Recombinant granulocyte-macrophage colony-stimulating factor restores deficient immune responses in mice with chronic Trypanosoma cruzi infections

Spleen cells from mice with chronic Trypanosoma cruzi infection generate a minimal plaque-forming response to SRBC in vitro. Addition of granulocyte-macrophage (GM)-CSF to cultures of spleen cells from chronically infected mice restored the plaque-forming cells (PFC) response to normal levels. Splenic adherent cells from chronically infected mice were deficient in their ability to reconstitute the PFC response of accessory cell-depleted normal spleen cells. Preincubation of splenic adherent cells from infected mice with GM-CSF restored their ability to reconstitute the PFC response of adherent cell depleted cultures. Ia Ag expression by splenic adherent cells from chronically infected mice was significantly lower compared to Ia Ag expression of cells from normal mice. Incubation of splenic adherent cells from chronically infected mice for 48 h with GM-CSF increased levels of Ia Ag expression to approximately those of uninfected mice. Peritoneal macrophages from infected mice produced IL-1 after incubation with GM-CSF at levels equivalent to those produced by similarly treated control macrophages. Spleen cells from chronically infected mice showed significant induction of IL-2 mRNA after GM-CSF treatment, and the addition of the anti-IL-2 mAb to GM-CSF supplemented cultures of spleen cells from infected mice blocked the restoration of the anti-SRBC PFC response. Thus, the ability of GM-CSF to restore the anti-PFC response to SRBC appears to involve the up-regulation of accessory cell function that includes increased Ia Ag expression and the induction of IL-1 production. These events also involve increased IL-2 production with resultant up-regulation of the response to SRBC by spleen cells from infected mice. Finally, it was shown that treatment of infected mice with rGM-CSF completely restored their depressed PFC production in vivo.     

The enhancement of the immune response by pain stimulation in mice. I. The enhancement effect on PFC production via sympathetic nervous system in vivo and in vitro

Effects of catecholamines and osmotical and physical stimuli on the induction of anti-sheep red blood cells (SRBC) plaque-forming cells (PFC) were investigated in (C57BL/6 X BALB/c)F1 mice in vivo and in vitro. The anti-SRBC PFC from mice immunized with 5 X 10(7) SRBC was markedly increased by daily s.c. injections of epinephrine. The enhancement of PFC by epinephrine was completely blocked by preadministration with propranolol and hexamethonium, but not with phentolamine. The PFC was increased by osmotic and physical stimuli given once a day for 4 days after immunization with SRBC. The enhancement of PFC by these stimuli was completely blocked by preadministration with propranolol and hexamethonium. The enhancement of PFC by physical stimuli was observed in nonimmunized mice when spleen cells from stimulated mice were cultured with SRBC in vitro. In normal mice, the enhancement of PFC was observed 2 hr after one physical stimulation. However, spleen cells from mice given two physical stimuli did not show the enhancement of PFC after treatment with anti-Thy-1.2 antibody and complement, nor after removal of nonadherent cells. Next, the serum obtained from mice 30 to 60 min after a physical stimulation enhanced PFC of normal mice spleen cells in vitro, but the enhancement was abolished by the addition of propranolol. The enhancement of anti-SRBC PFC by s.c. injection of epinephrine suggested that the autonomic nervous system, especially the sympathetic nervous system, was activated by a local stimulus effect of the injection. This enhancement of anti-SRBC PFC appear to be due to the activation of antigen non-specific helper T lymphocytes by the beta-actin of endogenous catecholamines from the adrenal gland.     

