Journal: Annals of Translational Medicine

Article Title: Long non-coding RNA LSINCT5 inactivates Wnt/β-catenin pathway to regulate MCF-7 cell proliferation and motility through targeting the miR-30a

doi: 10.21037/atm-20-7253

Figure Lengend Snippet: The primer sequence of genes

Article Snippet: Cell transfection The short hairpin RNA (shRNA) specifically targeting LSINCT5 (called sh-LSINCT5) was synthesized by GenePharma (Shanghai, China), and nonspecific shRNA was used as negative control (sh-NC).

Techniques: Sequencing

Journal: Annals of Translational Medicine

Article Title: Long non-coding RNA LSINCT5 inactivates Wnt/β-catenin pathway to regulate MCF-7 cell proliferation and motility through targeting the miR-30a

doi: 10.21037/atm-20-7253

Figure Lengend Snippet: LSINCT5 is highly expressed in BC. (A) LSINCT5 level in BC tissues and corresponding non-cancerous tissues. (B) Survival rate in BC patients based on LSINCT5 level. (C) LSINCT5 level in BC cell lines and normal breast epithelial cell line. *, P<0.05, **, P<0.01 vs. matched group. BC, breast cancer.

Article Snippet: Cell transfection The short hairpin RNA (shRNA) specifically targeting LSINCT5 (called sh-LSINCT5) was synthesized by GenePharma (Shanghai, China), and nonspecific shRNA was used as negative control (sh-NC).

Techniques:

Journal: Annals of Translational Medicine

Article Title: Long non-coding RNA LSINCT5 inactivates Wnt/β-catenin pathway to regulate MCF-7 cell proliferation and motility through targeting the miR-30a

doi: 10.21037/atm-20-7253

Figure Lengend Snippet: Relationship of LSINCT5 expression with clinicopathologic characteristics of BC

Article Snippet: Cell transfection The short hairpin RNA (shRNA) specifically targeting LSINCT5 (called sh-LSINCT5) was synthesized by GenePharma (Shanghai, China), and nonspecific shRNA was used as negative control (sh-NC).

Techniques: Expressing

Journal: Annals of Translational Medicine

Article Title: Long non-coding RNA LSINCT5 inactivates Wnt/β-catenin pathway to regulate MCF-7 cell proliferation and motility through targeting the miR-30a

doi: 10.21037/atm-20-7253

Figure Lengend Snippet: MiR-30a targeted LSINCT5 and negatively regulated LSINCT5 in MCF-7 cells. (A) The target site of miR-30a on LSINCT5 3'UTR (wt/mut). (B) LSINCT5 level in sh-LSINCT5 or NC transfected MCF-7 cells (**, P<0.01 vs. control). (C) qRT-PCR monitored miR-30a. MiR-30a level in sh-LSINCT5 or miR-30a inhibitors and co-transfected MCF-7 cells (**, P<0.01 vs. control, ##, P<0.01 vs. miR-30a inhibitor group). (D) miR-30a level in ApcDNA-LSINCT5 or miR-30a mimics and co-transfected MCF-7 cells (**, P<0.01 vs. control, ##, P<0.01 vs. miR-30a mimic group). (E) Luciferase activity of LSINCT5 was monitored using dual-luciferase reporter in LSINCT5 (wt/mut), and miR-30a mimic transfected MCF-7 cells. All experiments were conducted in triplicate. The images depicted a representative experiment, and the bar graph is a composite of three independent experiments (**, P<0.01 vs. control). wt, wild type; mut, mutant type; qRT-PCR, quantitative real-time polymerase chain reaction; shRNA, short hairpin RNA; NC, negative control.

Article Snippet: Cell transfection The short hairpin RNA (shRNA) specifically targeting LSINCT5 (called sh-LSINCT5) was synthesized by GenePharma (Shanghai, China), and nonspecific shRNA was used as negative control (sh-NC).

Techniques: Transfection, Control, Quantitative RT-PCR, Luciferase, Activity Assay, Mutagenesis, Real-time Polymerase Chain Reaction, shRNA, Negative Control

Journal: Annals of Translational Medicine

Article Title: Long non-coding RNA LSINCT5 inactivates Wnt/β-catenin pathway to regulate MCF-7 cell proliferation and motility through targeting the miR-30a

doi: 10.21037/atm-20-7253

Figure Lengend Snippet: Knockdown LSINCT5 inhibited cell proliferation by regulating miR-30a in MCF-7 cells. MCF-7 cells were transfected with sh-LSINCT5 or miR-30a inhibitor and in combination. (A) Clone formation images of MCF-7 cells in each group, images were magnified at 20× (stained with 0.2% crystal violet solution). (B) The protein levels of Ki-67 and cleaved-caspase-3 were monitored in each group by Western blot. The GAPDH protein was employed as an internal reference. All experiments were conducted in triplicate. The images depicted a representative experiment, and the bar graph is a composite of three independent experiments. **, P<0.01 vs. control, ##, P<0.01 vs. miR-30a inhibitor group. sh-LSINCT5, short hairpin LSINCT5.

