p53 regulates primordial follicle activation through the mTOR signaling pathway / 生理学报

This paper aimed to investigate the role and potential mechanism of p53 on primordial follicle activation. Firstly, the p53 mRNA expression in the ovary of neonatal mice at 3, 5, 7 and 9 days post-partum (dpp) and the subcellular localization of p53 were detected to confirm the expression pattern of p53. Secondly, 2 dpp and 3 dpp ovaries were cultured with p53 inhibitor Pifithrin-μ (PFT-μ, 5 μmol/L) or equal volume of dimethyl sulfoxide for 3 days. The function of p53 in primordial follicle activation was determined by hematoxylin staining and whole ovary follicle counting. The proliferation of cell was detected by immunohistochemistry. The relative mRNA levels and protein levels of the key molecules involved in the classical pathways associated with the growing follicles were examined by immunofluorescence staining, Western blot and real-time PCR, respectively. Finally, rapamycin (RAP) was used to intervene the mTOR signaling pathway, and ovaries were divided into four groups: Control, RAP (1 μmol/L), PFT-μ (5 μmol/L), PFT-μ (5 μmol/L) + RAP (1 μmol/L) groups. The number of follicles in each group was determined by hematoxylin staining and whole ovary follicle counting. The results showed that the expression of p53 mRNA was decreased with the activation of primordial follicles in physiological condition. p53 was expressed in granulosa cells and oocyte cytoplasm of the primordial follicles and growing follicles, and the expression of p53 in the primordial follicles was higher than that in the growing follicles. Inhibition of p53 promoted follicle activation and reduced the primordial follicle reserve. Inhibition of p53 promoted the proliferation of the granulosa cells and oocytes. The mRNA and protein expression levels of key molecules in the PI3K/AKT signaling pathway including AKT, PTEN, and FOXO3a were not significantly changed after PFT-μ treatment, while the expression of RPS6/p-RPS6, the downstream effectors of the mTOR signaling pathway, was upregulated. Inhibition of both p53 and mTOR blocked p53 inhibition-induced primordial follicle activation. Collectively, these findings suggest that p53 may inhibit primordial follicle activation through the mTOR signaling pathway to maintain the primordial follicle reserve.

Oenothein B inhibits proliferation and migration of breast cancer cells by regulating P53 / 中国中药杂志

The effects of oenothein B(OEB) on the proliferation, apoptosis, invasion, and migration of breast cancer MCF-7 and MDA-MB-231 cells were investigated by cell culture in vitro, network pharmacology, and molecular docking. In vitro cell experiments revealed that OEB inhibited the proliferation and colony formation ability, and promoted the apoptosis and formation of apoptotic bodies in breast cancer cells, as well as inhibited the invasion and migration of breast cancer cells. The targets of OEB were obtained using SwissTargetPrediction database and breast cancer targets were obtained from GeneCards. The targets of OEB and breast cancer were entered separately in Venny 2.1 software to obtain the Venn diagram of common targets of OEB and breast cancer. The common targets of OEB and breast cancer were input into STRING database to construct a protein-protein interaction(PPI) network, which was entered into Cytoscape 3.7.2 software for network topology analysis. Key targets were screened according to protein association strength, and analyzed for KEGG pathway enrichment. Molecular docking of OEB to key targets using AutoDock software revealed that OEB stably bound to the active pocket of P53, while OEB promoted the expression of P53 protein. MCF-7 and MDA-MB-231 cell viability and migration ability increased after silencing P53, and this change was reversed after treatment with OEB. Therefore, this study showed that OEB inhibited the proliferation, migration, and invasion of breast cancer MCF-7 and MDA-MB-231 cells, and promoted the apoptosis of breast cancer MCF-7 and MDA-MB-231 cells, which may be related to the targeted regulation of P53.

Expression and localization of endogenous C-reactive protein in THP-1 monocytes and LO2 hepatocytes / 南方医科大学学报

<p><b>OBJECTIVE</b>To observe the expression and localization of endogenous C-reactive protein (CRP) in cells from different tissues under different conditions.</p><p><b>METHODS</b>Macrophages differentiated from THP-1 monocytes with phorbol ester (PMA) induction and human LO2 hepatocytes were stimulated with lipopolysaccharide (LPS). The culture supernatant of the LPS-stimulated THP-1 cells was collected and added into LO2 cell culture, and after incubation, the cells were lysed to extract the proteins for SDS-PAGE and Western blotting. The stimulated cells were also examined immunocytochemically for CRP expression.</p><p><b>RESULTS</b>Western blotting detected CRP in both of the unstimulated cell lysates, but in neither of the two cell supernatants. After LPS stimulation, CRP expression was significantly increased in the cell lysate of THP-1 cells with also a small amount present in the supernatant, but CRP expression and release in the LO2 cells showed no significant variation. Treatment of the LO2 cells with the culture supernatant of LPS-stimulated THP-1 cells resulted in positivity of CRP in the cell lysate and the culture supernatant. Immunocytochemistry identified CRP expression throughout the THP-1 cell body (most obvious in the nuclei), which increased after LPS stimulation. In LO2 hepatocytes, CRP expression was found only outside the nuclei and increased after stimulation with the culture supernatant of LPS-treated THP-1 cells, especially obvious around the membrane.</p><p><b>CONCLUSION</b>CRP can not be up-regulated directly by LPS treatment in LO2 cells, but can be induced by certain cytokines (IL-6) secreted from LPS-stimulated THP-1 cells. The localization of CRP represents the characteristics of secreted protein in LO2 cells, but in THP-1 cells, CRP is found mainly in the cell nuclei.</p>