MiR-330-5p regulates tyrosinase and PDIA3 expression and suppresses cell proliferation and invasion in cutaneous malignant melanoma.

BACKGROUND: Increasing evidence has suggested that miR-330-5p can function as a tumor suppressor in different types of cancers. However, the effects and underlying mechanisms of miR-330-5p in the development of cutaneous malignant melanoma (CMM) remain largely unknown. The aim of the present study was to investigate the role of miR-330-5p in CMM and to determine the molecular mechanisms underlying its action. MATERIALS AND METHODS: The expression level of miR-330-5p was detected in 26 cases of primary CMM tissues and cell lines by real-time quantitative polymerase chain reaction. We also assessed whether overexpression of miR-330-5p influences in vitro cell proliferation, invasion, and migration. Western blotting analysis was used to detect the influence of miR-330-5p on the targets, and Pearson analysis was used to calculate the correlation between the expression of targets gene and miR-330-5p in CMM tissues. RESULTS: Our study showed that miR-330-5p was downregulated in CMM tissues (P = 0.010) and cell lines (P < 0.05), and patients with high mitotic activity showed lower miR-330-5p expression levels (P = 0.002). Enforced expression of miR-330-5p inhibits malignant CMM cells proliferation and migration and led to downregulation of the TYR and PDIA3 protein. Moreover, the expression level of miR-330-5p in CMM tissues showed inverse relationship with the expression level of TYR and PDIA3 protein. CONCLUSIONS: In conclusion, our findings suggested that miR-330-5p represents a potential tumor-suppressive miRNA and plays an important role in CMM progression by suppressing TYR and PDIA3 expression.

Circular RNA circSEC24A Promotes Cutaneous Squamous Cell Carcinoma Progression by Regulating miR-1193/MAP3K9 Axis.

BACKGROUND: Circular RNAs (circRNAs) have been increasingly demonstrated to play critical roles in cancer progression. However, the biological functions and underlying mechanism of circRNA SEC24 homolog A, COPII coat complex component (circSEC24A) in cutaneous squamous cell carcinoma (CSCC) have not been well elucidated yet. METHODS: The expression levels of circSEC24A, microRNA-1193 (miR-1193) and mitogen-activated protein kinase kinase kinase 9 (MAP3K9) were analyzed by quantitative real-time polymerase chain reaction (qRT-PCR) or Western blot. 3-(4,5-dimethylthiazol-2-yl)-2,5-diphenyltetrazolium bromide (MTT) assay and colony formation assay were used to assess cell proliferation ability. Flow cytometry and transwell assay were utilized to detect cell apoptosis and migration and invasion. Glycolytic metabolism was examined via the measurement of lactate production, glucose consumption, extracellular acidification rate (ECAR), hexokinase 2 (HK2) and Lactate dehydrogenase A (LDHA) expression. The interaction between miR-1193 and circSEC24A or MAP3K9 was predicted by starBase v2.0 and verified by dual-luciferase reporter, RNA Immunoprecipitation (RIP) and pull-down assay assays. The mice xenograft model was established to investigate the roles of circSEC24A in vivo. RESULTS: CircSEC24A and MAP3K9 were upregulated and miR-1193 was downregulated in CSCC tissues and cells. CircSEC24A knockdown inhibited the progression of CSCC cells by inhibiting cell proliferation, migration, invasion, and glycolysis and inducing apoptosis. Moreover, miR-1193 was a direct target of circSEC24A and its downregulation reversed the inhibitory effect of circSEC24A knockdown on the progression of CSCC cells. Furthermore, MAP3K9 was a downstream target of miR-1193 and its upregulation attenuated the anti-cancer role of miR-1193 in CSCC cells. Additionally, circSEC24A acted as a molecular sponge of miR-1193 to regulate MAP3K9 expression. Furthermore, interference of circSEC24A repressed tumor growth via upregulating miR-1193 and downregulating MAP3K9. CONCLUSION: CircSEC24A interference suppressed the progression of CSCC by regulating miR-1193/MAP3K9 axis, which might be a promising strategy for CSCC treatment.

LncRNA MIR205HG regulates melanomagenesis via the miR-299-3p/VEGFA axis.

