Dysregulation of microRNAs in metal-induced angiogenesis and carcinogenesis

Abstract

MicroRNAs (miRNAs) are small endogenous non-coding RNAs that regulate cancer initiation, development, angiogenesis, and therapeutic resistance. Metal exposure widely occurs through air, water, soil, food, and industrial contaminants. Hundreds of millions of people may have metal exposure associated with toxicity, serious health problems, and cancer occurrence. Metal exposure is found to induce oxidative stress, DNA damage and repair, and activation of multiple signaling pathways. However, molecular mechanisms of metal-induced carcinogenesis remain to be elucidated. Recent studies demonstrated that the exposure of metals such as arsenic, hexavalent chromium, cadmium, and nickel caused dysregulation of microRNAs that are implicated to play an important role in cell transformation, tumor growth and angiogenesis. This review focuses on the recent studies that show metal-induced miRNA dysregulation and underlined mechanisms in cell malignant transformation, angiogenesis and tumor growth.

1. Introduction

MicroRNAs (miRNAs) are small endogenous non-coding RNAs with the mature miRNA length of 18-23 base pairs. The first miRNA was discovered in the C. elegans as heterochronic gene lin-4 to control the gene expression of different coding genes by Ambros and colleagues in 1993[1]. Most miRNA genes are transcribed as long primary RNAs, called pre-miRNAs, and pri-miRNAs are initially processed in the nucleus by DROSHA, then released as hairpin-shaped precursors (pre-miRNA) and exported from nucleus to cytoplasm and cleaved by DICER, to generate a miRNA/miRNA* duplex [2–5]. The miRNA/miRNA* duplexes are loaded into an Argonaute (Ago) protein, which preferentially ejects the miRNA* strand, while retains the mature miRNA [6]. Additionally, Ago protein in cells may be associated with cofactors of the GW182/TNRC6 family proteins to form the RNA-induced silencing complex (RISC). Mature miRNA/Ago complexes recognize target mRNAs in cells by pairing miRNA seed regions with mRNAs [7]. Majority of human mRNAs are regulated by evolutionarily conserved miRNAs with the possibility of a particular miRNA which may post transcriptionally regulate hundreds of target mRNAs, and of each mRNA which may be suppressed by several different types of miRNAs [7]. miRNA stability and abundance can be altered during the pathological processes, and such multifaceted regulation of miRNAs ultimately influence gross changes in mRNA levels and gene expression that can regulate cancer development and numerous diseases [8].

Croce and colleagues initially discovered the importance of miRNAs in human cancer development when they found that miR-15a/16-1 cluster was frequently deleted in chronic lymphocytic leukemia (CLL), and miR-15a/16-1 loss was directly linked to Bcl2 overexpression [9]. After the discovery, alterations of miRNA expression have been found in all kinds of human cancers[10–12]. Most miRNAs have been found to be downregulated in human cancer tissues compared to normal tissue counterparts, and to function as tumor suppressor-like miRNAs by directly targeting oncogenes; while only a small portion of miRNAs that are upregulated in cancer tissues and function as oncogene-like miRNAs by directly suppressing tumor suppressor genes [13–15]. Huge body of information has been learned about the key roles of miRNAs in regulating cancer development, angiogenesis and drug resistance [16–19]. Several metals including hexavalent chromium, arsenic, cadmium, and nickel are known carcinogens through the exposure of contaminated air, soil, water, and food; and the human exposures are strongly associated with various types of cancer development [20–23]. In this review, we will summarize a growing list of recent studies that have shown that miRNA dysregulations are important in metal-induced cell transformation, tumor growth and angiogenesis.

