Benzo(a)pyrene Group Research Articles

MicroRNAs from Yishen Tongluo formula can repair sperm DNA damage caused by benzo(a)pyrene.

Plant microRNAs (miRNAs) present in Yishen Tongluo formula (YSTL, a traditional Chinese herbal medicine formula) are considered as potential therapeutic drugs for reducing the sperm DNA fragmentation index (DFI). To study the effectiveness of plant miRNAs in YSTL for repairing mouse sperm DNA damage caused by benzo(a)pyrene (BaP). Twenty-four male SPF ICR (CD1) mice were divided into control, BaP and YSTL groups. A BaP-induced (100 mg/kg) sperm DNA damage model was established in the BaP and YSTL groups, and the mice in the YSTL group were treated with YSTL (23.78 g/kg) for 8 weeks. Sperm DFI was determined via a sperm chromatin structure assay (SCSA). MicroRNAs in the testes of the mice were analysed via RNA-seq, and the top four plant miRNAs were screened, identified and overexpressed in GC cells. The effects of plant miRNAs on the viability and DNA integrity of GC cells exposed to benzo(a)pyrene diol epoxide (BPDE) (1 μM) were tested using CCK8 and comet assays. Compared with that of the BaP group, the DFI of the YSTL group decreased (9.57% vs. 18.54%, F = 18.645, p = 0.0236). miR166-y, miR894-x, miR822-x and miR396-x were screened. The CCK8 and comet assays revealed that the DFI of the mimic group was significantly lower than that of the BPDE (IC50 = 1.006 μM) group, with the most significant difference in the miR396-x group. Plant miRNAs such as miR396-x can penetrate the blood-testis barrier through the digestive system to repair sperm DNA.

Prenatal exposure to Benzo[a]pyrene affects maternal–fetal outcomes via placental apoptosis

Prenatal exposure to Benzo[a]pyrene (BaP) has been suggested to increase the risk of adverse pregnancy outcomes. However, the role of placental apoptosis on BaP reproductive toxicity is poorly understood. We conducted a maternal animal model of C57BL/6 wild-type (WT) and transformation-related protein 53 (Trp53) heterozygous knockout (p53KO) mice, as well as a nested case–control study involving 83 women with PB and 82 term birth from a birth cohort on prenatal exposure to BaP and preterm birth (PB). Pregnant WT and p53KO mice were randomly allocated to BaP treatment and control groups, intraperitoneally injected of low (7.8 mg/kg), medium (35 mg/kg), and high (78 mg/kg) doses of 3,4-BaP per day and equal volume of vegetable oil, from gestational day 10.5 until delivery. Results show that high-dose BaP treatment increased the incidence of preterm birth in WT mice. The number of fetal deaths and resorptions increased with increasing doses of BaP exposure in mice. Notably, significant reductions in maternal and birth weights, increases in placental weights, and decrease in the number of livebirths were observed in higher-dose BaP groups in dose-dependent manner. We additionally observed elevated p53-mediated placental apoptosis in higher BaP exposure groups, with altered expression levels of p53 and Bax/Bcl-2. In case–control study, the expression level of MMP2 was increased among women with high BaP exposure and associated with the increased risk of all PB and moderate PB. Our study provides the first evidence of BaP-induced reproductive toxicity and its adverse effects on maternal–fetal outcomes in both animal and population studies.

Benzo(a)pyrene exposure impacts cerebrovascular development in zebrafish embryos and the antagonistic effect of berberine

Polycyclic aromatic hydrocarbons (PAHs) widely present in the environment, but their effect on cerebrovascular development has been rarely reported. In this study, dechorionated zebrafish embryos at 24 hpf were exposed to benzo(a)pyrene (BaP) at 0.5, 5 and 50 nM for 48 h, cerebrovascular density showed a significant reduction in the 5 and 50 nM groups. The expression of aryl hydrocarbon receptor (AhR) was significantly increased. Transcriptomic analysis showed that the pathway of positive regulation of vascular development was down-regulated and the pathway of inflammation response was up-regulated. The transcription of main genes related to vascular development, such as vegf, bmper, cdh5, f3b, itgb1 and prkd1, was down-regulated. Addition of AhR-specific inhibitor CH233191 in the 50 nM BaP group rescued cerebrovascular developmental defects and down-regulation of relative genes, suggesting that BaP-induced cerebrovascular defects was AhR-dependent. The cerebrovascular defects were persistent into adult fish raised in clean water, showing that the relative area of vascular network, the length of vessels per unit area and the number of vascular junctions per unit area were significantly decreased in the 50 nM group. Supplementation of berberine (BBR), a naturally derived medicine from a Chinese medicinal herb, alleviated BaP-induced cerebrovascular defects, accompanied by the restoration of altered expression of AhR and relative genes, which might be due to that BBR promoted BaP elimination via enhancing detoxification enzyme activities, suggesting that BBR could be a potential agent in the prevention of cerebrovascular developmental defects caused by PAHs.

