Journal: Acta Pharmaceutica Sinica. B

Article Title: CRISPR-Cas9 gene editing strengthens cuproptosis/chemodynamic/ferroptosis synergistic cancer therapy

doi: 10.1016/j.apsb.2024.05.029

Figure Lengend Snippet: In vitro distribution and tumor-combatting effect of RNP@Cu 2 O@SPF. (A) The viability of cancer and normal cells treated with RNP@Cu 2 O@SPF for 24 h in the dark ( n = 6). (B) CLSM images of SW480 cells incubated with RNP@Cu 2 O@SPF or RNP@Cu 2 O@S (FITC-labelled RNP, RITC-labelled silica). Scale bar, 20 μm. (C) FCM analysis showcasing transfection rates in SW480 cells after 4 h of immersion with PBS, RNP@Cu 2 O@S, or RNP@Cu 2 O@SPF. WB assay (D) and fluorescence imaging (E) of ATP7A expression in SW480 cells treated with RNP@Cu 2 O@S or RNP@Cu 2 O@SPF. Scale bar, 100 μm. (F) Relative copper contents in SW480 cells after a 24 h treatment with Cu 2 O@SPF or RNP@Cu 2 O@SPF ( n = 6). (G) Fluorescence microscopy images of ROS generation in SW480 cells treated with PBS, Cu 2 O@SPF(40 μg/mL), RNP@Cu 2 O@SPF (40 μg/mL), and RNP@Cu 2 O@SPF (80 μg/mL) for 4 h and stained with DCFH-DA to indicate the ROS generation. Scale bar, 50 μm. (H) GSH/GSSG ratio in SW480 cells treated with different concentrations of RNP@Cu 2 O@SPF (25, 50, 100 μg/mL) for 6 h ( n = 6). The data are shown as mean ± SD. ns, no significant; ∗ P < 0.05, ∗∗ P < 0.01.

Article Snippet: The Cell Copper (Cu) Colorimetric Assay Kit (Elabscience Biotechnology, Wuhan, China) was selected to detect the copper ion content in SW480 cells, and the BCA Protein Colorimetric Assay Kit (Elabscience Biotechnology, Wuhan, China) was also used to determine the total protein concentration.

Techniques: In Vitro, Incubation, Transfection, Fluorescence, Imaging, Expressing, Microscopy, Staining

Journal: Acta Pharmaceutica Sinica. B

Article Title: CRISPR-Cas9 gene editing strengthens cuproptosis/chemodynamic/ferroptosis synergistic cancer therapy

doi: 10.1016/j.apsb.2024.05.029

Figure Lengend Snippet: Anti-tumor mechanisms studies of RNP@Cu 2 O@SPF. (A) The schematic diagram of cell death mechanisms of RNP@Cu 2 O@SPF nanomedicine-mediated synergetic therapy. (B) FCM analysis with Annexin V-FITC/PI dual labels on SW480 cells after 24 h of incubation with different nanoparticles. (C) The percentages of cells undergoing apoptosis (%) in different groups ( n = 3). (D) Viability of SW480 cells grown in media containing either glucose or galactose treated with RNP@Cu 2 O@SPF (ratio 1:1) ( n = 6). (E) Viability of SW480 cells pretreated with 0.1 mmol/L rotenone (Rot), 0.1 mmol/L antimycin A (anti-A), or 1 mmol/L FCCP and then treated with RNP@Cu 2 O@SPF ( n = 6). (F) Viability of SW480 cells post-treated with Antimycin A or UK5099 under different conditions (incubated with various nanoparticles) ( n = 6). (G) Viability of SW480 cells treated with tetrathiomolybdate in different concentrations of RNP@Cu 2 O@SPF (50, 100 μg/mL) ( n = 6). (H) Western blot analysis of DLAT-oligomers, DLAT, and Tubulin expression levels in SW480 cells after treatment with different nanoparticles. (I) Western blot analysis of lipoylated proteins, FDX1, LIAS, ACO-2, SDHB9, and Tubulin expression levels in SW480 cells after treatment with different nanoparticles. (J) Fluorescence depiction of GPX4 levels in SW480 cells post RNP@Cu 2 O@SPF treatment at different dosages (25, 50, and 100 μg/mL) for 24 h. Scale bar, 25 μm. (K) LPO contents in SW480 cells treated with RNP@Cu 2 O@SPF at different concentrations (25, 50, and 100 μg/mL) for 24 h ( n = 6). (L) Survival rates of SW480 cells after 24 h of incubation with solutions of PBS, RNP@Cu 2 O@SPF, PBS+DFO, or RNP@Cu 2 O@SPF+DFO ( n = 6). (M) WB assay of GPX4, ACSL4, and Tubulin expression in SW480 cells treated with RNP@Cu 2 O@SPF at varied concentrations (25, 50, and 100 μg/mL) for 24 h. The data are shown as mean ± SD. ns, not significant; ∗ P < 0.05, ∗∗ P < 0.01.

