HLA-DR antigens render resting T cells sensitive to interleukin-2 and induce production of the growth factor in the autologous mixed lymphocyte reaction

Monoclonal anti-HLA-DR (anti-Ia) antibodies inhibited autologous mixed lymphocyte reactions (AMLR) when added from the initiation of the cultures, but not 72 hr later. The suppressive principle was removed by the stimulator non-T cells, but not by the responding T cells. Antibody-treated non-T cells lost their ability to activate T cells, whereas antibody-treated T cells could still respond to untreated non-T cells. The anti-DR antibodies prevented T cells from acquiring responsiveness to Interleukin-2 (IL-2). However, T cells previously activated by AMLR responded to IL-2 even in the presence of the anti-DR antibodies. OKT4⁺ lymphocytes synthesized IL-2 in the AMLR while OKT8⁺ cells did not. Anti-DR antibodies caused OKT4⁺ cells to become unresponsive to Interleukin-1 stimulation and inhibited the production of IL-2. Interleukin-1 (IL-1) promoted the synthesis of IL-2 in non-anti-DR-treated AMLR cultures. Since resting T cells are unresponsive to IL-2 and resting OKT4⁺ lymphocytes are unable to produce IL-2 even in the presence of IL-1, it is concluded that HLA-DR antigens render resting T cells sensitive to IL-2 and enable OKT4⁺ lymphocytes to respond to IL-1 and subsequently, to produce Interleukin-2.     

Galectin-1 synthesis in type 1 diabetes by different immune cell types: reduced synthesis by monocytes and Th1 cells

Galectins are a group of β-galactoside-binding mammalian lectins that play important roles in the regulation of the immune response by promoting T cell tolerance, blunting Th1 and Th17 responses and suppressing autoimmune inflammation. However, the synthesis of these molecules by different T helper (Th) subsets and in the context of human type 1 diabetes (T1D) has not yet been studied. Our results show that Th17 polarising conditions induce the synthesis of higher levels of galectin-1 compared to Th1-polarised lymphocytes. In the context of human diabetes, peripheral blood mononuclear cells (PBMCs) from T1D patients, either unstimulated or after stimulation, secreted significantly lower amounts of galectin-1 in vitro compared to healthy donors. The reduced galectin-1 synthesis observed in this autoimmune disease occurs in a dominant pro-inflammatory cytokine milieu and it is mainly due to the lower synthesis by monocytes. Surprisingly, CD4⁺ T helper cells from these patients secreted similar levels of galectin-1 compared to healthy donors, probably mediated by Th17 cytokines. In conclusion, CD4⁺ T helper lymphocytes from T1D patients produce normal levels of the immunoregulator galectin-1 but its reduced synthesis by monocytes helps to maintain a skewed pro-inflammatory response.     

Synthesis of immunoglobulin by rabbit lymphocytes in vitro in response to anti-immunoglobulin antiserum

Stimulation of synthesis of immunoglobulin (Ig) in vitro by Con A and anti-Ig in cultures of rabbit lymphoid cells has been analyzed qualitatively using an assay that measures the incorporation of [³H]leucine into newly synthesized proteins, followed by the specific absorption of tritiated immunoglobulin by staphylococcal protein A. Whereas Con A stimulates Ig production by spleen cells only if T lymphocytes are present, anti-immunoglobulin serum enhances Ig synthesis in the absence of T lymphocytes. In contrast, neither Con A nor anti-immunoglobulin serum stimulates peripheral blood lymphocytes to produce enhanced levels of Ig. It is concluded that both Con A and anti-immunoglobulin serum do not activate resting B cells but drive differentiation of B cells which are already synthesizing Ig. Anti-Ig acts directly whereas stimulation of B-cell Ig synthesis by Con A occurs indirectly through stimulation of T cells.     

