Brain pericytes among cells constituting the blood-brain barrier are highly sensitive to tumor necrosis factor-α, releasing matrix metalloproteinase-9 and migrating in vitro

<p>Abstract</p> <p>Background</p> <p>Increased matrix metalloproteinase (MMP)-9 in the plasma and brain is associated with blood-brain barrier (BBB) disruption through proteolytic activity in neuroinflammatory diseases. MMP-9 is present in the brain microvasculature and its vicinity, where brain microvascular endothelial cells (BMECs), pericytes and astrocytes constitute the BBB. Little is known about the cellular source and role of MMP-9 at the BBB. Here, we examined the ability of pericytes to release MMP-9 and migrate in response to inflammatory mediators in comparison with BMECs and astrocytes, using primary cultures isolated from rat brains.</p> <p>Methods</p> <p>The culture supernatants were collected from primary cultures of rat brain endothelial cells, pericytes, or astrocytes. MMP-9 activities and levels in the supernatants were measured by gelatin zymography and western blot, respectively. The involvement of signaling molecules including mitogen-activated protein kinases (MAPKs) and phosphoinositide-3-kinase (PI3K)/Akt in the mediation of tumor necrosis factor (TNF)-α-induced MMP-9 release was examined using specific inhibitors. The functional activity of MMP-9 was evaluated by a cell migration assay.</p> <p>Results</p> <p>Zymographic and western blot analyses demonstrated that TNF-α stimulated pericytes to release MMP-9, and this release was much higher than from BMECs or astrocytes. Other inflammatory mediators [interleukin (IL)-1β, interferon-γ, IL-6 and lipopolysaccharide] failed to induce MMP-9 release from pericytes. TNF-α-induced MMP-9 release from pericytes was found to be mediated by MAPKs and PI3K. Scratch wound healing assay showed that in contrast to BMECs and astrocytes the extent of pericyte migration was significantly increased by TNF-α. This pericyte migration was inhibited by anti-MMP-9 antibody.</p> <p>Conclusion</p> <p>These findings suggest that pericytes are most sensitive to TNF-α in terms of MMP-9 release, and are the major source of MMP-9 at the BBB. This pericyte-derived MMP-9 initiated cellular migration of pericytes, which might be involved in pericyte loss in the damaged BBB.</p

New insights into the impact of neuro-inflammation in rheumatoid arthritis.

Rheumatoid arthritis (RA) is considered to be, in many respects, an archetypal autoimmune disease that causes activation of pro-inflammatory pathways resulting in joint and systemic inflammation. RA remains a major clinical problem with the development of several new therapies targeted at cytokine inhibition in recent years. In RA, biologic therapies targeted at inhibition of tumor necrosis factor alpha (TNFα) have been shown to reduce joint inflammation, limit erosive change, reduce disability and improve quality of life. The cytokine TNFα has a central role in systemic RA inflammation and has also been shown to have pro-inflammatory effects in the brain. Emerging data suggests there is an important bidirectional communication between the brain and immune system in inflammatory conditions like RA. Recent work has shown how TNF inhibitor therapy in people with RA is protective for Alzheimer's disease. Functional MRI studies to measure brain activation in people with RA to stimulus by finger joint compression, have also shown that those who responded to TNF inhibition showed a significantly greater activation volume in thalamic, limbic, and associative areas of the brain than non-responders. Infections are the main risk of therapies with biologic drugs and infections have been shown to be related to disease flares in RA. Recent basic science data has also emerged suggesting that bacterial components including lipopolysaccharide induce pain by directly activating sensory neurons that modulate inflammation, a previously unsuspected role for the nervous system in host-pathogen interactions. In this review, we discuss the current evidence for neuro-inflammation as an important factor that impacts on disease persistence and pain in RA

Suppression of PKC-α attenuates TNF-α-evoked cerebral barrier breakdown via regulations of MMP-2 and plasminogen–plasmin system

Ischaemic stroke, accompanied by neuroinflammation, impairs blood-brain barrier integrity through a complex mechanism involving both protein kinase C (PKC) and urokinase. Using an in vitro model of human blood-brain barrier (BBB) composed of brain microvascular endothelial cells (HBMEC) and astrocytes, this study assessed the putative roles of these elements in BBB damage evoked by enhanced availability of pro-inflammatory cytokine, TNF-α. Treatment of HBMEC with TNF-α significantly increased the mRNA and protein expressions of all plasminogen-plasmin system (PPS) components, namely tissue plasminogen activator, urokinase, urokinase plasminogen activator receptor and plasminogen activator inhibitor-1 and also the activities of urokinase, total PKC and extracellular MMP-2. Inhibition of urokinase by amiloride abated the effects of TNF-α on BBB integrity and MMP-2 activity without affecting that of total PKC. Conversely, pharmacological inhibition of conventional PKC isoforms dramatically suppressed TNF-α-induced overactivation of urokinase. Knockdown of PKC-α gene via specific siRNA in HBMEC suppressed the stimulatory effects of TNF-α on protein expression of all PPS components, MMP-2 activity, DNA fragmentation rates and pro-apoptotic caspase-3/7 activities. Establishment of co-cultures with BMEC transfected with PKC-α siRNA attenuated the disruptive effects of TNF- on BBB integrity and function. This was partly due to elevations observed in expression of a tight junction protein, claudin-5 and partly to prevention of stress fibre formation. In conclusion, specific inhibition of PKC-α in cerebral conditions associated with exaggerated release of pro-inflammatory cytokines, notably TNF- may be of considerable therapeutic value and help maintain endothelial cell viability, appropriate cytoskeletal structure and basement membrane

