Chromosome damage in underground coal miners: detection by conventional cytogenetic techniques and by submitting lymphocytes of unexposed individuals to plasma from at-risk groups

Chromosome abnormalities and the mitotic index in lymphocyte cultures and micronuclei in buccal mucosa cells were investigated in a sample of underground mineral coal miners from Southern Brazil. A decreased mitotic index, an excess of micronuclei and a higher frequency of chromosome abnormalities (fragments, polyploidy and overall chromosome alterations) were observed in the miners when compared to age-paired normal controls from the same area. An alternative assay for clastogenesis in occupational exposition was tested by submitting lymphocytes from non-exposed individuals to a pool of plasmas from the exposed population. This assay proved to be very convenient, as the lymphocytes obtained from the same individuals can be used as target as well as control cells. Also, it yielded a larger number of metaphases and of successful cultures than with common lymphocyte cultures from miners. A significantly higher frequency of chromatid gaps, fragments and overall alterations were observed when lymphocytes from control subjects were exposed to miner plasma pools. Control plasma pools did not significantly induce any type of chromosome alterations in the cultures of normal subjects, thus indicating that the results are not due to the effect of the addition of plasma pools per se.

Genetic variability in salt tolerance during germination of Stylosanthes humilis H.B.K. and association between salt tolerance and isozymes

Variation in salt tolerance of six natural populations of Stylosanthes humilis from three ecogeographic regions, Mata (wet tropical climate), Agreste and Sertão (semi-arid tropical climate) of Pernambuco State, Northeast Brazil, was evaluated on germination in 201 mM NaCl. There were significant differences among families of all populations for germination percentage and of five populations (except Tamandaré, from Mata) for germination rate. Populations from semi-arid regions presented high coefficients of genetic variation, those from Agreste being higher than those from Sertão. Populations from Mata showed low coefficients of genetic variation. The coefficients of genotypic determination were high for five populations, except Tamandaré, both for germination percentage ( > or = 0.89) and for germination rate ( > or = 0.79), indicating the possibility of selection for salt tolerance in these populations. An electrophoretic analysis of esterase and peroxidase isozymes was also performed in the six populations, and correlations were estimated between salt tolerance and allelic frequencies. The analysis of salt tolerant and salt sensitive families of populations from Agreste suggested an association of alleles of a peroxidase locus with salt tolerance during germination in the Caruaru population

A search for RFLP markers to identify genes for aluminum tolerance in maize

The objective of this study was to identify restriction fragment length polymorphism (RFLP) markers linked to QTLs that control aluminum (Al) tolerance in maize. The strategy used was bulked segregant analysis (BSA) and the genetic material utilized was an F2 population derived from a cross between the Al-susceptible inbred line L53 and Al-tolerant inbred line L1327. Both lines were developed at the National Maize and Sorghum Research Center - CNPMS/EMBRAPA. The F2 population of 1554 individuals was evaluated in a nutrient solution containing a toxic concentration of Al and relative seminal root length (RSRL) was used as a phenotypic measure of tolerance. The RSRL frequency distribution was continuous, but skewed towards Al-susceptible individuals. Seedlings of the F2 population which scored the highest and the lowest RSRL values were transplanted to the field and subsequently selfed to obtain F3 families. Thirty F3 families (15 Al-susceptible and 15 Al-tolerant) were evaluated in nutrient solution, using an incomplete block design, to identify those with the smallest variances for aluminum tolerance and susceptibility. Six Al-susceptible and five Al-tolerant F3 families were chosen to construct one pool of Al-susceptible individuals, and another of Al-tolerant, herein referred as "bulks", based on average values of RSRL and genetic variance. One hundred and thirteen probes were selected, with an average interval of 30 cM, covering the 10 maize chromosomes. These were tested for their ability to discriminate the parental lines. Fifty-four of these probes were polymorphic, with 46 showing codominance. These probes were hybridized with DNA from the two contrasting bulks. Three RFLPs on chromosome 8 distinguished the bulks on the basis of band intensity. DNA of individuals from the bulks was hybridized with these probes and showed the presence of heterozygous individuals in each bulk. These results suggest that in maize there is a region related to aluminum tolerance on chromosome 8