Transcriptomic and proteomic features of a mouse model of sperm DNA damage induced by benzo(a)pyrene

This study replicated a mouse model of sperm DNA damage induced by benzo(a)pyrene (BaP), and the transcriptomic and proteomic features of the model were examined to clarify the pathways related to BaP-induced damage to sperm DNA. Male mice in the BaP group were subjected to BaP at a dosage of 100 mg/kg/d or an equivalent quantity of saline solution in the control group for 60 days. Subsequently, the DNA fragmentation index (DFI) in sperm was assessed using a sperm chromatin structure assay (SCSA). RNA-seq and data-independent acquisition (DIA) were used to identify the mRNA and protein expression patterns in the testis. The sperm DFI significantly increased in the BaP group. Compared to the control group, the BaP group exhibited differential expression of 240 genes (referred to as DEGs) and 616 proteins (referred to as DEPs). These molecules included Aldh1a1, Cyb5r3, Fads1, Oxsm, Rcn3, and Prss45. Pathways in cancer, the PI3K-Akt signaling pathway, metabolic pathways, and the MAPK signaling pathway were the primary areas where these genes showed enrichment. BaP can damage the DNA of sperm and affect metabolism, the PI3K-Akt pathway, and pathways associated with cancer signaling.

Nitro substituent caused negative impact on occurrence and development of atherosclerotic plaque by PM2.5-bound polycyclic aromatic compounds

PM2.5 exposure is a significant risk factor for the occurrence and development of atherosclerosis. Polycyclic aromatic hydrocarbons (PAHs) play prominent roles in PM2.5-related toxicity. However, the nitrated derivatives of PAHs, nitrated polycyclic aromatic hydrocarbons (NPAHs), have strong oxidizing properties due to the nitro substituents. Thus, the in vivo and in vitro experiments exposure to benzo[a]pyrene (BaP) and 6-nitro benzo[a]pyrene (NBaP) were conducted to evaluate the effect of nitro substituent on the atherosclerosis due to (or attributable to) PAHs. The results showed that NBaP exposure induced the inhibition of human umbilical vein endothelial cells (HUVECs) viability and cell morphology damage via more severe oxidative stress than BaP exposure. Furthermore, exposure to PM2.5-bound NBaP caused dyslipidemia in the Apolipoprotein E-deficient (ApoE−/−) mice, including the increment of total cholesterol, triglycerides, low-density lipoprotein cholesterol, and malondialdehyde levels, and the decrement of high-density lipoprotein cholesterol levels, superoxide dismutase and glutathione peroxidase levels in serum and aorta. Furthermore, histology showed atherosclerotic plaque in the aorta of ApoE−/− mice. However, there were no significant differences of the physiological and pathological changes between BaP and control groups. Thus, NPAHs induced endothelial dysfunction and dyslipidemia via severe oxidative stress, and further accelerated the occurrence and development of atherosclerosis compared with the parent PAHs. Our findings provide the first evidence that nitro substituent caused much severer negative health impact of polycyclic aromatic compounds, which highlight the significance of NPAHs in health risk estimation of polycyclic aromatic compounds.

Cucumis sativus (Cucurbitaceae) seed oil prevents benzo(a)pyrene-induced prostate cancer in vitro and in vivo.

Despite enormous progress in modern medicine, prostate cancer (PCa) remains a major public health problem due to its high incidence and mortality. Although studies have shown in vitro antitumor effects of cucurbitacins from Cucumis sativus, the in vivo anticancer effect of the seed oil as a whole, has yet to be demonstrated. The present study evaluated the in vitro anticancer mechanisms of C. sativus (CS) seed oil and its possible chemopreventive potential on benzo(a)pyrene (BaP)-induced PCa in Wistar rat. In vitro cell growth, clone formation, cell death mechanism, cell adhesion and migration as well as expression of integrins β-1 and β-4 were assessed. In vivo PCa was induced in 56 male rats versus 8 normal control rats, randomized in normal (NOR) and negative (BaP) control groups which, received distilled water; the positive control group (Caso) was treated with casodex (13.5 mg/kg BW). One group received the total seed extract at the dose of 500 mg/kg BW; while the remaining three groups were treated with CS seed oil at 42.5, 85, and 170 mg/kg BW. The endpoints were: morphologically (prostate tumor weight and volume), biochemically (total protein, prostate specific antigen (PSA), oxidative stress markers such as MDA, GSH, catalase, and SOD) and histologically. As results, CS seed oil significantly and concentration-dependently reduced the DU145 prostate cancer cell growth and clone formation (optimum = 100 μg/mL). It slightly increased the number of apoptotic cells and inhibited the migration and invasion of DU145 cells, while it decreased their adhesion to immobilized collagen and fibrinogen. The expression of integrin β-1 and β-4 was increased in presence of 100 μg/mL CS oil. In vivo, the BaP significantly elevated the incidence of PC tumors (75%), the total protein and PSA levels, pro-inflammatory cytokines (TNF-α, IL-1, and IL-6) and MDA levels compared to NOR. CS seeds oil significantly counteracted the effect of BaP by decreasing significantly the PC incidence (12.5%), and increasing the level of antioxidant (SOD, GSH, and catalase) and anti-inflammatory cytokine IL-10 in serum. While in BaP group PCa adenocarninoma was the most representative neoplasm, rats treated with 85 and 170 mg/kg prevented it in the light of the casodex. It is conclude that CS may provide tumor suppressive effects in vitro and in vivo which makes it an interesting candidate to support the current treatment protocol.

