Journal: Thoracic Cancer

Article Title: microRNA ‐100 functions as a tumor suppressor in non‐small cell lung cancer via regulating epithelial‐mesenchymal transition and Wnt/β‐catenin by targeting HOXA1

doi: 10.1111/1759-7714.13459

Figure Lengend Snippet: miR‐100 inhibited NSCLC cell proliferation. ( a ) qRT‐PCR was used to measure miR‐100 expressions in NSCLC cells. ( b , c ) miR‐100 overexpression or inhibition was confirmed by qRT‐PCR. ( d , e ) MTT assays were performed to detect the functions of miR‐100 in NSCLC cell proliferation. * P < 0.05, ** P < 0.01, *** P < 0.001 (d: NC and miR‐100 mimics; e: NC and miR‐100 inhibitor).

Article Snippet: Human normal bronchial epithelium cell BEAS‐2B and NSCLC cells (NCI‐H460, NCI‐H1299, SPC‐A1 and A549) were purchased from ATCC and maintained in DMEM (Gibco; Thermo Fisher Scientifc, Inc., Waltham, MA, USA) which contained 10% FBS (Gibco; Thermo Fisher Scientifc, Inc.) in a humidified atmosphere with 5% CO 2 at 37°C.

Techniques: Quantitative RT-PCR, Over Expression, Inhibition

Journal: Thoracic Cancer

Article Title: microRNA ‐100 functions as a tumor suppressor in non‐small cell lung cancer via regulating epithelial‐mesenchymal transition and Wnt/β‐catenin by targeting HOXA1

doi: 10.1111/1759-7714.13459

Figure Lengend Snippet: miR‐100 suppressed the cell invasion and migration abilities of NSCLC cells. ( a , b ) The impacts of miR‐100 restoration on NSCLC cell invasion and migration were determined using transwell assays. ( c , d ) Transwell assay was performed to determine invasion and migration capacities of miR‐100 suppressed NSCLC cells. ** P < 0.01, *** P < 0.001.

Article Snippet: Human normal bronchial epithelium cell BEAS‐2B and NSCLC cells (NCI‐H460, NCI‐H1299, SPC‐A1 and A549) were purchased from ATCC and maintained in DMEM (Gibco; Thermo Fisher Scientifc, Inc., Waltham, MA, USA) which contained 10% FBS (Gibco; Thermo Fisher Scientifc, Inc.) in a humidified atmosphere with 5% CO 2 at 37°C.

Techniques: Migration, Transwell Assay

Journal: Thoracic Cancer

Article Title: microRNA ‐100 functions as a tumor suppressor in non‐small cell lung cancer via regulating epithelial‐mesenchymal transition and Wnt/β‐catenin by targeting HOXA1

doi: 10.1111/1759-7714.13459

Figure Lengend Snippet: HOXA1 was a direct target of miR‐100 in NSCLC cells. ( a )The WT and MUT binding sites of miR‐100 on HOXA1 3′‐UTR. ( b ) Dual‐luciferase reporter assay was used to confirm the association between miR‐100 and HOXA1 in NSCLC cells ( NC and miR‐100 mimics). ( c , d ) Regulatory effects of miR‐100 on HOXA1 expressions in NSCLC cells. * P < 0.05, ** P < 0.01.

Article Snippet: Human normal bronchial epithelium cell BEAS‐2B and NSCLC cells (NCI‐H460, NCI‐H1299, SPC‐A1 and A549) were purchased from ATCC and maintained in DMEM (Gibco; Thermo Fisher Scientifc, Inc., Waltham, MA, USA) which contained 10% FBS (Gibco; Thermo Fisher Scientifc, Inc.) in a humidified atmosphere with 5% CO 2 at 37°C.

