Journal: Acta pharmaceutica Sinica. B

Article Title: A cyclodextrin-based nanoformulation achieves co-delivery of ginsenoside Rg3 and quercetin for chemo-immunotherapy in colorectal cancer.

doi: 10.1016/j.apsb.2021.06.005

Figure Lengend Snippet: Figure 2 Ginsenoside Rg3 induced immunogenic cell death in CRC cells. (A) IC50 of Rg3 for CT26 and HCT116 cells at 24 h. Data are presented as mean SD (n Z 3). (B) Apoptosis (%) in CT26 and HCT116 cells following treatment of Rg3 at 6, 12 and 24 h. Data are presented as mean SD (n Z 3). *P < 0.05 and **P < 0.01 relative to DMSO. (C) The activity of UPR signaling pathways following treatment of Rg3 ([c] Z 30 mmol/L) at 6, 12 and 24 h. The quantification was demonstrated in Fig. S1. (D) The characterization of ICD in CRC cells following treatment of Rg3 ([c] Z 30 mmol/L), including CRT exposure (6 h), ATP secretion (12 h) and HMGB1 release (12 h). Data are presented as mean SD (n Z 3). *P < 0.05 relative to DMSO; scale bar Z 20 mm. (E) The expression of CD11c and CD86 in DCs stimulated with the supernatant from Rg3 ([c] Z 30 mmol/L) -treated cells (24 h). Data are presented as mean SD (n Z 3). **P < 0.01 relative to DMSO. (F) The in vivo vaccination assay using BALB/C and nude mice. Data are presented as mean SD (n Z 4). ***P < 0.001.

Article Snippet: DNA fragments were detected using the TransDetect Fluorescein TUNEL Cell Apoptosis Kit (TransGen Biotech), and nuclei were stained with DAPI (Beyotime Biotech), for confocal microscopic analysis (Olympus); 2) Measurement of immune cells.

Techniques: Activity Assay, Protein-Protein interactions, Expressing, In Vivo

Journal: Acta pharmaceutica Sinica. B

Article Title: A cyclodextrin-based nanoformulation achieves co-delivery of ginsenoside Rg3 and quercetin for chemo-immunotherapy in colorectal cancer.

doi: 10.1016/j.apsb.2021.06.005

Figure Lengend Snippet: Figure 3 Quercetin caused reactive oxygen species in CRC cells. (A) IC50 of QTN for CT26 and HCT116 cells at 24 h. Data are presented as mean SD (n Z 3). (B) Apoptosis (%) in CT26 and HCT116 cells following treatment of QTN ([c] Z 80 mmol/L) at 24 h. Data are presented as mean SD (n Z 3). **P < 0.01 relative to DMSO. (C) The activity of Bcl-2/BAX/caspase 9/caspase 3 signaling pathways following treatment of QTN ([c] Z 80 mmol/L) at 6, 12 and 24 h. The quantification was demonstrated in Fig. S2. (D) The ROS level in CT26 and HCT116 cells following treatment of QTN ([c] Z 80 mmol/L) at 6, 12 and 24 h. Data are presented as mean SD (n Z 3). **P < 0.01 and ***P < 0.001 relative to DMSO. (E) Cell viability (%) of CT26 and HCT116 cells with or without NAC prior to treatment of QTN ([c] Z 80 mmol/L) (24 h). Data are presented as mean SD (n Z 3). *P < 0.05 and **P < 0.01 relative to untreated control. (F) Apoptosis (%) in CT26 and HCT116 cells with or without NAC prior to treatment of QTN ([c] Z 80 mmol/L) (24 h). Data are presented as mean SD (n Z 3). *P < 0.05 and **P < 0.01 relative to untreated control.

Article Snippet: DNA fragments were detected using the TransDetect Fluorescein TUNEL Cell Apoptosis Kit (TransGen Biotech), and nuclei were stained with DAPI (Beyotime Biotech), for confocal microscopic analysis (Olympus); 2) Measurement of immune cells.