Selective Stimulation of IgM Synthesis in Mouse B Lymphocytes by Pokeweed Mitogen

THE division of lymphocytes into thymus-derived (T) cells and bursa-equivalent-derived (B) cells is well established (reviewed in refs. 1–3). The result of antigenic stimulation in the B line of lymphocytes is a differentiation process, involving clonal expansion and ultrastructural changes, to give a specialized population of cells which synthesize and secrete immunoglobulin. In the study of these processes a major problem is the small number of cells involved in response to antigen, usually less than 1% of the total lymphocyte population. Clearly a system for activating large numbers of lymphocytes into immunoglobulin synthesis would offer considerable advantages. This seems to occur when mouse B lymphocytes are stimulated by pokeweed mitogen (PWM). In our experimental conditions, however, IgM is the only immunoglobulin class to be synthesized. The rational basis for our experiments rests on three previous observations: (1) PWM-stimulated lymphocytes develop rough endoplasmic reticulum^4,5 and might therefore be expected to be secreting cells; (2) a small proportion of enlarged (“blast”) lymphoid cells in PWM-treated human blood lymphocyte cultures contain immunoglobulin demonstrated by immunofluorescence⁶ and (3) the recent demonstration that mouse B lymphocytes are activated by PWM⁷.     

Enzymatic relationship between human Tγ+ lymphocytes and thymocytes

The lactate dehydrogenase isoenzyme pattern has been determined in human thymocytes and in peripheral blood T lymphocytes, Tγ⁺, and Tμ⁺ lymphocytes. Thymocytes have a highly significant lower activity in LDH-1 and LDH-2 together with a highly significant higher activity in LDH-3 and LDH-4 than peripheral T lymphocytes. The same differences are seen when Tγ⁺ and Tμ⁺ cells are compared. On the contrary there are almost no differences in pattern between the peripheral T lymphocytes and Tμ⁺ cells on one hand and between thymocytes and Tγ⁺ cells on the other hand. These findings suggest that both thymocytes and Tγ⁺ cells exhibit a very immature LDH pattern with regard to the Tμ⁺ cell population.     

ADAR1 Facilitates HIV-1 Replication in Primary CD4+ T Cells

Unlike resting CD4⁺ T cells, activated CD4⁺T cells are highly susceptible to infection of human immunodeficiency virus 1 (HIV-1). HIV-1 infects T cells and macrophages without activating the nucleic acid sensors and the anti-viral type I interferon response. Adenosine deaminase acting on RNA 1 (ADAR1) is an RNA editing enzyme that displays antiviral activity against several RNA viruses. Mutations in ADAR1 cause the autoimmune disorder Aicardi-Goutieères syndrome (AGS). This disease is characterized by an inappropriate activation of the interferon-stimulated gene response. Here we show that HIV-1 replication, in ADAR1-deficient CD4⁺T lymphocytes from AGS patients, is blocked at the level of protein translation. Furthermore, viral protein synthesis block is accompanied by an activation of interferon-stimulated genes. RNA silencing of ADAR1 in Jurkat cells also inhibited HIV-1 protein synthesis. Our data support that HIV-1 requires ADAR1 for efficient replication in human CD4⁺T cells.     

Increased tumor-infiltrating CD8+Foxp3+ T lymphocytes are associated with tumor progression in human gastric cancer

Background

CD8⁺Foxp3⁺ T lymphocytes have been detected in tumors. However, the distribution, phenotypic features, and regulation of these cells in gastric cancer remain unknown.

Methods

The levels of CD8⁺Foxp3⁺ T lymphocytes in the peripheral blood, tumor-draining lymph nodes, non-tumor tissues, and tumor tissues of patients with gastric cancer were detected by flow cytometry. Foxp3 induction in CD8⁺Foxp3^? T cells was investigated in vitro. The suppressive function of CD8⁺Foxp3⁺ T lymphocytes was analyzed by their effect on CD4⁺ T-cell proliferation and IFN-γ production. The percentages of CD8⁺Foxp3⁺ T lymphocytes were evaluated for the association with tumor stage.

Results

The frequency of CD8⁺Foxp3⁺ T lymphocytes in tumor tissues was significantly higher than that in non-tumor tissues, and similar results were also observed in tumor-draining lymph nodes compared with peripheral blood. Most intratumoral CD8⁺Foxp3⁺ T lymphocytes were activated effector cells (CD45RA^?CD27^?). TGF-β1 levels were positively correlated with the frequency of CD8⁺Foxp3⁺ T lymphocytes in tumor tissues, and in vitro TGF-β1 could induce the generation of CD8⁺Foxp3⁺ T lymphocytes in a dose-dependent manner. Furthermore, intratumoral CD8⁺Foxp3⁺ T lymphocytes suppressed the proliferation and IFN-γ production of CD4⁺ T cells. Finally, intratumoral CD8⁺Foxp3⁺ T lymphocytes were significantly increased with tumor progression in terms of tumor-node-metastasis (TNM) stage.

