Youjian Zhang

Activation of the PI3K–AKT–mTOR signaling pathway promotes DEHP-induced Hep3B cell proliferation

Hep3B cells were treated with DEHP at various concentrations (62.5, 125.0, 250.0, 500.0 and 1000.0 μM). After 24 h exposure to DEHP only, increased Hep3B cell viability was observed (p<0.05 or p<0.01). However, after 24 h co-exposure to DEHP at indicated concentrations plus 50.0 μM LY294002 (PI3K inhibitor), cell viability was significantly decreased compared to the corresponding DEHP treated groups. DEHP increased mitochondrial membrane potential level and induced oxidative DNA damage in Hep3B cells, DEHP also increased DNA replication rate and accelerated the cell cycle. The PI3K inhibitor LY294002 could recover the mitochondrial membrane potential and attenuate the oxidative stress in Hep3B cells; however, it could not protect the cells from oxidation of DNA damage. The findings showed that LY294002 attenuated DEHP-induced up-regulation of the selected genes (pi3k, akt, mtor and p70s6k) involved in PI3K-AKT-mTOR signaling pathway at both mRNA and protein levels thus inhibited the cell abnormal proliferation.

Tris(2-chloroethyl)phosphate-induced cell growth arrest via attenuation of SIRT1-independent PI3K/Akt/mTOR pathway

Tris(2‐chloroethyl)phosphate (TCEP) as an organophosphorus flame retardant and plasticizer has been widely used in industrial and household products. It not only was detected in residential indoor air and dust, surface and drinking water, but also in human plasma and breast milk, and tissue samples of liver, kidneys and brain from rodents. TCEP is classified as carcinogenic category 2 and toxic for reproduction category 1B. Sufficient evidence from experimental animals indicated carcinogenicity of TCEP in the liver, and kidneys as well as cell loss in the brain. However, the underlying mechanisms of TCEP‐induced hepatotoxicity are mostly unknown. We investigated the in vitro effects of TCEP as well as TCEP‐induced cell growth in the L02 and HepG2 cells through the PI3K/Akt/mTOR pathway. We found that TCEP reduced cell viability of these cell lines, induced the cell growth arrest, upregulated mRNA and protein levels of SIRT1, and attenuated the PI3K/Akt/mTOR pathway. However, growth arrest of the L02 and HepG2 cells were aggravated after inhibiting the SIRT1 expression with EX‐527. The findings above suggested that TCEP induced the cell growth arrest of L02 and HepG2 cells via attenuation of the SIRT1‐independent PI3K/Akt/mTOR pathway. Copyright

Combined effect of tris(2-chloroethyl)phosphate and benzo (a) pyrene on the release of IL-6 and IL-8 from HepG2 cells via the EGFR-ERK1/2 signaling pathway

Tris(2-chloroethyl)phosphate (TCEP) and benzo (a) pyrene (BaP) coexist in the environment. Humans are exposed to them via multiple routes every day. Each of them induces hepatotoxicity, which may increase their risk to human health. However, the mechanism underlying the combined toxicity of both compounds in vitro is still unclear. The present study aimed to investigate the molecular mechanism underlying the inflammatory response in the cotreatment of HepG2 cells with TCEP and BaP. The cell viability, and the expression of interleukin (IL)-6 and IL-8 at the mRNA and protein levels were measured in HepG2 cells. The results indicated that TCEP plus BaP decreased HepG2 cell viability, and up-regulated the expression of IL-6 and IL-8 at the mRNA and protein levels. Additionally, the inhibitors of EGFR (AG1478), ERK1/2 (U0126) and p38 MAPK (SB203580) displayed anti-inflammatory properties in the inflammatory response elicited by TCEP plus BaP. The activation of ERK1/2, but not p38 MAPK was inhibited by AG1478. These results indicated that TCEP plus BaP may induce an inflammatory response in HepG2 cells by the activation of the EGFR-ERK1/2 signaling pathway.

Tris (2-chloroethyl) phosphate induces senescence-like phenotype of hepatocytes via the p21Waf1/Cip1-Rb pathway in a p53-independent manner

Tris (2-chloroethyl) phosphate (TCEP) has been widely used as a plasticizer and flame retardant. TCEP as a potential carcinogen is often detected in the occupational and nature environments. To investigate effects of TCEP on human hepatocytes, we assessed cell growth rate, cellular membrane integrity, senescence-associated β-galactosidase (SA-β-Gal) activity and analyzed expression of regulators involved in the p53-p21Waf1/Cip1-Rb pathway in TCEP-treated L02 cells. The results showed TCEP increased the percentage of SA-β-Gal positive cells, decreased IL-6 levels, down-regulated the regulators of p38MAPK-NF-κB pathways, but up-regulated the regulators of p21Waf1/Cip1-Rb pathway in L02 cells. Furthermore, we measured the SA-β-Gal activity and expression of regulators involved in the p53-p21Waf1/Cip1-Rb pathway in L02-p53 cells and p53-null Hep3B cells. Similar results were found in L02-p53 cells and Hep3B cells. The findings demonstrated that TCEP induced senescence-like growth arrest via the p21Waf1/Cip1-Rb pathway in a p53-independent manner, without activation of the IL-6/IL6R, p38MAPK-NF-κB pathways in hepatocytes.

