Enhancement of specific [3H]ouabain binding and ouabain sensitive 86rubidium influx in intact human lymphocytes by a dialysable factor in human and fetal calf serum

1987 ◽  
Vol 72 (1) ◽  
pp. 71-79 ◽  
Author(s):  
V. M. S. Oh ◽  
E. A. Taylor ◽  
J. L. Ding ◽  
N. A. Boon ◽  
J. K. Aronson ◽  
...  
Keyword(s):  
Protein Synthesis ◽  
Plasma Membrane ◽  
Human Serum ◽  
Major Part ◽  
Fetal Calf Serum ◽  
Human Lymphocytes ◽  
Calf Serum ◽  
Thymidine Uptake ◽  
Ouabain Binding ◽  
Intact Cells

1. We have measured specific [3H]ouabain binding and ouabain sensitive 86rubidium influx in intact human lymphocytes incubated for up to 7 days in media containing different concentrations of fetal calf serum and human serum. 2. Incubation for periods of up to 7 days with fetal calf serum and human serum produced increases in both specific [3H]ouabain binding and ouabain sensitive 86rubidium influx that were dependent on concentration and time. 3. Neither specific [3H]ouabain binding nor ouabain sensitive 86rubidium influx was altered when dialysed serum was used, suggesting that both fetal calf serum and human serum contain a dialysable factor or factors which stimulate specific [3H]ouabain binding and ouabain sensitive 86rubidium influx in intact human lymphocytes. 4. To further elucidate the mechanisms underlying these changes we also measured the activity of two other enzymes of the lymphocyte plasma membrane, 5′-nucleotidase and γ-glutamyltransferase, the uptake of [3H]thymidine by the intact cells, and the effects of cycloheximide, puromycin, and anisomycin, inhibitors of protein synthesis. 5. The activity of 5′-nucleotidase was increased after incubation of the lymphocytes in fetal calf serum for 72 h, but the activity of γ-glutamyltransferase was not changed, suggesting some selectivity of the stimulatory effect. 6. Measurements of [3H]thymidine uptake by the lymphocytes showed that the major part of the observed changes in specific [3H]ouabain binding and ouabain sensitive 86rubidium influx was not attributable to transformation of the lymphocytes to lymphoblasts. 7. All three inhibitors of protein synthesis prevented the increase in specific [3H]ouabain binding due to fetal calf serum.

Cyclooxygenase products formed by primary cultures of cells from human chorion laeve: influence of steroids

1988 ◽  
Vol 66 (6) ◽  
pp. 788-793 ◽  
Author(s):  
W. Gibb ◽  
L. Riopel ◽  
R. Collu ◽  
J. R. Ducharme ◽  
M. D. Mitchell ◽  
...  
Keyword(s):  
Protein Synthesis ◽  
Fetal Calf Serum ◽  
Thymidine Incorporation ◽  
Elective Caesarean Section ◽  
Primary Cultures ◽  
Calf Serum ◽  
Inhibitory Effect ◽  
Defined Media ◽  
Chorion Laeve ◽  
Low Levels

Cells were isolated from human chorion laeve obtained at term (38–40 weeks gestation) by elective caesarean section and were maintained in primary culture for 1 week in defined media supplemented with 10% fetal calf serum. The production of various cyclooxygenase products by the cultures was examined. Little or no prostaglandin (PG) F2α, 6-keto-PGF1α, thromboxane B2, or 13,14-dihydro-15-keto-PGF2α was found. In contrast, the cells produced PGE2 which was low on day 0, increased during culture to a maximum on day 1 or 2, then declined to low levels. When cells were grown in the presence of media containing cortisol, dexamethasone, progesterone, and estradiol (at 10−7 or 10−9 M), the glucocorticoids (at 10−7 and 10−9 M), but not estrogen or progesterone, markedly inhibited the increase in PGE2 output. There was no difference in the protein content and thymidine incorporation of cells grown in the presence of glucocorticoids when compared with controls. This inhibitory effect was not sensitive to cycloheximide (1 μg/mL) indicating protein synthesis may not be involved in the process. These studies indicate that PGE2 is the major prostaglandin formed by primary cultures of chorion laeve and that prostaglandin metabolism in the chorion is sensitive to glucocorticoid inhibition.