Conclusions

Our data have shown that increased intratumoral CD8⁺Foxp3⁺ T lymphocytes are associated with tumor stage and potentially influence CD4⁺ T-cell functions, which may provide insights for developing novel immunotherapy protocols against gastric cancer.     

Lysis of herpes simplex virus-infected targets: II. Nature of the effector cells

Peripheral blood lymphocytes (PBL) from patients with herpes simplex virus (HSV)-1 recurrences in the cornea only (Group I) exhibited reduced lysis of HSV-1-infected targets compared to PBL from patients with oral-facial and corneal HSV recurrences (Group II). The cytotoxic lymphocytes appeared to belong to a subpopulation of natural killer (NK-HSV) cells. Monoclonal antibodies to human lymphocyte differentiation antigens were used to define the surface phenotype of the NK-HSV cells. Most of the NK-HSV activity was mediated by lymphocytes expressing the surface markers Leu-7⁺ (HNK-1), OKT3⁺ (pan T), OKM1⁺ (myeloid and NK), Leu-2^? (cytotoxic/ suppressor T cell), and Leu-8^? (regulatory T cell). In contrast, lysis of K562 cells (NK-K562) was mediated by lymphocytes bearing the surface phenotype Leu-7⁺, OKT3^?, OKM1⁺, Leu-2^+/?, and Leu-8^?. The low level of NK-HSV activity in PBL from Group I donors appeared to be due to active suppression by suppressor T lymphocytes. Depletion of Leu-2⁺ cells from PBL of Group I donors resulted in significant augmentation of NK-HSV activity. Similar treatment of PBL from Group II donors either had no effect or slightly diminished the NK-HSV activity.     

Activation of T lymphocytes by the Fc portion of immunoglobulin

T lymphocytes are stimulated to release T-cell-replacing factors in response to Fc fragments of human IgG. Lyt 1⁺23^? T cells are directly triggered to factor production by Fc subfragments, derived from intact Fc fragments by macrophage-dependent enzymatic cleavage. These factor(s) replace T cell function in two Fc-mediated immune responses; induction of polyclonal antibody synthesis, and potentiation of anti-SRBC responses.     

Primary and secondary antibody responses of rabbits to bacteriophage T2: kinetic and quantitative analysis and suppression by antimacrophage globulin.

Antibody synthetic capacity of popliteal lymph node cells removed from rabbits at various times after immunization with bacteriophage T2 was assayed by radioimmunoassay of tissue culture fluid after incubation with ¹⁴C-leucine. Antibody synthesis began on day 2; IgM synthesis peaked on day 3; IgG synthesis peaked on day 5 and again on day 14. Reinjection of T2 one month later elicited an enhanced response which peaked sharply on day 2. The primary and secondary responses, but not priming for the secondary response, were suppressed by injection of goat antimacrophage globulin (AMG), but only when AMG was injected 1 to 3 days before T2. AMG reacted strongly with rabbit peritoneal macrophages and only slightly with rabbit lymphocytes or erythrocytes. Thus, macrophages appear to participate in the induction of antibody responses of rabbit lymph nodes to T2 and their function inhibited by AMG apparently operates only during the early phase of induction.     

T Lymphocyte Density and Distribution in Human Colorectal Mucosa,and Inefficiency of Current Cell Isolation Protocols

Mucosal tissues are critical immune effector sites containing complex populations of leukocytes in a tissue microenvironment that remains incompletely understood. We identify and quantify in human distal colorectal tissue absolute mucosal CD3⁺ lymphocytes, including CD4⁺ and CD8⁺ subsets, by direct visualization using immunohistochemistry (IHC), immunofluorescence (IF), and an automated counting protocol (r²=0.90). Sigmoid and rectal mucosal tissues are both densely packed with T lymphocytes in the mucosal compartment. Both compartments had similar densities of CD3⁺ T lymphocytes with 37,400 ± 2,801 cells/mm³ and 33,700 ± 4,324 cell/mm³, respectively. Sigmoid mucosa contained 57% CD3⁺CD4⁺ and 40% CD3⁺CD8⁺ T lymphocytes which calculates to 21,300 ± 1,476/mm³ and 15,000 ± 275/mm³ T lymphocytes, respectively. Rectal mucosa had 57% CD3⁺CD4⁺ and 42% CD3⁺CD8⁺ or 21,577 ± 332, and 17,090 ± 1,206 cells/mm³, respectively. By comparison, sigmoid mucosal biopsies subjected to conventional collagenase digestion, mononuclear cell (MMC) isolation and staining for flow cytometry yielded 4,549 ± 381/mm³ and 2,708 ± 245/mm³ CD4+ and CD8+ T lymphocytes. These data suggest only ~20.7% recovery compared to IHC results for these markers. Further studies will determine if this reflects a selective bias in only CD3⁺, CD4⁺ and CD8⁺ T cells or can be generalized to all flow-analyzed cells from mucosal tissues for phenotyping and functional testing.     

