Biphasic effect of transforming growth factor-beta on Epstein-Barr virus-induced activation of human tonsillar B cells.

Transforming growth factor-beta (TGF-beta) is known to inhibit mitogen-induced proliferation of human B lymphocytes. Earlier results showed that activation of B cells by Epstein-Barr virus (EBV) was also inhibited by TGF-beta. On the other hand, TGF-beta could enhance the transformation of EBV-infected B-cell cultures. In the present set of experiments, we have confirmed the inhibitory effect of TGF-beta on the EBV-induced blastogenesis and found lower expression of CD23 in the treated cultures. However, cells which escaped inhibition and entered in the blast stage expressed a higher level of CD23 molecules. The elevation of CD23 in the TGF-beta-treated cultures was more marked at a time when the cell size profiles of the control and treated cultures were similar. In view of the function of the CD23 molecule as an autocrine growth factor, its increased expression is consistent with previous findings on TGF-beta-mediated enhancement of the transformation of B-cell cultures. The occurrence of growth inhibitory and growth stimulatory effect of TGF-beta on the same cell type has been observed in several other systems as well.

Influence of transforming growth factor-beta (TGF-beta) on the immunoglobulin production by EBV-infected B cell cultures.

TGF-beta inhibits the proliferation of human B lymphocytes stimulated by a variety of activators, including EBV. However, EBV-immortalised cells are refractory to TGF-beta. The influence of TGF-beta on B cell maturation varies, apparently depending on the origin of the B lymphocytes and their maturation/activation state, the strength of the stimulus and the presence of cofactors. We investigated the effect of TGF-beta on immunoglobulin production by 5-day-old EBV-infected B cells. TGF-beta added at the initiation of the cultures inhibited IgM, IgG and IgA secretion by decreasing the numbers of secretory cells. The inhibition of IgM secretion was strongest. At the cytoplasmic level, TGF-beta reduced the expression of IgM heavy, lambda and kappa light chains but not IgG and IgA heavy chains. However, the IgM production by an established EBV-transformed B cell line was not affected by TGF-beta. Thus, TGF-beta inhibited EBV-induced maturation of the B cells until they acquired a transformed state. We discuss the relevance of these findings for the potential role of TGF-beta on EBV infection.

Effect of TGF-beta 1 on the EBV-induced transformation of human lymphocyte cultures.

We tested the effect of TGF-beta 1 on the EBV-induced activation and immortalization of human B lymphocytes. In lymphocyte cultures of EBV-seropositive individuals, T cells can inhibit the EBV-induced transformation of B cells. We found that in the presence of TGF-beta 1 the transformation was more efficient, due to the inhibition of this function. TGF-beta 1 inhibited the EBV-induced proliferation of purified B-cell cultures. Its effect was strongest when added at the beginning of the culture. Added later, the inhibition gradually decreased and was lost by the 4th day. After 10 to 14 days, in the cultures initiated with TGF-beta 1, the number of cells was higher and they formed larger aggregates as compared with control cultures. Thus, TGF-beta 1 modifies EBV-induced transformation in a complex way. It inhibits the activation of B cells but does not affect those already activated. Once they acquire the immortalized state, the B cells are even stimulated by TGF-beta 1.

Endogenous TGF-beta contributes to the induction of the EBV lytic cycle in two Burkitt lymphoma cell lines.

A low proportion of cells in the BL lines P3HR-I and Akata enter spontaneously into the EBV lytic cycle, detectable by the expression of early antigens (EA). We found that both lines produce the active and inactive forms of TGF beta. It was shown earlier that a larger number of cells can be induced to enter the lytic cycle by exposing P3HR-I to phorbol esters and n-butyrate and the surface IgG-positive Akata cells to anti-IgG. We now show that the same treatments raise the level of active TGF beta release. Exposure to anti-TGF beta antibodies reduced EA induction by 75-85%. Our results indicate that induction of the viral productive cycle by the above-mentioned reagents is at least partly dependent on the activation of endogenous TGF beta production.

Correlation between the growth-inhibitory effect of TGF-beta 1 and phenotypic characteristics in a panel of B-cell lines.