Techniques: Binding Assay, Luciferase, Reporter Assay

Journal: Biology Open

Article Title: Endoplasmic reticulum stress-induced cellular dysfunction and cell death in insulin-producing cells results in diabetes-like phenotypes in Drosophila

doi: 10.1242/bio.046524

Figure Lengend Snippet: Induction of UPR observed in wing imaginal discs and IPCs with ectopic Hsc70-3 DN expression. (A–B) Expression of Xbp1-GFP generated by ER stress-dependent splicing of xbp1*-GFP mRNA in wing imaginal discs. Phase contrast (A,B) and fluorescence (A′,B′) micrographs of wing imaginal discs. (A,A′) Control wing disc ( Bx>xbp1*-GFP ). (B,B′) Wing disc expressing a dominant-negative form of Hsc70-3 in the wing pouch region (arrow) ( Bx>hsc70-3 DN , xbp1*-GFP ). (C–E) Fluorescence micrograph of wing discs stained with DAPI (white). (C′–E′) Immunostaining of the wing discs with an anti-GRP78 antibody. (D″) Immunostaining of the wing disc with anti-HA antibody. (C,C′) Fluorescence micrograph of a control wing imaginal disc ( Bx-Gal4/+ ). (D–D″) Wing imaginal disc expressing control Hsc70-3 in the wing pouch region of the imaginal disc ( Bx>hsc70-3 ). (E,E′) Wing imaginal disc expressing a dominant-negative form of Hsc70-3 in the same region ( Bx>hsc70-3 DN ). Anti-GRP78 immunostaining is shown in white. Note that more intense immunofluorescence was observed exclusively in areas expressing Hsc70-3 DN , but not the control protein. (A–F) Relative intensity of anti-GRP78 immunostaining in wing imaginal discs. Immunofluorescence signal intensity in each wing imaginal disc with the control Hsc70-3 ( n =31) or Hsc70-3 DN ( n =25) expression was calculated and normalized to the control value, which was set as 1.0 ( Bx-Gal4/+ ) ( n =25; n.s., not significant, P >0.05; *** P <0.001, Student's t -tests). Error bars represent s.e.m. (G–I) Anti-GRP78 immunostaining of IPCs expressing GFPnls in brains from third-instar larvae. (G) Control IPCs ( ilp2>GFPnls ), (H) IPCs expressing the control Hsc70-3 ( ilp2>hsc70-3, GFPnls ), (I) IPCs expressing Hsc70-3 DN ( ilp2>hsc70-3 DN , GFPnls ). Anti-GRP78 immunostaining is colored in red (G–I; white in G′–I′). Nuclei of IPCs visualized by GFPnls expression are colored green (G–I; white in G″–I″). Arrows in H′ and H″ indicate positions of IPC cells. Note that remarkably higher immunostaining signal was observed in IPCs expressing Hsc70-3 DN , but not the control protein. (J) Relative intensities of anti-GRP78 immunostaining in larval IPCs. Immunofluorescence signal intensities in each IPC expressing Hsc70-3 ( n =25) or Hsc70-3 DN ( n =21) were calculated and normalized to the control value of 1.0 ( ilp2>GFPnls ) ( n =21, * P <0.05, *** P <0.001, Student's t -test). Error bars represent s.e.m. Scale bars: (A–E) 100 µm, (G–I) 50 µm.

Article Snippet: The following primary antibodies were used at the dilution described; rabbit anti-β-galactosidase (MP Biomedicals, #55976) at 1:1000, rabbit anti-GRP78 (Bip) (StressMarq Biosciences Inc., Cadboro Bay, Victoria, Canada) that could recognize Hsp70 family proteins including Hsc70-3 in Drosophila at 1:500, rabbit Cleaved Caspase-3 (Asp175) (#9661, Cell Signaling, Danvers, Massachusetts, USA) at 1:200 for larval brain immunostaining and at 1:150 for wing disc immunostaining, rabbit anti-Cleaved Drosophila Dcp-1 (Asp216) (Cell Signaling, antibody #9578) at 1:500, and rabbit anti-phospho-SAPK/JNK (pThr183, pTyr185) (Calbiochem, La Jolla, CA, USA) at 1:200.

