Oxidative LDL modification is increased in vascular dementia and is inversely associated with cognitive performance

It is not known whether the association between increased plasma homocysteine (Hcy) associated with LDL modification and propensity for LDL uptake by macrophages in cardiovascular disease patients holds true in vascular dementia (VaD). Plasma from 83 subjects diagnosed with Alzheimer's disease (AD), VaD, mild cognitive impairment (MCI) and from controls was analysed to examine (1) whether LDL isolated from the plasma of VaD is biochemically and functionally distinct from that isolated from AD, MCI or controls; and (2) whether such biomarkers of LDL phenotype are related to plasma folate levels, Hcy levels and/or to disease severity. Folate and vitamin B6 levels were significantly lower in VaD subjects than in controls. VaD-LDL showed increased protein carbonyl content (p <0.05) and was more susceptible to scavenging by macrophages (p <0.05) than AD- or control-LDL. Patients from the VaD cohort were more prevalent in the lowest tertile for HDL:LDL and the upper tertile for LDL oxidation; the combined parameters of HDL cholesterol, LDL oxidation and scavenging by macrophages show 87% sensitivity towards VaD detection. The association between folate deficiency, LDL modification and dysfunction in VaD but not in AD may provide a novel biomarker assessment to discriminate between the diseases.

Ocular and systemic vascular dysfunction in neurodegenerative disease

The important role played by vascular factors in the pathogenesis of neurodegenerative disease has been increasingly realised over recent years. The nature and impact of ocular and systemic vascular dysfunction in the pathogenesis of comparable neurodegenerative diseases such as glaucoma and Alzheimer’s disease (AD) has however never been fully explored. The aim of this thesis was therefore to investigate the presence of macro- and micro-vascular alterations in both glaucoma and AD and to explore the relationships between these two chronic, slowly progressive neurodegenerative diseases. The principle sections and findings of this work were: 1. Is the eye a window to the brain? Retinal vascular dysfunction in Alzheimer’s disease · Mild newly diagnosed AD patients demonstrated ocular vascular dysfunction, in the form of an altered retinal vascular response to flicker light, which correlated with their degree of cognitive impairment. 2. Ocular and systemic vascular abnormalities in newly diagnosed normal tension glaucoma (NTG) patients · NTG patients demonstrated an altered retinal arterial constriction response to flicker light along with an increased systemic arterial stiffness and carotid artery intima-media thickness (IMT). These findings were not replicated by healthy age matched controls. 3. Ocular vascular dysregulation in AD compares to both POAG and NTG · AD patients demonstrated altered retinal arterial reactivity to flicker light which was comparable to that of POAG patients and altered baseline venous reactivity which was comparable to that of NTG patients. Neither alteration was replicated by healthy controls. 4. POAG and NTG: two separate diseases or one continuous entity? The vascular perspective · POAG and NTG patients demonstrated comparable alterations in nocturnal systolic blood pressure (SBP) variability, ocular perfusion pressure, retinal vascular reactivity, systemic arterial stiffness and carotid IMT. · Nocturnal SBP variability was found to correlate with both retinal artery baseline diameter fluctuation and carotid IMT across the glaucoma groups.

Vascular endothelial growth factor promotes physical wound repair and is anti-apoptotic in primary distal lung epithelial and A549 cells

Objective: There is evidence to suggest a beneficial role for growth factors, including vascular endothelial growth factor (VEGF), in tissue repair and proliferation after injury within the lung. Whether this effect is mediated predominantly by actions on endothelial cells or epithelial cells is unknown. This study tested the hypothesis that VEGF acts as an autocrine trophic factor for human adult alveolar epithelial cells and that under situations of pro-apoptotic stress, VEGF reduces cell death. Design: In vitro cell culture study looking at the effects of 0.03% H2O2 on both A549 and primary distal lung epithelial cells.Measurement and Main Results: Primary adult human distal lung epithelial cells express both the soluble and membrane-associated VEGF isoforms and VEGF receptors 1 and 2. At physiologically relevant doses, soluble VEGF isoforms stimulate wound repair and have a proliferative action. Specific receptor ligands confirmed that this effect was mediated by VEGF receptor 1. In addition to proliferation, we demonstrate that VEGF reduces A549 and distal lung epithelial cell apoptosis when administered after 0.03% H2O2 injury. This effect occurs due to reduced caspase-3 activation and is phosphatidylinositol 3′–kinase dependent. Conclusion: In addition to its known effects on endothelial cells, VEGF acts as a growth and anti-apoptotic factor on alveolar epithelial cells. VEGF treatment may have potential as a rescue therapy for diseases associated with alveolar epithelial damage such as acute respiratory distress syndrome.

