Haemagglutinin activity in Acrididae (grasshopper) haemolymph

The haemolymph of Acrididae causes haemagglutination of human and animal erythrocytes. Thirteen of seventeen species tested had detectable activity and gave agglutination titres in the range 2–64, Melanoplus bivittatus, and M. sanguinipes showed greatest activity. Haemagglutinin activity is continuously present in male and female insects from 4th instar and throughout adulthood. Females contain slightly more activity than do males. M. sanguinipes haemolymph agglutinates rabbit, calf, human (all ABO types) guinea pig, mouse, chicken, cat, pig and sheep erythrocytes. Rabbit red cells are agglutinated most strongly and sheep and chicken cells least. M. sanguinipes haemolymph also agglutinates the protozoan Nosema locustae, a natural grasshopper pathogen. Preabsorption of haemolymph with different erythrocyte types selectively removes haemagglutinin activity suggesting the presence of multiple or heteroagglutinins. M. sanguinipes haemagglutinin is inhibited by glycoproteins, simple carbohydrates and carbohydrate derivatives. The inhibitory pattern is complex and among the sugars tested only mannose and derivatives of mannose are exclusively non-inhibitory. Haemolymph haemagglutinin activity is destroyed by heat and EDTA. It is totally precipitated by dialysis against water and may be partially recovered in phosphate or Tris buffer. Activity is stable in frozen haemolymph.     

Amino acid and protein changes in the haemolymph of developing fourth instar Chironomus tentans

The concentration of free amino acids, total soluble protein, and haemoglobin in the haemolymph of fourth instar Chironomus tentans was investigated.The concentration of the free amino acid pool increases between the early (15.7 mM/l) and mid-(33.9 mM/l) fourth larval stages followed by a decline during the late (16.9 mM/l) fourth larval period. Alanine, serine, and the amides of aspartic acid and glutamic acid are the predominant free amino acids at all stages. Physiological fluid analysis of late fourth instar haemolymph detected 32 ninhydrin positive components including 18 common amino acids plus homoarginine, ornithine, citrulline, β-alanine, α-aminoadipic acid, α-aminoisobutyric acid, and sarcosine.The concentration of total soluble protein steadily increases during fourth instar larval development to a maximum of $\text{9.3}\text{g}\text{100}\text{ml}$ followed by a decline during the pharate pupal period. A similar pattern of variation occurs in haemoglobin content which comprises from 51 to 66% of Chironomus tentans haemolymph protein.The mM percentage of individual amino acids of total haemolymph protein varies little during the fourth instar. At all stages alanine and aspartic acid are the predominant amino acids.     

Haemagglutinin activity in the haemolymph of individual Acrididae (grasshopper) specimens

Twenty-four individual grasshopper specimens representing four Melanoplus spp. contained similar broad-spectrum haemolymphatic haemagglutinin. The agglutinin activity showed highest titre toward human ABO and rabbit cells among nine types of erythrocytes tested. Titre values differed between individual insects but agglutination specificity toward different erythrocytes was similar. Agglutination of type-O red cells by individual grasshopper haemolymph was inhibited by 34 of 41 tested carbohydrates, carbohydrate derivatives, alcohols and chelating agents. Individual insects showed similar patterns of haemagglutination inhibition. Non-inhibitory compounds were mannose and mannose derivatives (excepting N-acetylneuraminate), several glucose derivatives, amino sugars and ethanol. The observations indicated that haemolymph from an individual grasshopper contained complex heteroagglutinin activity similar to that found in haemolymph pooled from several insects. Determination of minimal effective inhibitor concentrations confirmed the presence of heteroagglutinin activity primarily directed toward galactose and glucose and related α-linked glycosidic derivatives.     

The molecular basis for cold agglutination: effect of receptor density upon thermal amplitude of a cold agglutinin.

Cold agglutinin MKV is a Waldenström macroglobulin that agglutinates human erythrocytes in the cold by binding $\text{N}$-acetylneuraminosyl-containing carbohydrate chains on their surfaces. Neuraminidase-treated cells are not agglutinated but their reactivity can be restored by allowing them to adsorb hematoside (NeuNAcα2-3Galβ1-4Glcβ1-ceramide). When between 7 × 10⁴ and 10⁶ molecules are adsorbed per cell, the cells are agglutinated at 0° but not at 37°. When over 10⁶ molecules of hematoside are adsorbed, they are agglutinated at both 0° and 37°. The density of receptors on the erythrocyte surface can thus influence the thermal amplitude of cold agglutinins.     

