PFOI stimulates the motility of T24 bladder cancer cells: Possible involvement and activation of lncRNA malat1
Introduction
Perfluorinated chemicals (PFCs) have been widely used in a variety of applications, including adhesives, cosmetics, cleaners, coatings, and electronics (Lindstrom et al., 2011). Because of some uncertainty that surrounds the safety of PFCs with evidence suggesting that their application may lead to potential toxicity and risk to mammal and environmental health (Lau et al., 2007), perfluorooctane sulfonate (PFOS) and related compounds have been classified as persistent organic pollutants under Annex B of the Stockholm Convention in 2009 (UNEP, 2009). There is an increasing focus on the potential toxicity of PFCs in rodent, avian, reptilian, aquatic organism, mammalian, and non-mammalian wildlife in the last decades, including hepatotoxicity (Ojo et al., 2020), neurotoxicity (Chen et al., 2014), carcinogenicity (Chang et al., 2014), developmental toxicity (K. Wang et al., 2020), and immune toxicity (DeWitt et al., 2012). As an important raw material and intermediate to produce PFCs by telomerization, the demand for PFIs has increased significantly over the years (Prevedouros et al., 2010). The perfluorooctyl iodide (PFOI, C8) is the main product of PFIs and has been widely detected both in air and soil around a fluorochemical manufacturing plant (Ruan et al., 2010).
Long noncoding RNA (lncRNA) transcripts are generally longer than 200 nt in length and cannot be translated. LncRNAs play a significant role in the regulation of multiple cellular processes at the transcriptional, post-transcriptional, and epigenetic levels (Kondo et al., 2017). Among these lncRNAs, malat1 is expressed in the nucleus, with a length of 7000 nt, and can effectively promote tumor progression by regulating cancer cell proliferation, migration, metastasis, and permeability of the blood tumor barrier (Zhang et al., 2017). In addition, it has been found that malat1 can stimulate the progression of non-small cell lung cancer, ovarian cancer, endometrial cancer, pancreatic cancer, and other tumors (Jiao et al., 2014; Wu et al., 2018; Zhou et al., 2016).
Bladder cancer (BC) is considered as one of the most common malignant tumor of the urinary system. Around 75% of diagnosed bladder cancers are non-muscle invasive tumors, whereas one-quarter of patients with muscle invasive tumors are often at a high risk of distant metastasis (Burger et al., 2013). Despite an early diagnosis and multimodal therapy providing the best treatment effect for patients with metastatic tumor, the 5-year survival rate is only 77% (Arrieta-Cortes et al., 2017). The most common risk factors for bladder cancer are smoking, occupational exposure to carcinogens, environmental pollution, and an unhealthy diet regimen (Burger et al., 2013). For example, occupational exposure assessment has suggested that the excretion of PFAS is mainly caused through the urinary system, thereby indicating the exposure target of the bladder (Worley et al., 2017). An epidemiological study has revealed that exposure to PFOS can significantly increase the risk of bladder cancer (Arrieta-Cortes et al., 2017). In metastatic bladder cancer, malat1 expression level was up-regulated, and the overexpressed malat1 could effectively activate the Wnt (Wingless/Integrated) pathway, eventually promoting epithelial-mesenchymal transition (EMT) and human bladder cancer cell metastasis (Ying et al., 2012).
A number of studies have found that an abnormal expression of lncRNA is related to a variety of human diseases, especially tumorigenesis (Geisler and Coller, 2013). More evidences have revealed that PFCs could regulate lncRNA expression both in vitro and in vivo (Chen et al., 2018; J. J. Li et al., 2020). For instance, PFOS exposure was found to enhance the expression of lncRNA H19 in the mouse placenta and HTR-8/SV neo cells. Overexpression of H19 reduced the levels of miR-19a and miR-19b, and finally inhibited the growth of placental cells. Prenatal PFOS exposure altered the expression of lncRNA xist, which may primarily act as ceRNA by sponging miR-429 to suppress vegf-a, leading to an abnormal placental angiogenesis (Chen et al., 2018). These findings indicated that the toxic effects of PFCs may be closely correlated with lncRNAs. Perfluorooctanoic acid (PFOA) could also potentially promote the migration and invasion of ovarian cancer and endometrial carcinoma (Li et al., 2018; Ma et al., 2016). However, it remains to be clarified whether PFOI exposure also affects tumor cell motility. The impact of PFOI exposure on bladder cancer and T24 cells can serve as an ideal model to study the unknown toxicological effects of PFOI.
In this study, the effect of PFOI exposure on the migration ability of T24 cells was studied through cell migration and invasion experiments, and the role of lncRNA malat1 in this process was further explored, and it was found to regulate the genes such as MMPs and cadherins to affect cellular motility.
Section snippets
Chemicals
PFOI (purity, 97%), Triton X-100, dimethyl sulfoxide (DMSO) and isopropanol were purchased from Sigma-Aldrich (St. Louis, MO, USA). Certified fetal bovine serum (FBS) and hormone-free FBS were purchased from Biological Industries (Beit HaEmek, Israel). McCOY's 5A medium, trypsin solution (0.05%), and penicillin-streptomycin were obtained from Gibco (Grand Island, NE, USA). The transmembrane plate and metrigel were obtained from Corning Company (New York, NY, USA). The cell counting kit-8
PFOI exposure enhanced the migration and invasion of T24 cells
To minimize the interference of cytotoxicity in the assay, the concentration used was selected based on the cell viability test. The results revealed that PFOI at concentration range (0.1–1000 μM) did not significantly induce cytotoxicity after 24 h or 48 h exposure (Fig. 1. A). This suggested the low cytotoxicity of PFOI towards T24 cells. Based on the plate culture (wound healing assay) and transwell culture (migration), the results indicated that PFOI promoted the migration of T24 cells in a
Discussion
PFCs and their carcinogenic effects have gained significant attention in the recent years. A number of epidemiological studies have revealed that the serum levels of PFCs might be potential risk factors in the development of breast cancer (Bonefeld-Jorgensen et al., 2011; Mancini et al., 2020). For instance, an in vitro study has demonstrated that PFCs could effectively promote the carcinogenesis of breast epithelial cells by inducing cell migration and proliferation (Pierozan et al., 2020).
Declaration of competing interest
The authors declare that they have no known competing financial interests or personal relationships that could have appeared to influence the work reported in this paper.
Acknowledgment
We thank Dr. Ling Wang from Jianghan University for her kind assistance with cell culture and experiments. This work was financially supported by grants from the National Natural Science Foundation of China (21677062, 21607059, 21507155). We thank MJEditor (www.mjeditor) for its linguistic assistance during the preparation of this manuscript.
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These authors contributed equally to this work.