1344. CX3CR1-M280 Impairs Fractalkine-induced CX3CR1 Signaling in Human Monocytes and Results in Defective Monocyte Survival and Monocytopenia at Steady State
Session: Oral Abstract Session: Mycotic Diseases: Pathogenesis and Prevention
Saturday, October 10, 2015: 8:30 AM
Room: 32--ABC
Background: Systemic candidiasis is the leading cause of nosocomial bloodstream infection in the US with mortality that exceeds 40% despite antifungal therapy. We previously identified Cx3cr1, a monocyte/macrophage-targeted chemokine receptor that has a single ligand, fractalkine (FKN), as a critical mediator of host survival in a mouse model of systemic candidiasis by promoting macrophage survival, accumulation and fungal killing in infected tissues (Lionakis et al., JCI, 2013). Consistent with the mouse findings, the mutant CX3CR1 allele, CX3CR1-M280, was an independent risk factor for developing systemic candidiasis and worse outcome after infection in two patient cohorts. Yet, how this mutation predisposes humans to systemic candidiasis is unknown. To address this question, we investigated the effects of the CX3CR1-M280 mutation on the biological function of primary human monocytes.

Methods: We magnetically sorted CD14+monocytes from whole blood for functional assays including calcium flux to assess receptor signaling, flow cytometry to assess receptor expression and cell apoptosis/death, alamarBlue-based fungal killing, and cytokine production post-fungal stimulation measured by Luminex array. Data were analyzed using a Mann-Whitney or t-test where appropriate. 

Results: We screened 108 healthy Caucasian donors from the NIH Blood Bank and identified 67 wild-type (WT), 37 CX3CR1-M280 heterozygous, and 4 CX3CR1-M280 homozygous donors. We found that CX3CR1-M280 homozygotes exhibit a 50% decrease in FKN-induced receptor signaling (p=0.0002), which was not due to decreased CX3CR1 surface expression. Consistent with our prior mouse findings that Cx3cr1 mediates anti-apoptotic effects in macrophages, we found that FKN was able to rescue serum starvation-induced monocyte death in WT donors (p=0.0031) but not in CX3CR1-M280 homozygotes (p=ns). This survival defect in CX3CR1-M280 homozygote monocytes may account for the observed steady state monocytopenia seen in these donors (p=0.0178). Instead, CX3CR1-M280 did not impair monocyte fungal killing or production of pro-inflammatory cytokines in response to heat-killed Candida.

Conclusion: Together, these data show that the CX3CR1-M280 allele impairs FKN-induced CX3CR1 signaling in human monocytes and results in defective monocyte survival and monocytopenia.

Amanda Collar, B.S., Muthulekha Swamydas, PhD and Michail Lionakis, MD, ScD, Fungal Pathogenesis Unit, Laboratory of Clinical Infectious Diseases, Niaid, National Institutes of Health, Bethesda, MD

Disclosures:

A. Collar, None

M. Swamydas, None

M. Lionakis, None

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