control shrna shctrl lentiviral particles  (Santa Cruz Biotechnology)

Journal: BMC Cancer

Article Title: NSDHL contributes to breast cancer stem-like cell maintenance and tumor-initiating capacity through TGF-β/Smad signaling pathway in MCF-7 tumor spheroid

doi: 10.1186/s12885-024-13143-3

Figure Lengend Snippet: NSDHL knockdown suppresses tumor initiation and growth in tumor spheroid-injected xenograft tumor models. (A) Representative flow cytometry histogram of mCherry-positive cells sorted from NSDHL shRNA (shNSDHL) or control shRNA (shCtrl)-tagged mCherry-transduced MCF-7 cells. (B) Representative confocal images of mCherry-positive cells in shNSDHL- or shCtrl-spheroids. mCherry (red) and DAPI (blue). Scale bar: 100 μm C-E. NSDHL mRNA and protein levels and representative western blot images of NSDHL and mCherry in shNSDHL-MCF-7 spheroids relative to shCtrl-MCF-7 spheroids by qRT-PCR ( n = 6) and western blot analysis ( n = 6). F-G. Representative flow cytometry dot plots and quantification of CD44+/CD24 − cells in shCtrl and shNSDHL MCF-7 spheroids ( n = 3). H-I. Representative flow cytometry dot plots and quantification of ALDH + cells in shCtrl- and shNSDHL-MCF-7 spheroids using ALDEFLUOR assay ( n = 3). J. Tumor volume was measured weekly in shCtrl- or shNSDHL-MCF-7 spheroid-injected mice ( n = 6) for 56 days post-injection. K-L. Gross images and wet weights of tumors removed from shNSDHL- or shCtrl-MCF-7 spheroid-injected mice ( n = 6) at 56 days post-injection. M-N. Representative western blot images and quantification data of NSDHL proteins in shNSDHL-MCF-7 tumors compared to shCtrl-MCF-7 tumors ( n = 6). O. Representative NSDHL and mCherry immunohistochemistry images and H&E images of shNSDHL- or shCtrl-MCF-7 tumor tissues. Scale bar: 100 μm. The bar graph shows the mean ± standard deviation. * P < 0.05, ** P < 0.01, *** P < 0.001 as compared to shCtrl using unpaired t-tests

Article Snippet: shRNA lentiviral particles (shNSDHL) (LPP-HSH103352-LVRU6MH-c-100) and control shRNA lentiviral particles (shCtrl) (LPP-CSHCTR001-LVRU6MH-025-C) were purchased from GeneCopoeia (Rockville, MD, USA).

Techniques: Knockdown, Injection, Flow Cytometry, shRNA, Control, Western Blot, Quantitative RT-PCR, Immunohistochemistry, Standard Deviation

Journal: BMC Cancer

Article Title: NSDHL contributes to breast cancer stem-like cell maintenance and tumor-initiating capacity through TGF-β/Smad signaling pathway in MCF-7 tumor spheroid

doi: 10.1186/s12885-024-13143-3

Figure Lengend Snippet: Incidence of palpable tumors in mice injected with shCtrl or shNSDHL cells

Article Snippet: shRNA lentiviral particles (shNSDHL) (LPP-HSH103352-LVRU6MH-c-100) and control shRNA lentiviral particles (shCtrl) (LPP-CSHCTR001-LVRU6MH-025-C) were purchased from GeneCopoeia (Rockville, MD, USA).

