Journal: International journal of molecular sciences

Article Title: Porcine Kidney Organoids Derived from Naïve-like Embryonic Stem Cells.

doi: 10.3390/ijms25010682

Figure Lengend Snippet: Figure 5. Gene expression in 3D porcine kidney organoids. (A) Expression of renal progenitor cell markers in the organoids on day 21 and fetal porcine kidney on day 45. Renal progenitor cells markers (EYA1, SIX1) were measured by immunofluorescence test (EYA1 was localized in the cytoplasm, SIX1 was localized in the nucleus). (B) Expression of mature renal cell markers in the organoids on day 21 and fetal porcine kidney on day 45. Mature nephron components markers (PAX2, E-CAD, PODO) were measured by immunofluorescence test and immunohistochemistry test (the circular dotted lines are tubule-like structures; PAX2 was localized in the nucleus; and PODO and ECAD were localized to the cell membrane). (C) Whole-mount co-staining of the organoids for WT1 and CD31 on day 21 (WT1 was localized in the nucleus and CD31 was localized to the cell membrane) (scale bar: 50 µm).

Article Snippet: The primary antibodies included the following: EYA1 (Abcam), SIX1 (Proteintech), PAX2 (Santa Cruz Biotechnology), WT1 (Abcam), E-CAD (Abcam), PODO (Absin), CD31 (Santa Cruz Biotechnology), AQP1 (Proteintech), GLUT1 (proteintech), and KI67 (Abcam); the secondary antibodies included goat anti-rabbit IgG Alexa Fluor 546 (ABclonal), goat anti-rabbit IgG Alexa Fluor 488 (Thermo Fisher Scientific), goat anti-mouse IgG Alexa Fluor 546 (Thermo Fisher Scientific), goat anti-mouse IgG Alexa Fluor 488 (Thermo Fisher Scientific), goat anti-rabbit IgG H&L(HRP) (Abcam), and goat anti-mouse IgG(H + L) (KeyGEN BioTECH, Nanjing, China).

Techniques: Gene Expression, Expressing, Immunohistochemistry, Membrane, Staining

Journal: Frontiers in Oncology

Article Title: circ0093740 Promotes Tumor Growth and Metastasis by Sponging miR-136/145 and Upregulating DNMT3A in Wilms Tumor

doi: 10.3389/fonc.2021.647352

Figure Lengend Snippet: circ0093740 is upregulated in Wilms tumor compared to normal kidney tissue. (A) The heat map depicted the differentially expressed circRNAs of three paired Wilms tumor and normal kidney tissue. (B) The relative expression level of top five upregulated circRNA between Wilms tumor tissues and normal kidney tissues. (C) The relative expression level of circ0093740 in normal HEK293T cell line and Wilms tumor cell lines. (D) The circular structure of circ0093740 was tested RNase R assay in SKNEP1 Wilms tumor cell line. (E) Circular transcripts of circ0093740 were more stable than its linear PDCH15 mRNA transcripts determined by actinomycin D treated assay in SKNEP1 Wilms tumor cell line. **p < 0.01.

Article Snippet: SKNEP1 Wilms tumor cells were exposed to 5 ug/ml actinomycin D (MCE) to degrade the linear mRNA transcription at 0-, 8-, 16-, and 24-hour time point.

Techniques: Wilms Tumor Assay, Expressing

Journal: Frontiers in Oncology

Article Title: circ0093740 Promotes Tumor Growth and Metastasis by Sponging miR-136/145 and Upregulating DNMT3A in Wilms Tumor

doi: 10.3389/fonc.2021.647352

Figure Lengend Snippet: Suppression of circ0093740 inhibits the proliferation of Wilms tumor cells. (A) siRNA knockdown of circ0093740 was validated in SKNEP1 and G401 Wilms tumor cell line. (B) CCK-8 assays were conducted to evaluate cell proliferation in SKNEP1 Wilms tumor cell line. (C) Mouse xenograft models of SKNEP1 Wilms tumor cell line was established. Tumor volume was estimated in every four days. (D) The images of Ki-67 IHC expression are presented. **p < 0.01.

Article Snippet: SKNEP1 Wilms tumor cells were exposed to 5 ug/ml actinomycin D (MCE) to degrade the linear mRNA transcription at 0-, 8-, 16-, and 24-hour time point.

