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Pro-osteoporotic miR-320a impairs osteoblast function and induces oxidative stress.

ABSTRACT: MicroRNAs (miRNAs) are important regulators of many cellular processes, including the differentiation and activity of osteoblasts, and therefore, of bone turnover. MiR-320a is overexpressed in osteoporotic bone tissue but its role in osteoblast function is unknown. In the present study, functional assays were performed with the aim to elucidate the mechanism of miR-320a action in osteoblastic cells. MiR-320a was either overexpressed or inhibited in human primary osteoblasts (hOB) and gene expression changes were evaluated through microarray analysis. In addition, the effect of miR-320a on cell proliferation, viability, and oxidative stress in hOB was evaluated. Finally, matrix mineralization and alkaline phosphatase activity were assessed in order to evaluate osteoblast functionality. Microarray results showed miR-320a regulation of a number of key osteoblast genes and of genes involved in oxidative stress. Regulation of osteoblast differentiation and ossification appeared as the best significant biological processes (PANTHER P value = 3.74E-05; and P value = 3.06E-04, respectively). The other enriched pathway was that of the cellular response to cadmium and zinc ions, mostly by the overexpression of metallothioneins. In hOBs, overexpression of miR-320a increased cell proliferation and oxidative stress levels whereas mineralization capacity was reduced. In conclusion, overexpression of miR-320a increased stress oxidation levels and was associated with reduced osteoblast differentiation and functionality, which could trigger an osteoporotic phenotype.

SUBMITTER: De-Ugarte L

PROVIDER: S-EPMC6261634 | biostudies-literature | 2018

REPOSITORIES: biostudies-literature

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Pro-osteoporotic miR-320a impairs osteoblast function and induces oxidative stress.

De-Ugarte Laura L Balcells Susana S Nogues Xavier X Grinberg Daniel D Diez-Perez Adolfo A Garcia-Giralt Natalia N

PloS one 20181128 11

MicroRNAs (miRNAs) are important regulators of many cellular processes, including the differentiation and activity of osteoblasts, and therefore, of bone turnover. MiR-320a is overexpressed in osteoporotic bone tissue but its role in osteoblast function is unknown. In the present study, functional assays were performed with the aim to elucidate the mechanism of miR-320a action in osteoblastic cells. MiR-320a was either overexpressed or inhibited in human primary osteoblasts (hOB) and gene expres ...[more]

PMID: 30485349

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Project description:Introduction: Methoxychlor (MXC) is an organochlorine pesticide (OCP) that was formerly used worldwide as an insecticide against pests and mosquitoes. However, MXC is not biodegradable and has lipophilic characteristics; thus, it accumulates in organisms and affects reproductive function. MXC, as an estrogenic compound, promotes oxidative stress, induces oxidative stress damage to ovarian follicles, and causes miscarriages and stillbirths in females. In this research endeavor, our primary objective was to explore the ramifications of MXC regarding the developmental processes occurring during the initial stages of embryogenesis in pigs. Methods: In this study, we counted the blastocyst rate of early embryos cultured in vitro. We also examined the reactive oxygen species level, glutathione level, mitochondrial membrane potential, mitochondrial copy number and ATP level in four-cell stage embryos. Finally, apoptosis and DNA damage in blastocyst cells, as well as pluripotency-related and apoptosis-related genes in blastocyst cells were detected. The above experiments were used to evaluate the changes of MXC damage on early parthenogenetic embryo development. Results and Discussion: The results showed that early embryos exposed to MXC had a significantly lower cleavage rate, blastocyst rate, hatching rate, and total cell count compared with the control group. It was also of note that MXC not only increased the levels of reactive oxygen species (ROS), but also decreased the mitochondrial membrane potential (ΔΨm) and mitochondrial copy number during the development of early embryos. In addition, after MXC treatment, blastocyst apoptosis and DNA damage were increased, decreased cell proliferation, and the expression of pluripotency-related genes SOX2, NANOG, and OCT4 was down-regulated, while the expression of apoptosis-related genes BAX/BCL-2 and Caspase9 was up-regulated. Our results clearly show that MXC can have deleterious effects on the developmental processes of early porcine embryos, establishing the toxicity of MXC to the reproductive system. In addition, the study of this toxic effect may lead to greater concern about pesticide residues in humans and the use of safer pesticides, thus potentially preventing physiological diseases caused by chemical exposure.

Project description:Oxidative stress, generated by excessive reactive oxygen species (ROS) or decreased antioxidant defenses (and possibly both), is associated with male infertility. Oxidative stress results in redox-dependent protein modifications, such as tyrosine nitration and S-glutathionylation. Normozoospermic sperm samples from healthy individuals were included in this study. Samples were incubated with increasing concentrations (0-5 mM) of exogenous hydrogen peroxide, tert-butyl hydroperoxide, or diethylamine NONOate (DA-NONOate, a nitric oxide (NO∙) donor) added to the medium. Spermatozoa treated with or without ROS were incubated under capacitating conditions and then levels of tyrosine phosphorylation and percentage of acrosome reaction (AR) induced by lysophosphatidylcholine were determined. Modified sperm proteins from cytosolic, triton-soluble, and triton-insoluble fractions were analyzed by SDS-PAGE immunoblotting and immunocytochemistry with anti-glutathione and anti-nitrotyrosine antibodies. Levels of S-glutathionylation increased dose dependently after exposure to hydroperoxides (P<0.05) and were localized mainly to the cytosolic and triton-soluble fractions of the spermatozoa. Levels of tyrosine-nitrated proteins increased dose dependently after exposure to DA-NONOate (P<0.05) and were mainly localized to the triton-insoluble fraction. ROS-treated spermatozoa showed impaired motility without affecting viability (hypo-osmotic swelling test). These treated spermatozoa had tyrosine phosphorylation and AR levels similar to that of non-capacitated spermatozoa following incubation under capacitating conditions, suggesting an impairment of sperm capacitation by oxidative stress. In conclusion, oxidative stress promotes a dose-dependent increase in tyrosine nitration and S-glutathionylation and alters motility and the ability of spermatozoa to undergo capacitation.Free Spanish abstractA Spanish translation of this abstract is freely available at http://www.reproduction-online.org/content/149/1/113/suppl/DC1.

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Pro-osteoporotic miR-320a impairs osteoblast function and induces oxidative stress.

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Pro-osteoporotic miR-320a impairs osteoblast function and induces oxidative stress.

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