P17 – The Immunophenotype of Culture Positive Bronchoalveolar Lavage from Mechanically Ventilated Trauma Patients

Author(s):
Ryan Huebinger, Ashley Smith, Ding Chen, Sara Ireland, Christian Minshall, Joseph Minei, Steven Wolf, Michael Allen, Robert Barber, Nancy Monson, UT Southwestern Medical Center

Background: Mechanically ventilated trauma patients have a variety of bacterial flora that are often undetectable. To address this problem, we have used next generation sequencing to query the microbiome of such patients as a means to identify the source of infection. In addition, we have initiated a series of experiments to address the immunophenotype in bronchoalveolar lavage (BAL) samples from these same patients. We anticipate this data will allow us to characterize immune responses to particular bacteria in the lung that we identify by next generation sequencing.

Hypothesis: Different immune cell response and phenotypes exist that correlate with the type of bacterial pathogen(s) indentified in bronchoalveolar lavage

Methods: Bronchoalveolar lavage samples were collected in the SICU as a part of standard of care for intubated individuals that had a CPIS>6. BALs were filtered through a 70 micron filter and centrifuged to pellet cells. Cells were resuspended in human freezing media and stored at -80 until flow cytometric analysis. BAL samples were stained with an 11-color flowcytometry panel that identifies both innate and adaptive immune components. BAL cells were analyzed on a FACS Aria flow cytometer.

Results: Our initial analysis of culture positive BALs (CPIS >6) indicates that the vast majority of CD45+ lymphocytes in the BAL samples are CD16+ monocytes, followed by fairly equal frequencies of CD4+ T cells and CD8+ T cells. Some CD19+ B cells are also present, but the majority of these tend to be CD27- naïve B cells rather than the antigen experienced CD27+ B cells.

Conclusions: Understanding the immunophenotype of BAL samples from patients with pulmonary infections will provide a foundation for classifying the genetics of the adaptive immune response that emerge in response to particular bacterial infections of the lung.