Paracrine PDGF-B/PDGF-Rbeta signaling controls mesangial cell development in kidney glomeruli.

Kidney glomerulus mesangial cells fail to develop in mice carrying targeted null mutations in the platelet-derived growth factor (PDGF)-B or PDGF-Rbeta genes. We have examined the pattern of expression of these genes and smooth muscle markers during kidney development, to address the possible mechanisms underlying the mutant phenotypes. In wild-type embryos, PDGF-B was expressed in vascular endothelial cells, particularly in capillary endothelial cells in the developing glomeruli, whereas PDGF-Rbeta was found in perivascular mesenchymal cells in the developing renal cortex.

CD40 and IL-4 regulate murine CD27L expression.

It is well known that interactions between accessory molecules on T cells and their ligands on APC play a key role in regulating T cell effector activity. The factors controlling the expression of these molecules are thus important determinants in the outcome of T cell activation. We have examined the expression of the murine ligand for CD27, a costimulatory molecule on T cells.

Exit of major histocompatibility complex class II-invariant chain p35 complexes from the endoplasmic reticulum is modulated by phosphorylation.

The Iip35 isoform of the major histocompatibility complex (MHC) class II-associated invariant chain (Ii) contains an endoplasmic reticulum (ER) targeting motif, but in B cell lines the ER retention is ineffective and a fraction of Iip35 is transported through the Golgi complex associated with class II molecules. We found Iip35 (but not Iip33, the major form of Ii) to be phosphorylated in B cell lines, as well as in transfected HeLa cells. The phosphorylation of Iip35 was found to be necessary for the exit of Iip35-class II complexes out of the ER.

Altered antigen presentation in mice lacking H2-O.

HLA-DM catalyzes the release of MHC class II-associated invariant chain-derived peptides (CLIP) from class II molecules. Recent evidence has suggested that HLA-DO is a negative regulator of HLA-DM in B cells, but the physiological function of HLA-DO remains unclear. Analysis of antigen presentation by B cells from mice lacking H2-O (the mouse equivalent of HLA-DO), together with biochemical analysis using purified HLA-DO and HLA-DM molecules, suggests that HLA-DO/H2-O influences the peptide loading of class II molecules by limiting the pH range in which HLA-DM is active.

Factors influencing yields of progenitor cells for allogeneic transplantation: optimization of G-CSF dose, day of collection, and duration of leukapheresis.

Mobilization of hematopoietic progenitor cells by G-CSF was attempted on 89 occasions in 85 healthy donors. Three dose ranges of G-CSF were chosen for analysis: low (4-7.4 micrograms/kg), intermediate (7.

Short- and long-term oestradiol treatment inhibits growth and alters expression of p21WAF1/Cip1 in an endometrial adenocarcinoma cell line lacking functional p53.

Oestrogen is assumed to play a significant role in cell cycle regulation of cells expressing the oestrogen receptor, although its mechanism of action is not yet well defined. To examine this, a mutant p53-expressing human endometrial adenocarcinoma cell line of the oestradiol-inhibited growth phenotype was treated with oestradiol for 2 weeks (short-term) and 6 months (long-term). With short-term treatment, cells were treated with increasing doses of oestradiol.

Estradiol induces DNA fragmentation in a human endometrial adenocarcinoma with estradiol-inhibited growth phenotype.

A moderately differentiated human endometrial adenocarcinoma heterotransplanted into nude mice was investigated for morphological and molecular changes in the tumours after treating the animals with estradiol. The tumour growth was previously characterised as estradiol-independent but responsive (inhibited) without any changes in cell proliferation. In response to hormonal treatment rather the cell loss factor increased.

Alveogenesis failure in PDGF-A-deficient mice is coupled to lack of distal spreading of alveolar smooth muscle cell progenitors during lung development.

PDGF-A(-/-) mice lack lung alveolar smooth muscle cells (SMC), exhibit reduced deposition of elastin fibres in the lung parenchyma, and develop lung emphysema due to complete failure of alveogenesis. We have mapped the expression of PDGF-A, PDGF receptor-alpha, tropoelastin, smooth muscle alpha-actin and desmin in developing lungs from wild type and PDGF-A(-/-) mice of pre- and postnatal ages in order to get insight into the mechanisms of PDGF-A-induced alveolar SMC formation and elastin deposition. PDGF-A was expressed by developing lung epithelium.

