The effects of hypophysectomy and of growth hormone on the uptake of radioactive phosphorus by tissues

Normal and hypophysectomized animals, fed ad lib., both with and without growth hormone treatment, were injected with 4 microcuries of P³²/100 g. body weight, and then autopsied at varying time intervals following the injection.The plasma of hypophysectomized animals possesses a greater P³² activity than normal plasma at all intervals from 0.5 to 24 hr. after P³² injection. Maintenance of hypophysectomized animals with growth hormone results in a plasma P³²-activity level much smaller than that found in the plasma of hypophysectomized controls, yet greater than that found in the plasma of normal animals.The normal liver uptake of P³² shows a maximum between 0.5 and 1 hr. after intraperitoneal injection. Kidney shows a maximum in less than 0.5 hour, and thymus attains a maximum activity level in 2 hr. Hypophysectomy results in an increased liver, kidney, and thymus P³² uptake when compared to the normal. After hypophysectomy the time of maximal P³² activity in liver is shifted to 4 hr. after injection. Growth hormone therapy in the hypophysectomized animal lowers the P³²-uptake levels of kidney and liver toward normal, and brings down the P³² level in thymus to normal.Hypophysectomy with or without growth hormone therapy, results in a muscle P³² accumulation which is less than normal. Growth hormone administration to the normal animal causes an increased P³² accumulation by muscle.The P³² uptake by the tibia of the hypophysectomized rat is less than normal. Maintenance of the hypophysectomized rat with growth hormone prevents the decrease in bone P³² uptake due to hypophysectomy.     

Responsiveness of Thymus Cells to Syngeneic and Allogeneic Lymphoid Cells

THE thymus is necessary for the normal development of cell-mediated immunity in mice as shown by the immunological defects after neonatal thymectomy¹. Thymus cells themselves can be stimulated by allogeneic lymphoid cells in mixed leucocyte reaction (MLR)² and become killer cells or cytotoxic lymphocytes after stimulation with allogeneic spleen cells in vitro (H. Wagner and M. Feldmann, unpublished work) and in vivo^3,4. This suggests that the thymus as well as peripheral lymphoid tissues contain T cells which can be stimulated by foreign histocompatibility antigen to divide and differentiate into the cytotoxic lymphocytes which mediate cellular immunity. There have been suggestions that thymus cells might be stimulated to divide by “self” antigen, as well as foreign cells: incorporation of ³H-thymidine above background levels has been found in cultures with syngeneic spleen and thymus cells of adult rats⁵, although the experiments do not determine whether thymus or spleen cells have been stimulated. In contrast to these experiments, Howe et al. reported that only thymus cells of neonatal CBA mice reacted to allogeneic and syngeneic spleen cells of adult animals in “one way” MLR cultures^6,7. Whether the reaction of neonatal thymus cells to syngeneic adult spleen cells is recognition of “self” antigens is uncertain, since spleens of adult mice could carry antigens which do not occur in neonatal animals and are therefore “unknown” for neonatal thymus cells. We demonstrate here that neonatal thymus cells do not react to 4-day-old CBA spleen cells, but adult thymus cells do react against both allogeneic and syngeneic adult spleen cells.     

STUDIES ON THE REGULATION OF LYMPHOCYTE PRODUCTION IN THE MURINE THYMUS AND SOME EFFECTS OF A CRUDE THYMUS EXTRACT

Cell proliferation in the murine thymus was studied in vivo under normal conditions and from 0 to 24 hr after a single injection of a water-soluble extract from mouse thymus, mouse spleen, and mouse skin. The thymus extract reduced during the first 24 hr the mitotic activity 40%; the spleen extract had a weaker inhibitory effect. The skin extract had no such effect. The thymus extract and spleen extract inhibited the flux of cells into the S phase 0–8 hr after the injection of the extract. Initial labelling index was also reduced in this period. Eight hours after injection of the thymus or spleen extracts the inhibited cells initiated DNA synthesis. The rate of progression of blast cells through the cell cycle was normal 24 hr after the injection of the extracts. It was deduced from the analysis that the thymus extract inhibits processes triggering Go/Gi cells into DNA synthesis, the inhibition of G₂ efflux being of minor importance. Finally a model for the regulation of proliferating thymic blast cells and the emigration of small lymphocytes from the thymus is proposed.     

