Enhanced electrophysiological activity and neurotoxicity screening of environmental chemicals using 3D neurons from human neural precursor cells purified with PSA-NCAM.

The assessment of neurotoxicity for environmental chemicals is of utmost importance in ensuring public health and environmental safety. Multielectrode array (MEA) technology has emerged as a powerful tool for assessing disturbances in the electrophysiological activity. Although human embryonic stem cell (hESC)-derived neurons have been used in MEA for neurotoxicity screening, obtaining a substantial and sufficiently active population of neurons from hESCs remains challenging. In this study, we successfully differentiated neurons from a large population of human neuronal precursor cells (hNPC) purified using a polysialylated neural cell adhesion molecule (PSA-NCAM), referred to as hNPCPSA-NCAM+. The functional characterization demonstrated that hNPCPSA-NCAM+-derived neurons improve functionality by enhancing electrophysiological activity compared to total hNPC-derived neurons. Furthermore, three-dimensional (3D) neurons derived from hNPCPSA-NCAM+ exhibited reduced maturation time and enhanced electrophysiological activity on MEA. We employed subdivided population analysis of active mean firing rate (MFR) based on electrophysiological intensity to characterize the electrophysiological properties of hNPCPSA-NCAM+-3D neurons. Based on electrophysiological activity including MFR and burst parameters, we evaluated the sensitivity of hNPCPSA-NCAM+-3D neurons on MEA to screen both inhibitory and excitatory neuroactive environmental chemicals. Intriguingly, electrophysiologically active hNPCPSA-NCAM+-3D neurons demonstrated good sensitivity to evaluate neuroactive chemicals, particularly in discriminating excitatory chemicals. Our findings highlight the effectiveness of MEA approaches using hNPCPSA-NCAM+-3D neurons in the assessment of neurotoxicity associated with environmental chemicals. Furthermore, we emphasize the importance of selecting appropriate signal intensity thresholds to enhance neurotoxicity prediction and screening of environmental chemicals.

ToxSTAR: drug-induced liver injury prediction tool for the web environment.

SUMMARY: Drug-induced liver injury (DILI) is a challenging endpoint in predictive toxicology because of the complex reactive metabolites that cause liver damage and the wide range of mechanisms involved in the development of the disease. ToxSTAR provides structural similarity-based DILI analysis and in-house DILI prediction models that predict four DILI subtypes (cholestasis, cirrhosis, hepatitis and steatosis) based on drug and drug metabolite molecules. AVAILABILITY AND IMPLEMENTATION: ToxSTAR is freely available at https://toxstar.kitox.re.kr/. SUPPLEMENTARY INFORMATION: Supplementary data are available at Bioinformatics online.

Pathway-level multi-omics analysis of the molecular mechanisms underlying the toxicity of long-term tacrolimus exposure.

Tacrolimus (TAC)-based treatment is associated with nephrotoxicity and hepatotoxicity; however, the underlying molecular mechanisms responsible for this toxicity have not been fully explored. This study elucidated the molecular processes underlying the toxic effects of TAC using an integrative omics approach. Rats were sacrificed after 4 weeks of daily oral TAC administration at a dose of 5 mg/kg. The liver and kidney underwent genome-wide gene expression profiling and untargeted metabolomics assays. Molecular alterations were identified using individual data profiling modalities and further characterized by pathway-level transcriptomics-metabolomics integration analysis. Metabolic disturbances were mainly related to an imbalance in oxidant-antioxidant status, as well as in lipid and amino acid metabolism in the liver and kidney. Gene expression profiles also indicated profound molecular alterations, including in genes associated with a dysregulated immune response, proinflammatory signals, and programmed cell death in the liver and kidney. Joint-pathway analysis indicated that the toxicity of TAC was associated with DNA synthesis disruption, oxidative stress, and cell membrane permeabilization, as well as lipid and glucose metabolism. In conclusion, our pathway-level integration of transcriptome and metabolome and conventional analyses of individual omics profiles, provided a more comprehensive picture of the molecular changes resulting from TAC toxicity. This study also serves as a valuable resource for subsequent investigations aiming to understand the mechanism underlying the molecular toxicology of TAC.

Delineation of the molecular mechanisms underlying Colistin-mediated toxicity using metabolomic and transcriptomic analyses.

