Vascular inflammation is a key component for cerebrovascular disease and ischemic injury is suggested to be a significant contributor, resulting in either myocardial ischemia or stroke. A strong inflammatory response is characterized by the release of inflammatory cytokines, thus producing and/or activating pro-inflammatory proteins in the cell. Our previous studies have demonstrated that hypoxia plus glucose deprivation (HGD), an in vitro model of ischemia, increases the proinflammatory mediator, cyclooxygenase-2 levels (COX-2), in vascular tissues. Nuclear factor kappa B (NF-κB) activation is an upstream transcription factor of COX-2 and had been suggested to be involved in “sterile” inflammation in experimental stroke models. Mechanisms underlying the development and progression of inflammation in the cerebrovasculature following ischemic injury in human tissue has not been addressed. Thus, the purpose of this study was to examine the impact of HGD on NF-κB expression and activation in human brain vascular smooth muscle cells (HBVSMC). In addition, we assessed pro-inflammatory mediator levels of downstream NF-κB transcription products, COX-2 and iNOS, and level of its upstream receptor, TLR4. Primary HBVSMC at passage 7 were treated with normoxia (room air) or HGD (1% O2). Following exposure to HGD (3h), cells were isolated, homogenized, and total protein content determined. Lysates, either whole cell or nuclear and cytosolic fractions, were prepped for western blot and analysis. Anti-α-smooth muscle actin was used to verify HBVSMC origin and -actin was used as a loading control. NF-κBp65, phosphorylated NF-κBp65, COX-2, and TLR4 protein levels were all measured post HGD. NF-κBp65 total protein was expressed in HBVSMC and a trend for an increase in levels following HGD was observed. Indirect activation of pNF-kBp65 was assessed via nuclear fractionation studies and was increased following HGD. Lamin AC was used to verify nuclear fractionation. Additional findings suggested that HBVSMC expressed TLR4 however, total protein levels of TLR4 were not altered by HGD. COX-2 and iNOS protein levels were also increased following HGD. In conclusion, these studies indicate that HGD alters proinflammatory enzyme levels, potentially by altering NF-κBp65 activation in human vascular smooth muscle cells. Funding Support: University of Arizona Sarver Heart Center and University of Arizona Valley Research Project Grant VRP P1 (RG).

Cardiovascular disease (CVD) is characterized by impaired vasodilation and the development of atherosclerosis.78 A diet high in saturated fat, such as palmitate, contributes to this by promoting inflammation and oxidative stress in human vascular smooth muscle cells (VSMC). 11,12,84,88 The inflammation cascade that occurs increases pro-inflammatory cytokines, like tumor necrosis factor alpha (TNF-alpha) and increases proinflammatory enzymes like cyclooxygenase 2 (COX-2) contributing to inflammation, oxidative stress, blood pressure shifts, and atherosclerosis.11,12,69,84 Palmitate has been found to upregulate TNF-alpha,85 and COX-2. 11,12, 84

In various studies, sumac, a Mediterranean spice and known antioxidant,39,7,66,67 has been shown to have antioxidant properties through its ability to inhibit reactive oxygen species (ROS) such as superoxide.39,7,66,67 Sumac has also been found to reduce TNF-alpha.100 Results from a study of hypertensive human subjects fed a sumac supplement showed a decrease in blood pressure.59

In the current study, COX-2 levels were determined to evaluate the level of inflammation in response to palmitate when primary aortic human vascular smooth muscle cells (HAoVSM) were treated with sumac. The treatments included: vehicle (bovine serum albumin), 100 µM palmitate, and 10, 20, 40, 60, and 80 µg/mL sumac. Sumac did not alter COX-2 protein levels between vehicle and sumac groups. Additional studies were designed to examine whether 80 µg/mL sumac could reverse impaired vasodilation caused by 10 weeks of high fat intake, consisting of 60% of total calories from fat, in Sprague-Dawley rats. Mesenteric arteries were isolated and exposed to sumac. High fat diet (HFD) arteries had impaired vasodilation compared to arteries from chow-fed fats. HFD arteries exposed to sumac had similar endothelium-dependent vasodilation responses as those not exposed to sumac, however, there were trends for improved vasodilation. I suggest that sumac likely exhibits antioxidant capabilities that prevent superoxide from decreasing the bioavailability of nitric oxide in the vasculature, thus promoting endothelium-dependent vasodilation and preventing the creation of more harmful reactive oxygen species. Isolated arteries from chow fed rats developed irreversible vasodilation when exposed to sumac and were therefore not responsive to pre-constriction with phenylephrine (PE) likely related to nitrates and gallic acid naturally present in sumac whereby inhibiting PE.