Techniques: Activity Assay, Protein-Protein interactions, Control

Journal: Acta pharmaceutica Sinica. B

Article Title: A cyclodextrin-based nanoformulation achieves co-delivery of ginsenoside Rg3 and quercetin for chemo-immunotherapy in colorectal cancer.

doi: 10.1016/j.apsb.2021.06.005

Figure Lengend Snippet: Figure 4 Synergistic effects of Rg3 and QTN in CT26 cells. (A) IC50 of drug combination at 24 h. Data are presented as mean SD (n Z 3). CI values at IC50 were shown in Fig. S3. (B) Apoptosis (%) caused by drug combination at 24 h. Data are presented as mean SD (n Z 3). *P < 0.05 and **P < 0.01 relative to DMSO. (C) The CRT exposure with or without NAC before treatment of drug combination (6 h). Data are presented as mean SD (n Z 3). *P < 0.05 and **P < 0.01 relative to DMSO; scale bar Z 20 mm. (D) The ATP secretion with or without NAC before treatment of drug combination at 12 h. Data are presented as mean SD (n Z 3).*P < 0.05 and **P < 0.01, between NAC and No NAC. (E) The HMGB1 release with or without NAC before treatment of drug combination at 12 h. Data are presented as mean SD (n Z 3). *P < 0.05 and **P < 0.01, between NAC and No NAC. (F) The expression of CD11c and CD86 in DCs stimulated (24 h) by the supernatant from Rg3- treated cells with or without pretreatment of NAC. Data are presented as mean SD (n Z 3). *P < 0.05, **P < 0.01 and ***P < 0.001, relative to DMSO.

Article Snippet: DNA fragments were detected using the TransDetect Fluorescein TUNEL Cell Apoptosis Kit (TransGen Biotech), and nuclei were stained with DAPI (Beyotime Biotech), for confocal microscopic analysis (Olympus); 2) Measurement of immune cells.

Techniques: Expressing

Journal: Acta pharmaceutica Sinica. B

Article Title: A cyclodextrin-based nanoformulation achieves co-delivery of ginsenoside Rg3 and quercetin for chemo-immunotherapy in colorectal cancer.

doi: 10.1016/j.apsb.2021.06.005

Figure Lengend Snippet: Figure 8 Combination therapy of targeted co-formulation and Anti-PD-L1 for CRC. (A) Treatment schedule and IVIS images. (B) The CRC progression over a 35-day period. Data are presented as mean SD (n Z 5). *P < 0.05 and **P < 0.01; NS, no significance. (C) Animal survival (median survival: PBS ~38 days, Anti-PD-L1 ~40 days, targeted co-formulation ~62 days, and combination z 96 days). Data are presented as mean SD (n Z 5). **P < 0.01 and ***P < 0.001. (D) Immunofluorescent staining assay (green Z DNA fragments and blue Z nuclei) on Day 20 to assess apoptosis in the tumor (scale bar Z 50 mm). Data are presented as mean SD (n Z 3). *P < 0.05 and **P < 0.01, relative to PBS. (E) Level of immune cells in the tumor on Day 20 was analyzed using flow cytometry (BD). Data are presented as mean SD (n Z 4). *P < 0.05 and **P < 0.01; NS, no significance. (F) The mRNA expression of cytokines and chemokines in the tumor on Day 20 was analyzed using real time RT-PCR. Data are presented as mean SD (n Z 4). *P < 0.05 and **P < 0.01; NS, no significance. (G) Orthotopic CRC mice treated with targeted co-formulation following the removal of CD4 þ or CD8þ T cells. Data are presented as mean SD (n Z 4). *P < 0.05 and **P < 0.01; NS, no significance.

Article Snippet: DNA fragments were detected using the TransDetect Fluorescein TUNEL Cell Apoptosis Kit (TransGen Biotech), and nuclei were stained with DAPI (Beyotime Biotech), for confocal microscopic analysis (Olympus); 2) Measurement of immune cells.

