Human platelets efficiently kill IgG-opsonized E. coli

Abstract

Platelets are known contributors of hemostasis but have recently been shown to be important in inflammation and infectious diseases. Moreover, thrombocytopenia is often observed in patients with sepsis. We previously reported that platelets actively phagocytosed IgG-coated latex beads. In this study, the capacity of human platelets to participate in host defense against bacterial infections was determined by assessing their ability to kill Escherichia coli. Washed human platelets were incubated with unopsonized or IgG-opsonized E. coli and evaluated for binding and killing of E. coli. We found that although both unopsonized and IgG-opsonized E. coli were associated with platelets, only IgG-opsonized E. coli were efficiently killed unless platelets were activated by a potent agonist. The bactericidal activity was dependent on FcγRIIA, was sensitive to cytochalasin D, but was not due to reactive oxygen metabolites. These data suggest that platelets may play an important role in protection against infection.

Figure 1. Association of E. coli with human platelets

Washed human platelets were incubated with E. coli K12 strain at a MOI of 1 for 60 minutes. A: Plates were washed, stained with BacLight Live-Dead stain (Syto 9 and propidium iodide) and imaged using epifluorescence microscopy. Representative PI-positive E. coli are indicated by arrows and PI-negative E. coli by arrow-heads. B: Numbers of platelet-bound E. coli per field of view were counted and the % PI-negative E. coli was calculated. * p<0.05

Figure 2

Human platelet killing of E. coli. A: Washed human platelets were incubated with E. coli K12 strain at a MOI=1 for 60 minutes at 37°C in triplicate wells of 12-well plates. Platelets were lysed and samples were serially diluted and plated on duplicate LB agar, incubated overnight at 37°C and colony forming units were counted and expressed as CFU/ml. B: Experiment performed exactly as in (A) except that E. coli were opsonized with IgG. Bars are mean ± SE (n=3). * p<0.05

Figure 3

Killing of E. coli by activated platelets. IgG-opsonized or non-opsonized E. coli were incubated with 0.1 unit/ml thrombin in the presence or absence of platelets for 60 minutes at 37°C in triplicate wells of 12-well plates. Platelets were lysed and samples were serially diluted and plated on LB agar, incubated overnight at 37°C and colony forming units were counted and expressed as CFU/ml. Bars are mean ± SE (n=3). * p<0.05 NS=not significant