Nano-titanium Dioxide Exposure Induced Reproductive Toxicity And Alterations Of Gene Expression In Mice

The dynamical development of the nanotechnology industry has led to thewide-scale production and application of nanomaterials. Among the variousnanomaterials, customarily titanium dioxide nanoparticles （TiO₂NPs）, owing to theirlarger surface area to volume ratio and high reactivity, have been used as nontoxic,chemical inert and biocompatible pigment products or photocatalysts in industry,medicine and health, cosmetics, paint industries, and environmental protection.However, the potentially human toxicity and environmental impact of TiO₂NPs haveattracted considerable attention with their increased use in industrial applications. Inearly2012, the National Research Council of the National Academy of Sciences of USAreleased " environment, health and safety research strategy of engineered nanomaterials（A Research Strategy for Environmental, Health, and Safety Aspects of EngineeredNanomaterials）" report. The report presented the need for overall research program toavoid potential risks caused by fast development nanotechnology（http://www8.nationalacademies.org/onpinews/newsitem.aspx? RecordID=13347）.Recent studies confirmed that the TiO₂NPs exposure can cause liver, kidney, spleen,lung, and brain damages in animals, and clarify their damage mechanisms, particularly,kidney and brain damages are closely related with function of the reproductive system.Long-term use containing TiO₂NPs cosmetics and long-term living or working in theenvironment （such as workers in workshop） exposure to TiO₂NPs, the NPs canpenetrate through the skin, respiratory and digestive system into the body, and possiblythrough blood circulation and blood-testis barrier are accumulated in the ovaries andtestes, which led to damage of the reproductive system. However, the damage by TiO₂NPs exposure and its mechanism needs to be addressed. Thus, the aims of the presentstudy were to test the hypothesis that TiO₂NPs exerts a toxic effect in the reproductivesystem of mice, and elucidate the synergistic molecular mechanisms of multiple genesactivated by TiO₂NPs-induced toxicity of reproductive system in animals and humans.In this study, ovarian and testicular damages and alterations in gene expression profiles in female and male mice induced by intragastric administration of10mg/kg bodyweight of TiO₂NPs for90consecutive days were examined. This study can provideimportant theoretical basis for assessment of reproductive toxicity for long-termexposure to nanomaterials.Results are listed as follows:（1） In the current study, ovarian injury and gene-expressed characteristics in femalemice induced by intragastric administration of TiO₂NPs （10mg/kg） for90consecutivedays were investigated. Our findings indicated that TiO₂NPs can accumulate in theovary and result in ovarian damage including ovarian atrophy, disturbance of primaryand second follicle development, irregular arrangement of cells, and a shapelessfollicular antrum. TiO₂NPs can enter ovarian cells and even the nuclei of ovarian cells,thus leading to apoptosis. TiO₂NPs exposure caused increases of calcium, sodium,potassium and zinc contents, and decreases of magnesium, copper, and iron contents, animbalance of sex hormones such as decreased levels of serum progesterone, luteinizinghormone, testosterone and follicle-stimulating, and increased concentration of estradiolin the ovary, which resulted in significant decreases in the mating rate, pregnancy rate,and number of fetuses, and inhibition of growth and development of offspring. TiO₂NPsexposure caused severe ovarian oxidative damage, such as overproduction of ROS andincrease level of DNA oxidation （8-OHdg）. Microarray analysis showed that in ovariesfrom mice treated with TiO₂NPs compared to controls,223genes of known functionwere up-regulated（diffscore≥13, p <0.05）, while65ovarian genes weredown-regulated（diffscore-13, p <0.05）. In the present study, functions of differencegene in the ovaries were involved in cell differentiation, immune and inflammatoryresponse, ion transport, metabolic process, oxidative stress, the steroid metabolism,apoptosis, signal transduction, transcription, transport, etc. Especially Akr1c18,Cyp17a1and Lgmn that related to the steroids and sex hormone metabolism processsignificantly up-regulated, over-expression of these genes caused increased levels ofestradiol, progesterone, luteinizing hormone and decreased level of testosterone, therebyinhibited follicular development and reduced fertility of female mice. In addition, TiO₂NPs exposure resulted in a significant increase in the expression of ovarian Cyp17a1,which also can promote the biosynthesis of estradiol.（2） In this study, the reproductive toxicity and testicular gene expression profilinginduced by intragastric administration of10mg/kg body weight TiO₂NPs for90 consecutive days were investigated. The results showed that TiO₂NPs can be depositedin the testicles, and cause testicular damage and apoptosis, such as rare sperm, spermbreakages, rarefaction of Sertoli cell and androgone, Sertoli cell apoptosis, androgonenecrosis of the seminiferous tubules, decreased germinative layer thickness, vacuolation,and irregular arrangement of Sertoli cells of the seminiferous tubules, significantdecreases in sperm numbers and sperm motility, and increased abnormal sperm, spermwas severely damaged, such as fracture of the head and tail in the cauda epididymis, thefertility of male mice significantly decreased, such as significant decreases in the matingrate, pregnancy rate, and number of fetuses. TiO₂NPs exposure led to the decline ofcalcium level and increases of iron, zinc contents in testicular tissue. Similarly, TiO₂NPs exposure also caused an imbalance of sex hormones, e.g., estradiol andprogesterone significantly increased, and levels of follicle-stimulating hormone,luteinizing hormone and testosterone were significantly decreased. Microarray analysisshowed that in testes from mice treated with TiO₂NPs compared to controls,155geneswere found to be up-regulated（diffscore≥13, p<0.05）, while100genes were found to bedown-regulated （diffscore-13, p<0.05）. Functions of difference genes were involved inspermatogenesis, steroid metabolic processes, apoptosis, immune, cell differentiation,oxidative stress, redox activity, hormone activity, ion transport. Of the altered geneexpressions, significantly down-regulated expressions of Spata19, Tdrd6and Tnp2weredirectly related with the suppression of spermatogenesis, and Cyp2e1, Mvd, Sc4mol andSrd5a2were related with steroid metabolic process. Cyp2e1was downregulated, Mvd,Sc4mol and Srd5a2were upregulated. These alterations of gene expressions impairedbalance of sex hormones, and decreased fertility of male mice.