Major histocompatibility complex class II-dependent unfolding, transport, and degradation of endogenous proteins.

We have analyzed the ability of major histocompatibility (MHC) class II molecules to capture proteins in the biosynthetic pathway and whether this may be associated with MHC class II-dependent antigen processing. When coexpressed with HLA-DR 4 molecules in HeLa cells, influenza hemagglutinin was inhibited from folding and trimerization in the biosynthetic pathway, targeted to endosomal compartments, and rapidly degraded. Due to the interaction with MHC class II molecules, therefore, unfolded forms of hemagglutinin were bypassing the quality control mechanism of the secretory pathway.

Thymic selection by a single MHC/peptide ligand produces a semidiverse repertoire of CD4+ T cells.

The influence of individual peptides in thymic selection was examined in H2-M- mice, in which positive selection is directed to a single peptide, class II-associated invariant chain peptide (CLIP) bound to H2-A(b). Two sensitive in vivo approaches showed that 70%-80% of CD4+ T cells undergoing positive selection to CLIP+H2-A(b) have self-reactivity to the various peptides expressed on wild-type H2-M+ antigen-presenting cells. When these self-reactive T cells were depleted, the residual CD4+ cells displayed a polyclonal repertoire in terms of alloreactivity, responses to foreign protein antigens, and Vbeta usage.

Pharmacokinetics and effects of HI 6 in blood and brain of soman-intoxicated rats: a microdialysis study.

The bispyridinium oxime HI 6 (1-(((4-amino-carbonyl)pyridino)methoxy)methyl)-2-(hydroxyimino )methyl)-pyridinium dichloride monohydrate), combined with atropine, is effective for treating poisoning with organophosphate nerve agents. The protective action of HI 6 in soman poisoning has been attributed mainly to its peripheral reactivation of inhibited acetylcholinesterase. In the present study we investigated whether high intramuscular doses of HI 6 can reach the brain in a sufficient amount to reactivate inhibited brain acetylcholinesterase.

Decreased endosomal delivery of major histocompatibility complex class II-invariant chain complexes in dynamin-deficient cells.

Major histocompatibility complex class II molecules are heterodimeric cell surface molecules which acquire antigenic peptides in the endosomal/lysosomal system. Invariant chain (Ii), a third chain which is associated with class II molecules intracellularly mediates the endosomal targeting, but it is debated whether class II molecules reach the endosomal system mainly from the trans-Golgi network or via the cell surface. Dynamin is a cytosolic GTPase which is necessary for the formation of clathrin-coated vesicles from the plasma membrane, but which is not required for vesicle formation from the trans-Golgi network.

Quantitative study of intussusceptive capillary growth in the chorioallantoic membrane (CAM) of the chicken embryo.

In an attempt to sort out the respective contributions of sprouting and intussusceptive microvascular growth (IMG) during chicken chorioallantoic membrane (CAM) development, we analyzed the morphology and the quantitative growth of the capillary bed of the CAM by light microscopy. By perfusing the CAM microvasculature with highly concentrated colloidal gold particles, the capillaries could be unambiguously distinguished from the surrounding unlabelled tissue. This allowed us to identify, count and measure the intercapillary tissue profiles.

Antigen specificity of dual reactive T hybridomas determines the requirement for CD40 ligand-CD40 interactions.

CD40 ligand (CD40L) expression on T cells is known to play a crucial role in B cell responses. Some evidence also supports a role for CD40L-CD40 interactions in T cell responses, at least in vivo. Whether the T cell requirement for these interactions is an invariable finding, however, is less clear.

Selectivity of the major histocompatibility complex class II presentation pathway of cortical thymic epithelial cell lines.

Major histocompatibility complex (MHC) restriction of the immune response is established during positive selection of T cells in the thymus. This occurs mainly through interactions of T cell receptor of developing thymocytes with MHC/peptide ligands on cortical thymic epithelial cells (TEC). An ongoing controversy concerns the origin and the role of peptides involved in the positive selection of thymocytes.