The mechanisms underlying colistin-induced toxicity are not fully understood. This study used untargeted metabolomics and transcriptomics to elucidate the molecular processes occurring in the liver and kidney of rats after treatment with colistin methanesulfonate (CMS). Rats were treated with 50 mg/kg CMS (high-dose), 25 mg/kg CMS (low-dose), or vehicle control, either as a single dose or once daily for 1 or 4 weeks. We found that metabolic alterations were dose- and treatment duration-dependent in the kidney, whereas mild changes were noted in the liver. Metabolic profiles in the high-dose, low-dose, and control groups of both tissues could be classified using partial least-squares discriminant analysis. Metabolic alterations were associated with the citric acid cycle and related processes, disrupted balance between pro-oxidants and antioxidants, inflammatory responses, and amino acid and nucleic acid metabolism. Gene expression profiles further showed that high-dose treatment was associated with disrupted metabolism, oxidative stress, and proinflammatory signals in the kidney. The expression levels of genes related to the cell cycle, DNA replication, and programmed cell death were also predominantly upregulated. These findings suggested that high-dose treatment was associated with a dramatic increase in cellular kidney injury, while only minor effects were observed in the low-dose group. Almost no significant gene expression was changed in the liver, even with high-dose CMS. In conclusion, untargeted metabolomics and transcriptomics provided better insights into the biological mechanisms underlying colistin-induced nephrotoxicity.

Comparative omics analyses of hepatotoxicity induced by oral azole drugs in mice liver and primary hepatocytes.

Ketoconazole (KTZ) and itraconazole (ITZ) are antifungal agents that have a broad spectrum of activity against fungal pathogens. However, the therapeutic indications of many antifungal drugs, including those of the azole group, are restricted due to possible hepatotoxicity. We performed toxicogenomic analyses using in vivo and in vitro models to investigate the molecular mechanisms underlying the hepatotoxicity of two azole antifungal drugs. C57BL/6 male mice were treated daily with KTZ or ITZ, sacrificed at days 1 or 7, and the serum biochemistry and histopathology results showed that the KTZ-treated mice exhibited hepatotoxicity. Primary hepatocytes from C57BL/6 mice also exposed to KTZ or ITZ, and the cytotoxic effects of KTZ and ITZ were evaluated; KTZ exerted a greater cytotoxic effect than ITZ. The gene expression profiles in the livers of the 7-day-treated group and primary hepatocytes of the 24-h-treated group for both KTZ and ITZ were comparatively analyzed. Differentially expressed genes were selected based on the fold-changes and statistical significance, and the biological functions were analyzed using ingenuity pathways analysis. The results revealed that genes related to cholesterol synthesis were overexpressed in the liver in the KTZ-treated group, whereas expression of those related to acute phase injury was significantly altered in the ITZ-treated group. Causal gene analyses suggested that sterol regulatory element-binding transcription factors are key regulators that activate the transcription of target genes associated with the hepatotoxicity induced by oral KTZ. These findings enhance our understanding of the molecular mechanisms underlying the hepatotoxicity of azole drugs.

Increased serum bile acid concentration following low-dose chronic administration of thioacetamide in rats, as evidenced by metabolomic analysis.

A liquid chromatography/time-of-flight mass spectrometry (LC/TOF-MS)-based metabolomics approach was employed to identify endogenous metabolites as potential biomarkers for thioacetamide (TAA)-induced liver injury. TAA (10 and 30mg/kg), a well-known hepatotoxic agent, was administered daily to male Sprague-Dawley (SD) rats for 28days. We then conducted untargeted analyses of endogenous serum and liver metabolites. Partial least squares discriminant analysis (PLS-DA) was performed on serum and liver samples to evaluate metabolites associated with TAA-induced perturbation. TAA administration resulted in altered levels of bile acids, acyl carnitines, and phospholipids in serum and in the liver. We subsequently demonstrated and confirmed the occurrence of compromised bile acid homeostasis. TAA treatment significantly increased serum levels of conjugated bile acids in a dose-dependent manner, which correlated well with toxicity. However, hepatic levels of these metabolites were not substantially changed. Gene expression profiling showed that the hepatic mRNA levels of Ntcp, Bsep, and Oatp1b2 were significantly suppressed, whereas those of basolateral Mrp3 and Mrp4 were increased. Decreased levels of Ntcp, Oatp1b2, and Ostα proteins in the liver were confirmed by western blot analysis. These results suggest that serum bile acids might be increased due to the inhibition of bile acid enterohepatic circulation rather than increased endogenous bile acid synthesis. Moreover, serum bile acids are a good indicator of TAA-induced hepatotoxicity.