Inhibition of lipid peroxidation and iron (II)-dependent DNA damage by extracts of Pothomorphe peltata (L.) Miq

Leaves of Pothomorphe peltata (L.) Miq. (Piperaceae) are used locally as anti-inflammatory, antipyretic, hepatoprotective and diuretic infusions and to treat external ulcers and local infections in several parts of the Peruvian, Bolivian and Brazilian Amazon region. The antioxidant activity of different extracts of P. peltata was studied using the hydroperoxide-initiated chemiluminescence assay in liver homogenates, and the methanolic extract was found to have the highest antioxidant activity, with an IC50 = 4 µg/ml. Aqueous and dichloromethane extracts did not show antioxidant activity. The extracts were further evaluated using the thiobarbituric acid-reactive substances (TBARS) assay. Finally, an assay of DNA sugar damage induced by Fe (II) salt was used to determine the capacity of the extracts to suppress the oxidative degradation of DNA. All the extracts showed antioxidant activity in the latter two bioassays. The methanolic extract showed the highest activity in reducing oxidative damage to DNA, with an IC50 = 5 µg/ml. Since this extract was highly effective in reducing chemiluminescence and DNA damage, and because the latter activity could be due to the presence of compounds that bind to DNA, DNA-binding activity was studied using the DNA-methyl green (DNA-MG) bioassay. A 30% decrease in the initial absorbance of DNA-MG complex was observed in the methanolic extract at 1000 µg/ml, suggesting the presence of compounds that bind to genetic material. No DNA-binding activity was observed in the aqueous or dichloromethane extracts

Effects of naltrexone and cross-tolerance to morphine in a learned helplessness paradigm

Opiates have been implicated in learned helplessness (LH), a phenomenon known to be related to opiate stress-induced analgesia (SIA). In the present study, we investigated the role of opiates in the induction of LH and SIA under different conditions. Adult female Wistar rats were trained either by receiving 60 inescapable 1-mA footshocks (IS group, N = 114) or by confinement in the shock box (control or NS group, N = 92). The pain threshold of some of the animals was immediately evaluated in a tail-flick test while the rest were used 24 h later in a shuttle box experiment to examine their escape performance. The opiate antagonist naltrexone (0 or 8 mg/kg, ip) and the previous induction of cross-tolerance to morphine by the chronic administration of morphine (0 or 10 mg/kg, sc, for 13 days) were used to identify opiate involvement. Analysis of variance revealed that only animals in the IS group demonstrated antinociception and an escape deficit, both of which were resistant to the procedures applied before the training session. However, the escape deficit could be reversed if the treatments were given before the test session. We conclude that, under our conditions, induction of the LH deficit in escape performance is not opiate-mediated although its expression is opiate-modulated

Hepatic damage during acute pancreatitis in the rat

We studied the alterations in the metabolism of liver mitochondria in rats with acute pancreatitis. Male Wistar rats were allocated to a control group (group I) and to five other groups corresponding to 2, 4, 12, 24 and 48 h after the induction of acute pancreatitis by the injection of 5% sodium taurocholate into the pancreatic duct. Sham-operated animals were submitted to the same surgical steps except for the induction of acute pancreatitis. Mitochondrial oxidation and phosphorylation were measured polarographically by determining oxygen consumption without ADP (basal respiration, state 4) and in the presence of ADP (activated respiration, state 3). Serum amylase, transaminases (ALT and AST) and protein were also determined. Ascitic fluid, contents of amylase, trypsin and total protein were also determined and arterial blood pressure was measured in all groups. In ascitic fluid, trypsin and amylase increased reaching a maximum at 2 and 4 h, respectively. Serum amylase increased at 2 h reaching a maximum at 4 h. Serum transaminase levels increased at 12 and 24 h. After 2 h (and also 4 h) there was an increase in state 4 respiration (45.65 ± 1.79 vs 28.96 ± 1.50) and a decrease in respiration control rate (3.53 ± 0.09 vs 4.45 ± 0.08) and in the ADP/O ratio (1.77 ± 0.02 vs 1.91 ± 0.01) compared to controls (P<0.05). These results indicate a disruption of mitochondrial function, which recovered after 12 h. In the 48-h groups there was mitochondrial damage similar to that occurring in ischemic lesion. Beat-to-beat analysis (30 min) showed that arterial blood pressure remained normal up to 24 h (111 ± 3 mmHg) while a significant decrease occurred in the 48-h group (91 ± 4 mmHg). These data suggest biphasic damage in mitochondrial function in acute pancreatitis: an initial uncoupled phase, possibly secondary to enzyme activity, followed by a temporary recovery and then a late and final dysfunction, associated with arterial hypotension, possibly related to ischemic damage.

