莪术油对卵巢癌裸鼠移植瘤的抑制作用及其联合顺铂的协同作用研究

目的 探讨莪术油对卵巢癌裸鼠移植瘤的抑制作用,并探讨其与顺铂联合的协同作用。方法 建立卵巢癌细胞株HO-8910PM荷瘤裸鼠模型,随机分为模型对照组、莪术油组、顺铂组和联合给药组（莪术油+顺铂）4组,每组6只,分别腹腔注射莪术油（300 mg·kg^-1）、顺铂（2 mg·kg^-1）、莪术油联合顺铂（300 mg·kg^-1+2 mg·kg^-1）以及空白对照液（0.4 mL生理盐水）,给药3周后处死小鼠并测瘤体积变化,计算肿瘤生长抑制率,免疫组化检测各组肿瘤组织中NM23、TNF-α、VEGF、NF-KBP65、PCNA的蛋白表达。ELISA检测裸鼠血清中IL-2、IFN-γ水平。结果 与模型对照组比较,莪术油、顺铂及其联合给药后均能显著抑制裸鼠肿瘤体积的增长（P<0.05或P<0.01）,且联合给药的抑瘤率（55.06%）要高于莪术油（20.5%）和顺铂（52.52%）单独给药。免疫组化检测结果显示各组药物治疗后,能显著降低裸鼠肿瘤组织中NM23、TNF-α、VEGF、NF-KBP65、PCNA的阳性表达（P<0.05或P<0.01）,且降低程度从低到高依次为莪术油组、顺铂组、联合给药组。ELISA检测结果也表明顺铂以及莪术油联合顺铂给药3周后能显著提高裸鼠血清IL-2、IFN-γ水平（P<0.01）,且含量上升程度由低到高依次为莪术油组、顺铂组、联合给药组。结论 莪术油能够抑制卵巢癌荷瘤裸鼠移植瘤的增长,具有一定的抗肿瘤效果,其机制可能与调节肿瘤生长因子NM23、TNF-α、VEGF、NF-KBP65、PCNA及免疫相关因子IL-2、IFN-γ的表达有关。莪术油联合顺铂用药对卵巢癌肿瘤的抑制具有潜在的协同作用,但还需更深入实验研究。     

Toxicological evaluation of metal oxide nanoparticles and mixed exposures at low doses using zebra fish and THP1 cell line

Metal and metal oxide nanoparticles are being used in different industries now‐a‐days leading to their unavoidable exposure to humans and animals. In the present study, toxicological testing was done using nanoparticles of copper oxide, cerium oxide and their mixture (1:1 ratio) on zebra fish embryos and THP‐1 cell line. Zebrafish embryos were exposed to 0.01 μg/ml to 50 μg/ml concentrations of dispersed nanoparticles using a 96 well plate and their effects were studied at different hours post fertilization (hpf) i.e. 0 hpf, 24 hpf, 48 hpf, 72 hpf and 96 hpf respectively. Results showed that copper oxide nanoparticles has drastic effects on the morphology and physiology of zebra fish whereas cerium oxide nanoparticles and mixture of these nanoparticles did not show much of the effects. Comparable results were obtained from in vitro study using human monocyte cell line (THP‐1). It is concluded that these nanoparticles may cause toxicological effects to humans and environment.     

The size‐dependent genotoxic potentials of titanium dioxide nanoparticles to endothelial cells

Despite intensive research activities, there are still many major knowledge gaps over the potential adverse effects of titanium dioxide nanoparticles (TiO₂‐NPs), one of the most widely produced and used nanoparticles, on human cardiovascular health and the underlying mechanisms. In the present study, alkaline comet assay and cytokinesis‐block micronucleus test were employed to determine the genotoxic potentials of four sizes (100, 50, 30, and 10 nm) of anatase TiO₂‐NPs to human umbilical vein endothelial cells (HUVECs) in culture. Also, the intracellular redox statuses were explored through the measurement of the levels of reactive oxygen species (ROS) and reduced glutathione (GSH) with kits, respectively. Meanwhile, the protein levels of nuclear factor erythroid 2‐related factor 2 (Nrf2) were also detected by western blot. The results showed that at the exposed levels (1, 5, and 25 μg/mL), all the four sizes of TiO₂‐NPs could elicit an increase of both DNA damage and MN frequency in HUVECs in culture, with a positive dose‐dependent and negative size‐dependent effect relationship (T100 < T50 < T30 < T10). Also, increased levels of intracellular ROS, but decreased levels of GSH, were found in all the TiO₂‐NP‐treated groups. Intriguingly, a very similar manner of dose‐dependent and size‐dependent effect relationship was observed between the ROS test and both comet assay and MN test, but contrary to that of GSH assay. Correspondingly, the levels of Nrf2 protein were also elevated in the TiO₂‐NP‐exposed HUVECs, with an inversely size‐dependent effect relationship. These findings indicated that induction of oxidative stress and subsequent genotoxicity might be an important biological mechanism by which TiO₂‐NP exposure would cause detrimental effects to human cardiovascular health.     

