Consumption of the HFD caused an increased lipid deposition in the liver that was not observed in HFA40 mouse liver. The NAFLD activity score was significantly higher in the liver from HF compared to C, CA, and HFA40 mice . Several proteins involved in the inflammatory response were measured in liver by Western blot . The chemokine MCP-1, the cytokine TNFα, the macrophage marker F4/80, and the enzyme NOS2 were all upregulated in the liver of HF mice. No significant differences were observed between C and CA in markers of steatosis and liver inflammation.Chronic consumption of a HFD by mice led to the development of obesity, adiposity, dyslipidemia, steatosis, liver inflammation and insulin resistance. Simultaneous consumption of a diet rich in AC, i.e. cyanidins and delphinidins, attenuated all these adverse effects. In addition, the AC-associated improvement of inflammation, oxidative stress, and insulin sensitivity was in part associated with their capacity to modulate NF-κB and JNK. Diets rich in fat and carbohydrates are in part responsible for the increasing global burden of overweight and obesity. The dietary consumption of a HFD by mice mimics the consequences of Western style diets in humans. The amount of ACs provided are comparable in quality and amount to that achievable through food consumption and/or rational amounts of dietary supplements in humans. We observed that mice eating the HFD and AC gained less weight than those fed the HFD alone, despite consuming similar amounts of calories.
In line with these results, AC at the highest amount provided, i.e. 40 mg/kg body weight,plastic growers pots led to lower weight of brown, visceral, and retroperitoneal fat pads, but not epididymal or subcutaneous fat, compared to the non-supplementedHFD-fed mice. AC-mediated decrease in visceral fat is particularly relevant given the role of this fat pad in the development of systemic adverse effects through the release of adipokines, growth factors and inflammatory molecules. Thus, visceral fat accumulation is associated with the development of metabolic syndrome and associated diseases, e.g., NAFL and cardiovascular disease. AC supplementation also attenuated the hyperlipidemia and steatosis associated with HFD consumption. These results disagree with previous reports that pure AC but not AC in berry extracts have the capacity to improve dyslipidem. These differences may be related to differences in experimental design, but stress the relevance of the overall food matrix in AC absorption and metabolism, and subsequently on their biological actions. Part of the mentioned effects on obesity and steatosis could be due to the actions of AC at the gastrointestinal tract. In this regard, factors that can contribute to the capacity of AC to mitigate body weight gain and excess tissue lipid deposition may include the modulation of GLP-1, a hormone known to reduce adiposity, and/or a decreased fat absorption associated to the inhibition of pancreatic lipase, which is essential for dietary triglyceride absorption in the intestine. With regard to the latter, both cyanidin and cyanidin-3,5- diglucoside were shown to inhibit the enzyme in vitr. Pancreatic lipase inhibition would also be consistent with the current finding of high fecal triglyceride levels in the AC-supplemented and HFD-fed mice. The increasing incidence of T2D worldwide has paralleled that of overweight and obesity. As previously reported. HFD consumption by C57BL/6J mice led to insulin resistance as evidenced by high fasted plasma glucose and insulin levels, and impaired ITT and GTT tests. At the highest concentration tested, the AC blend improved all these parameters.
