Microcystin-LR (MC-LR), a prevalent cyanotoxin present in hazardous cyanobacterial blooms, is recognized as a neurotoxic environmental pollutant that induces brain damage and neurobehavioral deficits. However, the mechanisms underlying MC-LR-induced neurotoxicity remain unclear. This study aims to elucidate the role of mitophagy in MC-LR-induced neurotoxicity both in vitro and in vivo. We found that administration of 10 mu g/kg body weight (intraperitoneally) MC-LR impaired learning and memory abilities and induced neuronal damage and apoptosis in the CA1 region of the hippocampus in rats. Exposure to MC-LR (1 mu M-10 mu M) resulted in cellular damage and apoptosis in PC-12 and HT22 cells. MC-LR induced mitophagy through the PINK1/Parkin pathway but hindered mitophagy progression by repressing Scd2 transcription in neurons. These inhibitory effects were reversed by Scd2 overexpression. Furthermore, MC-LR was found to repress Scd2 transcription by directly binding to type 1 insulin-like growth factor receptor (IGF-1R) and competitively inhibiting its activation by Insulin-like growth factor 1 (IGF-1). Overexpression of IGF-1R and administration of exogenous IGF-1 mitigated the MC-LR-induced inhibition of Scd2 and the associated mitophagy defects. These findings indicate that IGF-1R is the direct target of MC-LR in neurons. MC-LR initiates mitophagy defects and apoptosis by inhibiting Scd2 transcription through binding to IGF-1R.

Objective To explore the role and mechanism of RNA demethylase fat mass and obesity-associated protein (FTO) in the ferroptosis in testicular interstitial cells induced by di (2-ethylhexyl) phthalate (DEHP). Methods Forty 3-week-old C57BL/6 male mice were randomly divided into a control group (corn oil) and 3 dosed DEHP treatment groups (5, 250 and 500 mg/kg), and received an intragastric infusion of corresponding agents for 35 d, respectively. After mouse testicular interstitial TM3 cells was treated with 0, 100, 200 and 400 μmol/L mono-2-ethylhexyl phthalate (MEHP) for 24 h, corresponding plasmids were transfected to construct Fto overexpressing TM3 cells. Serum testosterone level was detected by ELISA, expression of testicular proteins was detected with immunohistochemical assay, and contents of Fe2+, malondialdehyde (MDA) and lipid peroxides in the testicle were detected by colorimetry. Methylated RNA immunoprecipitation, RT-PCR, and Western blotting were used to detect the level of N6-methyladenosine (m6A) modification. Results In the mice exposed to 250 and 500 mg/kg DEHP, the serum testosterone level was significantly reduced (P < 0.01), contents of Fe2+, MAD and lipid peroxides in testicular tissue were obviously increased (P < 0.01), and protein levels of RNA demethylase FTO, and ferroptosis related molecules ferritin heavy chain 1 (FTH1) and glutathione peroxidase 4 (GPX4) were significantly down-regulated (P < 0.05), while those of transferrin receptor (TFRC), ferroportin (FPN), cyclooxygenase-2 (COX-2), and acyl-CoA synthetase long-chain family member 4 (ACSL4) were notably up-regulated (P < 0.05). MEHP treatment for 24 h resulted in remarkably decreased cell viability in the TM3 cells, increased production of intracellular reactive oxygen species (ROS), reduced mitochondrial membrane potential (MMP) (P < 0.01), down-regulated mRNA and protein levels of Fto (P < 0.01), and the changes in other ferroptosis related proteins were consistent with the trend in testicular tissue, indicating ferroptosis in testicular interstitial cells. Intervention with ferroptosis inhibitor Fer-1 or overexpression of Fto significantly inhibited MEHP-induced toxicity and ferroptosis in TM3 cells (P < 0.05), and overexpression of Fto reduced the m6A modification of Gpx4 and Fth1 mRNA (P < 0.05). Conclusion Abnormal m6A modification of Gpx4 and Fth1 caused by inhibiting FTO expression may be the mechanism of ferroptosis in testicular interstitial cells induced by DEHP.