Nonylphenol-induced apoptotic cell death in mouse TM4 Sertoli cells via the generation of reactive oxygen species and activation of the ERK signaling pathway.

Nonylphenol (NP), a representative endocrine disruptor, interferes with reproductive function in aquatic organisms and animals. Although many previous studies have focused on apoptotic cell death by NP, the fundamental mechanism of NP on apoptosis remains poorly understood. Here, we investigated the molecular mechanism on NP-induced apoptotic cell death in mouse TM4 Sertoli cells. To evaluate NP treatment on cell viability, formazan and lactate dehydrogenase (LDH) assays were performed. Results indicate that NP reduced cell viability and increased the release of LDH in dose- and time-dependent manners. The reduction of cell viability by NP treatment appeared to involve necrosis as well as apoptosis based on nuclear fragmentation, an increase in the sub G1 population, and the detection of poly(ADP ribose) polymerase and caspase-3 cleavage. Additionally, the anti-apoptotic protein Bcl-2 diminished, whereas the pro-apoptotic protein Bax increased in a time-dependent manner. Note that NP-induced apoptotic cell death was enhanced by the generation of reactive oxygen species (ROS) and activation of extracellular signal-regulated kinase (ERK) signaling. Pretreatment with N-acetylcysteine, an antioxidant, attenuated NP-induced apoptotic cell death. Moreover, NP caused a transient activation of the MAPK pathway. In particular, NP-induced cell death was significantly suppressed by U0126, a specific inhibitor of ERK. Taken together, our results suggest that NP induces apoptosis in mouse TM4 Sertoli cells via ROS generation and ERK activation.

Hepa-ToxMOA: a pathway-screening method for evaluating cellular stress and hepatic metabolic-dependent toxicity of natural products.

In the field of drug discovery, natural products have emerged as therapeutic agents for diseases such as cancer. However, their potential toxicity poses significant obstacles in the developing effective drug candidates. To overcome this limitation, we propose a pathway-screening method based on imaging analysis to evaluate cellular stress caused by natural products. We have established a cellular stress sensing system, named Hepa-ToxMOA, which utilizes HepG2 cells expressing green fluorescent protein (GFP) fluorescence under the control of transcription factor response elements (TREs) for transcription factors (AP1, P53, Nrf2, and NF-κB). Additionally, to augment the drug metabolic activity of the HepG2 cell line, we evaluated the cytotoxicity of 40 natural products with and without S9 fraction-based metabolic activity. Our finding revealed different activities of Hepa-ToxMOA depending on metabolic or non-metabolic activity, highlighting the involvement of specific cellular stress pathways. Our results suggest that developing a Hepa-ToxMOA system based on activity of drug metabolizing enzyme provides crucial insights into the molecular mechanisms initiating cellular stress during liver toxicity screening for natural products. The pathway-screening method addresses challenges related to the potential toxicity of natural products, advancing their translation into viable therapeutic agents.

Safety assessment and gastrointestinal retention of orally administered cerium oxide nanoparticles in rats.

Cerium oxide nanoparticles (CeO2 NPs, NM-212) are well-known for their catalytic properties and antioxidant potential, and have many applications in various industries, drug delivery, and cosmetic formulations. CeO2 NPs exhibit strong antimicrobial activity and can be used to efficiently remove pathogens from different environments. However, knowledge of the toxicological evaluation of CeO2 NPs is too limited to support their safe use. In this study, CeO2 NPs were orally administered to Sprague Dawley rats for 13 weeks at the doses of 0, 10, 100, and 1000 mg/kg bw/day, followed by a four week recovery period. The hematology values for the absolute and relative reticulocyte counts in male rats treated with 1000 mg/kg bw/day CeO2 NPs were lower than those in control rats. The clinical chemistry values for sodium and chloride in the treated male rat groups (100 and 1000 mg/kg/day) and total protein and calcium in the treated female rat groups (100 mg/kg/day) were higher than those in the control groups. However, these changes were not consistent in both sexes, and no abnormalities were found in the corresponding pathological findings. The results showed no adverse effects on any of the parameters assessed. CeO2 NPs accumulated in the jejunum, colon, and stomach wall of rats administered 1000 mg/kg CeO2 NPs for 90 days. However, these changes were not abnormal in the corresponding histopathological and immunohistochemical examinations. Therefore, 1000 mg/kg bw/day may be considered the "no observed adverse effect level" of CeO2 NPs (NM-212) in male and female SD rats under the present experimental conditions.

