Journal: BMC Immunology

Article Title: Comparative efficacy of glucocorticoid receptor agonists on Th2 cell function and attenuation by progesterone

doi: 10.1186/s12865-020-00383-8

Figure Lengend Snippet: A comparison of prednisolone- and dexamethasone-mediated reduction of type-2 cytokine mRNA levels in primary Th2 cells. Fold differences in mRNA level from corticosteroid (CS) treated cells compared to vehicle are provided ( n = 3). Quantification of IL-13 mRNA following steroid treatment ( a ). Comparison of CS ability to suppress IL-13 ( b ). The half maximal inhibitory concentration (IC 50 ) for prednisolone and dexamethasone required to suppress IL-13 mRNA expression ( c ). Quantification of IL-5 mRNA following CS treatment ( d ). Comparison of the ability of either CS to suppress IL-5 ( e ). A comparison of the IC 50 values for prednisolone and dexamethasone required to suppress IL-5 mRNA expression ( f ). Cell counts following culture in vehicle or increasing concentration of prednisolone or dexamethasone ( f ). Data represent mean and standard error. Pred, prednisolone; Dex, dexamethasone.* p < 0.05 determined by one-way ( a , d & g ) or two-way ( b & e ) RM ANOVA or t-test ( c & f )

Article Snippet: Fig. 4 Expression of the nuclear progesterone receptor (PGR) is undetectable in a Th2 cell line (CCRF-CEM) and primary Th2 cells, but present in a breast adenocarcinoma cell line (MCF-7) used as a positive control (n = 3; a ).

Techniques: Comparison, Concentration Assay, Expressing

Journal: BMC Immunology

Article Title: Comparative efficacy of glucocorticoid receptor agonists on Th2 cell function and attenuation by progesterone

doi: 10.1186/s12865-020-00383-8

Figure Lengend Snippet: A comparison of prednisolone- and dexamethasone-induced apoptosis in a Th2 cell line (CCRF-CEM). Data are expressed as percentage of 7AAD + cells ( a ) identifying dead cells. The half maximal dose (EC 50 ) for prednisolone and dexamethasone required to induce necrosis of cells ( b ). Percentage of Annexin V + 7AAD − cells, identifying apoptotic cells ( c ). A comparison of the EC 50 values for prednisolone and dexamethasone required to reach 50% maximal induction of apoptosis in cells ( d ; n = 11). Percentage of Annexin V + 7AAD − primary Th2 cells following treatment with dexamethasone, exhibiting a similar plateau effect as CCRF-CEM at 0.5 μM dexamethasone ( e ; n = 3). Comparison of CS-induced apoptosis in CCRF-CEM as fold-difference over vehicle ( f ). Data represent mean and standard error. Pred, prednisolone; Dex, dexamethasone. * p < 0.05 determined by two-way ( a & c ), t -test ( b & e ) or one-way RM ANOVA ( g )

Article Snippet: Fig. 4 Expression of the nuclear progesterone receptor (PGR) is undetectable in a Th2 cell line (CCRF-CEM) and primary Th2 cells, but present in a breast adenocarcinoma cell line (MCF-7) used as a positive control (n = 3; a ).

Techniques: Comparison

Journal: BMC Immunology

Article Title: Comparative efficacy of glucocorticoid receptor agonists on Th2 cell function and attenuation by progesterone

doi: 10.1186/s12865-020-00383-8

Figure Lengend Snippet: Corticosteroid-induced cell death is dampened by the female sex hormone progesterone. Dexamethasone-induced cell death ( a , % 7AAD + ) and apoptosis ( b , % Annexin V + 7AAD − ) of a Th2 cell line (CCRF-CEM) in the presence or absence of progesterone (2 μM, n = 5). Head-to-head comparison of apoptosis following treatment with prednisolone ( c ) or dexamethasone ( d ) with or without progesterone ( n = 4). The half maximal response (EC 50 ) for dexamethasone and prednisolone in the presence of progesterone ( e ). Influence of progesterone on the maximal response of prednisolone- or dexamethasone-induced apoptosis ( f ). Efficacy of prednisolone vs dexamethasone to induce apoptosis in the presence of prednisolone ( g ). Data represent mean and standard error. Pred, prednisolone; Dex, dexamethasone; Prog, progesterone; Pre, pretreatment. * p < 0.05 determined by one-way ( a - e ) or two-way ( f & g ) RM ANOVA

Article Snippet: Fig. 4 Expression of the nuclear progesterone receptor (PGR) is undetectable in a Th2 cell line (CCRF-CEM) and primary Th2 cells, but present in a breast adenocarcinoma cell line (MCF-7) used as a positive control (n = 3; a ).