Human tumor cells grown in fetal calf serum and human serum: Influences on the tests for lymphocyte cytotoxicity, serum blocking and serum arming effects

1976 ◽  
Vol 17 (4) ◽  
pp. 461-468 ◽  
Author(s):  
Hector L. Sulit ◽  
Sidney H. Golub ◽  
Reiko F. Irie ◽  
Rishab K. Gupta ◽  
Gary A. Grooms ◽  
...  
Keyword(s):  
Human Serum ◽  
Tumor Cells ◽  
Fetal Calf Serum ◽  
Calf Serum ◽  
Human Tumor ◽  
Human Tumor Cells ◽  
Lymphocyte Cytotoxicity

scholarly journals Assessment of LDL-Receptor Function in Human Lymphocytes with Lipoprotein Depleted Fetal Calf Serum and Pravastatin in the Culture Medium

1992 ◽  
Vol 20 (1) ◽  
pp. 53-56
Author(s):  
Seiji IWATA ◽  
Nagahiko SAKUMA ◽  
Hiroyuki HIRATA ◽  
Takayoshi ICHIKAWA ◽  
Yoshinori NOGUCHI ◽  
...  
Keyword(s):  
Culture Medium ◽  
Fetal Calf Serum ◽  
Human Lymphocytes ◽  
Calf Serum ◽  
Ldl Receptor ◽  
Receptor Function

scholarly journals Evidence for intracellular sodium pumps in permeabilized Xenopus laevis oocytes

1989 ◽  
Vol 260 (2) ◽  
pp. 395-399 ◽  
Author(s):  
G Schmalzing ◽  
S Kröner ◽  
H Passow
Keyword(s):  
Plasma Membrane ◽  
Xenopus Laevis ◽  
Binding Capacity ◽  
Intracellular Sodium ◽  
Xenopus Laevis Oocytes ◽  
Ouabain Binding ◽  
Intact Cells ◽  
Sodium Pumps

Ouabain binding was studied in Xenopus laevis oocytes permeabilized by detergents. The behaviour of markers showed that 10 microM-digitonin selectively disrupts the plasma membrane. In the presence of ATP, oocytes permeabilized at 10 microM-digitonin bound no more ouabain molecules than were required to abolish active 86Rb+ uptake in the intact cells. However, the ouabain binding capacity increased approx. 2-fold when inner membranes were disrupted by SDS or excess digitonin, as judged from the accompanying release of the lysosomal marker beta-hexosaminidase. The results suggest that oocytes have a large internal pool of functional sodium pumps.

scholarly journals Comparative Analysis of Mesenchymal Stem Cell Cultivation in Fetal Calf Serum, Human Serum, and Platelet Lysate in 2D and 3D Systems

2021 ◽  
Vol 8 ◽  
Author(s):  
Marline Kirsch ◽  
Jessica Rach ◽  
Wiebke Handke ◽  
Axel Seltsam ◽  
Iliyana Pepelanova ◽  
...  
Keyword(s):  
Human Serum ◽  
Mammalian Cells ◽  
Chondrogenic Differentiation ◽  
Fetal Calf Serum ◽  
Calf Serum ◽  
3D Culture ◽  
Cell Spreading ◽  
Platelet Lysate ◽  
Cell Cultivation