Techniques: Injection

Journal: BMC Cancer

Article Title: NSDHL contributes to breast cancer stem-like cell maintenance and tumor-initiating capacity through TGF-β/Smad signaling pathway in MCF-7 tumor spheroid

doi: 10.1186/s12885-024-13143-3

Figure Lengend Snippet: NSDHL knockdown reduced the BCSC population with CD44+/CD24 − phenotype and ALDH activity in tumor spheroid-injected xenograft tumor tissues, accompanied by decreased Smad2/3 phosphorylation and SOX2 expression. A-D. Representative flow cytometry histograms, dot plots, and quantification of CD44 + and CD24- cells in control (shCtrl) and NSDHL shRNA (shNSDHL)-transduced MCF-7 tumors ( n = 4). E-F. Representative flow cytometry dot plots and quantification of ALDH + cells in shCtrl- and shNSDHL MCF-7 tumors using ALDEFLUOR assay ( n = 4). G. Representative double immunohistochemistry images of CD44 + and ALDH1A1 + cells in shCtrl and shNSDHL MCF-7 tumors. CD44 (green), ALDH1A1(red). Double CD44+/ALDH1A1 + cells (arrow). H-O. Representative western blot images and quantification data of phopho-Smad2 and 3, Smad2, Smad3, SOX2, and NANOG levels in shNSDHL MCF-7 tumors relative to shCtrl MCF-7 tumors ( n = 5). All graphs show the mean ± standard deviation. * P < 0.05, compared with shCtrl using unpaired t-tests

Article Snippet: shRNA lentiviral particles (shNSDHL) (LPP-HSH103352-LVRU6MH-c-100) and control shRNA lentiviral particles (shCtrl) (LPP-CSHCTR001-LVRU6MH-025-C) were purchased from GeneCopoeia (Rockville, MD, USA).

Techniques: Knockdown, Activity Assay, Injection, Expressing, Flow Cytometry, Control, shRNA, Immunohistochemistry, Western Blot, Standard Deviation

Journal: BMC cancer

Article Title: NAD(P)-dependent steroid dehydrogenase-like is involved in breast cancer cell growth and metastasis.

doi: 10.1186/s12885-020-06840-2

Figure Lengend Snippet: Fig. 2 NSDHL knockdown decreases viability, proliferation, and colony and sphere formation abilities of BT-20 and MDA-MB-231 cells. a, b Data of cell viability and cell cycle in BT-20 and MDA-MB-231 cells transfected with NSDHL siRNA or control siRNA (20 nM); c, d Representative images and data analyzed in colony formation of BT-20 and MDA-MB-231 cells transfected with NSDHL siRNA or control siRNA (20 nM); e Representative image and data analyzed in 3D sphere formation of BT-20 and MDA-MB-231 cells transfected with NSDHL siRNA or control siRNA (20 nM). All data represent the means ± standard deviations of three independent experiments, each performed in triplicates. *p < 0.05, **p < 0.01, ***p < 0.001

Article Snippet: Short hairpin RNA (shRNA) lentiviral transduction Precisely, 4 × 105 MDA-MB-231 cells were seeded in each well of 12-well plate 24 h prior to viral infection and were replaced with media containing 5 μg/ml polybrene® (sc-134220, Santa Cruz) and 20 μl of NSDHLtargeting shRNA lentiviral particles (shNSDHL) (sc90849-V, Santa Cruz) or control shRNA lentiviral particles (shCtrl) (sc-108080, Santa Cruz).

Techniques: Knockdown, Transfection, Control

Journal: BMC cancer

Article Title: NAD(P)-dependent steroid dehydrogenase-like is involved in breast cancer cell growth and metastasis.

doi: 10.1186/s12885-020-06840-2

Figure Lengend Snippet: Fig. 3 NSDHL knockdown decreases the migration and invasion abilities of BT-20 and MDA-MB-231 cells. a,b,c Representative images and data analyzed in transwell migration assay, invasion assay and, wound healing assay of BT-20 and MDA-MB-231 cells transfected with NSDHL siRNA or control siRNA (20 nM). Data represent the means ± standard deviations of three independent experiments. *p < 0.05, **p < 0.01, ***p < 0.001

Article Snippet: Short hairpin RNA (shRNA) lentiviral transduction Precisely, 4 × 105 MDA-MB-231 cells were seeded in each well of 12-well plate 24 h prior to viral infection and were replaced with media containing 5 μg/ml polybrene® (sc-134220, Santa Cruz) and 20 μl of NSDHLtargeting shRNA lentiviral particles (shNSDHL) (sc90849-V, Santa Cruz) or control shRNA lentiviral particles (shCtrl) (sc-108080, Santa Cruz).

