Some histochemical observations on the effects of chorioallantoic grafts on the spleen, bursa, and peripheral blood of chicken embryos

Hatching eggs from inbred lines of chickens (inbreeding coefficient exceeds 95%) which show various degrees of resistance and susceptibility to Rous sarcoma, were used for experimentation. Adult tissues were grafted onto the chorioallantois on the tenth day of incubation and tissues of host and control embryos were harvested on the twentieth day of incubation. Enzymes were localized in tissues by histochemical procedures. Small pieces of tissue (thymus or bursa), when grafted onto the chorioallantois, increased the size of the spleen in host embryos although splenomegaly did not invariably occur. Two types of reactions were observed in the spleen, i.e., enlarged spleens with cysts or enlarged spleens which from a morphological point of view were normal. Grafts of either thymus or bursa decreased the size of the host embryo's bursa or were without effect. When weight of the bursa of host embryos was significantly less than that of control embryos on the twentieth day of incubation, this size relationship persisted in chicks four weeks post hatching. Intensity of dehydrogenase and acid phosphatase reactions in cysts of enlarged spleens and in the multinucleated giant cells investing them suggests that they consist of groups of degenerating cells. Intensity of enzyme reaction indicates that enlarged spleens of host embryos in which cysts were absent were normal. Enzyme reactions in the bursae of experimental embryos were more intense than those identified in the same tissues of control embryos. Catabolic reactions were the predominant type in grafts ten days subsequent to implantation. Grafts increased the number of erythrocytes in the peripheral blood of host embryos.     

Immune competence and spleen size scale with colony status in the naked mole-rat

Naked mole-rats (NM-R; Heterocephalus glaber) live in multi-generational colonies with a social hierarchy, and show low cancer incidence and long life-spans. Here we asked if an immune component might underlie such extreme physiology. The largest lymphoid organ is the spleen, which plays an essential role in responding to immunological insults and may participate in combating cancer and slowing ageing. We investigated the anatomy, molecular composition and function of the NM-R spleen using RNA-sequencing and histological analysis in healthy NM-Rs. Spleen size in healthy NM-Rs showed considerable inter-individual variability, with some animals displaying enlarged spleens. In all healthy NM-Rs, the spleen is a major site of adult haematopoiesis under normal physiological conditions. However, myeloid-to-lymphoid cell ratio is increased and splenic marginal zone showed markedly altered morphology when compared to other rodents. Healthy NM-Rs with enlarged spleens showed potentially better anti-microbial profiles and were much more likely to have a high rank within the colony. We propose that the anatomical plasticity of the spleen might be regulated by social interaction and gives immunological advantage to increase the lifespan of higher-ranked animals.     

Parasites and the avian spleen: helminths

A comparative analysis of the relationship between the spleen–a major organ of immunity and helminths was undertaken with bird species, using the phylogenetic regression technique. Species in which many individuals are infected with nematodes relative to the number of individuals examined for the presence of helminths (termed 'relative nematode presence') have significantly larger spleens, for a given body weight, in females (though not in males). There was little indication that this relationship depends on incidental ecological indices, the weights of other organs, or the 'relative presences' of trematodes, cestodes or haematozoa. Combined with previous, experimental, work it suggests that the avian spleen is important in resisting nematodes. Body weight is correlated with the relative presence of helminths; but even after removing body size effects, bird species which tend to be infected with trematodes are also more likely to be infected with cestodes. This paper indicates that the spleens of wild bird species are associated with macroparasites in the natural environment.     

Role of spleen in immune response to polyvalent pneumococcal vaccine.

The immune response of lymphocytes to subcutaneously administered pneumococcal vaccine was studied in five patients without spleens and in five healthy subjects. Seven days after immunisation circulating B cells synthesising IgG antipneumococcal capsular polysaccharides (anti-PCP) appeared in both groups. Twenty one days after vaccination this B cell population had disappeared and a B cell subset which secreted IgM and IgG anti-PCP in the presence of pokeweed mitogen was detected in the normal but not in the splenectomised subjects. In the splenectomised group polyclonal IgM synthesis induced by pokeweed mitogen was defective. It was concluded that the early events of the immune response to PCP may be mediated by lymph nodes but that, later, the spleen acquires a central role in producing lymphocyte subsets capable of synthesising specific antibodies and that this might explain the increased sensitivity of splenectomised subjects to pneumococcal infection.     

