Chronic immune thrombocytopenic purpura--immunological analyses of a patient pre- and post-HIV seroconversion

A seven year follow-up of immune parameters is reported for a patient with chronic immune thrombocytopenic purpura (ITP) pre and post human immunodeficiency virus (HIV) seroconversion. Therapies such as intravenous IgG, prednisone, vincristine, or Ciclosporin A had no clear-cut beneficial effect on platelet counts. A long-term normalization of platelet counts was achieved by splenectomy. At splenectomy the patient was seropositive for HIV, most likely transmitted by blood products received half a year prior to laparatomy. Mean plasma levels of the second component of complement, C2, were half of the normal values prior to and within the lowest normal range post HIV seroconversion. Nevertheless, the T cell-dependent B cell response to HIV, which is dependent on the activation of C3 via the classical pathway of complement, was normal: Western blot analysis of total IgG and of IgG subclass responses to individual HIV antigens proved to be unimpaired.     

The use of functional analysis of the ribosome as a tool to determine archaebacterial phylogeny

Forty different antibiotics with diverse kingdom and functional specificities were used to measure the functional characteristics of the archaebacterial translation apparatus. The resulting inhibitory curves, which are characteristic of the cell-free system analyzed, were transformed into quantitative values that were used to cluster the different archaebacteria analyzed. This cluster resembles the phylogenetic tree generated by 16S rRNA sequence comparisons. These results strongly suggest that functional analysis of an appropriate evolutionary clock, such as the ribosome, is of intrinsic phylogenetic value. More importantly, they indicate that the study of the nexus between genotypic and phenotypic (functional) information may shed considerable light on the evolution of the protein synthetic machinery.     

Inhibition of carbonic anhydrases in type I muscle fibers influences contractility

We tested the effects of inhibiting the carbonic anhydrase activity of rat soleus and extensor digitorum longus muscles on the isometric contractile properties and the resistance to fatigue. SOL and EDL muscles from female rats were incubated in vitro in the presence of methazolamide, a specific inhibitor of carbonic anhydrase, before determining their contractile properties. Methazolamide had no effects on the contractile properties of the soleus muscle (10(-5) or 10(-3) M) and extensor digitorum longus (10(-3) M), except for the half-relaxation time of the soleus muscle which increased significantly. Values for half-relaxation time were significantly increased with both concentrations of the inhibitor. Muscles were then submitted to a fatigue protocol lasting 30 min. During the fatigue test, no significant difference was observed between control and 10(-5) M methazolamide soleus muscles. In presence of 10(-3) M methazolamide however, the soleus muscle showed a significantly increased resistance to fatigue compared with control preparations. No significant effect was observed with the extensor digitorum longus muscle exposed to 10(-3) M methazolamide. Results are discussed in terms of the presence of two different isoforms of carbonic anhydrase that may be associated with calcium uptake and energy metabolic processes, respectively.     

Experimental autoimmune encephalomyelitis in "low" and "high" interleukin 2 producer rats. I. Cellular basis of induction

Albino Oxford (AO) rats in comparison to the Dark August (DA) strain exhibit lower susceptibility to the induction of experimental autoimmune encephalomyelitis (EAE), and interleukin 2 (IL-2) production by their spleen and lymph node cells is significantly lower. The cellular analysis of these differences in the outcome of the EAE induction, possibly related to the differences in the IL-2 production, revealed different changes in the T cell subsets in the draining lymph node (DLN) and different cellular composition of the mononuclear infiltrates in the central nervous system (CNS). After the encephalitogenic challenge, the frequency of CD8+ T cells was much higher and the expansion of CD4+ T cells was much lower in the DLN of "low" IL-2 producer rats. AO rats have not shown any clinical sign of EAE, although histological lesions in the early phases of EAE (Day 7-9) were similar to those seen in diseased DA rats. CD4/CD8 T cell ratios and the number of cells bearing receptor for IL-2 (IL-2-R+ cells) and cells bearing class II MHC antigens (Ia+) were significantly lower in the mononuclear cell infiltrates of AO rats. These data are compatible with the notion that CD4+ IL-2-R+ encephalitogenic T cells induce clinical signs of EAE in susceptible animals and show that CD8+ T cells are present in a higher percentage in the lesions of the symptom-free AO rats.     

