Platelets as Immune Cells in Infectious Diseases

Cornelia Speth; Jürgen Löffler; Sven Krappmann; Cornelia Lass-Flörl; Günter Rambach


Future Microbiol. 2013;8(11):1431-1451. 

In This Article

Interaction of Platelets With Bacterial Pathogens

Bacteria enter the bloodstream in response to infectious insults, via surgical procedures or indwelling catheters, and sepsis occurs in up to 6–30% of all intensive care unit patients.[94,95] The balance of pro- and anti-inflammatory reactions critically determines the severity and lethality of sepsis, a fact that turns the spotlight onto understanding the immune reaction in order to develop appropriate therapies.[96]

Bacterial contact with the platelets is a key event for the pathogenesis of sepsis, as deduced from the correlation between sepsis outcome and decreased platelet numbers: the more profound the thrombocytopenia the more severe the sepsis and the greater the mortality of affected patients.[97–99]

There are two main causes for the bacteria-induced thrombocytopenia:

  • Bacteria induce platelet activation. Activated platelets show shortened survival and are targets of phagocytic clearance;

  • Bacterial compounds induce apoptosis and cytotoxic effects in platelets. Both main mechanisms can complement and overlay each other and are described below in more detail (see also Box 2 & Figures 2 & 3).

Additional mechanisms are known for selected bacteria and listed in Box 2; however, exhaustive discussion goes beyond the scope of this article. Although they further augment and reinforce the thrombocytopenia in affected sepsis patients.

The mechanisms that finally lead to the loss of platelets and their attributed immune functions can decisively compromise the antibacterial immune defense (Box 1). The subsequent thrombocytopenia might be regarded as a normal consequence of immune exhaustion. However, it might also be regarded as targeted bacterial evasion strategies to eliminate a central innate immune cell from circulation with a kinetic that exceeds de novo generation in the bone marrow (Box 2 & Figure 3); thus, thrombocytopenia represents a status that helps the bacteria to survive in the bloodstream. There is yet a third aspect: those mechanisms of platelet activation or apoptosis that finally result in thrombocytopenia can also lead to thrombosis, thus contributing to tissue damage in the pathogenesis of bacterial infections.

Figure 3.

Mechanisms for pathogen-induced thrombocytopenia.
Pathogens can interfere with platelet production in the bone marrow by either infecting the precursor megakaryocytes (blue), by induction of autoimmune antibodies that trigger elimination of the megakaryocytes or by disturbance of thrombopoiesis via dysregulated cytokines or thrombopoietin. Pathogens can also target circulating platelets and induce apoptosis or cell lysis. In addition, platelet activation after contact with the microorganisms shortens their lifespan. Furthermore, pathogens can induce removal of platelets from the circulation by stimulating their sequestration in organs or by triggering their clearance by phagocytes (see text and Box 2 for further details).

Platelet Activation as a Putative Reason for Thrombocytopenia

Platelets have a broad spectrum of tools to sense the presence of bacteria and/or their secreted products and react on these signals with a multistep activation process. Consequences of this platelet activation might be their deposition in microvascular thrombi or their clearance from the circulation by phagocytes.[100,101] Both processes give rise to reduced platelet numbers in the circulation and can explain sepsis-induced thrombocytopenia (Figure 3).

Bacteria-induced platelet activation can be triggered by several mechanisms. One possibility is the adhesion of bacteria to platelet membrane receptors as described above, such as GPIIb-IIIa, GPIb, complement receptors, FcγRIIa or TLRs.[23] This adhesion can include a direct interaction between the bacterial surface structures and a platelet receptor, but also an indirect mechanism when bacteria are covered by plasma proteins (fibrinogen, fibronectin, vWF, complement factors and IgG) and attach to platelets via the corresponding receptors.[23] A bacterial species can also use both pathways to get in contact with the platelets, demonstrated, for example, by S. aureus and Streptococcus sanguinis.[23]

The platelet TLR4 is one important pattern recognition receptor and binds to lipopolysaccharides from Gram-negative bacteria. Studies using an animal model for bacterial sepsis revealed that TLR4 is particularly relevant for bacteria-induced platelet activation with subsequent thrombocytopenia.[102]

