Effects of Benzo[a]pyrene on Food Metabolism and Reproductive Endocrine and Ovarian Development in Female Scallop Chlamys farreri at Different Reproductive Stages.

Benzo[a]pyrene (B[a]P), a polycyclic aromatic hydrocarbon (PAH) with the most carcinogenic effects of all the PAHs, has multiple toxic effects on marine bivalves. We investigated the interference mechanism of B[a]P on food metabolism (sugars, proteins, and sugars), and on reproductive endocrine and ovarian development in female scallops (Chlamys farreri). Scallops were exposed to different concentrations of B[a]P concentrations of 0, 0.38, 3.8, and 38 µg/L throughout gonadal development. Total cholesterol and triglyceride contents in the digestive glands were increased, and their synthesis genes were upregulated. The plasma glucose contents decreased with the inhibition of glycogen synthesis genes and the induction of glycolysis genes in the digestive gland. The results showed that B[a]P had endocrine-disrupting effects on scallops, that it negatively affected genes related to ovarian cell proliferation, sex differentiation, and egg development, and that it caused damage to ovarian tissue. Our findings supplement the information on B[a]P disruption in gonadal development of marine bivalves. Environ Toxicol Chem 2024;43:748-761. © 2023 SETAC.

Insights into mechanism of DNA damage and repair-apoptosis in digestive gland of female scallop Chlamys farreri under benzo[a]pyrene exposure during reproductive stage.

As one of the most carcinogenic persistent organic pollutants (POPs), benzo[a]pyrene (B [a]P) brings high toxicity to marine bivalves. Digestive gland is the most important metabolism-related organ of aquatic animals. This study conducted the digestive gland transcriptome of Chlamys farreri under B[a]P treatment at reproductive stages. And the reproductive-stage dependence metabolism-DNA repair-apoptosis process of scallops under 0, 0.04, 0.4 and 4 µg/L B[a]P was studied by qRT-PCR. The results demonstrated that the detoxification metabolism was disturbed after ovulation except for CYP3A4. In antioxidant system, antioxidant enzyme CAT and GPX, and GGT1 (one of the non-enzymatic antioxidants synthesis gene) continuously served the function of antioxidant defense. Three types of DNA repair were activated under B[a]P stress, however, DNA strand breaks were still serious. B[a]P exposure weakened death receptor pathway as well as enhanced mitochondrial pathway, surprisingly suppressing apoptosis in scallops. In addition, ten indicators were screened by Spearman correlation analysis. This study will provide sound theoretical basis for bivalve toxicology and contribute to the biomonitoring of marine POPs pollution.

Integrated lipidomics and transcriptomics analysis reveal lipid metabolism disturbance in scallop (Chlamys farreri) exposure to benzo[a]pyrene.

Benzo[a]pyrene (B[a]P) commonly bioaccumulates in lipid-rich tissues due to its lipophilicity and further affects lipid metabolism. The present study systematically investigated the lipid metabolism disturbance in digestive glands of scallops (Chlamys farreri) exposure to B[a]P, based on lipidomics, transcriptomics, molecular and biochemical analysis. We exposed the scallops to environmentally relevant concentrations of B[a]P for 21 days. The bioaccumulation of B[a]P, lipid content and lipid peroxidation in digestive glands were measured. Integrated lipidomics and transcriptomics analysis, the differential lipid species were identified and key genes based on the pathways in which genes and lipid species involved together were selected in scallop exposure to 10 µg/L B[a]P. The changes of lipid profile showed that triglycerides (TGs) were accumulated after 21 days exposure, while the phospholipids (PLs) decreased demonstrated membrane structures were disrupted by B[a]P. In combination with the change of gene expression, we speculated that B[a]P could induce lipids accumulation by up-regulating lipid synthesis-related genes expression, down-regulating lipolysis-related genes expression and interfering with lipid transport. Overall, this study provides new insights into the mechanisms of lipid metabolism disturbance in bivalves exposed to PAHs, and establishes a foundation for understanding the bioaccumulation mechanism of B[a]P in aquatic organisms, which is of great importance for further ecotoxicological study.

