The adult mammalian CNS has a limited capacity for nerve regeneration and structural plasticity. The presence of glia-derived inhibitory factors myelin-associated glycoprotein (MAG) and Nogo-A have been suggested to provide a nonpermissive environment for elongating nerve fibers. In particular, Nogo-A, an integral membrane protein predominantly expressed by oligodendrocytes, has been demonstrated to impair neurite growth in vitro and in vivo. Structure function analysis revealed that Nogo-A protein contains at least two active domains, NiG and Nogo-66, with diverse effects on neurite outgrowth and cell spreading. We now provide evidence that these inhibitory domains mediate their effects via an antagonistic regulation of the small GTPases RhoA and Rac1, resulting in activation of RhoA and suppression of Rac1. By inactivating RhoA with C3 transferase or the downstream effector Rho-kinase ROCK with, the inhibitory effects of both Nogo-A fragments and MAG on neurite outgrowth and oligodendrocyte-mediated growth cone collapse were abolished. Furthermore, we show that the recently cloned receptor for Nogo-66 and MAG, NgR, is not necessary for either NiG- or MAG-induced RhoA activation.

The absence of fiber regrowth in the injured mammalian CNS is influenced by several different factors and mechanisms. Besides the nonconducive properties of the glial scar tissue that forms around the lesion site, individual molecules present in CNS myelin and expressed by oligodendrocytes, such as NI-35/NI-250, bNI-220, and myelin-associated glycoprotein (MAG), have been isolated and shown to inhibit axonal growth. Here, we report an additional neurite growth-inhibitory activity purified from bovine spinal cord myelin that is not related to bNI-220 or MAG. This activity can be ascribed to the presence of two chondroitin sulfate proteoglycans (CSPGs), brevican and the brain-specific versican V2 splice variant. Neurite outgrowth of neonatal cerebellar granule cells and of dorsal root ganglion neurons in vitro was strongly inhibited by this myelin fraction enriched in CSPGs. Immunohistochemical staining revealed that brevican and versican V2 are present on the surfaces of differentiated oligodendrocytes. We provide evidence that treatment of oligodendrocytes with the proteoglycan synthesis inhibitors beta-xylosides can strongly influence the growth permissiveness of oligodendrocytes. beta-Xylosides abolished cell surface presentation of brevican and versican V2 and reversed growth cone collapse in encounters with oligodendrocytes as demonstrated by time-lapse video microscopy. Instead, growth cones were able to grow along or even into the processes of oligodendrocytes. Our results strongly suggest that brevican and versican V2 are additional components of CNS myelin that contribute to its nonpermissive substrate properties for axonal growth. Expression of these CSPGs on oligodendrocytes may indicate that they participate in the restriction of structural plasticity and regeneration in the adult CNS.

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BackgroundTo explore the possibility that antibody-mediated complement lysis contributes to viremia control in HIV-1 infection, we measured the activity of patient plasma in mediating complement lysis of autologous primary virus.Methods and FindingsSera from two groups of patients—25 with acute HIV-1 infection and 31 with chronic infection—were used in this study. We developed a novel real-time PCR-based assay strategy that allows reliable and sensitive quantification of virus lysis by complement. Plasma derived at the time of virus isolation induced complement lysis of the autologous virus isolate in the majority of patients. Overall lysis activity against the autologous virus and the heterologous primary virus strain JR-FL was higher at chronic disease stages than during the acute phase. Most strikingly, we found that plasma virus load levels during the acute but not the chronic infection phase correlated inversely with the autologous complement lysis activity. Antibody reactivity to the envelope (Env) proteins gp120 and gp41 were positively correlated with the lysis activity against JR-FL, indicating that anti-Env responses mediated complement lysis. Neutralization and complement lysis activity against autologous viruses were not associated, suggesting that complement lysis is predominantly caused by non-neutralizing antibodies.ConclusionsCollectively our data provide evidence that antibody-mediated complement virion lysis develops rapidly and is effective early in the course of infection; thus it should be considered a parameter that, in concert with other immune functions, steers viremia control in vivo.

