| Mastitis, a disease of high incidence occurred among the postpartum women, is an acute purulent infection of the breast. It threatened the health of parturient and newborn. One of the major causes of mastitis is bacteria invasion. Mammary epithelial cells play a significant role in the process of bacteria invasion, for they act as the first defensive barrier against bacteria. There has been shown that the endotoxin firstly release from its intracellular after bacteria invasion and then bind with the Toll-like receptor family of the mammary epithelial cell membrane, activate downstream pathways, result in the proinflammatory cytokine release. Proinflammatory cytokine released into the blood can recruit leukocytes to the patient position to kill bacteria, while the massive release of proinflammatory cytokine can lead to excessive inflammatory response, directly or indirectly cause tissue and cell damage, and even result in the systemic inflammatory response.Caveolin-1(Cav-1) is the marker protein of Caveolae, which can participate in a variety of intracellular activities, such as cholesterol transport, membrane assembly, signal transduction, cell transformation and tumor formation. Our laboratory preliminary work have shown that Cav-1 can bind with ERαand Cav-1 gene silencing activates the PI3K/AKT and MEK/ERK signaling pathway to mediate mammary epithelia cell transformation. Recent studies have revealed that Cav-1 could take interaction with Toll-like receptor 4(TLR4), thereby inhibiting the inflammatory response. However, it is still not clear whether TLR4 could take part in the activation of pathways involved in the process of inflammatory.By using DNA microarrays, Western blot and immunofluorescence techniologies we observed the co-localization between Cav-1 and TLR4 and examined the effect of Cav-1 to the inflammation-related genes and the pathways of inflammatory response in human breast epithelial cell. We are aiming to investigate the mechanism of inflammatory pathway, and to explore whether Cav-1 could inhibit the pro-inflammation response induced by LPS by affecting TLR4 expression and the function of its downstream pathway in human mammary epithelial cells.The results are stated as follows:1. We successfully establish a stable low expression cell line MCF10ACE by using Gene capture and siRNA technologies. Using the DNA super microarray technology to comparatively analyze the different expression of inflammatory-related genes between MCF10A and MCF10ACE, we found that ten inflammatory-related genes expression were up-regulated when Cav-1 gene expressed at a low level. By using immunofluoescence technology we found Cav-1 co-localized TLR4, and their expression were negatively correlative in the MCF10A and MCF10ACE cells.4. By taking LPS stimulation method, our laboratory successfully established human breast epithelial pro-inflammatory response cell model, and found that LPS stimulation could activate JNK and P38 MAPK signaling pathways. When Cav-1 expressed at a low level, The stimulation of JNK and P38 MAPK signaling pathways were enhanced. Conclusion:In human breast epithelial cells, Cav-1 may play a significant role as a potential target to regulate TLR4-mediated pro-inflammatory response. Cav-1 co-located with TLR4 indicated that Cav-1 may down-regulated downstream signaling pathways such as JNK and P38 MAPK signaling pathways by means of influencing TLR4 expression and function and ultimately inhibit inflammatory response in human breast epithelial cell. |
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