Accordingly, an AC-rich blueberry extract was found to improve parameters of insulin sensitivity in HFD-fed mice, although under the tested conditions the ITT and GTT were not affected by fat consumption. The above beneficial effects could be in part related to the capacity of AC to modulate hormones that regulate different aspects of glucose homeostasis. Adipokines and incretins contribute to the regulation of satiety and/or glucose homeostasis. While plasma adiponectin levels were not affected, plasma leptin was increased because of HFD consumption, as it is observed in diet-induced obesity, The attenuation of hyperleptinemia by AC supplementation in HFD-fed mice may in part reflect a decreased fat pad mass and an improved capacity to modulate food intake and energy balance. In terms of the incretins, GIP and GLP- 1 increase insulin secretion after food consumption influencing glucose control. Furthermore, GLP-1 promotes satiety and improves basal and postprandial lipidemia. The prevention by AC of HFD-mediated GIP increase may reflect AC-mediated improved capacity to regulate glucose homeostasis. While AC supplementation did not affect HFD-mediated increase in plasma GLP-1, it increased GLP-1 plasma levels in mice fed the control diet. Consistently with the latter, delphinidin was found to increase GLP-1 secretion in GLUTag cells. The capacity of cyanidin and/or delphinidin to increase plasma GLP-1 may be an important mechanism underlying, in part, their anti-obesity and anti-T2D actions. In fact, GLP-1 is a relevant therapeutic target for the control of T2D. The liver is one of the central organs in the maintenance of glucose and lipid homeostasis. HFD consumption disrupted this homeostasis, and in parallel caused liver fat deposition and inflammation. The coordinated AC-mediated improvement of HFD-induced liver steatosis, inflammation, and systemic insulin resistance can be explained through the interplay among these adverse conditions. In terms of biochemical mechanisms, inflammation, oxidative stress and the activation of NF-κB are events that establish a self-feeding cycle that can be initiated by excess nutrient consumption.
The capacity of AC to mitigate HFD-triggered hepatic inflammation and NF-κB/JNK activation can be in part due to AC capacity to modulate liver oxidative stress. AC cannot exert direct antioxidant actions except at the gastrointestinal tract where they can reach high enough concentrations that allow to scavenge oxidants at a significant extent. On the other hand, in the liver and other organs, the regulation of oxidative stress is mainly due to indirect antioxidant actions exerted by AC, mainly their metabolites, through the modulation of the production of superoxide anion and nitric oxide. In fact, we observed that AC supplementation prevented HFD-mediated upregulation of NOX3 and NOX4, and of the pro-in- flammatory inducible nitric oxide synthase, NOS2. AC supplementation also prevented the high levels of 4-HNE-protein adducts in HFD-mouse liver. Consistently, nutrient overload can cause increased formation of 4-HNE which irreversibly form adducts with macromolecules , which can modify cell function, and contribute to T2D and NAFLD development. Inactivation of the redox sensitive transcription NF-κB can be central to AC beneficial actions. We observed that AC supplementation inhibited in the liver HFD-induced NF-κB activation, as evidenced by both decreased IKK phosphorylation, and NF-κB-DNA binding. Accordingly, we previously observed that cyanidin and delphinidin 3- O-glucosides inhibited TNFα-induced NF-κB activation and downstream loss of monolayer integrity in Caco-2 intestinal cells. Cyanidin inhibited the inflammatory response and redox imbalance triggered by TNFα in Caco-2 cells, which was attributed in part to NF-κB inhibition and Nrf2 upregulation. NF- κB and JNK pathways are key players in the development of insulin resistance. Activation of the NF-κB upstream kinase IKK and of JNK leads to the phosphorylation in serine residues of the insulin receptor substrate-1 causing a downregulation of the insulin cascade. In addition, NF-κB activation also induces the transcription of PTP1B, a tyrosine phosphatase which dephosphorylates and inactivates the insulin receptor and IRS1. Thus, disruption of the high fatdiet induced cycle of inflammation, oxidative stress and NF-κB/JNK activation can be central in the capacity of AC to mitigate HFD-induced insulin resistance. Furthermore, it can be argued that the effects of AC on insulin resistance are in part derived from an upstream regulation of oxidant levels and/or inflammation. In summary, blueberry in pot supplementation with a cyanidin and delphinidin-rich blend, mitigated the adverse consequences of HFD consumption, i.e. obesity, dyslipidemia, steatosis, and insulin resistance. Inhibition of inflammation, oxidative stress and NF-κB/JNK activation emerge as mechanisms underlying those AC-mediated benefits. In addition, AC could also act in part by inhibiting pancreatic lipase and subsequently dietary lipid absorption, and by modulating incretins involved in glucose and lipid homeostasis. Increased AC cyanidin/delphinidin consumption either through diet or by supplementation could be a plausible strategy to control the adverse effects of Western style diets.While the health benefits of fruits and vegetables are widely acknowledged, consumption of these foods among children and youth is at a low level. Fewer than 11% of school-aged children eat fruits and vegetables at the recommended levels ; as many as one-third of high school students eat vegetables less than once a day, and 28% eat fruit less than once a day .