Techniques: Formulation, Staining, Cytometry, Expressing, Quantitative RT-PCR

Journal: Frontiers in Physiology

Article Title: RvD2 mitigates TNFɑ-Induced mitochondrial reactive oxygen species through NRF2 signaling in placental trophoblasts

doi: 10.3389/fphys.2025.1547940

Figure Lengend Snippet: RvD2 upregulates the NRF2 signaling cascade in TNFɑ-induced JEG-3 cells. For cotreatment of TNFɑ + RvD2 (TR) groups, cells were pretreated with RvD2 for 16 h, followed by TNFɑ treatment for an additional 5 or 10 h (16 + 5 h or 16 + 10 h), resulting in total treatment durations of 21 and 26 h, respectively. For vehicle (V) or RvD2 (R) treatments, cells were treated for a total of 21 or 26 h. Cells treated with TNFɑ (T) were exposed for either 5 or 10 h. (A) Immunoblot analysis of NRF2 in 16 + 5 h and 16 + 10 h treatment strategies. The values below the immunoblot represent band intensity ratio of nNRF2/HDAC1. The same blot was used in . (B–G) Relative mRNA expression of 16 + 5 h treatment strategy of kelch-like ECH-associated protein 1 (KEAP1), hemoxygenase 1 (HOXO1), glutamate-cysteine ligase catalytic subunit (GCLC), glutamate-cysteine ligase modifier subunit (GCLM), NADPH quinone oxidoreductase 1 (NQO1) in trophoblasts; n = 3 per group. (H) Reduced glutathione was measured with the pretreatment of RvD2 (100 nM) for 16 h followed by a 1 h treatment of TNFɑ (100 ng/mL); n = 5–7 per group. Data presented as mean ± SEM; *p < 0.05 and **p < 0.01 compared against each treatment.

Article Snippet: The Cellular Glutathione Detection Assay Kit (Cell Signaling Technology, Catalog #13859) was used to quantify reduced glutathione (GSH) concentrations, following the manufacturer’s instructions.

Techniques: Western Blot, Expressing

Journal: International Journal of Biological Sciences

Article Title: NDRG1 regulates Filopodia-induced Colorectal Cancer invasiveness via modulating CDC42 activity

doi: 10.7150/ijbs.56694

Figure Lengend Snippet: Regulation of CDC42 activity by NDRG1 in CRC cells. A) Immunoblotting for total protein level or activated form of indicated Rho GTPase in NDRG1-modified HCT116 and RKO cells. Results are representative of at least three biological repeats, and the values in histograms are represented by mean ± S.D.; *P value <0.05, **P value <0.01, relative to the respective control cells. B) Confocal images were taken to show immunofluorescence staining of active-CDC42 (red) accompanied by the cell nucleus (blue) stained by DAPI in NDRG1 overexpression and NDRG1 knockdown HCT116 and RKO cells relative to the control cells, respectively. Fluorescence quantification was performed by comparing the integrated optical density (IOD)/area value of active-CDC42 to the IOD/area value of the nucleus (DAPI) in the same image. Results are representative of three to five images from different visual fields, and the histogram values are mean ±S.D. *P value <0.05, ***P<0.001, relative to the respective control cells. Scale bars: 25 µm.

Article Snippet: GST-pull down assay to detect active CDC42 and RAC1 was carried out as the protocol of Active CDC42 Detection Kit (Cat.8819, Cell Signaling Technology) and Active RAC1 Detection Kit (Cat.8815, Cell Signaling Technology).

Techniques: Activity Assay, Western Blot, Modification, Control, Immunofluorescence, Staining, Over Expression, Knockdown, Fluorescence

Journal: International Journal of Biological Sciences

Article Title: NDRG1 regulates Filopodia-induced Colorectal Cancer invasiveness via modulating CDC42 activity

