An involvement of cellular immunity in alveolar echinococcosis is strongly suggested by the intense granulomatous infiltrations observed around the hepatic parasite lesions. However, the basis of cellular immunoregulation in patients with alveolar echinococcosis is poorly understood. The present report shows a comparative analysis of lymphoid cell function in peripheral blood mononuclear cells (PBMC) of 16 patients with alveolar echinococcosis and of healthy individuals. Our in vitro restimulation studies with crude Echinococcus multilocularis antigen demonstrated that PBMC from patients with alveolar echinococcosis were responsive to challenge with parasitic antigen as measured by lymphoid cell proliferation. In this system, we also evaluated cytokine expression at the gene and protein levels after stimulation with E. multilocularis antigen. Analysis of cytokine mRNA expression revealed distinct patterns of cytokine expression in patients and normal donors. By using reverse transcriptase PCR, we could demonstrate that the T H 1 cytokine transcripts interleukin-2 (IL-2) and gamma interferon (IFN-␥) are present in PBMC from patients with alveolar echinococcosis. Moreover, it was found that stimulation with E. multilocularis antigen induced or enhanced the expression of the T H 2 cytokine IL-3, IL-4, IL-10, and especially IL-5 mRNAs in PBMC from 13 of 16 patients with alveolar echinococcosis. Two patients who were examined after radical surgery, as well as another patient with a stable course of the disease under continuous chemotherapy, were not able to generate the same pattern of cytokine response and had no evidence of IL-5 mRNA synthesis. In contrast to the frequent expression of T H 2 cytokine mRNAs observed in patients with alveolar echinococcosis, PBMC cultures from normal donors showed prominent IL-2 and IFN-␥ mRNA expression but weak IL-3, IL-4, and IL-10 mRNA expression. Most interestingly, IL-5 mRNA was substantially absent in PBMC from healthy individuals. In accordance with the mRNA studies, it was found that E. multilocularis antigen induced the secretion of large amounts of IL-5 and intermediate amounts of IFN-␥ in patients with alveolar echinococcosis, whereas large amounts of IFN-␥ and no or threshold amounts of IL-5 were detected in supernatants from healthy individuals. Collectively, the present study provides the first evidence that a T H 2 immune response is gradually activated during the course of E. multilocularis infection, indicating a critical role for IL-5 in the manifestation of human alveolar echinococcosis.

The isolation and characterization of human monoclonal antibodies (humAbs) against the hepatitis C Virus (HCV) glycoproteins E1 and E2 are described. B-cells from blood donors with anti-HCV were transformed with Epstein-Barr virus. The supernatants of the resulting lymphoblastoid clones were screened by ELISA with an extract of cells infected with a recombinant vaccinia virus RMPA95 expressing the envelope proteins E1 and E2 of an HCV genotype 1a virus (H strain). Positive clones were fused to the heteromyeloma cell line K6H6/B5. Fifteen heterohybridoma cell lines have been established. The specificity of the isolated humAbs was determined both by ELISA and Western blot assays. Several recombinant extracts expressing either the E1 or E2 protein or truncated forms were used in an attempt to map the epitopes on the viral glycoproteins. Some of the humAbs were used successfully for immunofluorescence investigation of transfected cells. Seven specific anti-E2 humAbs, which react with the envelope protein 2 of genotype 1a and 1b isolates, were characterized.

We assess the diagnostic value of the recombinant antigens E1/E2 (heterodimer) and E2 for the screening of blood donors. For this purpose an ELISA assay was developed which used these recombinant glycoproteins as solid antigens on the microtiter plates. The anti-E1/E2 assay was able to detect 80% of the positive samples by Matrix and 87% of the PCR-positive samples. Anti-E2 was less sensitive. Fifteen percent of the indeterminate samples were anti-E1/E2 and 8% anti-E2 positive. In the case of the indeterminate samples there was no coincidence of results between these assays and the PCR. Fifty percent of seroconverters showed an anti-E1/E2-positive result with a previous indeterminate donation. We conclude that the recombinant envelope proteins, mainly anti-E1/E2, might be used in future in the diagnostics of HCV infection to increase the sensitivity of the currently used assays.

The isolation and characterization of human monoclonal antibodies (humAbs) against the hepatitis C Virus (HCV) glycoproteins E1 and E2 are described. B-cells from blood donors with anti-HCV were transformed with Epstein-Barr virus. The supernatants of the resulting lymphoblastoid clones were screened by ELISA with an extract of cells infected with a recombinant vaccinia virus RMPA95 expressing the envelope proteins E1 and E2 of an HCV genotype 1a virus (H strain). Positive clones were fused to the heteromyeloma cell line K6H6/B5. Fifteen heterohybridoma cell lines have been established. The specificity of the isolated humAbs was determined both by ELISA and Western blot assays. Several recombinant extracts expressing either the E1 or E2 protein or truncated forms were used in an attempt to map the epitopes on the viral glycoproteins. Some of the humAbs were used successfully for immunofluorescence investigation of transfected cells. Seven specific anti-E2 humAbs, which react with the envelope protein 2 of genotype 1a and 1b isolates, were characterized.