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A serine protease isolated from the bristles of the Amazonic caterpillar, Premolis semirufa, is a potent complement system activator.

Villas Boas IM, Pidde-Queiroz G, Magnoli FC, Gonçalves-de-Andrade RM, van den Berg CW, Tambourgi DV - PLoS ONE (2015)

Bottom Line: The caterpillar of the moth Premolis semirufa, commonly named pararama, is found in the Brazilian Amazon region.The protease, named here as Ps82, reduced the haemolytic activity of the alternative and classical pathways and inhibited the lectin pathway.The use of Phenanthroline, metalloprotease inhibitor, in the reactions did not significantly interfere with the activity of the Ps82, whereas the presence of PMSF, serine protease inhibitor, totally blocked the activity.

View Article: PubMed Central - PubMed

Affiliation: Immunochemistry Laboratory, Butantan Institute, São Paulo, SP, Brazil.

ABSTRACT

Background: The caterpillar of the moth Premolis semirufa, commonly named pararama, is found in the Brazilian Amazon region. Accidental contact with the caterpillar bristles causes an intense itching sensation, followed by symptoms of an acute inflammation, which last for three to seven days after the first incident. After multiple accidents a chronic inflammatory reaction, called "Pararamose", characterized by articular synovial membrane thickening with joint deformities common to chronic synovitis, frequently occurs. Although complement mediated inflammation may aid the host defense, inappropriate or excessive activation of the complement system and generation of anaphylatoxins can lead to inflammatory disorder and pathologies. The aim of the present study was to evaluate, in vitro, whether the Premolis semirufa's bristles extract could interfere with the human complement system.

Results: The bristles extract was able to inhibit the haemolytic activity of the alternative pathway, as well as the activation of the lectin pathway, but had no effect on the classical pathway, and this inhibition seemed to be caused by activation and consumption of complement components. The extract induced the production of significant amounts of all three anaphylatoxins, C3a, C4a and C5a, promoted direct cleavage of C3, C4 and C5 and induced a significant generation of terminal complement complexes in normal human serum. By using molecular exclusion chromatography, a serine protease of 82 kDa, which activates complement, was isolated from P. semirufa bristles extract. The protease, named here as Ps82, reduced the haemolytic activity of the alternative and classical pathways and inhibited the lectin pathway. In addition, Ps82 induced the cleavage of C3, C4 and C5 and the generation of C3a and C4a in normal human serum and it was capable to cleave human purified C5 and generate C5a. The use of Phenanthroline, metalloprotease inhibitor, in the reactions did not significantly interfere with the activity of the Ps82, whereas the presence of PMSF, serine protease inhibitor, totally blocked the activity.

Conclusion: These data show that a serine protease present in the Premolis semirufa's bristles extract has the ability to activate the complement system, which may contribute to the inflammatory process presented in humans after envenomation.

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Related in: MedlinePlus

Terminal Complement Complex (TCC) generation induced by the Premolis semirufa’s bristles extract.Samples of NHS (50 μL) were incubated with the bristles extract (175 μg/mL) or PBS, in the presence or absence of 10 mM 1,10-Phenanthroline (Phen), at 37°C for 30 min, and SC5b-9 complex present in human serum samples was measured using the MicroVue SC5b-9 Plus EIA Kit. Data are representative for two separate experiments, performed in duplicate, and the results are expressed as concentration of SC5b-9 complex per mL of human serum (ng/mL) ± SD. (***) p < 0.001: significant differences between the mean values obtained with the buffer and between the treatments.
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pone.0118615.g003: Terminal Complement Complex (TCC) generation induced by the Premolis semirufa’s bristles extract.Samples of NHS (50 μL) were incubated with the bristles extract (175 μg/mL) or PBS, in the presence or absence of 10 mM 1,10-Phenanthroline (Phen), at 37°C for 30 min, and SC5b-9 complex present in human serum samples was measured using the MicroVue SC5b-9 Plus EIA Kit. Data are representative for two separate experiments, performed in duplicate, and the results are expressed as concentration of SC5b-9 complex per mL of human serum (ng/mL) ± SD. (***) p < 0.001: significant differences between the mean values obtained with the buffer and between the treatments.

Mentions: To evaluate whether activation/consumption of the complement triggered the activation of the Terminal complement pathway, we measured the concentration of SC5b-9 complex present in normal human serum samples incubated with the Premolis semirufa’s bristles extract in the presence or absence of 1,10-phenanthroline. As shown in Fig. 3, the bristles extract induced a significant generation of Terminal complement complexes in normal human serum samples that, in turn, was completely inhibited by 1,10-phenanthroline.


A serine protease isolated from the bristles of the Amazonic caterpillar, Premolis semirufa, is a potent complement system activator.