Electron microscopic investigation of benzo(a)pyrene-induced alterations in the rat kidney tissue and the protective effects of curcumin

ABSTRACT Benzo(a)pyrene (BaP) is a polycyclic hydrocarbon with carcinogenic and DNA damaging properties. Curcumin, primary yellow pigment in turmeric, has a wide range of biological, pharmacological properties in addition to being a powerful antioxidant. The aim of this study was to investigate protective effects of curcumin against benzo(a)pyrene damage in rat kidney. Thirty-six male Wistar albino rats were divided into six groups (n = 6) as: control, corn oil, Dimethyl sulfoxide (DMSO), BaP (10 mg/kg/day), Curcumin (100 mg/kg/day), Curcumin+BaP (100 mg/kg/day+10 mg/kg/day). Agents were daily and orally administered for six weeks. Kidney tissues were removed and examined ultrastructurally. Glomerular and tubular structures in control, corn oil, and DMSO groups demonstrated normal features. Glomerular capillary dilation, thickening, and folding of basement membrane and disruption of organelle contents were distinguished in BaP group. Deletion of podocyte cell and pedicels also sponge-like appearance of glomerular surface were remarkable in this group. Tissue components were protected in curcumin treated group. Proximal tubules and glomerular basement membrane exhibited normal features in Curcumin+BaP group. The abnormalities that accompanied BaP administration clearly revealed the detrimental effects of this agent. Therefore, this study provided substantial evidence that curcumin protects against benzo(a)pyrene nephrotoxicity.

Role and mechanism of benzo[a]pyrene in the transformation of chronic obstructive pulmonary disease into lung adenocarcinoma.

This experiment is explores the genes that play a key role, their expression changes and the biological processes in the transformation of chronic obstructive pulmonary disease (COPD) into lung adenocarcinoma (LAC). Meanwhile, identify the effects of Benzo[a]pyrene (BaP) in the conversion of COPD into LAC. 1. Differential expression genes of COPD and LAC were screened and analyzed by high-throughput microarray data between the two diseases and their respective control groups. 2. The screened genes were used for routine bioinformatics analysis such as functional analysis, expression verification, protein interaction analysis and functional enrichment. 3. Cigarette smoke extract (CSE) combined with lipopolysaccharide (LPS) was used to establish an in vitro COPD model. 4. MTT assay was used to detect the influence of B(a)P in effect on A549 cell proliferation. CCK-8, Transwell invasion test and scratch test were used to detect the cell proliferation, invasion and migration ability, while qPCR and Western Blot tests were used to observe the cell proliferation, apoptosis and changes in related indicators such as EMT. 5. Experimental method of separately adding agonists (tBHQ) and inhibitors (DIC) of NQO1 was used to confirm the effect of NQO1 on A549 cell proliferation, apoptosis, migration and invasion. 6. To further clarify whether BaP exerted effect on cell proliferation, apoptosis, migration and invasion through NQO1, we knocked down NQO1 gene and then infecting cells with BaP. 1. We screened genes of COPD and LAC using datasets from GSE151052, GSE118370, and GSE140797. After screening, the genes upregulated in COPD and downregulated in LAC were RTKN2, SLC6A4, and HBB, the gene downregulated in COPD and upregulated in LAC was NQO1, the genes downregulated in both COPD and LAC were FPR1, LYVE1 and PKHD1L1. 2. The main signaling pathways in which the target genes were enriched are cell cycle, EMT, PI3K/AKT, and apoptosis. In the data included GEPIA, PKHD1L1, FPR1, LYVE1, RTKN2, HBB, and SLC6A4 were significantly downregulated and NQO1 was upregulated in LAC relative to controls. In addition, there were 46 interaction proteins in the target genes, and the functions they enriched included hydrogen peroxide catabolism, etc. 3. When A549 cell was stimulated with 100ng/mL LPS+ 10% CSE, the COX-2 expression indicated that COPD model in vitro was successfully established. 4. The optimal dose and action time were screened which were 1μM and 24h. Compared to the control group, COPD and BaP group increased cell proliferation and invasion capabilities. On the basis of COPD, adding BaP could further increase the proliferation and migration capabilities. Interestingly, the levels of NQO1 decreased in COPD models, while increased by BaP. 5. tBHQ can increase the proliferation and migration capacity of A549 cells, which is inhibited by the addition of DIC. 6. The enhanced proliferation, migration and invasion of A549 cells by BaP were attenuated after knockdown of NQO1. Our study reveals that PKHD1L1, FPR1, LYVE1, RTKN2, HBB, SLC6A4 and NQO1 may play an important role in the conversion of COPD to LAC. High NQO1 expression may increase the proliferation and migration ability of A549 cells, and BaP may promote the EMT state by increasing the expression of NQO1, thereby making the COPD model in vitro expose the tumor characteristics.