Techniques: Comparison

Journal: BMC Immunology

Article Title: Comparative efficacy of glucocorticoid receptor agonists on Th2 cell function and attenuation by progesterone

doi: 10.1186/s12865-020-00383-8

Figure Lengend Snippet: Expression of the nuclear progesterone receptor (PGR) is undetectable in a Th2 cell line (CCRF-CEM) and primary Th2 cells, but present in a breast adenocarcinoma cell line (MCF-7) used as a positive control (n = 3; a ). Effect of progesterone with or without dexamethasone on expression of PIBF1 (n = 3, b ) . Progesterone reduced the dexamethasone-mediated increase in FKPB5 mRNA level, but had no effect when applied alone (n = 5, c ). Data represent mean and standard error. * p < 0.05 determined by one-way ANOVA

Article Snippet: Fig. 4 Expression of the nuclear progesterone receptor (PGR) is undetectable in a Th2 cell line (CCRF-CEM) and primary Th2 cells, but present in a breast adenocarcinoma cell line (MCF-7) used as a positive control (n = 3; a ).

Techniques: Expressing, Positive Control

Journal: BMC Immunology

Article Title: Comparative efficacy of glucocorticoid receptor agonists on Th2 cell function and attenuation by progesterone

doi: 10.1186/s12865-020-00383-8

Figure Lengend Snippet: RNA-sequencing of Th2 cells

Article Snippet: Fig. 4 Expression of the nuclear progesterone receptor (PGR) is undetectable in a Th2 cell line (CCRF-CEM) and primary Th2 cells, but present in a breast adenocarcinoma cell line (MCF-7) used as a positive control (n = 3; a ).

Techniques:

Journal: Pharmaceutics

Article Title: Synthesis and Anticancer Activity of Novel Dual Inhibitors of Human Protein Kinases CK2 and PIM-1 †

doi: 10.3390/pharmaceutics15071991

Figure Lengend Snippet: Viability of CCRF-CEM (acute lymphoblastic leukemia), K-562 (chronic myelogenous leukemia), MCF-7 (breast carcinoma), and non-cancerous Vero cells after treatment with the tested compounds. The IC 50 values were calculated using an MTT-based assay and the following equation: Y = 100/(1 + 10^((LogIC50 − X) ∗ HillSlope)).

Article Snippet: An acute lymphoblastic leukemia ALL cell line (named CCRF-CEM) was purchased from the European Collection of Authenticated Cell Cultures (ECACC), whereas MCF-7 (hormone-dependent breast adenocarcinoma), K-562 (human chronic myelogenous leukemia), and Vero cells ( Cercopithecus aethiops kidney) were purchased from the American Type Culture Collection (ATCC, Manassas, VA, USA).

Techniques: MTT Assay

Journal: BMC Cancer

Article Title: Evaluation of bioactive sphingolipids in 4-HPR-resistant leukemia cells

doi: 10.1186/1471-2407-11-477

Figure Lengend Snippet: Acquired resistance upon long-term 4-HPR exposure . (A, left) R cell lines were continuously exposed to different 4-HPR concentrations (0.5, 3, 5, and 10 μM; cell lines named R0.5, R3, R5, and R10, respectively) and resistance verified by comparison to 48 h drug-withdrawn R cells. Data are the average ± SD of three independent experiments performed in quadruplicates, n = 12. (A, right) Parental CCRF-CEM cells and developed R cell lines were exposed to 10 μM 4-HPR and viability estimated after 48 h of drug exposure. (n = 12). ** P < 0.01; ANOVA plus Tamhane post-hoc analysis. (B) Comparative cell proliferation was determined by BrdU labeling as specified in Materials and Methods. Data are average ± SD of four independent experiments. * P < 0.05; Kruskal Wallis and Mann-Whitney test for pair-wise comparison.