Reproductive toxicity induced by benzo[a]pyrene exposure: first exploration highlighting the multi-stage molecular mechanism in female scallop Chlamys farreri.

Reproductive toxicity induced by benzo[a]pyrene (B[a]P) exposure has received great ecotoxicological concerns. However, huge gaps on the molecular mechanism still exist in bivalves. In this study, reproduction-related indicators were investigated in female scallops Chlamys farreri during life cycle of proliferative, growth, mature, and spawn stages, under gradient concentrations of B[a]P at 0, 0.04, 0.4, and 4 µg/L. Meanwhile, a multi-stage ovarian transcriptome analysis under 4 µg/L B[a]P exposure was also conducted to elucidate the potential molecular mechanisms. The results indicated that life-cycle exposure to 0.4 and 4 µg/L B[a]P significantly decreased GSI and sex steroid levels. Even 0.04 µg/L B[a]P could play the adverse role in DNA integrity at the mature and spawn stages. Ovarian histological sections showed that B[a]P inhibited the maturation and release of oocytes. Through the functional enrichment analysis of differentially expressed genes (DEGs) from transcriptome data, 18 genes involved in endocrine disruption effects, DNA damage and repair, and oogenesis were selected and further determined by qRT-PCR. The downregulation of genes involved in steroidogenic and estrogen signaling pathways indicated that B[a]P could cause endocrine disruption through both receptor-dependent and receptor-independent pathways. The variations of gene expressions involved in DNA single-strand break and repair implied the presence of toxic mechanisms similar with vertebrates. Additionally, the changes of gene expressions of cell cycle, apoptosis, and cell adhesion suggested that exposure to B[a]P possibly caused the reproductive toxicity effects by affecting oogenesis. Taken together, this study was a pioneer in combining genome-wide transcriptomic analysis with its corresponding reproductive indicators (GSI, sex steroid levels, DNA single-strand break, and histological sections) to explore the bivalves' toxic mechanisms under B[a]P exposure. Meanwhile, some genes involved in estrogen signaling pathway and DNA damage were firstly analyzed in bivalves, and the expression data might be useful in establishing new hypotheses and discovering new biomarkers for marine biomonitoring.

Insights into disruption of lipid metabolism in digestive gland of female scallop Chlamys farreri under B[a]P exposure.

Lipids are the main energy support during gametogenesis. Digestive gland is the key organ of aquatic animal metabolism for storing nutrition and supplying energy. It participates in a variety of life activities (such as growth, digestion, immunity, and reproduction). Nutrients stored in digestive glands, especially lipids, provide energy for reproductive behaviors such as gametogenesis and ovulation. A large number of studies have confirmed the accumulation of lipids from digestive gland to gonad during gametogenesis. At present, the research on the interference mechanism of persistent organic pollutants (POPs) on lipid metabolism of aquatic animals and the adaptive response of aquatic animals to POPs stress focus on biochemical levels or a few genes. The potential molecular mechanism of lipid metabolism interference needs to be further studied. In addition, as an important stage of aquatic animals, the reproductive period is a vigorous period of lipid metabolism. However, at present, there is no report on the molecular mechanism of POPs interfering with the lipid metabolism of the digestive gland in the reproductive process of aquatic animals. In this study, female scallop C. farreri was cultured in natural seawater and exposed to 4 µg/L B[a]P in seawater. Transcriptome analysis of digestive glands at multiple stages (proliferative stage, growth stage, mature stage and spawn stage) was performed, and iPath pathway analysis was used to analyze lipid metabolism pathways and differential genes. The interference mechanism of lipid metabolism in bivalves during reproductive period was revealed. This study will provide valuable genomic information on the role of digestive glands in lipid metabolism and reproduction of C. farreri, and will contribute to further functional genomics of bivalves and other closely related species.

Gene expression and functional analysis of different heat shock protein (HSPs) in Ruditapes philippinarum under BaP stress.