Integrated transcriptomic analysis of liver and kidney after 28 days of thioacetamide treatment in rats.

Thioacetamide (TAA) was developed as a pesticide; however, it was soon found to cause hepatic and renal toxicity. To evaluate target organ interactions during hepatotoxicity, we compared gene expression profiles in the liver and kidney after TAA treatment. Sprague-Dawley rats were treated daily with oral TAA and then sacrificed, and their tissues were evaluated for acute toxicity (30 and 100 mg/kg bw/day), 7-day (15 and 50 mg/kg bw/day), and 4-week repeated-dose toxicity (10 and 30 mg/kg). After the 4-week repeated toxicity study, total RNA was extracted from the liver and kidneys, and microarray analysis was performed. Differentially expressed genes were selected based on fold change and significance, and gene functions were analyzed using ingenuity pathway analysis. Microarray analysis showed that significantly regulated genes were involved in liver hyperplasia, renal tubule injury, and kidney failure in the TAA-treated group. Commonly regulated genes in the liver or kidney were associated with xenobiotic metabolism, lipid metabolism, and oxidative stress. We revealed changes in the molecular pathways of the target organs in response to TAA and provided information on candidate genes that can indicate TAA-induced toxicity. These results may help elucidate the underlying mechanisms of target organ interactions during TAA-induced hepatotoxicity. Supplementary Information: The online version contains supplementary material available at 10.1007/s43188-022-00156-y.

Systems-level multi-omics characterization provides novel molecular insights into indomethacin toxicity.

The mechanism of indomethacin toxicity at the systemic level is largely unknown. In this study, multi-specimen molecular characterization was conducted in rats treated with three doses of indomethacin (2.5, 5, and 10 mg/kg) for 1 week. Kidney, liver, urine, and serum samples were collected and analyzed using untargeted metabolomics. The kidney and liver transcriptomics data (10 mg indomethacin/kg and control) were subjected to a comprehensive omics-based analysis. Indomethacin exposure at 2.5 and 5 mg/kg doses did not cause significant metabolome changes, whereas considerable alterations in the metabolic profile compared to the control were induced by a dose of 10 mg/kg. Decreased levels of metabolites and an increased creatine level in the urine metabolome indicated injury to the kidney. The integrated omics analysis in both liver and kidney revealed an oxidant-antioxidant imbalance due to an excess of reactive oxygen species, likely originating from dysfunctional mitochondria. Specifically, indomethacin exposure induced changes in metabolites related to the citrate cycle, cell membrane composition, and DNA synthesis in the kidney. The dysregulation of genes related to ferroptosis and suppression of amino acid and fatty acid metabolism were evidence of indomethacin-induced nephrotoxicity. In conclusion, a multi-specimen omics investigation provided important insights into the mechanism of indomethacin toxicity. The identification of targets that ameliorate indomethacin toxicity will enhance the therapeutic utility of this drug.

Gene expression analysis of toxicological pathways in TM3 leydig cell lines treated with Ethane dimethanesulfonate.

Ethane dimethanesulfonate (EDS), a well-known alkylating agent, selectively destroys Leydig cells. To clarify the molecular pathways underlying EDS action on Leydig cells, we analyzed gene expression profiles of an EDS-treated TM3 Leydig cell line. In this study, we analyzed the representative canonical pathways and toxicity pathways/gene lists using the Ingenuity Pathways Analysis program. In TM3 cells, 677 and 6756 genes were identified as being up- or downregulated after 3 and 24 h EDS treatments, respectively, (>1.3-fold changes, p < 0.05). Toxicological pathway analysis revealed that expression of genes related to Nrf2-mediated oxidative stress response showed remarkable changes in early or later stage of EDS-treated TM3 cells. Several genes related to steroidogenesis and apoptosis were also differentially expressed at 24 h in EDS-treated TM3 cells. Overall, toxicological pathway analysis using gene expression profiling showed that oxidative stress might be an important factor in cell death in TM3 cells affected by EDS treatment.

Inflammatory response in rat lungs with recurrent exposure to welding fumes: a transcriptomic approach.