Further, data collected by the Centers for Disease Control and Prevention’s National Health and Nutrition Examination Survey shows that the fruits and vegetables adolescents consume tend to be the less nutritious forms: Fruit juices and fried potatoes are major contributors . Children’s low consumption of fruits and vegetables has been documented in numerous studies. It is clearly addressed in the 2010 USDA Dietary Guidelines , which note that intakes of fried potatoes and fruit beverages have seen recent growth, while intakes of fresh fruits and vegetables have not.The United States is confronting an epidemic of poor nutrition among children. Schools can play an important role in addressing this epidemic, both by serving food directly to students and by using the power of role modeling to demonstrate healthy diets to students and their families. Despite educational efforts, at the population level fruit and vegetable intakes have changed very little, prompting some to suggest that alternative individual-, community- and population-level interventions are necessary . One promising approach is to provide more servings of fruits and vegetables in schools and youth-serving programs . Findings suggest that if children are provided with healthful, appealing foods, they will eat them. A European review of the literature found that availability and accessibility of fruits and vegetables and taste preferences were the determinants most consistently and positively related to consumption . Furthermore, a combination of increased access to fruits and vegetables at school with nutrition education in the curriculum has a considerably greater impact than nutrition education alone, although both are important . The USDA Fresh Fruit and Vegetable Program, which provides an extra serving of a fruit or vegetable as a between-meal snack to children at schools in low-income communities that apply for the program, is being evaluated and shows promise for increasing children’s consumption . The greatest room for improvement in children’s fruit and vegetable consumption is at school, where children consume up to half of their calories . The National Academy of Sciences Institute of Medicine has urged school action to increase fruit and vegetable intake , and federal policies resulting from the Healthy, Hunger-Free Kids Act of 2010 mandate this increase.The California Fresh Start Program was a pilot school breakfast program that informed state and federal policymakers about the opportunities, challenges and benefits of programs to increase produce consumption in schools. Lessons from the program are especially important now for two reasons: School districts will be increasing offerings of fruits and vegetables in the School Breakfast Program in July 2014 to meet the new school nutrition guidelines in the Healthy, Hunger-Free Kids Act; and childhood obesity has escalated, with the consequent risk of serious chronic conditions including type 2 diabetes and heart disease. Here, we highlight the results of the California Fresh Start Program, which was conducted during the 2006-2007 school year, and recommend promising strategies for increasing produce consumption by children in the school setting. The barriers we identify to program implementation can provide guidance to policymakers and administrators in school districts nationwide. Responding to the critical state of children’s nutritional health, California enacted Senate Bill 281, commonly known as the California Fresh Start Program , which was signed into law in 2005. It was the first statewide legislation to specifically address fresh and local produce in schools. The innovative pilot program offered a 10-cent per meal reimbursement to schools to increase the servings of fruits and vegetables they offered in the School Breakfast Program. Priority was given to serving fresh fruits and vegetables and, where possible, California-grown produce. The program goals were to promote the consumption of fresh fruits and vegetables, increase school breakfast participation and ultimately improve children’s lifelong eating habits and decrease the incidence of obesity. Supplementing fruits and vegetables in the breakfast program, which serves more than a million California students each day, was an important first step in reaching school-age children, nearly all of whom are at nutritional risk due to low produce consumption. Of California public school students who eat breakfast at school, 78% were reached by the California Fresh Start Program during the 2006-2007 school year.