2. MiRNA dysregulations in cancer development, angiogenesis and drug resistance

Since the discovery of the important role of miR-15a/16 cluster in chronic lymphocytic leukemia development, the field of miRNA dysregulation in cancer biology has attracted emerging and expanded interests, and huge number of new findings have been achieved to understand roles and mechanisms of miRNAs in cancer development, angiogenesis, diagnostics values, and therapeutic resistance [12, 24–26]. Tumor angiogenesis is vital to tumor growth and metastasis, which is the process that new vessels sprout from preexisting blood vessels, and miR-21 promoted tumor angiogenesis via inducing HIF-1α and VEGF expression as well as activation of AKT and ERK pathways[27]. MiR-145 was suggested as a tumor suppressor that inhibits angiogenesis and tumor growth by inhibiting HIF-1α and VEGF expression [28]. Functional studies have demonstrated that miRNAs can act as tumor suppressors by directly suppressing oncogenes such as Myc and Ras, or act as oncogenes by targeting tumor suppressors such as p53 and PTEN [29]. MiRNAs have been used as cancer biomarkers to develop therapeutic kits that have been used for clinical diagnostics [30]. MiRNA-based therapeutics have reached clinical application including miR-34 which reached phase I clinical trials and miR-122 which reached phase II trials for treating hepatitis[13]. Cancer drug resistance is one of the major hurdles for advanced cancer treatments. The growing list of evidence has showed microRNAs play important roles in different types of cancer drug resistance. A few examples are described here. MiR-21 upregulation increased breast cancer cell resistance to DOX treatment by downregulating PTEN expression [31]. MiR-152 expression levels were dramatically decreased in the cisplatin-resistant ovarian cancer cell lines, and forced expression of miR-152 made the resistant cells sensitive to cisplatin treatment [32]. Downregulation of miR-579-3p played an important role in melanoma cancer cells resistant to BRAF/MEK inhibitors, and overexpression of miR-579-3p targeted the 3′UTR of two oncoproteins: BRAF and MDM2 for the inhibition, and restored the therapy sensitivity [33]. Downregulation of miR-143 was involved in resistance of colorectal cancer cells to oxaliplatin resistance, and forced expression of miR-143 restored oxaliplatin chemosensitivity to oxaliplatin treatment [34].

3. Arsenic

Arsenic is considered to be among the leading environmental carcinogens in the United States and in the world. Approximately 200 million people in the world are exposed to high levels of arsenic [35]. Populations are exposed to arsenic through the arsenic-containing drinking water, food, and air-dust. The occupational exposure to arsenic may occur through the inhalation of arsenic-containing particles in the production and distribution processes. There are more than 1.5 million industrial workers that are potentially exposed to arsenic and arsenic compounds according to the NIOSH estimate [36]. Higher risks of cancer were associated with the occupational or environmental arsenic exposure leading to skin, lung, bladder, and liver cancers [37–40]. Although mechanisms of arsenic-induced carcinogenesis are still poorly understood, arsenic is considered to mainly alter epigenetic changes and gene expression, but not as a genotoxic carcinogen to induce gene mutations. Arsenic can increase oxidative stress, activate hypoxia and HIF-1 and other signaling pathways, and inhibit DNA repair that may contribute to the effect of arsenic in inducing angiogenesis and carcinogenesis [41, 42]. In human and rat bladder epithelial cells, chronic arsenic exposure upregulated HER2 expression, HER2 promoted cell proliferation, migration and angiogenesis by activating multiple signaling pathways, such as MAPK, PI3K/AKT and Src/STAT3 pathways in response to arsenic treatment [43]. Recent evidence showed that arsenic exposure caused DNA hypermethylation such as the hypermethylation of promoters of tumor suppressors p53 and p16 which inhibits their expression for inducing cell transformation and tumor growth [44, 45]. A list of recent studies found that dysregulations of miRNAs play an important role in arsenic-inducing carcinogenesis and angiogenesis (Table 1). The first evidence of critical role of arsenic-induced mi-RNA downregulation in cell transformation came from the study by the exposure of immortalized human bronchial epithelial cells (HBECs) with p53 knocked-down (p53lowHBECs) to low concentration of arsenic for 16 weeks, these p53lowHBECs cells became malignant transformation with morphology switch from epithelial to a spindle-like mesenchymal morphology, and greatly suppressed miR-200b/c expression [46]. The forced expression of miR-200b in the transformed cells completely inhibited the malignant transformation [46]. Similarly, downregulation of miR-200 family was observed in other cell types and human urine samples in response to arsenic exposure [47]. The downregulation of miR-199a-5p was observed in human bronchial epithelial BEAS-2B cells exposed to 1.0 μM arsenic for 26 weeks, and levels of miR-199a-5p in arsenic transformed cells were diminished to 100-fold lower than those in arsenic unexposed parental cells [48]. The suppression of miR-199a-5p greatly increased the expression levels of its direct targets HIF-1α and COX-2, and forced expression of miR-199a-5p diminished HIF-1α and COX-2 expression and impaired arsenic-induced angiogenesis and tumor growth [48]. Interestingly, miR-199a-5p expression was inhibited by ROS production in the cells through the promoter methylation of miR-199a gene by DNA methyltransferase 1[16]; while arsenic can induce ROS production in different cell types [49].

