Virginie Tassistro

Postnatal Programming of Glucocorticoid Metabolism in Rats Modulates High-Fat Diet–Induced Regulation of Visceral Adipose Tissue Glucocorticoid Exposure and Sensitivity and Adiponectin and Proinflammatory Adipokines Gene Expression in Adulthood

OBJECTIVE—Alterations of the perinatal environment, which lead to increased prevalence of the metabolic syndrome in adulthood, program an upregulation of systemic and/or adipose tissue glucocorticoid metabolism (11β-hydroxysteroid dehydrogenase type 1 [11β-HSD-1]-induced corticosterone reactivation). We hypothesized that postnatal programming could modulate high-fat diet–induced adipose tissue dysregulation in adulthood. RESEARCH DESIGN AND METHODS—We compared the effects of chronic (since weaning) high- or low-fat diet in postnatally normofed (control) or overfed (programmed) rats. RESULTS—Postnatal programming accentuated high-fat diet–induced overweight, insulin resistance, glucose intolerance, and decrease in circulating and epididymal adipose tissue adiponectin. Neither manipulation altered liver function. Postnatal programming or high-fat diet increased systemic corticosterone production, which was not further modified when both manipulations were associated. Postnatal programming suppressed high-fat diet–induced decrease in mesenteric adipose tissue (MAT) glucocorticoid sensitivity and triggered high-fat diet–induced increase in MAT glucocorticoid exposure, subsequent to enhanced MAT 11β-HSD-1 gene expression. MAT tumor necrosis factor (TNF)-α, TNF-receptor 1, interleukin (IL)-6, resistin, and plasminogen activator inhibitor-1 mRNAs were not changed by high-fat feeding in control rats and showed a large increase in programmed animals, with this effect further enhanced by high-fat diet for TNF-α and IL-6. CONCLUSIONS—Our data show for the first time that postnatal manipulation programs high-fat diet–induced upregulation of MAT glucocorticoid exposure, sensitivity, and inflammatory status and therefore reveal the pivotal role of the environment during the perinatal period on the development of diet-induced adipose tissue dysregulation in adulthood. They also urge the need for clinical trials with specific 11β-HSD-1 inhibitors.

Ultrastructural Interactions and Genotoxicity Assay of Cerium Dioxide Nanoparticles on Mouse Oocytes

Cerium dioxide nanoparticles (CeO2 ENPs) are on the priority list of nanomaterials requiring evaluation. We performed in vitro assays on mature mouse oocytes incubated with CeO2 ENPs to study (1) physicochemical biotransformation of ENPs in culture medium; (2) ultrastructural interactions with follicular cells and oocytes using Transmission Electron Microscopy (TEM); (3) genotoxicity of CeO2 ENPs on follicular cells and oocytes using a comet assay. DNA damage was quantified as Olive Tail Moment. We show that ENPs aggregated, but their crystal structure remained stable in culture medium. TEM showed endocytosis of CeO2 ENP aggregates in follicular cells. In oocytes, CeO2 ENP aggregates were only observed around the zona pellucida (ZP). The comet assay revealed significant DNA damage in follicular cells. In oocytes, the comet assay showed a dose-related increase in DNA damage and a significant increase only at the highest concentrations. DNA damage decreased significantly both in follicular cells and in oocytes when an anti-oxidant agent was added in the culture medium. We hypothesise that at low concentrations of CeO2 ENPs oocytes could be protected against indirect oxidative stress due to a double defence system composed of follicular cells and ZP.

DNA damage and oxidative stress induced by CeO2 nanoparticles in human dermal fibroblasts: Evidence of a clastogenic effect as a mechanism of genotoxicity

Abstract The broad range of applications of cerium oxide (CeO2) nanoparticles (nano-CeO2) has attracted industrial interest, resulting in greater exposures to humans and environmental systems in the coming years. Their health effects and potential biological impacts need to be determined for risk assessment. The aims of this study were to gain insights into the molecular mechanisms underlying the genotoxic effects of nano-CeO2 in relation with their physicochemical properties. Primary human dermal fibroblasts were exposed to environmentally relevant doses of nano-CeO2 (mean diameter, 7 nm; dose range, 6 × 10−5–6 × 10−3 g/l corresponding to a concentration range of 0.22–22 µM) and DNA damages at the chromosome level were evaluated by genetic toxicology tests and compared to that induced in cells exposed to micro-CeO2 particles (mean diameter, 320 nm) under the same conditions. For this purpose, cytokinesis-blocked micronucleus assay in association with immunofluorescence staining of centromere protein A in micronuclei were used to distinguish between induction of structural or numerical chromosome changes (i.e. clastogenicity or aneuploidy). The results provide the first evidence of a genotoxic effect of nano-CeO2, (while not significant with micro-CeO2) by a clastogenic mechanism. The implication of oxidative mechanisms in this genotoxic effect was investigated by (i) assessing the impact of catalase, a hydrogen peroxide inhibitor, and (ii) by measuring lipid peroxidation and glutathione status and their reversal by application of N-acetylcysteine, a precusor of glutathione synthesis in cells. The data are consistent with the implication of free radical-related mechanisms in the nano-CeO2-induced clastogenic effect, that can be modulated by inhibition of cellular hydrogen peroxide release.

