Studies on the function of Brutoll 's tyrosine kinase in B cell development

Each individual organism has to protect itself against a large variety of infectious microbial agents, such as bacteria, fungi and parasites to prevent pathological damage and death. In vertebrates, defense mechanisms against foreign substances, antigens, have evolved in the immune system, which has two functional divisions: the 'innate' immune system and the 'adaptive' immune system. The 'innate' immune system is aspecific and acts as a first line of defense, mediated by cells from thc myeloid lineage and soluble factors like complement and lysozyme. The main function of the 'innate' immune system is to avoid entering of micro or gall isms into the body and to clear it of killed pathogens. In contrast to the 'adaptive' immune system, repeated infection does not improve the resistance of the 'innate' immune system. If the first line of defense is defeated, the second line of defense, the 'adaptive' immune system, which is very specific and can develop memory to earlier accounted pathogens, is activated. The specific immune response is mediated by lymphocytes belonging to the B andlor T lineages (B and T cells). Both Band T cells express receptor molecules on their cell membrane, which specifically can bind antigens. The T ceH receptor (TCR) can only bind antigens if these are processed into small peptides, and presented by major histocompatibility complex (MHC) class I molecules on the surface of host cells and MHC class II molecules on the surface of antigen presenting cells. Intracellular antigens are processed into small peptides and presented on the surface by the MHC class I complex. Recognition of the MHC-class I-peptide complex by the TCR of cytotoxic T cells results in killing of the presenting host cells. MHC class II molecules present processed peptides, which are derived from external clldocytosed antigens. Recognition of MHC class II-peptide complexes by the TCR ofT helper (T H) cells results in the production of cytokines and stimulation of cells of the immune system ('cellular' immune response). The B cell receptor (BCR) binds to unprocessed antigens. Stimulation of the BCR results in a 'humoral' immune response, i.e. the secretion of soluble immunoglobulins (Ig)J with the same binding specificity as the BCR to the triggering antigen

Function of Bruton's tyrosine kinase during B cell development is partially independent of its catalytic activity

The Tec family member Bruton's tyrosine kinase (Btk) is a cytoplasmic protein tyrosine kinase that transduces signals from the pre-B and B cell receptor (BCR). Btk is involved in pre-B cell maturation by regulating IL-7 responsiveness, cell surface phenotype changes, and the activation of lambda L chain gene rearrangements. In mature B cells, Btk is essential for BCR-mediated proliferation and survival. Upon BCR stimulation, Btk is transphosphorylated at position Y551, which promotes its catalytic activity and subsequently results in autophosphorylation at position Y223 in the Src homology 3 domain. To address the significance of Y223 autophosphorylation and the requirement of enzymatic activity for Btk function in vivo, we generated transgenic mice that express the autophosphorylation site mutant Y223F and the kinase-inactive mutant K430R, respectively. We found that Y223 autophosphorylation was not required for the regulation of IL-7 responsiveness and cell surface phenotype changes in differentiating pre-B cells, or for peripheral B cell differentiation. However, expression of the Y223F-Btk transgene could not fully rescue the reduction of lambda L chain usage in Btk-deficient mice. In contrast, transgenic expression of kinase-inactive K430R-Btk completely reconstituted lambda usage in Btk-deficient mice, but the defective modulation of pre-B cell surface markers, peripheral B cell survival, and BCR-mediated NF-kappaB induction were partially corrected. From these findings, we conclude that: 1) autophosphorylation at position Y223 is not essential for Btk function in vivo, except for regulation of lambda L chain usage, and 2) during B cell development, Btk partially acts as an adapter molecule, independent of its catalytic activity

Sustainable Irrigation Management of Ornamental Cordyline Fruticosa “Red Edge” Plants with Saline Water