Background: To determine the effect of sumac on vasodilation and oxidative stress in vascular tissue. This study hypothesized that sumac would increase vasodilation and reduce vascular damage in vascular tissue taken from rats to improve symptoms and risk of vascular dementia.
Methods: Male Sprague-Dawley rats were fed a chow diet or a high fat diet (HFD) for ten weeks. Endothelium-dependent vasodilation was measured in isolated mesenteric arterioles that were treated with or without 80 µg/ml sumac in the superfusate throughout the experiment.
Results: Sumac did not improve vasodilation or in ex vivo arteries from rats fed a high fat diet. There were trends of improved vasodilation in sumac treated vessels from high fat diet rats, but sumac did not significantly improve vasodilation. In rats fed a chow diet, sumac prevented phenylephrine (PE) constriction in the vascular tissue. The most likely cause for this is the presence of Gallic acid in sumac. Another possible explanation is the presence of nitrates in sumac which may have prevented PE vasoconstriction.
Conclusions: Sumac did not significantly improve vasodilation in isolated arteries from rats fed a high fat diet. The results are inconclusive for the improvement of symptoms or risk of vascular dementia. In vivo treatment with sumac should be tested as results may differ.

A prominent aspect of Alzheimer’s disease (AD) is the presence of neuroinflammation is mediated by the activation of microglial cells, which are the immune cells in the central nervous system (CNS) that express an array of cytokines that may promote an inflammatory response. The main cytokines produced are: tumor necrosis factor-alpha (TNF-), interleukin-1β (IL-1β), and interleukin-6 (IL-6). The presence of these cytokines in the CNS may lead to neuronal death, to the production of toxic chemicals (such as nitric oxide), and to the generation of amyloid beta (a major pathological feature of AD). Previous studies have shown that modulation of the inflammatory response in the nervous system can potentially prevent and/or delay the onset of neurodegenerative diseases such as AD. Therefore, it is important to identify the process that induces CNS inflammation. For example, mitochondrial lysates have been found to produce an inflammatory response due to their ability to stimulate TNF-, Aβ, and APP mRNA [10]. Interestingly, extracellular mitochondria have been detected in the brain due to neurons degrading old mitochondria extracellularly. Therefore, we set out to study the effect of whole mitochondria isolated by differential centrifugation from human neuroblastoma cells (BE(2)-M17 cells) on the neuroinflammatory response in a human microglia model (THP-1 cells). Despite our best efforts, in the end it was unclear whether the mitochondrial fraction or other cellular components induced the inflammatory response we observed. Thus, further work with an improved mitochondrial isolation method should be carried out to address this issue.

Vascular inflammation plays a key role in the development and progression of cardiovascular disease. High fat diet has been associated with cardiovascular risk (1). Therefore, as poor nutrition and poor diet become more widespread, the number of people at risk to cardiovascular disease increases. We hypothesized that using the cancer drug lenalidomide would reverse the inflammation caused by high fat conditions. Human aortic vascular smooth muscle cells were used as an in vitro model to analyze the effect of lenalidomide on high fat diet induced inflammation. Palmitate, a saturated fatty acid was used to induce inflammation. Since lenalidomide has been shown to inhibit cytokine production and attenuate oxidative stress, we investigated whether lenalidomide alters select markers of vascular inflammation in vascular smooth muscle treated with high fat exposure using palmitate. These markers were cyclooxygenase-2 (COX-2) protein levels, TNF-α pro-inflammatory cytokine levels, and superoxide ions. Lenalidomide (5 µM) reversed COX-2 protein expression in cells exposed to high fat conditions (100 µM palmitate). In conclusion, high fat exposure elicits an inflammatory response in cultured primary human vascular smooth muscle, but this response appears to be independent of local cytokine or ROS production. Lenalidomide, although effective at reversing palmitate-induced COX-2, alone augments the pro-inflammatory mediators, COX-2 and TNF-α as well as promotes oxidative stress independent of high fat exposure in human vascular smooth muscle cells.