Techniques: Wilms Tumor Assay, Knockdown, CCK-8 Assay, Expressing

Journal: Frontiers in Oncology

Article Title: circ0093740 Promotes Tumor Growth and Metastasis by Sponging miR-136/145 and Upregulating DNMT3A in Wilms Tumor

doi: 10.3389/fonc.2021.647352

Figure Lengend Snippet: Suppression of circ0093740 inhibits the metastasis of Wilms tumor cells. (A) Wound healing assays were conducted in SKNEP1 and G401 Wilms tumor cell line. (B) Transwell assay was performed to investigate the migration ability of SKNEP1 and G401 Wilms tumor cell line. (C) The number of lung metastases was counted and recorded. HE-stained tumor sections of lung metastases were presented. **p < 0.01.

Article Snippet: SKNEP1 Wilms tumor cells were exposed to 5 ug/ml actinomycin D (MCE) to degrade the linear mRNA transcription at 0-, 8-, 16-, and 24-hour time point.

Techniques: Wilms Tumor Assay, Transwell Assay, Migration, Staining

Journal: Frontiers in Oncology

Article Title: circ0093740 Promotes Tumor Growth and Metastasis by Sponging miR-136/145 and Upregulating DNMT3A in Wilms Tumor

doi: 10.3389/fonc.2021.647352

Figure Lengend Snippet: circ0093740 serves as a sponge of miR-136/145 in Wilms tumor. (A) 18S, ACTIN, circ0093740 and PCDH15 expression in nuclear and cytoplasmic fractions were analyzed by RT-qPCR. (B) miRNA sequencing on three pairs of circ0093740 overexpression and vector samples was conducted in SKNEP1 cell line. (C) Predicted binding sites of miR-136/145 within the circ0093740 sequence. (D) miR-136 and miR-145 expression in Wilms tumor cell lines compared to HEK293T. (E) Luciferase reporter assay was conducted in SKNEP1 and G401 Wilms tumor cell. miR-136 and miR-145 mimics were cotransfected with circ0093740 wild type/mutant luciferase reporter. (F) MS2-based RIP assay transfected with MS2bs-circ0093740 vector, MS2bs-circ0093740-mt vector or Rluc control vector. **p < 0.01.

Article Snippet: SKNEP1 Wilms tumor cells were exposed to 5 ug/ml actinomycin D (MCE) to degrade the linear mRNA transcription at 0-, 8-, 16-, and 24-hour time point.

Techniques: Wilms Tumor Assay, Expressing, Quantitative RT-PCR, Sequencing, Over Expression, Plasmid Preparation, Binding Assay, Luciferase, Reporter Assay, Mutagenesis, Transfection, Control

Journal: Frontiers in Oncology

Article Title: circ0093740 Promotes Tumor Growth and Metastasis by Sponging miR-136/145 and Upregulating DNMT3A in Wilms Tumor

doi: 10.3389/fonc.2021.647352

Figure Lengend Snippet: circ0093740 facilitates Wilms tumor progression through circ0093740-miR-136/145-DNMT3A axis. (A) According to TargetScan online website, DNMT3A was the putative downstream target of both miR-136 and miR-145. (B) DNMT3A expression in Wilms tumor cell lines. (C) Expression of DNMT3A transcript was decreased after transfection with miR-136 and miR-145 mimics, detected by qPCR analysis. (D) Luciferase reporter assay of SKNEP1 and G401 Wilms tumor cells co-transfected with miR-136/145 mimics and the 3’-UTR of DNMT3A wild type/mutant luciferase reporter. (E) Enrichment of circ0093740, DNMT3A and miR-136/145 on AGO2 assessed by RIP assay. (F) Enrichment of DNMT3A to AGO2 was increased after silence of circ0093740. (G) Knockdown of circ0093740 contributed to the reduction of DNMT3A protein expression in SKNEP1 and G401 Wilms tumor cell. (H) The molecular mechanism of Circ0093740 Facilitates Wilms Tumor Progression Through circ0093740-miR-136/145-DNMT3A Axis diagram. **p < 0.01.

Article Snippet: SKNEP1 Wilms tumor cells were exposed to 5 ug/ml actinomycin D (MCE) to degrade the linear mRNA transcription at 0-, 8-, 16-, and 24-hour time point.

Techniques: Wilms Tumor Assay, Expressing, Transfection, Luciferase, Reporter Assay, Mutagenesis, Knockdown