in mice on a high-fat diet.

BACKGROUND: The rise in global obesity makes it crucial to understand how diet drives obesity-related health conditions, such as premature cognitive decline and Alzheimer's disease (AD). In AD hippocampal-dependent episodic memory is one of the first types of memory to be impaired. Previous studies have shown that in mice fed a high-fat diet (HFD) episodic memory is rapidly but reversibly impaired. METHODS: In this study we use hippocampal proteomics to investigate the effects of HFD in the hippocampus. Mice were fed either a low-fat diet (LFD) or HFD containing either 10% or 60% (Kcal) from fat for 3 days, 1 week or 2 weeks. One group of mice were fed the HFD for 1 week and then returned to the LFD for a further week. Primary hippocampal cultures were challenged with palmitic acid (PA), the most common long-chain saturated FA in the Western diet, and with the anti-inflammatory, n-3 polyunsaturated FA, docosahexaenoic acid (DHA), or a combination of the two to ascertain effects of these fatty acids on dendritic structure. RESULTS: HFD-induced changes occur in hippocampal proteins involved in metabolism, inflammation, cell stress, cell signalling, and the cytoskeleton after 3 days, 1 week and 2 weeks of HFD. Replacement of the HFD after 1 week by a low-fat diet (LFD) for a further week resulted in partial recovery of the hippocampal proteome. Microtubule-associated protein 2 (MAP2), one of the earliest proteins changed, was used to investigate the impact of fatty acids (FAs) on hippocampal neuronal morphology. PA challenge resulted in shorter and less arborised dendrites while DHA had no effect when applied alone but counteracted the effects of PA when FAs were used in combination. Dendritic morphology recovered when PA was removed from the cell culture media. CONCLUSION: This study provides evidence for the rapid and reversible effects of diet on the hippocampal proteome and the impact of PA and DHA on dendritic structure.

A high-fat diet induces rapid changes in the mouse hypothalamic proteome.

BACKGROUND: Prolonged over-consumption of a high-fat diet (HFD) commonly leads to obesity and insulin resistance. However, even 3 days of HFD consumption has been linked to inflammation within the key homeostatic brain region, the hypothalamus. METHODS: Mice were fed either a low-fat diet (LFD) or HFD containing 10% or 60% (Kcal) respectively from fat for 3 days. Mice were weighed, food intake measured and glucose tolerance calculated using intraperitoneal glucose tolerance tests (IPGTT). Proteomic analysis was carried out to determine if hypothalamic proteins were changed by a HFD. The direct effects of dietary fatty acids on mitochondrial morphology and on one of the proteins most changed by a HFD, dihydropyrimidinase-related protein 2 (DRP-2) a microtubule-associated protein which regulates microtubule dynamics, were also tested in mHypoE-N42 (N42) neuronal cells challenged with palmitic acid (PA) and oleic acid (OA). RESULTS: Mice on the HFD, as expected, showed increased adiposity and glucose intolerance. Hypothalamic proteomic analysis revealed changes in 104 spots after 3 days on HFD, which, when identified by LC/MS/MS, were found to represent 78 proteins mainly associated with cytoskeleton and synaptic plasticity, stress response, glucose metabolism and mitochondrial function. Over half of the changed proteins have also been reported to be changed in neurodegenerative conditions such as Alzheimer's disease. Also,in N42 neurons mitochondrial morphology and DRP-2 levels were altered by PA but not by OA. CONCLUSION: These results demonstrate that within 3 days, there is a relatively large effect of HFD on the hypothalamic proteome indicative of cellular stress, altered synaptic plasticity and mitochondrial function, but not inflammation. Changes in N42 cells show an effect of PA but not OA on DRP-2 and on mitochondrial morphology indicating that long-chain saturated fatty acids damage neuronal function.

Rapid and reversible impairment of episodic memory by a high-fat diet in mice.

Alzheimer's disease is a leading cause of morbidity and mortality with no cure and only limited treatment available. Obesity and type 2 diabetes are positively associated with the development of premature cognitive decline and Alzheimer's disease, linking diet with these conditions. Here we demonstrate that in mice episodic memory, together with spatial and contextual associative memory, is compromised after only one day of high-fat diet. However, object memory remains intact. This shows not only a more rapid effect than previously reported but also that more complex memories are at higher risk of being compromised by a high-fat diet. In addition, we show that these memory deficits are rapidly reversed by switching mice from a high-fat diet back to a low-fat diet. These findings have important implications for the contribution of nutrition to the development of cognitive decline and Alzheimer's disease.