Heat shock proteins (HSPs) are a class of highly conserved proteins which can protect cells against various types of stress. However, little information on the mechanism involved in the organic contaminants stress response of HSPs is available, especially in marine invertebrates. The present study was conducted to evaluate the responses of HSPs in clams (Ruditapes philippinarum) under Benzo[a] pyrene (BaP) exposure. The clams were exposed to BaP (concentrations: 0, 0.1, 1, 10 µg/L) for 15 days. 6 HSPs mRNA were classified, and the results of tissue distribution indicated that 4 HSPs gene expressed most in the digestive glands. The transcription level of 6 HSPs (HSP22-1, HSP22-2, HSP40A, HSP60, HSP70, HSP90) genes and the aryl hydrocarbon receptor signaling pathway-related genes, and detoxification system-related enzymes activities were analyzed at 0, 1, 3, 6, 10 and 15 days. The activities of phase II detoxification metabolic enzymes and signaling pathway related genes in clams were severely affected by BaP stress and presented significant difference. Our result suggested that HSPs were produced in the presence of BaP and participated in the process of detoxification metabolism to a certain extent. Additionally, the transcription of HSP40A gene may be used as a potential biomarker of BaP exposure due to its evident concentration- and time-dependent expression pattern. Overall, the study investigated the classification of HSPs in R. philippinarum, provided information about the expression profiles of various HSPs after BaP exposure and broadened the understanding mechanism of HSPs in detoxification defense system under PAHs stress in mollusks.

Impacts of benzo(a)pyrene exposure on scallop (Chlamys farreri) gut health and gut microbiota composition.

The gut tissue interacts with nutrients and pollutants which can impact gut health. Gut microbiota is essential to the host health, but is also easily affected by external environment. However, little is known about the toxicological assessment of environmental contaminants on gut health and microbiota, especially in marine invertebrates. In this study, we first explored the effect of benzo(a)pyrene (BaP) on the gut health and gut microbiota of scallops (Chlamys farreri). The scallops were exposed to different concentrations (0, 0.4, 2 and 10 µg/L) of BaP for 21 days. The histological morphology, immune- and oxidative enzyme-related gene expression, and lipid peroxidation of the scallops were analyzed at 7, 14 and 21 days. The results revealed that BaP could impair intestinal barrier function, increasing the intestinal permeability of scallops. Moreover, immune and antioxidant responses were induced in the gut tissue. After a 21-day exposure to different concentrations of BaP, the intestinal microbial community was analyzed based on 16S rRNA sequencing. Our results suggested that BaP exposure altered the gut microbial diversity and composition in scallops. Many beneficial genera declined after BaP treatment, while the potential pathogens were increased, such as Mycoplasma and Tenacibaculum. A series of hydrocarbon-degrading bacteria were recognized in BaP-treated groups, such as Pseudomonas, Polaribacter, Amphritea and Kordiimonas. Interestingly, the degrading bacteria present varied after exposure to different concentrations of BaP. Overall, this study provides new insights into gut health and gut microbiota in marine invertebrates following exposure to persistent organic pollutants.

Damages to biological macromolecules in gonadal subcellular fractions of scallop Chlamys farreri following benzo[a]pyrene exposure: Contribution to inhibiting gonadal development and reducing fertility.

Benzo[a]pyrene (B[a]P), a representative polycyclic aromatic hydrocarbon (PAH) compound in marine ecosystem, has great potential for chronic toxicity to marine animals. It is becoming increasingly apparent that reproductive system is the major target of B[a]P, but the adverse effects of B[a]P on subcellular fractions in bivalve gonads have not been elucidated. Scallops Chlamys farreri are used as the experimental species since they are sensitive to environmental pollutants. This study was conducted to investigate how B[a]P affected the gonadal subcellular fractions, including plasma membrane, nucleus, mitochondria and microsome in scallops, and whether subcellular damages were related to reproductive toxicity. The results showed that mature gametes' counts were significantly decreased in B[a]P-treated scallops. Three biological macromolecules (viz., DNA, lipids and proteins) in gonadal subcellular fractions obtained by differential centrifugation suffered damages, including DNA damage, lipid peroxidation and protein carbonylation in B[a]P treatment groups. Interestingly, mitochondria and microsome were more vulnerable to lipid peroxidation and protein carbonylation than plasma membrane and nucleus, meanwhile males were more susceptible to DNA damage than females under B[a]P exposure. In addition, histological analysis showed that B[a]P delayed gonadal development in C. farreri. To summarize, our results indicated that B[a]P caused damages to biological macromolecules in gonadal subcellular fractions and then induced damages to gonadal tissues of C. farreri, which further inhibited gonadal development and ultimately leaded to reduction in fertility. This study firstly reports the impacts of PAHs on subcellular fractions in bivalves and their relationship with reproductive toxicity. Moreover, exposure of reproductive scallops to B[a]P leads to defects in reproduction, raising concerns on the possible long-term consequences of PAHs for natural populations of bivalves.