doi: 10.7150/ijbs.56694

Figure Lengend Snippet: Inhibition of CDC42 prevents NDRG1 loss induced CRC cell filopodial protrusion formation through suppression of PAK1/Cofilin signaling. A) Immunoblotting analysis of the expression level of the total and phosphorylation form of PAK1 and Cofilin in indicated cell lines. B) Knockdown of CDC42 in HCT116 (left) and RKO (right) cells confirmed with immunoblotting analysis. Pool, combined siCDC42 sequences. C) Expression level of the total and phosphorylation form of PAK1 and Cofilin in indicated cell lines. D) Confocal images were taken to show immunofluorescence staining of MYO10 (green) and rhodamine-phalloidin (red) accompanied by the cell nucleus (blue) in colorectal cancer cells. Quantification of the MYO10-associated filopodial protrusions density and length is represented as mean ± S.D.; results are representative of 3-5 images from different visual fields, n>50 cells. *P value <0.05, **P value <0.01, ***P < 0.001, relative to the sh-Con/si-Con groups. # P value <0.05, ## P value <0.01, ### P < 0.001, relative to the sh-NDRG1/si-Con groups.

Article Snippet: GST-pull down assay to detect active CDC42 and RAC1 was carried out as the protocol of Active CDC42 Detection Kit (Cat.8819, Cell Signaling Technology) and Active RAC1 Detection Kit (Cat.8815, Cell Signaling Technology).

Techniques: Inhibition, Western Blot, Expressing, Phospho-proteomics, Knockdown, Immunofluorescence, Staining

Journal: International Journal of Biological Sciences

Article Title: NDRG1 regulates Filopodia-induced Colorectal Cancer invasiveness via modulating CDC42 activity

doi: 10.7150/ijbs.56694

Figure Lengend Snippet: NDRG1 suppresses CDC42 activity by stabilizing the RhoGDIα-CDC42 binding. A) The STRING network view of interactive proteins of CDC42 in humans. Gray lines between the nodes indicate various types of interaction evidence. B) Co-immunoprecipitation to examine the interaction of RhoGDIα and CDC42 in both HCT116 and RKO cell lines. C) Immunoblotting assay to evaluate the influence of NDRG1 modification on RhoGDIα expression in indicated cells. GAPDH was used as loading control. D) Double stained confocal immunofluorescence assay and co-localization analysis to confirm the interaction of RhoGDIα and CDC42 in indicated cells (red: CDC42, green: RhoGDIα, blue: DAPI, scale bar: 20 µm). Co-localization analysis on wide-field merged images was performed via Leica Application Suite X. Results are representative of five images from different visual fields.

Article Snippet: GST-pull down assay to detect active CDC42 and RAC1 was carried out as the protocol of Active CDC42 Detection Kit (Cat.8819, Cell Signaling Technology) and Active RAC1 Detection Kit (Cat.8815, Cell Signaling Technology).

Techniques: Activity Assay, Binding Assay, Immunoprecipitation, Western Blot, Modification, Expressing, Control, Staining, Immunofluorescence

Journal: International Journal of Biological Sciences

Article Title: NDRG1 regulates Filopodia-induced Colorectal Cancer invasiveness via modulating CDC42 activity

doi: 10.7150/ijbs.56694

Figure Lengend Snippet: Silence of NDRG1 promotes the peritoneal metastasis and correlates with upregulated CDC42 GTP expression. A) Peritoneal metastasis of CRC cells in BALB/c nude mice. Tumors in two groups were measured in situ and assessed by bioluminescence imaging in the fourth week. B) Statistical analysis of the bioluminescence in peritoneal foci of both groups. Results are shown as mean ± S.D. C) Tumors in two groups are demonstrated after laparotomy with hematoxylin-eosin staining of peritoneal foci on the lower panel. Scale bars are as indicated. D) Immunofluorescence staining of NDRG1 (left) or CDC42 GTP (right) accompanied by the cell nucleus stained by DAPI in peritoneal foci derived from sh-NDRG1 and control groups. Results are representative of 3-5 images from different visual fields and the histogram values are mean ± S.D.; *P value <0.05, ***P< 0.001, relative to the respective control groups. Scale bar: 50 µm.

Article Snippet: GST-pull down assay to detect active CDC42 and RAC1 was carried out as the protocol of Active CDC42 Detection Kit (Cat.8819, Cell Signaling Technology) and Active RAC1 Detection Kit (Cat.8815, Cell Signaling Technology).