Studies of the immunological capacity of germfree mouse radiation chimeras. III. In vitro reconstitution of the T-helper cell deficiency

The plaque-forming cell (PFC) response of long-term radiation induced allogeneic bone marrow chimeric (ABMC) mice has been shown to be markedly deficient. The nature of the cellular deficiency of the primary PFC response was investigated using in vitro culture techniques. Adherent spleen cells from ABMC or DBA/2 mice support equally well the development of PFC from nonadherent DBA/2 spleen cells. Nonadherent cells prepared from ABMC mice when cocultivated with DBA/2 adherent cells showed a minimal response. However, the addition of activated DBA/2 T cells to cultures containing adherent cells from DBA/2 mice and nonadherent cells from ABMC mice completely reconstituted the in vitro response to sheep erythrocytes. Therefore a cellular deficiency of the humoral immune system of ABMC mice was shown to be associated with the thymus-derived lymphocyte pool.     

In vivo immune response suppression by the supernatant from concanavalin A-activated spleen cells.

Supernatants from concanavalin A- (Con A) activated murine spleen cells have been shown to suppress the in vitro plaque-forming cell (PFC) response to sheep red blood cells (SRBC). The present study examined the effect of such Con A-activated spleen cell supernatants (herein termed CONS) on the in vivo immune response to SRBC in C57BL/6, BALB/c and CDF1 mice. CONS derived from BALB/c spleen cells suppressed direct PFC 4 and 8 days after immunization with 2 X 10(8) SRBC. CONS also suppressed indirect PFC 8 days after immunization, as well as serum hemagglutinins to SRBC. The PFC response of C57BL/6 (H-2b) mice was suppressed as much as that of BALB/c (H-2d) by CONS derived from BALB/c mice, indicating a lack of H-2 specificity of the CONS. In addition to suppression of the antibody response to SRBC, in vivo CONS administration resulted in reduction in spleen cell number. This reduction was not sufficient to explain the decreased PFC response. When the CONS was separated into less than 10,000 m.w. and greater than or equal to 10,000 m.w. fractions, the immunosuppressive activity was found in the less than 10,000 m.w. fraction. This observation suggests that intact interferon, SIRS, and MIF were not responsible for the results obtained.     

Immunosuppression induced by attenuated Salmonella. Reversal by IL-4

We previously demonstrated that an aroA- strain of Salmonella typhimurium, which provides excellent protection against virulent Salmonella challenge, also rendered immunized mice unable to mount in vivo and in vitro antibody responses to heterologous Ag. Coculture studies using transwell plates indicated that suppression was mediated by soluble factors. The suppressive cells were identified as belonging to the monocytic linkage. Macrophage precursors as well as mature adherent macrophages mediated the observed suppression. In the present study, the mechanism of immunosuppression was investigated. Suppression was found to be genetically nonrestricted as spleen cells from immunized C3HeB/FeJ mice (H-2k) suppressed the anti-SRBC plaque-forming cell (PFC) responses of normal spleen cells from two MHC noncompatible mouse strains, BALB/c (H-2d) and C57BL/6 (H-2b). Time course experiments demonstrated that the addition of spleen cells from immunized mice to normal splenocytes as late as day 4 of a 5-day assay was still markedly suppressive. Furthermore, suppression of the PFC responses was accompanied by a profound inhibition of the capacity of immune splenocytes to produce IL-2 in response to in vitro stimulation by Con A. Coculture studies showed that immune spleen cells were able to suppress IL-2 production by normal splenocytes in a dose-dependent fashion. However, the suppressed PFC responses of immune spleen cells could not be reversed by the exogenous addition of up to 200 U/ml of IL-2, suggesting that immune splenocytes are also defective in their ability to respond to IL-2. In marked contrast, suppression of PFC responses was reduced by more than 50% by the addition of as little as 1 U/ml of IL-4 and was completely abrogated when 5 U/ml of IL-4 were added to in vitro cultures of spleen cells from immunized mice. The antisuppressive action of IL-4 appeared to be via its inhibitory effect on activated macrophages. The implications of the above findings are discussed.     

