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Immunomodulatory effects of Newcastle disease virus AF2240 strain on human peripheral blood mononuclear cells.

Lam HY, Yusoff K, Yeap SK, Subramani T, Abd-Aziz S, Omar AR, Alitheen NB - Int J Med Sci (2014)

Bottom Line: Interestingly, the percentage of cells with activating markers CD16 and CD56 were increased significantly.Human PBMC treated with NDV titer 8 HAU was found to stimulate the highest level of cytokine production including interferon-γ, interleukin-2 and interleukin-12.The release of these proteins contributes to the antitumor effect of PBMC against MCF-7 breast cancer cells.

View Article: PubMed Central - PubMed

Affiliation: 1. Faculty of Biotechnology and Biomolecular Sciences, Universiti Putra Malaysia, 43400, Serdang, Selangor, Malaysia.

ABSTRACT
Immunotherapy has raised the attention of many scientists because it hold promise to be an attractive therapeutic strategy to treat a number of disorders. In this study, the immunomodulatory effects of low titers of Newcastle disease virus (NDV) AF2240 on human peripheral blood mononuclear cells (PBMC) were analyzed. We evaluated cytokine secretion and PBMC activation by cell proliferation assay, immunophenotyping and enzyme linked immunosorbent assay. The proliferation of the human PBMC was measured to be 28.5% and 36.5% upon treatment with 8 hemaglutinin unit (HAU) and 2 HAU of NDV respectively. Interestingly, the percentage of cells with activating markers CD16 and CD56 were increased significantly. Furthermore, the intracellular perforin and granzyme levels were also increased upon virus infection. Human PBMC treated with NDV titer 8 HAU was found to stimulate the highest level of cytokine production including interferon-γ, interleukin-2 and interleukin-12. The release of these proteins contributes to the antitumor effect of PBMC against MCF-7 breast cancer cells. Based on the 3-(4, 5-dimethylthiazol-2-yl)-2, 5-diphenyl tetrazolium bromide assay, activated human PBMC showed high cytolytic efficiency towards human breast tumor cells. In summary, NDV was able to stimulate PBMC proliferation, cytokine secretion and cytolytic activity.

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The granzyme B level in supernatant of MCF-7 cells (target) after being co-cultured for 24 hours with activated PBMC (effector) at 5 to 1 E/T ratios. The values were the means ± SE of three independent experiment. The differences between the control group and treated group were determined by one-way ANOVA. (* p < 0.05).
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Figure 6: The granzyme B level in supernatant of MCF-7 cells (target) after being co-cultured for 24 hours with activated PBMC (effector) at 5 to 1 E/T ratios. The values were the means ± SE of three independent experiment. The differences between the control group and treated group were determined by one-way ANOVA. (* p < 0.05).

Mentions: MTT assay was performed to determine the cytolysis effects of activated PBMC towards MCF-7 cell lines. After 72 hours of incubation of the human PBMC with NDV, the NDV was removed by washing with PBS and the activated PBMC cells were co-cultured with MCF-7 cells. We observed that the higher PBMC effector to MCF-7 target (E/T) ratio gave higher reduction of target cell viability (Figure 5); the MCF-7 viability dropped to 69.66% and 70.18% after being treated with activated PBMC by NDV at 8 and 2 HAU respectively at 5 to 1 E/T ratio. Furthermore, we carried out Annexin-V-FITC/PI flow cytometry apoptosis assay to determine the mode of cell death induced by NDV treated PBMC against MCF-7.The results showed that the activated PBMC induced around 30% of MCF-7 to undergo late apoptosis similar to the results of the MTT assay. At the same time, 8 HAU NDV treated PBMC also induced significant level of early apoptosis on MCF-7 compared to control (Table 4). Overall, the result indicated that 8 HAU NDV activated PBMC induced utmost cell death on MCF-7 cell by apoptosis. In addition, we analyzed the level of granzyme B with the cytolytic effect of NDV activated PBMC to MCF-7 cell. The level of granzyme B in the media of the co-culture was assayed using ELISA. In the media of MCF-7 cell co-cultured with 8 and 2 HAU NDV treated PBMC, 3.11 and 2.84 fold (which were 287 pg/ml and 261 pg/ml) increase of granzyme B was detected (Figure 6). On the other hand, minimum spontaneous release of granzyme B (<20 pg/ml) was detected in the NDV treated or untreated PBMC that were not co-cultured with MCF-7. This result indicated that elevated granzyme B expression in NDV treated PBMC may stimulate the apoptosis in MCF-7 cell.