The use of supercritical fluid extraction for sample preparation of pharmaceutical formulations.

This paper reviews the use of supercritical fluid extraction (SFE) as a sample work-up step in the analysis of drugs in various pharmaceutical formulations. Matrices studies include tablets, animal feed, creams, ointments and infusions. As in other fields of analytical chemistry, SFE has proven to be most suitable for comparatively non-polar compounds in solid matrices.

Tumour growth and cell kinetics in variants of a human endometrial adenocarcinoma expressing either wild-type or mutant p53.

We have compared the baseline cell proliferation and tumour growth in two variants of a human endometrial adenocarcinoma grown in nude mice. One of these tumour variants expressed wild-type p53 whereas the other had mutations of the p53 gene at codon 175 in both alleles and at codon 248 in one allele. There was no difference in growth rate between the tumour variants.

HLA-DO is a lysosomal resident which requires association with HLA-DM for efficient intracellular transport.

The murine MHC class II molecule H2-O is expressed in B-cells and in thymic epithelium but the human equivalent, HLA-DO (DO), has not been detected, though the corresponding genes, HLA-DNA and HLA-DOB, are well known. Here we show DO to be a lysosomal resident in B-cells. Surprisingly, DO forms stable complexes with HLA-DM (DM), another lysosomal class II-like molecule which is important for class II-restricted antigen presentation.

Simulation of re-entry in a piece of myocardial tissue: strong sensitivity to spatial and temporal conditions.

A simple model for the simulation of re-entrant excitation was created. The model consists of a matrix of 15x15 compartments, where each compartment has its own action potential that depends dynamically on four ion currents (INa, ICa, Ik and Ib) having time and voltage-dependent activation and inactivation kinetics. The compartments were combined with resistors to simulate electrotonic interaction.

Characterisation of a human endometrial adenocarcinoma established in vivo and in vitro.

A moderately differentiated human endometrial adenocarcinoma was heterotransplanted into nude mice and later established as a continuous in vitro cell line. Western blot analysis showed an accumulation of p53 protein in the cell line compared to the original tumour and heterotransplants. Sequential analysis of the p53 gene revealed point mutations in codons 175 and 248 in the cell line while no mutations were found prior to in vitro establishment.

Estradiol regulates tumor growth by influencing p53 and bcl-2 expression in human endometrial adenocarcinomas grown in nude mice.

This study analyzes the effects of estradiol on p53 and bcl-2 expression, tumor growth and cell kinetic parameters in three human endometrial adenocarcinomas grown in nude mice. The tumors used were estradiol receptor (ER) positive but differed in receptor concentration and hormone sensitivity. All three tumors expressed wild-type p53 protein.

Development of a Solid Phase Carbon Trap for Simultaneous Determination of PCDDs, PCDFs, PCBs, and Pesticides in Environmental Samples Using SFE-LC.

The purpose of this study was to find materials that can be used as a solid phase trap in a commercial supercritical fluid extraction instrument. PX-21 active carbon mixed with ODS proved to yield the best results. After SFE of the target compounds, the planar fraction containing the environmental pollutants PCDDs, PCDFs, and planar PCBs was successfully separated on a PX-21 solid phase trap from the nonplanar fraction containing the other PCBs and pesticides.

Antigen presentation and T cell development in H2-M-deficient mice.

HLA-DM (DM) facilitates peptide loading of major histocompatibility complex class II molecules in human cell lines. Mice lacking functional H2-M, the mouse equivalent of DM, have normal amounts of class II molecules at the cell surface, but most of these are associated with invariant chain-derived CLIP peptides. These mice contain large numbers of CD4+ T cells, which is indicative of positive selection in the thymus.

Genomic organization of a mouse MHC class II region including the H2-M and Lmp2 loci.

The region encompassing the Ma, Mb1, Mb2, and Lmp2 genes of the mouse class II major histocompatibility complex (MHC) was sequenced. Since this region contains clusters of genes required for efficient class I and class II antigen presentation, it was interesting to search for putative additional genes in the 21 kilobase gap between the Mb1 and Lmp2 genes. Computer predictions of coding regions and CpG islands, exon trapping experiments, and cross-species comparison with the corresponding human sequence indicate that no additional functional gene is present in that stretch.