Techniques: Expressing, In Situ, Imaging, Staining, Immunofluorescence, Derivative Assay, Control

Journal: International Journal of Biological Sciences

Article Title: NDRG1 regulates Filopodia-induced Colorectal Cancer invasiveness via modulating CDC42 activity

doi: 10.7150/ijbs.56694

Figure Lengend Snippet: CDC42 GTP is frequently upregulated in CRC tissues and correlated with NDRG1 expression and clinicopathological parameters. A) IHC staining of NDRG1 and active CDC42 expression in tumor and adjacent tissues in microarray. Magnification on the right with a scale bar of 100 µm. B) Heatmap illustrating different clinicopathological parameters between CDC42 GTP -high and -low-expression tumors of the 86 cases. Statistical significance was analyzed by the χ 2 test. P values are as indicated.

Article Snippet: GST-pull down assay to detect active CDC42 and RAC1 was carried out as the protocol of Active CDC42 Detection Kit (Cat.8819, Cell Signaling Technology) and Active RAC1 Detection Kit (Cat.8815, Cell Signaling Technology).

Techniques: Expressing, Immunohistochemistry, Microarray

Journal: International Journal of Biological Sciences

Article Title: NDRG1 regulates Filopodia-induced Colorectal Cancer invasiveness via modulating CDC42 activity

doi: 10.7150/ijbs.56694

Figure Lengend Snippet: Schematic diagram for the mechanism of NDRG1's regulation of CDC42/PAK1/Cofilin axis as a switch that modulates actin cytoskeleton rearrangement in human colorectal cancer invasion by stabilizing the RhoGDIα-CDC42 binding.

Article Snippet: GST-pull down assay to detect active CDC42 and RAC1 was carried out as the protocol of Active CDC42 Detection Kit (Cat.8819, Cell Signaling Technology) and Active RAC1 Detection Kit (Cat.8815, Cell Signaling Technology).

Techniques: Binding Assay

Journal: Cell Reports Methods

Article Title: A genetic tool for the longitudinal study of a subset of post-inflammatory reactive astrocytes

doi: 10.1016/j.crmeth.2022.100276

Figure Lengend Snippet: Expression of Lcn2CreERT2 following repeated low-dose LPS treatment (A) Immunostaining for tdTomato in Lcn2CreERT2 ; Ai9 brains. Animals were given one of the following four treatments for 5 consecutive days: Saline and corn oil, saline and tamoxifen, LPS and corn oil, LPS and tamoxifen. Sections were counterstained with DAPI. (B) Quantification of total number of red cells/section from animals in (A). ∗∗∗p < 0.001 ANOVA and Tukey’s post hoc test, n = 5 animals per treatment. (C) Immunostaining of brains from saline- (left) or LPS-treated (right) Lcn2CreERT2 ; Ai9 animals for tdTomato and the astrocytic marker GFAP (top), the endothelial and microglial marker lectin (second row), the neuronal marker NeuN (third row), and the microglial marker Iba1 (bottom). Sections were counterstained with DAPI. White arrowheads indicate co-positive cells for tdTomato and the panel maker. (D) Quantification of co-positive cells from (C). (E) RNAscope for the astrocytic marker Slc1a3 (top) and the endothelial marker Pecam1 (bottom) on brains from LPS- and saline-treated Lcn2CreERT2 ; Ai9 animals. Sections were immunostained for tdTomato and counterstained with DAPI. (F) Quantification of cells co-positive for tdTomato and RNA markers described in (E). Scale bars, (A) 250 μm, (C) 50 μm, (E) 25 μm. Error bars ± SEM. See also Figures S1 and .

Article Snippet: RNAScope Multiplex Fluorescent Detection kit v2 , Advanced Cell Diagnostics , 323110.

