Panoply™ Human CYP11A1 Over-expressing Stable Cell Line

During normal pregnancy, placental trophoblast cells secrete various steroid hormones that regulate maternal physiological functions and fetal development. The CYP11A1 gene encodes the cholesterol side-chain cleavage enzyme P450scc, which catalyzes the conversion of cholesterol into pregnenolone-the initial step in the synthesis of all steroid hormones. Under the influence of genetic predisposition and specific environmental factors such as drugs or toxins, CYP11A1 expression may be upregulated; this alters steroid metabolism and physiological functions within trophoblast cells and impacts fetal development. Here, researchers demonstrate that the upregulation of CYP11A1 in the BeWo cell line induces excessive mitochondrial oxidative stress, leading to mitochondrial damage and the release of interleukin-6 (IL-6), while also causing proliferation inhibition and DNA damage in neural stem cells (NSCs). Furthermore, vitamin D3 alleviates oxidative stress and inflammatory responses in a dose-dependent manner, thereby helping to rescue damaged neural stem cells.

Mitochondrial apoptosis is frequently accompanied by the generation of reactive oxygen species (ROS) and mitochondrial DNA (mtDNA) damage, and can be assessed by measuring mitochondrial membrane potential. JC-1 staining results showed that, compared with the vehicle and blank control groups, CYP11A1-overexpressing cells exhibited significantly reduced levels of JC-1 aggregates (red fluorescence) and significantly increased levels of JC-1 monomers (green fluorescence), indicating an alteration (depolarization) in mitochondrial membrane potential. Treatment with 5 μM vitamin D3 reversed this trend toward apoptosis (Figure 1A). To evaluate the impact of impaired mitochondrial function on the trophoblast cells themselves, the researchers measured the levels of a panel of inflammatory cytokines (including IL-1β, IL-4, IL-6, IL-8, IL-10, IL-13, IL-17α, IL-23, and TNF-α) in BeWo cells. The results showed that only IL-6 was detected in the cells. Subsequently, the researchers further verified IL-6 production using qPCR and ELISA. The results confirmed an alteration in the inflammatory state of the BeWo cells; specifically, both the expression and release levels of IL-6 were significantly higher in CYP11A1-overexpressing cells compared with the vehicle or blank control groups. Furthermore, the addition of vitamin D3 caused inflammation levels to gradually return to the normal range (Figure 1B, C).

Figure 1. Trophoblast mitochondrial apoptosis and inflammation induced by upregulation of CYP11A1 could be ameliorated by vitamin D3 addition. (Wang X, et al., 2021)