mouse th2 cell line d10 g4 1  (ATCC)

Journal:

Article Title: Differential Signaling of T Cell Generation of IL-4 by Wild-Type and Short-Deletion Variant of Type 2 G Protein-Coupled Receptor for Vasoactive Intestinal Peptide (VPAC 2 ) 1

doi:

Figure Lengend Snippet: Quantification of the levels of mRNA encoding WT and SD VPAC2 in D10G4.1 cell transfectants. Each column and bar depicts the mean ± SD of the results of three individual determinations. The corresponding values for mock-transfected D10G4.1 Th2 cells were marginally detectable and were indistinguishable from those of untransfected D10G4.1 Th2 cells. Peaks of mRNA encoding each type of VPAC2 receptor are evident at 24 h and persist without significant change at 48 h. None of the differences between levels of mRNA encoding WT and SD VPAC2 were significant.

Article Snippet: Cells, nucleofection procedures, and Abs A mouse Th2 cell line designated D10G4.1 was obtained from American Type Culture Collection.

Techniques: Transfection

Journal:

Article Title: Differential Signaling of T Cell Generation of IL-4 by Wild-Type and Short-Deletion Variant of Type 2 G Protein-Coupled Receptor for Vasoactive Intestinal Peptide (VPAC 2 ) 1

doi:

Figure Lengend Snippet: Western blot of membrane proteins extracted from mouse D10G4.1 Th2 cells transfected 48 h before with plasmids encoding mouse WT and SD VPAC2. The primary reagent for development was mouseanti-FLAG M2 mAb. The first pair of WT-SD lanes received 3 μg of extracted proteins, and the second pair 10 μg of extracted proteins; 10 μg of extract of control (C) untransfected D10G4.1 T cells were loaded into the last lane.

Article Snippet: Cells, nucleofection procedures, and Abs A mouse Th2 cell line designated D10G4.1 was obtained from American Type Culture Collection.

Techniques: Western Blot, Transfection

Journal:

Article Title: Differential Signaling of T Cell Generation of IL-4 by Wild-Type and Short-Deletion Variant of Type 2 G Protein-Coupled Receptor for Vasoactive Intestinal Peptide (VPAC 2 ) 1

doi:

Figure Lengend Snippet: Generation of VIP by mouse D10G4.1 Th2 cell transfectants. Each column and bar represents the mean ± SD of results from two studies analyzed in duplicate. None of the differences between levels of VIP generated by WT and SD transfectants was significant.

Article Snippet: Cells, nucleofection procedures, and Abs A mouse Th2 cell line designated D10G4.1 was obtained from American Type Culture Collection.

Techniques: Generated

Journal:

Article Title: Differential Signaling of T Cell Generation of IL-4 by Wild-Type and Short-Deletion Variant of Type 2 G Protein-Coupled Receptor for Vasoactive Intestinal Peptide (VPAC 2 ) 1

doi:

Figure Lengend Snippet: Dependence of the level of expression of mRNA encoding IL-4 by mouse D10G4.1 Th2 cell transfectants on endogenous VIP signaling through WT and SD VPAC2. The difference in corrected levels of mRNA encoding IL-4 in stimulated WT and SD VPAC2 transfectants without VIPase is significant with p < 0.05 (+) and that between WT transfectants without and with VIPase is significant with *, p < 0.01; n = 3, paired Student’s t test.

Article Snippet: Cells, nucleofection procedures, and Abs A mouse Th2 cell line designated D10G4.1 was obtained from American Type Culture Collection.

Techniques: Expressing

Journal:

Article Title: Differential Signaling of T Cell Generation of IL-4 by Wild-Type and Short-Deletion Variant of Type 2 G Protein-Coupled Receptor for Vasoactive Intestinal Peptide (VPAC 2 ) 1

doi:

Figure Lengend Snippet: Opposite signals from WT and SD VPAC2 to IL-4 secretion by D10G4.1 Th2 cell transfectants stimulated by adherent anti-CD3 plus anti-CD28 mAbs. Control secretion of immunoreactive IL-4 by transfectants in 24 h without VIP was set at 100%. Each column and bar depicts the mean ± SD of the results of three or more studies. The concentrations of VIP between those marked were 3 × 10−9, 3 × 10−8, and 3 × 10−7 M. The significance of differences between IL-4 secretion in the presence and absence of exogenous VIP for each set of transfectants was calculated by a standard paired t test;+, p < 0.05; and *, p < 0.01.

Article Snippet: Cells, nucleofection procedures, and Abs A mouse Th2 cell line designated D10G4.1 was obtained from American Type Culture Collection.

