Identification of effector T cells responsible for enhanced antitumor immunity induced by tumor vaccine and pyran copolymer

Summary The combination of concanavalin A-bound, but not concanavalin A-free, L1210 murine leukemia vaccine and pyran copolymer produced an enhanced therapeutic response in L1210-bearing mice. Immunoprophylactic experiments showed that the effective combination produced enhanced antitumor immunity as determined by refractoriness to the inoculation of live L1210 cells. In vitro antiproliferation and intraperitoneal adoptive transfer tests showed that antitumor effector cells were detected in the spleen and peritoneal cavity of primed mice after, but not before, live L1210 cell inoculation. These effector cells were identified as T cells on the basis of their non-adherence to plastic flasks and sensitivity to treatment with anti-mouse brain-associated T cell antigen antisera and complement. Although non-T cell populations, including macrophages of mice primed with L1210 vaccine and pyran copolymer, inhibited the in vitro proliferation of L1210 cells, we obtained evidence suggesting that they were not the primary antitumor effector cell populations responsible for the in vivo elimination of the inoculated L1210 cells.     

Development of host-dependent high-grade tumor-specific immunity through a novel mechanism triggered by the Lyt-2+ tumor-specific T cell clone (K7L) that induces temporal growth of L1210 leukemia-K7L-variant

When a murine leukemia L1210-specific Lyt-2+ T cell clone, K7L, was injected i.p. into CD2F1 mice together with L1210, the normal growth of L1210 in the peritoneal cavity of the mice at the early stage (days 0 to 5) was strongly inhibited, but L1210 grew progressively at the middle-stage (days 5 to 10), and then was rejected at the late stage (days 10 to 20). The mice thus survived for long times (more than 60 days), whereas the normal control injected with L1210 alone died within 14 days. The L1210 that grew at the middle stage in mice initially inoculated with L1210 together with K7L was a K7L-insensitive (K7L-) variant. All of eight tumor clones established from L1210-K7L- by limiting dilution was insensitive to the antitumor activity of K7L, and this property of tumor clones was stable after repeated in vitro passage. The initial depression of the L1210 growth by K7L followed by growth and rejection of the variant L1210-K7L- by the host T cell activity was then found to prepare a strong, long-lasting (more than 3 mo) immunity to protect mice against the high-dose (10(7) cells per mouse) challenge of original L1210. Corresponding to this result, definite tumor (L1210)-specific cytotoxic T lymphocyte (CTL) activity against both variant and original L1210 targets was developed by antigen (L1210) restimulation in the culture of spleen cells from these mice, but was not increased to a detectable level before L1210-K7L- variant started to grow. It was suggested that the 1210-K7L- variant and the original L1210 should have the common tumor-specific antigen that was independent of the K7L-reactive antigen, and that original L1210, whose growth was retarded by K7L, primed the host with the common antigen to be enormously boosted by the subsequently growing L1210-K7L- variant.     

Non-cytotoxic activity of pyran copolymer-induced macrophages associated with potentiation of tumour vaccine in recipient mice

Summary Mice inoculated with both L1210 murine tumour vaccine and pyran copolymer were more resistant to L1210 than those inoculated with either of these agents alone. Rabbit anti-mouse thymocyte globulin and silica reduced the augmented resistance of these mice, suggesting the involvement of activated anti-tumour T cells and macrophages in the augmented resistance. We studied the activation of these two cells separately and examined the possible contribution of pyran copolymer-induced peritoneal cells to the augmented resistance to an inoculation of live tumour. Pyran copolymer-induced peritoneal cells endowed the tumour vaccine-primed mice, but not unprimed mice, with resistance to implanted L1210 and, among those peritoneal cell populations, macrophages but not T cells were responsible for this effect since the activity was associated with a cell population which was (1) adherent to nylon wool columns, (2) sensitive to silica and (3) insensitive to anti-Thy 1.2 antibody plus complement. The pyran copolymer-induced peritoneal cells had very little antiproliferative activity when tested against L1210 in vitro and mice inoculated with these peritoneal cells did not survive a challenge of live L1210 cells much longer (<1 day) than L1210 inoculated control mice. Furthermore, the survival of L1210 vaccine-primed mice inoculated with one-tenth the amount of live L1210 (10²) was still much shorter than that of mice primed with L1210 vaccine plus pyran copolymer and challenged with ten times as many (10³) live L1210 cells. Therefore, direct tumouricidal activity was probably not a major factor in the in vivo immunological augmenting activity of the pyran copolymer-induced macrophages.     