Techniques: Expressing, Generated, Fluorescence, Dominant Negative Mutation, Staining, Immunostaining, Immunofluorescence

Journal: PLoS ONE

Article Title: The Proprotein Convertase Furin Contributes to Rhabdomyosarcoma Malignancy by Promoting Vascularization, Migration and Invasion

doi: 10.1371/journal.pone.0161396

Figure Lengend Snippet: A) mRNA levels of all nine proprotein convertases (PCs) were determined by qRT-PCR in 5 Ewing sarcoma, 8 osteosarcoma and 20 rhabdomyosarcoma (RMS) cell lines. Shown are levels relative to GAPDH expression. B) Protein levels of proform and mature furin were assessed by immunoblotting in 20 different RMS cell lines. C) Endogenous furin activity of selected RMS cell lines: Rh36, RD (eRMS), Rh3, Rh4, Rh30, Rh41 and RMS13 (aRMS). Furin activity deficient cells LoVo cells serve as negative control. Furin was captured from cell lysates on anti-furin antibody coated plates and furin activity was measured by addition of the fluorogenic substrate Boc-RVRR-AMC after 6h. Displayed are values normalized by background subtraction.

Article Snippet: Black FluoroNunc 96 well plates (MaxiSorp surface, Nunc, Thermo Scientific) were coated with goat anti human furin antibody (AF1503, R&D Systems) at 10 μg/ml in 50 mM Na 2 CO 3 , pH 9.6 (50 μl/ well) for 8h at RT, protected from light.

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

Journal: Cells

Article Title: Allele-Specific Epigenetic Regulation of FURIN Expression at a Coronary Artery Disease Susceptibility Locus

doi: 10.3390/cells12131681

Figure Lengend Snippet: MeCP2 knockdown increases monocyte proliferation and migration, and these effects are attenuated by a FURIN inhibitor. Results of migration ( A ), proliferation ( B ), and apoptosis ( C ) assays of THP1 monocytic cells transfected with either a MeCP2 siRNA or negative control siRNA for 48 h, with or without treatment with the FURIN inhibitor decanoyl-RVKR-CMK for 24 h. Data shown are mean (±standard deviation) values and p values from Mann-Whitney test; n = 5 ( A , C ) or n = 7 ( B ) in each group.

Article Snippet: In some assays, cells were incubated with the FURIN inhibitor decanoyl-RVKR-CMK (GLPBIO; GC15108) at a final concentration of 10 µM for 24 h. Cells were then subjected to a proliferation assay with the Cell Counting Kit-8 (MedChemExpress, HY-K0301).

Techniques: Migration, Transfection, Negative Control, Standard Deviation, MANN-WHITNEY

Journal: PLoS ONE

Article Title: Engineering, and production of functionally active human Furin in N . benthamiana plant: In vivo post-translational processing of target proteins by Furin in plants

doi: 10.1371/journal.pone.0213438

Figure Lengend Snippet: ( A): Western blot analysis of human furin, produced in N . benthamiana plants . N . benthamiana leaf samples were harvested at 6 dpi. Samples for western blot analysis were prepared as described in Materials and Methods. Proteins on the blot were probed with a purified mouse anti-His tag antibody. 1- crude extract from non-infiltrated N . benthamiana ; 2- crude extract from N . benthamiana plant infiltrated with pEAQ-Furin (truncated)-His-KDEL construct. M: MagicMark XP Western Protein Standard. ( B): SDS-PAGE analysis of Ni-NTA column purified plant produced recombinant human furin. 5 μg Ni-NTA column purified protein was loaded into well. ( C ): Western blot analysis of different dilutions of Ni-NTA column purified, plant produced recombinant human furin, along with protein standards. Partially purified, plant produced furin was diluted 2.5, 5, 10 and 25-fold and different amount of plant produced furin, as indicated, was run on SDS-PAGE, followed by western blot. Plant produced His tagged furin protein band was detected using a purified mouse anti-His tag antibody. The concentration of furin in Ni-NTA column purified samples was quantified using the gene tools software, Syngene Bioimaging. Plant produced Endo H deglycosylated, purified PA83 protein (dPA83) was used as a protein standard. M1: color prestained protein standard (NEB); M2: MagicMark XP Western Protein Standard (ThermoFisher Scientific). ( D, E ): schematic representation of the full length (D) and truncated furin (E) structures. SP- Signal peptide; PP- Propeptide; SD- Subtilisin-like catalytic domain; BD- Homo B domain; CD- Cysteine rich domain; TM- Transmembrane domain; CD- Cytoplasmic domain.

