Long non-coding RNA MEG3 promotes cataractogenesis by upregulating TP53INP1 expression in age-related cataract

https://doi.org/10.1016/j.exer.2020.108185Get rights and content

Highlights

  • MEG3 expression was up-regulated in the age-related cortical cataract (ARCC) lens capsules and positively correlated with the histological degree of ARCC.

  • TP53INP1 knockdown alleviated H2O2-induced lens turbidity.

  • MEG3 promoted ARC progression by up-regulating TP53INP1 expression through suppressing miR-223 and promoting P53 expression.

Abstract

Age-related cataract (ARC) is the leading cause of visual impairment or even blindness among the aged population globally. Long non-coding RNA (LncRNA) has been proven to be the potential regulator of ARC. The latest study reveals that maternally expressed gene 3 (MEG3) promotes the apoptosis and inhibits the proliferation of multiple cancer cells. However, the expression and role of MEG3 in ARC are unclear. In this study, we investigated the effects of MEG3 in ARC and explored the regulatory mechanisms underlying these effects. We observed that MEG3 expression was up-regulated in the age-related cortical cataract (ARCC) lens capsules and positively correlated with the histological degree of ARCC. The pro-apoptosis protein, active caspase-3 and Bax increased in the anterior lens capsules of ARCC tissue, while the anti-apoptotic protein Bcl-2 decreased compared to normal lens. Knockdown of MEG3 increased the viability and inhibited the apoptosis of LECs upon the oxidative stress induced by H2O2. MEG3 was localized in both nucleus and cytoplasm in LECs. MEG3 facilitated TP53INP1 expression via acting as miR-223 sponge and promoting P53 expression. Additionally, TP53INP1 knockdown alleviated H2O2-induced lens turbidity. In summary, MEG3 promoted ARC progression by up-regulating TP53INP1 expression through suppressing miR-223 and promoting P53 expression, which would provide a novel insight into the pathogenesis of ARC.

Introduction

Age-related cataract (ARC), an opacity in the lens substance, is the leading cause of visual impairment and even blindness among the elderly worldwide (Wang et al., 2016). The eye lens consists of a single layer of epithelial cells, which is the most active site of metabolism in the lens. At the equator, the epithelial cells elongate and differentiate into fiber cells. Damages to the lens epithelium and their components will ultimately result in ARC (Kantorow et al., 2004). Accumulating evidence indicates that oxidative stress is a leading cause of ARC (Crooke et al., 2017; Liu et al., 2017; Selin et al., 2014). Exposing the lens to oxidative stress of reactive oxygen species (ROS), such as H2O2, may in turn lead to apoptosis of the lens epithelium and then initiate cataract development, which eventually leads to the opacity of the lens (Ma et al., 2018; Smith et al., 2016; Yu et al., 2016). Therefore, oxidative stress-induced apoptosis of LECs becomes a common cellular basis for the occurrence and development of ARC. Although surgery has been considered the most effective treatment for cataracts, many postoperative complications still inevitably cause great damage to vision. Thus, strategies to prevent cataract formation should be determined based on the association between oxidative stress and cataract formation.

High-throughput transcriptome analyses reveal that non-coding RNA (ncRNA) accounts for the vast majority of transcripts, which constitute the regulatory molecules involved in response to stress and other environmental stimuli (Ponting et al., 2009). Long non-coding RNA (lncRNA) is a class of ncRNAs with lengths greater than 200 nt (Batista and Chang, 2013). A growing body of evidence has revealed that lncRNA is involved in oxidative stress and that aberrant expression of lncRNA is associated with several ocular diseases, including ARC (Li et al., 2016; Schmitz et al., 2016; Wang et al., 2019). LncRNA myocardial infarction-associated transcript (MIAT) functions as competitive endogenous RNA (ceRNA) and forms a feedback loop with miR-150-5p and Akt to regulate LEC proliferation, migration, and apoptosis (Shen et al., 2016) The H19/miR-675 axis participates in the regulation of LEC function by targeting CRYAA in ARC (Liu et al., 2018). These studies indicate that lncRNA plays an important role in the regulation of ARC, which needs to be further studied. However, only a few lncRNAs have been implicated in ARC.

