1201. Localized Cellular Immune Responses to HIV-associated Cryptococcal Meningitis in Cerebrospinal Fluid (CSF) during Antifungal Therapy and Immune Reconstitution Inflammatory Syndrome (IRIS) Events. 
Session: Oral Abstract Session: Biomarkers of Infectious Diseases
Saturday, October 5, 2013: 8:42 AM
Room: The Moscone Center: 300
Background: The high mortality of cryptococcal meningitis (CM) is well recognized but the contributing local leukocyte populations and their activation status at diagnosis, during CM and IRIS are not understood.

Methods: We tested CSF from 50 subjects with CM in Kampala, Uganda at diagnosis (day 0), at day 14 of antifungal therapy (n=15) and at IRIS (n=5). We characterized pan-leukocyte marker CD45+cells in CSF by lineage; natural killer (NK) cells based on CD16, CD56; monocytes (CD3-CD4+loCD56-) based on gating, CD14, CD16; and T cells by CD3+CD4+/-. We characterized activation (HLA-DR+ for CD3+ and NK cells, PDL-1+ for monocytes and NK cells). 

Results: Median CSF WBC percentages changed from days 0 to 14; of all CD45+ cells, CD3+CD4- T cells increased (94% to 97%;P=0.004) (Wilcoxon rank sum test). NK remained unchanged (4.4%), and monocytes decreased (0.6% to 0.2% P=0.03).

At CM diagnosis, T cells were highly activated with HLA-DR expressed on 77.5% of CD3+CD4-, 65% of CD3+4+, but only 7.6% of NK cells. Over 14 days, HLA-DR expression increased on CD3+CD4- (77.5% to 84.4%, P=0.03). Monocytes were predominantly of classical phenotype (88%) (CD14++16-) and activated (36.4% co-expressed PDL-1). NK cells were predominantly CD56++CD16+ and activated (66% expressed HLA-DR).  The percent NK cells correlated with cryptococcal colony forming units/mL on day 0 (r=0.42, P=0.01). Subjects who died had fewer NK cells (IQR) at day 0 compared with survivors (median 34% vs. 53%; P=0.05).

At IRIS events (median 15 weeks on ART), monocytes and T cells did not differ from those of 15 control subjects at screening, whereas NK cells were lower at IRIS (0.3% vs 2.1%, P=0.018). Non-classical monocytes (CD16++CD14+lo) predominated at IRIS (median 92%) vs. classical monocytes earlier.  At IRIS, activation (%HLA-DR+) was lower on CD3+CD4+ (50% vs 33%; P=0.049) but remained elevated on CD3+CD4- (66% vs 74%; P=0.103) compared with controls. 

Conclusion: Highly activated CD3+CD4- cells dominate initial pauci-cellular local leukocyte responses to Cryptococcus in CSF. NK cells are associated with microbiologic outcomes and mortality. Significant differences in T cell activation and monocyte differentiation in CSF highlight evolving immunopathologic mechanisms during IRIS events. 

David Meya, MMed1, Samuel Okurut2, Godfrey Zziwa2, Melissa Rolfes, MPH3, Kelsey Melander4, Proscovia Naluyima2, Brent Palmer4, Andrew Kambugu, MD5, Paul Bohjanen, MD, PhD6, Michael Eller, PhD2, Sharon Wahl7, David Boulware, MD, MPH3 and Edward Janoff, MD4, (1)Infectious Diseases Institute, Makerere University, Kampala, Uganda, (2)Makerere University Walter Reed Project, Makerere University, Uganda, Kampala, Uganda, (3)University of Minnesota, Minneapolis, MN, (4)University of Colorado, Anschutz Medical Center, Aurora, CO, (5)Infectious Diseases Institute, Makerere Univ, Kampala, Uganda, (6)Division of Microbiology, University of Minnesota, Minneapolis, MN, (7)National Institute of Dental and Craniofacial Research, National Institutes of Health, Bethesda, MD


D. Meya, None

S. Okurut, None

G. Zziwa, None

M. Rolfes, None

K. Melander, None

P. Naluyima, None

B. Palmer, None

A. Kambugu, None

P. Bohjanen, None

M. Eller, None

S. Wahl, None

D. Boulware, None

E. Janoff, None

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