Article Snippet: The Cell Copper (Cu) Colorimetric Assay Kit (Elabscience Biotechnology, Wuhan, China) was selected to detect the copper ion content in SW480 cells, and the BCA Protein Colorimetric Assay Kit (Elabscience Biotechnology, Wuhan, China) was also used to determine the total protein concentration.

Techniques: Incubation, Western Blot, Expressing, Fluorescence

Journal: American Journal of Translational Research

Article Title: CD44 expression enhances chemoresistance and implies occult micrometastases after conversion hepatectomy for initially unresectable colorectal liver metastases

doi:

Figure Lengend Snippet: CD44 protein is expressed in colon cancer cell lines and its downregulation attenuates cell growth. A. CD44 protein expression in nine colon cancer cell lines by western blotting. SW480 and Lovo strongly expressed CD44s protein, whereas HT29 expressed CD44v protein. β-actin is used as an internal control. B. Downregulation of Pan-CD44 mRNA using siRNA in SW480 cells at 24 h. C. Downregulation of CD44 protein expression in SW480 cells (with CD44s protein) by siRNA at 48 h. D. Downregulation of CD44 expression inhibits cell growth in SW480 cells compared to control. E. Downregulation of CD44 protein expression in HT29 cells (with CD44v protein) by siRNA at 48 h. F. Downregulation of CD44 expression inhibits cell growth in HT29 cells compared to control.

Article Snippet: The human colon cancer lines SW620, SW480, Lovo, HCT15, COLO201, COLO320, DLD1, COLO205, and HT29 were purchased from the Japanese Collection of Research Bioresources.

Techniques: Expressing, Western Blot, Control

Journal: American Journal of Translational Research

Article Title: CD44 expression enhances chemoresistance and implies occult micrometastases after conversion hepatectomy for initially unresectable colorectal liver metastases

doi:

Figure Lengend Snippet: Functional role of CD44 expression in chemoresistance in colon cancer cells. A. Downregulation of CD44 expression inhibits cell growth under 5-FU treatment (500 μM) in SW480 cells (with CD44s protein). B. Downregulation of CD44 expression inhibits cell growth under 5-FU treatment (500 μM) in HT29 cells (with CD44v protein). C. Downregulation of CD44 expression enhances cleaved-PARP and cleaved-caspase 3 expression in SW480 cells under 5-FU treatment (500 μM) by western blot analysis at 48 h. β-actin is used as an internal control. D. Downregulation of CD44 expression enhances cleaved-PARP and cleaved-caspase 3 expression in HT29 cells under 5-FU treatment (500 μM) by western blot analysis at 48 h. β-actin is used as an internal control. E. Downregulation of CD44 expression inhibits cell growth under L-OHP treatment (20 μM) in SW480 cells (with CD44s protein). F. Downregulation of CD44 expression inhibits cell growth under L-OHP treatment (20 μM) in HT29 cells (with CD44v protein). G. Downregulation of CD44 expression does not affect protein expression levels of cleaved-PARP, cleaved-caspase 3, DPD, or ERCC1 in SW480 cells under L-OHP treatment (20 μM) by western blot analysis at 48 h. H. Downregulation of CD44 expression does not affect protein expression levels of cleaved-PARP, cleaved-caspase 3, DPD, or ERCC1 in HT29 cells under L-OHP treatment (20 μM) by western blot analysis at 48 h. MIA Paca-2 cells were used as a positive control for DPD expression.

