Early cellular events in a systemic graft-vs.-host reaction. I. The migration of responding and nonresponding donor lymphocytes.

A systemic graft-vs.-host (GVH) reaction was initiated by the intravenous injection of parental strain thoracic duct lymphocytes (TDL) into irradiated F1 hybrid recipients with in-dwelling thoracic duct cannulae. The migration of the donor lymphocytes was followed by labeling them in vitro with either [3H] or [14C]uridine and measuring radioactivity by scintillation counting of the spleen and lymph nodes of the recipients removed 24 h after injection and in TDL collected throughout this period. The localization of labeled cells was always compared to that of a reference population of nonreactive lymphocytes, e.g. F1 hybrid, labeled with the alternative isotope (Fig. 1). A consistent surplus of the reactive label was found in the spleen which was balanced by a deficit of the reactive label in TDL; lymph nodes gave intermediate values. The same distribution pattern was noted when the reference population was a specifically unresponsive population of the parental strain. This differential distribution depends on recognition of the recipient's Ag-B antigens because when normal lymphocytes were injected together with specifically unresponsive lymphocytes into a "third party" F1 hybrid (against which both populations were reactive) there was no surplus of the normal cells in the spleen and no deficit in the lymph. Moreover in an Ag-B identical strain combination there was no detectable difference in the distribution of reactive and nonreactive populations. The distribution of a labeled reaction population can be accounted for if a substantial minority of cells are immobilized in the spleen and lymph nodes as a consequence of antigen recognition (Fig. 3). When the donor cells in the spleen were assayed 24 h after injection there was paradoxically a slight reduction in their specific GVH activity, which is at least partly because they are under-represented in a single cell suspension. The size of the splenic surplus (23%) and the thoracic duct deficit (12%) suggested that the minority of nonimmune lymphocytes which recognize each Ag-B complex carry 12% of the radioactive label in the original population. It is argued that this provides a near estimate of the frequency of T lymphocytes which can recognize each Ag-B antigenic complex.

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THE ROLE OF LYMPHOCYTES IN THE SENSITIZATION OF RATS TO RENAL HOMOGRAFTS

Journal of Experimental Medicine ◽

10.1084/jem.122.2.347 ◽

1965 ◽

Vol 122 (2) ◽

pp. 347-360 ◽

Cited By ~ 78

Author(s):

S. Strober ◽

J. L. Gowans

Keyword(s):

Lymph Nodes ◽

Thoracic Duct ◽

Femoral Vein ◽

F1 Hybrid ◽

Duct Cells ◽

Degree Of Sensitization ◽

Spleen And Lymph Nodes

In order to study the role of blood-borne small lymphocytes in the sensitization of rats to renal homografts 2 techniques for the perfusion of isolated rat kidneys were employed: (a) the in vitro perfusion of kidneys with thoracic duct cells suspended in either an artificial medium or in blood; the perfusates were then injected into rats syngeneic with the lymphocyte donors; (b) the in vivo perfusion of kidneys with blood issuing from the femoral artery and returning to the femoral vein of living rats. The degree of sensitization conferred on the recipients by the perfusates was assessed by applying a skin homograft from the kidney donor and scoring the epithelial necrosis at 6 days. The in vitro experiments indicated that parental strain thoracic duct cells, which had passed through an F1 hybrid kidney could confer upon a parental rat sensitivity to an F1 skin graft. Several perfusions with radioactively labelled lymphocytes showed that the injected cells migrated to the lymph nodes and spleen of the recipients Labelled large pyroninophilic cells were occasionally seen in the spleen and lymph nodes of recipients, and it was suggested that these had arisen from the injected cells. Although the in vitro perfusions with blood indicated that renal homografts might sensitize their hosts within 1 hour, the in vivo perfusions suggested that about 5 to 12 hours were required. The more rapid sensitization in vitro was possibly due to the more frequent opportunity for contact between lymphocytes and kidney vascular endothelium which was afforded by the conditions in vitro.

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Association of immunity and tolerance to host H-2 determinants in irradiated F1 hybrid mice reconstituted with bone marrow cells from one parental strain.

