renal proximal tubular epithelial cell line hk2  (ATCC)

Journal: Journal of Translational Medicine

Article Title: Exploring the oncogenic potential of circSOD2 in clear cell renal cell carcinoma: a novel positive feedback loop

doi: 10.1186/s12967-024-05290-9

Figure Lengend Snippet: Authentication and characteristics of circSOD2 in RCC. A circSOD2 was verified by Sanger sequencing. The arrow indicates the back-splicing site of circ SOD2. CircSOD2 is derived by reverse splicing of exons 5, 6 and 7 of the SOD2 gene. B The expression of circSOD2 in RCC cells (769P, 786-O, A498, ACHN, and Caki-1) and human renal cortical proximal convoluted tubule epithelial cells (HK2). C qRT-PCR analysis of circSOD2 in the reverse transcription products using random primers or oligo dT primers. D The presence of circSOD2 was detected in 786-O and Caki-1 cells by qRT-PCR with convergent or divergent primers and validated by gel electrophoresis. GAPDH served as a control. E Comparison of circSOD2 expression in the nucleus and cytoplasm. F FISH analysis confirmed that circSOD2 was predominantly located in the cytoplasm. The nuclei were stained with DAPI. U6, 18S, and circSOD2 were labelled with Cy3. G Stability of circSOD2 and linear SOD2 was assessed by Actinomycin D treatment, followed by qRT-PCR at different time points. H Stability of circSOD2 and linear SOD2 was assessed by RNase treatment, followed by qRT-PCR. (NS, nonsignificant; * P < 0.05; ** P < 0.01; *** P < 0.001; **** P < 0.0001)

Article Snippet: Specifically, Human RCC cell lines 786-O, 769P, Caki-1, A498, ACHN and human renal proximal tubular epithelial cell line HK2 were purchased from American Type Culture Collection (ATCC).

Techniques: Sequencing, Derivative Assay, Expressing, Quantitative RT-PCR, Reverse Transcription, Nucleic Acid Electrophoresis, Control, Comparison, Staining

Journal: Journal of Translational Medicine

Article Title: Exploring the oncogenic potential of circSOD2 in clear cell renal cell carcinoma: a novel positive feedback loop

doi: 10.1186/s12967-024-05290-9

Figure Lengend Snippet: miR-532-3p is one of the targets of circSOD2 in ccRCC cells. A, B Heat map and volcano plot showed the difference of miRNA expression with or without circSOD2 knockdown. C The expression of candidate miRNAs were verified in circSOD2 overexpressed HK2 cells and negative control cells by qRT-PCR. D, E Relative expression detected by qRT-PCR and gel electrophoresis of circSOD2 in ACHN and 769P lysates after RNA pull down with circSDHC specific probe or oligo probe. Expression levels were normalized to oligo probe. GAPDH was used as negative control. F Relative levels of candidate miRNAs were detected by qRT-PCR after being pull down by circSOD2 probe or oligo probe. G Relative levels of circSOD2 in ACHN and 769P lysates captured by biotinylated wild-type miRNA-532-3p or mutant miRNA-532-3p. H Schematic diagram model showed the putative binding sites of miRNA-532-3p associated with circSOD2. I Schematic diagram of circSOD2 wild-type (wt) and mutant (mut) luciferase reporter vectors. J Luciferase reporter assay in HEK293T with vector, circSOD2 wild-type sequence and circSOD2 mutant sequence transfected with miR-532-3p mimics. Vector group was utilized as normalization control. K Cellular localization of circSOD2 (Cy3) and miRNA-532-3p (FAM) detected by FISH. Nuclear was label with DAPI dye. (NS, nonsignificant; * P < 0.05; ** P < 0.01; *** P < 0.001; **** P < 0.0001)

Article Snippet: Specifically, Human RCC cell lines 786-O, 769P, Caki-1, A498, ACHN and human renal proximal tubular epithelial cell line HK2 were purchased from American Type Culture Collection (ATCC).