Bacteria can stimulate platelets not only as a consequence of adhesion, but also via secretion of soluble compounds that bind to platelet surface structures and trigger activation. This mechanism is described for S. aureus, Streptococcus pneumoniae, Streptococcus pyogenes and Porphyromonas gingivalis. The latter, a Gram-negative oral pathogen that is involved in the pathogenesis of periodontitis, secretes a family of cysteine proteases that stimulate the protease-activated receptors 1 and 4 on the platelet surface with subsequent increase in intracellular calcium and platelet aggregation.[103] Another example is Shiga toxin-producing Escherichia coli. Infection by this pathogen is associated with hemolytic uremic syndrome[104] and a key feature of this disease is thrombocytopenia. Culture filtrates of Shiga toxin-producing E. coli that contained all released bacterial compounds specifically induced downregulation of CD47, the receptor for thrombospondin-1, and reduced surface CD47 correlated with platelet activation and phagocytosis by macrophages.[105]

Activated platelets, as they occur in bacterial infection and infection-induced inflammation, represent a threat to homeostasis, since exposition of phosphatidylserine and release of granule contents might exaggeratedly trigger the coagulation. Therefore neutrophils attach to activated platelets by several surface receptors and clear them from circulation.[71] The initial binding event involves P-selectin exposed on stimulated platelets and the corresponding counter-receptor PSGL-1 on resting neutrophils. Subsequently, neutrophils upregulate the active form of αMβ2, which binds to fibrinogen on the surface of activated platelets. The final engulfment of the platelets by neutrophils is dependent on the recognition of platelet phosphatidylserine.[71,101]

Bacteria-induced Apoptosis & Disintegration of Platelets as a Putative Reason for Thrombocytopenia

The second putative reason for thrombocytopenia in sepsis is the damage of platelets, when bacteria either initiate the apoptotic program in platelets or disturb platelet integrity (see Figure 3).

The common sepsis inducers E. coli and S. aureus as well as their secretion products α-toxin and α-hemolysin, respectively, are capable of inducing apoptosis in platelets. Both bacteria and their secreted toxins can trigger the calpain-mediated degradation of the platelet protein Bcl-xL.[106] Since Bcl-xL represents a crucial factor for platelet survival, its proteolytic elimination is a key step for initiating the apoptotic program. Furthermore, the bacterial cell wall component peptidoglycan, purified from S. aureus, is also able to trigger apoptotic processes like mitochondrial depolarization, caspase-3 activation and membrane scrambling.[107]

Interestingly, E. coli and S. aureus can affect platelet integrity by another mechanism and thus enhance the level of thrombocytopenia. Their bacterial toxins α-toxin and α-hemolysin stimulate disturbances in the platelet membrane and thus act directly in a cytotoxic manner. α-toxin firmly binds to target membranes and forms ring-structured toxin oligomers, thus causing membrane damage and calcium influx and mimicking the effect of an ionophore.[108] Platelets might be lysed directly,[109] or may be affected by calcium-driven stimulation with subsequent clearance by neutrophils. A similar pore-forming mechanism was detected for the bacterial toxins streptolysin O from S. pyogenes, where resulting complexes between platelets and neutrophils impeded blood flow and lead to ischemia and tissue necrosis, and for pneumolysin from S. pneumonia.[110,111]

Platelets' Benefit in Bacterial Infection: Direct Attack & Guidance of Other Immune Cells

Bacteria-induced platelet activation, apoptosis or disintegration are all processes that result in the release of microbicidal peptides and thus contribute to pathogen elimination. Granule-stored PMPs are secreted by platelet activation, whereas apoptosis or disintegration lead to release of PMP stored in both granules and cytoplasm.

