935 MHz cellular phone radiation. An in vitro study of genotoxicity in human lymphocytes.

PURPOSE: The possibility of genotoxicity of radiofrequency radiation (RFR) applied alone or in combination with x-rays was investigated in vitro using several assays on human lymphocytes. The chosen specific absorption rate (SAR) values are near the upper limit of actual energy absorption in localized tissue when persons use some cellular telephones. The purpose of the combined exposures was to examine whether RFR might act epigenetically by reducing the fidelity of repair of DNA damage caused by a well-characterized and established mutagen. METHODS: Blood specimens from 14 donors were exposed continuously for 24 h to a Global System for Mobile Communications (GSM) basic 935 MHz signal. The signal was applied at two SAR; 1 and 2 W/Kg, alone or combined with a 1-min exposure to 1.0 Gy of 250 kVp x-rays given immediately before or after the RFR. The assays employed were the alkaline comet technique to detect DNA strand breakage, metaphase analyses to detect unstable chromosomal aberrations and sister chromatid exchanges, micronuclei in cytokinesis-blocked binucleate lymphocytes and the nuclear division index to detect alterations in the speed of in vitro cell cycling. RESULTS: By comparison with appropriate sham-exposed and control samples, no effect of RFR alone could be found for any of the assay endpoints. In addition RFR did not modify any measured effects of the x-radiation. CONCLUSIONS: This study has used several standard in vitro tests for chromosomal and DNA damage in Go human lymphocytes exposed in vitro to a combination of x-rays and RFR. It has comprehensively examined whether a 24-h continuous exposure to a 935 MHz GSM basic signal delivering SAR of 1 or 2 W/Kg is genotoxic per se or whether, it can influence the genotoxicity of the well-established clastogenic agent; x-radiation. Within the experimental parameters of the study in all instances no effect from the RFR signal was observed.

Cytogenetic study on children living in Southern Urals contaminated areas (nuclear incidents 1948-1967).

As a result of the activities of the first Soviet plutonium production reactor, large territories of the Southern Urals were exposed to radioactive contamination. Three different incidents occurring between 1948 and 1967 lead to major exposure. A total of 280,000 people residing on the contaminated territories were exposed both to external and internal contamination particularly due to the long-lived radionuclides 137Cs and 90Sr. The highest doses were received by 28,000 people living on the Techa riverside villages. In the present paper 15 presumably exposed children coming from the Muslyumovo village on the Techa river have been analyzed using conventional cytogenetic procedure in order to assess a radiation-induced damage. The data obtained have been compared to a group of matched unexposed controls. The results show a statistical difference between the two cohorts which suggests a possible residual contamination representing a continuous hazard for the new generations.

Lysosomal alpha-mannosidases of mouse tissues: characteristics of the isoenzymes, and cloning and expression of a full-length cDNA.

Lysosomal alpha-d-mannosidase from mouse tissues was separated into its constituent isoenzymes by DEAE-cellulose chromatography. Forms corresponding to the human isoenzymes B and A were present in testis, brain, spleen and kidney, whereas in epididymis and liver only the B form was present. Murine alpha-mannosidases A and B are glycoproteins and have pH optima, thermal stabilities and molecular masses similar to those of the human isoenzymes. A full-length cDNA (3.1 kb) containing the complete coding sequence for alpha-mannosidase was isolated from a mouse macrophage cDNA library. Comparison of the deduced amino acid sequences of human and mouse alpha-mannosidases showed that they had 75% identity and 83% similarity. Expression of this cDNA in COS cells showed that both the A and the B isoenzymes can arise from a single transcript. Northern blotting analysis showed a 10-fold range in the abundance of alpha-mannosidase mRNA in mouse tissues, with the highest levels found in epididymis, and the lowest in liver.

Constitutive expression of beta-N-acetylhexosaminidase in a microglial cell line: transcriptional modulation by lipopolysaccharide and serum factors.