Table 1.

Arsenic exposure-induced alterations of miRNAs and carcinogenesis

miRNA	Alteration	Role	Target(s)	Role in carcinogenesis	References
miR-217	Down	Tumor suppressor	EZH2	Inhibits cell cycle of human keratinocytes in carcinogenesis	[120]
miR-155	Up	Oncogene	Quaking	Activates STAT3 and induces IL6 and IL-8 in As-transformed liver cells; promotes CSC- like phenotype during the neoplastic transformation	[50, 121]
miR-21	Up	Oncogene	Spry1, PTEN, PDCD4	Enhances the neoplastic capacity of As-transformed cells; induces angiogenesis and EMT	[51, 56, 122, 123]
miR-182-5p	Down	Tumor suppressor	N/A	contributes to arsenic carcinogenesis via HIF-2α	[124]
miR-889	Up	Oncogene	DAB2IP	Promotes As-induced acquisition of CSC-like properties via DAB2IP/ZEB1	[125]
miR-455	Down	Tumor suppressor	ZEB1	Suppresses EMT during As-induced malignant transformation of human keratinocytes	[126]
miR-15b	Up	Oncogene	LATS1	inhibits the Hippo pathway and promotes cell proliferation, migration, and invasion	[127]
miR-191	Up	Oncogene	BASP1	HIF-2α promotes miR-191 expression to induce angiogenesis and EMT in As-transformed cells	[60, 128, 129]
miR-186	Up	Oncogene	BUB1	induces increased chromosome numbers and chromosome structural abnormalities	[130]
let-7a/b/c	Down	Tumor suppressor	N/A	Suppresses cancer stem cell-like properties by inhibiting Ras/NF-κB pathway	[131]
miR-181b and miR-9	Down	Tumor suppressor	neuropilin - 1	Inhibits arsenic- induced angiogenesis	[132]
miR-134, miR-373, miR-155, miR-138, miR-205, miR-181d/c, let-7, miR-143, miR-34c-5p, and miR-205	Down	Tumor suppressors	RAN, RAB27A, RAB22A, KRAS, NRAS, and RRAS	Suppress RAS activation and transformation	[133]
miR-21, miR-200a and miR-141	Up	Oncogenes	CDK6, BMPR2, ACVR2B, and phosphati dylinositol-3-phosphate/5-kinase genes	Affect MAPK, Wnt and the PI3K pathways	[58]
miR-200b	Down	Tumor suppressor	N/A	As inhibits miR-200b expression through inducing ZEB1 and ZEB2 and hypermethylation, which promotes EMT and malignant transformation	[46]
miR-410, miR-548ac, miR-3174	Down	Tumor suppressor	HMGB1, MDM2	Regulate TP53 regulatory network including MDM2 and HMGB1	[134]
miR-192b-5p, miR-15b-5p, and miR-33b-5p	Down	N/A	N/A	Downregulated in As-transformed HBC cells	[135]
miR-141-3p, miR-106b-5p, and miR-200b-3p	Up	N/A	N/A	Upregulated in As-transformed HBC cells	[135]
miR-221, miR-222 and miR-638	Up	N/A	RNF4	Upregulated upon 2 μM As treatment at 24 and 144 h	[59]
miR-190	Up	Oncogenes	PHLPP	cause Akt activation and increase VEGF expression	[54]