Purification of RNA-directed DNA polymerase from mouse spleen infected with Rauscher leukemia virus.

RNA-directed DNA polymerase was purified from spleens of Balb/c and NMRI mice infected with Rauscher murine leukemia virus. The method includes cell fractionation and lysis of microsomal fraction, chromtography on Sephadex G-200 and phosphocellulose. Estimation of molecular weight from the sedimentation rate of the purified enzyme in a glycerol gradient was consistent with a structure containing one polypeptide with a molecular weight of 70,000. Purified RLV DNA polymerase from spleen could transcribe purified DNA polymerase from purified virions. This simple preparation method offers a procedure for large scale preparation of the RNA-directed DNA polymerase which can be used for synthesis of DNA complementary to mRNA.     

The spleen of the cervidae (Gray, 1821). Quantitative-morphological studies on the spleens of stags (Cervus elaphus, L.1785) and of deers (Capreolus capreolus, L. 1758)]

The macroscopical and microscopical structure of 17 spleens of Cervus elaphus and 9 spleens of Capreolus capreolus is described. The spleens of both species exhibit structural characteristics which resemble those of the reticular "non-sinusoidal" type. These include: spleen arterial ramifications of the "magistral type" (SCHABADASCH), bilayered capsule, well developed smooth muscle cells containing trabecular networks, numerous muscle cells in the red pulp, poorly developed white pulp, splenic vein of large diameter, lack of veins associated with trabeculae, a thick tunica media in trabecular arteries and in arterial vessels of the hilus region, poorly developed SCHWEIGGER-SEIDEL sheaths, and splenic nerve trunks of considerable diameter. These structural features are comparable to those in other ruminant species having storage type spleens. However, there are differences in certain quantitative parameters between the spleen of Cervus elaphus and that of Capreolus capreolus, i.e., spleen weight versus body weight, relative volume of trabecular networks and capsular tissue, and relative amount of smooth muscle cells in trabecular tissue. On the basis of these quantitative parameters the spleen of Cervus elaphus and that of Capreolus capreolus can be classified into the system of spleen types as described by v. Herrath(1953) and should thus be ordered between the extreme storage type spleen and the extreme metabolic type spleen. The quantitative data observed in the spleen of Cervus elaphus are similar to those seen in the horse, whereas the data of the spleen of Capreolus capreolus can be compared to that of the sheep and the cow.     

The occurrence of two immunologically distinguishable beta-glucocerebrosidases in human spleen

The beta-glucosidase activity in spleen from control subjects and patients with different clinical phenotypes of Gaucher's disease was characterized. The occurrence of a soluble non-specific beta-glucosidase with a neutral pH optimum and two membrane-associated beta-glucocerebrosidases with an acid pH optimum was demonstrated. The two beta-glucocerebrosidases can be distinguished on the basis of their ability to react with anti-(placental beta-glucocerebrosidase) antibodies bound to protein-A--Sepharose 4B beads. One of the splenic beta-glucocerebrosidases (form I) is precipitated by the immobilized antibodies and the other (form II) is not. The two forms also differ in binding affinity to concanavalin A, degree of stimulation of enzymic activity by taurocholate and isoelectric point. In contrast, the Km values of the two beta-glucocerebrosidases for natural and artificial substrates are similar and both are inhibited by conduritol B-epoxide. In spleen from three patients with type 1, one patient with type 2 and one patient with type 3 Gaucher's disease form I beta-glucocerebrosidase was found to be clearly deficient, whereas the activity of form II was 25-50% of that in control spleen. The non-specific, neutral beta-glucosidase was not deficient in these Gaucher spleens. The distinct biochemical and immunological properties of non-specific beta-glucosidase and the fact that normal levels of the enzyme are present in patients with Gaucher's disease indicate, in confirmation of previous reports, that non-specific beta-glucosidase is not related to beta-glucocerebrosidase.     