Identification and quantification of juvenile hormones from different developmental stages of the cockroach Nauphoetacinerea

By the combined use of high-pressure liquid chromatography, $\text{Galleria}$ bioassay and gas chromatography/ chemical ionization/mass spectrometry we were able to isolate and identify the three known natural juvenile hormones (JHs) from haemolymph extracts of larval and adult females of the cockroach $\text{Nauphoeta}$$\text{cinerea}$. This is the first demonstration of the simultaneous occurrence of the three JHs in the same insect and the first time JH I and II have been identified in a hemimetabolous insect. Quantitative investigations show that the composition of the three JHs is different at different developmental stages. The haemolymph of larvae contains a high percentage of JH I and II, whereas the haemolymph of adult females in the oocyte maturation stage contains mostly JH III. This suggests more juvenilizing functions for JH I and II and more gonadotropic functions for JH III.     

Size-dependent haemagglutinating activity in the haemolymph from sub-adult blue shrimp (Penaeus stylirostris stimpson)

• 1.1. The blue shrimp (Penaeus stylirostris) haemolymph is capable of agglutinating the red blood cells of several vertebrates to different titres. However, the haemagglutinin is considered non-specific because it is incapable of differentiating erythrocytes of human blood types A, B and O.
• 2.2. Haemagglutinating activity and serum protein content were determined for male and female blue shrimp ranging in size from 8.5 to 16 cm. Haemagglutinating activity decreased significantly with animal size, while protein content was unaffected.
• 3.3. The above finding is probably related to maturation of the immune system and could explain the higher susceptibility of young shrimp to parasitic and viral diseases.

     

A new haemagglutinin from the amoebocytes of the horseshoe crab Carcinoscorpius rotundicauda. Purification and role in cellular aggregation.

The present paper describes the purification and function of a haemagglutinin from the amoebocyte lysate of the horseshoe crab Carcinoscorpius rotundicauda. The purified protein consisted of a single subunit of Mr 24 000 and agglutinated human blood-group-A+ erythrocytes. Its haemagglutinin activity was inhibited by purified lysate, coagulogen, but not by sugars. The haemagglutinin differed immunologically and in activity from the sialic-acid-binding lectin carcinoscorpin present in the haemolymph. It caused aggregation of forma-fixed amoebocytes, and on the basis of this observation its role in cell-cell adhesion is proposed. This new haemagglutinin promotes cell-cell aggregation in amoebocytes in a manner that shares some similarities with thrombospondin-mediated platelet aggregation in vertebrates [Jaffe, Leuang, Nachman, Levin & Moseher (1981) Nature (London) 295, 246-248].     

Concanavalin A-induced suppressor cell activity for T-cell proliferative responses: Autologous and allogeneic suppression in aging humans

Peripheral blood mononuclear cells from 48 healthy subjects of ages varying from 20 to 94 years were evaluated for the ability to generate suppressor cell activity following in vitro incubation with concanavalin A. The suppression of proliferative responses by autologous and young, allogeneic lymphocytes to phytohemagglutinin was assessed using suppressor/ responder cell ratios ($\text{S}\text{R}$) of 2:1 and 1:1 and by using a summation index. Inducible suppressor cell activity for autologous responder cells was comparable between 24 aged (76.0 ± 10.9 years) and 20 young (26.8 ± 4.6 years) subjects. However, aged subjects exhibited a significant decrease in suppressor cell activity ($\text{S}\text{R}\text{= 1}$) when allogeneic responder cells were utilized. Our results indicate that autologous inducible suppressor cell activity is preserved in the aged population, whereas allogeneic activity is impaired.     

The ferritin content of human red blood cells during the replacement of embryonic cells by fetal cells

Mature human embryonic erythrocytes (hemoglobin is ≥ 90% of the cellular protein) contained at least 20 times as much ferritin as human adult erythrocytes, suggesting the possibility that the embryonic red cells participate in iron storage as they do in other embryonic or larval vertebrates. The ferritin content of mature red cells in the circulation declined when fetal red cells replaced embryonic red cells; the cell replacement was monitored by the disappearance of embryonic ε-chains and the appearance of the fetal globin chains, γ^A and γ^G. A constant ratio of 0.67 was obtained for $\text{γ}{}^{\mathrm{<mtext>G</mtext>}}\text{γ}{}^{\mathrm{<mtext>A</mtext>}}\text{+ γ}{}^{\mathrm{<mtext>G</mtext>}}$ from the first detectable appearance (4 weeks after conception) until 13 weeks, a value which is similar to the value previously obtained at 20 weeks gestation and birth but higher than that observable in adults; thus, both γ^G and γ^A chains are produced in similar amounts throughout gestation. The high levels of ferritin in normal human embryonic erythrocytes emphasize the similarity of erythropoiesis in human embryos and other vertebrates. In addition, the results show that red cell ferritin can be used as a marker for studying the mechanism of induction of embryonic erythropoiesis in cultured cell lines, such as K562 from human chronic myelocytic leukemia, and that ferritin content may also serve as a marker for cellular transformations involving reversions to embryonic erythropoiesis.     