Article Snippet: To compare the in vitro activity of plant produced furin with commercial human furin, 5 μg plant produced, Ni-NTA column purified dPA83 was incubated with various concentrations (0, 1, 5, 25, 50, 100 ng) of commercial human furin (New England Biolabs, NEB), in 20 mM HEPES, 0.1% Triton X-100, 0.2 mM CaCl 2 , pH 7.5, at 25°C for 2 hours.

Techniques: Western Blot, Produced, Purification, Construct, SDS Page, Recombinant, Concentration Assay, Software

Journal: PLoS ONE

Article Title: Engineering, and production of functionally active human Furin in N . benthamiana plant: In vivo post-translational processing of target proteins by Furin in plants

doi: 10.1371/journal.pone.0213438

Figure Lengend Snippet: 5 μg recombinant APRIL protein was incubated with 25 ng of plant produced human furin or 25 ng of commercial human furin (NEB) at 25°C, for 2 h. 2.5 μg APRIL protein from each sample was loaded in each well. M: color prestained protein standard (NEB).

Article Snippet: To compare the in vitro activity of plant produced furin with commercial human furin, 5 μg plant produced, Ni-NTA column purified dPA83 was incubated with various concentrations (0, 1, 5, 25, 50, 100 ng) of commercial human furin (New England Biolabs, NEB), in 20 mM HEPES, 0.1% Triton X-100, 0.2 mM CaCl 2 , pH 7.5, at 25°C for 2 hours.

Techniques: Recombinant, Incubation, Produced

Journal: PLoS ONE

Article Title: Engineering, and production of functionally active human Furin in N . benthamiana plant: In vivo post-translational processing of target proteins by Furin in plants

doi: 10.1371/journal.pone.0213438

Figure Lengend Snippet: SDS-PAGE CBB (A, C) and Western blot (B) analysis of dPA83, cleaved with plant produced or commercial human Furin. (A) : 5 μg dPA83 (deglycosylated PA83) samples were treated with different concentrations (0, 1, 5, 20, 25, 50 and 100 ng) of plant produced human furin and then 4.5 μg samples were run on SDS-PAGE. dPA83: deglycosylated PA83; pp hFurin: plant produced, Ni-NTA column purified furin. M1: color prestained protein standard (NEB). ( B) : 5 μg of deglycosylated PA83 samples were treated with different concentrations of plant produced human furin or 50 ng commercial (NEB) human furin as indicated, and then 100 ng PA samples were loaded into the gel. dPA83: deglycosylated PA83; pp hFurin: plant produced, Ni-NTA column purified furin. M2: MagicMark XP Western Protein Standard. (C) : 5 μg dPA83 (deglycosylated PA83) samples were treated with different concentrations (0, 1, 5, 20, 25, 50 and 100 ng) of commercial human furin (NEB) as indicated, and then 4.5 μg samples were run on SDS-PAGE. ( D) : Schematic representation of PA83 protein structure. PA63 and PA20 (a 20-kDa amino-terminal fragment) are cleavage products of PA83 by furin. M1: color prestained protein standard (NEB).

Article Snippet: To compare the in vitro activity of plant produced furin with commercial human furin, 5 μg plant produced, Ni-NTA column purified dPA83 was incubated with various concentrations (0, 1, 5, 25, 50, 100 ng) of commercial human furin (New England Biolabs, NEB), in 20 mM HEPES, 0.1% Triton X-100, 0.2 mM CaCl 2 , pH 7.5, at 25°C for 2 hours.