Isolation of preparative amounts of polyribosomes from normal rabbit and guinea pig spleen]

Preparative amounts of polyribosomes were isolated from normal rabbit and guinea pig spleen; up to 40 optical units of the polyribosome preparation could be obtained by centrifugation in a Spinco L-2B centrifuge with SW-27 rotor. The amount of polyribosomes isolated from spleens of immune animals was 2-3 higher than that isolated from normal animal spleens. Concentration of polyribosomal preparations by lyophylization and the storage of dried preparations do not alter the sedimentation properties of the polyribosomes. The distribution pattern of normal rabbit spleen polyribosomes in a linear sucrose gradient and the sedimentation constants of the polyribosome peaks are in good agreement with data reported by some other authors for plasmocytome polyribosomes. Using electrophoresis in agarose-polyacrylamide gel the radioactive proteins synthesized in the cell culture of normal rabbit spleen it was shown that in normal spleen the average amount of globulins makes up to 35% of total protein synthesis, as reported by some authors.     

Climate,body condition and spleen size in birds

Climatic conditions may impact on the body condition of animals and thereby affect their survival prospects. However, climate may also impact directly on the survival prospects of animals by affecting the size of immune defence organs that are used for defence against parasites. We used a large long-term database on body condition and size of the spleen in birds to test for immediate and delayed relationships between climatic conditions as indexed by the North Atlantic Oscillation (NAO) and body condition and spleen mass, respectively. Across 14 species of birds, spleen mass was significantly positively correlated with the NAO index, while the delayed effect of NAO on spleen mass was not significant. Spleen mass was positively related to body condition, but body condition did not depend significantly on NAO or delayed NAO effects. Bird species with a strong positive effect of NAO on spleen mass tended to have small spleens for their body size, while species with a strong negative effect of NAO on spleen mass tended to have relatively large spleens. Since bird species with relatively large spleen have been shown to suffer more from the negative effects of parasites, we can infer that the effects of climate as indexed by NAO on the size of the spleen depends on the importance of parasite-mediated natural selection.Due to an error in the citation line, this revised PDF (published in December 2003) deviates from the printed version, and is the correct and authoritative version of the paper.     

Ligula intestinalis (L.) (Cestoda: Pseudophyllidea): plerocercoid-induced changes in the spleen and pronephros of roach, Rutilus rutilus (L.), and gudgeon, Gobio gobio (L.)

The effects of the plerocercoid of Ligula intestinalis (L.) (Cestoda: Pseudophyllidea) on the major lymphoid organs, the spleen and pronephros, of roach, Rutilus rutilus (L.), and gudgeon, Gobio gobio (L.), are described. The spleen of ligulosed roach showed a significant decrease in weight. Differential cell counts suggested this was due to a reduction in erythrocytes, despite significant increases in macrophages and vacuolated granulocytes. The spleen of gudgeon, which consisted almost entirely of erythrocytes, showed a slight reduction in weight in ligulosed fish. In both roach and gudgeon this decrease was independent of parasite burden. Differential cell counts of the pronephros from ligulosed roach revealed a significant decrease in neutrophils and increase in vacuolated granulocytes. In the pronephros of gudgeon, however, cell counts were unaffected by ligulosis. Ultrastructural observations included an apparent disintegration of vacuolated granulocytes and increased pinocytic activity in specialized endothelial cells in the spleens of ligulosed roach. Also, melano-macrophage centres and melano-macrophages increased in the spleen of ligulosed roach and gudgeon, respectively. The marked changes in spleen weight and differential cell counts in ligulosed roach and lack of such changes in ligulosed gudgeon correlate with the differential response to the parasite by these two fish species.     

Indigofera oblongifolia leaf extract regulates spleen macrophage response during Plasmodium chabaudi infection

Malaria is a major health problem that still affects numerous countries. The current study aimed to identify the role of Indigofera oblongifolia leaf extract in regulating mouse spleen macrophages during the progression of Plasmodium chabaudi infection. Three doses of the leaf extract (100, 200, and 300 mg/kg) were administered to mice inoculated with P. chabaudi infected erythrocytes. The weight of the infected mice improved after the treatment with I. oblongifolia. The infection causes disorganization of macrophage distribution in the spleen. After the mice had been treated with the leaf extract, the macrophages appeared to be reorganized in the white and red pulp areas. In addition, the I. oblongifolia leaf extract (IOLE) significantly increased the total antioxidant capacity of the mice spleens infected with P. chabaudi. The phagocytic activity of spleen macrophages was increased in the infected group as indicated by the significant decrease in the number of fluorescent particles in the spleen sections. This number increased in the mice spleens after treatment with IOLE. Based on these results, it is suggested that IOLE regulate macrophage response of the spleen during the blood stage of malaria in mice.     