CD161 Expression Defines a Th1/Th17 Polyfunctional Subset of Resident Memory T Lymphocytes in Bronchoalveolar Cells

Alveolar resident memory T cells (T_RM) comprise a currently uncharacterized mixture of cell subpopulations. The CD3⁺CD161⁺ T cell subpopulation resides in the liver, intestine and skin, but it has the capacity for tissue migration; however, the presence of resident CD3⁺CD161⁺ T cells in the bronchoalveolar space under normal conditions has not been reported. Bronchoalveolar cells (BACs) from healthy volunteers were evaluated and found that 8.6% (range 2.5%-21%) of these cells were CD3⁺ T lymphocytes. Within the CD3⁺ population, 4.6% of the cells (2.1–11.3) expressed CD161 on the cell surface, and 74.2% of the CD161⁺CD3⁺ T cells expressed CD45RO. The number of CD3⁺CD161⁺ T cells was significantly lower in the bronchoalveolar space than in the blood (4.6% of BACs vs 8.4% of peripheral blood mononuclear cells (PBMCs); P<0.05). We also found that 2.17% of CD4⁺ T lymphocytes and 1.52% of CD8⁺ T lymphocytes expressed CD161. Twenty-two percent of the alveolar CD3⁺CD161⁺ T lymphocytes produced cytokines upon stimulation by PMA plus ionomycin, and significantly more interferon gamma (IFN-γ) was produced compared with other cytokines (P = 0.05). Most alveolar CD3⁺CD161⁺ T cells produced interleukin-17 (IL-17) and IFN-γ simultaneously, and the percentage of these cells was significantly higher than the percentage of CD3⁺CD161⁻ T cells. Moreover, the percentage of alveolar CD3⁺CD161⁺ T lymphocytes that produced IFN-γ/IL-17 was significantly higher than those in the peripheral blood (p<0.05). In conclusion, Th1/Th17-CD3⁺CD161⁺ T_RM could contribute to compartment-specific immune responses in the lung.     

Abnormal activation of the Akt-GSK3β signaling pathway in peripheral blood T cells from patients with systemic lupus erythematosus

Systemic lupus erythematosus (SLE) is a chronic autoimmune disease accompanied by the activation and proliferation of T cells and B cells. In this study, we found that the distributions of lymphocytes obtained from patients with SLE or SLE with renal disease (RSLE) were reduced in the G₀/G₁ phase and were elevated in the S phase after phytohemagglutinin treatment. Increased expression of CDK2 and decreased expression of cyclin-dependent kinase inhibitors p27^Kip1 and p21^WAF1/CIP1 were observed in RSLE and SLE lymphocytes. The phosphorylation levels of Akt473 and GSK3β (ser9) were increased in lymphocytes from the patients. Moreover, inhibition of GSK3β with lithium chloride or SB216763 induced T cell proliferation, and the most significant effects were observed in RSLE lymphocytes. These results indicate that upregulation of CDKs and downregulation of p27^Kip1 and p21^WAF1/CIP1 increased the proliferation of T lymphocytes in SLE patients. Abnormal activation of the Akt–GSK3β signaling pathway increased the proliferation of lupus lymphocytes.     