Microcystin-leucine arginine (MC-LR) is a common cyantotoxin produced by hazardous cyanobacterial blooms, and eutrophication is increasing the contamination level of MC-LR in drinking water supplies and aquatic foods. MC-LR has been linked to colorectal cancer (CRC) progression associated with tumor microenvironment, however, the underlying mechanism is not clearly understood. In present study, by using GEO, KEGG, GESA and ImmPort database, MC-LR related differentially expressed genes (DEGs) and pathway- and gene set-enrichment analysis were performed. Of the three identified DEGs (CXCL1, GUCA2A and GDF15), CXCL1 was shown a positive association with tumor infiltration, and was validated to have a dominantly higher upregulation in MC-LR-treated tumor-associated macrophages (TAMs) rather than in MC-LR-treated CRC cells. Both CRC cell/macrophage co-culture and xenograft mouse models indicated that MC-LR stimulated TAMs to secrete CXCL1 resulting in promoted proliferation, migration, and invasion capability of CRC cells. Furtherly, IP-MS assay found that interaction between TAMs-derived CXCL1 and CRC cell-derived IGHG1 may enhance CRC cell proliferation and migration after MC-LR treatment, and this effect can be attenuated by silencing IGHG1 in CRC cell. In addition, molecular docking analysis, co-immunoprecipitation and immunofluorescence further proved the interactions between CXCL1 and IGHG1. In conclusion, CXCL1 secreted by TAMs can trigger IGHG1 expression in CRC cells, which provides a new clue in elucidating the mechanism of MC-LR-mediated CRC progression.

As a typical environmental endocrine disrupting chemical (EDC), di-(2-ethylhexyl) phthalate (DEHP) is thought to be related to reproductive disorders, especially in males. Growing evidence suggests that various EDCs may result in an impaired telomere structure and function, which is associated with male infertility. However, the adverse effect of DEHP on telomeres in male reproductive cells has rarely been studied, and the related mechanisms remain unclear. In this study, we tested the effects of mono-(2-ethylhexyl) phthalate (MEHP), the primary metabolite of DEHP, on telomere dysfunction in mouse spermatogonia-derived cells (GC-1) and the potential role of TERT and c-Myc in MEHP-induced spermatogenic cell damage. Results showed that MEHP induced cell viability inhibition, G0/G1 phase cell cycle arrest, and apoptosis in GC-1 cells in a dose-dependent manner. Shortened telomeres, reduced telomerase activity, and decreased expression of TERT, c-Myc, and upstream transcription factors of c-Myc were also observed in the MEHP-treated cells. In conclusion, it can be concluded that TERT-mediated telomere dysfunction may contribute to MEHP-induced G0/G1 phase cell cycle arrest and apoptosis in GC-1 cells through the impairment of c-Myc and its upstream transcription factors.

Objective To determine whether metformin(Met) inhibits the proliferation of colorectal cancer cells by inducing cell senescence, and to preliminarily explore the underlying mechanism. Methods LOVO and SW480 colorectal cancer cells were treated with different concentrations of Met(0, 2.5, 5.0, 10.0, 20.0 and 40.0 mmol/L). The cell viability was detected by CCK-8 assay, and apoptosis and cell cycle were detected by flow cytometry. According to the results of apoptosis, differences between control group (0 mmol/L) and 5.0 mmol/L Met group(n=3) were compared in following experiments. Cell proliferation was detected by CCK-8 assay, EdU assay, and clone formation assay, senescence metabolism was detected by senescence-associated β-galactosidase(SA-β-gal)staining, protein phosphatase 2A(PP2A) enzyme activity was measured with ELASA, and protein expression was detected with Western blotting. PP2A inhibitor LB-100 was used to treat the cells alone or combined with Met, and then the cells were divided into control group, Met group, Met+LB-100 group, and LB-100 group(n=3). Above experiments were performed again. Results Met treatment significantly inhibited LOVO and SW480 cells proliferation in a concentration- and time-dependent manner(P < 0.05). Under the treatment of low concentration(≤5.0 mmol/L) of Met, the cells presented no obvious apoptosis, but were obviously inhibited for proliferation and arrested at G0/G1 phase. The cells displayed a typical senescence-like morphology of large, flat and vacuolated, and the number of SA-β-gal positive cells was increased significantly. For PP2A, total protein expression showed no change, but phosphorylation level decreased obviously and PP2A activity increased statistically(P < 0.05), in the meantime, the phosphorylation level of downstream AKT protein decreased and senescence-related proteins p53 and P21 increased significantly. Treatment by PP2A inhibitor LB-100 combined with Met significantly reversed the inhibition of Met on cell proliferation and delayed Met-induced cell senescence. Conclusion Low concentration of Met can induce cell senescence through PP2A/AKT pathway and then inhibit the proliferation of colorectal cancer cells.