Article Snippet: Cell transfection The short hairpin RNA (shRNA) specifically targeting LSINCT5 (called sh-LSINCT5) was synthesized by GenePharma (Shanghai, China), and nonspecific shRNA was used as negative control (sh-NC).

Techniques: Knockdown, Transfection, Staining, Western Blot, Control

Journal: Annals of Translational Medicine

Article Title: Long non-coding RNA LSINCT5 inactivates Wnt/β-catenin pathway to regulate MCF-7 cell proliferation and motility through targeting the miR-30a

doi: 10.21037/atm-20-7253

Figure Lengend Snippet: Knockdown LSINCT5 suppressed cell motility by regulating miR-30a in MCF-7 cells. MCF-7 cells were transfected with sh-LSINCT5 or miR-30a inhibitor and in combination. (A) Cell invasion images of MCF-7 cells in each group, images were magnified at 200× (stained with 0.1% crystal violet solution). (B) Microscopic images of MCF-7 cells in each group, images were magnified at 100×. (C) The protein levels of epithelial-mesenchymal transition (Vimentin, N-cadherin, and E-cadherin) were monitored in each group by Western blot. The GAPDH protein was employed as an internal reference. All experiments were conducted in triplicate. The images depicted a representative experiment, and the bar graph is a composite of three independent experiments. *, P<0.05, **, P<0.01 vs. control, ##, P<0.01 vs. miR-30a inhibitor group. sh-LSINCT5, short hairpin LSINCT5.

Article Snippet: Cell transfection The short hairpin RNA (shRNA) specifically targeting LSINCT5 (called sh-LSINCT5) was synthesized by GenePharma (Shanghai, China), and nonspecific shRNA was used as negative control (sh-NC).

Techniques: Knockdown, Transfection, Staining, Western Blot, Control

Journal: Annals of Translational Medicine

Article Title: Long non-coding RNA LSINCT5 inactivates Wnt/β-catenin pathway to regulate MCF-7 cell proliferation and motility through targeting the miR-30a

doi: 10.21037/atm-20-7253

Figure Lengend Snippet: β-catenin/TCF4/c-Myc pathway took part in the roles of LSINCT5. MCF-7 cells were transfected with pcDNA-LSINCT5 or ICG-001 and in combination with ICG-001. (A) The protein levels of Wnt pathway-related proteins (β-catenin, TCF4, and c-Myc) were monitored in each group by Western blot. The GAPDH protein was employed as an internal reference. (B) Clone formation images of MCF-7 cells in each group, images were magnified at 20× (stained with 0.2% crystal violet solution). (C) Cell invasion images of MCF-7 cells in each group, images were magnified at 200× (stained with 0.1% crystal violet solution). All experiments were conducted in triplicate. The images depicted a representative experiment, and the bar graph is a composite of three independent experiments. *, P<0.05, **, P<0.01 vs. control, ##, P<0.01 vs. ICG-001 group. TCF4, transcription factor 4; c-Myc, cellular myelocytomatosis oncogene.

Article Snippet: Cell transfection The short hairpin RNA (shRNA) specifically targeting LSINCT5 (called sh-LSINCT5) was synthesized by GenePharma (Shanghai, China), and nonspecific shRNA was used as negative control (sh-NC).

Techniques: Transfection, Western Blot, Staining, Control

Journal: Annals of Translational Medicine

Article Title: Long non-coding RNA LSINCT5 inactivates Wnt/β-catenin pathway to regulate MCF-7 cell proliferation and motility through targeting the miR-30a

doi: 10.21037/atm-20-7253

Figure Lengend Snippet: Knockdown LSINCT5 restrained tumor growth and motility by inactivating the β-catenin/TCF4/c-Myc pathway in vivo. MCF-7 cells untreated or transfected with sh-LSINCT5 were injected into the ventral regions of mice to form tumors. Post successful modeling, mice were divided into three groups with 10 in each group: control group, sh-LSINCT5 group (mice were injected with MCF-7 cells transfected with sh-LSINCT5). (A) Tumor weight in 3 groups. (B) Tumor volume in three groups. (C) LSINCT5 and miR-30a were monitored by qRT-PCR. (D) The protein levels of Wnt pathway-related proteins (β-catenin, TCF4, and c-Myc) were monitored in each group by Western blot. The GAPDH protein was employed as an internal reference. (E) Expression of Ki-67 and Vimentin in tumors were detected by IHC assay, images were magnified at 400×. *, P<0.05, **, P<0.01 vs. control. All experiments were conducted in triplicate. The images depicted a representative experiment, and the bar graph is a composite of ten independent experiments. sh-LSINCT5, short hairpin LSINCT5; sh-NC, short hairpin negative control; TCF4, transcription factor 4; c-Myc, cellular myelocytomatosis oncogene.

