Decreased nitric oxide content mediated by asymmetrical dimethylarginine and proteinl-arginine methyltransferase 3 in macrophages induces trophoblast apoptosis: a potential cause of recurrent miscarriage

Abstract

STUDY QUESTION Is the protein L-arginine methyltransferase 3 (PRMT3)/asymmetrical dimethylarginine (ADMA)/nitric oxide (NO) pathway involved in the development of recurrent miscarriage (RM), and what is the potential mechanism? SUMMARY ANSWER Elevated levels of PRMT3 and ADMA inhibit NO formation in the decidua, thereby impairing the functions of trophoblast cells at the maternal–foetal interface. WHAT IS KNOWN ALREADY Decreased NO bioavailability is associated with RM. ADMA, an endogenous inhibitor of nitric oxide synthase (NOS), is derived from the methylation of protein arginine residues by PRMTs and serves as a predictor of mortality in critical illness. STUDY DESIGN, SIZE, DURATION A total of 145 women with RM and 149 healthy women undergoing elective termination of an early normal pregnancy were enrolled. Ninety-six female CBA/J, 24 male DBA/2 and 24 male BALB/c mice were included. CBA/J × DBA/2 matings represent the abortion group, while CBA/J × BALB/c matings represent the normal control group. The CBA/J pregnant mice were then categorised into four groups: (i) normal + vehicle group (n = 28), (ii) abortion + vehicle group (n = 28), (iii) normal + SGC707 (a PRMT3 inhibitor) group (n = 20) and (iv) abortion + SGC707 group (n = 20). All injections were made intraperitoneally on Days 0.5, 3.5 and 6.5 of pregnancy. Decidual tissues were collected on Days 8.5, 9.5 and 10.5 of gestation. The embryo resorption rates were calculated on Day 9.5 and Day 10.5 of gestation. PARTICIPANTS/MATERIALS, SETTING, METHODS NO concentration, ADMA content, NOS activity, expression levels of NOS and PRMTs in decidual tissues were determined using conventional assay kits or western blotting. PRMT3 expression was further analysed in decidual stromal cells, macrophages and natural killer cells. A co-culture system between decidual macrophages (DMs) and HTR-8/SVneo trophoblasts was constructed to study the roles of the PRMT3/ADMA/NO signalling pathway. Trophoblast apoptosis was analysed via Annexin V-fluorescein isothiocyanate/propidium iodide staining. CBA/J × DBA/2 mouse models were used to investigate the effects of SGC707 on embryo resorption rates. MAIN RESULTS AND THE ROLE OF CHANCE Our results show that NO concentration and NOS activity were decreased, but ADMA content and PRMT3 expression were increased in the decidua of RM patients. Moreover, compared with the normal control subjects, PRMT3 expression was significantly up-regulated in the macrophages but not in the natural killer cells or stromal cells of the decidua from RM patients. The inhibition of PRMT3 results in a significant decrease in ADMA accumulation and an increase in NO concentration in macrophages. When co-cultured with DMs, which were treated with SGC707 and ADMA, trophoblast apoptosis was suppressed and induced, respectively. In vivo experiments revealed that the administration of SGC707 reduced the embryo resorption rate of CBA/J × DBA/2 mice. LIMITATIONS, REASONS FOR CAUTION All sets of experiments were not performed with the same samples. The main reason is that each tissue needs to be reserved for clinical diagnosis and only a small piece of each tissue can be cut and collected for this study. WIDER IMPLICATIONS OF THE FINDINGS Our results indicate that the PRMT3/ADMA/NO pathway is a potential marker and target for the clinical diagnosis and therapy of RM. STUDY FUNDING/COMPETING INTEREST(S) This study was supported by the National Key Research and Development Program of China (2017YFC1001401), National Natural Science Foundation of China (81730039, 82071653, 81671460, 81971384 and 82171657) and Shanghai Municipal Medical and Health Discipline Construction Projects (2017ZZ02015). The authors have declared no conflict of interest. TRIAL REGISTRATION NUMBER N/A.

Endometrial stromal PRMT5 plays a crucial role in decidualization by regulating NF-κB signaling in endometriosis

“…Recently, some studies have reported increased expression of PRMT1 and PRMT3, two type I enzymes, in the endometrium of LPS-induced endometritis rats and the decidua of recurrent miscarriage patients, respectively [ 20 , 30 ]. By screening the expression levels of various PRMT mRNAs from several published gene expression profiles of endometriosis (GSE23339, GSE5108 and GSE7305), we observed that PRMT5 , a major type II enzyme, was decreased in the ectopic endometrium of endometriosis patients compared to the eutopic endometrium of healthy controls.…”

“…Depending on catalytic activity, PRMTs can be classified as a type I enzyme that catalyzes aDMA (PRMT1, PRMT2, PRMT3, PRMT4, PRMT6 and PRMT8) or a type II enzyme that generates sDMA (PRMT5, PRMT7 and PRMT9) [ 19 ]. A recent study found that aDMA content and PRMT3 expression were increased in the decidua of recurrent miscarriage patients, primarily in macrophages but not in natural killer cells or stromal cells of the decidua [ 20 ]. However, the localization and function of PRMT5, the main type II PRMT, is not clear in the endometrium.…”

Endometrial stromal PRMT5 plays a crucial role in decidualization by regulating NF-κB signaling in endometriosis

“…Recently, some studies have reported increased expression of PRMT1 and PRMT3, two type I enzymes, in the endometrium of LPS-induced endometritis rats and the decidua of recurrent miscarriage patients, respectively 20,32 . By screening the expression levels of various PRMT mRNAs from several published gene expression pro les of endometriosis (GSE23339, GSE5108 and GSE7305), we observed that PRMT5, a major type II enzyme, was sed in the ectopic endometrium of endometriosis patients compared to the eutopic endometrium of healthy controls.…”

“…Depending on catalytic activity, PRMTs can be classi ed as a type I enzyme that catalyzes aDMA (PRMT1, PRMT2, PRMT3, PRMT4, PRMT6 and PRMT8) or a type II enzyme that generates sDMA (PRMT5, PRMT7 and PRMT9) 19 . A recent study found that aDMA content and PRMT3 expression were increased in the decidua of recurrent miscarriage patients, primarily in macrophages but not in natural killer cells or stromal cells of the decidua 20 . However, the localization and function of PRMT5, the main type II PRMT, is not clear in the endometrium.…”

Tumor cell-derived asymmetric dimethylarginine regulates macrophage functions and polarization

“…Only a small proportion of ADMA is normally released into the circulation and excreted in urine. Interestingly, ADMA inhibits NOS activity, preventing nitric oxide production in macrophages in many animal models of disease [23][24][25]. However, unknown is whether inhibition of NOS by ADMA encourages the production of M2 macrophages.…”