Cellular requirements for the expression of IgM. Immunological memory in vitro

In vitro culture techniques have been used to compare the direct (IgM) plaqueforming cell (PFC) response to heterologous erythrocytes (RBC) by normal mouse spleen cells and spleen cells from mice injected intravenously with 5 × 10⁴ RBC ten days previously [low dose primed (LDP)]. Although LDP mice fail to undergo a significant primary PFC response, their spleen cells are capable of a secondary or enhanced PFC response in vitro. The secondary PFC response is shown to be a function of: (A) an increase in the frequency of immunocompetent cells or units (IU) due to in vivo priming, and (B) an increased number of PFC generated per IU subsequent to in vitro stimulation. The latter increase is shown to be mediated through a shorter PFC doubling-time during logarithmic expansion of the PFC population. Analysis of nonadherent spleen cell dose response experiments indicate that two nonadherent cell types interact in the secondary response. Subsequent cocultivation experiments suggested that both of these cell types must be “primed” to allow induction of a secondary response. Although adherent cells are required for the secondary response, normal splenic adherent cells serve as equivalent substitutes for LDP adherent cells.     

Interferon inhibition of the primary in vitro antibody response to a thymus-independent antigen

Partially purified and crude mouse L cell interferon preparations inhibited the in vitro plaque-forming cell (PFC) response of mouse C57B1/6 spleen cells to the T-cell independent lipopolysaccharide antigen of Escherichia coli 0127. PFC responses of 5-day cultures were inhibited approximately 70–90% by 100–200 NIH reference units of interferon/culture. A similar inhibitory effect was obtained with spleen cells from athymic (nude) mice homozygous for the nu/nu allele. Spleen cultures depleted of adherent cells were also inhibited in their anti-0127 PFC response by interferon. Interferon, then, appears capable of inhibiting the PFC response to E. coli 0127 via direct action on B cells. Heating experiments along with the use of interferon preparations of different specific activities suggest that the inhibition was due to the interferon in the preparations.     

Selective impairment of B cell function by Neisseria meningitidis

Spleen cells from CBA/J mice infected with Neisseria meningitidis displayed depressed in vitro plaque-forming cell (PFC) responses to T-dependent (sheep red blood cell; SRBC) and T-independent (TNP-LPS, TNP-Ficoll) antigens. The inhibition was observed over a wide range of antigen concentrations. The decreased responsiveness of splenocytes from infected mice was due to a selective impairment of B-cell function since helper-T-cell activity was intact in infected mice as shown by the ability of T-enriched lymphocytes to cooperate with normal B-enriched lymphocytes in the generation of an anti-SRBC response, accessory macrophage function was preserved since adherent spleen cells from bacteria-injected mice were shown to produce normal or increased levels of IL-1 and were able to cooperate with normal non-adherent spleen cells in the generation of PFC against SRBC. Addition of peritoneal cells from normal animals or extraneous IL-1 both failed to restore normal PFC responses in cultures of splenocytes from infected mice. Finally, B-enriched lymphocytes from infected mice produced poor anti-SRBC responses when cultured with either Con A supernatant or T-enriched lymphocytes from normal or infected mice. Cell-mixing experiments failed to detect the presence of suppressor cells in cultures of unfractionated spleen cells or B-enriched lymphocytes from infected mice. Therefore, the immunological unresponsiveness associated with a Neisseria meningitidis infection was attributed to a meningococcus-induced defect(s) in B-cell function. In vivo polyclonal B-cell activation leading to clonal exhaustion did not play a major role in the depression of humoral responses since meningococcal infection induced little or no polyclonal Ig secretion.     

