Immunologic properties of mouse thymus cells: membrane antigen patterns associated with various cell subpopulations

The membrane antigen components of mouse thymus cells and fractions derived from BSA density gradient centrifugation were assayed by quantitative cytotoxicity tests. Two subpopulations were identified on the basis of average density and antigen patterns. The major subpopulation consisted of small lymphoid cells and comprised 80%–90% of all cells, was of high relative density and rich in θ, TL, G_IX, Ly-A, Ly-B, and Ly-C, but contained little or no H-2. The minor subpopulation was chiefly large lymphoid cells, comprised 10%–15% of cells, was of low relative density, was relatively rich in H-2 but low in θ and Ly antigens, and contained no detectable TL or G_IX. This minor subpopulation was identical in density and antigen patterns to those cells remaining in the thymus after short-term cortisone treatment or whole-body irradiation. It could also be reproduced by treating whole thymus with anti-TL or anti-θ sera. The antigenic attributes of this minor subpopulation differed from those of spleen lymphocytes only with respect to average density.     

Albumin synthesis during induced and spontaneous metamorphosis in the bullfrog Rana catesbeiana

Treatment of premetamorphic tadpoles with triiodothyronine (T₃) alters the in vivo distribution of radioactive amino acids among serum protein fractions. The effects on the albumin fraction have been interpreted as reflections of the relative rate of synthesis. About 12 hr after intraperitoneal injection of 2.5 × 10^?10 mole of T₃ per gram, there is an increase in the relative rate of albumin synthesis. The effect peaks on day 3 at 5 × the untreated level and returns to near the untreated level by day 6. Continuous immersion in 1 × 10^?7M T₃ results in a similar stimulation of albumin synthesis, but with no decline after day 3. The timing of the response is independent of dose or route of T₃ administration. The effect of T₃ on the relative rate of albumin synthesis is also observed in froglets. There is a 6-fold increase in the relative rate of albumin synthesis during spontaneous metamorphosis peaking at stage XXI and returning to the premetamorphic level by stage XXV. The following was concluded: (1) The increase in the relative rate of albumin synthesis during metamorphosis results from increased endogenous thyroid levels. (2) Following a peak, the decline in albumin synthesis observed in induced and spontaneously metamorphosing animals is a result of decreasing thyroid hormone levels. (3) The effect of T₃ on albumin synthesis may be the summation of two effects, a direct effect of T₃ and a stimulation by amino acids from the resorbing tail. (4) A decreased relative rate of albumin degradation or a sparing of albumin is probably responsible for the elevated relative concentration of albumin in the serum of postmetamorphic animals.     

A double-blind evaluation of transfer factor therapy of HBsAg-positive chronic aggressive hepatitis: preliminary report of efficacy.

A controlled, prospective, double-blind trial of transfer factor in chronic aggressive hepatitis was carried out. This report presents the results obtained from study of the nine HB_sAg-positive subjects who were included in the trial. Transfer factor was prepared from adults who had recovered from acute hepatitis B or from acute non-B viral hepatitis and was administered to five HB_sAg-positive subjects. Four HB_sAg-seropositive subjects received placebo. Percutaneous liver biopsy was performed at the beginning and at the conclusion of the 10-week study period; biochemical and clinical parameters were monitored throughout. Cell-mediated immunity was assessed at the beginning and at the end of the study period. Four of five transfer factor recipients showed moderate or marked improvement in hepatic histology, clinical status, and serum transaminase levels, while none of four placebo recipients demonstrated improvement. The difference in response between the two groups was significant (P = 0.048). No consistent changes in in vivo or in vitro correlates of cell-mediated immunity were demonstrated. No adverse effects were noted among transfer factor recipients. These data suggest that transfer factor may be of benefit in chronic aggressive hepatitis associated with HB_sAg. Additional studies appear to be indicated to delineate the mode of action of transfer factor as well as the role that such immunotherapy should play in the management of patients with this disorder.     