Defective adipose tissue development associated with hepatomegaly in cathepsin E-deficient mice fed a high-fat diet

Cathepsin E is an intracellular aspartic proteinase, which is predominantly distributed in immune-related and epithelial cells. However, the role of the enzyme in adipose tissues remains unknown. In this study, we investigated the characteristics of cathepsin E-deficient (CatE-/-) mice fed a high-fat diet (HFD), as a mouse model of obesity. HFD-fed CatE-/- mice displayed reduced body weight gain and defective development of white adipose tissue (WAT) and brown adipose tissue (BAT), compared with HFD-fed wild-type mice. Moreover, fat-induced CatE-/- mice showed abnormal lipid accumulation in non-adipose tissues characterized by hepatomegaly, which is probably due to defective adipose tissue development. Detailed pathological and biochemical analyses showed that hepatomegaly was accompanied by hepatic steatosis and hypercholesterolemia in HFD-induced CatE-/- mice. In fat-induced CatE-/- mice, the number of macrophages infiltrating into WAT was significantly lower than in fat-induced wild-type mice. Thus, the impaired adipose tissue development in HFD-induced CatE-/- mice was probably due to reduced infiltration of macrophages and may lead to hepatomegaly accompanied by hepatic steatosis and hypercholesterolemia

Role of the Blood-Brain Barrier in the Formation of Brain Metastases

The majority of brain metastases originate from lung cancer, breast cancer and malignant melanoma. In order to reach the brain, parenchyma metastatic cells have to transmigrate through the endothelial cell layer of brain capillaries, which forms the morphological basis of the blood-brain barrier (BBB). The BBB has a dual role in brain metastasis formation: it forms a tight barrier protecting the central nervous system from entering cancer cells, but it is also actively involved in protecting metastatic cells during extravasation and proliferation in the brain. The mechanisms of interaction of cancer cells and cerebral endothelial cells are largely uncharacterized. Here, we provide a comprehensive review on our current knowledge about the role of junctional and adhesion molecules, soluble factors, proteolytic enzymes and signaling pathways mediating the attachment of tumor cells to brain endothelial cells and the transendothelial migration of metastatic cells. Since brain metastases represent a great therapeutic challenge, it is indispensable to understand the mechanisms of the interaction of tumor cells with the BBB in order to find targets of prevention of brain metastasis formation

Astroglial Inhibition of NF-κB Does Not Ameliorate Disease Onset and Progression in a Mouse Model for Amyotrophic Lateral Sclerosis (ALS)

Motor neuron death in amyotrophic lateral sclerosis (ALS) is considered a “non-cell autonomous” process, with astrocytes playing a critical role in disease progression. Glial cells are activated early in transgenic mice expressing mutant SOD1, suggesting that neuroinflammation has a relevant role in the cascade of events that trigger the death of motor neurons. An inflammatory cascade including COX2 expression, secretion of cytokines and release of NO from astrocytes may descend from activation of a NF-κB-mediated pathway observed in astrocytes from ALS patients and in experimental models. We have attempted rescue of transgenic mutant SOD1 mice through the inhibition of the NF-κB pathway selectively in astrocytes. Here we show that despite efficient inhibition of this major pathway, double transgenic mice expressing the mutant SOD1G93A ubiquitously and the dominant negative form of IκBα (IκBαAA) in astrocytes under control of the GFAP promoter show no benefit in terms of onset and progression of disease. Our data indicate that motor neuron death in ALS cannot be prevented by inhibition of a single inflammatory pathway because alternative pathways are activated in the presence of a persistent toxic stimulus

The restorative role of annexin A1 at the blood–brain barrier

Annexin A1 is a potent anti-inflammatory molecule that has been extensively studied in the peripheral immune system, but has not as yet been exploited as a therapeutic target/agent. In the last decade, we have undertaken the study of this molecule in the central nervous system (CNS), focusing particularly on the primary interface between the peripheral body and CNS: the blood–brain barrier. In this review, we provide an overview of the role of this molecule in the brain, with a particular emphasis on its functions in the endothelium of the blood–brain barrier, and the protective actions the molecule may exert in neuroinflammatory, neurovascular and metabolic disease. We focus on the possible new therapeutic avenues opened up by an increased understanding of the role of annexin A1 in the CNS vasculature, and its potential for repairing blood–brain barrier damage in disease and aging

Tissue plasminogen activator enhances the hypoxia/reoxygenation-induced impairment of the blood-brain barrier in a primary culture of rat brain endothelial cells.

Hemorrhagic transformation is a major complication associated with tissue plasminogen activator (tPA) therapy for ischemic stroke. We studied the effect of tPA on the blood-brain barrier (BBB) function with our in vitro monolayer model generated using rat brain microvascular endothelial cells subjected either to normoxia or to hypoxia/reoxygenation (H/R) with or without the administration of tPA. The barrier function was evaluated by the transendothelial electrical resistance (TEER), the permeability of sodium fluorescein and Evans\u27 blue-albumin (EBA), and the uptake of lucifer yellow (LY). The permeability of sodium fluorescein and EBA was used as an index of paracellular and transcellular transport, respectively. The administration of tPA increased the permeability of EBA and the uptake of LY under normoxia. It enhanced the increase in the permeability of both sodium fluorescein and EBA, the decrease in the TEER, and the disruption in the expression of ZO-1 under H/R conditions. Administration of tPA could cause an increase in the transcellular transport under normoxia, and both the transcellular and paracellular transport of the BBB under H/R conditions in vitro. Even in humans, tPA may lead to an opening of the BBB under non-ischemic conditions and have an additional effect on the ischemia-induced BBB disruption.The original publication is available at www.springerlink.co