Journal of Experimental Medicine ◽

10.1084/jem.142.2.321 ◽

1975 ◽

Vol 142 (2) ◽

pp. 321-331 ◽

Cited By ~ 87

Author(s):

J Sprent ◽

H V Boehmer ◽

M Nabholz

Keyword(s):

Bone Marrow ◽

T Cells ◽

Thoracic Duct ◽

Parental Strain ◽

Bone Marrow Cells ◽

Third Party ◽

F1 Hybrid ◽

Host Type ◽

Hybrid Mice ◽

Marrow Cells

Semiallogenetic radiation chimeras were prepared by injecting heavily irradiated F1 hybrid mice with bone marrow cells from one parental strain; the bone marrow cells were treated with anti-theta serum and complement to remove T cells and injected in large numbers (2 times 10-7 cells). The mice survived in excellent health until sacrifice 6 mo later. Thoracic duct cannulation at this stage showed that the mice possessed normal numbers of recirculating lymphocytes. Close to 100% of thoracic duct lymphocytes and lymph node cells were shown to be of donor strain origin. The capacity of lymphocytes from the chimeras to respond to host-type determinants was tested in mixed leukocyte culture and in an assay for cell-mediated lympholysis (CML). Mixed leukocyte reactions (MLR) were measured both in vitro and in vivo; tumor cells and phytohemmaglutinin-stimulated blast cells were used as target cells for measuring CML. While responding normally to third party determinants, cells from the chimeras gave a definite, though reduced MLR when exposed to host-type determinants. However, this proliferative response to host-type determinants, unlike that to third party determinants, was not associated with differentiation into cytotoxic lymphocytes. No evidence could be found that unresponsiveness in this situation was due to blocking serum factors or suppressor T cells. It is argued that the results support the concept that lymphocytes responsive in mixed leukocyte culture have a different specificity to those exerting cell-mediated lympholysis.

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The lymphocyte surface. II. Separation of Fc receptor, C'3 receptor and surface immunoglobulin-bearing lymphocytes

Proceedings of the Royal Society of London Series B Biological Sciences ◽

10.1098/rspb.1974.0061 ◽

1974 ◽

Vol 187 (1086) ◽

pp. 65-81 ◽

Cited By ~ 49

Keyword(s):

Lymph Nodes ◽

Thoracic Duct ◽

Large Population ◽

Fc Receptors ◽

Fc Receptor ◽

Substantial Proportion ◽

Separation Procedure ◽

Total Recovery ◽

Surface Immunoglobulin ◽

Spleen And Lymph Nodes

A one-step separation procedure is described for both depleting and obtaining in pure form Fc receptor (FcRL), C'3 receptor (CRL) and surface immunoglobulin bearing (IgL) lymphocytes from rat lymphoid populations. The method is a modification of the Bӧyum (1968) technique for separating lymphocytes from whole blood by sedimentation on Ficoll/Isopaque, and is based on the fact that when a lymphocyte forms a rosette with sensitized erythrocytes it will sediment with the red cells rather than float with the non-rosetting lymphocytes. The technique is > 99.5% efficient at depleting thoracic duct lymphocytes (TDL) of FcRL, CRL and IgL and these subpopulations can be recovered 93-98% pure. The total recovery of lymphocytes applied is usually > 90% and the separated lymphocytes are > 95% viable. This technique allowed the cellular distribution of Fc receptors, C'3 receptors and surface Ig to be determined. It was found that ( a ) Almost all CRL carry surface Ig, although a very small sub-population of CRL (0.2-0.8%) which lacks surface Ig could regularly be detected. ( b ) A substantial proportion of IgL (12-25%) lacks C'3 receptors. ( c ) IgL and CRL which lack Fc receptors are more frequent in spleen and lymph nodes than in TDL. The proportion of this subpopulation increases in TDL after prolonged thoracic duct drainage. ( d ) Some FcRL exist which lack both C'3 receptors and surface Ig. These cells are more evident in TDL after prolonged thoracic duct drainage and in lymph nodes (20-30% of FcRL) than in early TDL or spleen (5-10% of FcRL). ( e ) The thymus contains very few FcRL, CRL or IgL. ( f ) A large population of lymphocytes exists in B rats (32-42% of TDL) which is killed by an anti-B serum but which lacks surface Ig. These cells are much less frequent in normal TDL ( < 5%) and probably also lack Fc and C'3 receptors. ( g ) Large lymphocytes probably shed their Fc and C'3 receptors, but retain their surface Ig, during S-phase. ( h ) Studies on a secondary anti-DNP response showed that a substantial proportion of direct and indirect plaque forming cells (PFC) in the spleen express Fc receptors, whereas only indirect PFC carry C'3 receptors. Virtually all PFC ( > 98%) possess surface Ig.