Tobacco consumption and benzo(a)pyrene-diol-epoxide–DNA adducts in spermatozoa: in smokers, swim-up procedure selects spermatozoa with decreased DNA damage

OBJECTIVE To analyze the distribution of benzo(a)pyrene-diol-epoxide (BPDE)-DNA adducts in spermatozoa selected and nonselected by a swim-up procedure with relation to smoking habits. DESIGN Comparative study. SETTING Public university and public university hospital. PATIENT(S) Seventy-nine men (37 smokers and 42 nonsmokers) who visited an infertility clinic for diagnostic. INTERVENTION(S) Tobacco and environmental exposure assessment, semen sample analysis, swim-up procedure, BPDE-DNA adduct immunolabeling. MAIN OUTCOME MEASURE(S) BPDE-DNA adduct quantification in selected (SEL-SPZ) and nonselected (NONSEL-SPZ) spermatozoa. Data were normalized by using a normalized fluorescence value (NFV). RESULT(S) The mean NFV (±SD) in SEL-SPZ was significantly higher in smokers than in nonsmokers (18.9±11.5 vs. 10.5±10.4, respectively). Within smokers, a paired analysis (SEL-SPZ and NONSEL-SPZ) showed that NFV was significantly lower in SEL-SPZ than in NONSEL-SPZ (20.0±11.3 vs. 31.5±16.0, respectively). Conversely, within nonsmokers, the mean NFV was higher in SEL-SPZ than in NONSEL-SPZ (10.3±10.6 vs 4.3±7.1, respectively). CONCLUSION(S) Tobacco consumption is associated with BPDE-DNA adducts in spermatozoa. In smokers, semen processing by swim-up recovers potentially fertilizing spermatozoa that show a significantly lower amount of BPDE-DNA adducts compared with NONSEL-SPZ. Further study is needed to improve the spermatozoa selection in smoking patients requiring assisted reproductive technologies.

Cerium dioxide nanoparticles affect in vitro fertilization in mice

Abstract Due to their catalytic and oxidative properties, cerium dioxide nanoparticles (CeO2NPs) are widely used as diesel additive or as promising therapy in cancerology; yet, scarce data are available on their toxicity, and none on their reproductive toxicity. We showed a significant decrease of fertilization rate, assessed on 1272 oocytes, during in vitro fertilization (IVF) carried out in culture medium containing CeO2NP at very low concentration (0.01 mg.l−1). We also showed significant DNA damage induced in vitro by CeO2NP on mouse spermatozoa and oocytes at 0.01 mg.l−1 using Comet assay. Transmission Electron Microscopy did not detect any nanoparticles in the IVF samples at 0.01 mg.l−1, but showed, at high concentration (100 mg.l−1), their endocytosis by the cumulus cells surrounding oocytes and their accumulation along spermatozoa plasma membranes and oocytes zona pellucida. We did not observe any nanoparticles in the cytoplasm of spermatozoa, oocytes or embryos. This study demonstrates for the first time the impact of CeO2NP on in vitro fertilization, as well as their genotoxicity on mouse spermatozoa and oocytes, at low nanoparticle concentration exposure. Decreased fertilization rates may result from: (1) CeO2NP’s genotoxicity on gametes; (2) a mechanical effect, disrupting gamete interaction and (3) oxidative stress induced by CeO2NP. These results add new and important insights with regard to the reproductive toxicity of nanomaterials requesting urgent evaluation, and support several publications on metal nanoparticles reprotoxicity. Our data highlight the need for in vivo studies after low-dose exposure.

Poorly soluble cobalt oxide particles trigger genotoxicity via multiple pathways

BackgroundPoorly soluble cobalt (II, III) oxide particles (Co3O4P) are believed to induce in vitro cytotoxic effects via a Trojan-horse mechanism. Once internalized into lysosomal and acidic intracellular compartments, Co3O4P slowly release a low amount of cobalt ions (Co2+) that impair the viability of in vitro cultures.In this study, we focused on the genotoxic potential of Co3O4P by performing a comprehensive investigation of the DNA damage exerted in BEAS-2B human bronchial epithelial cells.ResultsOur results demonstrate that poorly soluble Co3O4P enhanced the formation of micronuclei in binucleated cells. Moreover, by comet assay we showed that Co3O4P induced primary and oxidative DNA damage, and by scoring the formation of γ-H2Ax foci, we demonstrated that Co3O4P also generated double DNA strand breaks.ConclusionsBy comparing the effects exerted by poorly soluble Co3O4P with those obtained in the presence of soluble cobalt chloride (CoCl2), we demonstrated that the genotoxic effects of Co3O4P are not simply due to the released Co2+ but are induced by the particles themselves, as genotoxicity is observed at very low Co3O4P concentrations.