Effects of dietary whole grain buckwheat and oat on benzo[a]pyrene-induced genotoxicity, oxidative and pyroptotic injury in liver of mice

Benzo[a]pyrene (BaP), as a highly toxic substance, exists in the surrounding and foods of people's life and poses a great threat to the human body. Our previous studies have shown that BaP caused oxidative and pyroptotic damage to the liver, which is an urgent problem to be solved. Whole grains not only contain a series of important nutrients, but also have important significance for preventing obesity, dyslipidemia and inflammation. Therefore, this work tried to use whole grain diet as a new method to inhibit the BaP-induced toxicity in mice. Compared with BaP group, the addition of whole grain buckwheat (WGB) or oat (WGO) diet could improve the appetite of mice, stabilize the levels of ALT, AST, TC and TG in serum, maintain the balance of H2O2, MDA, GSH-Px and SOD levels in liver tissues, reduce BPDE-DNA adduct content and the mRNA expression of BaP metabolites related enzyme (including CYP1A1, CYP1B1, GST-P1 and AhR). Furthermore, both WGB and WGO could inhibit the pyroptotic markers (NLRP3, Caspase-1, GSDMD) increase and inflammatory factors release, promote the synthesis of Short-chain fatty acids (SCFAs), and the effect of WGO was more excellent than WGB. This study fully verified that the reverse effect of the dietary whole grain on BaP-induced genotoxicity, oxidative and pyroptotic injury in liver of mice, and provided a powerful theoretical support for new toxicity control dietary methods in the future.

Long-term co-exposure DBP and BaP causes imbalance in liver macrophages polarization via activation of Notch signaling regulated by miR-34a-5p in rats

Humans are often exposed to complex mixtures of environmental pollutants over long periods of time. It is reported that Dibutyl phthalate (DBP) and benzo[a]pyrene (BaP) are typical environmental pollutants, which are associated with liver injury. Nevertheless, little is known about the effects of DBP and BaP combined exposure on liver. In the current study, rats were exposed to DBP alone (50, or 250 mg/kg), BaP alone (1, or 5 mg/kg), or DBP and BaP (50 + 1, or 250 + 5 mg/kg) for ninety days. More serious liver damage, including abnormal liver function, infiltration of inflammatory cells and disturbed secretion of inflammatory factors, were observed in long-term co-exposure to DBP and BaP group relative to those in single exposure group. Our data showed that long-term co-exposure to DBP and BaP induces macrophages to polarize toward M1 and inhibits polarization of M2 macrophages. Long-term co-exposure to DBP and BaP downregulated miR-34a-5p level and upregulated Notch signaling. These results indicated that imbalance in macrophages M1/M2 polarization mediated by activation of Notch signaling due to reduced miR-34a-5p level may contribute to additive effects on disorder of inflammatory factors secretion and subsequent liver injury following long-term DBP and BaP co-exposure.

Particulate matter 2.5 induced hyperpigmentation in reconstructed human epidermis model (MelaKutis®).

Particulate matter 2.5 induced hyperpigmentation in reconstructed human epidermis model (MelaKutis®).

Paeoniflorin improves the in vitro maturation of benzo(a)pyrene treated porcine oocytes via effects on the sonic hedgehog pathway

