Journal: Connective tissue research

Article Title: GREM1 inhibits osteogenic differentiation, senescence and BMP transcription of adipose-derived stem cells.

doi: 10.1080/03008207.2020.1736054

Figure Lengend Snippet: Figure 3. GREM1 overexpression inhibited ADSC senescence. (a) SA-β-gal staining showed fewer positive cells in GREM1 over- expressing ADSCs compared to vector control. Scale bar: 100 μm. (b) A quantitative analysis of SA-β-gal positive cells. (c) The relative telomerase activity was measured by the telomerase ELISA kit. (d, e) Real-time RT-PCR results showed that GREM1 overexpression decreased the expressions of P16 (d) and P53 (e). The control gene was GAPDH. The statistical significance was evaluated by Student’s t-test. The error bar was indicated by SD. (n = 3). *p ≤0.05.

Article Snippet: We measured the human telomerase reverse transcriptase levels in the cells by ELISA following the manufacturer’s protocol (Human Telomerase Reverse Transcriptase, TERT ELISA Kit, Cusabio, Wuhan, China).

Techniques: Over Expression, Staining, Expressing, Plasmid Preparation, Control, Activity Assay, Enzyme-linked Immunosorbent Assay, Quantitative RT-PCR

Journal: Connective tissue research

Article Title: GREM1 inhibits osteogenic differentiation, senescence and BMP transcription of adipose-derived stem cells.

doi: 10.1080/03008207.2020.1736054

Figure Lengend Snippet: Figure 4. GREM1 knockdown accelerated ADSC senescence. (a) SA-β-gal staining showed an increased number of positive cells in the ADSC-GREM1sh group compared to the control group. Scale bar: 100 μm. (b) The quantitative analysis of SA-β-gal positive cells. (c) Telomerase activity assay results showed that GREM1 knockdown significantly decreased ADSCs telomerase activity. (d, e) Real- time RT-PCR results showed that GREM1 knockdown upregulated the expressions of P16 (d) and P53 (e). The control gene was GAPDH. The statistical significance was evaluated by Student’s t-test. The error bar was indicated by SD. (n = 3). *p ≤0.05.

Article Snippet: We measured the human telomerase reverse transcriptase levels in the cells by ELISA following the manufacturer’s protocol (Human Telomerase Reverse Transcriptase, TERT ELISA Kit, Cusabio, Wuhan, China).

Techniques: Knockdown, Staining, Control, Telomerase Activity Assay, Activity Assay, Quantitative RT-PCR

Journal: bioRxiv

Article Title: The role of Cycloastragenol at the intersection of Nrf-2/ARE, telomerase, and proteasome activity

doi: 10.1101/2022.03.11.483898

Figure Lengend Snippet: (a) HEKn cells were transfected with either oligo duplex specific to Nrf-2, hTERT, or Control siRNA. After 48 h later, cells were harvested, and total protein levels of hTERT, Nrf-2, β1, and β5 were determined by IB. GAPDH was used as a loading control. ( b, c, d ) HEKn cells were transfected with either Nrf-2 or Control siRNA (each of them 2 nM). After 48 h later, cells were treated with indicated concentrations of CA for 24 h. ( b ) The total protein levels of Nrf-2, hTERT, β1, and β5 were determined by IB. ( c ) The telomerase enzyme activity was determined by using ELISA (n:3, *p ≤ 0.05, **p ≤ 0.001, ***p ≤ 0.005). ( d ) The proteasomal β1 and β5 subunit activities were determined by using fluorogenic substrates. The activity data were normalized to cellular total protein level and presented as a fold change compared to control cells treated with DMSO used as a solvent control (n:3, *p ≤ 0.05, **p. ≤ 0.001, ***p ≤ 0.005).

Article Snippet: When HEKn cells reached at confluency of 60-70%, cells were transfected with 2 and 15 nM siRNA against Nrf-2 (Origene, USA, SR321100) and hTERT (Origene, USA, SR322002), respectively.