Techniques: Expressing, Knockdown, Negative Control, Quantitative RT-PCR, Nucleic Acid Electrophoresis, Mutagenesis, Binding Assay, Luciferase, Reporter Assay, Plasmid Preparation, Sequencing, Transfection, Control

Journal: Journal of Translational Medicine

Article Title: Exploring the oncogenic potential of circSOD2 in clear cell renal cell carcinoma: a novel positive feedback loop

doi: 10.1186/s12967-024-05290-9

Figure Lengend Snippet: miR-532-3p plays an inhibitory role in ccRCC by targeting PAX5. A The expression of miR-532-3p in RCC cells (769P, 786-O, A498, ACHN, Caki-1) and human renal cortical proximal convoluted tubule epithelial cells (HK2). B CCK8 assays demonstrated that cell proliferation was inhibited after transfected with miR-532-3p mimics. C Transwell assays demonstrated that cell migratory and invasive capacity was inhibited after transfected with miR-532-3p mimics. D FACS assays demonstrated that cell apoptosis was enhanced after transfected with miR-532-3p mimics. E Luciferase reporter assay in HEK293T cells co-transfected mimics miR- miR-532-3p or mimics NC and candidate genes of Luc-wild-type or Luc-mutant. F Western blot analysis indicated that miR-532-3p could down-regulate PAX5 expression in RCC cells. G Binding site of miR-532-3p and PAX5. (NS, nonsignificant; *P < 0.05; **P < 0.01; ***P < 0.001; ****P < 0.0001)

Article Snippet: Specifically, Human RCC cell lines 786-O, 769P, Caki-1, A498, ACHN and human renal proximal tubular epithelial cell line HK2 were purchased from American Type Culture Collection (ATCC).

Techniques: Expressing, Transfection, Luciferase, Reporter Assay, Mutagenesis, Western Blot, Binding Assay

Journal: Kidney & blood pressure research

Article Title: MicroRNA-155-5p Targets NR3C2 to Promote Malignant Progression of Clear Cell Renal Cell Carcinoma.

doi: 10.1159/000521745

Figure Lengend Snippet: Fig. 1. MicroRNA-155-5p is highly expressed in ccRCC. a Expression level of microRNA-155-5p in cancer tissue and para-cancerous tissue of 42 patients with ccRCC detected by qRT-PCR. b Detection of microRNA-155-5p expression in HK2 and ccRCC cell lines. *denotes p < 0.05.

Article Snippet: Cell Culture and Transfection Human proximal renal tubular epithelial cell line HK2 (ATCC® CRL-2190) and human ccRCC cell lines 786-O (ATCC® CRL1932), A498 (ATCC® HTB-44), ACHN (ATCC® HTB-1611), and Caki-1 (ATCC® HTB-46) were purchased from ATCC.

Techniques: Expressing, Quantitative RT-PCR

Journal: Kidney & blood pressure research

Article Title: MicroRNA-155-5p Targets NR3C2 to Promote Malignant Progression of Clear Cell Renal Cell Carcinoma.

doi: 10.1159/000521745

Figure Lengend Snippet: Fig. 3. NR3C2 is the target gene of microR- NA-155-5p. A Venn diagram of target genes of microRNA-155-5p. B Volcano map of differential genes in the ccRCC mRNA data set in TCGA database between the normal group and the tumor group. Red and green dots represent upregulated and downregulated mRNA expression in ccRCC tissue, respectively. C Venn dia- gram of downregulated genes in ccRCC and predicted target genes of microRNA- 155-5p. D Pearson correlation heat map of microRNA-155-5p and its target genes. E Effect of NR3C2 expression on prognosis of ccRCC patients. F NR3C2 expression in TCGA database is downregulated in the tu- mor group. G Expression of NR3C2 mRNA and protein in HK2 and ccRCC cell lines. H Binding of microRNA-155-5p and NR3C2 detected by the dual-luciferase as- say. I, J Effects of silencing or overexpress- ing microRNA-155-5p on NR3C2 expres- sion in ccRCC cells. *denotes p < 0.05. NC, negative control.

Article Snippet: Cell Culture and Transfection Human proximal renal tubular epithelial cell line HK2 (ATCC® CRL-2190) and human ccRCC cell lines 786-O (ATCC® CRL1932), A498 (ATCC® HTB-44), ACHN (ATCC® HTB-1611), and Caki-1 (ATCC® HTB-46) were purchased from ATCC.

Techniques: Expressing, Binding Assay, Luciferase, Negative Control