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Mycoplasma genitalium lipoproteins induce human monocytic cell expression of proinflammatory cytokines and apoptosis by activating nuclear factor kappaB.

Wu Y, Qiu H, Zeng Y, You X, Deng Z, Yu M, Zhu C - Mediators Inflamm. (2008)

Bottom Line: LPs were also found to increase the DNA-binding activity of NF-kappaB, a possible mechanism for the induction of cytokine mRNA expression and the cell apoptosis.These effects were abrogated by PDTC, an inhibitor of NF-kappaB.Our results indicate that M. genitalium-derived LP may be an important etiological factor of certain diseases due to the ability of LP to produce proinflammatory cytokines and induction of apoptosis, which is probably mediated through the activation of NF-kappaB.

View Article: PubMed Central - PubMed

Affiliation: Pathogenic Biology Institute, University of South China, 421001 Hengyang, China. yimouwu@sina.com

ABSTRACT
This study was designed to investigate the molecular mechanisms responsible for the induction of proinflammatory cytokines gene expression and apoptosis in human monocytic cell line THP-1 stimulated by lipoproteins (LPs) prepared from Mycoplasma genitalium. Cultured cells were stimulated with M. genitalium LP to analyze the production of proinflammatory cytokines and expression of their mRNA by ELISA and RT-PCR, respectively. Cell apoptosis was also detected by Annexin V-FITC-propidium iodide (PI) staining and acridine orange (AO)-ethidium bromide (EB) staining. The DNA-binding activity of nuclear factor-kappaB (NF-kappaB) was assessed by electrophoretic mobility shift assay (EMSA). Results showed that LP stimulated THP-1 cells to produce tumor necrosis factor-alpha (TNF-alpha), interleukin-1beta (IL-1beta), and IL-6 in a dose-dependent manner. The mRNA levels were also upregulated in response to LP stimulation. LPs were also found to increase the DNA-binding activity of NF-kappaB, a possible mechanism for the induction of cytokine mRNA expression and the cell apoptosis. These effects were abrogated by PDTC, an inhibitor of NF-kappaB. Our results indicate that M. genitalium-derived LP may be an important etiological factor of certain diseases due to the ability of LP to produce proinflammatory cytokines and induction of apoptosis, which is probably mediated through the activation of NF-kappaB.

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The effect ofdifferent concentrations of LP on the production of TNF-α (a), IL-6 (b), andIL-1β (c) in THP-1 cells. Dose-dependent induction of TNF-α (a), IL-6 (b), and IL-1β (c) was seen in THP-1 cells after 24 hours of stimulation with 0.5 to 5 μg/mL of LP, 3 μg/mL of LP in combination with 25 μM PDTC, or 0.1 μg/mL LPS. In this experiment THP-1 cells stimulated with aqueous phase were used as the negative control. Proinflammatory cytokine levels were determined by ELISA as indicated in Section 2. The results were representative of three independent experiments.
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fig1: The effect ofdifferent concentrations of LP on the production of TNF-α (a), IL-6 (b), andIL-1β (c) in THP-1 cells. Dose-dependent induction of TNF-α (a), IL-6 (b), and IL-1β (c) was seen in THP-1 cells after 24 hours of stimulation with 0.5 to 5 μg/mL of LP, 3 μg/mL of LP in combination with 25 μM PDTC, or 0.1 μg/mL LPS. In this experiment THP-1 cells stimulated with aqueous phase were used as the negative control. Proinflammatory cytokine levels were determined by ELISA as indicated in Section 2. The results were representative of three independent experiments.

Mentions: LP prepared from M. genitalium stimulated THP-1 cells toproduce TNF-α, IL-1β, and IL-6 in dose-dependent manner (Figure 1). The dose-responsebar revealed a 3 μg/mL optimal concentration of LP for the induction of TNF-α [(1885.29 ± 58.62) pg/mL], IL-1β [(256.20 ± 16.030) pg/mL], and IL-6 [(35.29 ± 1.26) pg/mL], leading to cytokine concentrationsimilar to those obtained with 0.1 μg/mL LPS [TNF-α (1288.96 ± 34.34) pg/mL, IL-1β (469.36 ± 21.11) pg/mL, IL-6 (33.01 ± 1.81) pg/mL, P > .05].Interestingly, when the concentration of LP was increased from 3 μg/mL to 5 μg/mL,the level of cytokines decreased. The contaminationof LP by LPS was not responsible for this since polymyxin B pretreated LP hadno effect on TNF-α production (Figure 2).


Mycoplasma genitalium lipoproteins induce human monocytic cell expression of proinflammatory cytokines and apoptosis by activating nuclear factor kappaB.