Decrease in pool of T lymphocytes with surface phenotypes of effector and central memory cells under Influence of TCR transgenic β-chain expression

Peripheral T lymphocytes can be subdivided into naive and antigen-experienced T cells. The latter, in turn, are represented by effector and central memory cells that are identified by different profiles of activation markers expression, such as CD44 and CD62L in mice. These markers determine different traffic of T lymphocytes in the organism, but hardly reproduce real antigenic experience of a T lymphocyte. Mechanisms of homeostasis maintenance of T lymphocytes with different activation phenotypes remain largely unknown. To investigate impact of T cell receptor (TCR) transgenic chains on formation of T lymphocytes, their peripheral survival and activation surface phenotypes, we have generated the transgenic mouse strain expressing transgenic β-chain of TCR 1D1 (belonging to the Vβ6 family) on the genetic background B10.D2(R101). Intrathymic development of T cells in these transgenic mice is not impaired. The repertoire of peripheral T lymphocytes in these mice contains 70–80% of T cells expressing transgenic β-chain and 20–30% of T cells expressing endogenous β-chains. The ratio of peripheral CD4⁺CD8^? and CD4^?CD8⁺ T lymphocytes remained unchanged in the transgenic animals, but the percent of T lymphocytes with the “naive” phenotype CD44^?CD62L⁺ was significantly increased, whereas the levels of effector memory CD44⁺CD62L^? and central memory CD44⁺CD62L⁺ T lymphocytes were markedly decreased in both subpopulations. On the contrary, T lymphocytes expressing endogenous β-chains had surface phenotype of activated T cells CD44⁺. Thus, for the first time we have shown that the pool of T lymphocytes with different activation phenotypes depends on the structure of T cell receptors.     

Intracellular cytokine profiles and T cell activation in pulmonary sarcoidosis

In granulomatous inflammatory lung diseases such as sarcoidosis, the balance of cytokine production by activated T cells in the lungs may influence clinical disease outcome. To investigate the potential of T lymphocytes to produce cytokines and contribute to this process, T cells from bronchoalveolar lavage (BAL) and PB from 19 patients with active lung disease were stimulated, stained, and analysed by flow cytometry for intracellular production of cytokines and expression of the activation marker CD69. Higher proportions of BAL cells expressed CD69 compared with PB, in the absence of in vitro stimulation. The expression of IFN-γ was similar in unstimulated BAL and PB T cells, and there was no association between the expression of CD69 and IFN-γ. Following stimulation, there were increased numbers of IFN-γ⁺ T cells. A similar trend was found with IL-2⁺ T cells, but there were lower levels of IL-4⁺ T cells in BAL compared with PB, and similar levels of IL-10⁺ T cells. The presence of activated T lymphocytes in BAL samples from patients with sarcoidosis, with the potential to produce Th1 type 1 cytokines may contribute to the inflammatory processes in this granulomatous lung disease. The use of intracellular flow cytometry to investigate cytokine production by BAL T cells could help to indicate potential targets for future therapy.     

Improvement of a method to reproducibly immortalize human T cells by oncogene transfection

The method to immortalize human T cells efficiently and reproduciblyby oncogene transfection was improved. T cells were first grown selectively from peripheralblood lymphocytes population of healthy donors andatopic asthma patients, and from lymph nodelymphocytes population of lung cancer patients byactivating with mitogens (phytohemagglutinin andconcanavalin A) and recombinant human interleukin-2(rhIL-2) for five days. Plasmids expressingoncogenes, such as c-Ha-ras, c-myc,c-fos, v-myb and v-jun under the controlof human cytomegalovirus promoter, were then introducedinto these stimulated lymphocytes either separately orin various combinations by electropolation. Afterculturing these transfected lymphocytes for recoveryfor 1 day, they were fed every 3–4 days. Although all the control cells died within one month,oncogene-transfected lymphocytes continued toproliferate actively even for more than severalmonths, indicating that oncogene-transfectedlymphocytes were successfully immortalized. Flowcytometric analyses revealed that most of theimmortalized lymphocytes were T cells expressingCD3⁺ surface antigen. The ratios of CD4⁺and CD8⁺ subpopulations in immortalized T cellsderived from healthy donors varied, depending onthe kinds of oncogenes used. However, CD8⁺subpopulation in immortalized T cells derived fromcancer patients and atopic asthma patients weredominant, independent of the kinds of oncogenes. These immortalized T cells showed differentproliferative responses in the presence or absence ofexogenous human rhIL-2, depending on their origin ofdonors. Furthermore, immortalized T cells derivedfrom healthy donors showed stronger cytotoxicityagainst K562 cells, suggesting that MHC-nonrestrictedkiller T cells in T cell population were alsoimmortalized. Immortalized T cell lines, whichproliferate continuously without stimulation of amitogen or antigen in medium containing a lowconcentration of rhIL-2, have been maintained for morethan 2 years without any growth rate decrease.     