Impairment of primary in vitro response of New Zealand mouse spleen cells to a thymic-dependent antigen.

New Zealand Black (NZB) and NZB by New Zealand White (NZW) F₁ hybrid ($\text{B}\text{W}$) mice develop clinical signs of autoimmune disease between 6 and 10 months of age but spleen cells from these strains have a greatly reduced in vitro response to sheep erythrocytes (SRBC) as early as 5–6 weeks of age. This hyporesponsiveness can be only partially restored with 2-mercaptoethanol, allogeneic macrophages or spleen cells, or allogeneic factor. The response of NZB and $\text{B}\text{W}$ spleen cells to the thymic independent antigen DNP-Ficoll is nearly normal. The reduced in vitro SRBC response was found to be attributable to splenic T and B cells rather than macrophages. Macrophages from NZB mice were found to function normally. The in vitro behavior of NZB lymphocytes is very similar to non-autoimmune mice infected with common murine viral pathogens. NZB and $\text{B}\text{W}$ mice may be making an active immune response as early as 5 weeks of age.     

The enzymatic sulphation of heparan sulphate by hen's uterus

A sulphotransferase preparation from hen's uterus catalysed the transfer of sulphate from adenosine 3′-phosphate 5′-sulphatophosphate to N-desulphated heparan sulphate, heparan sulphate, N-desulphated heparin and dermatan sulphate. Heparin, chondroitin sulphate and hyaluronic acid were inactive as substrates for the enzyme. N-desulphated heparin was a much poorer substrate for the enzyme than N-desulphated heparan sulphate suggesting that properties of the substrate other than available glucosaminyl residues influenced enzyme activity. N-acetylation of N-desulphated heparin and N-desulphated heparan sulphate reduced their sulphate acceptor properties so it was unlikely that the N-acetyl groups of heparan sulphate facilitated its sulphatiion. Direct evidence for the transfer of [³⁵S]sulphate to amino groups of N-desulphated haparan sulphate was obtained by subsequent isolation of glucosamine $\text{N-[}{}^{35}\text{S}\text{]}\text{sulphate}$ from heparan [³⁵S]sulphate product. This was made possible through the use of a flavobacterial enzyme preparation which contained “heparitinase” activity but had been essentially freed of sulphatases. Attempts to transfer [³⁵S]sulphate to glucosamine or N-acetylglucosamine were unsuccessfull.     

Macrophages as effector cells of protective immunity in murine schistosomiasis: IV. Coincident induction of macrophage activation for extracellular killing of schistosomula and tumor cells

Peritoneal macrophages from Schistosoma mansoni-infected mice are activated both for nonspecific tumor cytotoxicity and for killing of skin-stage schistosomula in vitro. In the current study, mechanisms for induction of macrophage tumoricidal and schistosomulacidal activity have been compared. Examination of macrophages activated in vivo by BCG infection or C. parvum treatment, or in vitro by exposure to lymphokine prepared from antigen-stimulated BCG-immune spleen cells, showed that these effector functions were closely linked. Indeed, fractionation of lymphokine-rich supernatant fluids by Sephadex G-100 gel filtraction showed that activities responsible for induction of schistomula killing by inflammatory macrophages and for induction of tumoricidal activity cochromatographed as a single peak in the 50,000 MW region. Thus, development of macrophage-mediated cytotoxicity against these two extracellular (tumor cell or helminth) targets was coincident in several cell populations activated in vivo or in vitro. However, activation for tumoricidal and schistosomulacidal capacity appeared to be quantitatively dissociated in macrophages from mice with chronic schistosomiasis; those cells demonstrated low, yet significant, levels of larval killing ($\text{1}\text{3}$ to $\text{1}\text{5}$ those of BCG or lymphokine-activated cells) but maximal levels of tumor cell cytotoxicity. Furthermore, cytotoxicity by peritoneal cells from S. mansoni-infected mice was not increased in vitro by exposure to lymphokine. Identification of this functional alteration in S. mansoni-activated cells may help to clarify the role of macrophages in the partial immunity against challenge infection which is demonstrated by mice with chronic primary S. mansoni infection.     

Fatty acid hydroperoxide lyase in tobacco cells cultured in vitro

Fatty acid hydroperoxide lyase (HPO lyase) was found in green and non-green tobacco cells cultured in vitro. The HPO lyase activity in non-green cells was $\text{1}\text{3}$-$\text{1}\text{2}$ of that in green cells. When the cells were transferred from the light to dark conditions or vice versa, cells turned non-green or green according to the light conditions. The HPO lyase activity also changed according to the light conditions, but the changes in HPO lyase activities were not proportional to the changes in chlorophyll contents. These results suggest that at least two types of HPO lyases are present in the green cells. One type of HPO lyase is perhaps common both to the green and non-green cells; another one is chloroplastic. The fatty acid compositions of cells and substrate specificities of HPO lyase differed between green and non-green cells.     