BackgroundAlthough combination antiretroviral therapy (cART) initiated in the acute phase of HIV-1 infection may prevent expansion of the latent reservoir, its benefits remain controversial. In the current study, HIV-1 RNA transcription patterns in peripheral blood mononuclear cells (PBMC) were monitored during acute cART to assess the effect of early treatment on cellular viral reservoirs.Methodology/Principal FindingsAcutely HIV-1 infected patients (n = 24) were treated within 3–15 weeks after infection. Patients elected to cease treatment after ≥1 year of therapy. HIV-1 DNA (vDNA), HIV-1 RNA species expressed both in latently and productively infected cells, unspliced (UsRNA), multiply spliced (MsRNA-tatrev; MsRNA-nef), and PBMC-associated extracellular virion RNA (vRex), expressed specifically by productively infected cells, were quantified in PBMC by patient matched real-time PCR prior, during and post cART. In a matched control-group of patients on successful cART started during chronic infection (n = 15), UsRNA in PBMC and vDNA were measured cross-sectionally. In contrast to previous reports, PBMC-associated HIV-1 RNAs declined to predominantly undetectable levels on cART. After cART cessation, UsRNA, vRex, and MsRNA-tatrev rebounded to levels not significantly different to those at baseline (p>0.1). In contrast, MsRNA-nef remained significantly lower as compared to pretreatment (p = 0.015). UsRNA expressed at the highest levels of all viral RNAs, was detectable on cART in 42% of patients with cART initiated during acute infection as opposed to 87% of patients on cART initiated during chronic infection (Fisher's exact test; p = 0.008). Accordingly, UsRNA levels were 105–fold lower in the acute as compared to the chronic group.ConclusionEarly intervention resulted in profound depletion of PBMC expressing HIV-1 RNA. This is contrary to chronically infected patients who predominantly showed continuous UsRNA expression on cART. Thus, antiretroviral treatment initiated during the acute phase of infection prevented establishment or expansion of long-lived transcriptionally active viral cellular reservoirs in peripheral blood.

Recently, passive immunization of human immunodeficiency virus (HIV)-

Background: Mathematical models based on kinetics of HIV-1 plasma viremia after initiation of combination antiretroviral therapy (cART) inferred HIV-infected cells to decay exponentially with constant rates correlated to their strength of virus production. To further define in vivo decay kinetics of HIV-1 infected cells experimentally, we assessed infected cell-classes of distinct viral transcriptional activity in peripheral blood mononuclear cells (PBMC) of five patients during 1 year after initiation of cART

treatment of tree shrews in China 15,16 as well For the systematic analysis of various clinical and moas the easy adaptation of these animals to the laboratory lecular aspects of hepatitis B virus (HBV) infection, an environment, 17,18 we systematically analyzed the tree shrew experimental small animal system of HBV infection species tupaia belangeri for the study of HBV infection in would be a great advance. The susceptibility to HBV invitro and in vivo. fection, therefore, of hepatocytes from the tree shrew species tupaia belangeri was studied in vitro and in vivo. MATERIALS AND METHODS Primary hepatocytes isolated from livers of tupaias can be reproducibly infected with HBV. In vitro infectionTupaia Colony. For breeding purposes, animals (tupaia belangeri) were obtained from the German Primate Center, Göttingen, Gerresults in viral DNA and RNA synthesis in hepatocytes many. They were bred and maintained at the animal facilities of and secretion hepatitis B surface antigen (HBsAg) and the University Hospital Zü rich in accordance with institutionally hepatitis B e antigen (HBeAg) into culture medium. Tuapproved protocols. Under optimal conditions (room temperature, paias can also be infected with HBV in vivo, resulting 28ЊC; relative humidity, 70%; day-night light cycle, 12 and 12 hours), in viral DNA replication and gene expression in tupaia the mothers give birth to two to three animals every 8 to 10 weeks. livers. Similar to acute, self-limited hepatitis B in hu-At birth, the animals are about 6 to 7 cm long (without tail) and mans, HBsAg is rapidly cleared from serum, followed weigh 16 to 19 g. Within 10 to 12 weeks, they reach adulthood with by seroconversion to anti-HBe and anti-HBs. These data an average length of 12 to 18 cm (without tail) and 180 to 200 g body weight. All animals were negative for serological markers of HBV clearly show that HBV is infectious to tupaia hepato- problem worldwide and is associated with a wide spectrum of exclusion, was 90% to 95%. The cells were washed and seeded at a clinical presentations, ranging from the asymptomatic HBV density of 3 1 10 6 viable cells/10 mL medium on collagen-coated 100-carrier status to the development of cirrhosis and hepatocel-mm tissue culture plates. Plating efficiency was 85%. Cells were lular carcinoma. 1 HBV represents the prototypic member of maintained in Williams' E medium with Glutamax J, buffered with the hepadnaviruses 2,3 and many molecular and clinical as-15 mmol/L hydroxyethyl piperazine ethane sulfuric acid (Sigma), pH 7.4, and supplemented with 1.5% dimethyl sulfoxide (HybriMax, pects of HBV infection have been defined. 4,5 HBV naturally Sigma), 0.1 mmol/L insulin, 0.1 mmol/L dexamethasone, 100 mg/mL infects only humans and experimentally chimpanzees. While penicillin, and 100 mg/mL streptomycine.various aspects of the pathogenesis of HBV-related liver disTransient Transfection of Primary Tupaia Hepatocytes. Transfecease can be studied in transgenic animals 6-9 and animal modtion of primary tupaia hepatocytes with a replicatio...