Article Snippet: Cell transfection The short hairpin RNA (shRNA) specifically targeting LSINCT5 (called sh-LSINCT5) was synthesized by GenePharma (Shanghai, China), and nonspecific shRNA was used as negative control (sh-NC).

Techniques: Knockdown, In Vivo, Transfection, Injection, Control, Quantitative RT-PCR, Western Blot, Expressing, Negative Control

Journal: Frontiers in Immunology

Article Title: Prognostic stratification of sepsis through DNA damage response based RiskScore system: insights from single-cell RNA-sequencing and transcriptomic profiling

doi: 10.3389/fimmu.2024.1345321

Figure Lengend Snippet: Validation of characteristic genes in vivo (A) Relative expression levels of DDR-related genes in control and sepsis groups (n=4 in each group). (B) Relative expression levels of ARL4C in control, sepsis, sepsis+Ad-shNC, and sepsis+Ad-shARL4C group (n=8 in each group). (C) Expression levels of inflammatory factors (IL-1β, TNF-α, IL-10, and IL-18) in the peripheral blood of rat with sepsis+Ad-shNC and sepsis+Ad-shARL4C (n=8 in each group). (D) Survival status of rats in each group (n=10 in each group). * p < 0.05, ** p < 0.01, *** p < 0.001, **** p < 0.0001. NS, no significant difference.

Article Snippet: For knockdown experiments, shRNA plasmids targeting ARL4C and negative control shRNA were procured from RiboBio (Guangzhou, China), and the specified shRNA lentiviral vectors were generated in 293 T cells.

Techniques: Biomarker Discovery, In Vivo, Expressing, Control

Journal: Frontiers in Immunology

Article Title: Prognostic stratification of sepsis through DNA damage response based RiskScore system: insights from single-cell RNA-sequencing and transcriptomic profiling

doi: 10.3389/fimmu.2024.1345321

Figure Lengend Snippet: Validation of ARL4C in vitro . (A) Flow cytometry detected the apoptosis rate in the Control, Sepsis, Sepsis+Lv-shNC, and Sepsis+Lv-shARL4C groups. (n=4 in each group) (B) Flow cytometry detected the ROS production in the Control, Sepsis, Sepsis+Lv-shNC, and Sepsis+Lv-shARL4C groups. (n=4 in each group), *** p < 0.001, **** p < 0.001. NS, no significant difference.

Article Snippet: For knockdown experiments, shRNA plasmids targeting ARL4C and negative control shRNA were procured from RiboBio (Guangzhou, China), and the specified shRNA lentiviral vectors were generated in 293 T cells.

Techniques: Biomarker Discovery, In Vitro, Flow Cytometry, Control

Journal: Cell Cycle

Article Title: TRG-AS1 is a potent driver of oncogenicity of tongue squamous cell carcinoma through microRNA-543/Yes-associated protein 1 axis regulation

doi: 10.1080/15384101.2020.1786622

Figure Lengend Snippet: The correlations between patients’ clinicopathological parameters and TRG-AS1 expression in TSCC.

Article Snippet: Xenograft tumor model analysis Short hairpin RNA (shRNA) specifically targeting TRG-AS1 (sh-TRG-AS1) and scrambled control shRNA (sh-NC) were synthesized by Shanghai GenePharma Co, Ltd and inserted into the pLKO.1 vector (Biosettia, San Diego, CA, USA).

Techniques: Expressing

Journal: Cell Journal (Yakhteh)

Article Title: FHL1 Overexpression as A Inhibitor of Lung Cancer Cell Invasion via Increasing RhoGDIß mRNA Expression

doi: 10.22074/cellj.2022.8031

Figure Lengend Snippet: FHL1 ectopic expression promoted the protein and mRNA expression of RhoGDIβ . A, B. The Flag-tagged FHL1 plasmid was transfected into A549 and H1299 cell lines stably. The western blot assay was utilized to detect the expression of RhoGDIα and RhoGDIβ protein. β-Actin was used as a protein loading control. C, D. RhoGDIβ mRNA expression was determined by real-time polymerase chain reaction (PCR). The bars indicate the mean ± standard deviation (SD) of 3 independent experiments. The asterisk (*) represents a notable enhancement in comparison with vector control cells (P<0.05).