Rheological characteristics of desialylated erythrocytes in relation to fibrinogen-induced aggregation

The effect of fibrinogen and sialic acid content of erythrocytes on the aggregation of erythrocytes was quantitatively examined by using a rheoscope combined with a television image analyzer and a computer. (1) The electrophoretic mobility of erythrocytes was proportional to the sialic acid content of erythrocytes (the surface potential of erythrocytes could be expressed by the sialic acid content). (2) The aggregation of erythrocytes was accelerated by increasing fibrinogen concentration in the medium (due to the increased bridging force among erythrocytes) or by decreasing the sialic acid content (due to the reduction of the electrostatic repulsive force among erythrocytes). (3) An empirical equation expressing the velocity of aggregate formation (ν, in μm²/min) by the concentration of fibrinogen ($\text{F}$, in g/dl) and the sialic acid content ($\text{S}$, in μmol/ml red blood cells), log $\text{ν = ?0.065 F}{}^{\mathrm{?1.2}}\text{S + 2.2 F}{}^{0.35}$, was deduced. (4) The contribution of the bridging force of fibrinogen to the erythrocyte aggregation was much greater than that of the electrostatic repulsive force produced by sialic acid on the surface of erythrocytes.     

A study of the diffusion of compact particles in polymer solutions using quasi-elastic light scattering

This paper describes an investigation, using quasi-elastic light scattering, of the diffusion of polystyrene spheres through solutions of dextran. The diffusion coefficient, D, of the spheres is shown to vary inversely with the volume fraction, φ, of dextran according to $\text{D =}\text{D}{}_0\text{(1 + νφ + κφ}{}^2\text{)}$. Changes in the molecular weight of dextran are shown to reflect changes in the macromolecular shape parameter, ν, and the interaction parameter, κ. This result differs from previous studies which suggested an $\text{exp}\text{(?}\text{B}\text{φ}{}^{\mathrm{<mtext>1</mtext><mtext>2</mtext>}}\text{)}$ dependence and no molecular weight dependence [8].     

Studies on NADH-cytochrome b5 reductase activities in hemolysates of human and rabbit red cells by isoelectric focusing

NADH-cytochrome $\text{b}{}_5$ reductase activities in hemolysates of young and old human erythrocytes, and in hemolysates of rabbit reticulocytes and erythrocytes were measured after the separation of the enzyme from the bulk of hemoglobin only by isoelectric focusing. In any cases, a single main peak of the enzyme activity was detected after the electrophoresis in the fraction with pH 6.8 and 8.3 for human and rabbit red cells, respectively. The rabbit enzyme showed more than 30 times higher enzyme activity than that of human erythrocytes under the standard assay conditions. Significant differences of Micahelis constants for cytochrome $\text{b}{}_5$ of the enzyme were found between young and old human erythrocytes, and also between human and rabbit red cells.     

Changes in nucleotide concentrations in the erythrocytes of man, rabbit and rat during short-term storage

The $\text{ATP}\text{ADP}$ ratio, measured by high performance liquid chromatography, has been used as an indicator of stability of erythrocyte nucleotides. The nucleotides from human, rabbit and rat whole blood, but not separated erythrocytes were stable for maximum periods of 40, 20 and 15 min respectively after venepuncture. The ratios then declined rapidly from 9 to 5, 12 to 4 and 9 to 1 respectively during 2h storage at room temperature. Similar changes occurred in $\text{GTP}\text{GDP}$ ratios. The relevance of these observations to metabolic studies in intact cells, nucleotide analyses in the clinical situation and comparative studies in other species is discussed.     

Immunoregulation in MRL/Mp-lpr/lpr mice: evidence for decreased helper-T-cell and increased suppressor-T-cell function with age

In vivo and in vitro plaque-forming cell (PFC) responsiveness to sheep erythrocytes (SRBC) was used to assess immunoregulatory function in the autoimmune MRL mouse strain. MRL/Mp-lpr/lpr (MRL/l) mice had good primary and secondary IgM and IgG responses in vivo compared to MRL/Mp-+/+ (MRL/n) mice when young, but with age the MRL/l responses declined markedly. In vitro primary SRBC-specific PFC responses in MRL/l mice declined at the same time as in vivo responses, indicating that the in vivo autoimmune environment could not account for cellular dysfunction. When varied mixtures of T and B cells from MRL/l and MRL/n mice were cultured, abnormalities in MRL/l T-cell function became apparent. T-helper-cell (T_H) function declined rapidly with age, beginning by 2 to $\text{2}\text{1}\text{2}$ months of age. T cells from MRL/l mice 2 months of age and older also had increased suppressor activity when cultured with B cells and MRL/n T cells. The degree of suppressor activity increased with age. The correlation of these findings with results of previous studies by others and with autoimmune disease is discussed.