Article Snippet: 4-HPR-sensitive parental leukemia cell line (CCRF-CEM) and 4-HPR-resistant cell lines (R0.5 and R10) were challenged by different anticancer drugs, such as cisplatin (0.5, 1, 2.5, or 5 μg/ml), paclitaxel (1, 5, 10, 30, or 100 nM), or adriamycin (50, 100, or 500 nM) and other insults such as UV irradiation (1, 3, or 5 mJ/cm ) or direct oxidative stress (H O 2; 10, 50, or 100 μM).

Techniques: Comparison, Labeling, MANN-WHITNEY

Journal: BMC Cancer

Article Title: Evaluation of bioactive sphingolipids in 4-HPR-resistant leukemia cells

doi: 10.1186/1471-2407-11-477

Figure Lengend Snippet: Effect of selected insults on the viability of 4-HPR-sensitive and resistant leukemia cell lines . 4-HPR-sensitive (CCRF-CEM) and derived resistant cell lines (R0.5 and R10) were exposed to several drugs or UV radiation, and cytotoxicity tested after 48 h by the XTT assay. Data are average ± SD of three independent experiments performed in quadruplicates (n = 12); * P < 0.05, ** P < 0.01; ANOVA plus Tamhane post-hoc test.

Article Snippet: 4-HPR-sensitive parental leukemia cell line (CCRF-CEM) and 4-HPR-resistant cell lines (R0.5 and R10) were challenged by different anticancer drugs, such as cisplatin (0.5, 1, 2.5, or 5 μg/ml), paclitaxel (1, 5, 10, 30, or 100 nM), or adriamycin (50, 100, or 500 nM) and other insults such as UV irradiation (1, 3, or 5 mJ/cm ) or direct oxidative stress (H O 2; 10, 50, or 100 μM).

Techniques: Derivative Assay, XTT Assay

Journal: BMC Cancer

Article Title: Evaluation of bioactive sphingolipids in 4-HPR-resistant leukemia cells

doi: 10.1186/1471-2407-11-477

Figure Lengend Snippet: Comparative endogenous SL pattern in 4-HPR-sensitive and resistant leukemia cell lines . Cell were grown with (R cell lines) or without (CCRF-CEM) 4-HPR. Endogenous TOTAL (sum of the different fatty acid chain-length species) dhCer (A, left panel) and Cer (A, right panel) levels, as well as dhSph (B), TOTAL GluCer, and LactCer (C) levels were determined by LC/MS. Data are average ± SD of four (A, B) or two (C) independent experiments. * P < 0.05; ** P < 0.01; ANOVA plus Tamhane or Bonferroni post-hoc test.

Article Snippet: 4-HPR-sensitive parental leukemia cell line (CCRF-CEM) and 4-HPR-resistant cell lines (R0.5 and R10) were challenged by different anticancer drugs, such as cisplatin (0.5, 1, 2.5, or 5 μg/ml), paclitaxel (1, 5, 10, 30, or 100 nM), or adriamycin (50, 100, or 500 nM) and other insults such as UV irradiation (1, 3, or 5 mJ/cm ) or direct oxidative stress (H O 2; 10, 50, or 100 μM).

Techniques: Liquid Chromatography with Mass Spectroscopy

Journal: BMC Cancer

Article Title: Evaluation of bioactive sphingolipids in 4-HPR-resistant leukemia cells

doi: 10.1186/1471-2407-11-477

Figure Lengend Snippet: Comparative analysis of dihydroceramide desaturase (DES) activity . Cells were incubated with the unnatural pyridinium dhCer (C12-PyrdhCer) alone or with 4-HPR (5 μM) for the selected time points and conversion into C12-PyrCer measured by LC/MS. Data are average ± SD of two independent values. ** P < 0.01; ANOVA plus Bonferroni post-hoc test.

