Background: Knowledge of differences in the cellular physiology of malignant and non-malignant cells is a prerequisite for the development of cancer treatments that effectively kill cancer without damaging normal cells. Calcium is a ubiquitous signal molecule that is involved in the control of proliferation and apoptosis. We aimed to investigate if the endoplasmic reticulum (ER) Ca 2+ -homeostasis is different in lung cancer and normal human bronchial epithelial (NHBE) cells.

Background: The interleukins IL-4 and IL-13 play a key role in the pathophysiology of asthma. The interleukin receptor IL-13Rα2 is believed to act as a decoy receptor, but until now, the functional significance of IL-13Rα2 remains vague. Methods: Bronchial reactivity was quantified in murine lung slices by digital video microscopy and acetylcholine (ACH)-induced Ca²⁺ signaling was measured in human airway smooth muscle cells (ASMC) using fluorescence microscopy. Results: IL-4 or IL-13 up to 50 ng/ml induced bronchial hyperreactivity. But after incubation with 100 ng/ml this effect was lost and bronchial responsiveness was again comparable to the control level. The effects of IL-4 and IL-13 on bronchial reactivity were paralleled by the effects on ASMC proliferation. Fifty nanograms per milliliter of IL-4 and IL-13 increased the Ca²⁺ response of human ASMC to ACH. At 100 ng/ml, however, the effects of the cytokines on the Ca²⁺ response were no longer evident. The expression of IL-13Rα2 increased with increasing concentrations of IL-4 or IL-13, reaching its maximum at 100 ng/ml. Blocking IL-13Rα2, the loss of the effect of IL-4 and IL-13 at 100 ng/ml on human ASMC proliferation and the ACH-induced Ca²⁺ response were no longer present. Conclusions: IL-4 and IL-13 induce bronchial hyperreactivity by changing the Ca²⁺ homeostasis of ASMC. These effects are counteracted by IL-13Rα2. The biological significance of IL-13Rα2 might be a protective function by regulating IL-13- and IL-4-mediated signal transduction and thereby limiting pathological alterations in Th2-mediated inflammatory diseases.

Background: Airway smooth muscle cells (ASMC) play a key role in bronchial hyperresponsiveness (BHR). A major component of the signaling cascade leading to ASMC contraction is calcium. So far, agonist-induced Ca 2+ -signaling in asthma has been studied by comparing innate properties of inbred rat or mouse strains, or by using selected mediators known to be involved in asthma. T-bet knock-out (KO) mice show key features of allergic asthma such as a shift towards T H 2-lymphocytes and display a broad spectrum of asthma-like histological and functional characteristics. In this study, we aimed at investigating whether Ca 2+ -homeostasis of ASMC is altered in T-bet KO-mice as an experimental model of asthma.

Background: Asthma is characterized by airway remodeling, altered mucus production and airway smooth muscle cell (ASMC) contraction causing extensive airway narrowing. In particular, alterations of ASMC contractility seem to be of crucial importance. The elevation of the cytoplasmic Ca²⁺ concentration is a key event leading to ASMC contraction and changes in the agonist-induced Ca²⁺ increase in ASMC have been reported in asthma. Objective: The aim of this study was to investigate mechanisms underlying these changes. Methods: Murine tracheal smooth muscle cells (MTSMC) from T-bet KO mice and human bronchial smooth muscle cells (HBSMC) incubated with IL-13 and IL-4 served as asthma models. Acetylcholine-induced changes in the cytoplasmic Ca²⁺ concentration were recorded using fluorescence microscopy and the expression of Ca²⁺ homeostasis regulating proteins was investigated with Western blot analysis. Results: Acetylcholine-induced Ca²⁺ transients were elevated in both asthma models. This correlated with an increased Ca²⁺ content of the sarcoplasmic reticulum (SR). In MTSMC from T-bet KO mice, the expression of the SR Ca²⁺ buffers calreticulin and calsequestrin was higher compared to wild-type mice. In HBSMC incubated with IL-13 or IL-4, the expression of ryanodine receptors, inositol-3-phosphate receptors and sarcoplasmic/endoplasmic reticulum Ca²⁺ ATPases 2 was increased compared to HBSMC without incubation with interleukins. The enlarged acetylcholine-induced Ca²⁺ transients could be reversed by blocking inositol-3-phosphate receptors. Conclusions: We conclude that in the murine asthma model the SR Ca²⁺ buffer capacity is increased, while in the human asthma model the expression of SR Ca²⁺ channels is altered. The investigation of the Ca²⁺ homeostasis of ASMC has the potential to provide new therapeutical options in asthma.