Anti-theta Antisera may Contain Anti-allotype Contamination

THETA (θ) is a tissue-specific mouse allo-antigen present in the highest concentrations in thymus and brain^1–3. Anti-θ allo-antisera are produced after multiple injections of thymus cell suspensions into hosts differing at the θ locus, but not at the principal histocompatibility locus (H-2). Anti-θ antisera have been used by many investigators^3–9 to differentiate thymus derived lymphocytes from non-thymus derived lymphocytes and to describe the relative role of each class of cells in various immunological functions. Because it is possible that some thymocytes bear immunoglobulins bound to the cell surface, we tested the hypothesis that anti-θ allo-antisera contain antibodies directed against immunoglobulin allotype specificities of the donor, as well as antibodies to the donor θ allo-antigens.     

Early Effect of Adult Thymectomy

THYMECTOMY of the adult mouse or rat results in a very slow decrease in immunological responsiveness^1–4. This is compatible with the idea that thymus function depends on a long lived population of thymus (T) cells which derive from the thymus but are subsequently independent of it⁵. Removal of the thymus in the neonatal period in the mouse could therefore have a drastic effect on the development of immunological responsiveness, because the organ is taken before it has shed enough T cells. Our aim is to show here that removal of the thymus in the adult can have an effect which is perceptible within 5 days on a cell population which has been shown to be a significant component of the immunological apparatus⁶.     

Migratory patterns of B lymphocytes: II. Fate of cells from central lymphoid organs in the chicken

Studies were made of the fate of ³H-nucleoside-labeled chicken bursa, thymus and bone marrow cells after intravenous injection into 8-day-old, irradiated chickens. A higher rate of proliferation of bursa cells was indicated by the fact that bursa cells incorporated more ³H-thymidine and ³H-adenosine than thymus cells, while ³H-uridine incorporation was similar for these two cell types. Homing patterns in recipient spleen tissue were similar for all three nucleoside-labeled bursa cells. Bursa cells localized in follicular and periellipsoidal areas, whereas thymus cells preferentially homed to periarteriolar sheaths. Bone marrow cells were found in all these locations. Bursa cells from chickens aged 4 weeks and older showed a greater tendency to home to the spleen than did bursa cells from 8-day-old donor chickens. Homing to other tissues appeared independent of donor age.     

Properties of Educated T Cells for Rosette Formation and Cooperation with T Cells

IT has been shown that the humoral antibody response in mice to many antigens requires cooperation between thymus derived lymphocytes (T cells) and bone marrow derived lymphocytes (B cells)^1,2. The B cells are the direct precursors of antibody secreting cells and, although T cells react specifically with antigen, their role is unknown^2–6.     

Evidence for Subpopulation of Mature Lymphocytes within Mouse Thymus

THERE is increasing evidence that thymus cells migrate from the thymus to the peripheral lymphoid tissues where they make up most, if not all, of the thymus-dependent population of lymphocytes^1–3. The term “thymus-derived” is thus appropriately applied to this population. Yet most thymocytes are different from peripheral lymphocytes, both in immunological competence and in surface antigenic characteristics. For example, thymocytes have more theta (θ)⁴ and less H2 antigen⁵ than do peripheral lymphocytes and in TL-positive strains of mice only thymocytes normally express the TL antigen⁶. Recently, Lance et al.⁷ found that injected thymocytes which had migrated to lymph nodes and spleen were progressively less susceptible to anti-TL and more susceptible to anti-H2 serum over the first 24 h. I report here experiments in which thymus cells injected intravenously into irradiated syngeneic mice and harvested as early as 3 h later from the peripheral lymphoid tissues can be shown to have the surface antigenic properties of peripheral thymus-derived lymphocytes rather than thymocytes. A second experiment demonstrates that at least part of the differentiation from thymocyte to thymus-derived lymphocyte seems to occur within the thymus.     

Effect of thymus cell injections on germinal center formation in lymphoid tissues of nude (thymusless) mice

Nude mice, partially backcrossed to Balb/c or DBA/2, were injected iv with 5 × 10⁷ thymus cells from the respective inbred strain. The response of these mice to immunization with Brucella abortus antigen was studied, with respect to both antibody production and the formation of germinal centers in their lymphoid tissues. The results were compared to those obtained with nude mice to which no thymus cells were given, as well as to Balb/c, DBA/2, or +/? litter mate controls.Nude mice formed less 19S as well as 7S antibody than did litter mate controls and completely lacked germinal centers in lymph nodes and gut-associated lymphoid tissue. Those nude mice which had been injected with thymus cells made a much better secondary response, both for 19S and for 7S antibody, and had active germinal centers in their lymph nodes as early as 3 wk after thymus cell injection. Intestinal lymphoid tissue in nude mice showed only slight reconstitution of germinal center activity several months after thymus cell injection and none at earlier times. Irradiated (3000 R) thymus cells appeared as effective as normal cells in facilitating germinal center appearance and 7S antibody production in the nude mice.     