Article Snippet: 4-HPR-sensitive parental leukemia cell line (CCRF-CEM) and 4-HPR-resistant cell lines (R0.5 and R10) were challenged by different anticancer drugs, such as cisplatin (0.5, 1, 2.5, or 5 μg/ml), paclitaxel (1, 5, 10, 30, or 100 nM), or adriamycin (50, 100, or 500 nM) and other insults such as UV irradiation (1, 3, or 5 mJ/cm ) or direct oxidative stress (H O 2; 10, 50, or 100 μM).

Techniques: Activity Assay, Incubation, Liquid Chromatography with Mass Spectroscopy

Journal: BMC Cancer

Article Title: Evaluation of bioactive sphingolipids in 4-HPR-resistant leukemia cells

doi: 10.1186/1471-2407-11-477

Figure Lengend Snippet: Comparative SL profile analysis upon drug withdrawal . R10 cells were incubated in 4-HPR-free medium for 48 h (WD = without drug) or long-term (~3 weeks) and endogenous SL profiles analyzed by LC-MS. Significance was based on R10 values. Data are average ± SD of 2-4 independent values. * P < 0.05; ** P < 0.01; ANOVA plus Tamhane or Bonferroni post-hoc test.

Article Snippet: 4-HPR-sensitive parental leukemia cell line (CCRF-CEM) and 4-HPR-resistant cell lines (R0.5 and R10) were challenged by different anticancer drugs, such as cisplatin (0.5, 1, 2.5, or 5 μg/ml), paclitaxel (1, 5, 10, 30, or 100 nM), or adriamycin (50, 100, or 500 nM) and other insults such as UV irradiation (1, 3, or 5 mJ/cm ) or direct oxidative stress (H O 2; 10, 50, or 100 μM).

Techniques: Incubation, Liquid Chromatography with Mass Spectroscopy

Journal: BMC Cancer

Article Title: Evaluation of bioactive sphingolipids in 4-HPR-resistant leukemia cells

doi: 10.1186/1471-2407-11-477

Figure Lengend Snippet: Effect of drug withdrawal on cell resistance . (A) Parental sensitive CCRF-CEM cells, resistant R10, and long-term drug withdrawn resistant (R10 longWD) cells were treated and cell death determined by annexin V-FITC/propidium iodide staining at the selected time points. Values represent the percentage of cells in each quadrant. Representative data from two independent experiments are shown. (B) A portion of each sample for SL profile analysis (Figure 5) was saved for XTT assay estimation of resistance. Briefly, R10 cells after 48 h and long-term drug withdrawal were subjected to 48 h treatment with 3 or 10 μM 4-HPR. All data were related to the corresponding untreated cells. Statistical analysis was related to R10 data. Data are average ± SD of 12 values from to three independent experiments. * P < 0.05; ** P < 0.01; ANOVA plus Tamhane post-hoc test.

Article Snippet: 4-HPR-sensitive parental leukemia cell line (CCRF-CEM) and 4-HPR-resistant cell lines (R0.5 and R10) were challenged by different anticancer drugs, such as cisplatin (0.5, 1, 2.5, or 5 μg/ml), paclitaxel (1, 5, 10, 30, or 100 nM), or adriamycin (50, 100, or 500 nM) and other insults such as UV irradiation (1, 3, or 5 mJ/cm ) or direct oxidative stress (H O 2; 10, 50, or 100 μM).

Techniques: Staining, XTT Assay

Journal: BMC Cancer

Article Title: Evaluation of bioactive sphingolipids in 4-HPR-resistant leukemia cells

doi: 10.1186/1471-2407-11-477

Figure Lengend Snippet: Analysis of 4-HPR-mediated alterations in dhSph and Sph phosphorylation . Externally added unnatural dhSph (17C-dhSph) and Sph (17C-Sph) analogues were used to estimate the secondary effects of 4-HPR-induced DES inhibition. CCRF-CEM cells were exposed to 4-HPR (10 μM) for 1 h and unnatural 17C species added for an additional 30 min incubation, as described in Methods. The effect on endogenous SLs (A) and unnatural dhSph species (B,C) analyzed by LC/MS. Comparative exogenous and endogenous SL profiles among parental (CCRF-CEM) and resistant cell lines (R0.5, R10) (D, E) were analyzed by the same methodology. Data are average ± SD of an experiment performed in duplicate.