For characterization and determination of recombinant bovine GH (rbGH) eight monoclonal antibodies (MAb) were produced against rbGH from Monsanto. The various MAb showed different affinities to rbGH, pituitary bovine GH (pbGH), and pituitary ovine GH (poGH). With epitope analysis several MAb were shown to recognize different epitopes of rbGH. The MAb MUC-rbGH-3A11 and MUC-rbGH-1E5 were used to develop a Sandwich ELISA. By checking the specificity of the assay no cross reactivity was found with pituitary porcine GH, pituitary human GH, bovine or ovine prolactin and little cross reactivity with poGH could be found. The Sandwich ELISA detected various rbGH (Monsanto, Elanco, Cyanamid) with different N-terminal amino acids and discriminated between rbGH and pituitary bovine GH by an affinity factor of 2.0. The detection level was 2 ng rbGH per ml PBS buffer. The recovery was about 86% in bovine serum. It might therefore be possible to detect rbGH-treated cows using a Sandwich ELISA, but this would need a field study.

The influence of Echinacea angustifolia, Aconitum napellus, Lachesis mula, and Apis meUiJica, the contents of the combination product Influex (Steigerwald, Germany), on the phagocytosis of human granulocytes was investigated. Incorporation of latex microspheres was measured with a flow cytometric method. With regard to the constituents of the prescription, only vitamin C, lipopolysaccharide, and Echinacea angustifoh were able to enhance the phagocytosis rate. Aconitum, Apis and Lachesis, alone were not able to stimulate phagocytosis but showed an enhancement effect when combined with one another or with Echinacea. For example, a combination of Aconifum (lo-') and Lachesis (lo-') was able to stimulate phagocytosis up to 390%.

Summary In order to obtain high amounts of specific antibodies against the hapten methylphosphoric acid, para‐aminophenyl‐1,2,2‐trimethyl‐propyldiester (MATP) different animals were immunized with MATP coupled to the carrier protein human serum albumin (MATP12‐HSA) using several adjuvants. The best specific immune response in sheep, rabbit and chicken was reached with Freund's complete adjuvant with animal specific differences being tested by an ELISA. The adjuvants aluminium hydroxide (5% and 10%) and diphosphoryl lipid A showed no significant difference compared to the control group (NaCl with MATP12‐HSA). In rabbits and chickens MATP12‐HSA can be used to reach an immune response without the help of an adjuvant.

In this study, rabbit antisera to hapten-rabbit serum albumin conjugates were investigated regarding antibody titer, affinity, specificity, and affinity distribution. Methyl phosphonic acid p-amino-phenyl 1,2,2-trimethylpropyldiester (MATP) served as model hapten. Four MATP-rabbit serum albumin conjugates with various hapten densities (with and without spacer) were synthesized and used for immunization of rabbits. Antisera were collected over a 130 day-period and characterized with different ELISA methods. We found that immunogens with rabbit serum albumin gave antisera with lower titers, but similar affinity as compared to polyclonal or monoclonal antibodies obtained with non-rabbit protein as carrier protein. Immunogens with a low hapten density led to higher final titers without affecting antibody affinity or specificity. Immunogens containing a bridging group resulted in higher antibody affinity with a changed specificity. The pattern of antibody affinity distribution differed considerably among individual rabbits and showed a non-Gaussian subpopulation distribution.