Mercapturic acid biosynthesis: the separate identities of glutathione-S-aryl chloride transferase and ligandin

To examine if, as has been suggested, a peculiar proteolytic activity of thymus cell lysates might explain failures to detect immunoglobulin (Ig) biosynthesis by thymus cells, lysates of ¹⁴C leucine labelled mouse myeloma cells were incubated with a 10³ excess of unlabelled mouse thymus or spleen cell lysates, and then submitted to immune precipitation to isolate labelled Ig chains. Analysis of the immune precipitates by SDS polyacrylamide gel electrophoresis followed by radioautography failed to provide evidence for the purported proteolytic activity of the thymus cell lysates. Furthermore, thymus cell suspensions uncontaminated by plasma cells were biosynthetically labelled, then lysed in the presence or absence of Trasylol, an inhibitor of trypsin-like protesses. No labelled Ig chains could be detected under either condition of cell lysis. Evidence is presented that the detection of Ig chains synthesized by thymus cell suspensions might result from the contamination of these suspensions by plasma cells.     

Response of Mouse T and B Lymphocytes to Sheep Erythrocytes

THE primary immune response of mice to sheep red blood cells (SRBC) involves two types of lymphocytes: the antibody-forming cell series, whose precursors are found in bone marrow (B cells) and cooperating or “helper” cells of uncertain function whose precursors are found in the thymus (T cells)¹. Within 24 h of an intravenous injection of SRBC, an increase of haemolytic antibody plaque-forming cells (p.f.c.) occurs in the spleen, reaching a peak 5 days later. Part, but not all, of this increase is due to division among the B cells². T cells in the spleen also undergo a wave of mitosis, detectable from the second to the fifth day³.     

Regulation of bone marrow lymphocyte production: III. Increased production of B and non-B lymphocytes after administering systemic antigens

To examine the influence of exogenous stimuli on the genesis of lymphocytes in mouse bone marrow, the production rate and subsets of marrow lymphocytes were examined after a systemic injection of sheep red blood cells (SRBC). Radioautographic analysis after either pulse labeling or infusion of [³H]thymidine revealed a pronounced increase in the number of newly formed small lymphocytes appearing in the marrow, maximal 4–5 days after SRBC injection and dose related. The resulting expansion of the marrow lymphocyte population included both immature B cells and null cells, as shown by cell surface and cytoplasmic markers. Similar stimulation of marrow lymphocyte production followed an injection of either bovine serum albumin or mineral oil. No comparable stimulation occurred in either the thymus or the spleen. The results demonstrate that antigens and nonspecific irritants can exert a central effect in the bone marrow, producing a surge in the production of both primary B and non-B lymphocytes. The possible role of external stimulants in determining the normal rate of bone marrow lymphocyte production is discussed.     

Nuclear receptors for 1,25(OH)2 vitamin D3 in thymus reticular cells studied by autoradiography

Summary After injection of ³H 1,25(OH)₂ vitamin D₃ to rats fed a vitamin D-deficient diet, nuclear concentration and retention of radioactivity exists in reticular cells of the thymus medulla and cortex, as well as outer cells of developing Hassal's corpuscles. Lymphocytes do not show nuclear concentration of radioactivity. Nuclear concentration in reticular cells is prevented by prior injection of excess 1,25(OH)₂ vitamin D₃. The results indicate that reticular-endothelial cells contain nuclear receptors for 1,25(OH)₂ vitamin D₃ and suggest that effects of 1,25(OH)₂ vitamin D₃ on immune response and lymphocyte differentiation are indirect and mediated through genomic modulation of reticular cell functions such as messenger secretion.     

Autoradiographic Localization of <Superscript>3</Superscript>H-GABA in Rat Retina

γ-AMINOBUTYRIC acid (GABA) is present in all layers of vertebrate retinae^1–3: in the rabbit retina it seems to be most concentrated in the ganglion cell layers² while in the frog it is concentrated primarily in cell layers which are rich in amacrine cells¹. Recent autoradiographic studies of the distribution of ³H-GABA in rat brain slices after incubation in vitro suggest that the labelled amino-acid is selectively concentrated by certain neural elements^4,5. In a study of the distribution of ³H-GABA in rabbit retina after injection of the labelled amino-acid into the eye, Ehinger⁶ found that radioactivity was accumulated principally in the inner plexiform, inner nuclear and ganglion cell and nerve fibre layers. Labelling was also concentrated in some cells occupying the same position as amacrine cells and in some nerve cells of the ganglion cell layer.     