As chronic exposure to welding fumes causes pulmonary diseases, such as pneumoconiosis, public concern has increased regarding continued exposure to these hazardous gases in the workplace. In a previous study, the inflammatory response to welding fume exposure was analysed in rat lungs in the case of recurrent exposure and recovery periods. Thus using lung samples, well-annotated by histological observation and biochemical analysis, this study examines the gene expression profiles to identify phenotype-anchored genes corresponding to lung inflammation and the repair phenomenon after recurrent welding fume exposure. Seven genes (Mmp12, Cd5l, LOC50101, LOC69183, Spp1, and Slc26a4) were found to be significantly up-regulated according to the severity of the lung injury. In addition, the transcription and translation of Trem2, which was up-regulated in response to the repair process, were validated using a real-time polymerase chain reaction, Western blotting, and immunohistochemistry. The differentially expressed genes in the exposure and recovery groups were also classified using k-means and hierarchical clustering, plus their toxicological function and canonical pathways were further analysed using Ingenuity Pathways Analysis Software. As a result, this comprehensive and integrative analysis of the transcriptional changes that occur during repeated exposure provides important information on the inflammation and repair processes after welding-fume-induced lung injury.

Genome-wide gene expression analysis reveals molecular insights into the drug-induced toxicity of nephrotoxic agents.

Drug-induced nephrotoxicity is frequently reported. However, the mechanisms underlying nephrotoxic medications and their overlapping molecular events, which might have therapeutic value, are unclear. We performed a genome-wide analysis of gene expression and a gene set enrichment analysis to identify common and unique pathways associated with the toxicity of colistin, ifosfamide, indomethacin, and puromycin. Rats were randomly allocated into the treatment or control group. The treatment group received a toxic dose once daily of each investigated drug for 1 week. Differentially expressed genes were found in the drug-treated kidney and liver compared to the control, except for colistin in the liver. Upregulated pathways were mainly related to cell death, cell cycle, protein synthesis, and immune response modulation in the kidney. Cell cycle was upregulated by all drugs. Downregulated pathways were associated with carbon metabolism, amino acid metabolism, and fatty acid metabolism. Indomethacin, colistin, and puromycin shared the most altered pathways in the kidney. Ifosfamide and indomethacin affected molecular processes greatly in the liver. Our findings provide insight into the mechanisms underlying the renal and hepatic adverse effects of the four drugs. Further investigation should explore the combinatory drug therapies that attenuate the toxic effects and maximize the effectiveness of nephrotoxic drugs.

miRNA regulation of cytotoxic effects in mouse Sertoli cells exposed to nonylphenol.

BACKGROUND: It is known that some environmental chemicals affect the human endocrine system. The harmful effects of endocrine disrupting chemical (EDC) nonylphenol (NP) have been studied since the 1980s. It is known that NP adversely affects physiological functions by mimicking the natural hormone 17 beta-estradiol. In the present study, we analyzed the expression of miRNAs and their target genes in mouse Sertoli TM4 cells to better understand the regulatory roles of miRNAs on Sertoli cells after NP exposure. METHODS: Mouse TM4 Sertoli cells were treated with NP for 3 or 24 h, and global gene and miRNA expression were analyzed using Agilent mouse whole genome and mouse miRNA v13 arrays. RESULTS: We identified genes that were > 2-fold differentially expressed in NP-treated cells and control cells (P < 0.05) and analyzed their functions through Gene Ontology analysis. We also identified miRNAs that were differentially expressed in NP-treated and control cells. Of the 186 miRNAs the expression of which differed between NP-treated and control cells, 59 and 147 miRNAs exhibited 1.3-fold increased or decreased expression at 3 and 24 h, respectively. Network analysis of deregulated miRNAs suggested that Ppara may regulate the expression of certain miRNAs, including miR-378, miR-125a-3p miR-20a, miR-203, and miR-101a, after exposure to NP. Additionally, comprehensive analysis of predicted target genes for miRNAs showed that the expression of genes with roles in cell proliferation, the cell cycle, and cell death were regulated by miRNA in NP-treated TM4 cells. Levels of expression of the miRNAs miR-135a* and miR-199a-5p were validated by qRT-PCR. Finally, miR-135a* target gene analysis suggests that the generation of reactive oxygen species (ROS) following exposure to NP exposure may be mediated by miR-135a* through regulation of the Wnt/beta-catenin signaling pathway. CONCLUSIONS: Collectively, these data help to determine NP's actions on mouse TM4 Sertoli cells and increase our understanding of the molecular mechanisms underlying the adverse effects of xenoestrogens on the reproductive system.