Uptake of the yolk protein, lipovitellin, by developing crustacean oocytes

A variety of cytochemical techniques were used to demonstrate how crustacean lipovitellin accumulates within the egg. It was found that a protein serologically identical to the lipovitellin of yolk spheres was present in the hemolymph of vitellogenic crustaceans, but was absent from the hemolymph of males and immature females.In the three crustacean species studied (Uca pugilator, Cambarus clarkii, and Libinia emarginata), pinocytosis of fluorescein-conjugated lipovitellin and trypan blue occurred only during those periods when oocytes were accumulating yolk.It may be concluded from the present studies that yolk spheres develop in crustacean eggs primarily through micropinocytotic uptake of lipovitellin from the hemolymph, although other oocyte proteins appear to be made in the oocyte.     

Prostaglandin E inhibition of mitogen stimulation in patients with multiple sclerosis

Kirbey et al have reported that leukocyte function from patients with multiple sclerosis is not suppressed by PGE₂, as are normal leukocytes. We examined the ability of PGE₂ (0.01–0.5 μg/ml) to suppress Phytohemagglutinin induced ³H-thymidine incorporation in peripheral blood lymphocytes from multiple sclerosis patients and normals. There was no difference in sensitivity between the two groups. There was also no difference in activity of the prostaglandin producing suppressor cell between the multiple sclerosis patients and controls.     

Magnetic resonance studies of the mitochondrial divalent cation carrier

Measurements of water proton spin relaxation enhancements (ε) can be used to discriminate high-affinity binding of Mn²⁺ or Gd³⁺ to biological membranes, from low-affinity binding. In rat liver mitochondria, ε_b values of approx. 11 are observed upon binding of Mn²⁺ to the inner membrane, while internal or low-affinity binding remains invisible to this technique. Energy-driven Mn²⁺ uptake by liver mitochondria results in the subsequent decay of ε¹.Comparison of ε¹ with the initial velocity of Mn²⁺ uptake in rat liver mitochondria reveals a linear correlation, which holds at all temperatures between 0 °C and 40 °C, regardless of the mitochondrial protein concentration. Consequently, enhancement appears to reflect the binding of Mn²⁺ to the divalent cation pump.Binding of Mn²⁺ to blowfly flight muscle also results in substantial ε¹, which is associated with the glycerol-1-phosphate dehydrogenase instead of divalent cation transport. Consequently, no decay in ε¹ due to uptake occurs after Mn²⁺ is bound.Lanthanide ions are also bound and transported by mitochondria. Addition of Gd³⁺ to pigeon heart or rat liver mitochondria results in ε_b ≈ 5–6, which decays with similar kinetics in both systems. The uptake velocity of Gd³⁺ in rat liver mitochondria is about $\text{1}\text{6}$ the rate with which Mn²⁺ is transported. Lanthanides also diminish ε¹ due to the addition of Mn²⁺, and greatly retard the Mn²⁺ uptake kinetics. The presence of carbonylcyanide-p-trifluoromethoxyphenylhydrazone depresses ε¹ upon addition of Mn²⁺ or Gd³⁺ and also uncouples energy-driven uptake. On the other hand, prolonged anaerobic incubation in the presence of antimycin and rotenone exhausts the mitochondria of their energy stores, blocks the uptake of Mn²⁺, but does not affect ε¹ significantly. Evidently, the uncoupler-induced disappearance of divalent cation binding sites is not the result of “de-energization”.Measurements of ε¹ at several NMR frequencies indicate a correlation time (τ_b) for carrier-bound Mn²⁺ in rat liver mitochondria between 20 ns and 4 ns as one varies the temperature between 10 °C and 30 °C. The 13 Kcal/mole activation energy for τ_b suggests that the 11 ns time constant at room temperature represents the movement of the Mn^II-carrier complex. On the other hand, τ_b is probably approx. 100 times too short to represent the rotational motion of a carrier protein. Apparently, Mn²⁺ binds to a small arm of the carrier which moves independently of the main body of any protein.In addition to Mn(H₂O)₆²⁺, other complexes of Mn²⁺ may also be bound and transported by rat liver mitochondria. Only a small increase in ε¹ occurs upon addition of MnHPO₄, yet this species is accumulated by the mitochondria. Consequently, the carrier does not recognize divalent metal ions on the basis of charge.     

Male copulatory behavior and the induction of ovulation in female voles: a quest for species specificity.