In this study, we investigated the role of lncRNA MIR205HG in melanomagenesis. Quantitative real-time PCR (qRT-PCR) analysis showed that MIR205HG levels were significantly upregulated in melanoma cell lines compared to normal human melanocytes. Similarly, MIR205HG levels were significantly higher melanoma tissues than adjacent normal skin tissues (n=30). CCK-8 and flow cytometry assays showed that MIR205HG knockdown significantly decreased the viability of melanoma cells. Dual luciferase reporter and RNA pull-down assays confirmed that MIR205HG directly binds to microRNA (miR)-299-3p. Targetscan analysis and dual luciferase reporter assays showed that miR-299-3p directly binds to the 3'UTR of VEGFA mRNA. Wound healing and transwell invasion assays showed that MIR205HG knockdown decreased in vitro migration and invasiveness of melanoma cells, and these effects were reversed by treatment with miR-299-3p inhibitor. MIR205HG-silenced melanoma cells showed increased miR-299-3p expression and lower levels of both VEGFA mRNA and protein. Tumor volumes were significantly smaller in nude mice xenografted with MIR205HG knockdown melanoma cells than the controls. These results demonstrate that MIR205HG supports melanoma growth via the miR-299-3p/VEGFA axis. This makes MIR205HG a potential therapeutic target for the treatment of melanoma.

Long non-coding RNA PICSAR knockdown inhibits the progression of cutaneous squamous cell carcinoma by regulating miR-125b/YAP1 axis.

AIMS: The purpose of this study was to explore the precise role and mechanism of p38 inhibiting cutaneous squamous cell carcinoma associated lincRNA (PICSAR) in CSCC. MATERIALS AND METHODS: The expression levels of PICSAR, microRNA-125b (miR-125b) and yes-associated protein1 (YAP1) were determined by quantitative real-time polymerase chain reaction (qRT-PCR). Cell proliferation, apoptosis and invasion were evaluated by Cell Counting Kit-8 (CCK-8) assay, flow cytometry, transwell assay, respectively. The interaction between miR-125b and PICSAR or YAP1 was predicted by bioinformatics software and confirmed by dual-luciferase reporter and RNA immunoprecipitation (RIP) assays. Western blot was employed to detect the protein expression of YAP1. The mice xenograft model was established to investigate the role of PICSAR in vivo. KEY FINDINGS: PICSAR was upregulated in CSCC tissues and cells. PICSAR knockdown inhibited cell proliferation and invasion and induced apoptosis in CSCC cells. Moreover, miR-125b could directly bind to PICSAR and its inhibition reversed the effect of PICSAR knockdown on proliferation, invasion and apoptosis in CSCC cells. In addition, YAP1 was a direct target of miR-125b and its overexpression attenuated the anti-cancer role of miR-125b in CSCC cells. Furthermore, YAP1 expression was positively regulated by PICSAR and negatively regulated by miR-125b. Besides, interference of PICSAR suppressed tumor growth by upregulating miR-125b and downregulating YAP1. SIGNIFICANCE: PICSAR knockdown suppressed cell proliferation and invasion and promoted apoptosis in CSCC cells by regulating miR-125b/YAP1 axis, providing new sights for treatment of CSCC.

[Epinephrine combined with acupuncture therapy can treat anaphylactic shock mice through nuclear factor-κB signaling pathway: a rat experimental result].