Identification of biomarkers of chemically induced hepatocarcinogenesis in rasH2 mice by toxicogenomic analysis.

Toxicogenomic approaches have been applied to chemical-induced heptocarcinogenesis rodent models for the identification of biomarkers of early-stage hepatocarcinogenesis and to help clarify the underlying carcinogenic mechanisms in the liver. In this study, we used toxiciogenomic methods to identify candidate biomarker genes associated with hepatocarcinogenesis in rasH2 mice. Blood chemical, histopathologic, and gene expression analyses of the livers of rasH2 mice were performed 7 and 91 days after the administration of the genotoxic hepatocarcinogens 2-acetylaminofluorene (AAF) and diethylnitrosoamine (DEN), the genotoxic carcinogen melphalan (Mel), and the nongenotoxic noncarcinogen 1-naphthylisothiocynate (ANIT). Histopathologic lesions and a rise in accompanying serum marker levels were found in the DEN-treated rasH2 mice, whereas no neoplastic lesions were observed in the rasH2 mice. However, biological functional analysis using Ingenuity Pathways Analysis (IPA) software revealed that genes with comparable molecular and cellular functions were similarly deregulated in the AAF- and DEN-treated rasH2 mice. We selected 68 significantly deregulated genes that represented a hepatocarcinogen-specific signature; these genes were commonly deregulated in both the AAF- and DEN-treated rasH2 mice on days 7 and 91. Hierarchical clustering analysis indicated that the expression patterns of the selected genes in the hepatocarcinogen (AAF and DEN) groups were distinctive from the patterns in the control, Mel, and ANIT groups. Biomarker filter analysis using IPA software suggested that 28 of the 68 signature genes represent promising candidate biomarkers of cancer. Quantitative real-time PCR analysis confirmed that the deregulated genes, which exhibited sustained up- and down-regulation up to day 91, are likely involved in early-stage hepatocarcinogenesis. In summary, the common and significant gene expression changes induced by AAF and DEN may reflect early molecular events associated with hepatocarcinogenesis, and these "signature" genes may be useful as biomarkers of hepatocarcinogenesis in mice.

TNFα enhances trovafloxacin-induced in vitro hepatotoxicity by inhibiting protective autophagy.

Trovafloxacin (TVX) is associated with idiosyncratic drug-induced liver injury (iDILI) and inflammation-mediated hepatotoxicity. However, the inflammatory stress-regulated mechanisms in iDILI remain unclear. Herein, we elucidated the novel role of tumor-necrosis factor alpha (TNFα), an inflammatory stress factor, in TVX-induced in vitro hepatotoxicity and synergistic toxicity. TVX specifically induced synergistic toxicity in HepG2 cells with TNFα, which inhibits autophagy. TVX-treated HepG2 cells induced protective autophagy by inhibiting the expression of mTOR signaling proteins, while ATG5 knockdown in HepG2 cells, responsible for the impairment of autophagy, enhanced TVX-induced toxicity due to the increase in cytochrome C release and JNK pathway activation. Interestingly, the expression of mTOR signal proteins, which were suppressed by TVX, disrupted the negative feedback of the PI3K/AKT pathway and TNFα rebounded p70S6K phosphorylation. Co-treatment with TVX and TNFα inhibited protective autophagy by maintaining p70S6K activity, which enhanced TVX-induced cytotoxicity. Phosphorylation of p70S6K was inhibited by siRNA knockdown and rapamycin to restore TNFα-inhibited autophagy, which prevented the synergistic effect on TVX-induced cytotoxicity. These results indicate that TVX activates protective autophagy in HepG2 cells exposed to toxicity and an imbalance in negative feedback regulation of autophagy by TNFα synergistically enhanced the toxicity. The finding from this study may contribute to a better understanding of the mechanisms underlying iDILI associated with inflammatory stress.

Gene networking in colistin-induced nephrotoxicity reveals an adverse outcome pathway triggered by proteotoxic stress.

