Studies On The Mechanisms Of Sesamin Protecting The Acute Lung Injury By Inducing Transdifferentiation Of Alveolar Epithelial Type Ⅱ Cells

Acute lung injury(ALI)or acute respiratory distress syndrome(ARDS)refers to the acute progressive respiratory failure caused by various pathogenic factors other than cardiogenesis,and the damage of type Ⅰ alveolar epithelial cells is an important cause of ALI.In the stage of injury repair,type Ⅱ alveolar epithelial cells(ATⅡ)are transdifferentiated into type Ⅰ alveolar epithelial cells(ATⅠ).which can make up for the loss of ATⅠ cells in the stage of lung injury,and is essential for rebuilding and maintaining the complete structure and function of alveolar epithelial cells.At present,there are few reports on tools that can effectively promote the transformation of ATⅡ to ATⅠ,and the molecular mechanism of this transformation needs to be further explored.Sesamin is a lignan compound existing in many plants,which has antioxidant and anti-inflammatory effects,and also has good effects in treating cardiovascular diseases and cancers.It has been reported that sesamin can reduce the level of cytokines and inhibit lung injury in mice.However,the role of sesamin in the transdifferentiation of alveolar epithelial cells has not been reported.The purpose of this study is to explore the role of sesamin in the transdifferentiation from ATⅡ to ATⅠ and its related mechanism,so as to lay a foundation for the adjuvant treatment of ALI with sesamin.In this study,two acute lung injury models were successfully established by intraperitoneal and tracheal injection of LPS in mice.The lung tissue sections combined with HE staining and the detection of inflammatory factor mRNA level showed that sesamin treatment group significantly improved the lung edema,alveolar septum thickening and inflammatory cell infiltration caused by LPS,and reduced the level of inflammatory factors,which verified the protective effect of sesamin on acute lung injury.There are damaged and exfoliated ATⅠ cells in alveolar lavage fluid(BALF).In order to explore whether the protective effect of sesamin on lung injury is related to ATⅠ,we used the mouse model of LPS intratracheal injection.By collecting BALF and detecting the expression level of AT1 marker AQP5,it was found that the AQP5 level in BALF treated with sesamin was lower than that in LPS group.However,the results of immunofluorescence staining of AQP5 in lung tissue sections of mice showed that the AQP5 level in lung tissue of sesamin-treated mice increased.These results indicate that sesamin can protect the injury of type Ⅰ alveolar epithelial cells induced by LPS and maintain the number of ATⅠ cells.ATⅠ cells cannot proliferate,but ATⅡ cells can proliferate and differentiate into ATⅠcells to supplement damaged ATⅠ cells.Therefore,the number of ATⅠ cells is not only related to the death caused by its injury,but also affected by the proliferation and transdifferentiation of ATⅡ cells.To explore the reason why sesamin maintains the number of ATⅠ cells,we used type Ⅱ alveolar epithelial cell line A549 to study the effect of sesamin on its apoptosis,proliferation and transdifferentiation.The results showed that low concentration of sesamin had no effect on proliferation and apoptosis of A549 cells,while high concentration of sesamin inhibited cell proliferation and promoted cell apoptosis.Contrary to the results in vivo,it may not be the reason why sesamin maintained the number of AT1 cells.However,the study of its transdifferentiation found that sesamin promoted A549 cells from irregular scales to elongated ATⅠ cells,and the mRNA AQP5,RAGE,PDPN and protein expression of type Ⅰ alveolar epithelial cell markers increased.It is suggested that the protective effect of sesamin on acute lung injury is related to its promotion of the transdifferentiation from ATⅡto ATⅠ.Next,we will further explore the molecular mechanism of sesamin-induced transdifferentiation of ATⅡ cells.As AKT pathway is widely involved in the action process of sesamin,in order to further explore whether AKT is involved in the transdifferentiation process of ATⅡ,the expression level of P-AKT was detected,and the results showed that sesamin treatment decreased the activity of P-AKT S473.Pretreatment with AKT inhibitor LY294002 further promoted the transdifferentiation from ATⅡ to ATⅠ.AnnexinA1 and TRPV1 are two cell receptors on which sesamin functions.In order to explore whether they are the upstream proteins of AKT,it was found that the inhibition of P-AKT S473 by sesamin was reversed by using si-RNA transfection instead of AnnexinA1,which indicated that sesamin participated in the transdifferentiation process by regulating AKT phosphorylation through TRPV1 instead of AnnexinA1.Next,we studied the role of AnnexinA1 and TRPV1 in transdifferentiation.The si-RNA of AnnexinA1 and TRPV1 both reversed the transformation of ATⅡ to ATⅠ promoted by sesamin,which indicated that AnnexinAl and TRPV1 were the key factors of sesamin promoting A549 cell transformation.In this paper,sesamin protects acute lung injury induced by LPS by protecting type Ⅰepithelial cells in mice,and the function of sesamin inducing the transdifferentiation from ATⅡ to ATⅠ was found in vitro.This study provides a reference for the development of sesamin as a therapeutic drug and health product for acute lung injury,and provides a new tool and effective target for the study of alveolar type Ⅱ epithelial cell trans-differentiation.