Plasma membrane calcium ATPase isoform 4 inhibits vascular endothelial growth factor-mediated angiogenesis through interaction with calcineurin

Approach and Results - Using in vitro and in vivo assays, we here demonstrate that the interaction between PMCA4 and calcineurin in VEGF-stimulated endothelial cells leads to downregulation of the calcineurin/NFAT pathway and to a significant reduction in the subsequent expression of the NFAT-dependent, VEGF-activated, proangiogenic genes RCAN1.4 and Cox-2. PMCA4-dependent inhibition of calcineurin signaling translates into a reduction in endothelial cell motility and blood vessel formation that ultimately impairs in vivo angiogenesis by VEGF. Objective - Vascular endothelial growth factor (VEGF) has been identified as a crucial regulator of physiological and pathological angiogenesis. Among the intracellular signaling pathways triggered by VEGF, activation of the calcineurin/ nuclear factor of activated T cells (NFAT) signaling axis has emerged as a critical mediator of angiogenic processes. We and others previously reported a novel role for the plasma membrane calcium ATPase (PMCA) as an endogenous inhibitor of the calcineurin/NFAT pathway, via interaction with calcineurin, in cardiomyocytes and breast cancer cells. However, the functional significance of the PMCA/calcineurin interaction in endothelial pathophysiology has not been addressed thus far. Conclusions - Given the importance of the calcineurin/NFAT pathway in the regulation of pathological angiogenesis, targeted modulation of PMCA4 functionality might open novel therapeutic avenues to promote or attenuate new vessel formation in diseases that occur with angiogenesis.

Author response: can vascular function be assessed by the interpretation of retinal vascular diameter changes?

A protocol with repeated stimulation cycles should be analyzed stepwise, in that each stimulation is evaluated, and a reaction pattern is identified. No two subjects will react identically, in that dilation and recovery times can vary; however, this is not reason enough to abandon a multiple stimulation cycle with fixed recovery and stimulation times. Furthermore, it enables us to examine and determine the range in which a normal subject will be placed and can then be compared to different pathophysiological states (i.e., smokers and different diseases). The purpose of our paper was to highlight the importance of evaluating these different cycles and the danger of false interpretation when averaging results. There are many different ways of evaluating dilatory responses and elasticity, but each of them must be carefully evaluated and should not be overaveraged, which can result in a loss of sensitivity and specificity.

Investigating the Role of PDE4D1/2 in Vascular Smooth Muscle Cell Desensitization

Vascular smooth muscle cell (VSMC) behaviour and phenotypic modulation is critical to vessel repair following damage, and the progression of various cardiovascular diseases. The second messenger cyclic adenosine monophosphosphate (cAMP) plays a key role in VSMC function under the synthetic/activated phenotype, which is typically associated with unhealthy cell behaviour. Consequently, cAMP signaling is often targeted in attempts to impact several pathological diseases, including atherosclerosis, restenosis, and pulmonary arterial hypertension (PAH). The cyclic nucleotide phosphodiesterases (PDEs) catalyze hydrolysis of cAMP to an inactive form, and therefore directly regulate cAMP signaling. The PDE4D family dominates in synthetic VSMCs, and there is considerable interest in determining how distinct PDE4D isoforms affect cell function. Specifically, we are interested in the potential link between short isoforms of PDE4D and VSMC desensitization to pharmacological agents that impact cardiovascular disease via cAMP signaling. This study extends on previous work that assessed the expression of PDE4D splice variants in rat aortic VSMCs following prolonged challenge with cAMP-elevating agents. It was determined that PDE4D1 and PDE4D2 were uniquely expressed in synthetic VSMCs incubated with these agents, and that this upregulation impacted PDE activity and cAMP accumulation in these cells. Here, we report that PDE4D1 and PDE4D2 are markedly upregulated in synthetic human aortic smooth muscle cells (HASMCs) following prolonged challenge with cAMP-elevating agents. Using a combination of RNAi-based and pharmacological approaches, we establish that this upregulation is reflected in levels of cAMP PDE activity, and restricted to the cytosolic sub-cellular compartment. Our results suggest a role for localized PDE4D1 and PDE4D2 activity in regulating cAMP-mediated desensitization in HASMCs, and highlight their therapeutic potential in treating various cardiovascular diseases.

