Dual Effects of Cell Free Supernatants from Lactobacillus acidophilus and Lactobacillus rhamnosus GG in Regulation of MMP-9 by Up-Regulating TIMP-1 and Down-Regulating CD147 in PMADifferentiated THP-1 Cells.

Figures

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  Fig.1. Undifferentiated and PMA-differentiated THP-1 cells and the effect of lactobacilli supernatant on cell viability. A. Undifferentiated THP- 1 cells with round phenotype, B. Induction of monocyte-macrophage differentiation by PMA (50 ng/ml) after 24 hours accompanied by the adherence of cells to the surface of plates with amoeboid-like phenotype, and C. Effects of different concentrations of LAS, LRS, MRL and MRS (0, 2, 5, 10, 15, 20 and 50 % v/v) on the viability of PMA-differentiated THP-1 cells were determined using MTT assay. LAS; Lactobacillus acidophilus supernatant, LRS; Lactobacillus rhamnosus GG supernatant, MRS; De Man Rogosa Sharpe, MRL; MRS with lactic acid, MTT; 3-(4,5-Dimethylthiazol-2-yl)-2,5-diphenyltetrazolium bromide, and PMA; Phorbol myristate acetate.

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  Fig.2. Effects of lactobacilli supernatant on expression of MMP-2 and MMP-9 mRNAs. A. Effects of LAS, LRS, MRL and MRS on expression of MMP-2 mRNAs and B. Effects of LAS, LRS, MRL and MRS on expression of MMP-9 mRNAs in PMA-differentiated THP-1 cells. Relative quantification of gene expression was performed by the 2−ΔΔCt method. The results showed the means ± SEM from six independent experiments. Significant difference was compared with positive control: *; P<0.05, **; P<0.01, ***; P<0.001, ****; P<0.0001. NC; Negative control (undifferentiated THP-1 cells), PC; Positive control (PMA-differentiated THP-1 cells), LAS; Lactobacillus acidophilus supernatant, LRS; Lactobacillus rhamnosus GG supernatant, MRS; De Man Rogosa Sharpe, MRL; MRS with lactic acid, and MMPs; Matrix metalloproteinases.

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  Fig.3. Effects of lactobacilli supernatant on expression of TIMP-1 and TIMP- 2 mRNAs. A. Effects of LAS, LRS, MRL and MRS on expression of TIMP-1 mRNAs and B. Effects of LAS, LRS, MRL and MRS on expression of TIMP-2 mRNAs in PMA-differentiated THP-1 cells. Relative quantification of gene expression was performed by the 2−ΔΔCt method. The results showed the means ± SEM from six independent experiments. Significant difference was compared with positive control: ***; P<0.001, ****; P<0.0001. NC; Negative control (undifferentiated THP-1 cells), PC; Positive control (PMA-differentiated THP-1 cells), LAS; Lactobacillus acidophilus supernatant, LRS; Lactobacillus rhamnosus GG supernatant, MRS; De Man Rogosa Sharpe, MRL; MRS with lactic acid, and MMPs; Matrix metalloproteinases.

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  Fig.4. Effects of LAS, LRS, MRL and MRS on gelatinolytic activities of MMP- 2 and MMP-9. A. Representative gelatin zymography for MMP-2 and MMP-9, B. Relative activity of MMP-2, and C. Relative activity of MMP-9. The results showed the means ± SEM of three independent experiments. Significant difference was compared with positive control: ***; P<0.001, ****; P<0.0001. NC; Negative control (undifferentiated THP-1 cells), PC; Positive control (PMA-differentiated THP-1 cells), LAS; Lactobacillus acidophilus supernatant, LRS; Lactobacillus rhamnosus GG supernatant, MRS; De Man Rogosa Sharpe, and MRL; MRS with lactic acid, and MMPs; Matrix metalloproteinases.

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  Fig.5. Effects of LAS, LRS, MRL and MRS on expression of CD147 protein in PMAdifferentiated THP-1 cells. Cell surface expression of CD147 was quantified after treatment with phycoerythrin-labeled mouse anti-human CD147 antibodies. A. Representative PMA-differentiated THP-1 flow cytometry scatter diagram for cells, B. NC (undifferentiated THP-1 cells), C. PC (PMA-differentiated THP-1 cells), D. LAS, E. LRS, F. MRS, G. MRL, and H. MFI of the six groups shown above. Data are represented as means± SE of 3 independent experiments. Significant difference was compared with positive control: *; P<0.05, **; P<0.01, ***; P<0.001. NC; Negative control, PMA; Phorbol myristate acetate, PC; Positive control, LAS; Lactobacillus acidophilus supernatant, LRS; Lactobacillus rhamnosus GG supernatant, MRS; De Man Rogosa Sharpe, MRL; MRS with lactic acid, and MFI; Mean fluorescence intensity.