Immune dysfunction associated with graft-vs-host reaction in mice transplanted across minor histocompatibility barriers. II. Reversible defect in T-dependent antibody responses

B cell and Th cell functions were assessed in mice undergoing a graft-vs-host reaction (GvHR) in response to minor histocompatibility Ag by using the plaque-forming cell (PFC) response to the T-independent Ag TNP-Brucella abortus and the T-dependent Ag TNP-SRBC. Bone marrow plus spleen cells from B10.D2 mice were transplanted into lethally irradiated B10.D2 (syngeneic recipient) or H-2d-compatible BALB/c (allogeneic recipient) to produce a chronic form of GvHR. BALB/c recipients of an allogeneic transplant demonstrated a marked and proportional lymphoid depletion of the spleen with normal percentages of B cells, T cells, and CD4+ and CD8+ T cell subsets. Mice with GvHR made normal numbers of PFC/10(5) spleen cells in response to the T-independent Ag, but a significantly depressed number of PFC/10(5) spleen cells to the T-dependent Ag compared with normal B10.D2 mice and with irradiated B10.D2 recipients of syngeneic B10.D2 marrow plus spleen cells. Mice undergoing the minor Ag GvHR made significantly larger numbers of PFC/10(5) spleen cells after secondary immunization with TNP-SRBC compared with controls. In vitro assays demonstrated that B cells from mice with GvHR responded to T help from normal B10.D2 mice and that T cells from mice with GvHR provided help to normal B cells after in vivo immunization. These data demonstrate that radiation chimeras with GvHR in response to minor histocompatibility Ag have relatively normal B cell function and an apparent defect in T helper cell function that is reversible by immunization with appropriate Ag.     

Specific suppression of the in vitro parent anti-hybrid reaction. I. Characterization of a suppressor cell induced in hybrids by pretreatment with parent-strain spleen cells

Spleen cells from B6D2F1 hybrid mice pretreated with 5 X 10(7) B6 spleen cells iv 7 days earlier (B6-pretreated B6D2F1) exhibit a reduced capacity to stimulate the in vitro proliferative and anti-D2 CTL responses of B6 spleen cells. This inability of B6-pretreated B6D2F1 spleen cells to stimulate B6 spleen cells efficiently is due neither to the absence of stimulating cells bearing the D2 alloantigens nor to the destruction of B6 responding cells, but to the presence in the B6-pretreated B6D2F1 cell population of a suppressor mechanism, since the addition of B6-pretreated B6D2F1 spleen cells to a culture of normal B6 responding and irradiated B6D2F1-stimulating spleen cells can suppress the B6 anti-B6D2F1 response. This suppression is mediated by a nylon adherent, Thy-1-negative cell of parent-strain origin which is radioresistant at 2000 R. This suppressor cell is not induced by the injection to B6D2F1 hybrids of spleen cells from the other parent strain (D2) or an allogeneic strain (C3H). It does not suppress either the response of the other parent (D2) or an allogeneic strain (C3H) to B6D2F1 antigens, or the response of B6 cells to an allogeneic strain (C3H).     

In vitro immune response of cells of various lymphoid tissues in (NZB X NZW)F1 mice: evidence for abnormality of the mesenteric lymph node cells

Autoimmune-prone (NZB X NZW)F1 (B/W) mice have been shown to have a variety of immunologic perturbations. However, most studies have been performed with spleen cells. By using the Mishell-Dutton culture system, we examined the in vitro immune response of the various lymphoid tissue to determine whether an imbalance at a selective lymphoid site may exist in B/W mice. It was shown that the ability of mesenteric lymph node (MLN) cells of B/W mice to generate plaque-forming cells (PFC) in response to sheep red blood cells was consistently less than that of the spleen cells. This relationship held true in the aged mice. In contrast, the ability of the MLN cells of other strains not prone to develop autoimmunity to generate PFC was higher than that of the spleen cells. No significant difference in the mitogenic response of the lymphoid cells from various lymphoid tissue in the young B/W mice was seen, as compared with normal lymphoid cells from control mice. However, it was demonstrated that a relative decrease of B cells and immunoregulatory Lyt-123+ cells in the MLN in the B/W mice occurred early in life, and it was concluded that this abnormality may account for the low PFC response observed.     