[Improved Regression Kriging Prediction of the Spatial Distribution of the Soil Cadmium by Integrating Natural and Human Factors].

Mastering the spatial distribution of heavy metals in the soil plays an important supporting role in the scientific formulation of soil pollution risk management and control strategies. Few factors were considered and multiple collinearity between parallel variables existed,resulting in low prediction accuracy. OK (common Kriging method), NRK (regressive Kriging method based on natural factors only), and NARK (regressive Kriging based on natural-human factors)were used to simulate the spatial distribution of soil Cd by selecting 23 natural-artificial influencing factors. The prediction accuracy was evaluated based on an empirical study of the area around Shaoguan Qujiang smelter. The results showed that the above-standard rate of soil cadmium in this area reached 85.93%, and the effect on the spatial heterogeneity of soil cadmium was shown as air emissions from smelters > air emissions from steel plants > pH > organic matter > Euclidean distance to road > Euclidean distance to river. The root-mean-square error and average absolute error of NARK's prediction results for soil cadmium were 26.86% and 30.56% lower than that of the OK method, respectively. The model determination coefficient R2 increased from 0.78 to 0.88. Compared with that of NRK, it was reduced by 24.15% and 24.23% and R2 increased from 0.81 to 0.88. The NRK combining natural and human factors significantly improved the simulation accuracy of the spatial distribution of soil cadmium, and the addition of human factors as auxiliary variables, especially atmospheric point source pollution emissions, greatly contributed to the improvement of the model accuracy.

Targeting BRD4 proteins suppresses the growth of NSCLC through downregulation of eIF4E expression.

Lung cancer is the leading cause of cancer-related death worldwide. Bromodomain and extraterminal domain (BET) proteins act as epigenome readers for gene transcriptional regulation. Among BET family members, BRD4 was well studied, but for its mechanism in non-small cell lung carcinoma has not been elucidated. eIF4E regulates gene translation and has been proved to play an important role in the progression of lung cancer. In this study, we first confirmed that BET inhibitors JQ1 and I-BET151 suppressed the growth of NSCLCs, in parallel with downregulated eIF4E expression. Then we found that knockdown of BRD4 expression using siRNAs inhibited the growth of NSCLCs as well as decreased eIF4E protein levels. Moreover, overexpression of eIF4E partially abrogated the growth inhibitory effect of JQ1, while knockdown of eIF4E enhanced the inhibitory effect of JQ1. Furthermore, JQ1 treatment or knockdown of BRD4 expression decreased eIF4E mRNA levels and inhibited its promoter activity by luciferase reporter assay. JQ1 treatment significantly decreased the binding of eIF4E promoter with BRD4. Finally, JQ1 inhibited the growth of H460 tumors in parallel with downregulated eIF4E mRNA and protein levels in a xenograft mouse model. These findings suggest that inhibition of BET by JQ1, I-BET151, or BRD4 silencing suppresses the growth of non-small cell lung carcinoma through decreasing eIF4E transcription and subsequent mRNA and protein expression. Considering that BET regulates gene transcription epigenetically, our findings not only reveal a new mechanism of BET-regulated eIF4E in lung cancer, but also indicate a novel strategy by co-targeting eIF4E for enhancing BET-targeted cancer therapy.

AEG-1 induces gastric cancer metastasis by upregulation of eIF4E expression.