Arsenic can also upregulate miRNA expression such as miR-21, miR-222, miR-190, and miR-155[50–54]. MiR-21 expression levels were higher in human serum samples from individuals with arsenic exposure compared to those of the nonexposed subjects and were higher in HaCaT cells with arsenic exposure [55]. MiR-21 upregulation may directly target and inhibit tumor suppressors such as PTEN and programmed cell death 4 (PDCD4) expression [52, 56]. MiR-222 expression was significantly induced by arsenic exposure, and miR-222 in turn inhibited PTEN expression for inducing cell transformation and tumor growth [53]. MiR-190 levels were induced by arsenic to suppress the PH domain leucine-rich repeat protein phosphatase (PHLPP) expression, which is a negative regulator of Akt signaling [54]. MiR-155 expression levels were markedly increased by arsenic exposure through NF-κB induction [50]. Other miRNAs that are regulated by arsenic including Let 7 family, miR-141, miR-433, miR-184, miR-191, miR-151, miR-148b, and miR-183[55, 57–60].

4. Chromium

Chromium may be in three different states: Cr (0), Cr(III), and Cr(VI); and only Cr(VI) is known carcinogen. Environmental and occupational Cr(VI) exposure through water, air or landfills has emerged as a major public health concern, and is associated with human lung cancer[61–63]. Changes in signaling pathways and oxidative stress have been implicated as causal factors in response to Cr(VI) exposure[64–66]. The epigenome was reported to be altered by chromium, the chromatin state changes through histone modifications as well as the DNA methylation landscape [67, 68]. However, the mechanisms underlying Cr(VI)-induced carcinogenesis and angiogenesis remain to be elucidated. The miRNA dysregulation was recently shown to be important in Cr(VI)-induced cell transformation, carcinogenesis and angiogenesis (Table 2). Our group initially demonstrated that miR-143 down-regulation played an important role in Cr(VI)-transformed human bronchial epithelial BEAS-2B cell transformation and angiogenesis[69]. Repression of miR-143 led to the transformation and angiogenesis of the cell, induced by Cr(VI), through upregulation of insulin-like growth factor 1 (IGF-IR) receptor and insulin receptor substrate 1 (IRS1). Furthermore, interleukin-8 was induced by Cr (VI) treatment, which was a major upregulated angiogenesis inducer through activations of the IGF-IR/IRS1 axis and ERK/HIF-1α/NF-κB signal pathway. This result suggest that the miR-143/IL-6/HIF-1α signaling pathway plays a vital role in Cr(VI)-induced cell malignant transformation and carcinogenesis[70]. MiR-143 suppression was also implicated in arsenic-induced cell transformation of the human normal prostate stem/progenitor cell (SC) line, which was a causal effect due to loss of miR-143 expression during the formation of prostate CSCs (As-CSCs). The forced miR-143 over-expression in these CSCs led to a considerable decrease in various types of cancer [71].

Table 2.

Chromium exposure-induced alterations of miRNAs and carcinogenesis

miRNA	Alteration	Role	Target(s)	Role in carcinogenesis	References
miR-143	Down	Tumor suppressor	IGF-IR, IRS	Suppress cell transformation and tumor angiogenesis via IGF-IR/IRS1/ERK/IL-8 pathway	[69]
miR-222	Up	Oncogene		Promote MAPK and nerve growth factor pathways	[72]
miR-494	Down	Tumor suppressor	c-Myc	Inhibit CSC-like property and carcinogenesis by targeting c-Myc	[75]
miR-3940-5p	Down	Tumor suppressor	XRCC2, BRCC3	Protective effect in Cr(VI) induced DNA damage	[76]
miR-19a-3p, miR-19b-3p and miR-142-3p	Down	N/A	See Ref	Regulate MAPK, cAMP, cGMP-PKG, and FoxO signaling pathways	[77]
miR-590-3p	Up	N/A	See Ref	Regulate MAPK, cAMP, cGMP-PKG, and FoxO signaling pathways	[77]
miR-21 and miR-155	Down	N/A	N/A	N/A	[78]

