Abstract

ABSTRACTS

Endothelial Dysfunction in Cardiovascular Risk:
A Nanotechnology Approach

R. Preston Mason

Cardiovascular risk factors are inextricably linked to atherosclerosis, a systemic inflammatory process that manifests clinically as coronary and cerebrovascular diseases. During atherosclerosis, damage to the endothelium leads to reduced nitric oxide (NO) bioavailability, an important vasodilator that also inhibits platelet aggregation, leukocyte adhesion and smooth muscle cell proliferation. There is now evidence that multiple risk factors, such as hypertension and hyperlipidemia, lead to an accelerated loss of endothelial function through oxidative stress mechanisms. Using nanotechnology approaches such as x-ray diffraction, we have evaluated the specific contributions of oxidative stress to mechanisms of endothelial dysfunction. These observations provide a compelling rationale for aggressive and comprehensive management of risk factors in order to effectively reverse endothelial dysfunction.
A goal for therapy is the restoration of endothelial function, and specifically, NO synthase (NOS) coupling efficiency. Certain calcium channel blockers (CCBs), RAS-inhibitors, β-blockers and statins are known to directly improve endothelial function and NO bioavailability beyond risk factor management, as characterized by nanosensor techniques. Our studies with a charged dihydroypridine-type CCBs and statin indicate that these agents synergistically stimulate NO release from human endothelial cells in a fashion that reverses endothelial dysfunction, independently of their separate effects on blood pressure or LDL levels. These agents enhance endothelial-dependent NO release through changes in caveolin-1 expression, NOS coupling mechanisms and enhanced sensitivity to bradykinin. The implications of these basic science observations are discussed in light of recent clinical trials that indicate possible interactions between certain antihypertensive and lipid lowering agents on clinical events.

Adolescent Depression and Suicide

Donald E. Greydanus

Sadness is one of the basic human emotions and is usually associated with a current or past tragic event. A more sustained state of sadness is called depression which can either arise spontaneously or follow disruptive events in life. One percent to 2% of children will experience a depressive disorder, while during or after puberty, the prevalence of depression increases to 3%-8% of young adolescents. By the end of adolescence, about 20% of pediatric individuals will have had a diagnosable depressive disorder. Depression is a major precipitant to suicide in adolescents. Suicides are among the three leading causes of death for adolescents in the world and rates are rising faster in teens than in other age groups; at least 90,000 adolescents commit suicide each year in the world in the context of 4 million suicide attempts each year. Suicide rates globally are underreported because they may be classified as accidents or not classified at all. The ratio of suicide attempts to completed suicides is 3 to 1 (female to male) while three times as many males vs. females complete the suicide. Chronic illness, gender orientation concerns, and abuse are other risk factors for suicide. Clinicians caring for adolescents must be aware of the risk of teen death from suicide and must be prepared to screen all their patients for this potentially tragic but preventable phenomenon.

The Roles of β₁-integrins during Focal Cerebral Ischemia

Gregory J. del Zoppo

Focal cerebral ischemia initiates profound changes in the integrity of microvessels and in the neurovascular unit in the brain. These changes are very rapid: activation of endothelial cells, alterations in basal lamina matrix, and astrocyte changes occur simultaneously with injury to the neurons targetted by ischemia. The neurovascular unit is a conceptual construct that links the functions of cerebral microvessels with adjacent neurons/axons via their astrocytes. β₁-integrins appear to play prominent roles in the integrity of the microvasculature (endothelial cells and astrocyte end-feet), responses to hemorrhage by the cerebral tissues (microglia), and in new vessel formation (microvessels), in addition to other adhesion receptors (e.g. the integrins α₆β₄ and α_vβ₃, and the receptor αβ-dystroglycan). During focal cerebral ischemia the integrin β₁-subunit and β-dystroglycan rapidly disappear from the endothelial surface and astrocyte-end feet, respectively, which attach to the intervening matrix. Hemorrhage can accompany focal ischemia, and is correlated with the loss of microvessel matrix. The matrix metalloproteinase pro-MMP-9 is generated in ischemic tissue in response to hemorrhagic transformation. Pro-MMP-9 appears from the responses of activated microglia to plasma proteins, such as fibronectin, that extravasate into the tissue. This activation is β₁-integrin dependent. The generation of new vessels, in separate settings depends upon the expression of integrin α_vβ₃, and possibly α₅β₁. Because of their important roles in cell survival and structure, changes in β₁-integrin family expression by specific cells within the CNS imply major alterations in the neurovascular unit. They also imply roles in cell-cell signalling within the neurovascular unit.

