Increased ALA dehydratase activity and spleen weight in lead-intoxicated rats. A consequence of increased blood cell destruction

Lead was given in the diet (1%) to rats from birth and at different times the animals were studies for delta amino levulinic acid dehydratase (ALAD) activity, spleen weight, 59Fe incorporation in erythrocytes and 51Cr-labeled erythrocytes survival. The increased ALAD and spleen weight found after lead treatment is explained as a consequence of a shortened survival, which results in a younger age of circulating erythrocytes with higher ALAD activity.     

Tubulin pools in human erythrocytes: altered distribution in hypertensive patients affects Na<Superscript>+</Superscript>, K<Superscript>+</Superscript>-ATPase activity

The presence of tubulin in human erythrocytes was demonstrated using five different antibodies. Tubulin was distributed among three operationally distinguishable pools: membrane, sedimentable structure and soluble fraction. It is known that in erythrocytes from hypertensive subjects (HS), the Na⁺, K⁺-ATPase (NKA) activity is partially inhibited as compared with erythrocytes from normal subjects (NS). In erythrocytes from HS the membrane tubulin pool is increased by ~150%. NKA was found to be forming a complex with acetylated tubulin that results in inhibition of enzymes. This complex was also increased in erythrocytes from HS. Treatment of erythrocytes from HS with nocodazol caused a decrease of acetylated tubulin in the membrane and stimulation of NKA activity, whereas taxol treatment on erythrocytes from NS had the opposite effect. These results suggest that, in erythrocytes from HS, tubulin was translocated to the membrane, where it associated with NKA with the consequent enzyme inhibition.     

Direct inhibition of phospholipid scrambling activity in erythrocytes by potassium ions

The exposure of phosphatidylserine (PS) at the cell surface plays a critical role in blood coagulation and serves as a macrophage recognition moiety for the engulfment of apoptotic cells. Previous observations have shown that a high extracellular [K⁺] and selective K⁺ channel blockers inhibit PS exposure in platelets and erythrocytes. Here we show that the rate of PS exposure in erythrocytes decreases by ~50% when the intracellular [K⁺] increases from 0 to physiological concentrations. Using resealed erythrocyte membranes, we further show that lipid scrambling is inducible by raising the intracellular [Ca²⁺] and that K⁺ ions have a direct inhibitory effect on this process. Lipid scrambling in resealed ghosts occurs in the absence of cell shrinkage and microvesicle formation, processes that are generally attributed to Ca²⁺-induced lipid scrambling in intact erythrocytes. Thus, opening of Ca²⁺-sensitive K⁺ channels causes loss of intracellular K⁺ that results in reduced intrinsic inhibitory effect of these ions on scramblase activity. Received 11 September 2008; received after revision 17 October 2008; accepted 27 October 2008     

Evidence for suppression of immune function by insulin-like growth factor-1 in dwarf rats in vivo

These studies were undertaken to investigate the effects of increasing or decreasing IGF-1 levels on aspects of immune function in rats. Female dwarf rats were treated with recombinant human IGF-1 or with a potent sheep anti-IGF-serum. Body weight, thymus weight and spleen weight increased with IGF-1 treatment (p<0.001), while there was no effect of anti-IGF-1 treatment when compared with the appropriate normal sheep serum (NSS) treated controls. IGF-1 treatment significantly decreased WBC and RBC counts, but increased the ratio of CD4⁺:CD8⁺ T-cells. Anti-IGF-1 serum had no effect on these parameters compared with NSS. However anti-IGF-1 was associated with increased T-cell numbers, decreased natural killer cells, and enhancement of the animals' ability to produce specific IgG in response to injection of keyhole limpet haemocyanin (KLH). These results indicate that IGF-1 may suppress immune function although increasing the size of immune organs such as spleen.These studies were part of an M.Sc. at the University of Waikato, Hamilton, New Zealand.     

Incorporation of14C-thymidine by cultures of erythrocytes from rheumatoid arthritis patients and normal subjects,suggesting the presence of an L-form

Summary Cultures of washed erythrocytes from rheumatoid arthritis patients and normal subjects were found to incorporate¹⁴C-thymidine,suggesting the presence of an L-form. The extent of the incorporation by erythrocytes from rheumatoid patients was more variable than that by erythrocytes from normal subjects, and correlated negatively with IgG measurements and white cell counts performed on the patients' bloods, although not with the clinical activity of the patients.     