Intracellular localization of enzymes in spleen. I. Reduced diphosphopyridine nucleotide cytochrome c reductase,cytochrome c oxidase,and succinic dehydrogenase in the rat and guinea pig

1. The intracellular distribution of nitrogen, DPNH cytochrome c reductase, succinic dehydrogenase, and cytochrome c oxidase has been studied in fractions derived by differential centrifugation from rat and guinea pig spleen homogenates. 2. In the spleens of each species, the nuclear fraction accounted for 40 to 50 per cent of the total nitrogen content of the homogenate, and the mitochondrial, microsome, and supernatant fractions contained about 8, 12, and 30 per cent of the total nitrogen, respectively. 3. Per mg. of nitrogen, DPNH cytochrome c reductase was concentrated in the mitochondria and microsomes of both rat and guinea pig spleens. Seventy per cent of the total DPNH cytochrome c reductase activity was recovered in these two fractions. The reductase activity associated with the nuclear fraction was lowered markedly by isolating nuclei from rat spleens with the sucrose-CaCl(2) layering technique. The lowered activity was accompanied by the recovery of about 90 per cent of the homogenate DNA in the isolated nuclei, indicating that little, if any, of the reductase is present in spleen cell nuclei. 4. Per mg. of nitrogen, succinic dehydrogenase was concentrated about 10-fold in the mitochondria of rat spleen, and 65 per cent of the total activity was recovered in this fraction. 5. Cytochrome c oxidase was concentrated, per mg. of nitrogen, in the mitochondria of both rat and guinea pig spleens. The activity associated with the nuclear fraction was greatly diminished when this fraction was isolated from rat spleens by the sucrose-CaCl(2) layering technique. Only 50 to 70 per cent of the total cytochrome c oxidase activity of the original homogenates was recovered among the four fractions from both rat and guinea pig spleens, while the specific activities of reconstructed homogenates were only 55 to 75 per cent of those of the original whole homogenates. This was in contrast to the results with DPNH cytochrome c reductase and succinic dehydrogenase where the recovery of total enzyme activity approached 100 per cent, and the specific activities of reconstructed homogenates equalled those of the original homogenates. The recovery of cytochrome c oxidase was greatly improved when only the nuclei were separated from rat spleen homogenates. 6. Data were presented comparing the concentrations (ratio of activity per mg. of nitrogen of the fraction to activity per mg. of nitrogen of the homogenate) of DPNH cytochrome c reductase in mitochondria and microsomes derived from different organs of different animals. 7. Data were presented comparing the activities per mg. of nitrogen of DPNH cytochrome c reductase in homogenates from several organs of various animals.     

Strain-specific spleen remodelling in Plasmodium yoelii infections in Balb/c mice facilitates adherence and spleen macrophage-clearance escape

Knowledge of the dynamic features of the processes driven by malaria parasites in the spleen is lacking. To gain insight into the function and structure of the spleen in malaria, we have implemented intravital microscopy and magnetic resonance imaging of the mouse spleen in experimental infections with non-lethal (17X) and lethal (17XL) Plasmodium yoelii strains. Noticeably, there was higher parasite accumulation, reduced motility, loss of directionality, increased residence time and altered magnetic resonance only in the spleens of mice infected with 17X. Moreover, these differences were associated with the formation of a strain-specific induced spleen tissue barrier of fibroblastic origin, with red pulp macrophage-clearance evasion and with adherence of infected red blood cells to this barrier. Our data suggest that in this reticulocyte-prone non-lethal rodent malaria model, passage through the spleen is different from what is known in other Plasmodium species and open new avenues for functional/structural studies of this lymphoid organ in malaria.     