Article Snippet: 4-HPR-sensitive parental leukemia cell line (CCRF-CEM) and 4-HPR-resistant cell lines (R0.5 and R10) were challenged by different anticancer drugs, such as cisplatin (0.5, 1, 2.5, or 5 μg/ml), paclitaxel (1, 5, 10, 30, or 100 nM), or adriamycin (50, 100, or 500 nM) and other insults such as UV irradiation (1, 3, or 5 mJ/cm ) or direct oxidative stress (H O 2; 10, 50, or 100 μM).

Techniques: Phospho-proteomics, Analogues, Inhibition, Incubation, Liquid Chromatography with Mass Spectroscopy

Journal: BMC Cancer

Article Title: Evaluation of bioactive sphingolipids in 4-HPR-resistant leukemia cells

doi: 10.1186/1471-2407-11-477

Figure Lengend Snippet: Toxicity mediated by the combination of 4-HPR with SL modulators . Parental-sensitive (CCRF-CEM) and derived-resistant (R0.5, R5, R10) cells were co-treated with 4-HPR (1 μM), DHS (5 μM; unnatural dhSph analogue), PPMP (10 μM; glucosylceramide synthase inhibitor), SAF (4 μM; another unnatural dhSph analogue), and/or SKI-II (5-20 μM) and cytotoxicity estimated after 48 h exposure using the XTT assay. Data are average ± SD of at least two independent experiments performed in quadruplicate (n ≥ 8); ** P < 0.01; ANOVA plus Tamhane post-hoc test.

Article Snippet: 4-HPR-sensitive parental leukemia cell line (CCRF-CEM) and 4-HPR-resistant cell lines (R0.5 and R10) were challenged by different anticancer drugs, such as cisplatin (0.5, 1, 2.5, or 5 μg/ml), paclitaxel (1, 5, 10, 30, or 100 nM), or adriamycin (50, 100, or 500 nM) and other insults such as UV irradiation (1, 3, or 5 mJ/cm ) or direct oxidative stress (H O 2; 10, 50, or 100 μM).

Techniques: Derivative Assay, XTT Assay

Journal: BMC Immunology

Article Title: Comparative efficacy of glucocorticoid receptor agonists on Th2 cell function and attenuation by progesterone

doi: 10.1186/s12865-020-00383-8

Figure Lengend Snippet: A comparison of prednisolone- and dexamethasone-mediated reduction of type-2 cytokine mRNA levels in primary Th2 cells. Fold differences in mRNA level from corticosteroid (CS) treated cells compared to vehicle are provided ( n = 3). Quantification of IL-13 mRNA following steroid treatment ( a ). Comparison of CS ability to suppress IL-13 ( b ). The half maximal inhibitory concentration (IC 50 ) for prednisolone and dexamethasone required to suppress IL-13 mRNA expression ( c ). Quantification of IL-5 mRNA following CS treatment ( d ). Comparison of the ability of either CS to suppress IL-5 ( e ). A comparison of the IC 50 values for prednisolone and dexamethasone required to suppress IL-5 mRNA expression ( f ). Cell counts following culture in vehicle or increasing concentration of prednisolone or dexamethasone ( f ). Data represent mean and standard error. Pred, prednisolone; Dex, dexamethasone.* p < 0.05 determined by one-way ( a , d & g ) or two-way ( b & e ) RM ANOVA or t-test ( c & f )

Article Snippet: The Th2 cell line (CCRF-CEM) and primary Th2 cells had no PGR mRNA and no production of PIBF1 mRNA in response to progesterone, indicating that the effect was not due to activation of nuclear progesterone receptors [ ].

