Fungal dissemination into the bloodstream is a critical step leading to invasive fungal infections. Here, using intravital imaging, we show that Kupffer cells (KCs) in the liver have a prominent function in the capture of circulating Cryptococcus neoformans and Candida albicans, thereby reducing fungal dissemination to target organs. Complement C3 but not C5, and complement receptor CRIg but not CR3, are involved in capture of C. neoformans. Internalization of C. neoformans by KCs is subsequently mediated by multiple receptors, including CR3, CRIg, and scavenger receptors, which work synergistically along with C5aR signaling. Following phagocytosis, the growth of C. neoformans is inhibited by KCs in an IFN-γ independent manner. Thus, the liver filters disseminating fungi from circulation via KCs, providing a mechanistic explanation for the enhanced risk of cryptococcosis among individuals with liver diseases, and suggesting a therapeutic strategy to prevent fungal dissemination through enhancing KC functions.

MicroRNAs (miRNAs) are present in serum and have the potential to serve as disease biomarkers. As such, it is important to explore the clinical value of miRNAs in serum as biomarkers for ischemic stroke (IS) and cast light on the pathogenesis of IS. In this study, we screened differentially expressed serum miRNAs from IS and normal people by miRNA microarray analysis, and validated the expression of candidate miRNAs using quantitative reverse-transcriptase polymerase chain reaction assays. Furthermore, we performed gene ontology (GO) and Kyoto encyclopedia of genes and genomes (KEGG) pathway analyses to disclose functional enrichment of genes predicted to be regulated by the differentially expressed miRNAs. Notably, our results revealed that 115 miRNAs were differentially expressed in IS, among which miR-32-3p, miR-106-5p, and miR-532-5p were first found to be associated with IS. In addition, GO and KEGG pathway analyses showed that genes predicted to be regulated by differentially expressed miRNAs were significantly enriched in several related biological process and pathways, including axon guidance, glioma, MAPK signaling, mammalian target of rapamycin signaling, and ErbB-signaling pathway. In conclusion, we identified the changed expression pattern of miRNAs in IS. Serum miR-32-3p, miR-106-5p, miR-1246, and miR-532-5p may serve as potential diagnostic biomarkers for IS. Our results also demonstrate a novel role for miRNAs in the pathogenesis of IS.

BackgroundChemokines and chemokine receptors cooperate to promote immune cell recruitment to the central nervous system (CNS). In this study, we investigated the roles of CXCR2 and CXCL1 in leukocyte recruitment to the CNS using a murine model of neuroinflammation.MethodsWild-type (WT), CXCL1−/−, and CXCR2−/− mice each received an intracerebroventricular (i.c.v.) injection of lipopolysaccharide (LPS). Esterase staining and intravital microscopy were performed to examine neutrophil recruitment to the brain. To assess endothelial activation in these mice, the expression of adhesion molecules was measured via quantitative real-time polymerase chain reaction (PCR) and Western blotting. To identify the cellular source of functional CXCR2, chimeric mice were generated by transferring bone marrow cells between the WT and CXCR2−/− mice.ResultsExpression levels of the chemokines CXCL1, CXCL2, and CXCL5 were significantly increased in the brain following the i.c.v. injection of LPS. CXCR2 or CXCL1 deficiency blocked neutrophil infiltration and leukocyte recruitment in the cerebral microvessels. In the CXCR2−/− and CXCL1−/− mice, the cerebral endothelial expression of adhesion molecules such as P-selectin and VCAM-1 was dramatically reduced. Furthermore, the bone marrow transfer experiments demonstrated that CXCR2 expression on CNS-residing cells is essential for cerebral endothelial activation and leukocyte recruitment. Compared with microglia, cultured astrocytes secreted a much higher level of CXCL1 in vitro. Astrocyte culture conditioned medium significantly increased the expression of VCAM-1 and ICAM-1 in cerebral endothelial cells in a CXCR2-dependent manner. Additionally, CXCR2 messenger RNA (mRNA) expression in cerebral endothelial cells but not in microglia or astrocytes was increased following tumor necrosis factor-α (TNF-α) stimulation. The intravenous injection of the CXCR2 antagonist SB225002 significantly inhibited endothelial activation and leukocyte recruitment to cerebral microvessels.ConclusionsCXCL1 secreted by astrocytes and endothelial CXCR2 play essential roles in cerebral endothelial activation and subsequent leukocyte recruitment during neuroinflammation.