Observations on the gross changes in the secondary lymphoid organs of mice infected with Nematospiroides dubius

Gross changes in the size of the secondary lymphoid organs were studied during infection with the nematode parasite Nematospiroides dubius. In the strong responder NIH strain, the wet weight of the mesenteric lymph nodes (MLN) increased rapidly following infection with 400 larvae to peak on day 28 at approximately three times the resting weight. Enlargement of the spleens was also marked but regression to normal size took place when the MLN had achieved maximum size. In contrast in C57BL/10 mice, a slow responder strain, the enlargement of the MLN following infection was relatively slow, and there was no evidence of the regression of the spleen, once maximum enlargement had been achieved. When adult worms were removed by anthelmintic, the enlarged MLN and spleens returned rapidly to normal size. However, in mice infected with irradiated larvae (25 krad) the MLN stayed enlarged, despite the absence of adult worms but the spleens of these mice returned to normal size fairly rapidly. It was suggested that irradiated worms survive, perhaps as arrested larvae in the intestinal tissue, for a fairly long time, thereby providing a continual stimulus for the MLN.     

Ca2+, Mg2+-dependent endonuclease activity in different subpopulations of spleen cells from normal and erythroleukemic mice

We have detected Ca2+, Mg2+-dependent endonuclease activity in spleen cells of normal, Friend erythroleukemic, and phenylhydrazine-treated mice. When nuclei were isolated and incubated in the presence of Ca2+ and Mg2+ ions, the activity resulted in the production of 3'-OH termini in the cellular DNA and the release of chromatin due to internucleosomal DNA fragmentation. This enzyme activity was chromatin-bound and could be extracted from chromatin in an active form in 0.35 M KCl. The majority of endonuclease activity from erythroleukemic spleens was present in nuclei of precursor erythroid cells of low buoyant density (1.025-1.05 g/ml). Uninfected normal splenic tissue contained an endonuclease activity which was almost entirely confined to a B-lymphocyte population of high buoyant density (greater than 1.07 g/ml). Erythroid cell-enriched spleens from phenylhydrazine-treated mice exhibited a distribution of endonuclease activity in cells at low and high densities reflecting a mixture of erythroid and lymphoid cells. Cloned erythroleukemic cell lines propagated in vitro lacked cells of low density and showed no detectable endonuclease activity. However, nuclei from these cell lines were susceptible to exogenously added endonuclease extracted from erythroleukemic spleen cells. These same cell lines propagated as subcutaneous tumors contained endonuclease activity and a morphologically-similar low-density cell population which accounted for the endonuclease activity in these tumors. Nuclei from cloned lymphoid cell lines, representing different B-lymphocyte phenotypes, showed differences in the presence of endonuclease activity. Among the cell lines tested, only those expressing late B-cell markers showed detectable endonuclease activity.     

Parasite-associated growth enhancement in a fish-cestode system

Parasites impose an energetic cost upon their hosts, yet, paradoxically instances have been reported in which infection is associated with enhanced, rather than diminished, host growth rates. Field studies of these parasite effects are problematic, since the pre-infection condition of the hosts is generally unknown. Here, we describe a laboratory experiment in which the growth rate and body condition of 76 laboratory-reared three-spined stickleback fishes were examined before, during and after each fish was fed the infective stage of the parasitic cestode Schistocephalus solidus. Twenty-one of these fishes went on to become infected by the cestode. Fishes were individually housed and provided with an abundant food supply to eliminate the potentially masking effects of variable competitive ability. Infection occurred independently of fish gender, size, body condition or pre-exposure growth rate. After exposure to the cestode, infected fishes grew faster (excluding parasite weight) and maintained a similar or better body condition compared with uninfected fishes, despite developing enlarged spleens. The accelerated growth could not be explained by reduced gonadal development. This result, one of few demonstrations of parasite-associated growth enhancement in fishes, is discussed with respect to other such parasite systems.     

Differential expression of HLA class II antigens in fetal human spleen: relationship of HLA-DP, DQ, and DR to immunoglobulin expression