Na(+)-dependent, active nucleoside transport in mouse spleen lymphocytes, leukemia cells, fibroblasts and macrophages, but not in equivalent human or pig cells; dipyridamole enhances nucleoside salvage by cells with both active and facilitated transport

Formycin B influx studies have shown that P388 and L1210 mouse leukemia cells, mouse L929 cells, mouse RAW 309 Cr.1 cells, LK35.2 mouse B-cell hybridoma cells and cultured mouse peritoneal macrophages express both Na(+)-dependent, active and nonconcentrative, facilitated nucleoside transport systems. In the mouse cell lines, active transport represented only a minor nucleoside transport component and was detected only by measuring formycin B uptake in the presence of dipyridamole or nitrobenzylthioinosine, strong inhibitors of facilitated, but not of active, nucleoside transport. Inhibition of facilitated transport resulted in the concentrative accumulation of formycin B in cells expressing active nucleoside transport. Concentrative formycin B accumulation was abolished by treatment of the cells with gramicidin or absence of Na+ in the extracellular medium and strongly inhibited by ATP depletion or ouabain treatment. Mouse macrophages accumulated formycin B to 70-times the extracellular concentration in the absence of dipyridamole during 90 min of incubation at 37 degrees C. Thus active transport represents a major nucleoside transport system of these cells, similarly as previously reported for mouse spleen lymphocytes. In contrast to the various types of mouse cells, active formycin B transport was not detected in human HeLa cells, human H9, Jurkat and CEM T lymphoidal cells and pig spleen lymphocytes. These cells expressed only facilitated nucleoside transport with kinetic properties similar to those of the facilitated transporters of other mammalian cells.     

Suppressor T cells induced by soluble immune complexes can adoptively transfer inhibition of cytophilic antibody receptors on macrophages.

Immune complexes (soluble antigens of L1210 and antibody to L1210) when given to allogeneic C3H mice generated suppressor cells that inhibited receptors for cytophilic antibody on macrophages. Thymocytes or nylon-nonadherent splenic T cells (4 × 10⁷) from immune-complex-treated mice transferred this suppressive activity when injected into normal syngeneic mice. Maximal suppression of macrophages occurred 4 to 6 days after transfer. In contrast, even 5 × 10⁷ nylon-adherent, non-T spleen cells from immune-complex-treated (“suppressed”) mice failed to induce macrophage suppression in the syngeneic recipients. When T-cell-depleted “B” mice were used as recipients, neither thymocytes nor splenic T cells from suppressed mice were able to transfer suppressive activity. However, the admixture of 2 × 10⁷ normal syngeneic thymocytes with 4 × 10⁷ thymocytes from suppressed mice restored the latter's ability to elicit suppression of macrophages in T-cell-deprived recipients. Peritoneal monocytes from recipients of suppressor thymocytes (to L1210) could not attach cytophilic antibody to L1210 but could attach cytophilic antibody to EL-4 and sheep erythrocytes. Thus, suppressor T cells induced by immune complexes can transfer immunologically specific macrophage suppression (inhibition of cytophilic antibody receptors) to syngeneic recipients. The suppressor cells required the cooperation of normal T cells, suggesting either recruitment of suppressor cells from, or a helper effect by, the normal T cells, in order to produce their effect.     

