P-selectin contributes to severe experimental malaria but is not required for leukocyte adhesion to brain microvasculature

Abstract

Plasmodium berghei-infected mice, a well-recognized model of experimental cerebral malaria (ECM), exhibit many of the hallmarks of a systemic inflammatory response, with organ damage in brain, lung, and kidneys. Identification of the molecules mediating pathogenesis of the inflammatory response, such as leukocyte adhesion, may lead to new therapies. Indeed, mice lacking the cell adhesion molecule P-selectin were significantly (P = 0.005) protected from death due to P. berghei malaria compared with C57BL/6 controls despite similar parasitemia (P = 0.6) being found in both groups of mice. P-selectin levels assessed by the quantitative dual radiolabeled monoclonal antibody technique increased significantly (P < 0.05) in several organs in C57BL/6 mice infected with P. berghei, supporting the concept of a systemic inflammatory response mediating malarial pathogenesis. Intravital microscopic analysis of the brain microvasculature demonstrated significant (P < 0.001) leukocyte rolling and adhesion in brain venules of P. berghei-infected mice compared with those found in uninfected controls. The maximum leukocyte adhesion occurred on day 6 of P. berghei infection, when the mice become moribund and exhibit marked vascular leakage into the brain, lung, and heart. However, P-selectin levels were significantly (P < 0.005) increased in brain, lung, and kidneys during P. berghei malaria in ECM-resistant BALB/c mice compared with those found in uninfected BALB/c controls, indicating that increased P-selectin alone is not sufficient to mediate malarial pathogenesis. Leukocyte adhesion to brain microvessels of P-selectin-deficient mice with P. berghei malaria was similar to that observed in control mice. Collectively, these results indicate that P-selectin is important for the development of malarial pathogenesis but is not required for leukocyte adhesion in brain.

FIG. 1.

Effect of P-selectin deficiency (A) on the survival of groups of mice during P. berghei malaria. The average parasitemia and standard deviation of the groups of mice are shown (B).

FIG. 2.

P. berghei parasitemia in groups of either ECM-susceptible C57BL/6 mice (▪)or in ECM-resistant BALB/c mice (○) (A) and P-selectin in brain (B), lung (C), and kidney (D) during the course of P. berghei malaria in either ECM-susceptible C57BL/6 mice (filled bar) or in ECM-resistant BALB/c mice (open bar). For C57BL/6 mice, n = 8, 4, and 4 on days 0, 4, and 6, respectively; for BALB/c mice, n = 4 for all groups. Shown is the mean plus or minus standard deviation. ∼, P < 0.05; #, P < 0.005; and *, P < 0.0005 for the comparison with uninfected strain-matched controls. N.D., not done.

FIG. 3.

Analysis of P-selectin levels in brain in uninfected C57BL/6 mice (A) and on day 6 of P. berghei infection (B) by fluorescence microscopy. Magnification (A and B), ×8.8. Arrow marks blood vessels in each section.

FIG. 4.

Rolling (A) and adhesion (B) of leukocytes in brain microvasculature during the course of P. berghei malaria in ECM-susceptible C57BL/6 mice. Shown is the mean plus or minus SEM. At each time point, the brain circulation of five mice (five vessels) was assessed by intravital microscopy. A statistically significant (P < 0.05) difference in leukocyte rolling or adhesion in small (*) and large (#) postcapillary venules during the course of P. berghei infection was compared with leukocyte rolling and adhesion in uninfected C57BL/6 mice.

FIG. 5.

Rolling (A) and adhesion (B) of leukocytes to brain endothelium of mice lacking P-selectin on day 6 of P. berghei malaria. We assessed the mean plus or minus SEM of leukocyte rolling and adherence by using intravital microscopy in five brain venules of four infected and four uninfected P-selectin-deficient mice. The rolling and adherence of leukocytes in C57BL/6 controls on day 0 and day 6 of infection in panels A and B are repeated from Fig. 4 for comparison. Shown is a statistically significant (P < 0.05) difference in leukocyte rolling or adhesion from that found in P-selectin-deficient mice (*) or C57BL/6 controls (#) on day 6 of P. berghei infection (A and B).