We investigated the expression of the alpha- and beta-subunits of the lysosomal enzyme beta-N-acetylhexosaminidase in the BV-2 microglial cell line under different culture conditions. Beta-N-acetylhexosaminidase from BV-2 microglia cells was separated into its constituent isoenzymes on diethylaminoethyl (DEAE) cellulose, and its activity was monitored with 4-methylumbelliferyl-beta-N-acetylglucosamine and 4-methylumbelliferyl-beta-N-acetylglucosamine-6-sulphate substrates. Forms corresponding to the mouse isoenzymes A and B were present in the cells incubated in serum-supplemented medium as well as in serum-free medium. Lipopolysaccharide, a well-known activator of microglia in vitro, added to the BV-2 cells in serum-supplemented medium induced a decrease in the specific enzymatic activity determined with the 4-methylumbelliferyl-beta-N-acetylglucosamine substrate. Lipopolysaccharide had no effect on hexosaminidase isoenzyme pattern of BV-2 cells in serum-supplemented medium. The level of alpha-subunit mRNA was increased and the level of beta-subunit mRNA was decreased in BV-2 cells incubated in serum-supplemented medium plus lipopolysaccharide. In the cells incubated in a serum-free medium no significant changes in the hexosaminidase-specific activities towards the above substrates were observed. Interestingly, increased expression of alpha- and beta-subunit mRNA was evident in comparison with cultures in serum-supplemented medium. The present results suggest that the BV-2 cell line may be a useful tool to study the possible role of microglia in the metabolism of brain glycolipids.

Structural organization and expression of the gene for the mouse GM2 activator protein.

The GM2 activator protein is an essential component for the degradation of GM2 ganglioside by hexosaminidase A in vivo. Mutations in the human gene coding for the GM2 activator protein cause the AB variant of GM2-gangliosidosis, a condition that is clinically indistinguishable from Tay-Sachs disease. To understand better factors affecting the expression of the GM2 activator protein gene (Gm2a) in mouse tissues, we have determined its exon-intron organization and analyzed its promoter region. Gm2a is about 14 kb, has four exons, and the 5' flanking region contains a CAAT box, Sp1 binding sites, AP-1, AP-2 sites, and a pair of IRE sites. A 1.2-kb fragment upstream from the initiation codon was shown to have promoter activity in NIH 3T3 cells. Similarities between the elements present in Gm2a and Hexa promoters might in part explain their similar expression patterns in mouse tissues. The different levels of GM2 activator protein mRNA in liver, kidney, brain, and testis are not owing to the use of different transcription start sites, because a single start site was found 50 bp upstream from the initiation codon in each these tissues. Northern blot analysis demonstrated variation in the GM2 activator protein mRNA expression during mouse development. Gm2a was mapped to Chromosome (Chr) 11, where it co-segregated with Csfgm.

Cytogenetic effects in lymphocytes from children exposed to radiation fall-out after the Chernobyl accident.

In a previous paper we reported that a group of children exposed to ionizing radiation following the Chernobyl accident exhibited an appreciable number of chromosome breaks and rearrangements reflecting the persistence of a radiation-induced damage. The results suggested that the children were still exposed to radioactive contamination through consumer foodstuff and life styles. In the present paper, 31 exposed children have been considered together with a control group of 11 children with the aim to confirm previous results. All children underwent whole-body counter (WBC) measures and conventional cytogenetic analysis. The frequency of chromosome aberrations detected by conventional cytogenetics in the group of children chronically exposed to low doses of ionizing radiation resulted in significant differences with respect to the control group. The present work suggests that, for these groups of children, even if the frequency of aberrations is very low and the observation of statistically significant differences is consequently a problem, a persistently abnormal cytogenetic picture is still present several years after the accident.

Structural organization and expression of the gene for the mouse GM2 activator protein.