Factors responsible for immune resistance to L1210 murine leukemia in hyperimmune mice

Summary The serum of mice hyperimmune to L1210 leukemia was cytotoxic to L1210 cells and, to a much lesser extent, to P388 cells in the presence of complement. However, it did not suppress in vitro growth of L1210 cells, nor did it endow a recipient mouse with immunity to inoculated L1210 cells. This indicates that the serum did not play a significant role, if any, in immune protection of hyperimmune mice.Spleen and peritoneal exudate cells of hyperimmune mice suppressed the in vitro growth of L1210 but not of P388 cells. This is consistent with the fact that hyperimmune mice did not survive the inoculation of P388 cells. The immunocytes failed to suppress the in vitro growth of L1210 cells when preincubated with anti-Thy-1.2 antisera and complement. This, together with the finding that cell populations not adherent to a plastic dish suppressed in vitro growth of L1210 cells, indicates that T cells of immune spleen and peritoneal exudate cell populations were the effectors that suppressed in vitro growth of L1210 cells. Hyperimmune mice lost their immune protection in vivo following the administration of anti-thymocyte antisera, but not with carrageenan or silica, which resulted in the lethal growth of the inoculated L1210 cells. This indicates that T cells were in vivo effectors in immune protection.Hyperimmune spleen T cells endowed a recipient with immunity to L1210 leukemia when transferred in vivo. This confirmed the above results and suggests the applicability of immune cells in an adoptive immunotherapy approach.     

双歧杆菌和乳杆菌在诱发抗肿瘤免疫中的作用

双歧杆菌和乳杆菌给封闭群昆明小鼠腹腔注射,在体内激活后,胸腺细胞和脾细胞对ＣｏｎＡ刺激的增殖反应,脾贴附性细胞对ＹＡＣ－１,Ｌ９２９的细胞毒作用,以及脾贴附性细胞产生对上述二株瘤细胞的肿瘤坏死因子（ＴＮＦ）的活性都比对照动物明显增强。结果提示短双歧杆菌和嗜酸性乳杆菌给小鼠腹腔注射后,通过激活脾脏淋巴细胞和贴附性细胞（巨噬细胞）所介导的免疫功能而明显地增强宿主的抗肿瘤活性。     

Generation of specific helper cells and suppressor cells in vitro for the IgE and IgG antibody responses.

Normal splenic lymphocytes from BDF1 mice were cultured on ovalbumin (OA)-bearing syngeneic peritoneal adherent cells for 5 days and their subsequent helper function was tested by an adoptive transfer technique. Lymphocytes harvested from the culture were mixed with DNP-KLH-primed spleen cells and transferred into irradiated syngeneic mice followed by challenge with DNP-OA. The results showed that the cultured lymphocytes has helper function for both IgE and IgG anti-DNP antibody responses. Depletion of mast cells and T cells in the peritoneal adherent cell preparations did not affect the generation of helper cells in the culture. The helper function of the cultured lymphocytes was abolished by the treatment with anti-theta antiserum and complement and was specific for ovalbumin. The OA-specific helper T cells were generated in vitro by the culture of a T cell-rich fraction of normal spleen on T cell-depleted OA-bearing peritoneal cells. If the normal splenic lymphocytes or T cell-rich fraction were cultured with 10 mug/ml of OA in the absence of macrophages, cultured lymphocytes lacked helper function. The transfer of splenic lymphocytes or splenic T cells cultured with soluble OA to normal non-irradiated mice, however, suppressed both IgG and IgE antibody responses of the recipients to subsequent immunization with DNP-OA. The suppressor cells were sensitive to anti-theta antiserum and complement and their activity was specific for OA. The cultured cells transferred into normal mice did not suppress anti-hapten antibody response to DNP-KLH. Normal lymphocytes cultured on OA-bearing macrophages and had helper function in adoptive transfer experiments failed to suppress antibody response of non-irradiated recipients to DNP-OA. The results indicate that OA-bearing macrophages primed T cells and generated helper T cells, whereas the culture of normal lymphocytes with soluble OA in the absence of macrophages generated suppressor T cells.     

