Journal: International Journal of Medical Sciences

Article Title: The Prognostic Value of Cytokeratin and Sal-Like Protein 4 Expression in Hepatocellular Carcinoma and Intra-Hepatic Cholangiocarcinoma in Taiwan

doi: 10.7150/ijms.28440

Figure Lengend Snippet: Antibodies used in this study

Article Snippet: CK18 , Mouse monoclonal , TA500015 , IgG1 , OriGene , 1:200 , ER2 20 min.

Techniques:

Journal: Frontiers in Cell and Developmental Biology

Article Title: Honokiol Inhibits Melanoma Growth by Targeting Keratin 18 in vitro and in vivo

doi: 10.3389/fcell.2020.603472

Figure Lengend Snippet: Honokiol directly binds to KRT18, which is highly expressed in some melanoma cells and tissues. Honokiol directly binds to KRT18 (A) in melanoma cell lysate, (B) exogenously and (C) recombinant protein. Proteins were pulled down by Sepharose 4B-honokiol beads or with Sepharose 4B beads and then analyzed by western blotting. (D) Representative images of Immunohistochemical analysis of the melanoma tissue chip (left) and bar graphs showing the number of KRT18 stained cells by IHC staining through image scope (right). KRT18 protein level between normal skin and melanoma tissues (right, above) as well as between male and female melanoma tissues (right, down). (E) KRT18 expression between primary and metastatic melanoma tissues from TCGA database (SKCM: Skin Cutaneous Melanoma). (F) KRT18 protein level between melanoma cell lines and normal skin cell line. The asterisks indicate a significant difference (* p < 0.05 and ** p < 0.01 compared to control.

Article Snippet: Human KRT18 protein was purchased from OriGene (Maryland, United States).

Techniques: Recombinant, Western Blot, Immunohistochemical staining, Staining, Immunohistochemistry, Expressing, Control

Journal: Frontiers in Cell and Developmental Biology

Article Title: Honokiol Inhibits Melanoma Growth by Targeting Keratin 18 in vitro and in vivo

doi: 10.3389/fcell.2020.603472

Figure Lengend Snippet: KRT18 promotes melanoma cell growth. (A) Western blot analysis showing KRT18 expression in SK-MEL-5 and SK-MEL-28 cells stably transfected with sh-NT or sh-KRT18. (B) Western blot analysis showing KRT18 expression in MM200 cells overexpressed KRT18. (C) Effect of KRT18 knockdown on anchorage-independent growth of SK-MEL-5 and SK-MEL-28 melanoma cells was evaluated by soft agar assay. Clone count (above); Representative images (below). (D) Effect of KRT18 knockdown on the proliferation of SK-MEL-5 and SK-MEL-28 melanoma cells was evaluated at 0, 24, 48, 72, or 96 h by MTT assay. (E) Effect of KRT18 overexpression on anchorage-independent growth of MM200 melanoma cell was evaluated by soft agar assay. (F) Effect of KRT18 overexpression on the proliferation of MM200 melanoma cells was evaluated at 0, 24, 48, 72, or 96 h by MTT assay. The asterisk (** p < 0.01 and *** p < 0.001) indicates a significant difference compared to the control group.

Article Snippet: Human KRT18 protein was purchased from OriGene (Maryland, United States).

Techniques: Western Blot, Expressing, Stable Transfection, Transfection, Knockdown, Soft Agar Assay, MTT Assay, Over Expression, Control

Journal: Frontiers in Cell and Developmental Biology

Article Title: Honokiol Inhibits Melanoma Growth by Targeting Keratin 18 in vitro and in vivo

doi: 10.3389/fcell.2020.603472

Figure Lengend Snippet: Inhibitory effect of honokiol on melanoma cell growth is dependent upon KRT18 protein level. (A) Western blot analysis showing KRT18 expression in SK-MEL-5 and SK-MEL-28 cells stably transfected with sh-NT or sh-KRT18. (B) Effect of KRT18 knockdown on anchorage-independent growth of honokiol treated SK-MEL-5 and SK-MEL-28 melanoma cells was evaluated by soft agar assay. Clone count (above); Representative images (below). (C) KRT18 protein level after KRT18 overexpression in MM200 melanoma cell. (D) Effect of KRT18 overexpression on anchorage-independent growth of honokiol treated MM200 melanoma cells was evaluated by soft agar assay. Clone count (left); Representative images (right). The asterisks (* p < 0.05, *** p < 0.001) indicate a significant difference in treated cells compared to the control cells.

Article Snippet: Human KRT18 protein was purchased from OriGene (Maryland, United States).

Techniques: Western Blot, Expressing, Stable Transfection, Transfection, Knockdown, Soft Agar Assay, Over Expression, Control

Journal: Frontiers in Cell and Developmental Biology

Article Title: Honokiol Inhibits Melanoma Growth by Targeting Keratin 18 in vitro and in vivo

doi: 10.3389/fcell.2020.603472

Figure Lengend Snippet: Honokiol down-regulates KRT18 protein levels by inducing its ubiquitination in melanoma cells. (A) Effect of honokiol on KRT18 protein expression in SK-MEL-5 or SK-MEL-28 melanoma cells. Cells were treated with 0, 30, or 50 μM of honokiol for 48 h and KRT18 protein level was evaluated by western blotting. (B,C) KRT18 mRNA level after honokiol treatment for 48 h was determined by RT-qPCR and agarose gel electrophoresis. (D) Immunoprecipitation of cell lysates from HEK293T cells transfected with exogenous flag-tagged KRT18 and HA-tagged ubiquitin extracts of exogenous flag was followed by immunodetection of exogenous ubiquitin and flag-KRT18. IP Flag: immunoprecipitation with a flag antibody.

Article Snippet: Human KRT18 protein was purchased from OriGene (Maryland, United States).

Techniques: Ubiquitin Proteomics, Expressing, Western Blot, Quantitative RT-PCR, Agarose Gel Electrophoresis, Immunoprecipitation, Transfection, Immunodetection

Journal: Frontiers in Cell and Developmental Biology

Article Title: Honokiol Inhibits Melanoma Growth by Targeting Keratin 18 in vitro and in vivo

doi: 10.3389/fcell.2020.603472

Figure Lengend Snippet: Honokiol inhibits melanoma cell-derived tumor growth in vivo . SK-MEL-5 or SK-MEL-28 melanoma cell-derived xenograft mice models were established and treated with varying concentrations of honokiol. (A) Morphology of tumors after honokiol treatment. (B) Effect of honokiol treatment on melanoma growth. Tumor size was measured twice a week and volume was calculated, as indicated in materials and methods. (C) KRT18 protein level in random three tissues of vehicle and 50 mg/kg honokiol groups, as detected by western blotting (left) followed by densitometric quantification (right). (D) Expression of Ki-67 in vehicle group or 50 mg/kg honokiol treatment group tumors, as assessed by IHC staining. Quantitation of Ki-67 (left); Representative images (right). Bar graphs quantifying the number of Ki-67 stained cells by IHC staining through image scope. The asterisks indicate a significant difference (* p < 0.05,** p < 0.01, *** p < 0.001) between the vehicle and honokiol treated groups.

Article Snippet: Human KRT18 protein was purchased from OriGene (Maryland, United States).

Techniques: Derivative Assay, In Vivo, Western Blot, Expressing, Immunohistochemistry, Quantitation Assay, Staining