Techniques: Knockdown, Migration, Transwell Migration Assay, Invasion Assay, Wound Healing Assay, Transfection, Control

Journal: BMC cancer

Article Title: NAD(P)-dependent steroid dehydrogenase-like is involved in breast cancer cell growth and metastasis.

doi: 10.1186/s12885-020-06840-2

Figure Lengend Snippet: Fig. 4 NSDHL knockdown suppresses total cholesterol level and promotes erlotinib response in MDA-MB-231 cell. a Total cholesterol levels measured in BT-20 and MDA-MB-231 cells transfected with NSDHL siRNA or control siRNA (20 nM); b Dose-response curve of erlotinib in BT-20 and MDA-MB-231 cells transfected with NSDHL siRNA or control siRNA (20 nM); c Representative western blot images of NSDHL, EGFR, and precursor and mature SREBP-1 and data of relative expression levels of NSDHL, EGFR, and precursor SREBP-1 in BT-20 and MDA-MB-231 cells transfected with NSDHL siRNA or control siRNA (20 nM). Data represent the mean ± standard deviation of three independent experiments. *p < 0.05, ***p < 0.001

Article Snippet: Short hairpin RNA (shRNA) lentiviral transduction Precisely, 4 × 105 MDA-MB-231 cells were seeded in each well of 12-well plate 24 h prior to viral infection and were replaced with media containing 5 μg/ml polybrene® (sc-134220, Santa Cruz) and 20 μl of NSDHLtargeting shRNA lentiviral particles (shNSDHL) (sc90849-V, Santa Cruz) or control shRNA lentiviral particles (shCtrl) (sc-108080, Santa Cruz).

Techniques: Knockdown, Transfection, Control, Western Blot, Expressing, Standard Deviation

Journal: BMC cancer

Article Title: NAD(P)-dependent steroid dehydrogenase-like is involved in breast cancer cell growth and metastasis.

doi: 10.1186/s12885-020-06840-2

Figure Lengend Snippet: Fig. 5 NSDHL knockdown suppressed tumor growth and lung metastasis of MDA-MB-231 xenograft mice. a, b Data of relative NSDHL mRNA level analyzed by real-time RT-PCR and representative western blot images of NSDHL in MDA-MB-231 cells transduced with NSDHL shRNA or control shRNA lentivirus; c Data of tumor volume measured weekly in NSDHL shRNA or control shRNA mice; d Gross images and wet weight of tumors removed from NSDHL shRNA or control shRNA mice at 44 days post-injection; e Representative NSDHL immunohistochemistry image and scores analyzed from NSDHL shRNA or control shRNA tumor tissues; f Gross and H&E images of lungs and data of metastatic foci analyzed from NSDHL shRNA or control shRNA lung tissues. In vitro data represent the means ± standard deviations of three independent experiments. Animal data represent the means ± standard deviations of five mice per group. *p < 0.05, **p < 0.01

Article Snippet: Short hairpin RNA (shRNA) lentiviral transduction Precisely, 4 × 105 MDA-MB-231 cells were seeded in each well of 12-well plate 24 h prior to viral infection and were replaced with media containing 5 μg/ml polybrene® (sc-134220, Santa Cruz) and 20 μl of NSDHLtargeting shRNA lentiviral particles (shNSDHL) (sc90849-V, Santa Cruz) or control shRNA lentiviral particles (shCtrl) (sc-108080, Santa Cruz).

Techniques: Knockdown, Quantitative RT-PCR, Western Blot, Transduction, shRNA, Control, Injection, Immunohistochemistry, In Vitro