Bisphenol A alters transcript levels of biomarker genes for major depressive disorder in vascular endothelial cells and colon cancer cells

Bisphenol A (BPA) is capable of mimicking endogenous hormones with potential consequences for human health and BPA exposure has been associated with several human diseases including neuropsychiatric disorders. Here, quantitative Real-Time Polymerase Chain Reaction (qRT-PCR) results show that BPA at low concentrations (10 ng/mL and 1 μg/mL) induces differential transcript levels of four biomarker genes for Major Depressive Disorder (MDD) in HT29 human colon adenocarcinona cell line and Human Umbilical Vein Endothelial Cells (HUVEC). These results substantiate increasing concerns of BPA exposure in levels currently detected in humans.

The regulation of AMP-activated protein kinase by vascular endothelial growth factor

The function of the vascular endothelium is to maintain vascular homeostasis, by providing an anti-thrombotic, anti-inflammatory and vasodilatory interface between circulating blood and the vessel wall, meanwhile facilitating the selective passage of blood components such as signaling molecules and immune cells. Dysfunction of the vascular endothelium is implicated in a number of pathological states including atherosclerosis and hypertension, and is thought to precede atherogenesis by a number of years. Vascular endothelial growth factor A (VEGF) is a crucial mitogenic signaling molecule, not only essential for embryonic development, but also in the adult for regulating both physiological and pathological angiogenesis. Previous studies by our laboratory have demonstrated that VEGF-A activates AMP-activated protein kinase (AMPK), the downstream component of a signaling cascade important in the regulation of whole body and cellular energy status. Furthermore, studies in our laboratory have indicated that AMPK is essential for VEGF-A-stimulated vascular endothelial cell proliferation. AMPK activation typically stimulates anabolic processes and inhibits catabolic processes including cell proliferation, with the ultimate aim of redressing energy imbalance, and as such is an attractive therapeutic target for the treatment of obesity, metabolic syndromes, and type 2 diabetes. Metabolic diseases are associated with adverse cardiovascular outcomes and AMPK activation is reported to have beneficial effects on the vascular endothelium. The mechanism by which VEGF-A stimulates AMPK, and the functional consequences of VEGF-A-stimulated AMPK activation remain uncertain. The present study therefore aimed to identify the specific mechanism(s) by which VEGF-A regulates the activity of AMPK in endothelial cells, and how this might differ from the activation of AMPK by other agents. Furthermore, the role of AMPK in the pro-proliferative actions of VEGF-A was further examined. Human aortic and umbilical vein endothelial cells were therefore used as a model system to characterise the specific effect(s) of VEGF-A stimulation on AMPK activation. The present study reports that AMPK α1 containing AMPK complexes account for the vast majority of both basal and VEGF-A-stimulated AMPK activity. Furthermore, AMPK α1 is localized to the endoplasmic reticulum when sub-confluent, but translocated to the Golgi apparatus when cells are cultured to confluence. AMPK α2 appears to be associated with a structural cellular component, but neither α1 nor α2 complexes appear to translocate in response to VEGF-A stimulation. The present study confirms previous reports that when measured using the MTS cell proliferation assay, AMPK is required for VEGF-A-stimulated endothelial cell proliferation. However, parallel experiments measuring cell proliferation using the Real-Time Cell Analyzer xCELLigence system, do not agree with these previous reports, suggesting that AMPK may in fact be required for an aspect of mitochondrial metabolism which is enhanced by VEGF-A. Studies into the mitochondrial activity of endothelial cells have proved inconclusive at this time, but further studies into this are warranted. During previous studies in our laboratory, it was suggested that VEGF-A-stimulated AMPK activation may be mediated via the diacylglycerol (DAG)-sensitive transient receptor potential cation channel (TRPCs -3, -6 or -7) family of ion channels. The present study can neither confirm, nor exclude the expression of TRPCs in vascular endothelial cells, nor rule out their involvement in VEGF-A-stimulated AMPK activation; more specific investigative tools are required in order to characterise their involvement. Furthermore, nicotinic acid adenine dinucleotide phosphate (NAADP)-stimulated Ca2+ release from acidic intracellular organelles is not required for AMPK activation by VEGF-A. Despite what is known about the mechanisms by which AMPK is activated, far less is known concerning the downregulation of AMPK activity, as observed in human and animal models of metabolic disease. Phosphorylation of AMPK α1 Ser485 (α2 Ser491) has recently been characterised as a mechanism by which the activity of AMPK is negatively regulated. We report here for the first time that VEGF-A stimulates AMPK α1 Ser485 phosphorylation independently of the previously reported AMPK α1 Ser485 kinases Akt (protein kinase B) and ERK1/2 (extracellular signal-regulated kinase 1/2). Furthermore, inhibition of protein kinase C (PKC), the activity of which is reported to be elevated in metabolic disease, attenuates VEGF-A- and phorbol 12-myristate 13-acetate (PMA)-stimulated AMPK α1 Ser485 phosphorylation, and increases basal AMPK activity. In contrast to this, PKC activation reduces AMPK activity in human vascular endothelial cells. Attempts to identify the PKC isoform responsible for inhibiting AMPK activity suggest that it is one (or more) of the Ca2+-regulated DAG-sensitive isoforms of PKC, however cross regulation of PKC isoform expression has limited the present study. Furthermore, AMPK α1 Ser485 phosphorylation was inversely correlated with human muscle insulin sensitivity. As such, enhanced AMPK α1 Ser485 phosphorylation, potentially mediated by increased PKC activation may help explain some of the reduced AMPK activity observed in metabolic disease.