Techniques:

Journal:

Article Title: Differential Signaling of T Cell Generation of IL-4 by Wild-Type and Short-Deletion Variant of Type 2 G Protein-Coupled Receptor for Vasoactive Intestinal Peptide (VPAC 2 ) 1

doi:

Figure Lengend Snippet: Effect of VIP on anti-CD3 stimulated increases in the nuclear level of active Jun B protein in D10G4.1 Th2 cell transfectants. Each column and bar represents the mean ± range of two studies in duplicate. Suspensions of 106 transfectants in 0.6 ml of RPMI 1640 with 10% FBS were preincubated 60 min without or with 10−7 M VIP and then incubated with 1 μg of anti-CD3 Ab for 4 h. Each number in parentheses is the mean increase (+) or decrease (−) in active nuclear Jun B attributable to VIP. The significance of differences in levels of active nuclear Jun B between samples without and with VIP was calculated by a paired t test; *, p < 0.01.

Article Snippet: Cells, nucleofection procedures, and Abs A mouse Th2 cell line designated D10G4.1 was obtained from American Type Culture Collection.

Techniques: Incubation

Journal: Journal of Biological Chemistry

Article Title: Suppressor of Cytokine Signaling 3 Regulates Proliferation and Activation of T-helper Cells

doi: 10.1074/jbc.m300489200

Figure Lengend Snippet: FIG. 3. SOCS3 regulates proliferation of differentiated T-helper cells. A, Northern blot analysis using RNA (20 g/lane) from resting D10.G4.1 Th2 cells (lane 1) and D10.G4.1 cells stimulated with anti-CD3 and anti-CD28 Ab for 2 days as described under “Experimental Procedures.” B, detection of SOCS3 or -actin protein expression in D10.G4.1 Th2 cells with stable overexpression of SOCS3 (lane 2) or antisense SOCS3 (lane 3) or empty vector (lane 1) by Western blot analysis. C, D10.G4.1 Th2 cells transfected with vector alone, SOCS3 cDNA (S), or antisense SOCS3 cDNA (AS) were cultured for 1, 2, or 4 days, and cell proliferation was assessed by 3H-thymidine incorporation. Data are presented as mean CPM S.E. of responses of five replicate cultures. D, quantitative detection of SOCS1, SOCS2, or CIS mRNA transcripts in D10.G4.1 Th2 cells with stable overexpression of empty vector (Vector), SOCS3 (S), or antisense SOCS3 (AS) stimulated with Ag/APC for 2 days by real-time PCR. SOCS mRNA levels are presented as relative fluorescence units.

Article Snippet: The D10.G4.1 Th2 cell line (ATCC, Manassas, VA) was cultured in RPMI 1640 medium supplemented with 2 mM L-glutamine, 10 mM HEPES, 1 mM sodium pyruvate, 50 M 2-mercaptoethanol, 10 pg/ml IL-1 (R & D Systems), 10% rat T-STIM factor with concanavalin A (BD Biosciences) as recommended.

Techniques: Northern Blot, Expressing, Over Expression, Plasmid Preparation, Western Blot, Transfection, Cell Culture, Real-time Polymerase Chain Reaction, Fluorescence

Journal: Journal of Biological Chemistry

Article Title: Suppressor of Cytokine Signaling 3 Regulates Proliferation and Activation of T-helper Cells

doi: 10.1074/jbc.m300489200

Figure Lengend Snippet: FIG. 4. Transcriptional regulation of Gfi-1 or p27kip1 genes by TCR signaling correlates temporally with down-regulation of SOCS3 expression. A, RT-PCR analysis of Gfi-1, p27kip1, or -actin gene expression in naı¨ve cells (day 0) or primary T-helper cells stimulated with Ag (0.1, 1.0, or 10.0 g/ml HEL)/APC for 2 (Day 2) or 4 (Day 4) days. B, detection of Gfi-1 or -actin protein expression in primary T-helper cells stimulated with Ag (0.1, 1.0, or 10.0 g/ml HEL)/APC for 2 days by Western blot analysis. C, D10.G4.1 Th2 cells were transfected with a plasmid expressing the firefly luciferase gene under the control of the SOCS3 promoter (pGL3-SOCS3p), Gfi-1 cDNA mammalian expression vector (pcDNA-Gfi-1), and pRL-TK plasmid, which encodes Renilla luciferase as described under “Experimental Procedures.” Control cells were transfected with pcDNA vector alone or the pGL3-basic reporter plasmid (pGL3-Vector). Histograms represent SOCS3 promoter activity expressed in relative light units.

Article Snippet: The D10.G4.1 Th2 cell line (ATCC, Manassas, VA) was cultured in RPMI 1640 medium supplemented with 2 mM L-glutamine, 10 mM HEPES, 1 mM sodium pyruvate, 50 M 2-mercaptoethanol, 10 pg/ml IL-1 (R & D Systems), 10% rat T-STIM factor with concanavalin A (BD Biosciences) as recommended.

Techniques: Expressing, Reverse Transcription Polymerase Chain Reaction, Gene Expression, Western Blot, Transfection, Plasmid Preparation, Luciferase, Control, Activity Assay