Techniques: SDS Page, Western Blot, Produced, Purification

Journal: PLoS ONE

Article Title: Engineering, and production of functionally active human Furin in N . benthamiana plant: In vivo post-translational processing of target proteins by Furin in plants

doi: 10.1371/journal.pone.0213438

Figure Lengend Snippet: ( A ): Western blot analysis of Ni-NTA purified plant produced furin variants, i.e. glycosylated Endo H or PNGase F in vivo deglycosylated, as indicated. gfurin: plant produced furin, expressed alone (glycosylated); dfurin (E): plant produced furin co-expressed with Endo H; dfurin (P): plant produced furin co-expressed with PNGase F. Furin protein bands were detected using the purified anti-His tag antibody. M1: MagicMark XP Western Protein Standard. ( B ) SDS-PAGE CBB: 5.0 μg plant produced dPA83 was incubated with different amount (1.0, 5.0, 25, 50, 100 ng) of furin, which was co-expressed with PNGase F and then 2.5 μg PA83 protein from each sample was loaded into each well. G- positive control, 5.0 μg plant produced dPA83 was incubated with 50 ng of plant produced furin (glycosylated) and then 2.5μg dPA83 was loaded into a well and run on a SDS-PAGE. C-negative control, plant produced dPA83 protein, not incubated with furin. M2-color prestained protein standard (NEB); S- BSA standard. ( C ): 5.0 μg plant produced dPA83was incubated with different amount (1.0, 5.0, 25, 50, 100 ng) of furin co-expressed with Endo H and then 2.5 μg PA83 protein from each sample was loaded into each well. C-negative control, plant produced dPA83 protein, not incubated with furin. M2-color prestained protein standard (NEB); S- BSA standard. (D): 5.0 μg plant produced PA83 (deglycosylated) was incubated with commercial human furin, which was previously in vitro deglycosylated with commercial Endo H (lane 2) or PNGase F (Lane 3). Lane 1, positive control, 5.0 μg plant produced dPA83 was incubated with 50 ng commercial human furin (NEB) and 2.5μg PA83 was loaded into a well. C-negative control, plant produced dPA83 protein, not incubated with commercial furin. M2-color prestained protein standard (NEB); S- BSA standard.

Article Snippet: To compare the in vitro activity of plant produced furin with commercial human furin, 5 μg plant produced, Ni-NTA column purified dPA83 was incubated with various concentrations (0, 1, 5, 25, 50, 100 ng) of commercial human furin (New England Biolabs, NEB), in 20 mM HEPES, 0.1% Triton X-100, 0.2 mM CaCl 2 , pH 7.5, at 25°C for 2 hours.

Techniques: Western Blot, Purification, Produced, In Vivo, SDS Page, Incubation, Positive Control, Negative Control, In Vitro

Journal: PLoS ONE

Article Title: Engineering, and production of functionally active human Furin in N . benthamiana plant: In vivo post-translational processing of target proteins by Furin in plants

doi: 10.1371/journal.pone.0213438

Figure Lengend Snippet: ( A ): Lanes: 1- Co-expression of PA83 with PNGase F for the production of PNGase F deglycosylated PA83 protein; 2- Co-expression of PA83 with Endo H for the production of Endo H deglycosylated PA83 protein; 3-Expression of PA83 ( alone) for the production of glycosylated PA83 protein; 4- Co-expression of PA83 with furin and PNGase F for the production of furin cleaved and PNGase F deglycosylated PA83 protein; 4- Co-expression of PA83 with furin and Endo H for the production of furin cleaved and Endo H deglycosylated PA83 protein; 5- Co-expression of PA83 with furin for the production of furin cleaved and glycosylated PA83 protein. ( B ): WB analysis of N . benthamiana plant, infiltrated with PA83 and Endo H or infiltrated with PA83, Endo H and Furin constructs. 6-7-week-old N . benthamiana plant leaves, were infiltrated with the above constructs, were harvested and samples were processed for SDS-PAGE and western blot, as described in Materials and Methods. Boiled and un-boiled (raw) samples were diluted 5-fold and 10 μl from each sample was run on SDS-PAGE prior to western blotting. Proteins were probed with the purified anti-His tag antibody. The image was taken using a highly sensitive GeneGnome XRQ Chemiluminescence imaging system. An arrow indicates the protein bands corresponding to PA83 and PA63 and the formation of PA63 oligomers. M: MagicMark XP Western Protein Standard.

