Inhibition of Margination and Diapedesis of Neutrophils by Protein Synthesis Blockade

INHIBITION OF MARGINATION AND DIAPEDESIS OF NEUTROPHILS BY PROTEIN SYNTHESIS BLOCKADE By Phyllis V. Acquah, BS, M.Phil A thesis submitted in partial fulfillment of the requirements for the degree of Master of Science in Biochemistry at Virginia Commonwealth University. Virginia Commonwealth University, 2006 Major Director: Dr. Robert Diegelmann, Ph.D Professor, Department of Biochemistry Acute Respiratory Distress Syndrome (ARDS), an age-old clinical problem facing the Emergency Department and Intensive Care Units of all health systems, is a common debilitating lung condition consequent upon severe systemic inflammation. Although several studies have gone into understanding the epidemiology and pathogenesis of the disease thus making way for new advances in treatment strategies, there seems to be no known study tailored to its prevention. Neutrophil extravasation within the tissues during inflammation is the hallmark of this syndrome. Our study, sought to block excessive neutrophil infiltration by inhibiting the biosynthesis of some essential proteins necessary for the process. In .this initial study, neutrophil transmigration was successfully reduced by 66% using protein synthesis inhibitors, a combination of puromycin and anisomycin. Our strategy, if fine-tuned, could form the basis of a new clinical strategy for the prevention of ARDS. Funded by: VCURES (Dr. Kevin R. Ward) Literature Review Introduction Neutrophils, or polymorphonuclear leukocytes (PMN), are indispensable in mammalian host defense during infection. They are part of the innate immune system, which is very much involved in the first few hours of injury or infection before .the adaptive immune system is activated. Although, the role of neutrophils is vital at the site of inflammation, their activities have also been implicated in the parthenogenesis of certain clinical conditions following their activation. An exaggerated neutrophil activation due to extensive tissue injury or shock causes systemic inflammatory response syndrome, which could lead to acute respiratory distress syndrome (ARDS) and subsequently, multiple organ failure (MOF) [I]. Acute Respiratory Distress Syndrome (ARDS) Acute respiratory distress syndrome (ARDS) is a debilitating inflammatory condition resulting from an acute, severe lung injury to most or all of both lungs. Patients with ARDS experience severe shortness of breath and often require mechanical ventilation due to respiratory failure. The condition is characterized by fluid-filled alveoli and decreased lung compliance following extensive lung inflammation and small vessel injury[2]. ARDS is a syndrome not a specific disease, with predisposing factors like sepsis, trauma, and/or pneumonia. In ARDS, there is diffuse alveolar damage, resulting from structural changes in the alveolocapillary unit. Injury to the alveolocapillary membrane inevitably disrupts the endothelial barrier leading to the development of noncardiogenic pulmonary edema through increased vascular permeability. As the air spaces fill with fluid, the gas-exchange and mechanical properties of the lung deteriorate[3]. ARDS was first described in 1967 by Ashbaugh and collaborators, who observed 12 of 272 patients treated for respiratory failure, with the following unique features: cyanosis refractory to oxygen therapy, decreased lung compliance, and diffused infiltrates evident on chest radiograph that distinguished them from the others[4]. Initially, the condition was called adult respiratory distress syndrome but, in 1994, the AmericanEuropean Consensus Committee on ARDS standardized the definition and renamed it "acute" rather than "adult" because it occurs in all ages[5]. The incidence rate of ARDS ranges from 1.5 to 71 per 100,000 persons and affects about 150,000 each year in the United States. This syndrome is a major cause of morbidity, death, and cost in intensive care units [6]. The mortality rate of ARDS has been estimated to be approximately 30 40 %,[7-101 with the deaths usually resulting from multi-system organ failure rather than lung failure alone[2]. The majority of ARDS patients who survive will make a full recovery; however, some will have lasting damage to their lungs. According Herridge and colleagues, survivors of ARDS may experience persistent functional disability even one year after discharge from the intensive care unit, mostly muscle wasting and weakness [I I]. Thus, ARDS affects not only the patient, but also family members by way of stress and cost. Patients who experience severe trauma or sepsis are at increased risk for the development of ARDS with an onset between 24 and 72 hours of injury. In an event of severe infection or trauma, neutrophils are activated and recruited to the site of injury. This is the body's initial immune response to foreign invasion. The immune response to trauma is often exaggerated and the activated neutrophils extravasate not only to the site of infection, but also to distant tissues. Thus, the immunoactivation, though of benefit at the site of injury, has the potential to cause cell-mediated damage at remote organs[l]. In normal circulation, neutrophils constitute about 60% of leukocytes, but in ARDS total percentage can be as high as 90% in the lungs alone. The principal players involved in ARDS are cytokines, neutrophils, endothelium, and reactive oxygen species [12].