Use of size fractionation of in vitro-activated human B lymphocytes for studies of cell cycle-dependent growth regulation.
Cell cycle progression of in vitro-stimulated human B lymphocytes occurs asynchronously. In order to allow detailed studies of growth control in G1, B cells were stimulated with anti-mu and low molecular weight B-cell growth factor (LMW BCGF) for 50 h and subsequently separated into nine fractions of cells by means of centrifugal elutriation. As judged by volume profiles, activation antigen expression and DNA content, the cells in fractions 1-4 were in early to mid-G1, while fractions 5-7 mainly contained cells in late G1, and fractions 8-9 contained cells mainly in S and G2. Cells in fractions 5-7 had passed the commitment point, as demonstrated by a high spontaneous incorporation of [3H]thymidine when recultured in medium alone. Moreover, S-phase entry of these cells was largely unaffected by exogenous growth-promoting or growth-inhibitory signals. Cells in early (fractions 1-2) and intermediate fractions (fractions 3-4) showed a negligible spontaneous [3H]thymidine incorporation, but a significant proportion of these cells progressed to S phase upon restimulation. Moreover, while IL-4 or the anti-CD40 MoAb G28-5 potently stimulated cells in early and intermediate fractions, the responsiveness of LMW BCGF alone was obtained just prior to the commitment point.
Erikstein
B K
BK
Department of Tissue Culture, Norwegian Radium Hospital, Oslo.
Schwarze
P
P
Blomhoff
H K
HK
Funderud
S
S
Stokke
T
T
Smeland
E B
EB
D001402:B-Lymphocytes / Q000166:cytology* / Q000378:metabolism; D002453:Cell Cycle / Q000187:drug effects / Q000502:physiology*; D002458:Cell Fractionation / Q000379:methods; D005576:Colforsin / Q000494:pharmacology; D004257:DNA Polymerase II / Q000096:biosynthesis; D015870:Gene Expression; D006801:Humans; D066298:In Vitro Techniques; D007371:Interferon-gamma / Q000494:pharmacology; D008213:Lymphocyte Activation; D008222:Lymphokines / Q000494:pharmacology; D008233:Lymphotoxin-alpha / Q000494:pharmacology; D016212:Transforming Growth Factor beta / Q000494:pharmacology; D014409:Tumor Necrosis Factor-alpha / Q000494:pharmacology
D008222:Lymphokines; D008233:Lymphotoxin-alpha; D016212:Transforming Growth Factor beta; D014409:Tumor Necrosis Factor-alpha; C058162:low molecular weight-B cell growth factor; D005576:Colforsin; D007371:Interferon-gamma; D004257:DNA Polymerase II