In situ activation of syngeneic tumour-specific cytotoxic T lymphocytes: Intra-pinna immunization followed by restimulation in the peritoneal cavity

Summary Tumour-specific cytotoxic T lymphocytes (CTL) are usually obtained after immunization in vivo and restimulation of immune cells in vitro. We here describe the generation of syngeneic tumour-specific CTL within no more than 9 days by priming and restimulation in vivo. This is achieved only if the correct sites are used both for primary immunization (ear pinna) and for restimulation (peritoneal cavity). The kinetics of immune T cell induction and of the secondary response in vivo will be reported. While a secondary CTL response could be generated in the peritoneal cavity, this was not possible in the spleen, no matter which routes of antigen restimulation were used. Upon transfer of immune spleen cells into the peritoneal cavity but not into the spleen, a secondary response could be generated upon in situ restimulation, indicating the importance of the correct microenvironment for this type of response. The peritoneal effector cells were true T cells and recognized a tumour-associated antigen in association with the K^d major histocompatibility (MHC class I) antigen. Finally the activated tumour-specific peritoneal exudate cells were able to transfer protective immunity without exogenous interleukin-2 into normal syngeneic mice.     

Immunopotentiating Activity of Whole Cells and Mini-Cells of Salmonella typhimurium

Salmonella typhimurium strain 9 produces mini-cells during cell proliferation. Mini-cells are viable but cannot proliferate since they do not contain chromosomal DNA. Effects of whole cells and mini-cells of S. typhimurium on the immune responses were investigated, with the following results. Phagocytosis of peritoneal macrophages was enhanced by in vivo stimulation of both whole cells and mini-cells. Cellular immunity against L1210 cells (mouse leukemia cells) and Sarcoma 180 cells was also enhanced by both whole cells and mini-cells. Mini-cells slightly stimulated in vitro blast cell transformation of normal mouse lymphocytes. Whole cells of S. typhimurium induced antibody-forming cells to produce IgG of higher affinity but mini-cells did not. Mini-cells were not directly cytotoxic for normal lymphocytes or L1210 cells.     

Isolation of Mouse Peritoneal Cavity Cells

The peritoneal cavity is a membrane-bound and fluid-filled abdominal cavity of mammals, which contains the liver, spleen, most of the gastro-intestinal tract and other viscera. It harbors a number of immune cells including macrophages, B cells and T cells. The presence of a high number of naïve macrophages in the peritoneal cavity makes it a preferred site for the collection of naïve tissue resident macrophages (1). The peritoneal cavity is also important to the study of B cells because of the presence of a unique peritoneal cavity-resident B cell subset known as B1 cells in addition to conventional B2 cells. B1 cells are subdivided into B1a and B1b cells, which can be distinguished by the surface expression of CD11b and CD5. B1 cells are an important source of natural IgM providing early protection from a variety of pathogens (2-4). These cells are autoreactive in nature (5), but how they are controlled to prevent autoimmunity is still not understood completely. On the contrary, CD5⁺ B1a cells possess some regulatory properties by virtue of their IL-10 producing capacity (6). Therefore, peritoneal cavity B1 cells are an interesting cell population to study because of their diverse function and many unaddressed questions associated with their development and regulation. The isolation of peritoneal cavity resident immune cells is tricky because of the lack of a defined structure inside the peritoneal cavity. Our protocol will describe a procedure for obtaining viable immune cells from the peritoneal cavity of mice, which then can be used for phenotypic analysis by flow cytometry and for different biochemical and immunological assays.     

Studies on the regulation of lymphocyte reactivity by normal and activated macrophages.