Benzo(a)pyrene (BaP) is a toxic substance that people are often exposed to. It has serious harmful effects on the body, and has a destructive effect on oocytes and cumulus cells. Here, we found that paeoniflorin (Pae), a traditional Chinese medicine monomer with antioxidant effects, decreased BaP-induced meiotic failure by increasing the activity of the Sonic hedgehog (SHH) signaling pathway and reducing the level of reactive oxygen species (ROS). We found that the in vitro maturation (IVM) rate was significantly increased (P < 0.05) in the 0.1 μM Pae and BaP (co-treatment) group compared with BaP group due to reduced ROS levels and increased mitochondrial membrane potential (ΔΨ) and ATP content. The mRNA expression levels of oocyte maturation and cumulus cell expansion-related genes were also significantly higher in the co-treatment group. To demonstrate the quality of oocytes, the development capacity of parthenogenetically activated (PA) and in vitro fertilization (IVF) embryos from different treatment groups oocytes were determined.The blastocyst formation rate was significantly higher in PA and IVF embryos derived from oocytes in the co-treatment group than in those derived from oocytes in the BaP group. To further confirm that the SHH signaling pathway was involved in causing these effects of Pae, we treated oocytes with Pae and BaP in the presence or absence of cyclopamine (Cy), an inhibitor of this pathway. Cy abolished the effects of Pae in BaP treated porcine oocytes. In conclusion, Pae improves the IVM capacity of BaP-treated porcine oocytes by activating the SHH signaling pathway, inhibiting ROS production, and increasing ΔΨ.

Role of NF-κB p65 and related cytokines in rats with liver function injury induced by dibutyl phthalate and benzo (a) pyrene

Objective: To explore the role of nuclear factor-κB (NF-κB) p65 and related cytokines in rats with liver function injury induced by dibutyl phthalate (DBP) and benzo (a) pyrene (BaP) , in order to provide support for enriching the mechanism of liver injury induced by DBP and BaP. Methods: In September to December of 2019, a total number of 160 specific pathogen free Sprague Dawley rats were numbered in order of sex and body weight, then using the statistical table of random numbers, they were randomly divided into eight groups and each group consists of twenty animals (10 male and 10 female rats) , including blank control group, vehicle control group (given corn oil) , DBP 50 mg/kg (DBP(50)) group, DBP 250 mg/kg (DBP(250)) group, BaP 1 mg/kg (BaP(1)) group, BaP 5 mg/kg (BaP(5)) group, DBP 50 mg/kg plus BaP 1 mg/kg (DBP(50)+BaP(1)) group and DBP 250 mg/kg plus BaP 5 mg/kg (DBP(250)+BaP(5)) group, then DBP and BaP were administered to rats as a homogenous mixture in corn oil by gavage. After exposure for 90 days, liver was separated to test the mRNA and protein expression levels of NF-κB p65 by Real-time fluorescence quantitative polymerase chain reaction and Western blotting. Then serum of rats was collected to detect the levels of CXCL-13, interleukin-6 (IL-6) and tumor necrosis factor-alpha (TNF-α) by Enzyme-Linked Immunosorbent Assay, and the levels of alanine aminotransferase (ALT) , aspartate aminotransferase (AST) , albumin (ALB) and total protein (TP) were detected by Reitman-Frankel assay. Results: The protein expression of NF-κB p65 in BaP(1) group was not statistically significant, but the mRNA and protein expression levels of NF-κB p65 in the liver tissues of rats in other exposure group were higher than those in the blank control group (P<0.05) , and the expression levels of NF-κB p65 increased more obvious in the DBP and BaP co-exposed groups than those in the low and high dose groups that single-exposed to DBP and BaP (P<0.05) . The serum levels of CXCL-13 and IL-6 of rats in other group were obviously higher than those of the blank control group except for the BaP(1) group, and the increase was more obvious in the high-dose group that co-exposed to DBP and BaP (P<0.05) . While the level of TNF-α in each exposure group was higher than those in the blank control group and the levels of TNF-α in the DBP and BaP co-exposed groups were strongly augmented compared to those in the low and high dose groups that single-exposed to DBP and BaP (P<0.05) . What's more, compared with the blank control group, the level of ALT in each exposure group was increased significantly. Except for the BaP(1) group, the levels of AST in other exposed groups were increased (P<0.05) , and the levels of ALT and AST in the DBP and BaP co-exposed groups were significantly elevated in comparison to the low and high dose groups that single-exposed to DBP and BaP (P<0.05) . On the contrary, the level of ALB in each exposure group was significantly lower than that in the blank control group, especially decreased significantly in the DBP and BaP co-exposed group (P<0.05) . The level of TP decreased only in the high-dose group that single and co-exposed to DBP and BaP, and the decrease was more significant in the DBP and BaP co-exposed group (P<0.05) . When DBP exposed alone, Pearson correlation analysis showed that NF-κB p65 protein expression level was positively correlated with IL-6, TNF-α and ALT (r=0.762, 0.951, and 0.924, P<0.05) . After BaP exposed alone, the NF-κB p65 protein expression level was positively correlated with TNF-α and ALT (r=0.911 and 0.910, P<0.05) . When DBP and BaP exposed together, NF-κB p65 protein expression level was positively correlated with CXCL-13, IL-6, TNF-α, ALT and AST (r=0.711, 0.764, 0.955, 0.903 and 0.827, P<0.05) . In addition, Pearson correlation analysis showed a positive correlation between TNF-α and ALT (r=0.833 and 0.894, P<0.05) when DBP or BaP exposed alone. Furthermore, when DBP and BaP exposed together, CXCL-13, IL-6 and TNF-α were positively correlated with ALT (r= 0.871, 0.925 and 0.942, P<0.05) , and also positively correlated with AST (r=0.910, 0.892 and 0.890, P<0.05) . Conclusion: Single and co-exposed to DBP and BaP may regulate the abnormal secretion of related cytokines by upregulating the expression level of NF-κB p65 in rat liver tissue, thus leading to hepatocyte injury in rats, and the damage effect may be enhanced when DBP and BaP are exposed together.