Influence of age on the ability of thymic adherent cells to produce factors in vitro which modulate immune responses of thymocytes

Thymic adherent cells from BALB/c mice ranging in age from 1 day to 20 months were cultured in vitro for 1 month. The supernatants, collected at weekly intervals, were assessed for their ability to augment the antigen/mitogen responses of thymocytes from 2- or 4-week-old BALB/c mice and spleen cells from 3-month-old nude BALB/c mice. The results indicate that (a) the ability of thymic adherent cells to produce an augmenting factor(s) declines sharply between 2.5 and 5 months of age; (b) thymic adherent cells of 1-day-old mice synthesize an inhibitory factor(s) in addition to the augmenting factor, while those of young adult mice synthesize only the augmenting factor, and those of 20-month-old mice synthesize primarily the inhibitory factor; (c) supernatants containing the augmenting factor can be neutralized by mixing them with supernatants containing the inhibitory factor; (d) thymocytes which are responsive to the augmenting factor are immature as judged by their sensitivity to dexamethasone and by their ability to bind macrophages; and (e) spleen cells of normal and nude mice are not responsive to either the augmenting or the inhibitory factor of the supernatant.     

Immunogen structure and cell selection. I. Cellular requirements for the in vitro response to moderately haptenated forms of the solid-phase immunogen DNP-O-Bio-Gel-P.

The cellular requirements for the in vitro response to DNP-O-Bio-Gel-P were determined. Results of these experiments indicate that: 1) splenic adherent cells are required for generation of anti-hapten PFC to the solid phase immunogen; and 2) cultured nude-mouse spleen cells have far less capacity to respond to the immunogen than do cells from wild type mice. These experiments suggest that both T and B cells are required for responsiveness to the moderately haptenated form of DNP-O-Bio-Gel-P.     

Strain difference in the radiosensitivity of immunocompetent cells and its influence on the residual host-vs-graft reaction in lethally irradiated mice grafted with semiallogeneic bone marrow

A striking difference in radiosensitivity was noted between C3H/He (H-2k) and C57BL/6J (H-2b) strain mice when assessed by primary anti-SRBC PFC response of intact animals and primary cell-mediated lympholysis (CML) response of spleen cells to allogeneic cells in vitro, the C3H strain being more radioresistant. On the other hand, when C3H and B6 mice were exposed to 6.62 to 10.40 grays (Gy) of x-rays and then were transplanted with 2 X 10(6) bone marrow cells from B6C3F1 (H-2b/k) donor mice within 3 hr or at 24 hr after radiation exposure, the early mortality caused by residual host-vs-graft (HVG) reaction was much higher when C3H mice were used as recipients. Furthermore, the proportion of surviving animals manifesting host-type lymphohemopoiesis, i.e., host-type revertants, was much higher in B6C3F1 to C3H than in B6C3F1 to B6 combination. Spleen cells from such host-type revertants manifested strong anti-donor reactivity when assessed by mixed lymphocyte reaction (MLR) and/or CML in vitro. Increase of radiation doses to the recipients to 10.40 Gy resulted in 100% survival and 100% donor-type lymphohemopoiesis in both groups of chimeras. These results indicate strongly that a genetic difference in radiosensitivity of immune system of the recipients can greatly influence the magnitude of residual HVG reactions observed in hybrid to parental strain bone marrow transplantation in mice.     

In Vitro Proliferative Response and Polyclonal Antibody Production in Spleen Cells of Immunologically Defective CBA/N and C3H/HeJ Mice by Water-Soluble Adjuvant (Bu-WSA) Extracted from Bacterionema matruchotii