Flexible bronchoscopy contribution in the approach of diagnosis and treatment of children’s respiratory diseases: the experience of a unique pediatric unit in Tunisia.

Objective: Our study aimed at assessing the role of flexible bronchoscopy (FB) in improving diagnosis and management of children’s respiratory conditions in the pediatric unit of FB, newly created and unique in Tunisia. Methods: Retrospective study including all the FB achieved in our pediatric unit from 2009 to 2014. Results: We performed 365 FB in 333 patients aged 46 months on average (1 month - 15 years), often under conscious anesthesia (81.6%). FB was performed for diagnostic purposes in 341 cases and for therapeutic purposes in 24 cases. Eight anatomical abnormalities were revealed in 22 patients. An intraluminal bronchial obstruction was found in 71 FB, mainly due to a foreign body (n=36). A vascular anomaly was responsible for nine cases out of 17 extraluminal obstructions. Airways malacia was observed in 60 FB. Bronchoalveolar lavage was performed in 196 cases. It was determinant in 43.9% of the cases. FB was of great diagnostic value in 74.8% of the cases. It influenced the management of the patients in 58% of the cases. The FB for therapeutic purposes was beneficial in all cases. Few complications occurred (5.5%). Conclusion: FB is a safe tool providing precious diagnostic and/or therapeutic help for the clinician. Keywords:

Renal- and calcium-dependent vascular effects of Polybia paulista wasp venom

In the present study, the effects of Polybia paulista venom (PPV) on renal and vascular tissues were investigated. Isolated kidneys perfused with PPV (1 and 3 mu g/mL) had increased perfusion pressure, renal vascular resistance, urinary flow, and glomerular filtration rate; and reduced sodium tubular transport. Histological evaluation demonstrated deposits of proteins in Bowman's space and tubular lumen, and focal areas of necrosis. The venom promoted a cytotoxic effect on Madin-Darby canine kidney (MDCK) cells. A significant increase in lactic dehydrogenase levels was observed in response to venom exposure. In isolated mesenteric vascular beds, pressure and vascular resistance augmented in a dose-dependent manner. PPV increased the contractility of aortic rings maintained under basal tension. This contractile response was inhibited when preparations were maintained in Ca2+-free medium. Likewise, verapamil, a voltage-gated calcium channel blocker, also inhibited the contractile response. In this study, phentolamine, a blocker of a-adrenergic receptor blocker, significantly reduced the contractile effect of PPV in the aortic ring. In conclusion, PPV produced nephrotoxicity, which suggests a direct effect on necrotic cellular death in renal tubule cells. The vascular contractile effect of PPV appears to involve calcium influx through voltage-gated calcium channels via adrenergic regulation.

Investigating the Role of PDE4D1/2 in Vascular Smooth Muscle Cell Desensitization

Thesis (Master, Biochemistry) -- Queen's University, 2016-10-14 02:44:01.604