To determine the potential role of macrophages as regulators of the immune response, the effect of mouse peritoneal macrophages on transforming mouse spleen lymphocytes was investigated. Mitogen and antigen stimulated lymphocyte transformation, as measured by DNA synthesis, was enhanced by all concentrations of normal macrophages tested, but only by low concentrations of activated macrophages. High concentrations of activated macrophages markedly inhibited lymphocyte transformation. This inhibition occurred whether lymphocyte DNA synthesis was measured by incorporation of [³H]TdR or of ³²P. Activated macrophages cultured with lymphocytes within 4 hr of being removed from the peritoneal cavity inhibited lymphocyte transformation. When activated macrophages were cultured alone for 24 or more hours before addition of lymphocytes, enhancement of transformation was noted. Once lymphocytes were exposed to activated macrophages, they could not be induced to undergo transformation in the presence of Con A. Whereas heat-killed activated macrophages, which appeared intact morphologically, lost their capacity to inhibit lymphocyte transformation, macrophages treated with mitomycin C to inhibit DNA synthesis retained this capacity. Syngeneic and allogeneic macrophages had similar inhibitory ability. Supernatants from cultures of many cell types (including normal or activated macrophages, lymphocytes, lymphocytes plus macrophages, and L cells) inhibited [³H]TdR incorporation by both mitogen stimulated lymphocytes and tumor cells. These studies demonstrate the capacity of macrophages to regulate lymphocyte transformation in vitro and suggest a role for these cells as regulators of cell-mediated immunity in vivo.     

Immunological Properties of TtT/M-87 Cell Line Established from Murine Pituitary Tumor-Associated Macrophages

TtT/M-87 cell is a macrophage cell line established from thyrotropic pituitary tumor tissues in mouse. In this paper, we report the immunological properties of M-87 cells as a model of tumor-associated macrophage. Contrasting with resident peritoneal macrophages, M-87 cells constitutively secreted small but significant amounts of TNF-α and IL-1α, which were detectable in both biological assays (cytotoxic activity for L929 and co-mitogenic activity for Con A-induced T cell proliferation, respectively) and ELISA, and produced larger amounts of these cytokines upon stimulation with LPS. They expressed MHC class II molecules on their cell surface without stimulation by IFN-γ. The accessory or antigen-presenting cell activity in antibody-producing response of spleen lymphocytes to sheep red blood cells was shown to be much higher in M-87 cells than normal peritoneal macrophages. In addition, when normal spleen lymphocytes were cultured with allogeneic tumor cells, such as EL-4 and S-180, in the presence of M-87 cells, lymphocytes reactive to stimulator cells were activated to manifest inhibitory effect on the tumor cell growth and also to manifest specific cytotoxic effect on the allogeneic tumor cells. These results show that M-87 cells derived from tumor-associated tissue are activated macrophages and that they are inhibitory to tumor cell growth and augmentative in the induction of T-cell-mediated immune responses.     

Regulation of macrophage Ia expression in mice with severe combined immunodeficiency: induction of Ia expression by a T cell-independent mechanism

We have studied the expression of Ia molecules by macrophages from mice with severe combined immunodeficiency (CB-17 scid) that lack demonstrable T cell and B cell functions. CB-17 scid mice had approximately normal numbers of Ia-bearing macrophages in the peritoneal cavity, spleen, and liver. Peritoneal macrophages responded in culture to T cell-derived lymphokines with enhanced expression of Ia molecules. However, unlike immunocompetent controls, SCID mice could not enhance Ia expression in an antigen-specific T cell-dependent manner after secondary challenge in vivo with a conventional protein antigen such as hemocyanin. Further demonstration of their T cell deficiency was the failure of CB-17 scid spleen cells to proliferate and produce IL 2 in response to the T cell mitogen, concanavalin A. Upon infection with Listeria monocytogenes, CB-17 scid mice developed chronically high loads of bacteria, whereas CB-17 control mice eliminated all viable bacteria and became resistant to secondary infection. However, Listeria-infected CB-17 scid mice did show, in parallel with the CB-17 controls, an unexpected and striking increase of Ia-positive macrophages. These data indicate that induction of Ia expression in macrophages can occur via a mechanism that is independent of mature T cells.     