Obesity mediated the association of exposure to polycyclic aromatic hydrocarbon with risk of cardiovascular events

Exposure to polycyclic aromatic hydrocarbons (PAHs) could cause high blood pressure (BP) and increased risk for atherosclerotic cardiovascular disease (ASCVD). However, the mechanisms underlying the relationship between them were unclear. We investigated potential mediation effect of obesity on the association of exposure to PAHs with high BP and increased risk for ASCVD. In the repeated measures study, 106 community-dwelling residents in Wuhan, China finished the physical examination in the winter and summer seasons, eight urinary PAHs metabolites were measured. Associations of urinary PAHs with high BP and increased risk for ASCVD were assessed using either linear mixed effect models or generalized estimating equations models. Mediation analysis was performed to evaluate the mediating effect of obesity on the association of urinary PAHs metabolites with high BP or increased risk of ASCVD. We observed the positive association between urinary PAHs metabolites and BP or the odds ratios for high BP (all P<0.05). Additionally, each one-unit increase in ln-transformed urinary levels of 4-hydroxyphenanthrene or the total of PAH metabolites was associated with a 12.63% or 11.91% increase in the estimated 10-year ASCVD risk (both P<0.05). The waist-to-height ratio mediated 29.0% of the association of urinary 4-hydroxyphenanthrene with increased risk of ASCVD (P<0.05). The findings suggest that PAHs exposure may be associated with elevated BP and an increased risk of ASCVD. Obesity may partially mediate the association between PAHs exposure and higher BP or increased risk of ASCVD.

Seasonal variations of tris (2-chloroethyl) phosphate and cytotoxicity of organic extracts in water samples from Wuhan, China

Tris (2-chloroethyl) phosphate (TCEP) is a typical phosphate flame retardant. Its potential adverse health effects have recently aroused concern. We investigated the seasonal variations of TCEP concentrations in the raw, finished and tap water samples from two drinking water treatment plants (DWTPs) in China, and evaluated the cytotoxicity and apoptosis/necrosis of organic extracts (OEs) in water samples. We enriched TCEP and OEs in water samples by solid-phase extraction method. The TCEP concentrations in water samples were determined by gas chromatography-mass spectrometry. Normal human liver cell line L02 was treated with OEs in the water samples, and then the cytotoxicity and apoptosis/necrosis were measured by 3-(4, 5-dimethyithiazol-2-yl)-2,5-diphenyl-tetrazolium bromide assay and flow cytometry, respectively. The results showed that cytotoxicities of OEs in raw water samples from both DWTPs in summer and winter were stronger than those in spring and autumn, cytotoxicity of OEs in finished and tap water samples from both DWTPs in summer and autumn were stronger than those in spring and winter. In all seasons, the maximal concentrations (100 mL water/mL cell culture) of OEs in the raw water samples from both DWTPs induced late apoptosis/necrosis. The reasons for seasonal variations of TCEP in water samples and potential toxic effects of other pollutants in the water samples need to be further investigated.

Dose-response relationships between urinary phthalate metabolites and serum thyroid hormones among waste plastic recycling workers in China

Background Exposure to phthalates may affect thyroid hormone status. However, there were inconsistent observations for the associations of phthalates exposure with altered thyroid hormones. Objectives The aim of this study was to investigate effects of urinary excretion of phthalate metabolites on the levels of thyroid hormones among workers engaged in waste plastic recycling in China. Methods We measured serum levels of thyroid hormones and urinary levels of eight phthalate metabolites among 317 participants (165 workers engaged in waste plastic recycling and 152 farmers), analyzed relationships between urinary phthalate metabolites and thyroid function parameters by multivariate linear regression analysis and structural equation modelling as well as assessed the dose‐response relationships between them by restricted cubic spline functions. Results Maximum urinary values of eight phthalate metabolites in the occupational exposed workers were higher than the controls. Compared with the controls, the workers had higher levels of urinary monobenzyl phthalate (MBzP, 1.12 vs. 0.92 &mgr;g/g creatinine), mono (2‐ethyl‐5‐hydroxyhexyl) phthalate (MEHHP, 38.84 vs. 32.55 &mgr;g/g creatinine), mono‐n‐octyl phthalate (MOP, .11 vs. 0.09 &mgr;g/g creatinine), serum total triiodothyronine (T3, 1.04 vs. 0.92 ng/mL) and the T3 to thyroxine (T4) ratio (1.44 vs. 1.09) (all P < 0.05). The results from structural equation modelling analysis showed that phthalates metabolites were positively associated with total T3 (&bgr; = 0.044, SE = 0.021, P < 0.05) or the T3/T4 ratio (&bgr; = 0.053, SE = 0.022, P < 0.05) among all participants. Among the workers, there were the non‐monotonic dose‐response associations between urinary monomethyl phthalate (MMP) and serum total T3 or the T3/T4 ratio, as well as between urinary monoethyl phthalate (MEP) and the T3/T4 ratio (all P < 0.05). Conclusions The dose‐response relationships between urinary phthalate metabolites and thyroid hormone parameters may be non‐monotonic among the workers. Further investigations are needed to corroborate these findings. HighlightsMaximum urinary phthalate levels of the workers are higher than those of controls.A non‐linear association of urinary MMP with total T3 was found among the workers.The workers showed a non‐linear association of urinary MMP with the T3/T4 ratio.The workers exhibited a non‐linear association of urinary MEP with the T3/T4 ratio.