Alkaloids from Prosopis juliflora leaves induce glial activation, cytotoxicity and stimulate NO production.

Prosopis juliflora is used for feeding cattle and humans. Intoxication with the plant has been reported, and is characterized by neuromuscular alterations and gliosis. Total alkaloidal extract (TAE) was obtained using acid/basic-modified extraction and was fractionated. TAE and seven alkaloidal fractions, at concentrations ranging 0.03-30 microg/ml, were tested for 24h on astrocyte primary cultures derived from the cortex of newborn Wistar rats. The MTT test and the measure of LDH activity on the culture medium, revealed that TAE and fractions F29/30, F31/33, F32 and F34/35 were cytotoxic to astrocytes. The EC(50) values for the most toxic compounds, TAE, F31/33 and F32 were 2.87 2.82 and 3.01 microg/ml, respectively. Morphological changes and glial cells activation were investigated through Rosenfeld's staining, by immunocytochemistry for the protein OX-42, specific of activated microglia, by immunocytochemistry and western immunoblot for GFAP, the marker of reactive and mature astrocytes, and by the production of nitric oxide (NO). We observed that astrocytes exposed to 3 microg/ml TAE, F29/30 or F31/33 developed compact cell body with many processes overexpressing GFAP. Treatment with 30 microg/ml TAE and fractions, induced cytotoxicity characterized by a strong cell body contraction, very thin and long processes and condensed chromatin. We also observed that when compared with the control (+/-1.34%), the proportion of OX-42 positive cells was increased in cultures treated with 30 microg/ml TAE or F29/30, F31/33, F32 and F34/35, with values raging from 7.27% to 28.74%. Moreover, incubation with 3 microg/ml F32, 30 microg/ml TAE, F29/30, F31/33 or F34/35 induced accumulation of nitrite in culture medium indicating induction of NO production. Taken together these results show that TAE and fractionated alkaloids from P. juliflora act directly on glial cells, inducing activation and/or cytotoxicity, stimulating NO production, and may have an impact on neuronal damages observed on intoxicated animals.

Suppresion of the antivenom antibody response by serum therapy

1. The antivenom antibody response of mice injected with Bathrops jararaca venom and receiving specific serum therapy was studied under different experimental conditions. Balb/c mice (18-22g) injected with venom (1.75 mg/kg) presented the clinical symptoms observed in patients bitten by B. jararaca and a high and long-lasting antivenom antibody response. 2. Injection of 0.1 ml of horse antiserum to venom 15 min after venom administration abolished the symptoms induced by the venom and induced an almost completely suppressed production of mouse antivenom antibodies. The extent of suppression of the antivenom antibody response depended on the dose of horse antiserum administered and was greater the sooner the serum therapy was applied after envenomation. 3. Injection of antiserum into envenomed mice that received an unrelated antigen (KLH) did not suppress the antibody response to KLH antigen though it inhibited production of antivenom antibodies. 4. Envenomed mice receiving an equivalent dose of F(ab')2 fragments obtained by pepsin digestion of horse antiserum presented the same extent of suppression of the antivenom antibody response as mice injected with the non-treated antiserum. 5. Mice whose antibody response was suppressed, when rechallenged with venom, presented a primary antibody response. 6. These results suggest that suppression of the antivenom antibody response presented by envenomed patients submitted to serum therapy is due to the masking of the venom epitopes by horse antibodies as well as to the rapid elimination of the venom epitopes

Comparison between the antigenic composition of bloodstream and cell culture-derived trypomastigotes of Trypanosoma cruzi

Antigens of bloodstream and cell culture-derived trypomastigotes of T. cruzi were compared by western blotting using sera of chronic chagasic patient as a source of antiobodies. The immunoblots demonstrated that the two forms display extensive homology except for the 85 - and 52 - kDa bands. These antigens were more strongly stained in culture - derived trypomastigotes. Although the reported differences are not related to major antigens, these results might offer an explanation for previous studies showing that culture - derived trypomastigotes are more antigenic and infective in vitro than bloodstream trypomastigotes