control shrna lentiviral particles  (Santa Cruz Biotechnology)

Journal: Cancer Medicine

Article Title: Influence of Citrobacter freundii on NINJ2 Expression and Oxaliplatin Resistance in Colorectal Cancer

doi: 10.1002/cam4.70940

Figure Lengend Snippet: Comprehensive gene analysis associated with oxaliplatin resistance in colon cancer cells. (A) A volcano plot illustrates genes with altered expression levels in RKO cells treated with 50% bacterial culture filtrate compared to those in the RPMI group. The X‐axis represents log 2 fold‐change, while the Y‐axis indicates –log 10 ( p ‐value). Genes with a fold‐change exceeding 2 and a log worth surpassing 1.3 at a significance level of 0.05 were identified as candidate genes. Pseudogenes are included in the plot. (B) The MTT assay was conducted using siRNAs for the eight selected genes. Oxaliplatin (L‐OHP) was introduced 24‐h post‐transfection at a final concentration of 5 μM. Forty‐eight hours after oxaliplatin exposure, cell viabilities were assessed via the MTT assay. The vertical axis depicts the ratio of (cell viability following oxaliplatin treatment) to (cell viability after saline treatment) * p < 0.05. (C) Cell viability for siRNA‐transfected cells was determined using the MTT assay, with samples from NC siRNA, NINJ2 si#1, NINJ2 si#2, and NINJ2 si#3, respectively. The vertical axis shows the ratio of (cell viability post‐oxaliplatin treatment) to (cell viability post‐saline treatment) * p < 0.05. (D) RKO cells were transfected with NC siRNA, NINJ2 si#1, NINJ2 si#2, and NINJ2 si#3, respectively. Following transfection, RNA was extracted from each set of cells. To evaluate mRNA expression levels, qPCR was then employed. The vertical axis represents the mRNA expression level in cells transfected with NINJ2 si#1, NINJ2 si#2, or NINJ2 si#3 relative to the levels observed in NC siRNA‐transfected cells. (E) RNA was extracted from RKO, DLD1, HT29, and CO115 cells. The vertical axis represents the mRNA expression level in each cell line relative to that in RKO.

Article Snippet: Scramble shRNA (Control shRNA Lentiviral Particles‐A sc‐108080) and NINJ2 shRNA ( Ninjurin‐2 shRNA(h) Lentiviral Particles SC‐75917‐V) were procured from Santa Cruz (CA, USA).

Techniques: Expressing, MTT Assay, Transfection, Concentration Assay, Saline

Journal: Nature Communications

Article Title: Pharmacological and physiological activation of TGR5 in the NTS lowers food intake by enhancing leptin-STAT3 signaling

doi: 10.1038/s41467-025-60331-1

Figure Lengend Snippet: a Cumulative food intake of NTS CMC (n = 15; data replotted from Fig. ), leptin (n = 10; data replotted from Fig. ), CCDC (n = 10; data replotted from Fig. ) or CCDC+leptin (n = 11) HF rats 24 h after food was given back. b Representative western blot images of p-STAT3, t-STAT3, and GAPDH protein levels in NTS CCDC 5 min acute infusion, followed by a 3 h wait, then NTS CMC or leptin 5 min acute infusion, and a subsequent 30 min waiting time in HF rats. c Representative images (scale bar: 100 μm) of immunohistological staining for p-STAT3 with DAPI (top) and cFOS (bottom) in NTS CCDC 5 min acute infusion, followed by a 3 h wait, then NTS CMC or leptin 5 min acute infusion, and a subsequent 30 min waiting time in HF rats. d Relative NTS Tgr5 mRNA expression of NTS shMM (n = 6) or shTgr5 (n = 6) rats. e Relative NTS PKA activity of NTS CMC shMM (n = 6), CCDC shMM (n = 6), CMC shTgr5 (n = 6), or CCDC shTgr5 (n = 6) HF rats, measured 3 h after 5 min acute infusions. f Cumulative food intake of NTS CMC shMM (n = 5), CCDC+leptin shMM (n = 10), CMC shTgr5 (n = 5), or CCDC+leptin shTgr5 (n = 8) HF rats 24 h after food was given back. g Relative NTS LepRb mRNA expression of NTS shMM (n = 6) or shLepR (n = 6) rats. h Cumulative food intake of NTS CMC shMM (n = 7), leptin shMM (n = 7), CMC shLepR (n = 7), leptin shLepR (n = 7) chow rats 24 h after food was given back. i Representative western blot images of p-STAT3, t-STAT3, and GAPDH protein levels in the NTS of NTS shMM or shLepR HF rats receiving a NTS CCDC 5 min acute infusion, followed by a 3 h wait, then NTS leptin 5 min acute infusion, and a subsequent 30 min waiting time. j Cumulative food intake of NTS CCDC+leptin shMM (n = 8) or CCDC+leptin shLepR (n = 8) HF rats 24 h after food was given back. k Cumulative food intake of NTS CMC (n = 15, data replotted from Fig. ), STAT3-PI (n = 7), CCDC + STAT3-PI (n = 11), CCDC+leptin+STAT3-PI (n = 6), or CCDC+leptin (n = 11, data replotted from Fig. 2a) HF rats 24 h after food was given back. a , e , f , h , k P values were calculated by one-way ANOVA with Tukey’s multiple comparison. d , g , j P values were calculated by two-tailed t tests. Data presented as mean ± SEM.

