Journal: iScience

Article Title: Type 2 diabetes is associated with increased circulating levels of 3-hydroxydecanoate activating GPR84 and neutrophil migration

doi: 10.1016/j.isci.2022.105683

Figure Lengend Snippet:

Article Snippet: Receptor plasmid, human HCA2 (GPR109A) , Origene , N/A.

Techniques: RNAscope, Recombinant, Enzyme-linked Immunosorbent Assay, Isolation, Reporter Gene Assay, Plasmid Preparation, Software

Journal: Cell reports

Article Title: In vivo RNA-seq and ChIP-seq analyses show an obligatory role for the C terminus of p53 in conferring tissue-specific radiation sensitivity

doi: 10.1016/j.celrep.2023.112216

Figure Lengend Snippet: KEY RESOURCES TABLE

Article Snippet: Rabbit monoclonal antibody to PUMA , Cell Signaling , 14570S; RRID:AB_2798517.

Techniques: Recombinant, Protein Extraction, Membrane, Staining, Protease Inhibitor, Plasmid Preparation, TUNEL Assay, Purification, ChIP-qPCR, Software

Journal: Oncotarget

Article Title: P73 tumor suppressor and its targets, p21 and PUMA, are required for madin-darby canine kidney cell morphogenesis by maintaining an appropriate level of epithelial to mesenchymal transition

doi:

Figure Lengend Snippet: A. Knockdown of TAp73 decreases, whereas knockdown of ΔNp73 increases, the expression of p21 and PUMA. The levels of p21, PUMA, and actin were measured in parental MDCK, MDCK-TAp73-KD, and MDCK-ΔNp73-KD cells by Western blotting. B. Generation of MDCK cell lines in which p21 was stably knocked down. Parental MDCK and MDCK-p21-KD cells were mock-treated or treated with camptothecin for 18h and the level of p21 and actin protein was measured by Western blotting. C. Representative microscopic images of parental MDCK and MDCK-p21-KD cells in 2-D cultures. D. Top panel: colony formation assay was performed with parental MDCK or MDCK-p21-KD cells. Bottom panel: the number of colonies was counted and presented as Mean ± S.D. from three separate experiments. E. Wound healing assay was performed with parental MDCK and MDCK-p21-KD cells. Top panel: cell migration was determined by visual assessment of cells migrating into the wound for 24 h using a phase-contrast microscopy. The dashed lines indicate the wound edge. Bottom panel: the time required for would closure was measured and presented as Mean ± s.d. from three separate experiments. F. Representative images of parental MDCK and MDCK-p21-KD cells in 3-D culture for 6 d or 12 d. Scale bar: 50 μM.

Article Snippet: Antibodies used were purchased from Bethyl (anti-p73), ProSci (anti-PUMA), Santa Cruz Biotechnology (anti-p21 (H164), anti-β-catenin (E-5), anti-Snail-1, anti-Twist), BD Transduction Laboratories (anti-E-cadherin, San Jose, CA), Sigma (anti-actin, St. Louis, MO), and BioRad (secondary antibodies against rabbit or mouse IgG conjugated with HRP, Life Science Research, Hercules, CA).

Techniques: Expressing, Western Blot, Stable Transfection, Colony Assay, Wound Healing Assay, Migration, Microscopy

Journal: Oncotarget

Article Title: P73 tumor suppressor and its targets, p21 and PUMA, are required for madin-darby canine kidney cell morphogenesis by maintaining an appropriate level of epithelial to mesenchymal transition

doi:

Figure Lengend Snippet: A. - D. The levels of β-catenin, E-cadherin, Snail, Twist, and actin were determined by Western blotting with extracts from parental MDCK (A-D), MDCK-TAp73-KD A. , MDCK-p21-KD B. , MDCK-PUMA-KD C. , and MDCK-ΔNp73-KD D. cells. E. The levels of E-cadherin, Snail and Twist transcripts were measured by qRT-PCR in parental MDCK cells and MDCK cells with knockdown of TAp73, ΔNp73, p21 or PUMA. The level of Actin was measured as an internal control. F. A model for the role of p73, p21 and PUMA in MDCK cell morphogenesis.

Article Snippet: Antibodies used were purchased from Bethyl (anti-p73), ProSci (anti-PUMA), Santa Cruz Biotechnology (anti-p21 (H164), anti-β-catenin (E-5), anti-Snail-1, anti-Twist), BD Transduction Laboratories (anti-E-cadherin, San Jose, CA), Sigma (anti-actin, St. Louis, MO), and BioRad (secondary antibodies against rabbit or mouse IgG conjugated with HRP, Life Science Research, Hercules, CA).

Techniques: Western Blot, Quantitative RT-PCR

Journal: Oxidative Medicine and Cellular Longevity

Article Title: MicroRNA-29b Enhances Osteoclast Survival by Targeting BCL-2-Modifying Factor after Lipopolysaccharide Stimulation

doi: 10.1155/2019/6018180

Figure Lengend Snippet: Identification of target for LPS-induced miR-29b in OCs. BMMs were incubated with M-CSF (30 ng/ml) and RANKL (40 ng/ml) for 40 h, washed thoroughly, and incubated further with LPS (50 ng/ml) in the presence of M-CSF (30 ng/ml) for 48 h. (a) Cells were transfected with 30 nM of miR-29b mimic or con mimic in the presence of M-CSF (30 ng/ml) for 6 h. Total RNA was analyzed by qPCR to quantify the expression of BMF, PUMA, BAK1, BIM, and HRK. Expression levels with con mimic treatment were set at 1. Cell lysates were subjected to Western blot analysis with antibodies against BMF, PUMA, BAK1, BIM, HRK, and BCL-2. Antibodies against β -actin were used for normalization. (b) Without transfection, total RNA was analyzed by qPCR to quantify the expression of BMF, and cell lysates were subjected to Western blot analysis with anti-BMF Ab. (c) Total RNA and tissue lysate of tibiae from LPS-treated or vehicle-treated (V, PBS) mice were analyzed by qPCR to quantify the expression of BMF and were subjected to Western blot analysis with antibodies against BMF. (d) Cells were thoroughly washed, transfected with 30 nM of anti-miR-29b or con inh, and stimulated with LPS (50 ng/ml) in the presence of M-CSF. After 48 h, total RNA was analyzed by qPCR to quantify BMF expression. Expression levels with con inh treatment were set at 1.0. Cell lysates were subjected to Western blot analysis with anti-BMF and anti-BCL-2 Ab. (e) Cells were thoroughly washed, transfected with 50 nM of scRNA or siBMF, stimulated with LPS (50 ng/ml) in the presence of M-CSF for 48 h, and analyzed to measure TRAP-positive MNCs and annexin V-positive cells. siRNA-mediated silencing of BMF was confirmed by RT-PCR and qPCR. The Ct values of the genes were widely distributed between 17.33 and 30.92. ∗ p < 0.05; ∗∗ p < 0.01; ∗∗∗ p < 0.001 compared with each corresponding control. Similar results were obtained from three independent experiments.

Article Snippet: The antibodies used were acquired as follows: primary antibodies against BMF (NBP1-76658) and PUMA (NBP1-76639) from Novus Biologicals (Cambridge, UK), BAK (12105) and BCL-2 (3498) from Cell Signaling (Danvers, MA, USA), BIM (559685) from BD Pharmingen (San Diego, CA, USA), HRK (AHP1178) from Bio-Rad (Oxford, UK), and β -actin (A5441) from Sigma Chemical.

Techniques: Incubation, Transfection, Expressing, Western Blot, Reverse Transcription Polymerase Chain Reaction, Control