Aging and immunoglobulin isotype patterns in oral tolerance

In the present review we address oral tolerance as an important biological phenomenon and discuss how it is affected by aging. Other factors such as frequency of feeding and previous digestion of the antigen also seem to influence the establishment of oral tolerance. We also analyze immunoglobulin isotypes of specific antibodies formed by tolerant and immunized animals of different ages submitted to different conditions of oral antigen administration. Isotypic patterns were studied as a parameter for assessing the pathways of B and T cell interactions leading to antibody production

Oral tolerance induction with altered forms of ovalbumin

As a T cell-dependent phenomenon, oral tolerance is not expected to depend necessarily on native configuration of antigens. We investigated the induction of oral tolerance with modified ovalbumin (Ova). Oral administration of heat-denatured (HD-Ova) and cyanogen bromide-degraded ovalbumin was less effective than native Ova in inducing oral tolerance in B6D2F1 mice. HD-Ova was effective in suppressing delayed-type hypersensitivity (DTH) reactions but did not suppress specific antibody formation. Injection of Ova directly into the stomach, but not into the ileum or cecum, suppressed subsequent immunization to DTH reactions. Gavage with protease inhibitors (aprotinin or ovomucoid) before gavage with Ova was ineffective in blocking tolerance induction. Treatment with hydroxyurea to destroy cycling cells 24 h before gavage with Ova blocked oral tolerance induction and also the possibility to passively transfer tolerance to naive recipients with the serum of mice gavaged with Ova 1 h before. The implications of these findings about oral tolerance induction are discussed

Interrelationship between adipocytes and fibroblasts during acute damage to the subcutaneous adipose tissue of rats: an ultrastructural study

Ultrastructural phenotypic transitional features were noted between adult adipocytes and fibroblasts in the subcutaneous adipose tissue of the dorsal pad of normal adult Wistar rats of both sexes, weighing 180-260 g, after acute injury either by the implantation of small (1.8 x 1 x 0.4 cm) perforated plastic boxes or by local heat application. Soon after the inflicted damage, fat-containing cells presented variable shapes. Early after damage, some of these cells were round, adipocyte-like, with numerous and large cytoplasmic fat droplets. A few days later, fat-containing cells became elongated, with the fat droplets in their cytoplasm becoming smaller and less numerous. The cells also showed a prominent active rough endoplasmic reticulum and newly formed collagenous matrix accumulated in the interstices. Although current views consider adult adipocytes to be terminal cells, the present findings, in their time sequence, strongly suggest the transformation of adipocytes into fibroblasts after acute injury to adipose tissue.

Coupling of palmitate to ovalbumin inhibits the induction of oral tolerance

Oral tolerance is a phenomenon that may occur in animals exposed to protein antigens for the first time by the oral route. They become unable to produce immune responses at the levels normally observed when they are immunized parenterally with antigen in the presence of adjuvants. Lipids have been used as adjuvants for both parenteral and oral immunization. In the present study we coupled ovalbumin with palmitate residues by incubating the protein with the N-hydroxysuccinimide palmitate ester and tested the preparation for its ability to induce oral tolerance. This was performed by giving 20 mg of antigen to mice by the oral route 7 days prior to parenteral immunization in the presence of Al(OH)3. Mice were bled one week after receiving a booster that was given 2 weeks after primary immunization. Specific antibodies were detected by ELISA. Despite the fact that the conjugates are as immunogenic as the unmodified protein when parenterally injected in mice, they failed to induce oral tolerance. This discrepancy could be explained by differences in the intestinal absorption of the two forms of the antigen. In fact, when compared to the non-conjugated ovalbumin, a fast and high absorption of the lipid-conjugated form of ovalbumin was observed by "sandwich" ELISA.