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The route of re-circulation of lymphocytes in the rat

Proceedings of the Royal Society of London Series B Biological Sciences ◽

10.1098/rspb.1964.0001 ◽

1964 ◽

Vol 159 (975) ◽

pp. 257-282 ◽

Cited By ~ 724

Keyword(s):

Lymph Nodes ◽

Thoracic Duct ◽

Large Scale ◽

Plasma Cells ◽

Tritiated Thymidine ◽

Autoradiographic Study ◽

Total Output ◽

White Pulp ◽

Radioactive Label

The experiments presented in this paper support the idea that the output of small lymphocytes from the thoracic duct of the rat (about 10 9 /day) is normally maintained by a large-scale re-circulation of cells from the blood to the lymph. It has been shown that the main channel from blood to lymph lies with in the lymph nodes and that small lymphocytes enter the nodes by crossing the walls of a specialized set of blood vessels, the post-capillary venules. In order to trace the fate of small lymphocytes, cells from the thoracic duct of rats were incubated for 1 h in vitro with tritiated adenosine. This labelled the RNA of about 65% of the small lymphocytes and more than 95% of the large lymphocytes; it also labelled the DNA of a proportion of the large lymphocytes. The mixture of small and large labelled lymphocytes was transfused into the blood of two groups of rats which belonged to the same highly inbred strain as the cell donors. At various times after the transfusions the thoracic ducts in one group of rats were cannulated to determine the proportion of labelled cells which could be recovered in the lymph; at corresponding times, the rats in the other group were killed and autoradiographs prepared from their tissues to determine the location of the labelled cells. The radioactive label in the RNA of small lymphocytes was stable enough to ensure that the labelled small lymphocytes which were recovered in the lymph several days after a transfusion were those which had originally been transfused into the blood. When the thoracic duct was cannulated 20 to 27 h after a transfusion, about 70% of the labelled small lymphocytes which had been transfused into the blood could be recovered from the thoracic duct over a 5-day period of lymph collection. During the first 36 to 48 h after cannulation, while the total output of small lymphocytes was falling rapidly, the proportion of labelled cells in the lymph remained approximately constant. The pool of the animal’s own cells with which the labelled cells had mixed contained between 1·5 and 2 × 10 9 small lymphocytes; this was identified as the re-circulating pool. An autoradiographic study showed that after their transfusion into the blood the labelled small lymphocytes ‘homed’ rapidly and in large numbers into the lymph nodes, the white pulp of the spleen and the Peyer’s patches of the intestine. The concentration of labelled cells in other tissues was trivial in comparison. Labelled small lymphocytes were seen penetrating the endothelium of the post-capillary venules in the lymph nodes within 15 min of the start of a transfusion; they were traced into the cortex of the nodes and finally into the medullary lymph sinuses. Labelled small lymphocytes did not migrate into the adult thymus but a few entered the thymus of newborn rats. It was concluded that the re-circulating pool of small lymphocytes was located in the lymphoid tissue, the thymus excepted, and that the rapid ‘homing’ of cells into the lymph nodes had its basis in the special affinity of small lymphocytes for the endothelium of the post-capillary venules. The interpretation of these experiments was not complicated by the presence of large, as well as of small lymphocytes in the suspensions of labelled cells which were transfused. Other experiments, in which the large lymphocytes alone were labelled with tritiated thymidine, showed that most of them migrated from the blood into the wall of the gut where they assumed the appearance of primitive plasma cells; very few divided to form small lymphocytes.

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STUDIES ON ANEMIA IN F1 HYBRID MICE INJECTED WITH PARENTAL STRAIN LYMPHOID CELLS

Journal of Experimental Medicine ◽

10.1084/jem.113.6.1095 ◽

1961 ◽

Vol 113 (6) ◽

pp. 1095-1113 ◽

Cited By ~ 6

Author(s):

Eileen Harriss ◽

Cicely Currie ◽

Joseph P. Kriss ◽

Henry S. Kaplan

Keyword(s):

Lymph Nodes ◽

Parental Strain ◽

Lymphoid Cells ◽

Severe Anemia ◽

Wasting Disease ◽

F1 Hybrid ◽

Sudden Loss ◽

Donor Tissue ◽

Immunologic Reaction ◽

Hybrid Mice

The survival of 51Cr-labeled erythrocytes has been studied in F1 hybrid mice in which wasting disease was produced by injection of parental lymphoid cells taken either from lymph nodes and thymus or from the spleen. Coincident with the development of the disease syndrome, there occurred a severe anemia accompanied by a sudden loss of circulating labeled erythrocytes, whether host or parental. This finding suggests that the anemia is not due solely to specific immunologic reaction of donor tissue against host erythrocytes.

