T cell help is required to induce idiotypic-anti-idiotypic autoantibody network after immunization with complementary epitope 289-308aa of La/SSB autoantigen in non-autoimmune mice.

Immunotherapies against autoimmune diseases have been of limited success. Preventive vaccines could be developed on the basis to abrogate unwanted immune responses to defined autodeterminants. In this study it is shown that immunization of BALB/c mice with two linear T and B cell epitopes of the human La/SSB autoantigen (spanning the regions 289-308aa and 349-364aa) and their complementary forms specified by the complementary mRNA, results in characteristic B and T cell responses. Mice immunized with the 289-308aa epitope or its complementary peptide elicited specific antibodies against both epitopes. In contrast, mice immunized with the 349-364aa epitope or its complementary peptide mounted antibody titres against the immunizing peptide only. According to these data, the 289-308aa epitope and its complementary form were capable to generate an idiotypic-anti-idiotypic response, which were cross-regulated. Peptide-specific T cell proliferation and cytokine production in vitro revealed the induction of a two-stage T helper response (Th1-->Th2 type) after immunization with either the epitope 289-308 or its complementary peptide. IgG1 was the predominant subclass after immunization with the two forms of epitopes 289-308 and 349-364, while a response of the IgG2b > IgG2a was obtained after the immunization with the complementary form of 349-364 epitope reflecting the TH2/TH1 polarization, respectively. Our data suggest that the complementary peptides of two immunodominant epitopes of human LaSSB can mimic the autoantibodies against these epitopes and establish an active idiotypic-anti-idiotypic network.

Interleukin-1beta and interleukin-1alpha may affect the implantation rate of patients undergoing in vitro fertilization-embryo transfer.

OBJECTIVE: To investigate whether interleukin-1beta (IL-1beta) and interleukin-1alpha (IL-1alpha) affect the implantation rate of patients undergoing IVF-ET. DESIGN: Follicular fluid and serum were obtained on the day of hCG administration, the day of oocyte retrieval, and the day of embryo transfer. SETTING: Cellular immunology laboratory in a research institute, a high technology IVF unit in a medical center, and a university hospital. PATIENT(S): Thirty-three women who were undergoing IVF-ET. MAIN OUTCOME MEASURE(S): IL-1beta and IL-1alpha were measured by specific ELISA and their levels were correlated with the implantation rate. RESULT(S): Classification of IVF-ET patients according to their implantation rate revealed significantly higher amounts of follicular fluid IL-1beta in the implantation versus nonimplantation cycles (68.5+/-24.6 pg/mL versus 20.5+/-13.4 pg/mL); The difference between the level of IL-1alpha in the two groups was not statistically significant(11.6+/-5.1 pg/mL versus 7.3+/-1.9 pg/mL). In parallel, systemic FSH/hMG-dependent IL-1beta and IL-1alpha production was observed in implantation cycles but not in nonimplantation cycles. Statistically significant IL-1beta and IL-1alpha production was observed after administration of hCG. CONCLUSION(S): Gonadotropins used during IVF-ET induce local and systemic production of IL-1beta and IL-1alpha. In addition, the implantation rate for IVF-ET patients who have detectable serum concentrations of IL-1beta and IL-1beta on the day of hCG administration could be higher than the rate for IVF-ET patients who do not have detectable concentrations of these cytokines.

Alteration in peripheral blood mononuclear cell function and serum cytokines in oral lichen planus.

Different activation parameters of peripheral blood mononuclear cells (PBMC) from 31 patients with oral lichen planus (OLP) were examined and compared with 23 healthy donors. Impaired spontaneous (450 +/- 241 vs 1290 +/- 480 cpm) and mitogen-induced (39580 +/- 14470 vs 67000 +/- 11810 cpm) lymphocyte blastogenesis was observed in OLP patients. Furthermore, reduced cytokine production was found after phytohemagglutinin A (PHA) stimulation for all cytokines studied-tumour necrosis factor alpha (TNF alpha, 432.2 +/- 73.4 vs 979.8 +/- 46.3 units/ml), interleukin 2 (IL-2, 156.2 +/- 14.9 vs 572.6 +/- 12.9 pg/ml), interferon gamma (IFN gamma, 48.5 +/- 11.9 vs 82.6 +/- 12.4 pg/ml) and interleukin 6 (IL-6, 253.6 +/- 57.7 vs 1,419.0 +/- 279.6 units/ml)-except for interleukin 1 beta (IL-1 beta) and lymphotoxin (LT). In contrast, unstimulated culture supernatants showed increased TNF alpha (38.2 +/- 13.1 vs 8.0 +/- 0.2 units/ml), LT (10.2 +/- 2.2 units/ml vs < 0.4) and IL-6 (18.5 +/- 5.6 units/ml vs < 0.5) activity. Similarly, elevated concentrations of TNF alpha (19.6 +/- 6.3 units/ml) and IL-6 (22.9 +/- 4.7 units/ml) were detected in the sera of OLP patients. Combination of PHA and phorbol myristate acetate (PMA) could restore OLP proliferative T cell response and cytokine production to the level of healthy donors, whereas exogenous recombinant human IL-2 (rhuIL-2) plus PMA did not seem to be an effective stimulant for OLP T cells. These results indicate an alteration in the immune condition of OLP patients and an impairment in T lymphocyte function.

Interleukin-3 production as a sensitive measure of T-lymphocyte activation in the mouse.

Interleukin-3 activity is produced by T cells at an early phase after antigenic stimulation, and its quantitation provides a sensitive and non-destructive method for assaying T-cell activation. It can be detected before increased uptake of [3H]thymidine into DNA occurs, and is more sensitive than measuring [3H]thymidine uptake for low levels of T-cell activation. This method has been shown to be useful in measuring activation of T-cell clones and in detecting antigens that have been blotted onto nitrocellulose.

A fluorometric assay for determining cell growth in lymphocyte proliferation and lymphokine assays.

A microplate method for assessing cell growth and viability based on the hydrolysis of fluorogenic substrates by cell esterases has been investigated. Living cells incubated with fluorescein diacetate or 4-methylumbelliferyl heptanoate generate a fluorescent product which is proportional to the number of cells. This can be used as a simple and economical readout for various bioassays such as, for example, the assessment of IL-2 and IL-3 on factor-dependent cell lines, and on antigen- and mitogen-stimulated proliferation of lymphocytes. This fluorometric assay has similar sensitivity to the measurement of [3H]thymidine uptake and greater sensitivity than standard colorimetric assays. Incubation with MUH for a period of 30-60 min at 22 degrees C is adequate.