09:45-10:00 Down regulation of CD24 in preeclampsia and its In-Vitro STOX1 model - a new target for immunotherapy of the syndrome?

During pregnancy, an immune shield is created between the fetus and the mother to suppress fetal and placental rejection. CD24, a 30-70 kDa GPI- anchored and highly glycosylated cell surface protein, which can bind to P-selectin and Siglec-10 on haematopoitic cells. The CD24-Siglec-10 axix mediates immune tolerance, and, in association with signal transduction of DAMP, immunoregulation and anti-inflammatory effects. In solid tumours, the high expression of CD24 correlates with bad prognosis which can be explained by “ Don’t eat me” signals mediated by CD24-Siglec-10 axis [1]. First trimester CD24-Siglec-10 co-expression in close proximity to extravillous trophoblasts and endometrial glands, induces an immune suppression rendering the decidual invasion by the trophoblasts possible, as well as the blastocyst implantation in the uterine wall, thus avoiding maternal rejection of the pregnancy [2]. Previously, we explored the potential involvement of CD24 in immune protection. Initially, we have found that during the first trimester, CD24 is expressed in the maternal uterine glands, the villous and extravillous cytotrophoblasts, with a milder expression in stromal cells. While placental CD24 mRNA expression was increased from the first trimester to normal term controls by 2.27fold, a decreased expression was found in early preeclampsia cases as compared with unrelated preterm control and term deliveries. Lower labeling of the syncytiotrophoblasts was found in early preeclampsia compared to preterm and term control [3]. Altogether, a link between down regulation of CD24 and preeclampsia was implied. To gain more mechanistical insight into the CD24 link to preeclampsia, we studied its expression in cell lines trophoblast models reproducing some features of preeclampsia linked to over expression of the STOX-1 transcription factor, under its two major isoforms, STOX1A and STOX1B, that share the same gene target but have often opposite transcriptional effects [4]. Using qPCR, Western-Blot and flow cytometry, we studied CD24 in BeWo and JEG-3 cell lines stably overexpressing either of the two isoforms of the STOX1 transcription factor STOX 1A or STOX1B versus mock control CD24 gene expression was significantly down regulated by almost 90% in BeWo and ~70% in JEG-3 cells compared to the mock control by the expression or either STOX1A or STOX1B. Similar reduction level of total cellular CD24 protein was determined by Western Blotting in this in-vitro model. Flow cytometry analysis showed a somewhat lower level of reduction of cell surface expression of CD24, mainly in BeWo cells and less in JEG-3 cells. In all cases STOX1A was more potent in CD24 reduction than STOX1B isoform. In sum, using either actual preeclampsia placental samples or cell models mimicking some aspects of preeclampsia, an overall suppression of CD24 RNA and protein expression was demonstrated. Given the known link between CD24-Siglec-10 complex and immune suppression, we may hypothesize that CD24 reduction reflects a loss of immune tolerance in preeclampsia. Thus, attempts to elevate CD24 might be a novel innate immune checkpoint and a potential novel target for a possible immunotherapy of preeclampsia, a pathology where inflammation and cytokine play an important role, reminding the possible ways of alleviating the cytokine storm in COVID-19 infection.