Villas Boas IM, Pidde-Queiroz G, Magnoli FC, Gonçalves-de-Andrade RM, van den Berg CW, Tambourgi DV - PLoS ONE (2015)

Terminal Complement Complex (TCC) generation induced by the Premolis semirufa’s bristles extract.Samples of NHS (50 μL) were incubated with the bristles extract (175 μg/mL) or PBS, in the presence or absence of 10 mM 1,10-Phenanthroline (Phen), at 37°C for 30 min, and SC5b-9 complex present in human serum samples was measured using the MicroVue SC5b-9 Plus EIA Kit. Data are representative for two separate experiments, performed in duplicate, and the results are expressed as concentration of SC5b-9 complex per mL of human serum (ng/mL) ± SD. (***) p < 0.001: significant differences between the mean values obtained with the buffer and between the treatments.
© Copyright Policy
Related In: Results  -  Collection

License
Show All Figures
getmorefigures.php?uid=PMC4356561&req=5

pone.0118615.g003: Terminal Complement Complex (TCC) generation induced by the Premolis semirufa’s bristles extract.Samples of NHS (50 μL) were incubated with the bristles extract (175 μg/mL) or PBS, in the presence or absence of 10 mM 1,10-Phenanthroline (Phen), at 37°C for 30 min, and SC5b-9 complex present in human serum samples was measured using the MicroVue SC5b-9 Plus EIA Kit. Data are representative for two separate experiments, performed in duplicate, and the results are expressed as concentration of SC5b-9 complex per mL of human serum (ng/mL) ± SD. (***) p < 0.001: significant differences between the mean values obtained with the buffer and between the treatments.
Mentions: To evaluate whether activation/consumption of the complement triggered the activation of the Terminal complement pathway, we measured the concentration of SC5b-9 complex present in normal human serum samples incubated with the Premolis semirufa’s bristles extract in the presence or absence of 1,10-phenanthroline. As shown in Fig. 3, the bristles extract induced a significant generation of Terminal complement complexes in normal human serum samples that, in turn, was completely inhibited by 1,10-phenanthroline.

Bottom Line: The caterpillar of the moth Premolis semirufa, commonly named pararama, is found in the Brazilian Amazon region.The protease, named here as Ps82, reduced the haemolytic activity of the alternative and classical pathways and inhibited the lectin pathway.The use of Phenanthroline, metalloprotease inhibitor, in the reactions did not significantly interfere with the activity of the Ps82, whereas the presence of PMSF, serine protease inhibitor, totally blocked the activity.

View Article: PubMed Central - PubMed

Affiliation: Immunochemistry Laboratory, Butantan Institute, São Paulo, SP, Brazil.

ABSTRACT

Background: The caterpillar of the moth Premolis semirufa, commonly named pararama, is found in the Brazilian Amazon region. Accidental contact with the caterpillar bristles causes an intense itching sensation, followed by symptoms of an acute inflammation, which last for three to seven days after the first incident. After multiple accidents a chronic inflammatory reaction, called "Pararamose", characterized by articular synovial membrane thickening with joint deformities common to chronic synovitis, frequently occurs. Although complement mediated inflammation may aid the host defense, inappropriate or excessive activation of the complement system and generation of anaphylatoxins can lead to inflammatory disorder and pathologies. The aim of the present study was to evaluate, in vitro, whether the Premolis semirufa's bristles extract could interfere with the human complement system.

Results: The bristles extract was able to inhibit the haemolytic activity of the alternative pathway, as well as the activation of the lectin pathway, but had no effect on the classical pathway, and this inhibition seemed to be caused by activation and consumption of complement components. The extract induced the production of significant amounts of all three anaphylatoxins, C3a, C4a and C5a, promoted direct cleavage of C3, C4 and C5 and induced a significant generation of terminal complement complexes in normal human serum. By using molecular exclusion chromatography, a serine protease of 82 kDa, which activates complement, was isolated from P. semirufa bristles extract. The protease, named here as Ps82, reduced the haemolytic activity of the alternative and classical pathways and inhibited the lectin pathway. In addition, Ps82 induced the cleavage of C3, C4 and C5 and the generation of C3a and C4a in normal human serum and it was capable to cleave human purified C5 and generate C5a. The use of Phenanthroline, metalloprotease inhibitor, in the reactions did not significantly interfere with the activity of the Ps82, whereas the presence of PMSF, serine protease inhibitor, totally blocked the activity.

Conclusion: These data show that a serine protease present in the Premolis semirufa's bristles extract has the ability to activate the complement system, which may contribute to the inflammatory process presented in humans after envenomation.

Show MeSH
Related in: MedlinePlus