Induction of HLA-unrestricted and HLA-class-II-restricted cytotoxic T lymphocytes against MUC-1 from patients with colorectal carcinomas using recombinant MUC-1 vaccinia virus

We recently reported that immunization with a recombinant MUC-1 vaccinia virus (rVMUC-1) protected C57BL/6 mice from challenge with DF3/MUC-1-positive syngeneic tumors. To elucidate whether anti-MUC-1 tumor immunity, especially MUC-1-specific cytotoxic T lymphocytes (CTI), can be induced in cancer patients by rVMUC-1, we stimulated the peripheral blood lymphocytes from patients with DF3/MUC-1⁺ or DF3/MUC-1 colon carcinomas using the autologous monocytes infected with rVMUC-1 (rVAMN). The stimulated T lymphocytes from two patients with DF3/MUC-1-positive colorectal carcinomas (rVPY+T and rVPW+T) demonstrated HLA-unrestricted cytotoxicity against MUC-1, whereas those from the patient with DF3/MUC-1-negative colon carcinoma (rVPA-T) did not. The HLA-unrestricted cytotoxicity was demonstrated by the CD8⁺ T cells possibly recognizing an epitope present on the tandem repeats. Adoptive immunotherapy who performed three times with patient PY, at 4-week intervals. The adoptive transfer of the first stimulated lymphocytes, demonstrating a high level of HLA-unrestricted cytotoxicity against MUC-1, resulted in the significant reduction of the liver metastasis of patient PY. However, HLA-unrestricted cytotoxicity against MUC-1 was extremely reduced at the second transfer and finally eliminated at the third, whereas the CD4⁺ T cells demonstrating HLA-class-II-restricted cytotoxicity against MUC-1 predominantly proliferated at the third adoptive immunotherapy treatment. The liver metastasis and the serum levels of tumor markers (carcinoembryonic antigen CA19-9) demonstrated a rapid and marked increment after the second transfer and especially after the third. These results suggest that the HLA-unrestricted cytotoxic CD8⁺ T cells against MUC-1, induced in patients with DF3/MUC-1⁺ colorectal carcinomas using rVMUC-1, correlate with the antitumor activity in vivo. Received: 22 October 1997 / Accepted: 24 April 1998     

Signaling signatures and functional properties of anti-human CD28 superagonistic antibodies

Superagonistic CD28 antibodies (CD28SAs) activate T lymphocytes without concomitant perturbation of the TCR/CD3-complex. In rodents these reagents induce the preferential expansion of regulatory T cells and can be used for the treatment of autoimmune diseases. Unexpectedly, the humanized CD28 superagonist TGN1412 caused severe and life threatening adverse effects during a recently conducted phase I clinical trail. The underlying molecular mechanisms are as yet unclear. We show that TGN1412 as well as the commercially available CD28 superagonist ANC28.1 induce a delayed but extremely sustained calcium response in human naïve and memory CD4⁺ T cells but not in cynomolgus T lymphocytes. The sustained Ca⁺⁺-signal was associated with the activation of multiple intracellular signaling pathways and together these events culminated in the rapid de novo synthesis of high amounts of pro-inflammatory cytokines, most notably IFN-γ and TNF-α. Importantly, sustained transmembranous calcium flux, activation of Src-kinases as well as activation of PI3K were found to be absolutely required for CD28SA-mediated production of IFN-γ and IL-2. Collectively, our data suggest a molecular basis for the severe side effects caused by TGN1412 and impinge upon the relevance of non-human primates as preclinical models for reagents that are supposed to modify the function of human T cells.     

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.     

Quantification of Antigen Specific T and B Lymphocytes in Mouse Spleens

SPECIFIC T and B lymphocytes can be killed by radioactive antigen indicating that both B and T cells must have antigen specific receptors^{1, 2}. Thus, the labelled cells observed by autoradiography after incubation of lymphocytes with radioactive antigen of high specific activity (reviewed in ref. 3) should represent a mixture of specific T and B cells.