OBJECTIVE: To explore the therapeutic effect of epinephrine combined with acupuncture on anaphylactic shock and its mechanism. METHODS: Sixty male Kunming mice were randomly divided into normal saline (NS) group, anaphylactic shock model group, and integrated traditional Chinese and Western medicine treatment group with 20 mice in each group. The anaphylactic shock model was reproduced by egg albumin infusion: intraperitoneal injection of 0.25 mL egg albumin (0.01 mmol/L), repeated injection 1 week later, and intravenous injection of 0.5 mL egg albumin through caudal vein on the 3rd week to induce anaphylactic shock. The mice in the NS group were injected with NS. The mice in the treatment group were immediately subcutaneously injected with 0.2 µg of 0.1% epinephrine, and intraperitoneally injected with aminophylline 0.2 mg, combined with acupuncture at Shuigou, Neiguan and Hegu points. Number of died mice in each group were observed at 1, 6, and 12 hours after model reproduction. The mice were sacrificed at 12 hours, the blood was harvested, and the serum tryptase, immunoglobulin E (IgE), tumor necrosis factor-α (TNF-α), interleukins (IL-1 and IL-6) were determined by enzyme linked immunosorbent assay (ELISA). The lung tissues were harvested, and the protein expressions of p65, phosphorylation of p65 (p-p65), and phosphorylation of nuclear factor-κB inhibitor α (p-IκBα) were determined by Western Blot. RESULTS: No mice died in the NS control group at 12 hours. In the treatment group, the mortality at 12 hours was significantly lower than that in the model group (10% vs. 80%, P < 0.01). The levels of tryptase and IgE in the model group were significantly higher than those in the NS control group [tryptase (µg/L): 1.53±0.28 vs. 0.91±0.23, IgE (µg/L): 33.3±3.1 vs. 21.3±1.9, both P < 0.01], both levels in the treatment group were significantly lower than those in the model group [tryptase (µg/L): 1.31±0.26 vs. 1.53±0.28, IgE (µg/L): 25.6±2.2 vs. 33.3±3.1, both P < 0.05]. The levels of TNF-α, IL-1 and IL-6 in the model group were significantly higher than those in the NS control group [TNF-α (ng/L): 35.3±4.7 vs. 16.4±3.5, IL-1 (ng/L): 13.8±3.3 vs. 4.2±1.8, IL-6 (ng/L): 15.3±4.8 vs. 5.5±2.1, all P < 0.01]. The serum inflammatory factors of the treatment group were significantly higher than those of the model group [TNF-α (ng/L): 26.1±4.3 vs. 35.3±4.7, IL-1 (ng/L): 7.2±2.7 vs. 13.8±3.3, IL-6 (ng/L): 8.8±3.8 vs. 15.3±4.8, all P < 0.05]. It was shown by Western Blot results that there was no significant difference in p65 protein expression of the lung tissue among the three groups. In the NS control group, the expression of p-p65 protein in the nucleus of the lung tissue was extremely low but was significantly increased in the model group, and p-p65 protein in the treatment group was significantly decreased as compared with that in the model group. The expression tendency of p-IBα protein was consistent with that of p-p65. CONCLUSIONS: Epinephrine combined with acupuncture plays a therapeutic role in mice with anaphylactic shock by inhibiting the activation of NF-κB signaling pathway.

MiR-186 inhibits cell proliferation and invasion in human cutaneous malignant melanoma.

AIMS: MicroRNA-186 (miR-186) has been shown to be involved in various types of cancer. The purpose of this study was to investigate the expression level and functional role of miR-186 in human cutaneous malignant melanoma cells. SUBJECTS AND METHODS: Expression of miR-186 was analyzed in human cutaneous malignant melanoma (CMM) cell lines SK-MEL-1, G-361, A375 and A875, and human normal epidermal melanocytes cell line HEMn-LP by quantitative reverse transcription polymerase chain reaction (qRT-PCR). Additionally, the functional role of miR-186 on melanoma cells was investigated by transfection of miR-186 mimic followed by analyses of cell proliferation, apoptosis, and metastasis. RESULTS: We found that the expression levels of miR-186 were decreased in CMM cell lines compared with normal epidermal melanocytes cell line. Moreover, overexpression of miR-186 inhibited cells proliferation through abrogating the G1-S transition, and reduced cells migration and invasion. CONCLUSIONS: Our findings suggested that miR-186 exhibit an inhibitory effect on CMM cell proliferation, migration, and invasion; thus, may serve as a potential therapeutic target for human CMM intervention.

Mitochondrial genome of a melanoma disease model rat strain (Muridae; Rattus).

We sequenced the complete mitochondrial genome sequencing of an important melanoma disease model inbred rat strain for the first time. The total length of the mitogenome was 16,309 bp and coding 13 protein-coding genes, 2 ribosomal RNA genes, 22 transfer RNA genes.