Morbid obesity is associated with cardiovascular and metabolic disorders. A major contributor to the pathogenesis of these diseases is impaired vasodilation resulting from elevated reactive oxygen species (ROS). This is because certain ROS such as superoxide are raised with obesity and scavenge the endogenous vasorelaxant nitric oxide, resulting in hypertension. The objective of this study was to measure the ability of genistein to quench superoxide in the vasculature of ob/ob mice, an animal model of obesity and type 2 diabetes. Genistein is an isoflavonic phytoestrogen naturally found in soy products. While genistein has documented antioxidant and anti-inflammatory properties, it is not known whether this protects the vasculature from oxidative stress. Genistein was hypothesized to reduce superoxide in arteries from female ob/ob mice. The superoxide indicator dihydroethidium (DHE) [2µL/mL HEPES buffer] was added to isolated aortae and mesenteric arteries from mice fed either a control (standard rodent chow containing 200-300 mg genistein/kg) or genistein-enriched (600mg genistein/kg rodent chow) diets for 4 weeks. Frozen tissues sections were collected onto glass microscope slides and examined using confocal microscopy. Contrary to the hypothesis, a diet containing twice the amount of genistein found in standard chow did not significantly reduce superoxide concentrations in aortae (p=0.287) or mesenteric arteries (p=0.352). Superoxide dismutase, an antioxidant enzyme that breaks down superoxide, was significantly upregulated in the genistein-enriched diet group (p=0.004), although this elevation did not promote the breakdown of superoxide. In addition, the inflammatory marker iNOS was not downregulated in the genistein-enriched diet group (p>0.05). The results indicate that high amounts of isoflavones, like genistein, may not exhibit the purported antioxidant effects in the vasculature of obese or diabetic subjects. Further studies examining arteries from ob/ob mice fed a genistein-free diet are needed to elucidate the true effects of genistein on oxidative stress.

Background: Despite the reported improvements in glucose regulation associated with flaxseeds (Linum usitatissimum) few clinical trials have been conducted in diabetic participants. Objective: To evaluate the efficacy of ground flaxseed consumption at attenuating hyperglycemia, dyslipidemia, inflammation, and oxidative stress as compared to a control in adults with non-insulin dependent type 2 diabetes (T2D). Design: In a randomized parallel arm controlled efficacy trial, participants were asked to consume either 28 g/d ground flaxseed or the fiber-matched control (9 g/d ground psyllium husk) for 8 weeks. The study included 17 adults (9 male, 8 females; 46±14 y; BMI: 31.4±5.7 kg/m2) with a diagnosis of T2D ≥ 6 months. Main outcomes measured included: glycemic control (HbA1c, fasting plasma glucose, fasting serum insulin, and HOMA-IR), lipid profile (total cholesterol, LDL-C, HDL-C, total triglycerides, and calculated VLDL-C), markers of inflammation and oxidative stress (TNF-alpha, TBARS, and NOx), and dietary intake (energy, total fat, total fiber, sodium). Absolute net change for measured variables (week 8 values minus baseline values) were compared using Mann-Whitney U non-parametric tests, significance was determined at p ≤ 0.05. Results: There were no significant changes between groups from baseline to week 8 in any outcome measure of nutrient intake, body composition, glucose control, or lipid concentrations. There was a modest decrease in TNF-alpha in the flaxseed group as compared to the control (p = 0.06) as well as a mild decrease in TBARS in the flaxseed as compared to the control group (p = 0.083), though neither were significant. Conclusions: The current study did not detect a measurable association between 28 g/d flaxseed consumption for 8 weeks in T2D participants and improvements in glycemic control or lipid profiles. There was a modest, albeit insignificant, decrease in markers of inflammation and oxidative stress in the flaxseed group as compared to the control, which warrants further study.