Language：English   Publishing type：Research paper (scientific journal)  

The protein phosphatase inhibitor microcystin-LR (MC-LR), a hepatocyte-selective cyanotoxin, induces phenotypic changes in HEK293 OATP1B3-expressing (HEK293-OATP1B3) cells, which include cytoskeletal reorganization (HEK293-OATP1B3-AD) and anoikis resistance (HEK293-OATP1B3-FL) transformed cells, respectively. These cells acquire resistance to MC-LR and partial epithelial-mesenchymal transition (EMT) characteristics. In cancer cells, EMT is generally involved in multi-drug resistance. Here, we focused on the multi-drug resistance of HEK293-OATP1B3-AD and HEK293-OATP1B3-FL cells. The MTT assay and immunoblotting were conducted to examine the responses of HEK293-OATP1B3, HEK293-OATP1B3-AD, and HEK293-OATP1B3-FL cells to multiple toxins and drugs that function as substrates for OATP1B3, including MC-LR, nodularin (Nod), okadaic acid (OA), and cisplatin (CDDP). HEK293-OATP1B3-AD and HEK293-OATP1B3-FL cells were more resistant to MC-LR, Nod, and OA than HEK293-OATP1B3 cells. Conversely, the three cell types were equivalently sensitive to CDDP. By using protein phosphatase assay, the reduction of the inhibitory effect of MC-LR and Nod on phosphatase activity might be one reason for the resistance to MC-LR and Nod in HEK293-OATP1B3-AD and HEK293-OATP1B3-FL cells. Furthermore, the parental HEK293-OATP1B3 cells showed enhanced p53 phosphorylation and stabilization after MC-LR exposure, while p53 phosphorylation was attenuated in HEK293-OATP1B3-AD and HEK293-OATP1B3-FL cells. Moreover, in HEK293-OATP1B3-AD and HEK293-OATP1B3-FL cells, AKT phosphorylation was higher than that of the parental HEK293-OATP1B3 cell line. These results suggest that the multi-toxin resistance observed in HEK293-OATP1B3-AD and HEK293-OATP1B3-FL cells is associated with AKT activation and p53 inactivation.

DOI： 10.1016/j.toxicon.2023.107592

Scopus

PubMed

Language：Japanese   Publishing type：Research paper (scientific journal)   Publisher：Georg Thieme Verlag KG  

Abstract

The hepatotoxin microcystin-LR is a strong inhibitor of serine/threonine protein phosphatase (PP) 1 and PP2A. The onset of its cytotoxicity depends on its selective uptake via the hepatocyte uptake transporters, organic anion transporting polypeptide (OATP) 1B1 and OATP1B3. Understanding and preventing the cytotoxicity of microcystin-LR is crucial to maintain human health. This chemoprevention study demonstrates that the herbal plant extract of iwajisha (20 µg/mL) reduced microcystin-LR cytotoxicity in OATP1B3-expressing cells by approximately six times. In addition, 20 µM acteoside, which is one of the major compounds in iwajisha, reduced microcystin-LR cytotoxicity by approximately 7.4 times. Acteoside could also reduce the cytotoxicity of other compounds, such as okadaic acid and nodularin, which are both substrates of OATP1B3 and inhibitors of PP1/PP2A. To investigate the mechanism by which the cytotoxicity of microcystin-LR is attenuated by acteosides, microcystin-LR and microcystin-LR-binding proteins in cells were examined after microcystin-LR and acteosides were co-exposed. Thus, acteoside noncompetitively inhibited microcystin-LR uptake by OATP1B3-expressing cells. Furthermore, acteoside inhibited the intracellular interaction of microcystin-LR with its binding protein(s), including the 22 kDa protein. Furthermore, using immunoblot analysis, acteoside induced the phosphorylation of extracellular signal-regulated kinase (ERK), which is one of the survival signaling molecules. These results suggest that acteoside reduces microcystin-LR cytotoxicity through several mechanisms, including the inhibition of microcystin-LR uptake via OATP1B3, and decreased interaction between microcystin-LR and its binding protein(s), and that ERK signaling activation contributes to the attenuation effect of acteoside against microcystin-LR cytotoxicity.

DOI： 10.1055/a-1978-8768

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PubMed

The present study was designed to examine genotoxicity induced by 10-40 nm zinc oxide (ZnO) nanoparticles using the in vitro system. The frequency of micronuclei was significantly increased in a dose-dependent manner when cultured Chinese hamster lung (CHL/IU) cells were exposed to ZnO nanoparticles for 24, 48 and 72 h in the continuous treatment method. The maximal frequency of micronuclei was observed in exposure of CHL/IU cells to ZnO nanoparticles at a concentration of 125 μM. The frequency of micronuclei was profoundly enhanced when CHL/IU cells were exposed to ZnO nanoparticles in the presence vs. absence of S9 mixture including metabolic enzymes in the short-term treatment method, demonstrating an increase in the formation of micronuclei by S9 mixture. The maximal frequency of micronuclei was seen in exposure of CHL/IU cells to ZnO nanoparticles at a concentration of 140 μM. Similar results were obtained in chromosome aberrations, particularly structural aberrations. Surprisingly, administration of the superoxide radical scavenger, tempol, completely abolished an increase in the frequency of micronuclei in the presence or absence of S9 mixture, indicating a central role of superoxide radical in the formation of micronuclei. Indeed, reactive oxygen species (ROS) generation was elevated by simultaneous incubation of S9 mixture and ZnO nanoparticles and by exposure of CHL/IU cells to ZnO nanoparticles in the presence or absence of S9 mixture. An electron microscopic examination revealed mitochondrial damage in CHL/IU cells exposed to ZnO nanoparticles, indicating the participation of mitochondrial dysfunction in ROS generation in this setting. These observations suggest that ZnO nanoparticles evoke genotoxicity through superoxide radical-induced oxidative stress derived from mitochondrial damage in CHL/IU cells. S9 mixture appears to contribute to a further increase in genotoxicity through the generation of superoxide radical by metabolic activation of ZnO nanoparticles.

DOI： 10.1016/j.mrgentox.2018.07.003

Scopus

PubMed

Language：English   Publishing type：Research paper (scientific journal)  

Language：English   Publishing type：Research paper (scientific journal)  

Total pages：335   Responsible for pages：233-242   Language：Japanese Book type：Textbook, survey, introduction

Total pages：xiii, 335p   Language：Japanese

CiNii Books

Language：English

Language：Japanese

Total pages：331   Responsible for pages：232-241   Language：Japanese Book type：Textbook, survey, introduction

Language：Japanese   Publisher：(一社)日本衛生学会  

Language：Japanese   Publisher：(一社)日本衛生学会  

Authorship：Corresponding author  

Language：English  

J-GLOBAL

Event date： 2019.12

Language：English   Presentation type：Poster presentation  

Event date： 2018.12

Language：Japanese   Presentation type：Oral presentation (general)  

Event date： 2019.12

Language：Japanese   Presentation type：Oral presentation (general)  

Event date： 2019.12

Language：English   Presentation type：Poster presentation  

Event date： 2019.12

Language：English   Presentation type：Poster presentation