Macrophage Iron Signaling and Myofibroblast Cell Fate Regulation

Hide Tsukamoto

Our research pursues two simplistic yet central questions concerning the genesis of steatohepatitis and cirrhosis: 1) why the liver becomes chronically inflamed; and 2) why the liver becomes progressively scarred. For the former, we entertain the notion that macrophage iron accumulation in chronic inflammation positively feeds back to promote inflammation via accentuated iron signaling for IKK activation. This notion is based on our novel discovery on intracellular signaling of catalytically active iron in endotoxin or cytokine-stimulated macrophages characterized by a peroxynitrite-elicited transient rise in low molecular weight-iron complexes as an essential immediate event for IKK and NF-κB activation, and pro-inflammatory cytokine expression (J Biol Chem 278:17646, 2003). An increment in the chelatable pool of iron as seen in macrophages from chronically damaged livers or artificially produced by injection of iron dextran, heightens the signaling and downstream IKK and NF-κB activation and aggravates liver disease. Reduced iron activates IKK in a manner dependent on p21Ras, TAK1, PI3K, NIK, and MEKK and via protein-protein interactions among the former three effector molecules in caveosomes and endosomes (J Biol Chem 282:5582, 2007). The iron signaling is a function acquired by differentiated macrophages and relevant to human, and should be considered as a molecular target for chronic inflammation. For the latter question, our research demonstrates the loss of adipogenic transcriptional regulation underlies transdifferentiation of hepatic stellate cells (HSCs) to myofibroblasts in liver fibrogenesis. This anti-adipogenic transdifferentiation is mediated at least by TNF and Wnt, and restoration of adipogenic regulation by forced expression of PPARγ or SREBP-1c or by down-regulation of canonical Wnt signaling by Dickkopf-1, results in a phenotypic reversal of myofibroblasts to quiescent HSCs (J Biol Chem 279:11392, 2004, 280:4959, 2005; Am J Physiol 294:G39, 2008). This cell fate regulation is similarly regulated by transcriptional and epigenetic mechanisms to mesenchymal cell type determination and differentiation from pluripotent stem cells.

Mechanisms Controlling Type I Interferon Responses by Plasmacytoid Dendritic Cells

Wei Cao and Yong-Jun Liu

Plasmacytoid dendritic cells (pDCs) are rare but special immune cells important for antiviral immunity by producing large amounts of type I interferons (IFNs) upon viral infection. Such extraordinary ability is mediated through unique signal activation machinery including Toll-like receptor (TLR)-7 and TLR9, intracellular sensors to single stranded RNA and DNA, respectively. Since type I IFNs have profound effect on a wide range of innate and adaptive immune cells, the IFN response has to be under tight control; otherwise, autoimmune diseases or lymphopenia may arise. We and others have reported that pDCs express an array of surface receptors, such as ILT7, BDCA2, Siglec-H, high affinity IgE receptor, most of which couple with transmembrane signaling adaptors FcγR or DAP12 to form receptor complexes. Activation of these pDC receptors triggers a novel signaling pathway similar to that downstream of B cell receptor activation, but distinct from that mediates T cell receptor activation, which negatively modulates type I IFN and cytokine responses by pDCs upon TLR activation. The physiological context of such receptor-mediated regulation is revealed when the biological ligands for some of the receptors are taken into consideration. Therefore, pDCs, as potent innate immune responders as they are intrinsically, are constantly controlled by complex mechanisms to ensure appropriate responses.

Progress towards the Development of an Inner Ear Drug Delivery System

Michael McKenna, Jeffrey Borenstein, William Sewell, Sharon Kujawa, Jason Fiering, Mark Mescher, Zhijang Chen, Erin Leary-Swan, Brian Murphy, Marcello Peppi and Sara Tao

Advances in molecular biology of the auditory and vestibular systems are beginning to uncover opportunities for the treatment of degenerative inner ear disorders. The clinical exploitation of these discoveries will require the delivery of biological molecules and drugs to the inner ear in controlled and sustained amounts. Current methods for inner ear drug delivery are inefficient and often unpredictable. We have developed a drug delivery device with precision control of flow rates and volumes capable of delivering biologically active compounds for sustained periods of time. We have begun testing this device in guinea pigs, and have demonstrated the ability to distribute drug a significant distance from the site of entry without adversely effecting hearing. We have begun to design and fabricate the components for a fully implantable device that may ultimately be implanted in humans. In it's current design, perilymph is recirculated through the device, including a reservoir for the mixing of stored and stabilized drug. The device contains pressure and flow sensors that allow for programmed adjustment in pump output to maintain desired flow rates and concentration of delivered drug. In addition, the device is telemetrically controlled to allow for external adjustments to maximize desired response.