In vitro two-dimensional (2D) and three-dimensional (3D) cultivation of mammalian cells requires supplementation with serum. Mesenchymal stem cells (MSCs) are widely used in clinical trials for bioregenerative medicine and in most cases, in vitro expansion and differentiation of these cells are required before application. Optimized expansion and differentiation protocols play a key role in the treatment outcome. 3D cell cultivation systems are more comparable to in vivo conditions and can provide both, more physiological MSC expansion and a better understanding of intercellular and cell-matrix interactions. Xeno-free cultivation conditions minimize risks of immune response after implantation. Human platelet lysate (hPL) appears to be a valuable alternative to widely used fetal calf serum (FCS) since no ethical issues are associated with its harvest, it contains a high concentration of growth factors and cytokines and it can be produced from expired platelet concentrate. In this study, we analyzed and compared proliferation, as well as osteogenic and chondrogenic differentiation of human adipose tissue-derived MSCs (hAD-MSC) using three different supplements: FCS, human serum (HS), and hPL in 2D. Furthermore, online monitoring of osteogenic differentiation under the influence of different supplements was performed in 2D. hPL-cultivated MSCs exhibited a higher proliferation and differentiation rate compared to HS- or FCS-cultivated cells. We demonstrated a fast and successful chondrogenic differentiation in the 2D system with the addition of hPL. Additionally, FCS, HS, and hPL were used to formulate Gelatin-methacryloyl (GelMA) hydrogels in order to evaluate the influence of the different supplements on the cell spreading and proliferation of cells growing in 3D culture. In addition, the hydrogel constructs were cultivated in media supplemented with three different supplements. In comparison to FCS and HS, the addition of hPL to GelMA hydrogels during the encapsulation of hAD-MSCs resulted in enhanced cell spreading and proliferation. This effect was promoted even further by cultivating the hydrogel constructs in hPL-supplemented media.

scholarly journals Optimization of conditions for ex vivo expansion of CD34+ cells from patients with stage IV breast cancer

Blood ◽  
1994 ◽  
Vol 84 (10) ◽  
pp. 3567-3574 ◽  
Author(s):  
F Shapiro ◽  
TJ Yao ◽  
G Raptis ◽  
L Reich ◽  
L Norton ◽  
...  
Keyword(s):  
Breast Cancer ◽  
Human Serum ◽  
Fetal Calf Serum ◽  
Ex Vivo ◽  
Calf Serum ◽  
Stage Iv ◽  
Ex Vivo Expansion ◽  
Autologous Plasma ◽  
Prior Chemotherapy ◽  
Stage Iv Breast Cancer

Abstract Multiple cycles of high-dose chemotherapy can be hematologically supported by repeated administration of peripheral blood progenitors obtained after mobilization using cytokine alone or in combination with chemotherapy. We have explored the quality of such cells and their potential to undergo ex vivo expansion. Twenty-five leukapheresis samples from 19 patients who had received extensive prior chemotherapy for stage IV breast cancer were subjected to CD34+ cell selection using immunoaffinity columns of immunomagnetic bead separation. Cells were cultured in suspension in the presence of c-kit ligand, interleukin-3, interleukin-6, erythropoietin, and granulocyte colony-stimulating factor. Ten experiments were performed using weekly exchange of media and cytokines (Delta assay). Median myeloid and erythroid progenitors expanded 15-fold at 7 days (range, 7 to 43), 40-fold at 14 days (range, 18 to 470), 46-fold at 21 days (range, 0 to 118), and 21-fold at 28 days (range, 0 to 61). In a system using gas-permeable bags without exchange of media or cytokine, median progenitors expanded 13-fold at 7 days (range, 7 to 36), 14-fold at 10 days (range, 4 to 61), 14-fold at 12 days (range, 3 to 46), and 10-fold at 14 days (range, 1 to 35). Progenitor expansion less than 10-fold occurred in 8% of experiments at day 7, in 17% at day 10, in 43% at day 12, and in 50% at day 14. When autologous plasma, autologous plasma processed (removal of cryoprecipitate, centrifugation, then filtration), or human serum were substituted for 20% fetal calf serum, the ratio of progenitor expansion at 7 days relative to 20% fetal calf serum for 10% human serum, 20% human serum, and 1% autologous plasma processed was 1.01 (range, 0.62 to 1.33), 0.88 (range, 0.61 to 1.20), and 0.96 (range, 0.55 to 1.64), respectively. These findings support the feasibility of ex vivo expansion in a system free of nonhuman proteins of CD34(+)-derived progenitors obtained from the peripheral blood of patients who have received prior chemotherapy.

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