Abstract
ObjectivesAlthough many nanomaterials are being used in academia, industry and daily life, there is little understanding about the effects of nanoparticles on the reproductive health of vertebral animals, including human beings. An experimental study was therefore performed here to explore the effect of calcium phosphate nanoparticles on both steroid hormone production and apoptosis in human ovarian granulosa cells.MethodsCalcium phosphate nanoparticles uptaking was evaluated by transmission electron microscopy (TEM). The cell cycle was assessed with propidium iodide-stained cells (distribution of cells in G0/G1, S, and G2/M phases) by flow cytometry. The pattern of cell death (necrosis and apoptosis) was analyzed by flow cytometry with annexin V-FITC/PI staining. The expression of mRNAs encoding P450scc, P450arom and StAR were determined by RT-PCR. Progesterone and estradiol levels were measured by radioimmunoassay.ResultsTEM results confirmed that calcium phosphate nanoparticles could enter into granulosa cells, and distributed in the membranate compartments, including lysosome and mitochondria and intracellular vesicles. The increased percentage of cells in S phase when cultured with nanoparticles indicated that there was an arrest at the checkpoint from phase S-to-G2/M (from 6.28 +/- 1.55% to 11.18 +/- 1.73%, p < 0.05). The increased ratio of S/(G2/M) implied the inhibition of DNA synthesis and/or impairment in the transition of the S progression stage. The apoptosis rate of normal granulosa cells was 7.83 +/- 2.67%, the apoptotic rate increased to 16.53 +/- 5.56% (P < 0.05) after the cells were treated with 100 microM calcium phosphate nanoparticles for 48 hours. Treatment with calcium phosphate nanoparticles at concentrations of 10-100 microM didn't significantly change either the progesterone or estradiol levels in culture fluid, and the expression levels of mRNAs encoding P450scc, P450arom and StAR after 48 h and 72 h period of treatment.ConclusionCalcium phosphate nanoparticles interfered with cell cycle of cultured human ovarian granulosa cells thus increasing cell apoptosis. This pilot study suggested that effects of nanoparticles on ovarian function should be extensively investigated.
Highlights
Nanoparticles possess nanostructure-dependent properties due to their small size, chemical composition, surface charge, solubility and/or shape [1]
We firstly investigated the effect of calcium phosphate nanoparticles itself on hormone production and apoptosis in human granulosa cells cultured in vitro
Effect of HA nanoparticles on granulosa cells transmission electron microscopy (TEM) examination was performed on granulosa cells post 2 h, 4 h, 24 h, 48 h and 72 h exposure to HA nanoparticles
Summary
Nanoparticles possess nanostructure-dependent properties due to their small size, chemical composition, surface charge, solubility and/or shape [1]. Despite the wide applications of nanomaterials, there is a serious lack of information concerning the impact of manufactured. Hydroxyapatite (HA, a kind of calcium phosphate) nanoparticles are similar to human bone in chemical composition and have long been appreciated for their biocompatibility. Calcium phosphate is the primary mineral phase of human and animal bone and tooth. Such a mineral phase, with its plate-like or needle-like shape, typically varies in size from a very few to hundreds of nanometers. HA nanomaterial has been studied for various applications, including orthopedics, dentistry, and food science, with many research topics involving the mineral's interaction with cells [10]
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