The Role And Mechanism Of Monocytes In Mice Brain After Cryptococcus Neoformans Infection

Cryptococcus neoformans(C.neoformans)is an encapsulated yeast that causes a life-threatening illness in immunocompromised individuals,such as patients with AIDS,organ transplant recipients,and those treated with immunosuppressive therapies.Although the infection starts in the lung,cryptococcosis commonly presents as meningoencephalitis,which is one of the most common infections of the central nervous system.Transmigration of C.neoformans across the blood-brain barrier(BBB)is believed to be one of the most critical steps in the development of cryptococcal meningoencephalitis.Three hypotheses in BBB crossing field:paracellular pathway,transcellular pathway and Trojan horse pathway.The barrier,however,is open to some immune cells under physiological or pathological conditions allowing neurotropic pathogens to gain access to the brain parenchyma within immune cells(Trojan horse theory).However,there is little direct evidence demonstrating Trojan horse in vivo.Questions still remain as to how C.neoformans migrates to the brain across the BBB in vivo and what is the underlying mechanism(s).Answering these questions is fundamental for understanding cryptococcal pathogenesis,because brain invasion is the hallmark feature of this disease and meningoencephalitis is the major and most lethal complication of cryptococcosis.In contrast to previous in vitro studies,we have developed a novel in vivo model system based on multiple novel approaches to directly investigate the brain invasion by C.neoformans in vivo such as the subset of the monocyte,adhesion molecules and cytokines.Our study was based on mice model with C.neoformans infection.In vivo system was used to study traversal of the BBB by C neoformans,in combination with flow cytometry,qRT-PCR,immunohistochemical method and etc.Firstly,C.neoformans H99 strain was used to confirm the existence of "Trojan horse" mechanism in the process of brain invasion.Secondly,it was showed monocytes from the mouse brain were recruited into the postcapillary venules of the brain to roll and adhere to the vascular endothelial cells after infection.Thirdly,the monocyte recruitment was mainly dependent on VCAM-1/VLA-4 mediated rolling and ICAM-1/CD11a mediated adhesion,and the TNFR signaling pathway played a crucial role in monocyte recruitment by up-regulating VLA-4 molecules on monocytes.Fourthly,through in vivo fluorescence microscopy observation,it was found that the most of the mononuclear cells phagocytic C.neoformans in the brain of mice were the Ly6Clow subgroup,and a large number of Ly6Clow cells engulfed C.neoformans,crawled with the bacteria and adhered to the cerebral vascular endodermis,and gradually migrated to the brain parenchyma.Collectively,current study provides direct evidence for the Trojan horse hypothesis suggesting that Ly6Clow monocytes as the major player mediating this process,which can be considered as a potential immunotherapy target for treating meningoencephalitis caused by C.neoformans.