Pregnenolone enhanced proliferation of neural stem cells and reduced Oxidant and inflammatory marker after lipopolysaccharide induction of in vitro inflammation

Background: Pregnenolone as a precursor of steroid hormones, can be effect on inflammatory condition to maintain immune homeostasis. The previous study shows that pregnenolone levels are reduced in neuro inflammatory disease, which emphasizes its role in neuroprotection and nerve regeneration and its anti-inflammatory role  So the aim of this study try to investigated effect of pregnenolone on inflammatory marker and oxidant agent after induction of inflammatory lipopolysaccharide in the neural stem cells (NSCs) media. Methods: NSCs were isolated from the embryonic cortex of E14 mice with standard protocol and incubated for 5 days. And then after neurosphere formation and propagation for second passage the survival of the cells was done after pregnenolone combined treatment with lipopolysaccharide (LPS) inflammatory model. The number of neurospheres and cells derived from neurospheres were counted after 5 days of incubation in the inflammatory model. The supernatant of the cells was removed and the levels of oxidant and antioxidant markers MDA, NO FRAP, and inflammatory markers IL6 and TNFα were measured by ELISA method. Data were analyzed by one-way variance statistical method and Tukey's post hoc test. Results: in the inflammatory model, the survival of neural stem cells by LPS showed a significant decrease compared to the control group, while pregnenolone caused a significant increase of these cells compared to the control group, and the concentration of 10 μM showed the highest survival, also in Combined treatment of LPS and pregnenolone with a concentration of 10 μM, the number of neurospheres and cells derived from it increased significantly compared to the LPS group. Pregnenolone significantly reduced the amount of NO resulting from inflammation induced by lipopolysaccharide, in addition, a significant decrease in the amount of NO was observed in the combined treatment of pregnenolone and LPS in different concentrations compared to the LPS group. On the other hand, this neurosteroid alone as well as in combination with LPS caused a significant decrease in the level of IL-6 compared to the LPS group. And the treatment with 15 μM concentration of pregnenolone significantly reduced the amount of TNF-α. Conclusion: Pregnenolone increases the proliferation of neural stem cells by affecting inflammatory factors and on the other hand, decreases the inflammatory factors TNF-α and IL-6 and oxidative stress factors (NO, MDA) and increases the antioxidant marker (FRAP) in the model. Inflammation caused by lipopolysaccharide in the culture medium of mouse neural stem cells.