This PMP secretion is a key contribution of platelets to the immune defense against bacterial pathogens. Its relevance was unambiguously proven in vivo using animal models for bacteria-induced infective endocarditis (IE), where thrombocytopenic animals were more susceptible to IE than controls.[112] Detailed in vivo studies directly underlined the efficiency of PMP for antibacterial defense, since rabbits where PMP had been neutralized by antibodies were more susceptible to streptococcal IE than controls.[51] Further experiments revealed an inverse correlation between bacterial virulence and the stimulation of PMP release: those S. aureus variants that activated platelets to a high extent showed reduced virulence. In particular, the microbicidal proteins thrombin-induced platelet microbicidal protein (tPMP)-1, -2 and -3 were supposed to participate in this effect.[109] Strains of S. aureus that were insusceptible to the thrombocidin tPMP caused more severe IE and more effective hematogenous dissemination than did tPMP-susceptible strains.[113]

Beside tPMP, the thrombocidins TC-1 and TC-2 also have the potency to inhibit the growth of bacteria in vitro, such as E. coli, Bacillus subtilis and S. aureus.[51,114] The IE model confirmed these results in vivo, demonstrating that TC-1 and TC-2 are involved in the clearance of Streptococcus viridans from the endocardial surface; bacterial strains with low TC-1/TC-2 susceptibility could persist on the surface, whereas highly susceptible bacteria were rapidly eliminated.[115,116]

Human platelets have recently been described to express human β-defensins (hBD). The defensin hBD-1 is capable if impairing the growth of S. aureus, but also inducing NET formation by neutrophils. Interestingly, the β-defensins are stored in extragranular compartments, and platelet activation with release of the granule content does not result in secretion of defensins. Instead, bacterial toxins such as α-toxin by S. aureus, which induces apoptosis and permeabilizes the cell membrane, efficiently trigger the release of hBD-1 from the platelets.[53]

The relevance of PMP in general is further underlined by the fact that many bacteria developed corresponding resistance mechanisms and thus acquired protection against inactivation (Box 2 & Figure 2). Since PMPs have to cross the bacterial envelope to reach the cytoplasmic target membrane, the permeability and the physicochemical properties of the envelope are key determinants of bacterial susceptibility. Accordingly, countermechanisms against PMP attack include: limitation of attachment by covalent modification of anionic molecules in the membrane; active transport of the peptides out of the bacterial cell; alteration of membrane fluidity; and proteolytic cleavage of the peptides.[117,118] Reduced susceptibility of clinical S. aureus strains against tPMP-1 is associated with increased membrane fluidity and with the QacA efflux pump.[118,119] Furthermore, the hydrophilic–lipophilic balance and the net reduction in surface-positive charge of bacteria correlate with enhanced killing by tPMPs.[120,121]

Figure 2.

Mechanisms for platelet evasion and exploitation exerted by microorganisms.
Pathogens can induce elimination of platelets (red) from the bloodstream or impair their activation and/or aggregation. Some pathogens also developed resistance against platelet-derived microbicidal peptides. Platelets can even be exploited by pathogens and mediate their immune evasion or support their dissemination (see text for further details).

Beside PMP release, further effects of platelets on neutrophil stimulation, T- and B-cell activation and maturation of dendritic cells were demonstrated (Box 1 & Figure 1), can be postulated or have been described to decisively contribute to an efficient antibacterial immune defense.

The 'Dark Side' of Platelets in Bacterial Infections

Platelet activation after contact with bacterial pathogens or their secreted compounds can result in detrimental events such as thrombosis and thus might critically contribute to pathogenesis. Thrombotic manifestations can be localized or occur widespread in the blood vessels of different organs with organ dysfunction as putative consequence. The most severe form is a disseminated intravascular coagulation, characterized by intense microvascular thrombosis, consumption of platelets and coagulation factors, and bleeding.[122] Bacteria-induced platelet activation directly leads to aggregation, and thrombosis is further aggravated by the fact that platelets augment NET formation with deposition of platelets and neutrophils.[123]

Another negative effect of platelets in bacterial infections is the enhancement of biofilm formation. Platelets were demonstrated in vitro and in vivo to be essential for biofilm generation by streptococci.[124] The biofilms on injured heart valves were composed of bacteria embedded in platelet aggregates; the bacteria in the biofilm were able to induce further platelet aggregation, which facilitates the formation of multilayer biofilms.[124] Thus, platelets are hijacked by the streptococci to support biofilm formation and to subsequently favor survival of the bacteria in the host. In addition, bacteria in this platelet-containing biofilm were refractory to antibiotic treatment.[124]