Careful consideration of the effects induced by glutathione depletion in rat liver and heart. The involvement of cytosolic and mitochondrial glutathione pools

One of the most widely used mechanisms by which the role of glutathione (GSH) in cellular functions has been withdrawn, is to deplete GSH intracellularly. The importance of the procedure and xenobiotic chosen to get it is discussed. Mitochondrial GSH plays certainly an important role in maintaining cellular homeostasis. This contribution varies depending on the tissue and the conclusions obtained about the functions of this GSH pool in one organ may not be applied to others. Original data on the subcellular distribution of GSH in myocardial tissue of the rat are presented, and the effect of phorone on both cardiac GSH pools is compared with the effect in liver. The mechanical failure of myocardium after ischemic or reperfusion damage might involve mitochondrial GSH, in view of the literature data referring to the role of thiol groups in energy transfer from mitochondria to cytosol.     

Identification of the spI products of Balbiani ring genes in Chironomus thummi

The spI fraction of high molecular weight secretory proteins was analysed in Chironomus thummi. These proteins are encoded by giant Balbiani ring (BR) genes which develop specifically in salivary gland cells. Each component of the spI fraction was studied electrophoretically from early and middle 4th instar larvae and prepupae, as well from galactose-treated larvae where changes in the relative puffing pattern of BR1 and BR2 are known to occur. The spI fraction consists of at least two bands with electrophoretic mobilities slower than those of the spI components of Camptochironomus. The slow migrating component remains throughout the 4th larval instar, while the amount of the faster component changes, being abundant in early 4th instar and prepupae, but not present (or very weak) in middle 4th instar. The correlated shifts in BR puffing pattern during these developmental stages suggest that the slow and fast components are encoded by BR2 and BR1. The spI fraction is modified by galactose treatment, the fast component being induced in parallel with a decrease in the slow component. These changes are correlated with changes in the steady-state levels of RNA: an increase in BR1 RNA and a decrease in BR2 RNA, and of proteins. These proteins could correspond to the spIb and spIa fractions allocated to BR2 and BR1, respectively, in Camptochironomus. After galactose treatment a new faster band sometimes appears, that could correspond to the spIc fraction of Camptochironomus. A possible spId equivalent was also identified. In conclusion the main features of the spI family in C. thummi are similar to those of spI in Camptochironomus.Abbreviations BR Balbiani ring - spI family of M_r=10⁶ secretory polypeptides     

Effect of glycosaminoglycans and divalent cations on the thermal properties of dipalmitoylphosphatidylcholine

The effect of glycosaminoglycans (GAG) and divalent cations on the thermal properties of dipalmitoyl-phosphatidylcholine (DPPC)-water systems was examined in order to model some interactions taking place on low density lipoprotein (LDL) surfaces. The thermal properties of these systems were measured by differential scanning calorimetry (DSC). According to the results, all three glycosaminoglycans used (chondroitin-4-sulfate, chondroitin-6-sulfate and heparin) were effective but to a different extent. Calcium ions enhance the interaction more than magnesium ions, probably because divalent cations form bridges between the negatively charged groups of GAGs and the headgroups of lipids. It is conceivable that similar processes might occur in the case of LDL.     

Characteristics of the pulmonary transport functions for heat and dye in pulmonary edema and orthostasis

The aim of this study was to investigate whether changes in the distribution of pulmonary blood flow and disturbances of the pulmonary microcirculation can be detected by use of inflow-outflow indicator-dilution measurements. In 18 anesthetized (N2O-piritramide) mongrel dogs 221 thermal-indocyanine green dye indicator dilution kinetics were recorded in the pulmonary artery and aorta after central venous indicator injection. The lagged normal density function was used as a model for the pulmonary transport functions for heat and dye. The parameters of the lagged normal density function were computed by a non-linear least squares procedure by iterative convolution. After baseline measurements, in nine dogs, pulmonary edema was induced by central venous application of oleic acid. In nine other dogs, measurements were performed before and after postural changes. Our data show that both the microvascular injury caused by oleic acid edema and the perfusion heterogeneity caused by orthostasis can be detected by the indicator dilution technique since the both relative dispersion and skewness of the transport functions for heat and dye were significantly increased after these interventions.     