Activation of T and B thymus cells to recognize histocompatibility antigens

Lethally irradiated (A × CBA) F₁ or (A × C57BL/6) F₁ mice were injected with 10⁷ A strain thymus cells in attempts to activate donor cells to recognize CBA or C57BL/6 histocompatibility antigens, respectively. Activation could be revealed by injecting activated thymus cells (day 5 irradiated F₁ hybrid spleen cells) into corresponding unirradiated F₁ hybrid hosts. The alloantibody titers formed by these cells and the antirecognition structure (anti-RS) antibody titers induced by them were similar to those observed after injection of normal parental strain spleen cells, indicating that thymus cells had become endowed with recognition structures (RS). Alloantibodies, but no anti-RS antibodies, were present in the serum of F₁ mice given activated thymus cells treated with anti-θ and complement. It, therefore, appeared that activated thymus cells contained sufficient B cells differentiated into antibody-forming cells to give a measurable alloantibody response. On the other hand, receptors responsible for anti-RS antibody induction presumably were located on T cells. Specificity and restriction of antigenic recognition were revealed by negative results obtained when activated thymus cells were injected into F₁ hosts not containing the antigens against which activation had been directed.     

Relationship between the expression of class I antigen and reactivity of chicken thymocytes

The expression of major histocompatibility complex (MHC) class I antigens in ontogenesis and the distribution of B-F⁺ cells, defined by means of a monoclonal antibody, were studied by indirect membrane immunofluorescence tests on suspensions of thymus, bursa, spleen, peripheral blood lymphocytes (PBL) and red blood cells (RBC) from 18-day-old chicken embryos and chickens from 1–90 days after hatching. At 18 days of incubation and at the first day after hatching, RBC, PBL, and the cells from bursa and thymus are negative. The percentage of positive PBL and bursal cells increases up to 9 days after hatching. By 2 weeks after hatching almost 100 % of the RBC, PBL, bursa, and spleen cells were positive whereas the thymus showed only 20% positive cells. Analysis on 4-m-thick, frozen acetone-fixed tissue sections of thymus showed that medullary cells are positive, while the cortical area is negative. The graft-versus-host (GvH) competence of these thymus subpopulations was compared after sorting by the fluorescence-activated cell sorter and injection into MHC incompatible embryos. GvH reactivity was associated primarily with the B-F⁺ population. Double staining studies with peanut agglutinin (PNA)-fluorescein isothiocyanate and a rabbit-anti-Ig tetramethyl isothiocyanate-conjugate proved that the PNA^– thymocytes are identical with B-F⁺ thymocytes.Abbreviations used in this paper: FACS fluorescence-activated cell sorter - FCS fetal calf serum - FITC-Ig fluorescein isothiocyanate-conjugated immunoglobulin - GvH graft-versus-host - HAT hypoxanthineaminopterin-thymidine - HBSS Hanks' balanced salt solution - IIF indirect immunofluorescence - MCA monoclonal antibody - MHC major histocompatibility complex - NWL normal white Leghorn - OS Obese strain - PBL peripheral blood lymphocytes - PBS phosphate-buffered saline - PNA peanut agglutinin - RBC red blood cells - TRITC-Ig tetramethyl isothiocyanate-conjugated immunoglobulin     

Experimental study of the evaporation and expansion of a solid pellet in a plasma heated by an electron beam

Results are presented from experiments on the injection of solid pellets into a plasma heated by an electron beam in the GOL-3 device. For this purpose, two pellet injectors were installed in the device. The target plasma with a density of ～10¹⁵ cm^?3 was produced in a solenoid with a field of 4.8 T and was heated by a highpower electron beam with an electron energy of ～1 MeV, a duration of ～7 s, and a total energy of 120–150 kJ. Before heating, the pellet was injected into the center of the plasma column transversely to the magnetic field. The injection point was located at a distance of 6.5 or 2 m from the input magnetic mirror. Polyethylene pellets with a mass of 0.1–1 mg and lithium-deuteride pellets with a mass of 0.02–0.5 mg were used. A few microseconds after the electron beam starts to be injected into the plasma, a dense plasma bunch is formed. In the initial stage of expansion, the plasma bunch remains spherically symmetric. The plasma at the periphery of the bunch is then heated and becomes magnetized. Next, the dense plasma expands along the magnetic field with a velocity on the order of 300 km/s. A comparison of the measured parameters with calculations by a hydrodynamic model shows that, in order to provide such a high expansion velocity, the total energy density deposited in the pellet must be ～1 kJ/cm². This value substantially exceeds the energy density yielded by the target plasma; i.e., the energy is concentrated across the magnetic field onto a dense plasma bunch produced from the evaporated particle.     