Article Snippet: For the NTS lentiviral infection, after the NTS-targeted cannulation surgery and while the rats were still anaesthetized, lentivirus (1.0 × 10 6 IFU) expressing shTGR5 (sc-270404-V), shLepR (sc-270697-V, Santa Cruz Biotechnology Inc.), or shMM (sc-108080, Santa Cruz Biotechnology Inc.) was injected via the NTS bilateral cannula (3 μl/site over 10 min) .

Techniques: Western Blot, Staining, Expressing, Activity Assay, Comparison, Two Tailed Test

Journal: Nature Communications

Article Title: Pharmacological and physiological activation of TGR5 in the NTS lowers food intake by enhancing leptin-STAT3 signaling

doi: 10.1038/s41467-025-60331-1

Figure Lengend Snippet: a Total bile acid levels in the plasma of chow rats during fasting (n = 5) or at 0.5 h after refed (n = 5). b Unconjugated bile acid levels in the plasma of chow rats during fasting (n = 5) or at 0.5 h after refed (n = 5). c Total bile acid levels in the plasma of HF rats during fasting (n = 6) or at 0.5 h after refed (n = 5). d Unconjugated bile acid levels in the plasma of HF rats during fasting (n = 6) or at 0.5 h after refed (n = 5). e DCA levels in the plasma of chow and HF rats during fasting (chow n = 5; HF n = 6) or at 0.5 h after refed (chow n = 5; HF n = 5). f Unconjugated bile acid levels in the NTS (pmol/g of tissue) of chow rats at 0.5 h (n = 6) or 2 h (n = 5) after refed. g Total bile acid levels in the NTS (pmol/g of tissue) of chow rats at 0.5 h (n = 6) or 2 h (n = 5) after refed. h Unconjugated bile acid levels in the NTS (pmol/g of tissue) of HF rats at 0.5 h (n = 5) or 2 h (n = 6) after refed. i Total bile acid levels in the NTS (pmol/g of tissue) of HF rats at 0.5 h (n = 5) or 2 h (n = 6) after refed. j DCA levels in the NTS (pmol/g of tissue) of chow and HF rats at 0.5 h (chow n = 6; HF n = 5) or 2 h (chow n = 5; HF n = 6) after refed. k Cumulative food intake of NTS saline shMM (n = 8), DCA shMM (n = 8), saline shTgr5 (n = 7), DCA shTgr5 (n = 8) HF rats 24 h after food was given back. l Representative western blot images of p-STAT3, t-STAT3, and GAPDH protein levels in the NTS of HF rats receiving a NTS DCA 5 min acute infusion, followed by a 3 h wait, then NTS CMC or leptin 5 min acute infusion, and a subsequent 30 min waiting time. m Illustration of TGR5 and leptin-STAT3 signaling in feeding regulation and associated experimental approaches. a–d , f–i P values were calculated by two-tailed t tests; e , j , k P values were calculated by one-way ANOVA with Tukey’s multiple comparison. Data presented as mean ± SEM. DCA deoxycholic acid, HDCA hyodeoxycholic acid, CDCA chenodeoxycholic acid, CA cholic acid, UDCA ursodeoxycholic acid, α-MCA α-muricholic acid, β-MCA β-muricholic acid.

Article Snippet: For the NTS lentiviral infection, after the NTS-targeted cannulation surgery and while the rats were still anaesthetized, lentivirus (1.0 × 10 6 IFU) expressing shTGR5 (sc-270404-V), shLepR (sc-270697-V, Santa Cruz Biotechnology Inc.), or shMM (sc-108080, Santa Cruz Biotechnology Inc.) was injected via the NTS bilateral cannula (3 μl/site over 10 min) .