SerpinA3N is a novel hypothalamic gene upregulated by a high-fat diet and leptin in mice.

BACKGROUND: Energy homeostasis is regulated by the hypothalamus but fails when animals are fed a high-fat diet (HFD), and leptin insensitivity and obesity develops. To elucidate the possible mechanisms underlying these effects, a microarray-based transcriptomics approach was used to identify novel genes regulated by HFD and leptin in the mouse hypothalamus. RESULTS: Mouse global array data identified serpinA3N as a novel gene highly upregulated by both a HFD and leptin challenge. In situ hybridisation showed serpinA3N expression upregulation by HFD and leptin in all major hypothalamic nuclei in agreement with transcriptomic gene expression data. Immunohistochemistry and studies in the hypothalamic clonal neuronal cell line, mHypoE-N42 (N42), confirmed that alpha 1-antichymotrypsin (α1AC), the protein encoded by serpinA3, is localised to neurons and revealed that it is secreted into the media. SerpinA3N expression in N42 neurons is upregulated by palmitic acid and by leptin, together with IL-6 and TNFα, and all three genes are downregulated by the anti-inflammatory monounsaturated fat, oleic acid. Additionally, palmitate upregulation of serpinA3 in N42 neurons is blocked by the NFκB inhibitor, BAY11, and the upregulation of serpinA3N expression in the hypothalamus by HFD is blunted in IL-1 receptor 1 knockout (IL-1R1 -/- ) mice. CONCLUSIONS: These data demonstrate that serpinA3 expression is implicated in nutritionally mediated hypothalamic inflammation.

High Salinity Relaying to Reduce Vibrio parahaemolyticus and Vibrio vulnificus in Chesapeake Bay Oysters (Crassostrea virginica).

Cases of Vibrio infections in the United States have tripled from 1996 to 2009 and these infections are most often associated with the consumption of seafood, particularly oysters (Crassostrea virginica). Information is needed on how to reduce numbers of Vibrio parahaemolyticus and Vibrio vulnificus in bi-valve molluscan shellfish (for example, oysters). The purpose of this study was to evaluate the effectiveness of high salinity relaying or treatment in recirculating aquaculture systems (RASs) as methods to reduce the abundance of V. parahaemolyticus and V. vulnificus in oysters. For relaying field trials, oysters were collected from approved harvest waters, temperature abused outside under a tarp for 4 h, and then transferred to high (29 to 33 ppt.) and moderate (12 to 19 ppt.) salinities. For RAS treatment trial, oysters were transferred to 32 to 34 ppt. salinity at 15 °C. After 7, 14, 21, and in some instances 28 d, oysters were collected and analyzed for V. parahaemolyticus and V. vulnificus levels using multiplex real-time PCR. Initial levels of V. parahaemolyticus and V. vulnificus ranged from 3.70 to 5.64 log10 MPN/g, and were reduced by 2 to 5 logs after 21 to 28 d in high salinity water (29 to 34 ppt.). Oyster mortalities averaged 4% or less, and did not exceed 7%. Relaying of oysters to high salinity field sites or transfer to high salinity RAS tanks was more effective in reducing V. vulnificus compared with V. parahaemolyticus. These results suggest that high salinity relaying of oysters is more effective in reducing V. vulnificus than V. parahaemolyticus in the oyster species used in this study.

Dietary supplementation with conjugated linoleic acid plus n-3 polyunsaturated fatty acid increases food intake and brown adipose tissue in rats.

The effect of supplementation with 1% conjugated linoleic acid and 1% n-3 long chain polyunsaturated fatty acids (CLA/n-3) was assessed in rats. Food intake increased with no difference in body weights. White adipose tissue weights were reduced whereas brown adipose tissue and uncoupling protein-1 expression were increased. Plasma adiponectin, triglyceride and cholesterol levels were reduced while leptin, ghrelin and liver weight and lipid content were unchanged. Hypothalamic gene expression measurements revealed increased expression of orexigenic and decreased expression of anorexigenic signals. Thus, CLA/n-3 increases food intake without affecting body weight potentially through increasing BAT size and up-regulating UCP-1 in rats.