Growth hormone alters lymphocyte sub-populations and antibody production in dwarf rats in vivo

Female dwarf rats at different ages were treated with recombinant porcine GH or with a potent sheep anti-rat GH serum. Body weight and spleen weight increased with GH and decreased with anti-GH treatment (p<0.001). Neither GH nor anti-GH treatment resulted in a change in circulating WBCs, but GH decreased, while anti-GH increased, RBC counts (p<0.001). Similarly, GH treatment tended to decrease the ratio of CD4⁺:CD8⁺ T-cells while anti-GH increased (p<0.05) the ratio. Anti-GH treatment also enhanced the animals' ability to produce specific IgG in response to KLH injection. These results indicate that GH may have a physiological role in suppressing humoral immune function but may enhance cell-mediated immunity.     

Purification and some properties of hemagglutinin from the Myxomycete,Physarum polycephalum

A new hemagglutinin was isolated from the plasmodium ofPhysarum polycephalum by salting out with ammonium sulphate followed by chromatography on DE-32, DEAE-Toyopearl and hydroxyapatite. This hemagglutinin, named physarumin, was purified 1000-fold over crude extracts. The molecular weight of physarumin was determined to be 22,000 by size exclusion chromatography on Bio-Gel P-60 and 8,700 by SDS-polyacrylamide gel electrophoresis. Physarumin agglutinated rabbit, guinea pig, horse and human erythrocytes. Physarumin-induced hemagglutination was inhibited by fetuin and ₁ glycoprotein, but not by commercially available mono-and disaccharides. Hemagglutinating activity was blocked by EDTA, and was restored by adding Ca²⁺ but not by Mg²⁺.     

Agglutination of leukemic cells and daunomycin entrapped erythrocytes with lectin in vitro and in vivo

Summary Wheat germ agglutinin (WGA) agglutinated the L₁₂₁₀ leukemic cells and daunomycin entrapped erythrocytes in vitro. Comparing with control preparations, the greatest increase in survival was obtained in vivo when the daunomycin entrapped erythrocytes and WGA were given to BDF₁ mice bearing L₁₂₁₀ cells.     

An alternative decomplementation method for the measurement of the survival of human erythrocytes transfused into rats

Summary An alternative technique for measuring the survival of⁵¹Cr-labelled human erythrocytes transfused into rats is described, in which aggregated human gamma-globulin is substituted for cobra venom factor as a decomplementing agent.Acknowledgments. This work was supported by Fundação de Amparo à Pesquisa do Estado de São Paulo and Conselho Nacional de Desenvolvimento Cientifico e Tecnológico.     

Inhibition of glucose transport in human erythrocytes by 2,3-dioxoindole (Isatin)

10 mM isatin (2,3-dioxoindole) inhibited glucose influx into human erythrocytes by over 30%. The inhibition is of the competitive type, where the affinity constant (K_t) was increased from 5.71 (control) to 11.11 mM in the presence of isatin with no change in V_max (130 nmol/min/ml packed cells). The observed inhibition of sugar transport by isatin was not mediated through membrane–SH groups accessible to iodoacetate, iodoacetamide, DTNB, DNP or sodium arsenite. Isatin inhibited sugar transport in the presence of 2 mM harmaline, an alkaloid inhibitor of Na⁺, K⁺–ATP_ase activity. The inhibition was non additive which suggests that these two compounds interact with the same or a similar site on the erythrocyte membrane.     

Effect of 15(R)-15-methyl-prostaglandin E2 on iron absorption in the rat

Summary The administration of 15(R)-15-methyl prostaglandin E₂ (15(R)-15-M-PGE₂) in vivo significantly diminished the uptake of⁵⁹Fe into blood, spleen, liver, femur and dried intestine of rats, whereas acetylsalicylic acid (ASA) increased the counts significantly. This effect of ASA was counteracted by 15(R)-15-M-PGE₂. It is suggested that prostaglandins (PGs) might play an important role in inhibiting iron absorption at the intestinal level.This work was supported by grant No.6638 from CONICET (Argentina). The technical assistance of Mrs María E. Castro and Norma Rizzo is gratefully acknowleged.     

Possible role of intestinal alkaline phosphatase activity in thiamine transport

Summary Thiamine deficiency caused a marked decrease of intestinal alkaline phosphatase (al-Pase) activity, but had no effect on the Ca⁺⁺-ATPase activity and Ca⁺⁺-absorption in rats. The al-Pase activity was significantly decreased 1 h after oral administration of ethanol at 0.5 and 2.5 g/kg. In contrast, Mg⁺⁺-, Ca⁺⁺- and (Na⁺+K⁺)-ATPase activities did not change after the administration of ethanol. These findings show that the al-Pase activity, unlike the Ca⁺⁺-ATPase activity, is not related to Ca⁺⁺-absorption. A possible role of al-Pase activity in the active transport of thiamine in the intestine was discussed.     