Techniques: Clinical Proteomics, Saline, Western Blot, Two Tailed Test, Comparison

Journal: Cancer medicine

Article Title: FUT8 Is a Critical Driver of Prostate Tumour Growth and Can Be Targeted Using Fucosylation Inhibitors.

doi: 10.1002/cam4.70959

Figure Lengend Snippet: FIGURE 2 | FUT8 protein levels are increased in the blood of patients with aggressive prostate cancer. (A-D) Detection of FUT8 protein in blood samples from patients with prostate cancer using sandwich ELISA assays. (A) FUT8 levels were 3.02-fold higher in plasma samples from patients with prostate cancer compared to patients given a no-cancer diagnosis (n = 27, unpaired t test, p = 0.109). (B) The levels of FUT8 protein were 2.09- fold higher in plasma samples from men with prostate cancer compared to men diagnosed with BPH (n = 319, unpaired t test, p = 0.0457, *). (C) FUT8 levels were 1.86-fold increased in serum samples from patients with high grade prostate cancer (Gleason grade 8–9) compared to patients with low grade prostate cancer (Gleason grade 6–7) (n = 200, unpaired t test, p < 0.0218, *). (D) Analysis of FUT8 levels in matched serum samples from 7 men with prostate cancer taken before and after ADT. FUT8 serum levels significantly increase after ADT (n = 14, paired t test, p = 0.047, *).

Article Snippet: For FUT8 knockdown, shRNA lentiviral particles were purchased from Santa Cruz (FUT8 shRNA sc- 45757- V and Control shRNA sc- 108080).

Techniques: Sandwich ELISA, Clinical Proteomics, Biomarker Discovery

Journal: Cancer medicine

Article Title: FUT8 Is a Critical Driver of Prostate Tumour Growth and Can Be Targeted Using Fucosylation Inhibitors.

doi: 10.1002/cam4.70959

Figure Lengend Snippet: FIGURE 3 | Upregulation of FUT8 in prostate cancer cells promotes tumour growth, migration and invasion. (A) Upregulation of FUT8 in CWR22Rv1 cells increases the growth of subcutaneous xenograft tumours. 1 × 107 cells were injected into the flank of CD-1 nude mice. Tumour size was measured every 3–4 days using callipers. Over 15 days the CWR22V1 tumours with overexpression of FUT8 were 2.23 folf bigger (n = 16, un- paired t test, p = 0.1993). Representative tumour images from each group are shown. (B) Knockdown of FUT8 using shRNA significantly reduces the growth of PC3 tumours in a subcutaneous xenograft model. 3 × 106 PC3 cells were injected into the flank of NMRI mice. Tumour size was measured every 3–4 days using callipers. Over 40 days, the growth of PC3 tumours with knockdown of FUT8 was significantly reduced (n = 12, unpaired t test, p = 0.0055, **). (C-F) Upregulation of FUT8 in CWR22Rv1 cells promotes cell migration (unpaired t text, p = 0.0092, **) and invasion (unpaired t test, p = 0.0156, *). Knockdown of FUT8 in PC3 cells decreases prostate cancer cell migration (unpaired t test, p = 0.0102, *) and invasion (unpaired t test, p = 0.0113, *). Scale bar is 20 μm.

Article Snippet: For FUT8 knockdown, shRNA lentiviral particles were purchased from Santa Cruz (FUT8 shRNA sc- 45757- V and Control shRNA sc- 108080).

Techniques: Migration, Injection, Over Expression, Knockdown, shRNA

Journal: Cancer medicine

Article Title: FUT8 Is a Critical Driver of Prostate Tumour Growth and Can Be Targeted Using Fucosylation Inhibitors.

doi: 10.1002/cam4.70959

Figure Lengend Snippet: FIGURE 5 | FUT8 regulates oncogenic genes and proteins in prostate cancer cells. RNA-sequencing analysis of CWR22Rv1 cells with overex- pression of FUT8 identified 381 differentially expressed genes (adjusted p-value < 0.05, Log2FC 0.58) (Table S2). (A) Heatmap to illustrate the top 10 upregulated and 10 ten downregulated differentially expressed genes. (B, C) Gene Ontology and gene set enrichment analyses of genes regulated by FUT8 revealed CWR22Rv1 cells overexpressing FUT8 have enrichment in ‘ossification’, ‘bone mineralisation’ and ‘regulation of osteoblast dif- ferentiation’. (D–F) Validation at the protein level using immunocytochemistry shows (D) IGFBP5 is upregulated when FUT8 is overexpressed in CWR22Rv1 cells and (E, F) Knockdown of FUT8 downregulates IL1B and PTGES3 in PC3 cells. Scale bar is 20 μm. (G) Analysis of the TCGA PRAD cohort shows a significant correlation between the FUT8 gene and levels of IGFBP5, IL1B and PTGES3 in clinical prostate cancer tissue.

Article Snippet: For FUT8 knockdown, shRNA lentiviral particles were purchased from Santa Cruz (FUT8 shRNA sc- 45757- V and Control shRNA sc- 108080).

Techniques: RNA Sequencing, Biomarker Discovery, Immunocytochemistry, Knockdown