Co-exposure to different sized polystyrene microplastics and benzo[a]pyrene affected inflammation in zebrafish andbronchial-associated cells

In recent years, emerging research has revealed that microplastics (MPs) can pollute the air. Cellulose and plastic fibers have been found in the lungs of patients with lung cancer, indicating that MPs can “travel” through the atmosphere into the respiratory tract. Due to the hydrophobicity and relatively large surface area of MPs, the biotoxicity of MPs is greatly enhanced by their adsorption and enrichment in persistent organic pollutants (POPs). With the rapid development of the economy, emissions from coal burning and automobile exhaust have made polycyclic aromatic hydrocarbon (PAHs) one of the key pollutants in urban air that cannot be ignored. Epidemiological studies have shown that PAH exposure is associated with the development of respiratory diseases, such as bronchitis and asthma, in children. The aim of this study was to investigate the inflammatory effects of co-exposure to different size polystyrene MPs (PS-MPs) and benzo[a]pyrene (BaP) on adult zebrafish gills and bronchial-related cells to provide basic information for assessment of the potential health risks of combined exposure to the human respiratory system. PS-MPs (with size of 0.5 and 5 μm) and BaP were co-administered to embryos for 7 d, adult zebrafish for 14 d, and HBE cells and HBMSCs for 48 h. The relative expression levels of inflammatory cytokine genes in the different exposure groups were determined by real-time polymerase chain reaction (PCR). The ingestion of microplastics into fish bodies and cells was observed by stereoscope and confocal microscopy, respectively. Result shows that microplastics of two sizes could be ingested by zebrafish embryos. When exposed only to 0.5-μm PS-MPs, relative expression of the TNF-α , IL-6 , and IL-8 genes was significantly increased compared with that in the control; relative expression of the IFN-γ gene was largely increased in the gills of zebrafish in 5-μm PS-MP-exposed group. Moreover, relative expression of the IFN-γ , IL-6 and IL-8 genes in the group co-exposed to BaP and 0.5-μm PS-MPs was higher than that in BaP group. In addition, 0.5-μm PS-MPs could be ingested by both cell lines. Upon exposure to 0.5-μm PS-MPs alone, relative expression of genes TNF-α , IL-6 , and IL-8 was considerably increased compared with that in the control HBE cells. In HBSMCs, upon exposure to 0.5-μm PS-MPs alone, relative expression of the TNF-α , IL-8 , and IL-1β genes was substantially increased compared with that in the control group. Compared with that following exposure to BaP alone, relative expression of the IFN-γ and IL-6 genes in HBSMCs in 0.5-μm PS-MPs and BaP co-exposure group was remarkably increased, indicating a possible combined effect; however, co-exposure of 5-μm PS-MPs and BaP weakened the proinflammatory function of BaP and especially influenced relative expression of the TNF-α and IL-8 genes. In addition, none of the co-exposure groups of HBE cells showed obvious combined effects. The corollary is that particle size may affect the ingestion of PS-MPs in cells. In addition, the size-dependent effects of PS-MPs influence the accumulation of organic pollutants and their biotoxicity.

The protective effect of melatonin on benzo(a)pyrene-induced brain injury: role of apoptosis and autophagy pathways.

Benzo(a)pyrene (BaP), a toxic polycyclic aromatic hydrocarbon, is spread in different ways as an environmental pollutant. It has been proposed that BaP can induce toxicity through oxidative stress and apoptosis in vital organs. The present study evaluated the protective effect of melatonin, a circadian hormone of the pineal gland, on BaP-induced neurotoxicity focused on oxidative stress, autophagy, and apoptosis pathways. Thirty male mice in 5 groups were treated daily for 28 consecutive days: (I) control group (BaP and melatonin solvent), (II) BaP (75mg/kg, orally), (III) and (IV) BaP + melatonin (10 and 20mg/kg, i.p.), (V) melatonin (20mg/kg). The oxidative stress markers were determined in the brain. Western blot was conducted for the level of LC3 II/I and Beclin1, as autophagy markers, caspase3 and Bcl2, as apoptosis proteins, and Sirt1 in the brain. The exposure of mice to BaP caused a marked increase in the malondialdehyde (MDA) level and decrease of glutathione (GSH) content in the brain. Furthermore, the Sirt1 level upregulated as well as LC3 II/I, Beclin1, and cleaved caspase3 proteins, while the level of Bcl2 did not change. Melatonin at 20mg/kg concurrently with BaP restored the BaP alteration in the brain compared with the BaP group. In conclusion, BaP induced brain toxicity via the induction of oxidative stress, apoptosis, and autophagy, whereas melatonin afforded neuroprotection against BaP due to inhibition of these mechanisms.