Wu Y, Qiu H, Zeng Y, You X, Deng Z, Yu M, Zhu C - Mediators Inflamm. (2008)

The effect ofdifferent concentrations of LP on the production of TNF-α (a), IL-6 (b), andIL-1β (c) in THP-1 cells. Dose-dependent induction of TNF-α (a), IL-6 (b), and IL-1β (c) was seen in THP-1 cells after 24 hours of stimulation with 0.5 to 5 μg/mL of LP, 3 μg/mL of LP in combination with 25 μM PDTC, or 0.1 μg/mL LPS. In this experiment THP-1 cells stimulated with aqueous phase were used as the negative control. Proinflammatory cytokine levels were determined by ELISA as indicated in Section 2. The results were representative of three independent experiments.
© Copyright Policy - open-access
Related In: Results  -  Collection

Show All Figures
getmorefigures.php?uid=PMC2366083&req=5

fig1: The effect ofdifferent concentrations of LP on the production of TNF-α (a), IL-6 (b), andIL-1β (c) in THP-1 cells. Dose-dependent induction of TNF-α (a), IL-6 (b), and IL-1β (c) was seen in THP-1 cells after 24 hours of stimulation with 0.5 to 5 μg/mL of LP, 3 μg/mL of LP in combination with 25 μM PDTC, or 0.1 μg/mL LPS. In this experiment THP-1 cells stimulated with aqueous phase were used as the negative control. Proinflammatory cytokine levels were determined by ELISA as indicated in Section 2. The results were representative of three independent experiments.
Mentions: LP prepared from M. genitalium stimulated THP-1 cells toproduce TNF-α, IL-1β, and IL-6 in dose-dependent manner (Figure 1). The dose-responsebar revealed a 3 μg/mL optimal concentration of LP for the induction of TNF-α [(1885.29 ± 58.62) pg/mL], IL-1β [(256.20 ± 16.030) pg/mL], and IL-6 [(35.29 ± 1.26) pg/mL], leading to cytokine concentrationsimilar to those obtained with 0.1 μg/mL LPS [TNF-α (1288.96 ± 34.34) pg/mL, IL-1β (469.36 ± 21.11) pg/mL, IL-6 (33.01 ± 1.81) pg/mL, P > .05].Interestingly, when the concentration of LP was increased from 3 μg/mL to 5 μg/mL,the level of cytokines decreased. The contaminationof LP by LPS was not responsible for this since polymyxin B pretreated LP hadno effect on TNF-α production (Figure 2).

Bottom Line: LPs were also found to increase the DNA-binding activity of NF-kappaB, a possible mechanism for the induction of cytokine mRNA expression and the cell apoptosis.These effects were abrogated by PDTC, an inhibitor of NF-kappaB.Our results indicate that M. genitalium-derived LP may be an important etiological factor of certain diseases due to the ability of LP to produce proinflammatory cytokines and induction of apoptosis, which is probably mediated through the activation of NF-kappaB.

View Article: PubMed Central - PubMed

Affiliation: Pathogenic Biology Institute, University of South China, 421001 Hengyang, China. yimouwu@sina.com

ABSTRACT
This study was designed to investigate the molecular mechanisms responsible for the induction of proinflammatory cytokines gene expression and apoptosis in human monocytic cell line THP-1 stimulated by lipoproteins (LPs) prepared from Mycoplasma genitalium. Cultured cells were stimulated with M. genitalium LP to analyze the production of proinflammatory cytokines and expression of their mRNA by ELISA and RT-PCR, respectively. Cell apoptosis was also detected by Annexin V-FITC-propidium iodide (PI) staining and acridine orange (AO)-ethidium bromide (EB) staining. The DNA-binding activity of nuclear factor-kappaB (NF-kappaB) was assessed by electrophoretic mobility shift assay (EMSA). Results showed that LP stimulated THP-1 cells to produce tumor necrosis factor-alpha (TNF-alpha), interleukin-1beta (IL-1beta), and IL-6 in a dose-dependent manner. The mRNA levels were also upregulated in response to LP stimulation. LPs were also found to increase the DNA-binding activity of NF-kappaB, a possible mechanism for the induction of cytokine mRNA expression and the cell apoptosis. These effects were abrogated by PDTC, an inhibitor of NF-kappaB. Our results indicate that M. genitalium-derived LP may be an important etiological factor of certain diseases due to the ability of LP to produce proinflammatory cytokines and induction of apoptosis, which is probably mediated through the activation of NF-kappaB.

Show MeSH
Related in: MedlinePlus