Rashmi Singh,1 Jay Bullard,1 Krystal Vonfeldt,1 SenaitAssefa,1 Mamta Kalra,1 Anil Kaul,1 Robert Conrad,1 KhalidKhan,2 Harvey Sharp,2 Rashmi Kaul.1 1OSU-Center for HealthSciences, Tulsa, OK; 2University of Minnesota, Minneapolis,MN

Increased systemic LPS levels may cause bacterialtranslocation and inflammation in the liver. Epidemiologicdata of bacterial translocation, colonization and inflamma-tion in normal livers is lacking. TLR4 is LPS receptor involvedin liver homeostasis modulating innate and adaptive immu-nity by activation of NFκβ. We hypothesize liver homeostasiswill decrease the ability of translocating commensals toactivate TLR4 and NFκβ. Colonization by aerobic andfacultative anaerobic bacteria in normal, primary biliarycirrhosis (PBC), and nonalcoholic steatohepatitis, NASHexplant livers was studied and bacteria speciated by APIsystem. TLR4 and pIkKα protein expression analyzed bywestern blotting and TLR4 mRNA levels by real-time RT-PCR.Incidence of positive culture among normals (12/14), PBC (5/9), and NASH (3/6) was 86%, 56% and 50%. Both gram positiveand negative bacilli and cocci were isolated. Relative mRNAlevels for TLR4 varied significantly (pb0.0008). TLR4 mRNAlevels in normals were comparable to PBC (pN0.05) butlower compared to NASH (Pb0.0001). TLR4 protein levels innormals (0.39±0.02, Mean±SEM) were significantly lowerthan PBC (0.58±0.06, pb0.05) and NASH (0.59±0.03). pIkKαprotein levels in normals were low (0.83±0.07) compared toPBC (1.04±0.06) and NASH (1.04±0.09) (pN0.05). This is thefirst clinical evidence showing bacterial colonization innormal human livers comparable to cirrhotic livers withoutactivation of TLR4 and IkKα/NFκβ in normals highlighting theimportance of bacterial colonization in liver homeostasis. Ina susceptible host or in a liver transplant patient the residentbacteria in the liver may lead to break in tolerance resultingin re-infections, fibrosis, cirrhosis or cancer.

doi:10.1016/j.clim.2008.03.079

OR.73. Mycobacterium Tuberculosis DerivedToll-like Receptor 2 Ligand Modulate the Function ofCD4+ T Cells DirectlyXinchun Chen, Boping Zhou, Mingxia Zhang. ShenzhenDonghu Hospital, Shenzhen, China

Recent studies showed that Toll-like receptor 2(TLR2),which is previously known to be expressed in innate immunecells, also expressed on activated CD4 T cells. The objectiveof this study was to investigate whether and how mycobac-terium tuberculosis derived TLR2 ligand modulate thefunction of CD4 T cells in vivo and in intro. TLR2 expressionof CD4 Tcells from 33 cases of pulmonary tuberculosis and 79healthy donors was analyzed by real-time RT-PCR and flowcytometry. Human-TLR2 transduced 293 cell line with IL-8production as a reporter was used to determine TLR2 ligandactivity in serum samples. The effect of BCG and mycobac-terium tuberculosis derived TLR2 ligands (19 kDa lipoprotein,ESAT-6) on cytokine secretion and proliferation of CD4 T cellwas assessed in vitro by ELISA and CFSE labeling, respec-tively. Our results showed that expression of TLR2 on

activated CD4 T cells was significantly increased in patientswith pulmonary tuberculosis compared to healthy donors(pb0.05). Similarly, the TLR2 ligand activity was alsosignificantly increased in circulation of patients withpulmonary tuberculosis. In vitro assay showed that 19 kDalipoprotein, live BCG and ESAT-6 significantly enhanced theinterferon-gamma secretion and proliferation of CD4 T cellsactivated by anti-CD3 and anti-CD28. Blocking TLR2 receptorby antibody against TLR2 abrogate these effects indicatedthat the dependence on TLR2. These results suggested thatdirect modulating function of CD4 T cells through TLR2 is amechanism of immunopathogenesis of tuberculosis sincemycobacterium tuberculosis contains high proportion oflipoprotein, the natural TLR2 ligand.

doi:10.1016/j.clim.2008.03.080

OR.74. GCF2/LRRFIP1 Signals Influenza Infectionand Drives a Type I Interferon ResponseKathleen Sullivan, Asen Bagashev. CHOP, Philadelphia, PA

Uptake of virus and the cell autonomous response toinfection is carefully regulated to induce type I interferonswhich in turn, induce the establishment of an anti-viral state.We studied the role of a proteinwhose expression was found toinduce type I interferon expression. LRRFIP1/GCF2 is recruitedspecifically to virus-containing early endosomes within a fewminutes. It is also recruited to RNA-containing structures butnot endosomes containing transferring or E. coli. Overexpres-sion drives type I interferon expression at baseline andincreases type I interferon transcripts after influenza infec-tion. GCF2 co-immunprecipitates with p38, and Akt, proteinsknown to be involved in innate anti-viral responses. Inhibitionof p38 abrogates colocalization of GCF2 with virus-containingendosomes and impairs phosphorylation of GCF2. GCF2 alsocolocalizeswith sites of replicating virus and RNA suggesting anability to recognize pathogen-associated patterns. Over-expression of GCF2 diminished the ability of the cells tosupport influenza infection. Taken together, thesedata suggestthat GCF2 acts early in the viral entry process to signal viralinfection and plays a role in initiating antiviral responses.Recruitment as early as 3 minutes after infection suggests thatGCF2 is a very early sensor of viral infection.

doi:10.1016/j.clim.2008.03.081

OR.75. Identification of Cytotoxic Human LeukocyteAntigen (HLA)-DR-Restricted CD4+ T-Cell Epitopesfrom HSV-1 Glycoprotein B that are FrequentlyRecognized by Seropositive Asymptomatic PatientsAziz Alami Chentoufi,1 Alex Nguyen,1 Noureddine Berka,2

Ilham Bettahi,1 Steven Wechsler,1 Anthony Nesburn,1

Lbachir BenMohamed.1 1University of California Irvine,School of Medicine, Irvine, CA 92697-4375, CA; 2CalgaryLaboratory Services, Calgary, AB T2L2K8, AB, Canada

The identification of “protective” (i.e. “asymptomatic”)epitopes recognized by T cells from HSV-infected asympto-

S31Abstracts