Two experiments were conducted to investigate species specificity in the neuroendocrine responsiveness of female prairie voles to the copulatory patterns of males. In Experiment 1, prairie vole males mated for one ejaculatory series were not significantly more effective in inducing ovulation in prairie vole females than montane voles mated with prairie vole females for one series, two series, or to satiety. Mating with conspecific males did result in significantly more implanted embryos than did heterospecific matings. In Experiment 2, it was found that, when the amount of vaginal stimulation was both low and equated across groups, prairie vole males were significantly more effective in triggering ovulation in female prairie voles than were either meadow voles or montane voles. Although there appears to be some species specificity to the “vaginal codes” of these congeneric species, its biological significance is unclear.     

Ovulation and implantation in montane voles (Microtus montanus) as a function of varying amounts of copulatory stimulation

Estrous female voles (Microtus montanus) received (a) no copulation, (b) one ejaculatory series, (c) two ejaculatory series, or (d) more than two series. Copulation was shown to be essential for ovulation, and increasing amounts of copulatory stimulation were demonstrated to increase the probability of ovulation. Only 25% of females receiving one ejaculatory series ovulated, whereas 100% of those receiving more than two series ovulated. Thus, multiple ejaculations play an important role in successful reproduction in this species.     

Germination of Schizosaccharomyces pombe spores separated by zonal centrifugation

Spores from the fission yeast Schizosaccharomyces pombe (H90 strain) were separated from residual vegetative cells into distinct size classes by zonal density centrifugation. Spores were sized photographically and with a Coulter counter. The kinetics of germination were followed by time-lapse photomicrography. The duration of the pre-germination interval was size dependent. Large spores (˜50 μm³) germinated as early as 5 h after resuspension in nutrient media, smaller ones (˜20 μm³) did so at 11 h. The first cell division occurred 4–5 h later regardless of spore size. The large spores divided more synchronously as shown by the occurrence of peaks in the cell plate index at approximately one doubling time intervals.     

A case report of the successful treatment of recurrent aphthous stomatitis with some preparations of orally administered transfer factor

A patient with severe disabling recurrent aphthous stomatitis (RAS) was treated with four different preparations of oral human transfer factor (TF), as well as placebo, following a double-blind protocol. Two of the TF preparations had a significant effect upon the course of the patient's illness by prolonging the interval between attacks and decreasing the severity of attacks. No side effects attributable to any of the preparations were noted by the patient. Thus, some but not all preparations of human transfer factor given orally are an effective therapy for RAS.     

Methods for microcarrier culture of bovine pulmonary artery endothelial cells avoiding the use of enzymes

Enzymes situated along the luminal surface of pulmonary endothelial cells interact with circulating solutes, notably with vasoactive substances, to regulate the hormonal composition of systemic arterial blood. However, it is becoming clear that the range and complexity of reactions occurring at or near the surface of endothelial cells are greater than previously recognized. In addition, evidence indicates that the quality of cell cultures used to define specific endothelial functions must be carefully controlled, together with development of improved understanding of the effects of long-term culture on pulmonary endothelial cells. We have developed new techniques for the culture of pulmonary endothelial cells which avoid exposure to proteolytic enzymes at both the isolation step and during subculture. A combination of mechanical harvest and culture on microcarrier beads has provided a system for the long-term, large-scale culture of pulmonary endothelial cells, features which to a large extent determine the scope of biochemical studies which can be undertaken.     

Synthesis of LDH-1 during mamalian oogenesis and early development.

Nuclear multiplication stage embryos were punctured in either the anterior, midlateral, or posterior regions. Both embryos and adults were examined for defects, and the defects were correlated with whether there had been any leakage of cytoplasmic material from the egg at the time of puncturing. Embryonic defects were found, correlated to the site of damage, in all three regions. A number of embryos was followed through development and it was found that 15.1% of the embryos which leaked cytoplasm hatched into larvae, compared to 82.3% of those which did not leak any cytoplasm. Morphological defects arising as a result of lateral puncture only were observed in adults. Many sterile adults were obtained from eggs in which the posterior region had been punctured. The results show that nuclear multiplication embryos are well able to tolerate the disturbance of the cortical cytoplasm created by puncture, but only rarely are they able to compensate for the actual loss of material by regulation. The results were similar to those observed after puncturing Drosophila embryos at the cellular blastoderm stage.     

