Antioxidant enzymatic systems in pigment tissue of amphibia

Superoxide dismutase, catalase, glutathione peroxidase, and glutathione reductase activities in pigmented and unpigmented liver tissues of frog and albino rat, respectively, were studied. Our results show that pigmented tissue is lacking in manganese superoxide dismutase activity and that the main enzymatic activity utilized in the cytosol by pigmented cells to reduce the hydrogen peroxide to water is represented by catalase; on the contrary, for the same reaction, the cells of albino rat liver primarily utilize the glutathione peroxidase activity. Both a low glutathione peroxidase activity and a low glutathione reductase activity were found in pigmented tissue of frog liver when compared with unpigmented tissue of rat liver. In light of our results, we also report a hypothetical interrelationship between melanin and reduced glutathione: We believe that in pigmented cells the melanin could act as a reducing physiological agent replacing the glutathione in the reduction of hydrogen peroxide. This reducing action of melanin could cause a diminished need for GSH and therefore could provoke the low glutathione peroxidase and reductase activities in pigmented tissue.     

Ultrastructural organization of vegetative hyphae of Tuber Albidum Pico

The ultrastructural organization of vegetative hyphae ofTuber albidum, an ectomycorrhizal fungus is described. Three zones are recognized: 1) an apical zone with cytoplasmic vesicles; 2) a subapical zone with protoplasmic organelles; 3) a vacuolated zone. This organization corresponds in its main features to the pattern just described for rapidly growing fungi.     

Calmodulin in mouse male germ cells: a qualitative and quantitative study

Isolated male germ cells of the mouse possess a heat-stable stimulatory activity of Ca2+-dependent, calmodulin-free phosphodiesterase. Ionic exchange chromatography allowed partial purification of the activator and the isolation of multiple forms of phosphodiesterase stimulation inhibitor. The activator has been identified as calmodulin on the basis of chromatographic behaviour and electrophoretic mobility. Quantitative analysis showed variations of calmodulin levels at different stages of spermatogenesis. Quantitative analysis of cyclic nucleotide hydrolysis in germ cell cytosol showed that the activity of Ca2+-dependent phosphodiesterase is different in meiotic and post-meiotic mouse male germ cells. These data suggest that calcium-dependent pathway and a Ca2+-dependent regulation of cyclic nucleotides are present in developing germ cells.     

Isoelectric focusing studies of human pancreatic secretion

Pure bile, pancreatic and duodenal human juices have been analyzed by isoelectric focusing, either at rest or upon stimulation with caerulein. In rats, stimulation has also been performed with secretin. Twenty bands have been resolved and quantified in the pancreatic secretion. By developing zymograms, a number of isozymes have been identified: 6 isoamylases [pI's 7.2, 7.1 and 6.6 (major) and pI's 7.4, 6.7 and 5.8 (minor)], 3 lipases [pI's 7.0 and 6.8 (major) and 6.4 (minor)], two major alkaline proteases (pI's 9.8 and 8.4) and one major acidic protease (pI 4.3) and one band of RNAase activity (pI 8.6). The stimulation kinetics follow a mechanism according to Palade, indicating uniform response to secretogogues, parallel intracellular transport and parallel discharge of pancreatic exocrine proteins.     

Synthesis and crystal structure of the MgCl2(CH3COOC2H5)2· (CH3COOC2H5) adduct

The reaction of α-MgCl₂ with boiling ethyl acetate affords MgCI₂(CH₃COOC₂H₅)₂· (CH₃COOC₂H₅), which is obtained as crystals suitable for X-ray analysis only from the mother liquor. M=315.5, orthorhombic, space group P2₁22₁ (No. 18), a=25.077(3), b=8.616(1), c=7.345(1) Å, V=1587.0(3) ų, Z=4, D_x=1.32 g cm⁻³,λ A(Mo Kα)=0.71069 Å, μ=4.17 cm⁻¹, F(000)=664, T=298 K, observed reflections: 1667, R=0.059 and R_w=0.069. The structure is composed of polymeric chains of MgCl₂(CH₃COOC₂H₅₎₂ and the ethyl acetate molecules occupy a mutually trans position.     