In vitro genotoxic and cytotoxic effects of some paraben esters on human peripheral lymphocytes

Parabens (PBs) are p-hydroxybenzoic acid ester compounds commonly employed as antimicrobial preservatives, mainly in food, cosmetic, and pharmaceutical products. The aim of the present study was to investigate the genotoxic and cytotoxic effects of some paraben esters (butyl paraben, propyl paraben, isobutyl paraben, and isopropyl paraben) on human peripheral lymphocytes, using in vitro sister chromatid exchange (SCE), chromosome aberration (CA), and cytokinesis-block micronucleus (CBMN) tests. Lymphocyte cultures were treated with four concentrations of PBs (100, 50, 25 and 10?µg/mL) for 24 and 48?h. Paraben esters significantly induced MN formations as compared to solvent control. Furthermore, butyl paraben and propyl paraben increased MN formations a concentration-dependent manner at 24 and 48?h. PBs increased the CA at 24 and 48?h. However, this increase was not meaningful for butyl paraben and isopropyl paraben at 48?h when compared with solvent control. Butyl, isobutyl, and isopropyl paraben significantly increased the SCE at 24 and 48?h. However, propyl paraben did not induce SCE meaningfully in both treatment periods. A significant decrease in the cytokinesis-block proliferation index and mitotic index was observed in cells exposed to all concentrations of PBs at 24 and 48?h. However, proliferation index was not affected at all concentrations of PBs after 24?h treatment, although it was decreased at the highest concentration of PBs at 48?h. It is concluded that all of the paraben esters used in this study have highly genotoxic and cytotoxic effects on human lymphocytes cells in vitro.     

Mechanisms of genotoxicity. A review of in vitro and in vivo studies with engineered nanoparticles

Engineered nanoparticles (NPs) are widely used in different technologies but their unique properties might also cause adverse health effects. In reviewing recent in vitro and in vivo genotoxicity studies we discuss potential mechanisms of genotoxicity induced by NPs. Various factors that may influence genotoxic response, including physico-chemical properties and experimental conditions, are highlighted. From 4346 articles on NP toxicity, 112 describe genotoxicity studies (94 in vitro, 22 in vivo). The most used assays are the comet assay (58 in vitro, 9 in vivo), the micronucleus assay (31 in vitro, 14 in vivo), the chromosome aberrations test (10 in vitro, 1 in vivo) and the bacterial reverse mutation assay (13 studies). We describe advantages and potential problems with different methods and suggest the need for appropriate methodologies to be used for investigation of genotoxic effects of NPs, in vitro and in vivo.     

GSH-responsive curcumin/doxorubicin encapsulated Bactrian camel serum albumin nanocomposites with synergistic effect against lung cancer cells

The aim of this study was to prepare camel serum albumin (CSA) nanoparticles using a self-assembly strategy to co-immobilize curcumin (CCM) and doxorubicin (Dox) which was in favor of combined chemotherapy and biomedical applications of bactrian (Camelus bactrianus) CSA. The constructed CSA nanoparticles (CSA-NPs) with the size around 200 nm displayed a high degree of polydispersity and further encapsulation of CCM and Dox caused no apparent morphological changes to the nanocomposite (CCM/Dox CSA-NPs). The synergistic cytotoxic effect of CCM and Dox on cancer cell A549 was observed with the calculated combination index less than 1.0. Moreover, the release kinetic profile of encapsulated drugs showed a concentration dependence of glutathione (GSH) originating from the GSH used in nanoparticle formation to break the intramolecular disulfide bonds. In vitro cytotoxicity evaluations also revealed that CCM/Dox CSA-NPs showed higher cytotoxicity than that of single drug loaded CSA-NPs, which was also validated by high content screen assay. Taken together, the CCM/Dox CSA-NPs with redox-responsive attributes provided an integrated protein-based combinational drug-delivery matrix to exert synergistic effects.     

Genotoxicity and cytotoxicity of sevoflurane in two human cell lines in vitro with ionizing radiation

Objective:

To determine the in vitro toxicity of different concentrations of sevoflurane in cells exposed to X-ray.

