| Objective:This study aim to investigate the effect of high/low molecular weight hyaluronan on lymphatic vessel barrier function and lymphangiogenesis. To explore the role of sphingosine 1-phosphate receptor in different molecular weight HA regulated lymphatic vessel barrier function and lymphangiogenesis.Methods:1. The effect of HMW-HA/LMW-HA on the permeability and integrity of lymphatic endothelial cell monolayer was determined by fluorescein sodium permeability experiment and immunofluorescence staining. 2. Lymphangiogenesis induced by HMW-HA/LMW-HA were obeserved through cell proliferattion,migration and tube formation. 3. The localization and combination of LYVE-1 and S1P3 receptor in LEC were detected by immunofluorescence staining and immunoprecipitation. 4. Silenced of LYVE-1 and S1P3 receptor to investigate their roles in HMW-HA/LMW-HA-induced lymphatic vessel barrier function and lymphangiogenesis.Results:1. LMW-HA produced a significant increase of Na-F permeability in LEC monolayer, and opened intercellular junction and promoted actin stress fiber formation. On the contrary, HMW-HA protected lymphatic barrier function. 2.LMW-HA promoted lymphangiogenesis and activated the downstream Src/ERK1/2signaling pathways. 3. LMW-HA/HMW-HA increased the expression of S1P3 receptor and S1P1 receptor, respectively, si RNA down-regulted S1P1 receptor blocked the effect of HMW-HA on cell-cell junction, si RNA down-regulted S1P3 receptor inhibited the disruption of the junction integrity caused by LMW-HA. 4.LYVE-1 and S1P3 receptor is a pair of "partner", LMW-HA promote S1P3 receptor and LYVE-1 combine with each other. Silencing of either LYVE-1 or S1P3 receptor significantly inhibited LMW-HA-induced lymphangiogenesis.Conclusions : LMW-HA could bind to its receptor LYVE-1 disrupted lymphatic vessel barrier function and promoted lymphangiogenesis through S1P3 receptor.HMW-HA bind to LYVE-1 protected lymphatic vessel barrier function through S1P1 receptor. |
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