Nonrecirculating memory T lymphocytes in cellular resistance to infection

Acquired resistance of rats to intracellular infection with Listeria monocytogenes rests on the cooperation between sensitized mediator lymphocytes and effector macrophages. Large numbers of specific T lymphoblasts, capable of transferring resistance to recipients, appear in central lymph shortly (3–6 days) after subcutaneous infection of rats. In contrast, late-phase immunity is poorly transferrable with thoracic duct lymphocytes (TDL) despite high levels of specific resistance observed in spleen, liver, and testes of actively immunized animals. That late-phase immunity is mediated partly by resident, nonrecirculating T cells is attested to by the ineffective transfer of resistance from preinfected to normal partners of parabiotic rats. Transfer studies with thoracic duct cells and peritoneal cells from the stimulated and unstimulated peritoneal cavity seem to suggest that resident T cells mediating late-phase resistance are the progeny of lymphoblasts that extravasated during early phase. Assays measuring the proliferative response upon antigen stimulation in vitro support the concept of a gradual redistribution within the animal of memory T cells.     

Cellular origin and induction of pregnancy-associated alpha 2-glycoprotein synthesis in pregnant rats

The synthesis of alpha 2-PAG was measured and compared in tissues and cells from normal non-pregnant females, and maternal and fetal rats in vitro to define the target cells hormonally regulated during pregnancy. Synthesis was measured by [L-14C]leucine incorporation into immunochemically isolated alpha 2-PAG and confirmed by radioimmunodiffusion. alpha 2-PAG synthesis was demonstrated in maternal peripheral blood leucocytes, placenta, breast, spleen, liver and fetal peripheral blood leucocytes and liver. Maternal and fetal liver were the most active tissue producers and fetal liver synthesized 4 times more alpha 2-PAG than did maternal liver. Furthermore, fetal peripheral blood leucocytes synthesized 2 times more alpha 2-PAG per cell than did these same maternal cells. A direct comparison of synthesis by cells from pregnant and non-pregnant female rats revealed that (1) maternal peripheral blood leucocytes synthesized 5 times more alpha 2-PAG per cell than did normal leucocytes, although these same cells synthesized approximately equal amounts of total cell protein per cell, (2) maternal peritoneal exudate macrophages also synthesized 5 times more alpha 2-PAG per cell than did macrophages obtained from normal female rats, and total protein synthesis by these cells also closely paralleled each other, (3) maternal and fetal plastic-adherent peripheral blood monocytes synthesized 22 and 58 times more alpha 2-PAG per cell respectively than did normal monocytes, (4) maternal and fetal non-adherent lymphocytes synthesized 8 and 16 times more alpha 2-PAG per cell respectively than did normal lymphocytes and (5) fetal monocytes and lymphocytes synthesized 3 and 2 times more alpha 2-PAG per cell than did maternal monocytes and lymphocytes.(ABSTRACT TRUNCATED AT 250 WORDS)     

Suppressive effect of adrenalectomy on growth of L1210 leukemic cells in ascites

This study was designed to evaluate the effect of adrenalectomy on growth of L1210 leukemic cells in ascites of BDF1 mice. Varying doses of 1.5 x 10(4), 5.0 x 10(5), and 1.5 x 10(6) viable tumour cells were inoculated intraperitoneally into groups of either adrenalectomized or sham-operated mice. At days 4 to 7 after the inoculation, adrenalectomized mice inoculated with 1.5 x 10(4) or 5.0 x 10(5) tumour cells had a smaller number of tumour cells in ascites than sham-operated controls. However, after inoculation of 1.5 x 10(6) cells, no significant differences were found at days 2 to 4 between adrenalectomized and sham-operated mice. The growth retardation by adrenalectomy was not observed in adrenalectomized mice supplemented with 4 or 6 micrograms dexamethasone per day per mouse. It suggested that the ablation of glucocorticoids was at least partially responsible for the growth retardation observed in adrenalectomized mice. Cell kinetic analysis revealed that the difference in a potential doubling time could not explain these results. Tumour retention in the peritoneal cavity was measured using [125I]-iododeoxyuridine-labelled tumour cells as a tracer. At days 4 to 6 after inoculation of 5.0 x 10(5) labelled cells, radioactivity in the peritoneal cavity in adrenalectomized mice was about 70 per cent of that in sham-operated mice. This ratio was almost equivalent to the ratio of the number of cells in ascites of adrenalectomized mice to that of sham-operated ones. Consequently, growth retardation observed in adrenalectomized mice resulted from an increase in tumour cell migration and/or in tumour cell death, but not from an increase in doubling time.     

