Expression of GRIM-19 in unexplained recurrent spontaneous abortion and possible pathogenesis.

STUDY QUESTION: Is aberrant expression of gene associated with retinoid-interferon-induced mortality-19 (GRIM-19) associated with unexplained recurrent spontaneous abortion (URSA)?

SUMMARY ANSWER: GRIM-19 deficiency may regulate regulatory T cell/T helper 17 cell (Treg/Th17) balance partly through reactive oxygen species (ROS)-mammalian target of rapamycin (mTOR) signaling axis in URSA.

WHAT IS KNOWN ALREADY: Immunological disorders may cause impaired maternal immune tolerance to the fetus and result in fetal rejection. The differentiation of Treg and Th17 cells is controlled by phosphoinositide 3-kinase (PI3K)/Akt/mTOR signaling pathway. GRIM-19 participates in the immune response, but its role in URSA is largely unknown.

STUDY DESIGN, SIZE, DURATION: The current study included 28 URSA patients and 30 non-pregnant healthy women.

PARTICIPANTS/MATERIALS, SETTING, METHODS: The proportion of Treg and Th17 cells in peripheral blood of URSA patients and control subjects were assessed with flow cytometry. The expression of GRIM-19 in peripheral blood lymphocytes (PBLs) was measured with quantitative real-time PCR and western blot analysis. Furthermore, the ROS level in the PBLs of URSA patients and control subjects were assessed by 2',7'-dichlorodihydrofluorescein diacetate (DCFH-DA) staining. Then, Akt/mTOR expression in the PBLs was measured. Downregulation of GRIM-19 in Jurkat cells was performed by specific siRNA. Then, intracellular ROS production and the expression of p-mTOR, which is known to enhance Th17 differentiation and decrease Treg cell differentiation, were detected. Finally, N-acetylcysteine (NAC) was used to decrease the intracellular ROS level, and the expression of p-mTOR was measured.

MAIN RESULTS AND THE ROLE OF CHANCE: The proportion of Treg cells was reduced in URSA patients, whereas the proportion of Th17 cells was increased. The expression of GRIM-19 was significantly lower in PBLs of URSA patients. Furthermore, there is a considerable increase in intracellular ROS production and a high level of p-Akt and p-mTOR expression in the PBLs of URSA patients compared with the control subjects. In parallel to this, downregulation of GRIM-19 in the Jurkat cells by siRNA results in an increased ROS production and an increased expression of p-mTOR. Importantly, the upregulation of p-mTOR resulting from GRIM-19 loss was significantly reversed in the cells treatment with ROS inhibitor N-acetyl-l-cysteine (NAC), indicating that ROS was indeed required for GRIM-19 depletion induced p-mTOR expression.

LARGE SCALE DATA: None.

LIMITATIONS, REASONS FOR CAUTION: A large number of researches have confirmed that the differentiation of Treg and Th17 cells is controlled by PI3K/Akt/mTOR signaling pathway. We have not shown the regulatory role of ROS and PI3K/Akt/mTOR in Treg and Th17 differentiation in this study.

WIDER IMPLICATIONS OF THE FINDINGS: Our study has demonstrated that GRIM-19 deficiency may play a role in regulating Treg/Th17 balance partly through ROS-mTOR signaling axis in URSA. The present study offers a new perspective to the roles of GRIM-19 in immunoregulation.

STUDY FUNDING AND COMPETING INTEREST(S): This work was supported by the National Natural Science Foundation of China (Grant numbers 81571511, 81701528, 81370711 and 30901603), the Shandong Provincial Natural Science Foundation (Grant numbers ZR2017PH052 and ZR2013HM090) and the Science Foundation of Qilu Hospital of Shandong University, Fundamental Research Funds of Shandong University (Grant numbers 2015QLQN50 and 2015QLMS24). The authors declare that there is no conflict of interest that could prejudice the impartiality of the present research.