Although neutrophils are typically the first immune cells attracted to an infection site, little is known about how neutrophils dynamically interact with invading pathogens in vivo. Here, with the use of intravital microscopy, we demonstrate that neutrophils migrate to the arrested Cryptococcus neoformans, a leading agent to cause meningoencephalitis, in the brain microvasculature. Following interactions with C. neoformans, neutrophils were seen to internalize the organism and then circulate back into the bloodstream, resulting in a direct removal of the organism from the endothelial surface before its transmigration into the brain parenchyma. C. neoformans infection led to enhanced expression of adhesion molecules macrophage 1 antigen on neutrophils and ICAM-1 on brain endothelial cells. Depletion of neutrophils enhanced the brain fungal burden. Complement C3 was critically involved in the recognition of C. neoformans by neutrophils and subsequent clearance of the organism from the brain. Together, our finding of the direct removal of C. neoformans by neutrophils from its arrested site may represent a novel mechanism of host defense in the brain, in addition to the known, direct killing of microorganisms at the infection sites. These data are the first to characterize directly the dynamic interactions of leukocytes with a microbe in the brain of a living animal.

Extrapulmonary dissemination of Cryptococcus neoformans is one of the most critical steps in the development of meningoencephalitis. Here, we report that clearance of the disseminating C. neoformans occurs within the brain microvasculature. Interestingly, the efficiency of the intravascular clearance in the brain is not as great as that in the lung. Intravascular clearance is mainly mediated by neutrophils, and complement C5a receptor signaling is crucial via mediating neutrophil recruitment in the vasculature. C. neoformans-stimulated actin polymerization of neutrophils is critically involved in their recruitment to the lung, which is associated with the unique vascular structure detected in the lung. The relative lower efficiency of fungal clearance in the brain vasculature correlates with less efficient recruitment of neutrophils. Accordingly, intravascular clearance of C. neoformans in the brain could be remarkably improved by increasing the recruitment of neutrophils. We conclude that neutrophils have the ability to eliminate C. neoformans arrested in the vasculature. However, insufficient recruitment of neutrophils limited the optimal clearance of the organism in the brain. These results imply that a therapeutic strategy aimed at enhancing the accumulation of neutrophils could help prevent cryptococcal meningoencephalitis.

Acute lung injury (ALI) is the leading cause of morbidity and mortality in critically ill patients. Neutrophil extracellular traps (NETs) have been well documented in the ALI model of bacterial infection. In the present study, we demonstrated that poly I:C could induce pulmonary NETs. Upon poly I:C intratracheal inoculation, neutrophil infiltration in the bronchoalveolar lavage fluid (BALF) was significantly increased. Furthermore, the inflammatory cytokines IL-1β, IL-6, and TNF-α in the lung were also significantly elevated. Neutrophil depletion abolished NETs and decreased both neutrophil infiltration and IL-1β in the lung. As expected, DNase I, an inhibitor of MPO and NADPH, decreased pulmonary inflammation and NETs. Blocking of the poly I:C receptor TLR3 reduced lung inflammation and NETs. The MAPK kinase inhibitor p38 diminished the formation of NETs and restored the expression of the tight junction protein claudin-5 in the mouse lung when challenged with poly I:C. In summary, poly I:C induced the formation of pulmonary NETs and ALI, which may be associated with the activation of p38 MAPK and the decreased expression of claudin-5.

No abstract

No abstract