Techniques: Expressing, Immunostaining, Saline, Marker, RNAscope

Journal: Cell Reports Methods

Article Title: A genetic tool for the longitudinal study of a subset of post-inflammatory reactive astrocytes

doi: 10.1016/j.crmeth.2022.100276

Figure Lengend Snippet: Astrocytes that were initially reactive remain in the brain after resolution of inflammation (A) Lcn2CreERT2 ; Ai9 brain sections immunostained for tdTomato and counterstained with DAPI. Animals were given tamoxifen and saline or LPS and collected at either 24 h or 1 month after the last treatment. (B) Quantification of tdTomato-positive cells in the thalamic region. p = 0.38, ANOVA followed by Tukey’s HSD post hoc, n = 5 animals. (C) RNAScope on Lcn2CreERT2 ; Ai9 brain sections for the astrocyte marker Aldh1l1 24 h and 1 month after low-dose LPS treatment, followed by immunostaining for tdTomato and counterstaining with DAPI. (D) Quantification of Aldh1l1 and tdTomato co-positive cells in (C). p = 0.16, paired t test. n = 18 pooled sections across three animals. (E) Co-labeling of tdTomato and EdU-positive astrocytes in Lcn2CreERT2 ; Ai9 animals after LPS treatment. (F) High magnification image of labeled astrocytes and EdU-positive cells at 1 month. (G) Quantification of tdTomato and EdU co-positive astrocytes at 24 h, 7 days, and 1 month following LPS treatment. Error bars ± SEM. Scale bars, (A) 400 μm, (C, E) 200 μm, (F) 20 μm. See also Figure S4 .

Article Snippet: RNAScope Multiplex Fluorescent Detection kit v2 , Advanced Cell Diagnostics , 323110.

Techniques: Saline, RNAscope, Marker, Immunostaining, Labeling

Journal: Cell Reports Methods

Article Title: A genetic tool for the longitudinal study of a subset of post-inflammatory reactive astrocytes

doi: 10.1016/j.crmeth.2022.100276

Figure Lengend Snippet: Reactive marker expression decreases 1 month after inflammation (A and B) Analysis of reactive marker expression within tdTomato-positive cells. RNAScope for brain sections 24 h after LPS, or 1 month after LPS treatment to detect expression of the reactive markers Lcn2 (A) and GFAP (B), followed by immunostaining for tdTomato and counterstaining with DAPI. RNAScope for reactive markers Lcn2 (A) and GFAP (B) was quantified by looking at both number of fluorescent puncta (A, B, top panels) and integrated fluorescent density (A, B, bottom panels). ∗∗∗p < 0.001, paired t test. n = 29 astrocytes pooled from three animals. Error bars ± SEM. Scale bars, 50 μm. See also Figure S5 .

Article Snippet: RNAScope Multiplex Fluorescent Detection kit v2 , Advanced Cell Diagnostics , 323110.

Techniques: Marker, Expressing, RNAscope, Immunostaining

Journal: Cell Reports Methods

Article Title: A genetic tool for the longitudinal study of a subset of post-inflammatory reactive astrocytes

doi: 10.1016/j.crmeth.2022.100276

Figure Lengend Snippet: Reactive markers return to baseline levels in astrocytes that were initially reactive (A–C) RNAScope staining (green) for Slc1a2 (A), Slc1a3 (B), and Lrrc8a (C) in Lcn2CreERT2 ; Ai9 animals treated with saline or LPS. Sections were immunostained for tdTomato and counterstained with DAPI. Bottom: quantification of RNAScope puncta per tdTomato + astrocyte. ∗∗p < 0.01, ∗∗∗p < 0.001, ∗∗∗∗p < 0.0001, ANOVA followed by Tukey’s honest significant difference post hoc test. n = 30 pooled astrocytes across three animals. Error bars ± SEM. Scale bars, 20 μm.

Article Snippet: RNAScope Multiplex Fluorescent Detection kit v2 , Advanced Cell Diagnostics , 323110.

Techniques: RNAscope, Staining, Saline

Journal: Cell Reports Methods

Article Title: A genetic tool for the longitudinal study of a subset of post-inflammatory reactive astrocytes

doi: 10.1016/j.crmeth.2022.100276

Figure Lengend Snippet:

Article Snippet: RNAScope Multiplex Fluorescent Detection kit v2 , Advanced Cell Diagnostics , 323110.

Techniques: Virus, Recombinant, Plasmid Preparation, RNAscope, Multiplex Assay