Human B-cell lines established from Burkitt lymphoma (BL) and normal B cells immortalized in vitro by EBV (LCLs) differ in phenotype. While the BL correspond to resting B cells, the LCLs resemble activated B cells. When BLs which have the EBV genome are carried in vitro, they acquire some of the features of LCLs, such as expression of B-cell activation markers and the tendency to form aggregates. Comparison of several B-cell lines for sensitivity to TGF-beta showed that the growth of BLs (with few exceptions), but not of the LCLs, was inhibited. The results suggested that the sensitivity to TGF-beta correlates with the cellular phenotype. In the present work, this assumption is even more critically substantiated by studying 2 sublines of an EBV-genome carrying BL line, Mutu, which were selected for single cells and aggregates. The former (with resting phenotype) was inhibited, while the subline of aggregated cells, which also expressed B-cell activation markers, was not inhibited. Somatic-cell hybrids between BLs, LCLs and non-B cells provided lines with phenotypic differences. Results with a panel of such hybrid lines also showed that those which express the activated B-cell phenotype are not inhibited by TGF-beta. Differences in the levels of expression of activation markers did not influence the response to TGF-beta.

Effect of transforming growth factor-beta 1 and -beta 2 on the proliferation of Burkitt lymphoma and lymphoblastoid cell lines.

We tested the effect of transforming growth factor (TGF)-beta 1 and TGF-beta 2 on the proliferation of human B cell lines. The panel was selected to give information whether (1) their origin, (2) their phenotype, (3) their Epstein-Barr virus (EBV) carrier state, influence their responsiveness. The growth of lymphoblastoid cell lines (LCL) was not inhibited by TGF-beta 1. The EBV-carrying Burkitt lymphoma (BL) lines, Daudi, Jijoye, Rael but not Raji were inhibited. Three EBV-negative BL lines and the majority of their converted sublines were sensitive. The cell lines tested expressed TGF-beta receptors and TGF-beta 1 transcripts. The proliferation of EBV-infected B cells was inhibited by TGF-beta, their sensitivity decreased, however, after 3 days. The results suggest that the activation state of the B cells is decisive for TGF-beta sensitivity and EBV influences it indirectly by changing the cell phenotype.

Influence of viral load and alanine aminotransferase on viral genetic heterogeneity in patients with chronic hepatitis C virus infection.

BACKGROUND/AIM: Hepatitis C virus (HCV) populations in vivo consist of genetically different heterogeneous mixtures defined as 'quasispecies', which vary in the hypervariable region 1 (HVR1) mostly. To further address the role of quasispecies diversity in hepatitis C infection, this study aimed to evaluate the influence of ALT, viral load and genotypes on quasispecies heterogeneity in patients with HCV infection. METHODS: Thirty-six chronic hepatitis C patients with high levels of alanine aminotransferase (ALT) were studied. None of them received any antiviral therapy. HCV RNA serum levels, genotype and genetic heterogeneity were determined by branched-chain DNA assay, restriction fragment length patterns and RT-PCR single-strand conformational polymorphism analysis of HVR1, respectively. RESULTS: Twenty-eight patients had genotype 1b (28/36; 78%), 6 patients had genotype 1a (6/36; 17%), 1 patient was 2a (1/36; 3%) and genotype could not be determined in 1 patient. The patients were categorized into two groups according to the number of bands representing the dominant strains in the circulation: group A with 2 bands having 1 strain (14/36 patients; 39%) and group B with more than 2 bands indicating more than 1 strain (22/36 patients; 61%). The serum viremia and ALT levels for these groups were 11 +/- 8.8 and 5.3 +/- 4.6 mEq/ml (p < 0.05), and 79 +/- 20, and 127 +/- 80 IU/l (p < 0.05), respectively. CONCLUSION: The results of this study suggest that hepatitis C patients having 1 dominant strain in the circulation may show a relatively weaker immune response resulting in lower ALT and higher viremia levels, whereas patients with high degrees of virus quasispecies diversity have higher ALT levels and a more active immune response causing the selection of new genome variants and depressing viral replication partly.