Article Snippet: In this study, shRNA specific targeting RhoGDIβ (shRhoGDIβ) were obtained from BioVector NTCC Inc. (Cat No. 58897, Shanghai, China).

Techniques: Expressing, Plasmid Preparation, Transfection, Stable Transfection, Western Blot, Control, Real-time Polymerase Chain Reaction, Standard Deviation, Comparison

Journal: Cell Journal (Yakhteh)

Article Title: FHL1 Overexpression as A Inhibitor of Lung Cancer Cell Invasion via Increasing RhoGDIß mRNA Expression

doi: 10.22074/cellj.2022.8031

Figure Lengend Snippet: RhoGDIβ acted as a FHL1 downstream mediator responsible for the FHL1-inhibited human lung cancer invasion. A. The RhoGDIβ knockdown constructs were transfected into A549 (Flag-FHL1) cell lines stably. B. The invasion abilities of A549 (Flag-FHL1/Nonsense), A549 (Flag-FHL1/shRhoGDIβ#1), and A549 (Flag-FHL1/shRhoGDIβ#2) cell lines were detected (scale bar: 200 µm). C. The bars indicate mean ± SD of 3 independent experiments. Student’s t test was used to detect the P value. The asterisk (*) represents a significant increase as compared to A549 (Flag-FHL1/Nonsense) transfectants (P<0.05).

Article Snippet: In this study, shRNA specific targeting RhoGDIβ (shRhoGDIβ) were obtained from BioVector NTCC Inc. (Cat No. 58897, Shanghai, China).

Techniques: Knockdown, Construct, Transfection, Stable Transfection

Journal: Biology Open

Article Title: Downregulated brain and muscle aryl hydrocarbon receptor nuclear translocator-like protein-1 inhibits osteogenesis of BMSCs through p53 in type 2 diabetes mellitus

doi: 10.1242/bio.051482

Figure Lengend Snippet: Downregulated BMAL1 promoted p53 expression in T2DM. (A) Protein expression of BMAL1, p53 and p21 in WT Wistar BMSCs, diabetic GK BMSCs and GK-BMAL1 BMSCs as determined by western blot. (B) qRT-PCR analysis of BMAL1, p53 and p21 in WT Wistar BMSCs, diabetic GK BMSCs and GK-BMAL1 BMSCs. (C) Immunofluorescence staining of p53 in WT Wistar BMSCs, diabetic GK BMSCs and GK-BMAL1 BMSCs. Scale bars: 100 µm. All data are mean±s.e.m. and representative of three independent experiments. n =3. β-actin was used as a loading control. Statistical significance was defined as * P <0.05; ** P <0.01.

Article Snippet: The lentiviral vector carrying specific p53-targeting shRNA was obtained from GenePharma Co., Ltd. (Shanghai, China; Gene ID: 24842).

Techniques: Expressing, Western Blot, Quantitative RT-PCR, Immunofluorescence, Staining

Journal: Biology Open

Article Title: Downregulated brain and muscle aryl hydrocarbon receptor nuclear translocator-like protein-1 inhibits osteogenesis of BMSCs through p53 in type 2 diabetes mellitus

doi: 10.1242/bio.051482

Figure Lengend Snippet: Inhibition of osteogenic differentiation of BMSCs by BMAL1 downregulation in T2DM was mediated by p53. (A) Western blot analysis of BMAL1, Runx2, Alp, Osx and p53 expression in WT Wistar BMSCs, diabetic GK BMSCs and GK-BMAL1 BMSCs with or without 20 μM PFT-α treatment after 7 days of osteogenic differentiation. (B) The mRNA expression of Runx2, Osx and Alp in WT Wistar BMSCs, diabetic GK BMSCs and GK-BMAL1 BMSCs were determined by qRT-PCR, following 7 days of osteogenic differentiation, with or without 20 μM PFT-α treatment. (C) Osteogenic differentiation ability of WT Wistar BMSCs, diabetic GK BMSCs and GK-BMAL1 BMSCs with or without 20 μM PFT-α treatment as determined by alkaline phosphatase activity after osteogenic differentiation for 7 days. Data are from five independent experiments and are expressed as mean±s.e.m.. n =5. Statistical significance was defined as * P <0.05; # P <0.03. n.s: not significant.

Article Snippet: The lentiviral vector carrying specific p53-targeting shRNA was obtained from GenePharma Co., Ltd. (Shanghai, China; Gene ID: 24842).

Techniques: Inhibition, Western Blot, Expressing, Quantitative RT-PCR, Activity Assay