In the study of foundry workers in an electric-furnace steel plant, expression levels miR-222 were increased in post-exposition samples collected after 3 working days compared to the baseline samples [72]. MiR-222 expression levels were greatly increased in non-current smokers in response to chromium exposure, which may be regulated by MAPK and Nerve growth factor pathways [72]. Exposure to Cr(VI) was reported to induce CSC-like property through c-Myc which was significantly higher in human bronchial epithelial cells transformed by Cr(VI) compared with the control cells [73, 74]. Through using c-Myc shRNA, they found the significant decrease in the CSC-likes property of the Cr(VI)-transformed cells by downregulation of c-Myc expression, as evidenced by their decreased ability to form suspension spheres and tumorigenicity in nude mice. miR-494 expression was significantly down-regulated in Cr(VI)-transformed cells, and overexpression of miR-494 in Cr(VI)-transformed cells reduced c-Myc protein level, decreased Cr(VI)-induced CSC-like property and tumorigenesis[75]. In chromate manufacturing workers, lower levels of miR-3940-5p were associated with the lower blood Cr level, and miR-3940-5p mediated XRCC2 signal to promote DNA repair on DNA double-strand breaks induced by Cr(VI) exposure [76].

Other study showed that miR-19a-3p, miR-19b-3p and miR-142-3p were downregulated and miR-590-3p and miR-941 were upregulated in the plasma of workers occupationally exposed to Cr (VI) [77]. Target gene analysis showed that the miRNAs may participate in the regulation of DNA damage-related signaling pathways including cAMP, MAPK, FoxO, and cGMP-PKG pathways [77]. Welding fumes contain gases and metal oxides of chromium, cadmium, lead, nickel and others, which could be a threat on the reproductive system and renal function. In the study of welders, the correlations of blood and/or urine metal levels and expression levels of miR-21 and miR-155 were observed [78].

5. Nickel

Nickel is known as a common environmental pollutant and potent carcinogen. At the beginning of life, Nickel existed as a metal cofactor in the metabolism of methanogenic archaea [79, 80]. Nickel also functions as an essential element in biosynthesis of enzymes for bacteria and plant[81]. Ni compounds have been widely used in various human activities such as fossil fuel combustion and disposal from industries that can release almost 180,000 tons of nickel every year, which is a big burden to the environment. Studies suggested that workers under chronic Ni compounds exposure exhibited increased risk of nasal sinus cancer as well as lung cancer[82]. Thus, Nickel compounds has been classified as Class I human carcinogens by IARC [64, 83, 84].

Ni was found to induce gene expression which was associated with cancer and hypoxia, and Ni inhibited HIF-prolyl and asparaginyl-hydroxylases for stabilization of HIF-1α and HIF-1-dependant transcription[85]. Nickel may increase deletion mutations, DNA protein linkage and chromosomal aberrations in several report [86–88]. Further investigation showed that Ni may preferably induce epigenetic change, rather than mutation [89]. Chronic Ni exposure was showed to induce changes in methylation of histone H3, histone H4 acetylation, phosphorylation of H3S10, and histones H2A/H2B ubiquitination[89, 90]. Nickel may decrease SLBP mRNA expression by regulating SLBP promoter region, such as declining histone acetylation or increasing DNA methylation, then promote SLBP protein degradation, finally cause H3.1 polyadenylation[91]. NiCl2 increased VEGF expression through activation of Akt, ERK, NF-κB and suppression of AMPK[43].

Nickel has been showed to induce dysregulation of several kinds of miRNAs (Table 3). miR-152 downregulation may be involved in cell transformation [92]. C57BL/6J wildtype mice treated with Nano-Ni increased miR-21 expression and proinflammatory cytokines such as IL-6, TNFα, TGF-β1, COL1A1 and COL3A1 [93], and human cohort study showed that higher miR-21 levels were associated with Ni exposure, EGFR mutation and lung tumor invasion[94]. MiR-4417 levels were the most upregulated miRNA by NiCl₂ treatment in BEAS-2B and A549, and miR-4417 induction induced fibrogenesis, EMT and tumor progression through its target TAB2 [95].