Curcumin, an Active Constituent of Turmeric Spice: Implication in the Prevention of Lung Injury Induced by Benzo(a) Pyrene (BaP) in Rats.

Benzo(a)pyrene (BaP) is a well-known carcinogen and enhances oxidative stress and apoptosis and also alters several molecular pathways. Curcumin is an active ingredient of Curcuma longa, and it has potent anti-inflammatory, antioxidant activity that defends cells from oxidative stress and cell death. The objectives of the present study were to explore the protective effects of curcumin against long-term administration of BaP induced disturbances in lungs of rats. Male rats were randomly divided into four groups: saline control, BaP only, BaP + curcumin, and curcumin only. Lung histopathology, electron microscopy, inflammatory cytokine release, antioxidant levels, apoptosis, and cell cycle were examined. Instillation of BaP significantly increased infiltration of inflammatory cells in alveolar space and inflammatory cytokine in blood. BaP induced lung tissue alterations including mild bronchitis, scant chronic inflammatory cell infiltrate in the wall of the respiratory bronchiole, and mild intra-alveolar haemorrhage. However, these alterations were found to be significantly less as mild inflammatory cell infiltrate in curcumin plus BaP treated group. Furthermore, electron microscopy results also showed necrotic changes and broken cell membrane of Type-II epithelial cell of alveoli in BaP group, which was reduced after adding curcumin treatment. In addition, we found BaP plus curcumin treatment effectively reduced inflammatory cytokines Tumour Necrosis Factor alpha (TNF-α), Interleukin 6 (IL-6), and C-reactive protein (CRP) levels in blood serum. Moreover, the levels of tunnel staining and p53 expression were significantly increased by BaP, whereas these changes were noticeably modulated after curcumin treatment. BaP also interferes in normal cell cycle, which was significantly improved with curcumin treatment. Overall, our findings suggest that curcumin attenuates BaP -induced lung injury, probably through inhibiting inflammation, oxidative stress and apoptosis in lung epithelial cells, and improving cell proliferation and antioxidants level. Thus, curcumin may be an alternative therapy for improving the outcomes of Benzo(a)pyrene-induced lung injury.

A40 Antioxidant, Anti-Inflammatory, and Antiapoptotic Potential of Curcumin in Benzo(a)pyrene (BaP)-Induced Lung Injury in Rats

Benzo(a)pyrene (BaP) is a well-known pollutant that directly induces inflammatory microenvironment in the lung. It also enhances oxidative stress and apoptosis and interferes with several other molecular pathways including cell death, survival, and proliferation that disturb normal homeostasis of the lung. Curcumin (Cur) has potent anti-inflammatory, antioxidant activity that defends cells from oxidative stress and cell death. The objectives of the present study were to explore the protective effects of curcumin against long-term administration of BaP-induced disturbances in lungs of rats. Male rats were randomly divided into four groups: saline control, BaP only, BaP + Cur, and Cur only. Lung injury histopathology, electron microscopy, inflammatory cytokine release, antioxidant levels, apoptosis, and cell cycle were examined. Instillation of BaP significantly increased infiltration of inflammatory cells in alveolar space and inflammatory cytokine in blood. Histopathologic examination found BaP-induced pulmonary inflammatory changes were improved after administration of curcumin as evident by less infiltration of macrophages and neutrophils in alveolar space, less deposition of collagen, and edema. Furthermore, electron microscopy results also showed necrotic changes and broken cell membrane of Type II epithelial cell (T2E) of alveoli in BaP group, which was reduced after addition of curcumin treatment. In addition, we found BaP plus curcumin treatment effectively reduced inflammatory cytokines TNF-α and IL-6 in blood serum, but no significant changes were found in CRP levels. Moreover, the levels of tunnel staining and p53 expression were significantly increased by BaP, whereas these changes were noticeably modulated after curcumin treatment. Bap also interferes in normal cell cycle, which was markedly improved with curcumin treatment. Overall. these findings suggest that curcumin attenuates BaP-induced lung injury, probably through inhibiting inflammation, oxidative stress, and apoptosis in lung epithelial cells, and improving cell proliferation and antioxidants' level. Thus, curcumin may be an alternative therapy for improving the outcomes of benzo(a)pyrene-induced lung injury.