Expression and nutritional regulation of the (pro)renin receptor in rat visceral adipose tissue

Background: Early life nutritional environment plays an important role in the development of visceral adipose tissue and interacts with nutritional regulations in adulthood, leading to metabolic dysregulations. Aim: We hypothesized that the renin-angiotensin system may play a role in the programming-induced development of visceral adipose tissue. Material and methods: We studied, using a model of programming of overweight and glucose intolerance, obtained by post-natal overfeeding with consecutive high-fat diet, the status of plasma renin activity and mesenteric adipose renin-angiotensin system, including the recently identified (pro)renin receptor, in adult rats. Results: Post-natal overfeeding or high-fat feeding lead to overweight with increased visceral fat mass and adipocytes surface. When both paradigms were associated, adipocytes surface showed a disproportionate increase. A strong immunoreactivity for (pro)renin receptor was found in stromal cells. Plasma renin activity increased in programmed animals whereas (pro)renin receptor expressing cells density was stimulated by high-fat diet. There was a positive, linear relationship between plasma renin activity and (pro)renin receptor expressing cells density and adipocytes surface. Conclusions: Our experiments demonstrate that association of post-natal overfeeding and high-fat diet increased plasma renin activity and adipose (pro)renin receptor expression. Such phenomenon could explain, at least in part, the associated disproportionate adipocyte hypertrophy and its accompanying increased glucose intolerance.

A new approach for the oocyte genotoxicity assay: adaptation of comet assay on mouse cumulus–oocyte complexes

Conventional genotoxicity tests are technically difficult to apply to oocytes, and results obtained on somatic cells cannot be extrapolated to gametes. We have previously described a comet assay (original–CA) on denuded mouse oocytes, but, in vivo, oocytes are not isolated from their surrounding follicular cells. Our objective was to develop a comet assay on cumulus–oocyte complexes (COC–CA) for a more physiological approach to study the genotoxicity of environmental factors on oocytes. For COC–CA, whole COC were exposed directly to exogenous agents after ovulation and removal from oviducts. Three conditions were studied: a negative control group, and two positive control groups, one of which was exposed to hydrogen peroxide (H2O2) and the other group was incubated with cerium dioxide nanoparticles (CeO2 NPs). With both tests, DNA damage was significant in the presence of both H2O2 and CeO2 NPs compared with the negative control. COC–CA offers an interesting tool for assaying the genotoxicity of environmental agents towards germinal cells. Furthermore, COC–CA is less time-consuming and simplifies the protocol of the original–CA, because COC-CA is easier to perform without the washing-out procedure.

Immediate Postnatal Overfeeding in Rats Programs Aortic Wall Structure Alterations and Metalloproteinases Dysregulation in Adulthood.

BACKGROUND Alterations in the nutritional perinatal environment, such as intrauterine growth retardation with subsequent postnatal catch-up growth, program cardiovascular disease in adulthood, possibly through alterations in matrix metalloproteinase (MMP)-2 and -9. However, experimental evidences demonstrating that changes in the nutritional perinatal environment can program MMP-2 and -9 with subsequent alterations of vessel wall are lacking. AIM The current study evaluated whether immediate postnatal overfeeding is able to alter vascular morphological indexes and circulating and/or vascular MMP2-2 and -9 status. METHODS Aortic morphology (wall thickness and percentage of incomplete elastin lamellae) and circulating and aortic MMP-2 and -9 activity (measured by gelatin zymography) and aortic MMP-2 and -9 mRNA (measured by reverse transcription polymerase chain reaction (RT-PCR)) were studied in adult male rats overfed (OF) or normofed (NF) during the immediate postnatal period. RESULTS Postnatal overfeeding induced early onset obesity. Adult OF rats presented with increased blood pressure and circulating MMP-2 and -9 activity. In the thoracic aorta, postnatal overfeeding increased wall thickness and decreased elastin integrity (as demonstrated by an increased percentage of incomplete elastin lamellae). OF rats showed enhanced aortic MMP-2 activity and MMP-9 mRNA levels. Circulating and aortic MMP-2 activity correlated positively with the percentage of incomplete elastin lamellae and aortic wall thickness, respectively. CONCLUSION Our data demonstrate for the first time that immediate postnatal nutritional programming induces increases in circulating and aortic MMP-2 activity with parallel aortic wall alterations, such as decreased elastin integrity and enhanced thickening, showing that this experimental model is suitable for the study of perinatal nutritional programming of vascular functions.