Techniques: Comparison, Concentration Assay, Expressing

Journal: BMC Immunology

Article Title: Comparative efficacy of glucocorticoid receptor agonists on Th2 cell function and attenuation by progesterone

doi: 10.1186/s12865-020-00383-8

Figure Lengend Snippet: A comparison of prednisolone- and dexamethasone-induced apoptosis in a Th2 cell line (CCRF-CEM). Data are expressed as percentage of 7AAD + cells ( a ) identifying dead cells. The half maximal dose (EC 50 ) for prednisolone and dexamethasone required to induce necrosis of cells ( b ). Percentage of Annexin V + 7AAD − cells, identifying apoptotic cells ( c ). A comparison of the EC 50 values for prednisolone and dexamethasone required to reach 50% maximal induction of apoptosis in cells ( d ; n = 11). Percentage of Annexin V + 7AAD − primary Th2 cells following treatment with dexamethasone, exhibiting a similar plateau effect as CCRF-CEM at 0.5 μM dexamethasone ( e ; n = 3). Comparison of CS-induced apoptosis in CCRF-CEM as fold-difference over vehicle ( f ). Data represent mean and standard error. Pred, prednisolone; Dex, dexamethasone. * p < 0.05 determined by two-way ( a & c ), t -test ( b & e ) or one-way RM ANOVA ( g )

Article Snippet: The Th2 cell line (CCRF-CEM) and primary Th2 cells had no PGR mRNA and no production of PIBF1 mRNA in response to progesterone, indicating that the effect was not due to activation of nuclear progesterone receptors [ ].

Techniques: Comparison

Journal: BMC Immunology

Article Title: Comparative efficacy of glucocorticoid receptor agonists on Th2 cell function and attenuation by progesterone

doi: 10.1186/s12865-020-00383-8

Figure Lengend Snippet: Corticosteroid-induced cell death is dampened by the female sex hormone progesterone. Dexamethasone-induced cell death ( a , % 7AAD + ) and apoptosis ( b , % Annexin V + 7AAD − ) of a Th2 cell line (CCRF-CEM) in the presence or absence of progesterone (2 μM, n = 5). Head-to-head comparison of apoptosis following treatment with prednisolone ( c ) or dexamethasone ( d ) with or without progesterone ( n = 4). The half maximal response (EC 50 ) for dexamethasone and prednisolone in the presence of progesterone ( e ). Influence of progesterone on the maximal response of prednisolone- or dexamethasone-induced apoptosis ( f ). Efficacy of prednisolone vs dexamethasone to induce apoptosis in the presence of prednisolone ( g ). Data represent mean and standard error. Pred, prednisolone; Dex, dexamethasone; Prog, progesterone; Pre, pretreatment. * p < 0.05 determined by one-way ( a - e ) or two-way ( f & g ) RM ANOVA

Article Snippet: The Th2 cell line (CCRF-CEM) and primary Th2 cells had no PGR mRNA and no production of PIBF1 mRNA in response to progesterone, indicating that the effect was not due to activation of nuclear progesterone receptors [ ].

Techniques: Comparison

Journal: BMC Immunology

Article Title: Comparative efficacy of glucocorticoid receptor agonists on Th2 cell function and attenuation by progesterone

doi: 10.1186/s12865-020-00383-8

Figure Lengend Snippet: Expression of the nuclear progesterone receptor (PGR) is undetectable in a Th2 cell line (CCRF-CEM) and primary Th2 cells, but present in a breast adenocarcinoma cell line (MCF-7) used as a positive control (n = 3; a ). Effect of progesterone with or without dexamethasone on expression of PIBF1 (n = 3, b ) . Progesterone reduced the dexamethasone-mediated increase in FKPB5 mRNA level, but had no effect when applied alone (n = 5, c ). Data represent mean and standard error. * p < 0.05 determined by one-way ANOVA

Article Snippet: The Th2 cell line (CCRF-CEM) and primary Th2 cells had no PGR mRNA and no production of PIBF1 mRNA in response to progesterone, indicating that the effect was not due to activation of nuclear progesterone receptors [ ].

Techniques: Expressing, Positive Control

Journal: BMC Immunology

Article Title: Comparative efficacy of glucocorticoid receptor agonists on Th2 cell function and attenuation by progesterone

doi: 10.1186/s12865-020-00383-8

Figure Lengend Snippet: RNA-sequencing of Th2 cells

Article Snippet: The Th2 cell line (CCRF-CEM) and primary Th2 cells had no PGR mRNA and no production of PIBF1 mRNA in response to progesterone, indicating that the effect was not due to activation of nuclear progesterone receptors [ ].

Techniques: