Journal: BioMed Research International

Article Title: LINC00518 Promotes Cell Malignant Behaviors via Influencing EIF4A3-Mediated mRNA Stability of MITF in Melanoma

doi: 10.1155/2022/3546795

Figure Lengend Snippet: LINC00518 is upregulated in melanoma, and LINC00518 inhibits multiple malignant behaviors of melanoma cells. (a) The level of LINC00518 in melanoma tumor tissues and noncancerous tissues was projected on the GEPIA database. (b) The GEPIA database demonstrated the overall survival of patients in the high/low LINC00518 group. (c) The level of LINC00518 in melanoma cell lines (B16, A2058, and A375) and the human epidermal melanocytes (HEMn) was examined by RT-qPCR. (d) RT-qPCR assays measured knockdown efficiency of sh-LINC00518#1/2. (e) CCK-8 assays were done to examine proliferative ability of cells upon LINC00518 knockdown. (f) Number of formed colonies was detected by colony formation assays after LINC00518 deficiency. (g) Transwell assays evaluated the migration of melanoma cells upon LINC00518 depletion. (h) The influences of LINC00518 knockdown on the expression of key proteins related to invasion, migration, and EMT were assessed in western blot assays. ∗ P < 0.05; ∗∗ P < 0.01.

Article Snippet: Human melanoma cell lines (B16, A2058, and A375) and human epidermal melanocytes (HEMn) were provided by the American Type Culture Collection (ATCC; VA, USA).

Techniques: Quantitative RT-PCR, Knockdown, CCK-8 Assay, Migration, Expressing, Western Blot

Journal: BioMed Research International

Article Title: LINC00518 Promotes Cell Malignant Behaviors via Influencing EIF4A3-Mediated mRNA Stability of MITF in Melanoma

doi: 10.1155/2022/3546795

Figure Lengend Snippet: MITF has a significantly high expression in melanoma tissues and cells, and its expression positively correlates with LINC00518 expression. (a) MITF expression in melanoma tissues and adjacent noncancer tissues was projected on the GEPIA database. (b) The relationship between patients in the high MITF group and their overall survival was obtained from the GEPIA database. (c) The GEPIA database provided the correlation between LINC00518 expression and MITF expression. (d) The expression of MITF in melanoma cell lines (B16, A2058, and A375) and the human epidermal melanocytes (HEMn) was quantified by RT-qPCR and western blot. (e) RT-qPCR analysis measured the effect of LINC00518 knockdown on MITF mRNA expression. (f) Western blot assay examined the impact of LINC00518 knockdown on MITF protein level. ∗ P < 0.05; ∗∗ P < 0.01.

Article Snippet: Human melanoma cell lines (B16, A2058, and A375) and human epidermal melanocytes (HEMn) were provided by the American Type Culture Collection (ATCC; VA, USA).

Techniques: Expressing, Quantitative RT-PCR, Western Blot, Knockdown

Journal: PLoS ONE

Article Title: The Chick Embryo as an Experimental System for Melanoma Cell Invasion