Molecular cloning of the imidazoleglycerolphosphate dehydratase gene of Trichoderma harzianum by genetic complementation in Saccharomyces cerevisiae using a direct expression vector

Summary The Trichoderma harzianum imidazoleglycerolphosphate dehydratase gene (igh) has been isolated by complementation of a Saccharomyces cerevisiae his3 mutant using a direct expression vector. This Escherichia coli-yeast shuttle vector was developed to allow efficient cloning and expression of cDNA libraries. The cDNA is 627 nucleotides long and codes for a protein of 209 amino acids with an apparent molecular mass of 22 466 daltons. The predicted protein sequence showed 63.6%, 58.7%, and 38.4% identity respectively to the corresponding enzymes from S. cerevisiae, Pichia pastoris and E. coli. Northern analysis showed that the expression of the igh gene in T. harzianum is not inhibited by external histidine and the level of igh mRNA was about threefold higher in cells starved of histidine.     

Involvement of TRAIL/TRAIL-receptors in human intestinal cell differentiation

Despite the fact that tumor necrosis factor (TNF)-related apoptosis inducing ligand (TRAIL) and its receptors (TRAIL-Rs) are expressed in intestinal mucosa, little is known about the biological role of this system in intestinal cell physiology. The expression of surface TRAIL and TRAIL-R1, -R2, -R3, -R4 were examined by flow cytometry in the immortalized human cell line tsFHI under culture conditions promoting growth or growth arrest and expression of differentiated traits. A progressive increase of surface TRAIL expression paralleled tsFHI differentiation, consistently with immunohistochemistry analysis showing an increase of TRAIL immunostaining along the crypt-villus axis in normal jejuneal mucosa. In spite of the presence of TRAIL-R1 and TRAIL-R2 "death receptors," recombinant TRAIL was not cytotoxic for tsFHI cells. Exposure of tsFHI to recombinant TRAIL rather increased/anticipated the expression levels of the cyclin-dependent kinase inhibitors p21 and p27, which mediate the induction of growth arrest and the stabilization of differentiated traits, respectively, as well as of the canonical differentiation marker DPPIV. The differentiation inducing activity of TRAIL was abolished by pre-incubation with a Fc-TRAIL-R2 chimera. On the other hand, TRAIL did not significantly modulate the levels of osteoprotegerin (OPG), CXCL8/IL-8, CXCL9/MIG, and CXCL10/IP10 spontaneously released or induced by inflammatory cytokines. Taken together, these data suggest that TRAIL might act as a paracrine trophic cytokine on intestinal epithelium, promoting intestinal cell differentiation.     

PLC-beta2 monitors the drug-induced release of differentiation blockade in tumoral myeloid precursors

The differentiation therapy in treatment of acute promyelocytic leukemia (APL), based on the administration of all-trans retinoic acid (ATRA), is currently flanked with the use of As2O3, a safe and effective agent for patients showing a resistance to ATRA treatment. A synergy between ATRA and As3O3 was also reported in inducing granulocytic differentiation of APL-derived cells. We have demonstrated that phospholipase C-beta2 (PLC-beta2), highly expressed in neutrophils and nearly absent in tumoral promyelocytes, largely increases during ATRA treatment of APL-derived cells and strongly correlates with the responsiveness of APL patients to ATRA-based differentiating therapies. Here we report that, in APL-derived cells, low doses of As3O3 induce a slight increase of PLC-beta2 together with a moderate maturation, and cooperate with ATRA to provoke a significant increase of PLC-beta2 expression. Remarkably, the amounts of PLC-beta2 draw a parallel with the differentiation levels reached by both ATRA-responsive and -resistant cells treated with ATRA/As2O3 combinations. PLC-beta2 is not necessary for the progression of tumoral promyelocytes along the granulocytic lineage and is unable to overcome the differentiation block or to potentiate the agonist-induced maturation. On the other hand, since its expression closely correlates with the differentiation level reached by APL-derived cells induced to maturate by drugs presently employed in APL therapies, PLC-beta2 represents indeed a specific marker to test the ability of differentiation agents to induce the release of the maturation blockade of tumoral myeloid precursors.