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Effect of recent antigen priming on adoptive immune responses. III. Antigen-induced selective recruitment of subsets of recirculating lymphocytes reactive to H-2 determinants.

Journal of Experimental Medicine ◽

10.1084/jem.143.3.585 ◽

1976 ◽

Vol 143 (3) ◽

pp. 585-600 ◽

Cited By ~ 34

Author(s):

J Sprent ◽

J F Miller

Keyword(s):

Parental Strain ◽

Suppressor Cells ◽

Third Party ◽

Spleen Cells ◽

Skin Allograft ◽

Produce Cell ◽

F1 Hybrid ◽

Activated State

Information was sought on the reactivity of thoracic duct lymphocytes (TDL) from parental strain mice injected intravenously with large numbers of irradiated semiallogeneic spleen cells. TDL collected at 1 day after spleen cell injection were almost totally depleted of lymphocytes able to produce cell-mediated lympholysis (CML), a graft-versus-host (GVH) reaction, and skin allograft rejection against the H-2 determinants on the injected spleen cells. Normal or near normal responses were observed against third-party determinants. In the case of CML, there was no evidence that the unresponsiveness was due to suppressor cells. In marked contrast, the capacity of TDL to exert a specific mixed lymphocyte reaction (MLR) against the injected determinants was reduced by no more than two to fourfold; this applied whether MLR were measured in vivo or in vitro. Injection of normal rather than irradiated semiallogeneic spleen cells gave similar results. Complete and specific removal of MLR-producing lymphocytes was achieved, however, in a different system in which parental strain T cells were filtered from blood to lymph through irradiated F1 hybrid mice. Since this system presumably provided a much higher concentration of H-2 determinants to the responding lymphocytes, it is suggested that the differing results obtained with these two systems may indicate that certain cells reactive to H-2 determinants are of low affinity, their reactivity being detected in the MLR, but not by other parameters. With both systems, MLR-producing lymphocytes reappeared in the lymph after 2-3 days; the cells collected at this stage gave an MLR of altered kinetics. The present data, in toto, suggest that under certain conditions of antigen presentation, virtually all recirculating lymphocytes reactive to a given set of H-2 determinants can be induced to leave the circulation for a period of 1-2 days. After responding to the injected determinants (presumably in organs such as the spleen), the cells re-enter the circulation in an activated state after 2-3 days.

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Soufeng sanjie formula alleviates collagen-induced arthritis in mice by inhibiting Th17 cell differentiation

Chinese Medicine ◽

10.1186/s13020-021-00448-9 ◽

2021 ◽

Vol 16 (1) ◽

Author(s):

Di Hua ◽

Jie Yang ◽

Qinghai Meng ◽

Yuanyuan Ling ◽

Qin Wei ◽

...

Keyword(s):