doi: 10.1371/journal.pone.0053970

Figure Lengend Snippet: (A) Schematic drawing depicting ventral and medial neural crest migration pathways. n.c. neural crest; n.t. neural tube; s.t. sympathetic trunk. (B) Chick embryo 24 h after transplantation of SKMel28 melanoma cells into the neural tube. Melanoma cells (visualized by HMB45 immunoreactivity) spontaneously resuming neural crest migration have a stretched, mesenchymal-like morphology (arrows). (C) At the site of destination along the ventral migration pathway (para-aortic sympathetic ganglia) melanoma cells undergo apoptosis, visualized by TUNEL staining. (D,E) Chick embryo 24 h after transplantation of benign primary human melanocytes into the neural tube. Melanocytes (showing a compact, epithelial-like morphology) are encountered only in the lumen of the neural tube and, in part, integrated into the roof plate with no neural crest migration. (F) Melan A immunoreactivity confirms the melanocytic origin of the cells. (G) Schematic drawing of chick embryo 72 h after transplantation of B16-F1 melanoma cells into the optic cup. (H) Histological correlate of schematic drawing. Already in H&E staining the transplanted, invasively migrating melanoma cells are visible (arrows). (I) Single melanoma cells (identified by HMB45 immunoreactivity) form a tumor, and single melanoma cells invade the choroid of the optic cup (arrows). (J) Chick embryo 96 h after transplantation of human metastatic melanoma cells into the brain vesicle at the hindbrain (rhombencephalon). The cells form a large tumor in the dorsal neuroepithelium with (K) single HMB45 positive cells infiltrating the surrounding brain tissues. (L) MIB1 immunohistochemistry (proliferation marker not cross-reacting with chick cells) identifies melanoma cells during haematogenous spreading in blood vessels among host erythrocytes and lymphocytes, and in the surrounding neural tissue.

Article Snippet: For each series of transplantation, one of the following cells were used as aggregates or cell suspensions: Mouse B16-F1 metastatic melanoma cells (gifted from ); human SKMel28 metastatic melanoma cells (purchased as part of the NCI60 panel of cancer cells from the NCI); human 451LU metastatic melanoma cells (gifted from Meenhard Herlyn, Wistar Institute, Philadelphia, USA ), or human melanocytes (human epidermal melanocytes neonatal (HEMn), CellSystems, Troisdorf, Germany, cultivated in Lifeline's DermaLife M medium (CellSystems)).

Techniques: Migration, Transplantation Assay, TUNEL Assay, Staining, Immunohistochemistry, Marker

Journal: PLoS ONE

Article Title: The Chick Embryo as an Experimental System for Melanoma Cell Invasion

doi: 10.1371/journal.pone.0053970

Figure Lengend Snippet: Untreated, BMP-2 or nodal pre-treated melanocytes were injected into the optic cup of the chick embryo (stage 20 HH). After 72 h of further incubation, the embryos were analyzed for tumor growth and invasion. Untreated melanocytes formed loosely aggregated tumors adjacent to the hyaloid vessels (left image in upper row), in the developing vitreous body and behind the lens (right image in upper row) without invasion. The BMP-2 and nodal groups formed tumors in similar locations. In the BMP-2 group single melanocytes invaded the lens epithelium (insert in left image in middle row), the retina, the hyaloid vessels, and the choroid (right image in middle row; arrows pointing at melanocytes). In the nodal group single melanocytes invaded the choroid (lower row, arrows in right image) and the hyaloid vessels.

Article Snippet: For each series of transplantation, one of the following cells were used as aggregates or cell suspensions: Mouse B16-F1 metastatic melanoma cells (gifted from ); human SKMel28 metastatic melanoma cells (purchased as part of the NCI60 panel of cancer cells from the NCI); human 451LU metastatic melanoma cells (gifted from Meenhard Herlyn, Wistar Institute, Philadelphia, USA ), or human melanocytes (human epidermal melanocytes neonatal (HEMn), CellSystems, Troisdorf, Germany, cultivated in Lifeline's DermaLife M medium (CellSystems)).

Techniques: Injection, Incubation

Journal: Molecular and Cellular Biology

Article Title: MicroRNA 211 Functions as a Metabolic Switch in Human Melanoma Cells

doi: 10.1128/MCB.00762-15

Figure Lengend Snippet: Expression and cellular phenotypes of PDK4 in human melanoma cells with and without miR-211. (A) Putative miR-211 target genes. (B) Western blot analysis of PDK4 expression in HEM-l primary melanocytes as well as A375/VO and A375/211 cells (using GAPDH as a loading control). (C) qPCR analysis of PDK4 expression in HEM-l primary melanocytes, A375/VO, and A375/211 cells. All analyses were performed in triplicate. (D and E) Effects of miR-211 and PDK4 on cell invasion. Results in panel D are normalized to A375/VO + Scr siRNA. The assay was performed in triplicate. (F) Effects of miR-211 and PDK4 on melanoma cell motility. Wound-healing assay of A375/VO cells with negative-control siRNA versus PDK4 siRNA and A375/211 with negative-control plasmid versus PDK4 expression plasmid. Representative images of wound-healing assays for cells at a magnification of ×20 cover a 24-hour period. Assays were performed in triplicate.