Table 3.

Nickel exposure-induced alterations of miRNAs and carcinogenesis

miRNA	Alteration	Role	Target(s)	Role in carcinogenesis	References
miR-4417	Up	Oncogene	TAB2	miR-4417/TAB2 involved in nickel-induced fibronectin, EMT and carcinogenesis	[95]
miR-210	Up	Oncogene	ISCU1/2	Regulate n nickel induced aerobic glycolysis	[98]
miR-21	Up	Oncogene	c-Myc	Activate EGFR/NF-κB signaling pathway and promote carcinogenesis and invasiveness	[94]
miR-203	Down	Tumor suppressor	ABL1	Inhibit carcinogenesis in vitro and in vivo by targeting ABL1	[99]
miR-152	Down	Tumor suppressor	DNMT1	Inhibit cell growth, malignant transformation and DNMT1 via a feedback mechanism	[92]
miR-222	Up	Oncogene	CDKN1B, CDKN1C	promote carcinogenesis and regulate target genes	[136]

Aerobic glycolysis was significantly increased in cells treated with NiCl₂, and was considered to play an important role in nickel-induced cell transformation and carcinogenesis [96, 97]. During glycolysis process, phosphofructokinase (PFK) catalyzes fructose-6-phosphate to fructose-1,6-diphosphate, which also considered a rate-limiting step. To investigate the mechanism of nickel in inducing carcinogenesis, nickel was showed to upregulate miR-210 and HIF-1α expression, ROS scavenger, N-acetylcysteine(NAC) and melatonin attenuated HIF-1α and miR-210 expression, and HIF-1α/miR210/ISCU axis was important in aerobic glycolysis Ni-induced tumorigenesis [97, 98]. MiR-203 was reported to be significantly downregulated in Ni₃S₂-transformed 16HBE cells though the hypermethylation of miR-203 promoter region, miR-203 overexpression inhibited cell proliferation, cell migration and anchorage-independent growth [99]. Nickel induced miR-203 suppression, and increased direct target gene ABL1 expression to promote tumor development [99].

6. Cadmium

Cadmium(Cd) is a metal that is rare in earth’s crust, Cadmium is typically functioned in a diatom for living in the marine environment that utilize the Cd-enzyme[100]. Cadmium accumulates in animals and plants with a long half-life of about 25-30 years, usually released into the environment as a well-known environmental and occupational pollutant in the world due to anthropogenic activities, which has been a significant public health concern for decades[101]. In 1993, cadmium and its compounds are classified as a Group 1 carcinogen by IARC[102]. Recent studies indicated that cadmium exposure increased the risk of several human cancers such as thyroid cancer, prostate cancer, breast bladder and lung cancers [103–106].

Cadmium has a broad range of cell and molecular effects, both genetic and epigenetic, that may affect all stages of carcinogenic activity. The major mechanisms of cadmium-induced carcinogenesis include the induction of oxidative stress, ROS levels, decreased DNA repair capacity as well as epigenetic changes and alteration in gene expression[107–109]. Cadmium induces many biochemical changes in experimental systems, including E-cadherin dysfunction, DNA methylation inhibition as well as associated with apoptosis by increasing p53 protein and mRNA levels[110]. After a low-dose Cd treatment, the signaling molecules associated with tumor angiogenesis of both tumor cells and epithelial cells directly were altered. Cadmium treatment increased HIF-1α and VEGF expression, and induced ROS, ERK and AKT signaling pathways [111]. Low-dose exposure to Cd also promoted the proliferation activity of lung cancer cells A549 to induce angiogenesis[112].