Immunomodulatory effects of Newcastle disease virus AF2240 strain on human peripheral blood mononuclear cells.

Lam HY, Yusoff K, Yeap SK, Subramani T, Abd-Aziz S, Omar AR, Alitheen NB - Int J Med Sci (2014)

The granzyme B level in supernatant of MCF-7 cells (target) after being co-cultured for 24 hours with activated PBMC (effector) at 5 to 1 E/T ratios. The values were the means ± SE of three independent experiment. The differences between the control group and treated group were determined by one-way ANOVA. (* p < 0.05).
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Related In: Results  -  Collection

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Figure 6: The granzyme B level in supernatant of MCF-7 cells (target) after being co-cultured for 24 hours with activated PBMC (effector) at 5 to 1 E/T ratios. The values were the means ± SE of three independent experiment. The differences between the control group and treated group were determined by one-way ANOVA. (* p < 0.05).
Mentions: MTT assay was performed to determine the cytolysis effects of activated PBMC towards MCF-7 cell lines. After 72 hours of incubation of the human PBMC with NDV, the NDV was removed by washing with PBS and the activated PBMC cells were co-cultured with MCF-7 cells. We observed that the higher PBMC effector to MCF-7 target (E/T) ratio gave higher reduction of target cell viability (Figure 5); the MCF-7 viability dropped to 69.66% and 70.18% after being treated with activated PBMC by NDV at 8 and 2 HAU respectively at 5 to 1 E/T ratio. Furthermore, we carried out Annexin-V-FITC/PI flow cytometry apoptosis assay to determine the mode of cell death induced by NDV treated PBMC against MCF-7.The results showed that the activated PBMC induced around 30% of MCF-7 to undergo late apoptosis similar to the results of the MTT assay. At the same time, 8 HAU NDV treated PBMC also induced significant level of early apoptosis on MCF-7 compared to control (Table 4). Overall, the result indicated that 8 HAU NDV activated PBMC induced utmost cell death on MCF-7 cell by apoptosis. In addition, we analyzed the level of granzyme B with the cytolytic effect of NDV activated PBMC to MCF-7 cell. The level of granzyme B in the media of the co-culture was assayed using ELISA. In the media of MCF-7 cell co-cultured with 8 and 2 HAU NDV treated PBMC, 3.11 and 2.84 fold (which were 287 pg/ml and 261 pg/ml) increase of granzyme B was detected (Figure 6). On the other hand, minimum spontaneous release of granzyme B (<20 pg/ml) was detected in the NDV treated or untreated PBMC that were not co-cultured with MCF-7. This result indicated that elevated granzyme B expression in NDV treated PBMC may stimulate the apoptosis in MCF-7 cell.

Bottom Line: Interestingly, the percentage of cells with activating markers CD16 and CD56 were increased significantly.Human PBMC treated with NDV titer 8 HAU was found to stimulate the highest level of cytokine production including interferon-γ, interleukin-2 and interleukin-12.The release of these proteins contributes to the antitumor effect of PBMC against MCF-7 breast cancer cells.

View Article: PubMed Central - PubMed

Affiliation: 1. Faculty of Biotechnology and Biomolecular Sciences, Universiti Putra Malaysia, 43400, Serdang, Selangor, Malaysia.

ABSTRACT
Immunotherapy has raised the attention of many scientists because it hold promise to be an attractive therapeutic strategy to treat a number of disorders. In this study, the immunomodulatory effects of low titers of Newcastle disease virus (NDV) AF2240 on human peripheral blood mononuclear cells (PBMC) were analyzed. We evaluated cytokine secretion and PBMC activation by cell proliferation assay, immunophenotyping and enzyme linked immunosorbent assay. The proliferation of the human PBMC was measured to be 28.5% and 36.5% upon treatment with 8 hemaglutinin unit (HAU) and 2 HAU of NDV respectively. Interestingly, the percentage of cells with activating markers CD16 and CD56 were increased significantly. Furthermore, the intracellular perforin and granzyme levels were also increased upon virus infection. Human PBMC treated with NDV titer 8 HAU was found to stimulate the highest level of cytokine production including interferon-γ, interleukin-2 and interleukin-12. The release of these proteins contributes to the antitumor effect of PBMC against MCF-7 breast cancer cells. Based on the 3-(4, 5-dimethylthiazol-2-yl)-2, 5-diphenyl tetrazolium bromide assay, activated human PBMC showed high cytolytic efficiency towards human breast tumor cells. In summary, NDV was able to stimulate PBMC proliferation, cytokine secretion and cytolytic activity.

Show MeSH
Related in: MedlinePlus