Suppression of in vitro primary immune response by L1210 cells and their culture supernatant: Evidence for cytotoxic effects

L1210 cells and their culture supernatants were found to inhibit the generation of PFC in the in vitro primary immune response of spleen cells to SRBC. As few as 1% of L1210 cells and 1% of culture fluid were inhibitory. Inhibition of DNA or protein synthesis of L1210 cells did not abolish their immunosuppressive activity, excluding exhaustion of culture medium as a possible mechanism of inhibition of PFC. Heating of the supernatant completely abrogated the suppressive effect and resulted in a marked increase of PFC. Daily evaluation of cell viability in the cultures revealed that, in the presence of L1210 and supernatants, the fraction of surviving cells is markedly reduced. We conclude that a direct cytotoxic effect on splenic lymphocytes and macrophages is the predominant immunosuppressive mechanism of L1210 cells and their culture supernatants.     

3H-uridine incorporation as a T lymphocyte indicator in the rat

Although about 70% of rat thoracic duct small lymphocytes labeled readily in vitro with ³H-uridine, only 3–38% of peritoneal exudate lymphocytes labeled. Since exudate cells are mostly B lymphocytes, ³H-uridine in concentrations used were presumed to label the T lymphocyte. Percentages of small lymphocytes that labeled in cell suspensions from various tissues were consistent with other estimates of T cells in those sources: 74.7% in thoracic duct, 70.2% in blood and 65.6% in spleen. When lymphopenia was induced by polyethylene ³²P strips applied to the spleen, a procedure that depletes mostly small recirculating lymphocytes, both labeled (T) and nonlabeled (B) cells were depleted in similar time sequence. Both cell types recovered at a similar rate after the spleen strips were removed. Induction of peritoneal inflammation by PPD in tubercle-bacilli immune rats caused an enhanced lymphocytic exudation but no increase in percentage of labeled (T) lymphocytes.The defect in ³H-uridine incorporation that characterizes the rat B lymphocyte seemed to be relatively specific for that RNA precurser; ³H-cytidine labeled the majority of lymphocytes in peritoneal exudate.     

The effect of low exposure-rate gamma irradiation on T and B lymphocyte function in the mouse

The effect of chronic irradiation on T and B cell numbers and function was studied in mice. Cobalt 60 gamma radiation at 6 R/hour reduced the numbers of anti-SRBC PFC in the spleen, with minimal levels recorded after total exposures of 1000-2000 R. Recovery was incomplete after 1000 R, reaching only 40-50 per cent of normal in four months and remaining at that level for the animal's lifetime. The long-term deficiency in PFC formation was not due to a quantitative lack of T or B cells since normal cell numbers were observed in the spleen 60-144 days after 1000 R. Adoptive transfer studies with combinations of bone marrow and thymus cells, or of splenic T and B cells, from normal and irradiated mice, revealed functional defects in both cell compartments during the first two months. Normal and near normal function of T and B cells occurred 100 days postirradiation, a time when the splenic in vivo response was still only 50 per cent of the controls. The latter observation suggests that the microenvironment of the chronically irradiated spleen alters factors regulating T and B cell interactions in response to a T-dependent antigen.