Article Snippet: To compare the in vitro activity of plant produced furin with commercial human furin, 5 μg plant produced, Ni-NTA column purified dPA83 was incubated with various concentrations (0, 1, 5, 25, 50, 100 ng) of commercial human furin (New England Biolabs, NEB), in 20 mM HEPES, 0.1% Triton X-100, 0.2 mM CaCl 2 , pH 7.5, at 25°C for 2 hours.

Techniques: Expressing, Construct, SDS Page, Western Blot, Purification, Imaging

Journal: Cell reports

Article Title: Activation of endoplasmic reticulum stress in premature aging via the inner nuclear membrane protein SUN2

doi: 10.1016/j.celrep.2023.112534

Figure Lengend Snippet: KEY RESOURCES TABLE

Article Snippet: Primary antibodies used for immunofluorescence were: α-Hsp110 rabbit polyclonal (1:100, StressMarq, SPC-195), α-Hsp90 rat monoclonal (1:200, Enzo, ADI-SPA-835), α-Hsc70 rat monoclonal (1:500, Abcam, ab19136), α-Hsp40 rabbit polyclonal (1:500, StressMarq, SPC-100), α-GRP94 rabbit polyclonal (1:200, Abcam, ab3674), α-GRP78/BiP rabbit polyclonal (1:500, Abcam, ab21685), α-GRP78 mouse monoclonal (1:50, StressMarq, SMC-195), α-PDI P4HB mouse monoclonal (1:500, Abcam, ab2792), α-Calnexin rabbit polyclonal (1:100, Cell Signaling, 2433S), α-SUN1 rabbit polyclonal (1:100, Sigma, HPA008346), α-SUN2 rabbit polyclonal (1:500, Sigma, HPA001209), α -Nesprin 1 rabbit monoclonal (1:100, Abcam, ab192234),α-Nesprin 2 rabbit monoclonal (1:250, described in ), α-LAP2alpha rabbit polyclonal (LAP2a, 1:500, Abcam, ab5162), α-Lamin B1 rabbit polyclonal (1:200, Abcam, ab16048), α-Histone H3 (tri methyl K27) rabbit monoclonal (1:400, Abcam, ab192985), α-acetyl-Histone H4 (Ac-Lys16) rabbit polyclonal (1:400, Sigma-Aldrich, # H9164), α-Ki67 mouse monoclonal (1:200, DB Biosciences, #610968), and α-PCNA mouse monoclonal (1:200, Abcam, ab29).

Techniques: Produced, Marker, Recombinant, SYBR Green Assay, Transgenic Assay, Expressing, Knock-In, Negative Control, Real-time Polymerase Chain Reaction, Derivative Assay, Plasmid Preparation, Construct, Software, Imaging, Light Microscopy, Laser-Scanning Microscopy, Super-Resolution Microscopy, FACS

Journal: Cell reports

Article Title: Activation of endoplasmic reticulum stress in premature aging via the inner nuclear membrane protein SUN2

doi: 10.1016/j.celrep.2023.112534

Figure Lengend Snippet: KEY RESOURCES TABLE

Article Snippet: Rabbit polyclonal anti-Hsp40 , StressMarq Biosciences , Cat#SPC-100; RRID:AB_2261861.