Article Snippet: Cell lines examined in this study included the melanoma cell lines A375 (melanoma stage 4; American Type Culture Collection) and WM1552C (melanoma stage 3; ATCC CRL-2808), as well as the human epidermal melanocyte cell line HEM-l (catalog no. 2200; ScienCell).

Techniques: Expressing, Western Blot, Wound Healing Assay, Negative Control, Plasmid Preparation

Journal: Redox Biology

Article Title: Photoprotection by dietary phenolics against melanogenesis induced by UVA through Nrf2-dependent antioxidant responses

doi: 10.1016/j.redox.2015.12.006

Figure Lengend Snippet: Effects of Nrf2 knockdown on melanogenesis in HEMn and B16F10 cells in response to UVA irradiation. (A) HEMn and B16F10 cells were transfected with 5 nM Nrf2-siRNA (siNrf2) or non-silencing siRNA control (siCtrl) for 48 h. mRNA levels of Nrf2 and its target antioxidants (GCLC, GCLM, GST and NQO1) of HEMn and B16F10 cells transfected with siNrf2 were evaluated by real-time RT-PCR. * P <0.05; ** P <0.01; *** P <0.001 versus siCtrl-transfected cells. (B) Melanin content and (C) tyrosinase activity were measured in HEMn and B16F10 cells transfected with siNrf2 or siCtrl at 1 h following UVA (8 J/cm 2 ) irradiation. (D) Tyrosinase protein expression was measured in HEMn and B16F10 cells transfected with siNrf2 or siCtrl at 24 h post-irradiation. Data was expressed as mean±SD. The statistical significance of differences was evaluated by one-way ANOVA followed by Dunnett’s test. # P <0.05; ## P <0.01 versus siCtrl-transfected cells without UVA irradiation. ** P <0.01; *** P <0.001 versus siNrf2-transfected cells irradiated with UVA.

Article Snippet: Primary human epidermal melanocytes (HEMn) (Lonza, Basel, Switzerland) were grown in Medium 254 (#M-254-500) supplemented with human melanocyte growth supplement (HMGS) according to the manufacturer's instructions.

Techniques: Knockdown, Irradiation, Transfection, Control, Quantitative RT-PCR, Activity Assay, Expressing

Journal: International Journal of Molecular Sciences

Article Title: The Effects of Vitamin D on the Expression of IL-33 and Its Receptor ST2 in Skin Cells; Potential Implication for Psoriasis

doi: 10.3390/ijms222312907

Figure Lengend Snippet: Vitamin D modulates the expression of IL-33 , ST2 , TNF , IL-1B , IFNG and IL-17A in skin cell lines. Keratinocytes (HPEKp), melanocytes (HEMas), fibroblasts (HDF) and basal cell carcinoma cells (A431) were stimulated with 1,25(OH)2D3 (100 nM) for 4, 8 and 24 h. The relative IL-33 ( A ), ST2 ( B ), TNF ( C ), IL-1B ( D ), IFNG , ( E ) and IL-17A ( F ) mRNA levels were analyzed by real-time PCR. * p < 0.05, ** p < 0.01, *** p < 0.001.

Article Snippet: Pooled juvenile Human Epidermal Keratinocyte Progenitors (HPEKp) and Single donor Adult Epidermal Melanocytes (HEMas) were acquired from CELLnTEC (Bern, Switzerland).

Techniques: Expressing, Real-time Polymerase Chain Reaction