The Influence Of Ifn-λ₁on Peripheral Blood Source Dendritic Cell In Adolescent Patients Of Chronic Hbv Infection

Objective: It is accepted that hepatitis B virus(HBV) mainly inducedhepatocytes injury by immune response and immunereaction of the host,anddifferent immune response induced different clinical manifestations andoutcomes after HBV infection. Accordance to immune characteristics, naturalprocess of chronic HBV infection in infant can be clinically categorized intofour periods,①immune tolerant phase,②immune clearance phase,③inactiveHepatitis B (or asymptomatic),④reactivation phase. The mechanisms for thepersistent HBV infection is not clear. Professional antigen-presenting cells(APCs), especially dendritic cells (DCs), play an important role in bridging theresponses of innate and acquired immunity. The antigen-presentinghypofunction of dendritic cell may be one of the important factors. As thebeginner, modulator and effect of immune response, DC can activate CD8+CTL and CD4+T helper cells, control the process of immune response, whichhas become the central link of host immune response. Otherwise Macrophagesand B cells could only stimulate activated and memory T cell, so DCs areconsidered to be the initiation of organism immunity, which play a unique rolein inducing the immunity. The CD80, CD83, CD86and HLA-DR highexpression of modified DC. Interferon λ (IFN-λ) was a new type of interferon,including IFN-λ₁(IL-29) and IFN-λ2(IL-28A) and IFN-λ3(IL-28B),which wasdiscovered though bioinformatics means from the human genome in2003byU.S. scientist Kotenko and Sheppard. Recent studies have shown thatinterferon λ mainly came from DC, so it played an important role in regulatingthe maturation and function of DC. Studies have shown that IFN-λ₁caninduce DC maturation and enhance its antigen-presenting function. Interferonλ has been widely reported on hepatitis C, but rarely about chronic HBVinfection in china and abroad. In this study, our aim to explore the influence and clinical significance of IFN-λ₁on peripheral blood source DC in immunetolerant phase and immune clearance phase of chronic HBV infection patients.Methods: All of34adolescent chronic HBV infection subjects including18patients in immune clearance phase and16patients in immune tolerantphase were enrolled in this study. All patients underwent liver biopsy,inflammation graded as G0^G4, fibrosis was divided into S0^S4.10healthypersons were enrolled in this study.1The Cultivation of Dendritic Cells: The PBMC was isolated by densitygradient centrifugation separation from individual peripheral blood, culturedby serum-free suspension and then collected attached cells. They were dividedinto three groups: The alone group was added recombinant humaninterleukin-29singly; the normal control group was added recombinant humaninterleukin-4and recombinant human granulocyte-macrophagecolony-stimulating factor; the IFN-λ₁group was recombinant humaninterleukin-4and recombinant human granulocyte-macrophage colony-stimulating factor and recombinant human interleukin-29cocultured. Above all groups cultured for7days.2The cells' morphologic changes were evaluated by microscopy during cellculture3The collected mature DC were stained with PE-DC80, PE-CD83, PE-CD86and the PE-HLA-DR antibody and then detected by flow cytometry.4The cell culture supernatant was harvested for the determination of INF-γ,IL-12.Results:1DC morphological observation: The majority visibled part of the cellsuspension were B cells and peripheral blood DC when PBMC were adherentcultured by2-4hours later. In the alone group added recombinant humaninterleukin-29singly, adherent cell shape had no changed after the2,5,7daysand extended to11days yet no change. In the other two groups normal controland IL-29group, the number of adherent cells decreased after onovernightcultured, and cells size was small; the stretching adherent cells can be seen after2days; a typical DC morphology was observed when cultured5days,which is irregular and not in same size and thickness; the DC surfaceprotrusions were more pronounced after cultured7days, and in addition, alarge number of protrusions intertwined were observed at in adding IL-29group.2The results of DC surface markers detected by flow cytometry: There's havesignificant difference of DC surface molecules of CD80, CD83, CD86andHLA-DR expression in immune tolerance group and immune clearance groupand normal control group (p<0.01); There's have significant differencebetween immune tolerance group and immune clearance group(p<0.01);There's have significant difference between IL-29group and no IL-29group(p<0.05).3The results of IFN-γ and IL-12determination in the cell culture supernatant:There's have significant difference of IFN-γ and IL-12concentration inimmune clearance group stronger than immune tolerance group butsignificantly lower than those normal control group (p <0.05). There's havesignificant difference between IL-29group and no IL-29group. It cansignificantly improve the function of DC's secretion of IL-12and IFN-γcytokines when co-cultivation with IL-29(p<0.05)Conclusions:1DC can be successfully obtained by adherent culture for removing ofsuspended cells in the PBMC that was isolated from peripheral blood ofpatients. Adding IL-29singly can not induce maturation of DC. Added IL-4,GM-CSF cultured together can significantly promote and induce maturation ofDC from peripheral blood.2DC function in patients at immune tolerance phase with chronic HBVinfection was significantly lower than that in both normal people and patientsat immune clearance phase. It is suggested that lack of DC function is one ofthe reasons for chronic HBV infection of adolescent.3DC function is significantly enhanced through IFN-λ₁treatment in vitro inchronic HBV infected person at immune tolerance and immune clearance phase. IFN-λ₁can promote DC maturation and regulate antigen-presentingfunction of DC in vitro.The IFN-λ1may become the new choice for thetreatment of chronic hepatitis B. It is suggested that IFN-λ₁has a strongimmunomo-dulatory capacity.