Frozen sections of human fetal spleen from 12 to 20 wk gestation were examined by using polyclonal antibodies to Ig isotypes, monoclonal antibodies to HLA class II subregion locus products, B and T cells, and follicular dendritic cells. Scattered lymphoid cells in spleen sections from fetuses of 12 to 13 wk gestational age expressed IgM but not IgD. The appearance of lymphoid cells expressing IgD occurred at 14 to 15 wk before the formation of loose clusters of B cells at 16 wk. IgD expression was associated mainly with cells in these clusters, which by 17 wk had become definite follicles. Follicular dendritic cells were not detectable until 20 wk. OKT3-positive T cells were not detected until 17 wk, and at 20 wk constituted 5% of the nucleated cell population. HLA-DR- and DP-positive lymphocytes and macrophages were detectable in fetal spleen from 12 wk onward; DR was expressed on more cells than DP, and the numbers of cells stained by HLA-DR-specific monoclonal antibodies exceeded the number of Ig-positive cells in all spleens examined. HLA-DQ was expressed by consistently fewer cells than HLA-DR and -DP in all spleens tested. The small number of DQ-positive cells in spleens from 12- to 13-wk fetuses had the morphology of macrophages; HLA-DQ expression by lymphoid cells followed a similar pattern to IgD expression and was associated mainly with follicular lymphocytes. It could be demonstrated by double-labeling experiments that all follicular IgM-positive cells in 17- to 20-wk spleens expressed HLA-DP, DQ, and DR antigens: IgM-positive cells in 12- to 16-wk spleens and interfollicular IgM-positive cells in 17- to 20-wk spleens all expressed HLA-DR, but only 59% and 43% expressed DP and DQ, respectively. Ninety-one to 100% of IgD-positive cells in all spleens examined expressed HLA-DQ in addition to DR and DP. In these experiments IgD-negative, DQ-positive cells had the morphologic appearance characteristic of macrophages. These data suggest that class II antigens are differentially expressed on developing lymphoid cells; DR and DP expression occurring in the earliest spleens examined, with expression of DP on a subpopulation of DR-positive cells; IgD and DQ expression appears to be coincident on maturing B cells as they begin to form follicles. An immunoregulatory role for HLA-DQ in B cell development is implicated and remains to be fully investigated.     

Myelopoiesis related to perinatal spleen

Adult murine spleen is known to have a major role in the development of dendritic cell (DC) subsets, including conventional DC and plasmacytoid DC. In this lab, long-term cultures (LTCs) established from murine spleen support continuous production of novel dendritic-like cells, termed LTC-DC. An in vivo equivalent subset also exists in spleen, namely L-DC. As co-cultures using LTC-derived splenic stroma support the outgrowth of L-DC from spleen and bone marrow sources, it is likely that spleen represents an important niche for DC development. To investigate the appearance of L-DC during ontogeny, spleen was isolated from embryonic and neonatal mice of different ages for analysis of myeloid and DC subsets. Perinatal spleen was also used to establish co-cultures for identification of progenitors, and LTCs were established from spleens for assessment of stromal competence. Although spleen from 16-day embryos (E16.5) contained myeloid cells, DC subsets did not appear until day 4 after birth (D4). However, murine spleen at D0 contained progenitors, which could seed co-cultures for L-DC production. LTC could not be established from spleen until D4. The appearance of L-DC after D4 in spleen is dependent on the formation of the appropriate stromal microenvironment which occurs in the early postnatal period.     

Contact sensitivity to picryl chloride: the occurrence of B suppressor cells in the lymph nodes and spleen of immunized mice.

Mice were immunized for contact sensitivity and antibody production by painting the skin with picryl chloride. Lymph node and spleen cells taken 4 days later transferred contact sensitivity. However, cells taken at 7–8 days failed to transfer but were able to block the transfer by 4 day immune cells. These suppressor cells occurred in the regional lymph nodes, spleen and thymus. The suppressor activity of lymph node and spleen cells was due to B cells as shown by the effect of anti-θ serum and complement, nylon wool filtration and separation of EAC positive and negative cells by centrifugation on a discontinuous gradient. The transfer of fractions rich or poor in macrophages showed that the suppressor cell in the transferred population was not a macrophage. Separation using EAC rosettes suggested that B cells were responsible for the suppressor activity in the thymus.T cells isolated from the lymph nodes and spleen 7–8 days after immunization transferred contact sensitivity although the initial population was inactive. This indicates that passive transfer cells are present in the regional lymph nodes and spleen at later times after immunization but cannot be demonstrated because of the presence of suppressor B cells. However, no passive transfer cells were found in the thymus. The production of B suppressor cells required little or no T cell help and following immunization the spleens of reconstituted (B) mice were at least as active as control cells in causing suppression. There are several different suppressor cells which act in the picryl system and the B suppressor cells in immunized mice described here are distinct from the T suppressor cells in mice injected with picryl sulphonic acid.