Alkaline phosphatase in cholestatic and cirrhotic rats. A biochemical and histochemical study

Alkaline phosphatase (ALP) was studied by enzyme histochemical methods and by biochemical quantitations in rat livers with chronic bile duct obstruction and experimental cirrhosis. The most evident ALP increase was histochemically found in portal tracts of rats with bile duct obstruction and localized to the walls of proliferating blood vessels. Furthermore, a slight canalicular membrane enzyme increase was histochemically found in both groups, most evident in cirrhosis, whereas the biochemical assay of ALP in serum and liver from both pathological groups showed 3 times higher values compared to controls. The portal tracts did not seem to contribute to the serum increase, since the rise of serum ALP was similar in chronic bile duct obstruction and in experimental cirrhosis without changes of the portal tracts. It is concluded that the increase ALP activity in serum from rats with bile duct obstruction and cirrhosis mainly has a hepatocytic origin.     

N-linked oligosaccharide changes with oncogenic transformation require sialylation of multiantennae

Glycopeptides derived from NIH 3T3 fibroblasts and these cells transformed by transfection with human DNA containing oncogene H-ras were analyzed by 500-MHz 1H-NMR spectroscopy and binding to immobilized lectins. The cells were metabolically labeled with D-[3H]glucosamine or L-[3H]fucose and the glycopeptides included in Bio-Gel P-10 (Mr 5000-3500) were separated into neutral and charged fractions on DEAE-cellulose. The major portion (80%) of these [3H]fucose glycopeptides from the non-transformed NIH 3T3 fibroblasts were neutral or contained one or two charged residues, whereas 90% of the glycopeptides from the transformed cells contained two or more charged residues. The structure of the predominant neutral glycopeptide from the non-transformed NIH 3T3 cells was determined by 1H-NMR spectroscopy to be tetraantennary containing terminal Gal alpha 1----3. (formula; see text) This structure was verified by binding to the immobilized alpha-Gal-specific lectin, Griffonia simplicifolia I and leukoagglutinating phytohemagglutinin from Phaseolus vulgaris (L-PHA), which binds certain tri- or tetraantennary glycopeptides. In contrast, the structure derived by NMR spectroscopy of one of the predominant charged glycopeptides from the transformed cells was triantennary containing terminal NeuNAc alpha 2----3 in addition to Gal alpha 1----3. (formula; see text) In attempting to verify this structure by lectin-binding properties it was found that removal of NeuNAc alpha 2----3 reduced the affinity to L-PHA - agarose. The other major glycopeptides of the transformed cells which were more charged also cotained NeuNAc alpha 2----3 but no NeuNAc alpha 2----6 or Gal alpha 1----3. A tentative structure was proposed for the major glycopeptide of the first charged class from NIH 3T3 cells on the basis of lectin-binding properties and the NMR spectrum which showed, in addition to NeuNAc alpha 2----3, the presence of NeuNAc alpha 2----6 and Gal alpha 1----3. On the basis of the NMR spectrum and other results, it is concluded that the presence of tetraantennary oligosaccharides are not sufficient for the transformed oligosaccharide phenotype. Rather, the tri- or tetraantennae must be sialylated in alpha 2----3 linkage, on more than one antennae, when properties of transformation are expressed in NIH 3T3 cells. Prior to transformation the tetraantennary oligosaccharides of these cells are terminated in alpha-Gal residues, whereas after transformation alpha-Gal residues appear to be replaced by NeuNAc alpha 2----3.(ABSTRACT TRUNCATED AT 400 WORDS)