Measurement of serum ferritin by a 2-site immunoradiometric assay

A 2-site immunoradiometric assay (2-site IRMA) for human serum ferritin is carried out by reaction of the ferritin solution with a solid-phase anti(human ferritin), followed by a second reaction in which the insoluble product is incubated with purified, radioactively labeled anti(human ferritin). Unreacted labeled antibodies remain in solution and are washed away. As the amount of ferritin increases, the radioactivity in the solid-phase increases. Factors affecting the assay were evaluated including (a) concentration and stability of solid-phase antibody, (b) variation in temperature, reaction times, and reagent concentrations, (c) stability and storage of antibody coated tubes, (d) effect of tube washing cycles, (e) effect of serum proteins and anticoagulants, (f) organ specificity of ferritin.A paradoxical fall in dose-response was seen at high dose. Statistical analysis, dose interpolation, and automatic data processing were carried out by a generalization of the logit/log method and computer programs used in conventional radioimmunoassay. The dependence of the variance on the position on the dose-response curve is different than that seen in radioimmunoassay systems and is reflected in a greater effective assay range. 2-Site IRMA may also have advantages in reagent stability, specificity, antigen protection, and suitability for automation. The properties of 2-site IRMA are closely related to the usual IRMA assay system, but 2-site IRMA is more economical in antigen, is unlikely to be subject to deleterious allosteric reactions, and has a lower zero dose-response (0.5–2% of the total radioactivity in the system).     

The effects of desiccation and rehydration on the lone star tick

This study describes the effects of desiccation and rehydration on the water content, haemolymph volume (per cent), osmolarity, and concentrations of Na, K, Mg, and Ca in the haemolymph of the lone star tick, Amblyomma americanum.The water content percentages of ‘severely desiccated’, ‘moderately’ and ‘fully hydrated’ ticks were 46·0, 52·8, and 60·3 per cent respectively. The lowest and highest of these were near the minimum and maximum possible.The haemolymph volume (per cent) of ‘severely desiccated’ ticks was regulated near the level of ‘moderately hydrated’ ticks despite significant decreases in total body water content and increases in osmolarity and concentration of sodium. Conversely, the change from ‘severely desiccated’ to ‘moderately hydrated’ ticks can be viewed as causing an increase in total body water, decrease in blood osmolarity and sodium, but little change in haemolymph volume (per cent).Most of the water taken up by ‘moderately hydrated’ ticks (while becoming ‘fully hydrated’) was added to the haemolymph. At the same time, there was little change in the blood osmolarity or haemolymph concentration of sodium. Conversely, the change of ‘fully’ to ‘moderately hydrated’ ticks was marked by a substantial loss of haemolymph volume (per cent) but little change in osmolarity and concentration of sodium.The concentration of potassium was regulated over the full range of desiccating and hydrating conditions. The lone star tick appeared less able to regulate its haemolymph concentrations of Ca and Mg; both fluctuated at the same rate, but inversely as the haemolymph volume (per cent).It appears that a carefully controlled movement of solutes (Na the predominant cation) between haemolymph and non-haemolymph tissue is intimately linked with haemolymph volume regulation and movement of water into the haemolymph during hydration.     

Activation of protein kinase in the guinea pig fundic gastric mucosa by histamine.

The effect of histamine, 1,4-methylhistamine and ethanol on cyclic AMP levels and protein kinase activation was measured in tissue strips from the fundic region of guinea pig gastric mucosa. Histamine induced a significant elevation of tissue cyclic AMP levels and also $\text{in situ}$ activation of the protein kinase. 1,4-methylhistamine, an inactive analog of histamine, and ethanol had no effect on these two parameters. Results suggest that protein kinase activation is involved in the cyclic AMP-mediated action of histamine on the gastric fundic mucosa.     

A rapid technique to evaluate two high-affinity binding proteins in the same preparation

A modified DEAE-cellulose filtration assay was developed to estimate the capacities of two types of high-affinity progesterone-binding sites. The method was tested using a mixture containing rat corticosteroid-binding globulin and progesterone-binding protein from pregnant guinea pig plasma and by simulation. The assay has proved useful in studying the progesterone-binding proteins in uterine cytosol of the rat and other species and in following the separation of the two types of binding sites in rat uterine cytosol by ammonium sulfate fractionation or differential hydroxylapatite adsorption.