Article Snippet: The human colon cancer lines SW620, SW480, Lovo, HCT15, COLO201, COLO320, DLD1, COLO205, and HT29 were purchased from the Japanese Collection of Research Bioresources.

Techniques: Functional Assay, Expressing, Western Blot, Control, Positive Control

Journal: American Journal of Translational Research

Article Title: CD44 expression enhances chemoresistance and implies occult micrometastases after conversion hepatectomy for initially unresectable colorectal liver metastases

doi:

Figure Lengend Snippet: Downregulation of CD44 expression does not affect chemoresistance under irinotecan (CPT-11) treatment in colon cancer cell lines. A. Downregulation of CD44 expression does not affect cell growth under CPT-11 treatment (20 μM) in SW480 cells (with CD44s protein). B. Downregulation of CD44 expression does not affect cell growth under CPT-11 treatment (20 μM) in HT29 cells (with CD44v protein). β-actin is used as an internal control.

Article Snippet: The human colon cancer lines SW620, SW480, Lovo, HCT15, COLO201, COLO320, DLD1, COLO205, and HT29 were purchased from the Japanese Collection of Research Bioresources.

Techniques: Expressing, Control

Journal: International Journal of Molecular Sciences

Article Title: PI3K-AKT Pathway Modulation by Thymoquinone Limits Tumor Growth and Glycolytic Metabolism in Colorectal Cancer

doi: 10.3390/ijms23042305

Figure Lengend Snippet: Thymoquinone induces cell death and hampers cell proliferation in CRC cells. HCT116 and SW480 CRC cells were treated with thymoquinone as indicated. ( A ) Thymoquinone induced cell death in a dose and time-dependent manner, statistical significance was calculated by two-way ANOVA, considering both time and concentration-dependent observations. ( B ) Thymoquinone treatment-induced cell death/inhibited cell doublings per day over a time period of 1–4 days. ( C ) Pre-treatment of thymoquinone to HCT116 (21.71 µM) and SW480 (20.53 µM) led to a significant reduction in cell proliferation over time (0–4 days), statistical significance was calculated by employing two way ANOVA following appropriate post hoc test (Bonferroni test). The results are expressed mean ± SEM ( n = 3 or 4). ** p < 0.01, *** p < 0.001.

Article Snippet: The study was conducted using two CRC cell lines, HCT116 and SW480, acquired from National Centre for Cell Sciences, Pune India (NCCS, Pune) India.

Techniques: Concentration Assay

Journal: International Journal of Molecular Sciences

Article Title: PI3K-AKT Pathway Modulation by Thymoquinone Limits Tumor Growth and Glycolytic Metabolism in Colorectal Cancer

doi: 10.3390/ijms23042305

Figure Lengend Snippet: Thymoquinone reduces glycolytic metabolism in CRC. HCT116 and SW480 CRC cells were treated with thymoquinone as indicated. Thymoquinone treatment led to a significant reduction in—( A ) glucose fermentation rate/s (glucose production and/or lactate production), ( B ) ATP production, and ( C ) redox state (NADPH) in these cells. Statistical significance was calculated by employing unpaired t-test, data expressed as mean ± SEM ( n = 3). * p < 0.05 and ** p < 0.01.

Article Snippet: The study was conducted using two CRC cell lines, HCT116 and SW480, acquired from National Centre for Cell Sciences, Pune India (NCCS, Pune) India.