Techniques: Transfection, Activity Assay, Enzyme-linked Immunosorbent Assay

Journal: bioRxiv

Article Title: The role of Cycloastragenol at the intersection of Nrf-2/ARE, telomerase, and proteasome activity

doi: 10.1101/2022.03.11.483898

Figure Lengend Snippet: HEKn cells were transfected with either oligo duplex specific to hTERT or Control siRNA. After 48 h later, cells were treated with CA for 24 h. ( a ) The total protein levels of Nrf-2, hTERT, β1, and β5 were determined by IB. ( b ) Before performing the nuclear Nrf-2 activity, cellular fractionation was performed. The Nrf-2 activity was determined by ELISA. The Nuclear activity data was normalized to nuclear protein level and presented as a fold change compared to control cells treated with DMSO used as solvent control. Error bars are presented as standard deviations (n = 3; *p ≤ 0.05, **p ≤ 0.001, ***p ≤ 0.005). ( c ) The proteasomal β1 and β5 subunit activities were evaluated via fluorogenic substrates. The data was normalized to cellular total protein level and presented as a fold change compared to control cells treated with DMSO used as solvent control. Error bars are presented as standard deviations (n = 3; *p ≤ 0.05, **p ≤ 0.001, ***p ≤ 0.005).

Article Snippet: When HEKn cells reached at confluency of 60-70%, cells were transfected with 2 and 15 nM siRNA against Nrf-2 (Origene, USA, SR321100) and hTERT (Origene, USA, SR322002), respectively.

Techniques: Transfection, Activity Assay, Cell Fractionation, Enzyme-linked Immunosorbent Assay

Journal: bioRxiv

Article Title: The role of Cycloastragenol at the intersection of Nrf-2/ARE, telomerase, and proteasome activity

doi: 10.1101/2022.03.11.483898

Figure Lengend Snippet: ( a, b, c ) HEKn cells were transfected with Nrf-2, hTERT, and Control siRNA. After 48 h later, cells were pre-treated with the desired concentration of CA for 8 h, then treated with 6-OHDA for 16 h. The cell viability was assessed by MTT reagent. The absorbance value of cells treated with DMSO as a solvent control for each experiment was considered 100%, and the viabilities of others were calculated. This assay was performed by triplicate samples. Student’s t-test was used to determine the significance of the differences Error bars are presented as standard deviations (n = 3; *p ≤ 0.05, **p ≤ 0.001, ***p ≤ 0.005).

Article Snippet: When HEKn cells reached at confluency of 60-70%, cells were transfected with 2 and 15 nM siRNA against Nrf-2 (Origene, USA, SR321100) and hTERT (Origene, USA, SR322002), respectively.

Techniques: Transfection, Concentration Assay

Journal: Journal of Experimental & Clinical Cancer Research : CR

Article Title: RIF1 promotes human epithelial ovarian cancer growth and progression via activating human telomerase reverse transcriptase expression

doi: 10.1186/s13046-018-0854-8

Figure Lengend Snippet: Clinical relevance of RIF1 and hTERT in EOC. a GSEA plot showing that RIF1 expression was positively correlated with TERT-activated gene signatures and TERT downstream PI3K/AKT signaling in the GEO data set (NCBI/GEO/GSE7463; n = 43). b Bioinformatics analysis based on TCGA database showed the mRNA expression levels of RIF1 positively correlated with hTERT expression level in ovarian cancer tissues. c and d Immunohistochemical staining suggesting that RIF1 expression correlated positively with hTERT expression in 75 clinical EOC specimens. Percentage of EOC specimens showing low or high RIF1 expression relative to the expression levels of hTERT. ** P < 0.01. e The relative expression of RIF1 and hTERT was used to perform the correlation analysis

Article Snippet: The shRNA sequence targeting human TERT cDNA was synthesized by GenePharma: hTERT KD: 5’-CCGAAGAAGCCACCUCUUUTT-3′, Lentiviral vectors pGLV3/H1/GFP were acquired from GenePharma.

Techniques: Expressing, Immunohistochemical staining, Staining