Maternally expressed gene 3 (MEG3) is a tumour suppressor gene that is located on human chromosome 14q32 (Balik et al., 2013; Zhou et al., 2012). The latest study revealed that MEG3 promotes apoptosis and inhibits the proliferation of cancer cells (Jia et al., 2014; Luo et al., 2015). For example, its downregulation has been documented in multiple myeloma (Shen et al., 2018), renal cell carcinoma (He et al., 2018), and bladder cancer (BC) (Feng et al., 2018). MEG3 overexpression in human proximal tubule epithelial cells restrains cell viability and promotes cell apoptosis by regulating miR-7/RASL11B signalling (He et al., 2018). Moreover, treatment with the chemotherapy drug cisplatin can significantly induce cell apoptosis in MEG3-overexpressing BC cells. In addition, MEG3 and p53 can stimulate the expression of each other in BC cells, suggesting a potential positive feedback loop between MEG3 and p53. Recently, dysregulation of MEG3 has also been reported in ocular diseases. MEG3 promotes apoptosis and suppresses proliferation of retinoblastoma cells by negatively regulating the activity of the Wnt/β-catenin pathway (Gao and Lu, 2016). Silencing of MEG3 protects photoreceptor cells against light stress-induced apoptosis through p53-mediated transactivation (Zhu et al., 2018). However, the exact role of MEG3 in the pathogenesis of ARC remains unknown.

In our study, we detected the expression of MEG3 in human anterior lens capsules of normal and ARC crystals. We found that MEG3 expression was elevated in the ARC group. MEG3 knockdown could promote the viability and inhibit the apoptosis of LECs. Furthermore, we also investigated the underlying molecular mechanisms by which MEG3 plays a regulatory role in lens epithelium.

Section snippets

Human lens epithelium samples

A total of 30 patients (14 males and 16 females) aged 50–70 years were diagnosed with ARC and underwent cataract surgery at the Affiliated Hospital of Nantong University, where capsulorhexis specimens of anterior lens capsules were collected. According to the opacity region of the lens, ARC was classified into three subtypes: age-related cortical cataract (ARCC), age-related nuclear cataract (ARNC) and age-related posterior subcapsular cataract (ARPC). The type and degree of lenticular

MEG3 and cell apoptosis increase in anterior lens capsules of ARCC patients

MEG3 was observed to be elevated in ARC patient lens capsules compared to control (Fig. 1A). MEG3 was significantly upregulated in the ARCC group but remained unchanged in the ARNC and ARPC groups (Fig. 1B). MEG3 expression was positively correlated with the histological degree of ARCC (Fig. 1C). The pro-apoptotic protein active caspase-3 and Bax increased in the lens capsules of the ARCC group, while the anti-apoptosis protein Bcl-2 decreased (Fig. 1D–F). The data suggested that MEG3 and cell

Discussion

Recent studies have demonstrated the important role of lncRNAs in cataracts (Li et al., 2017; Liu et al., 2018; Shen et al., 2016). In our study, we reported that MEG3 was involved in the pathology of ARC. MEG3 was highly expressed in the anterior lens capsules of ARCC patients and in H2O2-treated SRA01/04 cells. MEG3 knockdown inhibited SRA01/04 cell apoptosis and promoted cell viability under oxidative stress. Furthermore, MEG3 exerted a pro-apoptotic role by upregulating TP53INP1 expression

Declaration of competing interest

All authors declare that they have no conflict of interest.

Acknowledgements

The study was supported by the National Natural Science Foundation of China (No. 81770906), the Suzhou Science and Technology Bureau (No. SYS2018005) and the Suzhou Commission of Health and Family Planning (No. KJXW2018076).

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      Another report also found that lncKCNQ1OT1 is the sponge of miR-223–3p and BCL2L2 is the direct target of miR-223–3p, thus promoting apoptosis and oxidative stress of H2O2-treated LECs through the KCNQ1OT1/miR-233–3p/BCL2L2 axis [206]. In addition to lncKCNQ1OT1, several novel promotive lncRNAs have been described only in the past few years, including taurine upregulated 1 (TUG1) [207], myocardial infarction association transcript (MIAT) [208], maternally expressed gene 3 (MEG3) [209], nuclear paraspeckle assembly transcript 1 (NEAT1) [210] and Opa interacting protein 5 antisense RNA 1 (OIP5-AS1) [211], and this list will undoubtedly continue to grow (Table 1). Some lncRNAs act as safeguards against ARC development by inhibiting LEC apoptosis, alleviating oxidative damage or maintaining cell viability.

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      Tang et al. declared that inhibition of miR-223-3p ameliorated hypoxia-induced injury via inhibiting oxidative stress and cardiomyocyte apoptosis via regulating KLF15 (Tang et al., 2018). Besides, Tu et al. revealed that miR-223 abundance was decreased in cataract patients and H2O2-treated SRA01/04 cells, and miR-223 overexpression suppressed cell apoptosis in H2O2-induced SRA01/04 cells by downregulating TP53INP1 (Tu et al., 2020). Here, we also uncovered that miR-223-3p was downregulated in cataract anterior lens capsule samples and H2O2-exposed SRA01/04 cells, and its expression was inversely correlated with KCNQ1OT1 abundance in cataract anterior lens capsule tissues.

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