Mitogenicity and the polyclonal plaque forming cell(PFC)-inducing property of a water soluble-adjuvant extracted from Bacterionema matruchotii by butanol (Bu-WSA) were examined in vitro in the spleen cells of hybrid (CBA/N female × BALB/c male) F1 mice and C3H strain of mice. The hybrid F1 male cells which expressed a CBA/N-defect were unable to respond to Bu-WSA, when assessed by the incorporation of [³H] thymidine into the cells and the generation of anti-trinitrophenyl (TNP)-PFC or autoantibody PFC defined by the anti-bromel-ain-treated mouse erythrocyte PFC assay. However, hybrid F1 female cells with normal traits responded to Bu-WSA. Cultured spleen cells of bacterial lipopolysaccharide (LPS)-nonresponsive C3H/HeJ mice responded to Bu-WSA as in the case of cells of LPS-responsive C3H/He mice, and the [³H]thymidine-uptakes and the numbers of PFC in these culture cells increased. Re-extraction of Bu-WSA by phenol did not affect its activities, while the activity of butanol-extracted LPS on C3H/HeJ cells decreased after re-extraction by the same procedure with phenol.     

Polyclonal Antibody Production in Murine Spleen Cells Induced by Staphylococcus

Polyclonal plaque-forming cell (PFC) responses in murine spleen cells induced by Staphylococcus aureus and S. epidermidis were studied. Injection of Balb/c mice with S. aureus strain 248βH resulted in the generation of anti-trinitrophenyl (TNP) and anti-sheep red blood cell PFC in their spleens. Cultures of Balb/c spleen cells in the presence of S. aureus 248βH, Cowan I, or a protein A-deficient mutant yielded many anti-TNP PFC. The larger the number of organisms that were added to the cultures, the better was the PFC response. Both living and killed organisms, were capable of inducing the response, but an excess of living 248βH organisms in the cultures abrogated the response. All of the organisms (12 strains of S. aureus and 11 strains of S. epidermidis) freshly isolated from patients had the ability to induce the polyclonal PFC response in cell cultures. These organisms stimulated cultured C3H/HeJ mouse spleen cells, which were unresponsive to bacterial lipopolysaccharide (LPS). Cultured cells from the spleens of athymic nu/nu mice also responded to these organisms, and the number of PFC in nu/nu cell cultures was always greater than that in nu/+ cells prepared from a haired litter mate. Moreover, the responses of nu/nu spleen cell cultures to which nylon wool column-filtered splenic nu/+ T cells were added were lower than expected. These findings suggest that the polyclonal PFC response to staphylococci is thymus independent, but that the magnitude of the response is regulated by mature T cells. Cultures of macrophage-depleted spleen cells responded to the organisms to an extent similar to that of the control. The 248βH organisms were less capable of stimulating spleen cells of 2-week-old mice (i.e., early maturing B cells) than LPS. However, spleen cells from adult (7-week-old) and aged (9-month-old) mice responded well to both the organisms and LPS. Previous sensitization with the organisms in vivo did not affect any polyclonal responses of spleen cells in vitro to either the organisms or LPS. The role of staphylococcal protein A in the polyclonal PFC response to staphylococci is discussed.     

Interleukin-6 production in Mycobacterium bovis BCG-infected mice.

Mycobacterium bovis BCG and its subcellular components (bacterial extract, culture filtrate, purified protein derivative, and muramyl dipeptide MDP) are potent in vitro IL-6 inducers in spleen cell cultures from uninfected and BCG-infected BALB/c mice. Both plastic adherent and nonadherent spleen cells are capable of producing IL-6. Athymic nude mice produce more IL-6 than euthymic mice, suggesting that monocyte/macrophages are the main IL-6-producing cells in response to BCG. Finally, IL-6 production seems to be controlled to some extent by T lymphocytes, as down-regulation of CD4+ cells resulted in a marked increase in IL-6 production. Interferon-gamma does not seem to be involved in this regulation.     