MRNA expression of CYP4 in marine polychaete Marphysa sanguinea in response to benzo[a]pyrene

Rapid amplification of cDNA ends (RACE) and real-time polymerase chain reaction (RT-PCR) were carried out to analyze the CYP4 gene expression in polychaete Marphysa sanguinea exposed to benzo[a]pyrene (BaP) in this study. The full length of MsCYP4 cDNA was 2 470 bp, and it encoded 512 amino acids. The deduced amino acid sequence showed 47% identity with CYP4F from frog Xenopus tropicalis and shared high homology with other known CYP4 sequences. To analyse the role of CYP4 in protecting M. sanguinea from BaP exposure, three BaP groups were established: 0.5, 5 and 50 μg/L. Polychaetes were sampled after 3, 7 and 12 d. At 0.5 μg/L, the effect of BaP on MsCYP4 gene expression increased with time prolonged. MsCYP4 gene expression curve showed U-shaped trend with time in 5 and 50 μg/L BaP groups. Therefore, MsCYP4 gene may play an important role in maintaining the balance of cellular metabolism and protecting M. sanguinea from BaP toxicity.

Benzo(a)pyrene inhibits endometrial cell apoptosis in early pregnant mice via the WNT5A pathway.

Benzo(a)pyrene (BaP) is an endocrine-disrupting pollutant present in various aspects of daily life, and studies have demonstrated that BaP exerts reproductive toxicity. We previously showed that BaP damages endometrial morphology and decreases the number of implantation sites in early pregnant mice, but the mechanisms underlying these effects remain unclear. The endometrial function is crucial for implantation, which is associated with endometrial cell apoptosis. In this study, we focused on the effect of BaP on endometrial cell apoptosis and the role of WNT signaling during this process. Pregnant mice were gavaged with corn oil (control group) or 0.2 mg·kg-1 ·day -1 BaP (treatment group) from Days 1 to 6 of pregnancy. BaP impaired endometrial function by decreasing the expression of HOXA10 and BMP2, two markers of receptivity and decidualization. WNT5A and β-catenin were activated in the BaP group. BaP affected the expression of apoptosis-related proteins and inhibited the apoptosis of endometrial stromal cells. In vitro, human endometrial stromal cells (HESCs) were treated with different concentrations of BaP (dimethyl sulfoxide (DMSO); 5, 10 µM). WNT5A and β-catenin were also upregulated in the BaP treatment group. HESC apoptosis was restrained by BaP. Inhibiting WNT5A by SFRP5 partially restored the effect of BaP on apoptosis. In summary, these results suggested that BaP exposure during early pregnancy activates WNT5A/β-catenin signaling pathway, which inhibits the endometrial cell apoptosis and potentially destroys endometrial function.

Alteration of benzo(a)pyrene biotransformation by resveratrol in ApcMin/+ mouse model of colon carcinogenesis.

Epidemiological surveys have revealed that environmental and dietary factors contribute to most of the human cancers. Our earlier studies have shown that resveratrol (RVT), a phytochemical reduced the tumor number, size and incidence of dysplasias induced by benzo(a)pyrene (BaP), an environmental toxicant in the ApcMin/+ mouse model of colon cancer. In this study we investigated to ascertain whether the preventive effects of RVT on BaP-induced colon carcinogenesis is a result of altered BaP biotransformation by RVT. For the first group of mice, 100μg BaP/kg bw was administered in peanut oil via oral gavage over a 60day period. For the second group, 45μg RVT/kg bw was co-administered with BaP. For the third group, RVT was administered for 1week prior to BaP exposure. Blood, colon and liver were collected from control and BaP/RVT-treated mice at 60days post-BaP & RVT exposure. We have assayed activities and expression (protein & mRNA) of drug metabolizing enzymes such as cytochrome P4501A1 (CYP1A1), CYP1B1, and glutathione-S-transferase (GST) in colon and liver samples from the treatment groups mentioned above. An increased expression of CYP1A1 in liver and colon and of CYP1B1 in liver of BaP-treated mice was seen, while RVT inhibited the extent of biotransformation mediated by these enzymes in the respective tissue samples. In the case of GST, an increased expression in colon of BaP alone-treated mice was noted when RVT was administered prior to BaP or simultaneously with BaP. However, there is no change in liver GST expression between BaP and RVT treatment groups. The concentrations of BaP aqueous (phase II) metabolites were found to be greater than the organic (phase I) metabolites, suggesting that RVT slows down the phase I metabolism (metabolic activation) of BaP, while enhancing phase II metabolism (detoxification). Additionally, the BaP-DNA adduct concentrations measured in colon and liver of BaP + RVT-treated mice were low relative to their BaP counterparts. Taken together, our findings strongly suggest that RVT alleviates BaP-induced colon carcinogenesis by impairing biotransformation pathways and DNA adduct formation, and therefore holds promise as a chemopreventive agent.