Protection of LPS-induced Intercellular Hyperpermeability in Caco-2 Cells by Polypeptide from Moringa oleifera
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Abstract:
The protective effect of polypeptide from Moringa oleifera (MOPP) on lipopolysaccharide (LPS, 2 μg/mL) induced hypepermeability in Caco-2 cells was investigated. The cell survival rate was measured by MTT assay. The cellular level of lactate dehydrogenase (LDH) was determined by kit. Enzyme linked immunosorbent assay (ELISA) was used to determine the level of Interleukin (IL)-1β, IL-8, and tumor necrosis factor (TNF)-α. The transepithelialelectricalresistance (TEER) and fluorescein dextran (FD40) permeability were used to evaluate the permeability of cells. Quantitative real-time PCR (qRT-PCR) was used to detect the expression of IL-1β, IL-8, TNF-α, Occludin, claudin-1, ZO-1, and myosin light chain kinase (MLCK). A high concentration of MOPP (150 μg/mL) treatment significantly increased the cell survival rate (to 83.8%) and inhibited the spillover of LDH (50.3%) compared with that in model cells. MOPP treatment effectively inhibited the secretion of inflammatory cytokines (IL-1β: 19.8%, IL-8: 43.7% and TNF-α: 37.9%) and reduced their mRNA transcription (IL-1β: 44.3%，IL-8: 35.0% and TNF-α: 33.5%)in model cells. In addition, MOPP also significantly increased the mRNA levels of Occludin, claudin-1, ZO-1, and inhibit the 31.3% of MLCK mRNA transcription to improve the intercellular permeability in LPS treated Caco-2 cells. These results suggest that the MOPP exhibited a strong anti-inflammatory activity, and can improve the LPS induced high permeability of Caco-2 cells may associate with regulating the mRNA transcription of tight junction related factors.
