Angela Panoskaltsis

Interference with Fc signals increases an antibody response by T-cell-deprived cultures to a T-dependent antigen

Affinity column purified goat anti-mouse immunoglobulin antibodies specific for the Fc portion of IgG increased an in vitro antibody response to a T-dependent antigen when T cells were limiting. Picogram amounts of specific anti-Fc antibody at culture initiation and nanogram quantities up to 3 days were required to demonstrate this effect. The demonstration of reconstitution by anti-Fc antibodies requires that the cultures be T-cell depleted and stimulated by antigen. These results support the concept that anti-Fc antibody and T cells block endogenously generated negative Fc signals.

Rheumatoid factor and Fc signaling: A tale of two Cinderellas

Signaling to lymphocytes depends not only upon the interaction of receptors with specific antigen, but also upon antigen nonspecific receptors which receive input from two classes of molecules, immunologically specific end products and lymphokines. These represent the central elements in the physiologic stimulation of immune responses to both self and foreign antigens. This review is dedicated primarily to describing the function of physiologically active receptors for end product which are central to end product feedback in normal immune responses. The emergence of autoimmunity as a pathologic process resultant from a defective end product signaling mechanism is described.

Rheumatoid factor blocks regulatory Fc signals

Immunosuppressed cultures of murine spleen cells, partly deprived of T cells and antigen-stimulated, can be reconstituted to near full activity in their antibody-forming cell response with murine rheumatoid factors (RF). The dose of RF required for recovery of 50% of the reconstitutable immune response was 10-100 ng and reconstitution was blocked by intact murine IgG added to the cultures. IgG subclass specificity of RF was demonstrated; RF specific for IgG2a was more potent than RF specific for IgG1 in reconstituting the response. Synergy was observed between RF added at culture initiation and late-acting B-cell differentiation factors. The greater the degree of T-cell deprivation, the more stringent the conditions needed for reconstitution. Suitable conditions for reconstitution with profound T-cell depletion included the limited reconstitution by specific RF, the synergistic action of RF with late-acting T-cell-replacing supernatants, and multiple additions of a number of RFs to the cultures on Days 0, 1, and 2. RF was also shown to block Fc-dependent immunosuppression by added antigen-antibody complexes. These results are interpreted as favoring the hypothesis put forward previously that the normal production of RF acts to reduce T-cell dependency by preventing negative Fc signal transmission by immune complexes on the B-cell surface. Abnormal production of RF may be a primary destabilizer of the immune responses leading to autoimmunity.

Regulation of an anti-self response : lack of influence of exogenous DNA on the in vitro anti-DNA response

Anti-DNA antibodies occur in outwardly normal individuals as well as in various forms of autoimmune disease. A number of publications have reported on the ability of added DNA to either induce or inhibit the in vitro production of anti-DNA antibody. In this study, the in vitro production of IgM anti-single stranded DNA (alpha ssDNA) antibody by spleen cells from normal or autoimmune mice neither depends upon, nor is inhibited by, the addition of high molecular weight DNA to the culture. The decrease in antibody forming cell plaques, reported previously, is due solely to the artifactual carryover of inhibitory material into the assay system, where it interferes with the expression of plaques by preventing anti-DNA antibody from reaching the DNA-coated erythrocytes. Similarly, plaque forming cell (PFC) methods have not detected alpha ssDNA antibody producing cells in murine spleen cells without culturing, but various other systems for measuring antibody normally detect anti-DNA antibodies in vivo. This discrepancy is also due to inadequate washing of freshly harvested cells to rid them of inhibitory substances which prevent them from registering as PFC. While S1 nuclease was able to prevent PFC interference by purified DNA, it did not remove the inhibitory substances from the culture supernatants; therefore substances other than ssDNA are able to interfere with alpha ssDNA PFC, suggesting that the alpha ssDNA PFC detected are polyspecific. Levels of alpha ssDNA PFC in spleen cells from non-autoimmune mice begin at one-quarter of the peak in vitro response, decrease to one-tenth in the first day and then reach peak values after 3 to 5 days of culture, suggesting that spleen cells are actively producing alpha ssDNA antibodies an in vivo and that then in vitro response is observed. Despite this evidence for an in vitro alpha ssDNA response, this response was not inhibited markedly by 1000 rad gamma-irradiation, while the response to sheep erythrocytes (SRBC) was profoundly suppressed. These findings suggest that anti-self B lymphocytes are resistant to interphase, possibly apoptotic, lymphocyte death due to gamma-irradiation, while anti-nonself B lymphocytes remain sensitive.

Immunoregulatory characteristics of the in vitro anti-ssDNA response

Continuous blockade of B-cell antigen receptors (BCRs) with Fab αsIg prevents the anti-ssDNA response of high, but not low, density B cells. Signaling via the BCRs, by prior exposure to crosslinking F(ab′)2 αsIg, had no effect on the spontaneous anti-DNA response, but prevented a lipopolysaccharide-induced anti-DNA response. Pretreatment with intact αsIg, which provides exogenously derived Fc signals, reduced the response. An Fc-signal-blocking agent, F(ab′)2 anti-IgG-Fc antibody, increased the number of anti-DNA antibody-forming cells produced in the absence of exogenous IgG anti-ssDNA antibody. Thus, activation is dependent on the availability of the BCRs, prior BCR crosslinking does not interfere with activation, and endogenous IgG anti-ssDNA antibody limits the activation of anti-ssDNA-specific B cells most of which are T-cell independent. These results indicate that the anti-ssDNA response is driven through the BCR.