The aim of this work was to analyze the influence of the salinity of the nutrient solution on the transpiration and growth of Cordyline fruticosa var. “Red Edge” plants. A specific irrigation management model was calibrated with the experimental data. An experiment was performed with four treatments. These treatments consisted of the application of four nutrient solutions with different electrical conductivity (ECw) levels ranging from 1.5 dS m−1 (control treatment) to 4.5 dS m−1. The results showed that day-time transpiration decreases when salt concentration in the nutrient solution increases. The transpiration of the plant in the control treatment was modelled by applying a combination method while the effect of the salinity of the nutrient solution was modelled by deriving a saline stress coefficient from the experimental data. The results showed that significant reductions in plant transpiration were observed for increasing values of ECw. The crop development and yield were also affected by the increasing salinity of the nutrient solution. A relationship between the ECw and the relative crop yield was derived

Serotyping, ribotyping, PCR-mediated ribosomal 16S-23S spacer analysis and arbitrarily primed PCR for epidemiological studies on Legionella pneumophila

Fifty clinical and environmental isolates of Legionella pneumophila were typed serologically and by DNA fingerprinting using arbitrarily primed polymerase chain reaction (AP-PCR). Furthermore, variability in and around ribosomal operons was assessed by conventional ribotyping and PCR-mediated amplification of the spacer region separating the 16S and 23S genes. It appears that serotyping suffers from low resolution capabilities, and ribotyping and spacer PCR display intermediate resolving capabilities, whereas AP-PCR is more discriminating. Results from AP-PCR and both forms of ribotyping analysis correlate with epidemiological and environmental data. It is suggested that AP-PCR typing may be the method of choice for rapidly determining clonality among L. pneumophila isolates

Coronary vasodilatory action of elgodipine in coronary artery disease

The effects of intravenous elgodipine, a new second-generation dihydropyridine calcium antagonist, on hemodynamics and coronary artery diameter were investigated in 15 patients undergoing cardiac catheterization for suspected coronary artery disease. Despite a significant decrease in systemic blood pressure, elgodipine infused at a rate of 1.5 micrograms/kg/min over a period of 10 minutes did not affect heart rate and left ventricular end-diastolic pressure. The contractile responses during isovolumic contraction showed a slight but significant increase in maximum velocity (56 +/- 10 to 60 +/- 10 seconds-1; p less than 0.005), whereas the time constant of early relaxation was shortened from 49 +/- 11 to 44 +/- 9 ms (p less than 0.05). Coronary sinus and great cardiac vein flow increased significantly by 15 and 26%, respectively. As mean aortic pressure decreased, a significant decrease in coronary sinus (-27%) and great cardiac vein (-28%) resistance was observed, while the calculated myocardial oxygen consumption remained unchanged. In all, 69 coronary segments (including 13 stenotic segments) were analyzed quantitatively using computer-assisted quantitative coronary angiography. A significant increase in mean coronary artery diameter (2.27 +/- 0.53 to 2.48 +/- 0.53 mm; p less than 0.000001), as well as in obstruction diameter, (1.08 +/- 0.29 to 1.36 +/- 0.32 mm; p less than 0.02), was observed. The results demonstrate that elgodipine, in the route and dose described, induces significant vasodilatation of both coronary resistance and epicardial conductance vessels, without adverse effects on heart rate, myocardial oxygen demand and contractile indexes

A Dual Reporter Mouse Model of the Human β-Globin Locus: Applications and Limitations

The human β-globin locus contains the β-like globin genes (i.e. fetal γ-globin and adult β-globin), which heterotetramerize with α-globin subunits to form fetal or adult hemoglobin. Thalassemia is one of the commonest inherited disorders in the world, which results in quantitative defects of the globins, based on a number of genome variations found in the globin gene clusters. Hereditary persistence of fetal hemoglobin (HPFH) also caused by similar types of genomic alterations can compensate for the loss of adult hemoglobin. Understanding the regulation of the human γ-globin gene expression is a challenge for the treatment of thalassemia. A mouse model that facilitates high-throughput assays would simplify such studies. We have generated a transgenic dual reporter mouse model by tagging the γ- and β-globin genes with GFP and DsRed fluorescent proteins respectively in the endogenous human β-globin locus. Erythroid cell lines derived from this mouse model were tested for their capacity to reactivate the γ-globin gene. Here, we discuss the applications and limitations of this fluorescent reporter model to study the genetic basis of red blood cell disorders and the potential use of such model systems in high-throughput screens for hemoglobinopathies therapeutics