Induction of lymphokine-activated killer cells from nude mouse spleen cells by interleukin-4

The induction of lymphokine-activated killer (LAK) cells from natural killer (NK) lineage cells by interleukin-4 (IL-4) was studied in vitro. Activation of nude mouse spleen cells by IL-4 generated cytotoxic cells, capable of killing NK-sensitive as well as NK-resistant tumor cells. The induction of peak lytic activity was demonstrated after 3 days of culture with IL-4. Surface marker analysis indicated that the majority of precursor cells were aGM1⁺, Thy1^–, and the majority of effector cells were aGM1⁺, Thy1⁺, suggesting that IL-4 induced LAK cells from nude mouse spleen cells were similar to those from normal mouse spleen cells. The induction of nude mouse LAK cells by IL-4 was partially inhibited by anti-IL-4 or anti-interferon (IFN)-, antibody, and it was further inhibited by the combination of two antibodies, suggesting that IFN-, production was associated with LAK induction of NK lineage cells by IL-4.     

Röntgenbestrahlung und Hypothermie. Der Einfluss auf Körpergewicht,Hirngewicht, Kleinhirnentwicklung und Blutbild bei neugeborenen Hausmäusen

Summary New-born mice were X-rayed under extreme hypothermia in order to estimate the radioprotective effect of hypothermia on body weight, brain weight, development of cerebellum and the hematopoietic system. A protective effect was found in the rate of survival, in body and brain weight, in the cerebellum and in the number of erythrocytes. After hypothermia, the repair of damage in the cerebellum and the erythropoietic system starts earlier. The protected mature cerebellum shows an approximately normal status. Hypothermia does not prevent the initial decrease of lymphocytes caused by X-rays, and it produces a severe disturbance of hemoglobin content.     

Aging of the erythrocyte. III. Cation content

Summary Studies on the main cation content of density-separated bovine erythrocytes showed a progressive decrease in the levels of K⁺ and Mg²⁺ with increasing cell density (and age) accompanied by an increase in the level of Na⁺. The magnitude of net cation loss corresponded to that of red cell volume decrease, but could not account for the total increase in the microviscosity of the erythrocyte interior.     

Mitogenicity of autolysates ofTrypanosoma congolense

Summary Autolysates ofTrypanosoma congolense, in subcytotoxic amounts, were found to be highly mitogenic in vitro for the spleen cells of normal mice. Significant amounts of [³H]-thymidine were also incorporated by the responding spleen cells of nu/nu (athymic) mice. In contrast, the spleen cells of cyclophosphamide-treated mice were unresponsive. The findings suggest that a potent B-cell-mitogen is generated by the autolysingT. congolense organism.Acknowledgments. This investigation was supported by the International Development Research Centre, Canada.     

Lack of Na+,K+-ATPase expression in intercalated cells may be compensated by Na+-ATPase: A study on MDCK – C11 cells

The lack of Na⁺,K⁺-ATPase expression in intercalated cells (IC) is an intriguing condition due to its fundamental role in cellular homeostasis. In order to better understand this question we compared the activities of Na⁺,K⁺-ATPase and Na⁺-ATPase in two MDCK cell clones: the C11, with IC characteristics, and the C7, with principal cells (PC) characteristics. The Na⁺,K⁺-ATPase activity found in C11 cells is far lower than in C7 cells and the expression of its β-subunit is similar in both cells. On the other hand, a subset of C11 without α-subunit expression has been found. In C11 cells the Na⁺-ATPase activity is higher than that of the Na⁺,K⁺-ATPase, and it is increased by medium alkalinization, suggesting that it could account for the cellular Na⁺-homeostasis. Although further studies are necessary for a better understanding of these findings, the presence of Na⁺-ATPase may explain the adequate survival of cells that lack Na⁺,K⁺-ATPase. Received 09 July 2008; received after revision 03 August 2008; accepted 12 August 2008     

Liver homing in the rat of111Indium labelled beef and neuraminidase-treated rat erythrocytes (Studies with a Gamma Camera)

Summary The rate of liver homing of¹¹¹In-labelled erythrocytes has been measured under a Gamma Camera. Homing of neuraminidase-treated or xenogeneic erythrocytes is delayed by preinjection of glycolipids or glycopeptides.We thank Miss Unverhau for excellent technical assistance.     

Semisynthesis and biological properties of the [B24-leucine]-, [B25-leucine]- and [B24-leucine,B25-leucine]-analogues of human insulin

Summary Trypsin-catalyzed coupling of porcine desoctapeptide-insulin with synthetic octapeptides produced the [Leu^B24]-(I), [Leu^B25]- (II) and [Leu^B24, Leu^B25]- (III)analogues of human insulin. I, II and III displayed respectively 20–30%, 1–2% and 0.5% of the receptor binding activity of the normal hormone. Biological activities of these analogues seemed to be proportional to their binding potencies when assayed in vitro, while in an in vivo assay analogue I was fully active and II exhibited 10–20% of normal activity. III was less active than II in all assays tested.