Techniques:

Journal: International Journal of Molecular Sciences

Article Title: PI3K-AKT Pathway Modulation by Thymoquinone Limits Tumor Growth and Glycolytic Metabolism in Colorectal Cancer

doi: 10.3390/ijms23042305

Figure Lengend Snippet: Thymoquinone inhibits Hexokinase-2 via modulating the PI3K-AKT pathway. HCT116 and SW480 CRC cells were treated with thymoquinone or LY294002 as indicated. ( A ) Thymoquinone treatment led to inhibition in HK2 protein levels. Such inhibition in HK2 was concomitant to inhibition in—( B ) the PI3K-AKT pathway. ( C ) Inhibiting the PI3K-AKT pathway using selective PI3K pharmacologic inhibitor LY294002 led to an inhibition of HK2 protein levels.

Article Snippet: The study was conducted using two CRC cell lines, HCT116 and SW480, acquired from National Centre for Cell Sciences, Pune India (NCCS, Pune) India.

Techniques: Inhibition

Journal: International Journal of Molecular Sciences

Article Title: PI3K-AKT Pathway Modulation by Thymoquinone Limits Tumor Growth and Glycolytic Metabolism in Colorectal Cancer

doi: 10.3390/ijms23042305

Figure Lengend Snippet: Inhibition of PI3K-AKT/ HK2 pathway impedes tumorigenic potential of CRC cells. HCT116 and SW480 CRC cells were treated as indicated. Inhibition of PI3K with its pharmacologic inhibitor LY294002 led to a significant inhibition in—( A ) cell proliferation, and ( B ) clonogenic propensity of these cells. Genetic (siRNA) or pharmacologic (3-Bromopyruvate) inhibition of HK2 led to a significant reduction in—( C ) proliferation, and ( D ) Clonogenic potential of these cells. Statistical analyses were carried out employing two-tailed paired t -test (proliferation assays) and two-way ANOVA-(Clonogenic assays) followed by appropriate post hoc tests (Bonferroni). Pharmacologic (LY294002) inhibition of PI3K led to an inhibition in ( E ) glucose fermentation rate/s (glucose uptake and lactate excretion), ( F ) ATP production and ( G ) redox state (NADPH). Genetic (siRNA) ablation of HK2 led to a significant abrogation in ( H ) glucose fermentation rate/s (glucose uptake and lactate excretion), ( I ) ATP production and ( J ) redox state (NADPH). Statistical significance was calculated by employing unpaired t-test, data expressed as mean ± SEM ( n = 3 or 4). * p < 0.05, ** p < 0.01, *** p < 0.001.

Article Snippet: The study was conducted using two CRC cell lines, HCT116 and SW480, acquired from National Centre for Cell Sciences, Pune India (NCCS, Pune) India.

Techniques: Inhibition, Two Tailed Test

Journal: International Journal of Molecular Sciences

Article Title: PI3K-AKT Pathway Modulation by Thymoquinone Limits Tumor Growth and Glycolytic Metabolism in Colorectal Cancer

doi: 10.3390/ijms23042305

Figure Lengend Snippet: Thymoquinone inhibits cell migration and invasion via modulating glucose metabolic reprogramming. HCT116 and SW480 CRC cells were treated as indicated. Treating these cells with thymoquinone led to a significant inhibition in—( A ) wound healing, and ( B ) invasion potential. ( C ) Thymoquinone also led to induction in E cadherin with a concomitant inhibition in N cadherin levels. Genetic (siRNA) ablation of HK2 led to a significant inhibition in—( D ) wound healing, and ( E ) cell invasion. ( F ) HK2 ablation also led to induction in E cadherin and inhibition in N cadherin levels. Statistical analyses were carried out by employing two-tailed paired t -test, data expressed as mean ± SEM ( n = 3). * p < 0.05, ** p < 0.01.

Article Snippet: The study was conducted using two CRC cell lines, HCT116 and SW480, acquired from National Centre for Cell Sciences, Pune India (NCCS, Pune) India.

Techniques: Migration, Inhibition, Two Tailed Test