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. 2015:2015:479412.
doi: 10.1155/2015/479412. Epub 2015 Mar 16.

Preliminary Studies of the Immunomodulator Effect of the Bougainvillea xbuttiana Extract in a Mouse Model

Lluvia Arteaga Figueroa  1 , Leticia Barbosa Navarro  1 , Martin Patiño Vera  2 , Vera L Petricevich  1
Affiliations

Preliminary Studies of the Immunomodulator Effect of the Bougainvillea xbuttiana Extract in a Mouse Model

Lluvia Arteaga Figueroa et al. Evid Based Complement Alternat Med. 2015.

Abstract

Bougainvillea xbuttiana is used as an analgesic in folk medicine in Mexico. The purpose of the present study was to determine the effects of the ethanolic extract from B. xbuttiana on macrophages activities. The phytochemical screening was performed for determine the presence of alkaloids, flavonoids, triterpenes, and saponins. The effects of B. xbuttiana were analyzed using the macrophages activities as determined by the H2O2 release, spreading and phagocytic index, vacuoles formation percentage, and mediators production. The viability percentage was determined in live cells after fixing and staining with crystal violet. The presence of H2O2 in macrophages was performed by using the peroxidase-phenol red solution. The cytokine production was determined by two assays, ELISA for detection of IL-6, IL-10, and IFN-γ and biological assay for TNF detection. The results showed that the Bxb extract dose-dependent manner produces (a) an increase in levels of H2O2 and spreading and vacuoles formation percentages, (b) a decrease in phagocytic index and in the amounts of TNF, IL-6, and IFN-γ, and (c) an increase significant in IL-10 and NO production. This study indicates that the ethanolic extract from Bougainvillea xbuttiana was able to activate macrophages. The combination of these results suggests that this extract has an immunomodulator effect.

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Figures

Figure 1
Figure 1
Effect of Bxb on peritoneal macrophage viability. Peritoneal macrophages were obtained and treated in vitro with different concentrations of Bxb as described above. The viability percentage was observed at 48 h after extract exposition. Each bar represents the mean value of samples from three experiments in different groups of five mice. Statistical differences between the treatments were P < 0.05.
Figure 2
Figure 2
Cytotoxic activity of Bxb. Peritoneal macrophages were obtained and exposed in vitro to 2.9, 29, and 290 μg/mL of Bxb as described above. The cytotoxic percentage was observed at different times after Bxb exposition. Each point represents the mean value of samples from four experiments in different groups of five mice. Statistical differences between the treatments were P < 0.05.
Figure 3
Figure 3
Phagocytosis index. Peritoneal macrophages were obtained and exposed to 2.9, 29, and 290 μg/mL of Bxb and the phagocytic index was determined as described above. Each point represents the mean value of samples from four experiments in different groups of five mice. Statistical differences between the treatments were P > 0.05.
Figure 4
Figure 4
Vacuole formation. Peritoneal macrophages were obtained and exposed to 2.9, 29, and 290 μg/mL of Bxb as described above. After different times of incubation at 37°C in an atmosphere of 5% CO2, the cells were stained with neutral red for 5 min. The absorbance was determined at 540 nm, and the results were expressed as described above. Each point represents the mean value of samples from four experiments in different groups of four mice. Statistical differences between the treatments were P < 0.05.
Figure 5
Figure 5
Spreading index. Peritoneal macrophages were obtained and exposed to 2.9, 29, and 290 μg/mL of Bxb and the spreading percentage was determined as described above. Each point represents the mean value of samples from four experiments in different groups of five mice. Statistical differences between the treatments were P < 0.05.
Figure 6
Figure 6
Hydrogen peroxide production. Peritoneal macrophages were obtained and exposed to 2.9, 29, and 290 μg/mL of Bxb and the hydrogen peroxide production was determined directly on cells as described above. Each point represents the mean value of samples from four experiments in different groups of five mice. Statistical differences between the treatments were P < 0.05.
Figure 7
Figure 7
Proinflammatory cytokines production. Peritoneal macrophages were obtained and exposed in vitro to 2.9, 29, and 290 μg/mL of Bxb as described above. At different times, the levels of IL-6 and IFN-γ were determined by ELISA assay using monoclonal antibodies as the probe. The levels of TNF were determined by standard assay with L929 cells. Each point represents the mean value of samples from four experiments in different groups of five mice. Statistical differences between the treatments were P < 0.05.
Figure 8
Figure 8
Anti-inflammatory cytokines production. Peritoneal macrophages were obtained and exposed to 2.9, 29, and 290 μg/mL of Bxb and at different times the levels of IL-10 were assayed by ELISA assay using monoclonal antibodies as the probe. Each point represents the mean value of samples from four experiments in different groups of five mice. Statistical differences between the treatments were P < 0.05.
Figure 9
Figure 9
NO production. Peritoneal macrophages were obtained and exposed to 2.9, 29, and 290 μg/mL of Bxb and at different times the levels of NO were determined by Griess colorimetric reaction. Each point represents the mean value of samples from four experiments in different groups of five mice. Statistical differences between the treatments were P < 0.05.
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