A list of evidence showed that cadmium-mediated microRNA dysregulations were important in cell proliferation, EMT, transformation, and cancer invasion (Table 4). Recent investigations showed that Cd induced the downregulation of miR-30 expression in lung epithelial cells, and miR-30 overexpression inhibited SNAIL gene expression to immediate Cd-induced EMT progression[106]. Dysregulations of hsa-miR-27b-3p, hsa-miR-1265, hsa-miR-944, hsv1-miR-H6-5p, hsa-miR-3960, hsa-miR-1261, hsa-miR-877-5p, and hsa-miR-4708-3p were observed in cadmium-induced cell transformation[113]. miR-101 expression levels were significantly decreased by CdCl₂ treatment, and miR-101 overexpression directly targeted and inhibited COX-2 to decease ER stress/COX-2/VEGF pathway and angiogenesis [114].Other miRNAs were also affected by Cd treatment [115]. Interestingly, miRNA-21 expression was also upregulated in bladder cancer, the miRNA-21 expression levels were significantly higher in the samples obtained from occupationally metal exposed workers than those from the nonexposed subjects [116], and miRNA-21 would play a vital role in metal-induced bladder carcinogenesis[117].

Table 4.

Cadmium exposure-induced alterations of miRNAs and carcinogenesis

miRNA	Alteration	Role	Target(s)	Role in carcinogenesis	References
miR-21	Up	Oncogene	N/A	miR-21 in cancerous tissues was significantly associated with blood concentration of Cd	[117]
miR-122-5p, miR-326-3p	Up	N/A	PLD1	enhance cadmium-induced apoptosis in NRK-52E cell through inhibiting PLD1 expression	[119]
miR-125a/b	Down	N/A	Bak, Caspase3	regulate the suppression of Cd-induced apoptosis by Se via the mitochondrial pathway in LLC-PK1 cells	[137]
miR-30a	Down	N/A	N/A	Cd triggered an miR-30a-GRP78 signaling axis disorder, increasing ER stress and activating the IRE-1-JNK pathway	[138]
miR-30 family(miR-30a/b/c/d/e)	Down	Tumor suppressor	SNAIL	inhibit SNAIL levels, acquisition of EMT phenotype and carcinogenesis	[106]
miR-363-3p	Up	N/A	PI3K	promote apoptosis by down-regulating the expression of PI3K	[139]

MiRNA microarray was used to test expression levels of miRNAs in Cd-exposed rat ovarian granulosa cells(OGCs), 19 miRNAs were altered such as miR-133, miR-204-5p, miR-195, let-7 family, miR-105 and miR-135 in response to Cd treatment, and miR-204-5p/Bcl2 were involved in Cd-induced damage and apoptosis in OGCs[118]. In addition, miR-122-5p and miR-326-3p were involved in cadmium-induced apoptosis in NRK-52E cells through inhibiting PLD1 expression[119].

7. Concluding remarks and future direction

Chronic metal exposure may affect several hundreds of millions of people around the globe. MiRNA dysregulations have recently demonstrated to play an important role in metal-induced angiogenesis and carcinogenesis. Majority of miRNAs are downregulated by arsenic, Cr(VI), nickel and Cd, and these miRNAs function as tumor suppressor-like miRNAs that directly target and inhibit oncogene expression; while some miRNAs are upregulated by metal exposure and may function as oncogene-like miRNAs such as miR-21 and miR-122 that directly target and inhibit tumor suppressor PTEN expression. MiRNA expression is mainly mediated through epigenetic regulation including DNA methylation and transcription factors in the cells in response to metal exposure. Though significant progress has been made over the past decade, further studies are required to define the mechanisms for miRNA dysregulation induced by metal exposure. Further study is also required to understand the regulation mechanisms of key miRNAs that are important in regulating carcinogenesis and angiogenesis. The functional effects of miRNAs are through their target proteins, the expression levels of multiple potential target proteins are important to be analyzed for us to fully understand role and mechanism of miRNAs in metal-induced carcinogenesis. In addition, most results have been obtained through cell models, the future studies using animal models and human blood and urine samples are important for us to obtain new biomarkers, knowledge and findings that will be used to develop early diagnostic strategy, new preventive methods or/and treatment options for metal exposure-induced human diseases in the future.