The Role Of MR In Immune Evasion Of Cryptococcal Infection And S100A10in Promoting Cryptococcus Neoformans Traversal Across Blood-brain Barrier

Cryptococcus neoformans(C.neoformans) is an opportunistic fungal pathogen thatcauses central nervous system and pulmonary disease among immunocompromisedpatients. Despite the availability of antifungal agents with anticryptococcal activity, themortality and treatment failure rates associated with cryptococcosis remain unacceptablyhigh. In cryptococcal infection, one of two things happens: either the host immuneresponse clears the infection, or C.neoformans successfully evades this response and,without treatment, will disseminate with potentially fatal consequences. An additionalpossible outcome is that cryptococcal infection may be contained but not cleared andbecomes latent. There are several ways for C.neoformans evade the body’s immuneresponse:1,"Phagosome extrusion" and intracellular replication;2, Lysis of macrophagesor inducing macrophage apoptosis;3, Avoiding being engulfed by macrophages;4,Avoiding immune recognition: shielding PAMPs and physical barriers;5,Inducing immunesuppression.Mannose receptor, also known as CD206, mainly expressed by macrophages anddendritic cells, are the C type lectin like receptor and pattern recognition receptors.MR cannot only recognize and bind to glycoprotein of pathogens and induce innate immuneresponse, but also can promote T cell activation and cytokine release, starting the adaptiveimmune response. C.neoformans mannoprotein(MP) is the ligand of MR, MP caneffectively activate the T cell immune response and enhance the immunity againstcryptococcal infection. MP immunized animals exhibited prolonged survival and reducedbrain and kidney fungal loads following intravenous challenge with C. neoformans, anddendritic cells can phagocytize C. neoformans via mannose receptors and Fc receptor IIand present antigens to T lymphocytes, inducing the antifungal immune response.Researchs on C.neoformans evading the immune response are mainly concentrated oncryptococcal capsular genes and intracellular replication, yet there is no report aboutC.neoformans evading immune response by regulating some important receptors expression.Because of the importance of MR in anti cryptococcal infection, we observed howC.neoformans regulate the expression of MR and its pathophysiological significance.C. neoformans is encapsulated yeast responsible for life threatening central nervoussystem (CNS) infections primarily in immunocompromised patients. How Cryptococcus addict to the central nervous system may involve the following factors:1, Trojanmechanism which depends on the the monocytes;2, Activating Rho GTPases;3,Activating urokinase plasminogen system;4, C. neoformans CPS1gene hyaluronicacid CD44;5, Urease. Our previous studies found that Cryptococcus can up regulate theexpression of S100A10in human umbilical vein endothelial cells, and suppression ofS100A10expression was associated with significantly reduced rates of phagocytosis andbudding of C. neoformans, the capsule of intracellular C. neoformans also became thickerafter S100A10was knocked down, indicating that S100A10may involved in C.neoformans invading brain microvascular endothelial cells. C. neoformans can invade theblood brain barrier though plasminogen system which was activated by UPA, and UPAactivating the plasminogen needs the expression of S100A10on endothelial cells,therefore, we intend to clarify the relationship between Cryptococcus upregulating theexpression of S100A10and its traversal across the blood brain barrier.1、The mechanisms of Cryptococcus regulating the expression of MRFirst, we found that Cryptococcus can down regulate the expression of MR in DCsand macrophages at both gene level and protein level. The downregulation of MR was notdue to its sheding into soluble MR(SMR). Interestingly, when using the Transwellchamber to separate the fungus and cells we did not observe such a change, indicating thatCryptococcus down regulate MR expression depending on the direct contact between thefungus and cells, and regardless of the vitality of the fungus. Further studies showed thatthe ligand MP can down regulate MR expression,it was similar to LPS down regulatingTLR4expression which was considered to be responsible for endotoxin tolerance. Somaybe this is the new mechanism used by Cryptococcus to evade the immune response.2、 The signal pathway of MRBecause the MR cytoplasmic tail does not contain intracellular ITAM signaling motifs, wehypothesized that direct interaction of other receptors and MR may happen,these receptorsmay form a functional complex, whereby MR may capture MP at the cell membrane,andthe other receptors signaling motif of the associated receptors may provide the necessaryintracellular signal transduction via NF κB and MAPKs to promote cytokine release. First,we observed that MP can activate MAPK and NF κB pathway, it’s shown that the MP canpromote p38, ERK, JNK and p65phosphorylation, knockout of TLR2significantlyaffected the phosphorylation of MAPK and NF κB, demonstrating that MR signaltransduction need the expression of TLR2. And it’s shown that knockout of TLR2 significantly reduced the secretion of pro inflammatory cytokines such as IL6,TNF α andIL1β and affected the ability of macrophages in killing Cryptococcus.Finally,we foundthat p38inhibitors can suppress the down regulation of MR induced by Cryptococcus andMP,therefore, Cryptococcus and MP down regulate the expression of MR depends on theactivation of p38MAPK.3、The significance of down regulation of MR induced by CryptococcusFirst,we found that overexpression of MR enhanced the ability of DCs inpresenting MP antigen and significantly enhanced the activation of T cells, demonstratingthat the antifungal immunity of DCs was positively correlated with the level of MR. Next,we observed that silencing MR reduced the ability of macrophage in phagocytizing andkilling C. neoformans, indicating that the antifungal immunity of macrophages was alsopositively correlated with the level of MR. Therefore, we supposed that thedown regulation of MR expression induced by Cryptococcus suppressed the antifungalimmunity,which may provided new mechanism for Cryptococcus immune evasion.4、S100A10with cryptococcal addicted central MechanismOur previous studies showed that Cryptococcus can up regulate the expression ofS100A10in human umbilical vein endothelial cells,and silencing S100A10significantlyreduced the ability of endothelial cells in phagocytizing Cryptococcus. Additionally,suppression of S100A10expression was associated with significantly reduced rate ofphagocytosis and budding of C. neoformans. The capsule of intracellular C. neoformansalso became thicker after S100A10was knocked down, indicating that S100A10may playan important role in transpenetration of C. neoformans across the vascular endothelia.Inthis research,we found that C. neoformans can up regulate the expression of S100A10inmouse brain microvascular endothelial cells, silencing S100A10reduced the ability ofplasminogen coated C. neoformans in invading the blood brain barrier, but it didn’t affectthe ability of C. neoformans not coated with plasminogen in crossing the blood brainbarrier. Further we found that silencing S100A10reduced the activation of urokinase andplasminogen, therefore, the activated S100A10urokinase plasminogen system may beinvolved in C. neoformans addicting to central nervous system.In summary, we found that C. neoformans can downregulate the expression of MRand the reduced MR expression may be responsible for C. neoformans immuneevasion.The upregulation of S100A10may be involved in C. neoformans addicting tocentral nervous system.