(1021) Developing an In Vitro Infectious Cell Culture Model for Acute Lung Injury

Introduction: Inflammation is a key characteristic of acute lung injury (ALI), or its clinical correlate, acute respiratory distress syndrome (ARDS). The majority of ARDS is caused by infections such as sepsis or pneumonia. MRSA is a serious pathogen contributing to pulmonary infections and is commonly encountered in critically ill patients. Animal models of ALI, while critical for basic and translational research, are costly and time intensive. Complementary in vitro cell culture models are a useful tool to study basic mechanisms of ALI, as they are comparatively easy to manipulate, for example by knock out or knock in studies.

Methods: Alveolar epithelial (A549) cells were treated with the following final concentrations of Methicillin-resistant Staphylococcus aureus (MRSA) USA300 strain from a 108 diluted solution in cell culture medium: 1ul/ml, 5ul/ml, and 10ul/ml. The A549 cells were plated on a 6-well plate with 300,000 cells each. Cells were harvested at 0h, 2h, 4h, and 6h. To determine the efficiency of the model: cell death rate and expression of proinflammatory cytokines were utilized.

Results: Cell death percentages increased as time and concentration of MRSA increased. A concentration of 5ul/ml at 4 hours demonstrated the closest cell death percentage to 25-35%, and pro-inflammatory cytokines were increased, which were comparable to cytokines levels measured in a murine model of MRSA pneumonia.

Conclusions: Overall, the data supports that a system was established that resulted in significant cell death and proinflammatory cytokine expression.. This model will be a valuable tool in investigating acute lung injury caused by infection as it allows for easy manipulation and will enable high throughput screening experiments.