Cell Differentiation ◽

Lymph Nodes ◽

Inflammatory Cytokines ◽

Th17 Cell ◽

Collagen Induced Arthritis ◽

Expression Levels ◽

Th17 Cell Differentiation ◽

Spleen And Lymph Nodes ◽

Seed Coats

Abstract Background Rheumatoid arthritis (RA) is a chronic autoimmune disease. Soufeng sanjie formula (SF), which is composed of scolopendra (dried body of Scolopendra subspinipes mutilans L. Koch), scorpion (dried body of Buthus martensii Karsch), astragali radix (dried root of Astragalus membranaceus (Fisch.) Bge), and black soybean seed coats (seed coats of Glycine max (L.) Merr), is a traditional Chinese prescription for treating RA. However, the mechanism of SF in treating RA remains unclear. This study was aim to investigate the anti-arthritic effects of SF in a collagen-induced arthritis (CIA) mouse model and explore the mechanism by which SF alleviates arthritis in CIA mice. Methods For in vivo studies, female DBA/1J mice were used to establish the CIA model, and either SF (183 or 550 mg/kg/day) or methotrexate (MTX, 920 mg/kg, twice/week) was orally administered to the mice from the day of arthritis onset. After administration for 30 days, degree of ankle joint destruction and serum levels of IgG and inflammatory cytokines were determined. The balance of Th17/Treg cells in the spleen and lymph nodes was analyzed using flow cytometry. Moreover, the expression levels of retinoic acid receptor-related orphan nuclear receptor (ROR) γt and phosphorylated STAT3 (pSTAT3, Tyr705) in the spleen were detected by immunohistochemistry. Furthermore, the effect of SF on Th17 cells differentiation in vitro was investigated in CD4+ T cells under Th17 polarization conditions. Results SF decreased the arthritis score, ameliorated paw swelling, and reduced cartilage loss in the joint of CIA mice. In addition, SF decreased the levels of bovine collagen-specific IgG in sera of CIA mice. SF decreased the levels of inflammatory cytokines (TNF-α, IL-6, and IL-17A) and increased the level of IL-10 both in the sera and the joint of CIA mice. Moreover, SF treatment rebalanced the Th17/Treg ratio in the spleen and lymph nodes of CIA mice. SF also reduced the expression levels of ROR γt and pSTAT3 (Tyr705) in the spleen of CIA mice. In vitro, SF treatment reduced Th17 cell generation and IL-17A production and inhibited the expression of RORγt, IRF4, IL-17A, and pSTAT3 (Tyr705) under Th17 polarization conditions. Conclusions Our results suggest that SF exhibits anti-arthritic effects and restores Th17/Treg homeostasis in CIA mice by inhibiting Th17 cell differentiation.

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EXTH-14. A NOVEL LONG-ACTING INTERLEUKIN-7 AGONIST, NT-I7, INCREASES CYTOTOXIC CD8 CELLS AND ENHANCES SURVIVAL IN MOUSE GLIOMA MODELS

Neuro-Oncology ◽

10.1093/neuonc/noaa215.368 ◽

2020 ◽

Vol 22 (Supplement_2) ◽

pp. ii89-ii89

Author(s):

Subhajit Ghosh ◽

Ran Yan ◽

Sukrutha Thotala ◽

Arijita Jash ◽

Anita Mahadevan ◽

...

Keyword(s):

T Cells ◽

Lymph Nodes ◽

Cd8 T Cells ◽

Peripheral Blood ◽

Cervical Lymph Nodes ◽

Cd8 Cells ◽

Interleukin 7 ◽

Long Acting ◽

Spleen And Lymph Nodes ◽

Mouse Glioma

Abstract BACKGROUND Patients with glioblastoma (GBM) are treated with radiation (RT) and temozolomide (TMZ). These treatments can cause prolonged severe lymphopenia, which is associated with shorter survival. NT-I7 (efineptakin alfa) is a long-acting recombinant human IL-7 that supports the proliferation and survival CD4+ and CD8+ cells in both human and mice. We tested whether NT-I7 would protect T cells from treatment-induced lymphopenia and improve survival. METHODS C57BL/6 mice bearing intracranial tumors (GL261 or CT2A) were treated with RT (1.8 Gy/day x 5 days), TMZ (33 mg/kg/day x 5 days) and/or NT-17 (10 mg/kg on the final day of RT completion). We followed for survival and profiled CD3, CD8, CD4, FOXP3 in peripheral blood over time. In parallel, we assessed cervical lymph nodes, bone marrow, thymus, spleen, and the tumor 6 days after NT-I7 treatment. RESULTS Median survival in mice treated with NT-I7 combined with RT was significantly better than RT alone (GL261: 40d vs 34d, p< 0.0021; CT2A: 90d vs 40d, p< 0.0499) or NT-I7 alone (GL261: 40d vs 24d, p< 0.008; CT2A: 90d vs 32d, p< 0.0154). NT-17 with RT was just as effective as NT-I7 combined with RT and TMZ in both GL261 (40d vs 47d) and CT2A (90d vs 90d). NT-I7 treatment significantly increased the amount of CD8+ cells in the peripheral blood and tumor. NT- I7 rescued CD8+ T cells from RT induced lymphopenia in peripheral blood, spleen, and lymph nodes. NT-I7 alone or NT-I7 in combination with RT increased the CD8+ T cells in peripheral blood and tumor while reducing the FOXP3+ T-reg cells in the tumor microenvironment. CONCLUSIONS NT-I7 protects T-cells from RT induced lymphopenia, improves cytotoxic CD8+ T lymphocytes systemically and in the tumor, and improves survival. Presently, a phase I/II trial to evaluate NT-I7 in patients with high-grade gliomas is ongoing (NCT03687957).

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