Suppression of lymphokine production: I. Macrophage-mediated inhibition of MIF production

Macrophages have been found to suppress the in vitro production by stimulated T lymphocytes of a lymphokine, migration inhibitory factor. When macrophages isolated from primary MSV-induced tumors were added to antigen-stimulated MSV-immune spleen cells, a complete suppression of MIF production was observed. This suppression was nonspecific, since MIF production by antigen-stimulated alloimmune spleen cells and by PHA-stimulated normal spleen cells was also inhibited. Suppressor macrophages could also be induced by inoculation with Corynebacterium parvum, whereas light mineral oil-induced peritoneal macrophages had no detectable effect on MIF production. The failure to detect MIF in the supernatants of stimulated cultures containing activated macrophages appeared to be due to inhibition of lymphokine production rather than to absorption or inactivation of MIF or to interference with the assay for detection of MIF. Macrophages were able to suppress MIF production only when added during the first 4–5 hr of culture and they had no effect when added later. These data show that activated macrophages can nonspecifically suppress lymphokine production and that this appears to be due to inhibition of an early step in lymphocyte stimulation.     

Co-oligopeptides containing two aromatic residues spaced by glycyl residues. X. Proton magnetic resonance study of co-oligopeptides of tryptophan and glycine

The ¹H-nmr spectra of co-oligopeptides of tryptophan and glycine with structure H-Gly-Trp-(Gly)_n-Trp-Gly-OH (n = 0–2) and those of several di- and tripeptides have been recorded at 360 MHz with CD₃OD solutions containing 0.1N NaOD. The assignment of resonance signals was generally possible by comparing the spectra of structurally related peptides with each other. In order to solve the remaining ambiguities in the assignment, H-(αL,βS)(α,β-d₂)Trp-OH, H-Trp-(αL,βS)(α,β-d₂)Trp-OH, and H-Trp-(δ¹,ε²,ζ²,ζ³,η²-d₅)Trp-OH have been prepared and their spectra compared with those of the undeuterated compounds. The distribution of rotamers around the χ₁ and (in two cases) χ₂ torsion angles of the side chains has been obtained from the vicinal coupling constants ³J and from the long-range coupling constants ⁴J. These data and an analysis of the chemical shifts of the Gly-C^α protons suggest that the orientation of the aromatic side chain is influenced by the following order of decreasing interaction with the functional groups at N- and C-side: -NH₂ > –NHCO– > –CONH–> –COO^?. This rule does not hold for the second Trp residue of di- and tripeptides containing the -Trp-Trp- sequence, which has tentatively been attributed to steric effects.     

Three-dimensional models of chick embryo somites obtained by mathematical methods applied to ultrastructural data

Summary A model of a thoracolumbar somite of a chick embryo at the 53rd incubation hour was obtained by mathematical methods, after identification of somite cell types by means of electron microscopy.Each specific district occupied by the cell types was precisely determined.On the basis of these observations, the somite was three-dimensionally reconstructed and the spatial positions of the primitive myotome, dermatome, sclerotome, undifferentiated mesoderm and myocele were precisely identified.     

On the fungitoxicity of some new thiocyanatopyrazole derivatives: Electron microscopical study in trichophyton mentagrophytes

Four thiocyanatopyrazole derivatives were synthesized and their fungistatic activity was demonstrated in vitro against a number of dermatophytic fungi. In Trichophyton mentagrophytes, the most active compound induced an unusual increase of the plasma membrane with production of intra and extracytoplasmic complexes, a deterioration of nuclear and mitochondrial membranes and a formation of autophagic-like vacuoles. Plasmolysis, accompanied by an almost complete disorganization of cytoplasmic structures, seemed to be the final event. A possible mechanism of action of the compounds was discussed.Investigation supported by a grant from Consiglio Nazionale delle Ricerche of Italy (Contract No. 7500536).     