Reversal of Con A-induced suppression by a platelet-derived factor which binds to activated suppressor T cells

A platelet-derived factor found in serum as well as in platelet releasate prepared either with calcium ionophore or with thrombin was shown to reverse Con A-induced suppression of the plaque forming cell (PFC) response to sheep erythrocytes (SRBC) in vivo in (CB6)F1 mice. In addition, as shown previously, lymphoma cell-induced suppression in SJL mice was similarly reversed. The factor could be injected prior to Con A on the day before SRBC injection, or on the same day as antigen with comparable results. It also enhanced PFC responses in the absence of Con A. Suppressor cell induction by Con A in vivo, as demonstrated by assay on PFC responses of normal spleen cells in vitro, was abrogated by simultaneous injection of the platelet factor. Cells from mouse spleen and lymph node, but not from thymus could absorb the factor from human serum at 4 degrees C. The phenotype of the relevant spleen cells was L3T4-, Ly1-, Ly2+, Thy1+, Ly22+, Qa1+, Qa4+, Qa5+, and Ly6.IE+. These results suggest that this factor binds to activated peripheral T cells of the suppressor cell phenotype.     

Modulation of lymphoproliferation and oxidative burst by herpes-transformed tumors.

In this study, herpes simplex virus Type 2 (HSV-2)-transformed cells (H238) and conditioned medium (CM) from H238 cell cultures were studied with respect to their effects on lymphoproliferation and the chemiluminescent oxidative burst of phagocytic cells. The H238 cells expressed a nuclear antigen detectable by fluorescent antibody testing using pooled sera from tumor-bearing mice, but no HSV-1 or HSV-2 cell membrane antigens could be found using specific monoclonal antibodies. BALB/c mice subcutaneously injected with 1 X 10(6) H238 cells developed progressively growing fibrosarcomas and depressed T lymphocyte blastogenesis in response to phytohemagglutinin (PHA) by 6 weeks post-injection when compared to non-injected controls. In contrast, oxygen radical production was increased by nearly 28-fold in the tumor-bearing subjects at this time. Incubation of normal mouse spleen cells in 100 microliters to 500 microliters of CM/ml resulted in significant dose-dependent suppression of PHA-induced lymphoproliferation. This was seen when the total spleen cell population was used, as well as after removal of the adherent cells, thereby suggesting that the inhibitory effect was not due to activation of adherent suppressor cells by the CM. However, the oxidative burst of total and adherent spleen cells from normal mice was significantly enhanced by the presence of either the H238 cells or their CM. In contrast, oxygen radical production by J774A.1 cells (a BALB/c mouse macrophage cell line) was depressed by H238 cells. Our results show that H238 tumors can alter lymphocyte as well as phagocytic cell functions both in vivo and in vitro. These tumor-induced modulations may occur via secretion of soluble factors or direct cell-to-cell interactions and, thus, may influence the outcome of immunotherapy in the tumor-bearing host.     

The role of cellular proliferation in the induction of experimental autoimmune thyroiditis (EAT) in mice

Experimental autoimmune thyroiditis (EAT) can be induced in susceptible strains of mice by injection of mouse thyroglobulin (MTg) and adjuvant. Lymphocytes from immunized mice develop a proliferative response to MTg which generally correlates with the development of EAT. We utilize a cell transfer system wherein spleen cells from CBA/J mice primed with MTg and lipopolysaccharide (LPS) in vivo are activated by culture with MTg in vitro to transfer EAT to naive recipients. In vivo priming of CBA/J mice is required to develop an antigen specific proliferative response to MTg. This response is optimal between 48 and 90 hr of culture at an MTg concentration of 125-250 micrograms/ml. The correlation between proliferation and transfer of EAT is not absolute as primed Balb/c X CBA/J F1 and AKR lymphocytes do not proliferate detectably in response to MTg but can be activated to transfer EAT; primed Balb/c lymphocytes neither proliferate nor transfer EAT. Proliferation per se is not sufficient to activate cells to transfer EAT as culture with nonspecific mitogens is not effective in activating primed CBA/J spleen cells to transfer EAT. However, lymphoblasts generated during in vitro culture of primed CBA/J spleen cells with MTg are responsible for transfer of EAT; small lymphocytes are ineffective. We conclude that antigen specific proliferation in response to MTg is essential in activating lymphocytes in vitro to transfer EAT.