High-mannose glycosylation of ITGAM regulates the development and differentiation of trophoblast in the placenta

Abstract

Background Development of placenta and differentiation of trophoblast is a hallmark event for successful pregnancy. Trophoblast stem (TS) cells proliferate and differentiate into TS-like cytotrophoblasts (CTBs), further formulate the differentiated subtypes, syncytiotrophoblasts (STBs) and invasive extravillous tropholoblasts (EVTs). Defective differentiation of TS-like CTBs is associated with severe gestational diseases. Protein glycosylation is an essential form of posttranslational modification. However, glycosylation-related mechanism controlling TS-like CTBs differentiation remains unknown. This study aims to investigate the function of high-mannose type glycans and ALG3 on TS-like CTBs differentiation. Furthermore, the mechanism of high-mannose and ITGAM during TS-like CTBs differentiation were explored.Methods Employing lectin microarray, the glycosylation expression traits were compared in the villi of miscarriage patients and healthy women. The expression of high-mannose and ALG3 were investigated by immunoblotting and immunofluorescence assays. The glycosylation proteins were screened by pull down and LC-MS/MS detection. Signaling pathway were screened by the human phosphokinase antibody array. The differentiation of TS-like CTBs were measured by immunoblotting and immunofluorescence assays.Results Lectin microarray results revealed that increased level of high-mannose type glycans on the TS-like CTBs of miscarriage patients compared with normal pregnancy women. Meantime, ALG3 levels increased in TS-like CTBs of miscarriage patients. Upregulating high-mannose type glycans by ALG3 hampered TS-like CTBs differentiated into STBs and EVTs, and arrested TS-like CTBs in the property stage. Furthermore, high level of high-mannose type glycans on ITGAM inhibited the binding of ITGAM and Fn, inactivating the p-STAT1 signaling pathway, further inhibiting TS-like CTBs differentiation potential.Conclusions These findings reveal that high-mannose type glycans, especially on ITGAM, hampered binding of ITGAM and Fn, which leads to the impaired TS-like CTBs differentiation by p-STAT1 signaling pathway. The present study provides novel insight into the function and mechanism of α1,3-linked high-mannose type glycans in TS-like CTBs differentiation in human placenta, which can also be used as a glycol molecular target for the treatment of miscarriage.