Fluorogenic detection of primary amines in plant histochemistry with fluorescamine: A comparative study on the effects of coagulant and non-coagulant fixatives

Summary The new highly sensitive method of fluorescamine reaction for the topochemical detection of primary amino groups was studied as a substitude of ninhydrin-Schiff's reaction for the localisation of total proteins in plant tissues. The influence of various coagulant and non-coagulatn fixatives on the induction of fluorescamine fluorescence was examined: ethanol, formaldehyde gas and solution, glutaraldehyde, acrolein, osmium tetroxide, Bouin, Rossman, Clarke and Zenker's fluids and FMA were employed. It was found that the use of the fluorogenic method is conditioned by the fixative ability to keep the amino groups disposable and by its capability to reduce the natural autofluorescence of plant material. A detailed account of the fixation methodology demonstrated that non-coagulant acrolein and coagulant mercuric chloride are the most promising fixatives for the use of the fluorescamine reaction in plant histochemistry.     

Analytical and preparative isoelectric focusing of peptides in immobilized pH gradients

A new method for peptide analysis and purification is described, based on isoelectric focusing in immobilized pH gradients. On the analytical scale, the peptide zones can now be revealed by an stain for primary and secondary amino group (e.g. ninydrin, fluorescamine, dansyl chloride) since the buffering species, unlike conventional carrier ampholytes, contain only carboxyl and tertiary amino groups. For preparative purposes, conditions have been described to remove most contaminants (e.g. unreacted monomers, non-cross-linked, short polyacrylamide chains) from the gel matrix before the electrophoretic run. However, ca. 2% of the gel dry mass is still present as extractable material. The focused peptides can be recovered in higly yields (ca. 90%) with a fairly high degree of purity (75%), the contaminants being mostly components eluted from the polyacrylamide gel.     

Glycoalkaloid Profile in Potato Haploids Derived from Solanum tuberosum–S. bulbocastanum Somatic Hybrids

Cultivated and wild potato species synthesize a wide variety of steroidal glycoalkaloids (GA) that may affect either human health or biotic stress resistance. Therefore, GA composition must be a major criterion in the evaluation of breeding products when species genomes are merged and/or manipulated. This work reports the results of GA analysis performed on unique haploid (2n=2x=24) plants obtained from tetraploid (2n=4x=48) Solanum bulbocastanum–S. tuberosum hybrids through in vitro anther culture. Glycoalkaloids were extracted from tubers and analyzed by HPLC. Haploids generally showed the occurrence of parental GA. However, in several cases loss of parental GA and gain of new GA lacking in the parents was observed. It may be hypothesized that new GA profiles of our haploids is the result of either genetic recombination or combinatorial biochemistry events. To highlight differences between haploids and parents, soluble proteins and antioxidant activities were also determined. Both were always higher in haploids compared to their parents. The nature of the newly formed GAs will be further investigated, because they may represent new metabolites that can be used against pest and diseases, or are useful for human health.     

Plasmid transformation of Ruminococcus albus by means of high-voltage electroporation

To apply recombinant DNA techniques to the genetic manipulation of cellulolytic ruminal bacteria, a plasmid vector transformation system must be available. The objective of this work was to develop a system for plasmid transformation of Ruminococcus albus. Using high voltage electrotransformation, pSC22 and pCK17 plasmid vectors, derived from lactic acid bacteria plasmids and replicating via single-stranded DNA intermediate, were successfully introduced into three freshly isolated R. albus strains and into R. albus type strain ATCC 27210. The optimization of the electrotransformation condition raised the electroporation efficiency up to 3 x 10(5) transformants per microgram of pSC22 plasmid.     

Effects of 50 Hz magnetic fields on mouse spermatogenesis monitored by flow cytometric analysis

Flow cytometry (FCM) was performed to monitor the cellular effects of extremely-low-frequency magnetic field on mouse spermatogenesis. Groups of five male hybrid F1 mice aged 8–10 weeks were exposed to 50 Hz magnetic field. The strength of the magnetic field was 1.7 mT. Exposure times of 2 and 4 h were chosen. FCM measurements were performed 7, 14, 21, 28, 35, and 42 days after treatment. For each experimental point, a sham-treated group was used as a control. The possible effects were studied by analyzing the DNA content distribution of the different cell types involved in spermatogenesis and using the elongated spermatids as the reference population. The relative frequencies of the various testicular cell types were calculated using specific software. In groups exposed for 2 h, no effects were observed. In groups exposed for 4 h, a statistically significant (P < 0.001) decrease in elongated spermatids was observed at 28 days after treatment. This change suggests a possible cytotoxic and/or cytostatic effect on differentiating spermatogonia. However, further studies are being carried out to investigate the effects of longer exposure times. © 1995 Wiley-Liss, Inc.