Methods:

The genotoxic effects of sevofluorane were studied by means of the micronucleus test in cytokinesis-blocked cells of irradiated human lymphocytes. Subsequently, its cytotoxic effects on PNT2 (normal prostate) cells was determined using the cell viability test (MTT) and compared with those induced by different doses of X-rays.

Results:

A dose- and time-dependent cytotoxic effect of sevofluorane on PNT2 cells was determined (p >0.001) and a dose-dependent genotoxic effect of sevofluorane was established (p >0.001). Hovewer, at volumes lower than 30 μL of sevofluorane at 100%, a non-toxic effect on PNT2 cells was shown.

Conclusion:

Sevofluorane demonstrates a genotoxic capacity as determined in vitro by micronucleus test in cytokinesis-blocked cells of irradiated human lymphocytes.     

Use of the fluorescent micronucleus assay to detect the genotoxic effects of radiation and arsenic exposure in exfoliated human epithelial cells

The exfoliated cell micronucleus (MN) assay using fluorescent in situ hybridization (FISH) with a centromeric probe is a rapid method for determining the mechanism of MN formation in epithelial tissues exposed to carcinogenic agents. Here, we describe the use of this assay to detect the presence or absence of centromeric DNA in MN induced in vivo by radiation therapy and chronic arsenic (As) ingestion. We examined the buccal cells of an individual receiving 6,500 rads of photon radiation to the head and neck. Exfoliated cells were collected before, during, and after treatment. After radiation exposure a 16.6-fold increase in buccal cell MN frequency was seen. All induced MN were centromere negative (MN −) resulting from chromosome breakage. This finding is consistent with the clastogenic action of radiation and confirmed the reliability of the method. Three weeks post-therapy, MN frequencies returned to baseline. We also applied the assay to exfoliated bladder cells of 18 people chronically exposed to high levels of inorganic arsenic (In-As) in drinking water (average level, 1,312 μg As/L) and 18 matched controls (average level, 16 μg As/L). The combined increase in MN frequency was 1.8-fold (P = 0.001, Fisher's exact test). Frequencies of micronuclei containing acentric fragments (MN −) and those containing whole chromosomes (MN+) both increased (1.65-fold, P = 0.07, and 1.37-fold, P = 0.15, respectively), suggesting that arsenic may have both clastogenic and weak aneuploidogenic properties in vivo. After stratification on sex, the effect was stronger in male than in female bladder cells. In males the MN-frequency increased 2.06-fold (P = 0.07) while the frequency of MN+ increased 1.86-fold (P = 0.08). In addition, the frequencies of MN − and MN+ were positively associated with urinary arsenic and its metabolites. However, the association was stronger for micronuclei containing acentric fragments. By using FISH with centromeric probes, the mechanism of chemically induced genotoxicity can now be determined in epithelial tissues. © 1996 Wiley-Liss, Inc.     

RIP3/MLKL-mediated neuronal necroptosis induced by methamphetamine at 39℃

Methamphetamine is one of the most prevalent drugs abused in the world.Methamphetamine abusers usually present with hyperpyrexia (39℃),hallucination and other psychiatric symptoms.However,the detailed mechanism underlying its neurotoxic action remains elusive.This study investigated the effects of methamphetamine + 39℃ on primary cortical neurons from the cortex of embryonic Sprague-Dawley rats.Primary cortex neurons were exposed to 1 mM methamphetamine + 39℃.Propidium iodide staining and lactate dehydrogenase release detection showed that methamphetamine + 39℃ triggered obvious necrosis-like death in cultured primary cortical neurons,which could be partially inhibited by receptor-interacting protein-1 (RIP1) inhibitor Necrostatin-1 partially.Western blot assay results showed that there were increases in the expressions of receptor-interacting protein-3 (RIP3) and mixed lineage kinase domain-like protein (MLKL) in the primary cortical neurons treated with 1 mM methamphetamine + 39℃ for 3 hours.After pre-treatment with RIP3 inhibitor GSK’872,propidium iodide staining and lactate dehydrogenase release detection showed that neuronal necrosis rate was significantly decreased;RIP3 and MLKL protein expression significantly decreased.Immunohistochemistry staining results also showed that the expressions of RIP3 and MLKL were up-regulated in brain specimens from humans who had died of methamphetamine abuse.Taken together,the above results suggest that methamphetamine + 39℃ can induce RIP3/MLKL regulated necroptosis,thereby resulting in neurotoxicity.The study protocol was approved by the Medical Ethics Committee of the Third Xiangya Hospital of Central South University,China (approval numbers: 2017-S026 and 2017-S033) on March 7,2017.