Gastric cancer is the third leading cause of cancer-related deaths worldwide, and patients with lymph node, peritoneal and distant metastasis have a poor prognosis. Overexpression of Astrocyte-elevated gene-1 (AEG-1) has been reported to be correlated with the progression and metastasis of gastric cancer. However, its mechanisms are quite unclear. In this study, we found that elevated expression of AEG-1 was correlated with metastasis in human gastric cancer tissues. Moreover, gain- or loss-of-function of AEG-1, respectively, promoted or suppressed epithelial-mesenchymal transition (EMT), migration and invasion of gastric cancer cells. AEG-1 positively regulated eIF4E, MMP-9 and Twist expression. Manipulating eIF4E expression by transfection of overexpression constructs or siRNAs partially eliminated AEG-1-regulated EMT, cell migration and invasion. In addition, overexpression or knockdown of eIF4E promoted or suppressed EMT, cell migration and invasion in parallel with upregulation of MMP-9 and Twist expression, while manipulating eIF4E expression partially abrogated AEG-1-induced MMP-9 and Twist. Finally, silencing of AEG-1 expression not only inhibited tumour growth in parallel with downregulation of eIF4E, MMP-9 and Twist expression in a xenograft nude mouse model, but also suppressed lymph node and peritoneal metastasis of gastric cancer in an orthotopic nude mouse model. These findings suggest that AEG-1 promotes gastric cancer metastasis through upregulation of eIF4E-mediated MMP-9 and Twist, which provides new diagnostic markers and therapeutic targets for cancer metastasis.

Bioprocessing of tea oil fruit hull with acetic acid organosolv pretreatment in combination with alkaline H2O2.

BACKGROUND: As a natural renewable biomass, the tea oil fruit hull (TOFH) mainly consists of lignocellulose, together with some bioactive substances. Our earlier work constructed a two-stage solvent-based process, including one aqueous ethanol organosolv extraction and an atmospheric glycerol organosolv (AGO) pretreatment, for bioprocessing of the TOFH into diverse bioproducts. However, the AGO pretreatment is not as selective as expected in removing the lignin from TOFH, resulting in the limited delignification and simultaneously high cellulose loss. RESULTS: In this study, acetic acid organosolv (AAO) pretreatment was optimized with experimental design to fractionate the TOFH selectively. Alkaline hydrogen peroxide (AHP) pretreatment was used for further delignification. Results indicate that the AAO-AHP pretreatment had an extremely good selectivity at component fractionation, resulting in 92% delignification and 88% hemicellulose removal, with 87% cellulose retention. The pretreated substrate presented a remarkable enzymatic hydrolysis of 85% for 48 h at a low cellulase loading of 3 FPU/g dry mass. The hydrolyzability was correlated with the composition and structure of substrates by using scanning electron microscopy, confocal laser scanning microscopy, and X-ray diffraction. CONCLUSION: The mild AAO-AHP pretreatment is an environmentally benign and advantageous scheme for biorefinery of the agroforestry biomass into value-added bioproducts.

p70S6K promotes IL-6-induced epithelial-mesenchymal transition and metastasis of head and neck squamous cell carcinoma.

Head and neck squamous cell carcinoma (HNSCC) is the fifth most common cancer worldwide and a common cause of cancer-related death, with a 5-year survival rate of less than 60%. IL-6 has been suggested to play an important role in cancer metastasis, but its mechanism in HNSCC has not been fully clarified. p70S6K has been reported to induce epithelial-mesenchymal transition (EMT) of ovarian cancer, but its role in HNSCC remains unknown. In this study, we found that p70S6K and IL-6 were upregulated in high-metastatic HNSCC cell lines that underwent EMT when compared to paired low-metastatic cell lines. Overexpression of p70S6K promoted EMT and migration of HNSCC cells, while downregulation of p70S6K attenuated IL-6-induced EMT and cell migration. Furthermore, IL-6-induced p70S6K activation was attenuated by inhibitors of the PI3K/Akt/mTOR, MAPK/ERK, and JAK/STAT3 signaling pathways, suggesting that it located downstream of these pathways. These findings suggest that p70S6K promotes IL-6-induced EMT and metastasis of HNSCC. Targeting p70S6K for HNSCC therapy may benefit patients through the inhibition of tumor growth, as well as metastasis.