Recruitment of null cells during graft versus host reactions in the chicken

Untreated SC (B2/B2) chicken spleen or thymus cells (2 × 10⁷) caused significantly increased [³H]thymidine incorporation in spleens of heavily irradiated FP (B15/B21) recipient chicks on Day 4 after iv injection. Mitomycin-treated SC spleen cells or spleen cells treated with rabbit anti-T-cell serum and complement failed to raise the [³H]thymidine incorporation over that in uninjected, bursa cell-injected or FP spleen cell-injected controls. However, the combination of mitomycin-treated spleen or thymus cells and anti-T-treated spleen cells caused an increased [³H]thymidine uptake, suggesting the recruitment of non-T cells into proliferation by alloreactive mitomycin-treated T cells. Bursa cells did not proliferate during GVH reactions even though they could be shown to undergo proliferation in vivo upon mitogen (lipopolysaccharide and dextran sulfate) stimulation. In contrast, anti-T-treated spleen cells from agammaglobulinemic chickens were recruited into proliferation, suggesting that the recruited cell was not only not a T cell, but also no pre-B or B cell and most likely represented a cell of the monocyte-macrophage series.     

In vivo 7 5 Se binding to human plasma proteins after administration of 7 5 SeO32−

Binding of ^{7 5} Se to plasma proteins was studied in four cancer patients who received 200–250 μCi of ^{7 5} SeO₃^2? (1.25 μg of selenium) intravenously for tumor scans. During the hour after injection, the ^{7 5} Se disappeared rapidly from the plasma. Gel filtration chromatography and dialysis experiments indicated that a large amount of the ^{7 5} Se returned to the plasma bound to protein between 1 and 6 h after injection. Up to 16% of the plasma ^{7 5} Se was found in very-low-density lipoproteins and low-density lipoproteins as early as 3 mikn after injection but very little ^{7 5} Se was found in high density lipoproteins. The very low density lipoprotein ^{7 5} Se activity declined very rapidly whereas low density lipoprotein ^{7 5} Se activity fell more slowly. Zonal ultracentrifugal studies of one subject's lipoproteins revealed a continuum of ^{7 5} Se-binding proteins from the very low density lipoprotein peak to the low density lipoprotein peak. Most of the ^{7 5} Se could be removed from the lipoproteins by denaturation with 8 M urea or treatment with 0.5 M mercaptoethanol. These treatments removed very little of the ^{7 5} Se from plasma collected 48 h after injection indicating a different type of binding of selenium in lipoproteins than in other plasma proteins.     

A Study of the Proliferative Response of Rabbit T Cells Using the Brdu-Hoechst Method

Con-A- and PHA-induced proliferation of cells from rabbit thymus, spleen and mesenteric lymph node was studied with the DNA-fluorescent probe 33258 Hoechst. the fluorescence of this probe is quenched when 5-bromo-2′-deoxy-uridine is incorporated into nascent DNA during the S phase. Fluorescence decreased with increasing content of newly formed DNA per cell. Proliferation kinetics and the number of Con-A- and PHA-reactive cells (C⁺ and P⁺ cells) were determined cytofluorometrically. Lymphocytes from control and dexamethasone (DX)-treated animals start their proliferation early: after 42 hr about 25% of the control and the majority of the DX-resistant cells finished their second cell division. Small numbers of C⁺ (12.0%) and P⁺ (3.5%) cells were found in control thymus, while these percentages were enhanced in DX thymus: 32.5 and 27.0% respectively; 50% of the spleen T cells in control and DX animals are C⁺ or P⁺ and 75% of the lymph-node T cells are C⁺ (after DX 45%) and 50% are P⁺ (after DX also 50%). It is concluded that in thymus and lymph nodes, a steroid sensitive (Ss) C⁺P^-, and in lymph nodes a Ss C⁺P⁺ cell pool is present. A mitogen non-proliferative cell pool (C^-P^-) is present in control and DX thymus.