The GM2 activator protein is an essential component for the degradation of GM2 ganglioside by hexosaminidase A in vivo. Mutations in the human gene coding for the GM2 activator protein cause the AB variant of GM2-gangliosidosis, a condition that is clinically indistinguishable from Tay-Sachs disease. To understand better factors affecting the expression of the GM2 activator protein gene (Gm2a) in mouse tissues, we have determined its exon-intron organization and analyzed its promoter region.Gm2a is about 14 kb, has four exons, and the 5' flanking region contains a CAAT box, Spl binding sites, AP-1, AP-2 sites, and a pair of IRE sites. A 1.2-kb fragment upstream from the initiation codon was shown to have promoter activity in NIH 3T3 cells. Similarities between the elements present in Gm2a and Hexa promoters might in part explain their similar expression patterns in mouse tissues. The different levels of GM2 activator protein mRNA in liver, kidney, brain, and testis are not owing to the use of different transcription start sites, because a single start site was found 50 bp upstream from the initiation codon in each these tissues. Northern blot analysis demonstrated variation in the GM2 activator protein mRNA expression during mouse development. Gm2a was mapped to Chromosome (Chr) 11, where it co-segregated with Csfgm.

Do human lymphocytes exposed to the fallout of the Chernobyl accident exhibit an adaptive response? 1. Challenge with ionizing radiation.

Several studies suggest that cells appear to become less susceptible to the induction of radiation damage, and in particular of chromosome and chromatid aberrations in short-term cultures of human lymphocytes, when a challenge exposure to ionizing radiation is preceded by a low 'adaptive' dose. Contradictory results have been reported on the conditions under which the phenomenon can be evidenced. In the present work, circulating lymphocytes of 13 children contaminated from the fallout after the Chernobyl accident were tested for their capability to exhibit an adaptive response in experiments in which the challenge dose was administered to stimulated lymphocytes in the S-G2 phase. Furthermore, the possible influence of 3-aminobenzamide, an inhibitor of poly(ADP-ribose) polymerase, was also investigated. Our results indicate that, at least in the instance of the end-point here used (chromosome and chromatid aberrations, the former resulting possibly from the Cs burden), human lymphocytes, chronically exposed to low doses from fallout, do not exhibit any decreased susceptibility to ionizing radiation. However, as reported in the accompanying paper, the same samples appear to show an 'adaptive' response when exposed to a challenge treatment with bleomycin (B. Tedeschi et al., 1995, this issue).

Do human lymphocytes exposed to the fallout of the Chernobyl accident exhibit an adaptive response? 2. Challenge with bleomycin.

The present study concerns the possible adaptive response, induced in vivo by a continuous exposure to ionizing radiations, to a challenge treatment with the radiomimetic glycopeptide bleomycin (BLM). Lymphocytes from children contaminated as a consequence of Chernobyl accident were treated for the last 5 h of culture with 2.5 micrograms/ml BLM. The induced chromosome damage was significantly lower than that found with the same treatment in lymphocytes from control children. This hyposensitivity to BLM was still present if, 1 h after the addition of the drug, inhibitors of the enzymes involved in DNA repair, such as 3-aminobenzamide (2 mM), or aphidicolin (0.4 microM) or 3-dideoxythymidine (5 mM) were added to the cultures. The resistance to BLM in lymphocytes from contaminated children seems to be related to a mechanism upstream in respect to the activities of enzymes involved in the DNA repair and specifically linked to the action of this drug. This is consistent with the different response found when the cells were challenged with ionizing radiation in vitro, as reported in the accompanying paper (L. Padovani, L. et al. (1995) Mutation Res., this issue).

Ultrastructural and cytochemical analysis for adenyl cyclase in frog choroid plexus.

An ultrastructural and cytochemical analysis of the anterior choroid plexus in adult Rana esculenta was undertaken. The epithelial cells are implicated in the production of cephalorachidian liquid by transporting metabolites from the blood and by synthesizing and secreting activity. The epithelial cells are also capable of re-absorbing catabolites from the cephalorachidian liquid. The presence of adenylate cyclase, along the basal and lateral membranes in some epithelial cells and along the apical membranes of others, leads us to hypothesize that the epithelium of the plexus is made up of two cell types, one type with a secretory function and another type with an absorption function.