Journal: Laboratory investigation; a journal of technical methods and pathology

Article Title: SSeCKS sequesters cyclin D1 in glomerular parietal epithelial cells and influences proliferative injury in the glomerulus.

doi: 10.1038/labinvest.2011.199

Figure Lengend Snippet: Figure 2 Expression and interaction of Src-suppressed protein kinase C substrate (SSeCKS) and cyclin D1 in cultured parietal epithelial cells (PECs). (a) Fluorescence detection of SSeCKS (green), cyclin D1 (red), and nuclei (blue) during increasing cell–cell contact between PECs from 50 to 100% cell confluency showing induction of SSeCKS in the cytoplasm with concomitant nuclear to cytoplasmic redistribution of cyclin D1, compartmentalization that remains in fully differentiated (FD) PECs. (b) Western immunoblots (IB) showing downregulation of cyclin D1 and its immunoprecipitation (IP) with cytoplasmic SSeCKS during increasing cell–cell contact between PECs from 50 to 100% cell confluency, binding that remains in fully differentiated PECs.

Article Snippet: Total protein from isolated glomeruli was immunoblotted for SSeCKS (1:200), cyclin D1 (1:500), cyclin D3 (1:500; mouse monoclonal; Lab Vision, Fremont, CA, USA), and PAX8 (1:500; rabbit polyclonal; ProteinTech Group, Chicago, IL, USA), using tubulin-a (1:500; mouse monoclonal mouse; Lab Vision) as a loading control.

Techniques: Expressing, Cell Culture, Fluorescence, Western Blot, Immunoprecipitation, Binding Assay

Journal: Laboratory investigation; a journal of technical methods and pathology

Article Title: SSeCKS sequesters cyclin D1 in glomerular parietal epithelial cells and influences proliferative injury in the glomerulus.

doi: 10.1038/labinvest.2011.199

Figure Lengend Snippet: Figure 3 Phosphorylation of Src-suppressed protein kinase C substrate (SSeCKS) by activated protein kinase C (aPKC) in cultured parietal epithelial cells (PECs). (a) Western immunoblots showing rapid phosphorylation of SSeCKS at serine 283 (phospho-SSeCKS ser283) within 5 min of activation of PKC in PECs by phorbol 12,13-diacetate (PDA) or tumor necrosis factor-a (TNF-a), signaling abrogated by staurosporine aglycone (inhibitor). (b) Control western immunoblots showing the same signaling as in panel a in mouse fibroblasts expressing SSeCKS. (c) Fluorescence detection of phospho-SSeCKS ser283 (green), cyclin D1 (red), and nuclei (blue) in fully differentiated PECs showing phosphorylation of SSeCKs and cytoplasmic-to-nuclear redistribution of cyclin D1 following activation of PKC.

Article Snippet: Total protein from isolated glomeruli was immunoblotted for SSeCKS (1:200), cyclin D1 (1:500), cyclin D3 (1:500; mouse monoclonal; Lab Vision, Fremont, CA, USA), and PAX8 (1:500; rabbit polyclonal; ProteinTech Group, Chicago, IL, USA), using tubulin-a (1:500; mouse monoclonal mouse; Lab Vision) as a loading control.

Techniques: Phospho-proteomics, Cell Culture, Western Blot, Activation Assay, Control, Expressing, Fluorescence

Journal: Scientific Reports

Article Title: Novel pH-Sensitive Cyclic Peptides

doi: 10.1038/srep31322

Figure Lengend Snippet: ( a – c ) Concentration- and pH-dependent inhibition of HeLa cells proliferation was monitored at 48 hours after incubation of cells within ( a ) cleavable c [(WE) 4 WC]-S-S-amanitin, ( b ) cleavable c [E 4 W 5 C]-S-S-amanitin and ( c ) non-cleavable c [E 4 W 5 C]-amanitin constructs for 3 hours at normal (pH 7.4) and low (pH 6.0) pHs followed by constructs removal and keeping cells in DMEM with 10% FBS at pH 7.4. ( d ) HeLa cells were treated with FITC-labelled c [E 4 W 5 C] peptide conjugate (5 μM) for 30 min at pH 7.4 or 6.2, followed by washing at pH 7.4 in both cases, addition of Trypan Blue for 5 min and live cell imaging. ( e ) Cellular uptake of Alexa546-labelled c [E 4 W 5 C] peptide conjugates (5 μM) treated with HeLa cells for 6 hours in L-15 media at pH 6.2 in absence and presence of 4% of FBS, followed by washing and counting of fluorescent signal from cells using cellometer. Data are presented as mean ± St.D. The two-tailed unpaired Student’s t-test was used to calculate p-levels.

Article Snippet: The cellular uptake of the constructs was measured by fluorescent signal from cells counted using cellometer (Cellometer Vision CBA, Nexcelom).

Techniques: Concentration Assay, Inhibition, Incubation, Construct, Live Cell Imaging, Two Tailed Test

Journal: Frontiers in Endocrinology

Article Title: Expression and Functional Analysis of the BCL2-Associated Agonist of Cell Death ( BAD ) Gene in the Sheep Ovary During the Reproductive Cycle

doi: 10.3389/fendo.2018.00512

Figure Lengend Snippet: Overexpression of Bad gene in ovarian granulosa cells (A–D) . Effects of BAD on the cell apoptosis and gonadal steroids secretion in primary sheep granulosa cells were investigated. (A) Fluorescence image of granulosa cells after lentiviral particles infection. (B) Flow cytometry sorting results of GFP-positive cells. (C) mRNA expression of BAD gene among blank control, negative control and overexpressed groups. (D) Cell images of overexpressed groups, negative control and blank control under light and fluorescence microscopy, respectively. Arrows indicates the apoptotic cells arrows indicates the apoptotic cells.

Article Snippet: Annexin V-FITC Apoptosis Detection kits (K101; Bio-Vision Inc., Mountain View, CA, USA) were used for measuring apoptotic cell death in a dual-staining protocol according to the manufacturer's instructions.

Techniques: Over Expression, Fluorescence, Infection, Flow Cytometry, Expressing, Negative Control, Microscopy

Journal: Frontiers in Endocrinology

Article Title: Expression and Functional Analysis of the BCL2-Associated Agonist of Cell Death ( BAD ) Gene in the Sheep Ovary During the Reproductive Cycle

doi: 10.3389/fendo.2018.00512

Figure Lengend Snippet: Apoptosis - related genes expression and the P4 and E2 levels secreted by granulose cells after overexpressed (A,B) . Expressions of BAD gene and apoptosis-related genes ( BAX and Bcl-2 ) were traced during the differentiation of sheep granulosa cells in our BAD -overexpression cell line. mRNA levels of three genes were detected at 4 time points (5, 7, 11, and 13 days) after virus infection. (A) Relative expression levels of BAD , BAX, and Bcl-2 genes at 4 time points. A:5 days; B:7 days; C:11 days; D:13 days; D (B) The concentrations of progesterone (P4) and estradiol (E2) secreted into cultured medium were detected by radioimmunoassay. * Significant differences ( P < 0.05). ** Significant differences ( P < 0.01).

Article Snippet: Annexin V-FITC Apoptosis Detection kits (K101; Bio-Vision Inc., Mountain View, CA, USA) were used for measuring apoptotic cell death in a dual-staining protocol according to the manufacturer's instructions.

Techniques: Expressing, Over Expression, Infection, Cell Culture, RIA Assay