Colistin has been widely used for the treatment of infections of multidrug­resistant Gram­negative bacteria, despite the fact that it induces serious kidney injury as a side effect. To investigate the mechanism underlying its nephrotoxicity, colistin methanesulfonate sodium (CMS; 25 or 50 mg/kg) was administered via intraperitoneal injection to Sprague­Dawley rats daily over 7 days. Serum biochemistry and histopathology indicated that nephrotoxicity occurred in the rats administered with CMS. Whole­genome microarrays indicated 894 differentially expressed genes in the group treated with CMS (analysis of variance, false discovery rate <0.05, fold­change ≥1.3). Gene pathway and networking analyses revealed that genes associated with proteotoxic stress, including ribosome synthesis, protein translation, and protein folding, were significantly associated with the nephrotoxicity induced by CMS. It was found that colistin inhibited the expression of the target genes heat shock factor 1 and nuclear factor erythroid­2­related factor­2, which are associated with proteostasis, and that nephrotoxicity of CMS may be initiated by proteotoxic stress due to heat shock response inhibition, leading to oxidative stress, endoplasmic reticulum stress, cell cycle arrest and apoptosis, eventually leading to cell death. A putative adverse outcome pathway was constructed based on the integrated gene networking data, which may clarify the mode of action of colistin­induced nephrotoxicity.

Cerium Aminoclay-A Potential Hybrid Biomaterial for Anticancer Therapy.

In this study, novel biomedical properties of Ce-aminoclay (CeAC) were investigated through in vitro and in vivo assays. CeAC (≥500 µg/mL) can selectively kill cancer cells (A549, Huh-1, AGS, C33A, HCT116, and MCF-7 cells) while leaving most normal cells unharmed (WI-38 and CCD-18Co cells). Notably, it displayed a high contrast of simultaneous imaging in HeLa cells by blue photoluminescence without any fluorescence dye. Its anticancer mechanism has been fully demonstrated through apoptosis assays; herein CeAC induced high-level apoptosis (16%), which promoted the expression of proapoptotic proteins (Bax, p53, and caspase 9) in tumor cells. Besides, its biological behavior was determined through antitumor effects using intravenous and intratumoral administration routes in mice implanted with HCT116 cells. During a 40 day trial, the tumor volume and tumor weight were reduced by a maximum of 92.24 and 86.11%, respectively. The results indicate that CeAC exhibits high bioavailability and therapeutic potential based on its unique characteristics, including high antioxidant capacity and electrostatic interaction between its amino functional groups and the mucosal surface of cells. In summary, it is suggested that CeAC, with its high bioimaging contrast, can be a promising anticancer agent for future biomedical applications.

Immunogenomics reveal molecular circuits of diclofenac induced liver injury in mice.

Diclofenac is a non-steroidal anti-inflammatory drug and its use can be associated with severe adverse reactions, notably myocardial infarction, stroke and drug-induced liver injury (DILI). In pursue of immune-mediated DILI mechanisms an immunogenomic study was carried out. Diclofenac treatment of mice at 30 mg/kg for 3 days caused significant serum ALT and AST elevations, hepatomegaly and degenerative changes including hepatic glycogen depletion, hydropic swelling, cholesterolosis and eosinophilic hepatocytes with one animal presenting subsegmental infarction due to portal vein thrombosis. Furthermore, portal/periportal induction of the rate limiting enzyme in ammonia detoxification, i.e. carbamoyl phosphate synthetase 1 was observed. The performed microarray studies informed on > 600 differential expressed genes of which 35, 37 and 50 coded for inflammation, 51, 44 and 61 for immune and 116, 129 and 169 for stress response, respectively after single and repeated dosing for 3 and 14 days. Bioinformatic analysis defined molecular circuits of hepatic inflammation with the growth hormone (Ghr)- and leptin receptor, the protein-tyrosine-phosphatase, selectin and the suppressor-of-cytokine-signaling (Socs) to function as key nodes in gene regulatory networks. Western blotting confirmed induction of fibronectin and M-CSF to hallmark tissue repair and differentiation of monocytes and macrophages. Transcript expression of the macrophage receptor with collagenous structure increased > 7-fold and immunohistochemistry of CD68 evidenced activation of tissue-resident macrophages. Importantly, diclofenac treatment prompted strong expression of phosphorylated Stat3 amongst individual animals and the associated 8- and 4-fold Soc3 and Il-6 induction reinforced Ghr degradation as evidenced by immunoblotting. Moreover, immunohistochemistry confirmed regulation of master regulatory proteins of diclofenac treated mice to suggest complex pro-and anti-inflammatory reactions in immune-mediated hepatic injury. The findings encourage translational research.