Techniques: Produced, Marker, Recombinant, SYBR Green Assay, Transgenic Assay, Expressing, Knock-In, Negative Control, Real-time Polymerase Chain Reaction, Derivative Assay, Plasmid Preparation, Construct, Software, Imaging, Light Microscopy, Laser-Scanning Microscopy, Super-Resolution Microscopy, FACS

Journal: Molecular Cancer

Article Title: Down-regulation of miR-675-5p contributes to tumor progression and development by targeting pro-tumorigenic GPR55 in non-small cell lung cancer

doi: 10.1186/s12943-015-0342-0

Figure Lengend Snippet: Down-regulation of miR-675-5p is inversely associated with advanced stage and lymph node metastasis of NSCLC. (A) miR-675-5p expression level was significantly lower in NSCLC tissues than in their matched normal tissues. (B) Low-level expression of miR-675-5p was associated with high tumor stage of NSCLC (P < 0.05). (C) Low-level expression of miR-675-5p was related with lymph node metastasis of NSCLC (P < 0.05). ( D) miR-675-5p expression in NSCLC cell lines and normal human bronchial epithelial cell line (HBE). Expression levels of miR-675-5p were determined by qRT-PCR and normalized against an endogenous control (U6 RNA). Data were represented as the mean ± SEM of three independent experiments. *P < 0.05, **P < 0.01.

Article Snippet: Six NSCLC cell lines (HTB-182, A549, SPC-A-1, H1299, 95-D, Ltep-a-2) were obtained from the American Type Culture Collection.

Techniques: Expressing, Quantitative RT-PCR, Control

Journal: Molecular Cancer

Article Title: Down-regulation of miR-675-5p contributes to tumor progression and development by targeting pro-tumorigenic GPR55 in non-small cell lung cancer

doi: 10.1186/s12943-015-0342-0

Figure Lengend Snippet: Overexpression of miR-675-5p inhibited proliferation and colony formation of NSCLC cells. (A) the level of miR-675-5p in A549 and HTB-182 cells was significantly up-regulated after transfection with miR-675-precursor. (B) miR-675-5p reduced cell proliferation in NSCLC cells. Cell proliferation was determined using MTT assays. (C) miR-675-5p induced cell cycle arrest at the G1/S phase. (D) miR-675-5p suppressed colony formation compared with controls. The number of colonies were calculated and depicted by the ban graph. Data were represented as the mean ± SEM of three independent experiments. Negative control: pGCsil-GFP Vector. *P < 0.05, **P < 0.01.

Article Snippet: Six NSCLC cell lines (HTB-182, A549, SPC-A-1, H1299, 95-D, Ltep-a-2) were obtained from the American Type Culture Collection.

Techniques: Over Expression, Transfection, Negative Control, Plasmid Preparation

Journal: Molecular Cancer

Article Title: Down-regulation of miR-675-5p contributes to tumor progression and development by targeting pro-tumorigenic GPR55 in non-small cell lung cancer

doi: 10.1186/s12943-015-0342-0

Figure Lengend Snippet: Requirement of GPR55 for miR-675-5p induced suppression of NSCLC cell proliferation, migration and invasion. (A) the level of miR-675-5p in the cells was significantly down-regulated after transfection with miR-675-5p inhibitor. (B) GPR55 was knockdowned in Ltep-a-2 miR-675-5p inhibition cells and analyzed by western blot analysis. Knockdown of GPR55 significantly inhibited proliferation (C) , inhibited colony formation (D) , induced cell cycle arrest at the G1/S phase (E) , inhibited migration and invasion (F) , and decreased the wound healing rate (G) in Ltep-a-2 cells. Data were represented as the mean ± SEM of three independent experiments. Negative control: pGC FU-RNAi-NC-LV. *P < 0.05, P* < 0.01.

Article Snippet: Six NSCLC cell lines (HTB-182, A549, SPC-A-1, H1299, 95-D, Ltep-a-2) were obtained from the American Type Culture Collection.

Techniques: Migration, Transfection, Inhibition, Western Blot, Knockdown, Negative Control