Cell culture

About: Cell culture is a research topic. Over the lifetime, 133361 publications have been published within this topic receiving 5364150 citations. The topic is also known as: cell culture techniques.

STAT5 as a molecular regulator of proliferation, differentiation and apoptosis in hematopoietic cells

Abstract: Signal transducers and activators of transcription (STATs) play key roles in growth factor‐mediated intracellular signal transduction. In the present study using a constitutively active STAT5 mutant, we show that STAT5 has pleiotropic functions regulating cell proliferation, differentiation and apoptosis in an IL‐3‐dependent Ba/F3 cell line. The mutant STAT5 possessed constitutive tyrosine phosphorylation and DNA binding activity, induced expression of bcl‐xL and pim‐1 in the absence of IL‐3 in Ba/F3 cells, and rendered Ba/F3 cells factor‐independent. Unexpectedly, IL‐3 treatment of the factor‐independent Ba/F3 cells expressing the constitutively active STAT5 resulted in apoptosis within 24 h, or differentiation followed by cell death. In these cells, mRNA expression of growth inhibitory genes downstream of STAT5 such as CIS, JAB/SOCS‐1/SSI‐1, and p21 WAF1/Cip1 was highly induced, correlating with prolonged hyper‐phosphorylation of the mutant STAT5 after IL‐3 stimulation. Of the STAT5‐regulated genes, we found that constitutive expression of JAB/SOCS‐1/SSI‐1 was sufficient to induce apoptosis of Ba/F3 cells, while p21 WAF1/Cip1 could induce differentiation of these cells. In contrast, constitutive expression of pim‐1 was sufficient to induce IL‐3‐independent growth of Ba/F3 cells. These findings suggest that a single transcription factor regulates cell fate by varying the intensity and duration of the expression of a set of target genes.

Differential Uptake of Functionalized Polystyrene Nanoparticles by Human Macrophages and a Monocytic Cell Line

Abstract: Tumor cell lines are often used as models for the study of nanoparticle-cell interactions. Here we demonstrate that carboxy (PS-COOH) and amino functionalized (PS-NH2) polystyrene nanoparticles of ∼100 nm in diameter are internalized by human macrophages, by undifferentiated and by PMA-differentiated monocytic THP-1 cells via diverse mechanisms. The uptake mechanisms also differed for all cell types and particles when analyzed either in buffer or in medium containing human serum. Macrophages internalized ∼4 times more PS-COOH than THP-1 cells, when analyzed in serum-containing medium. By contrast, in either medium, THP-1 cells internalized PS-NH2 more rapidly than macrophages. Using pharmacological and antisense in vitro knockdown approaches, we showed that, in the presence of serum, the specific interaction between the CD64 receptor and the particles determines the macrophage uptake of particles by phagocytosis, whereas particle internalization in THP-1 cells occurred via dynamin II-dependent endocytosis. PMA-differentiated THP-1 cells differed in their uptake mechanism from macrophages and undifferentiated THP-1 cells by internalizing the particles via macropinocytosis. In line with our in vitro data, more intravenously applied PS-COOH particles accumulated in the liver, where macrophages of the reticuloendothelial system reside. By contrast, PS-NH2 particles were preferentially targeted to tumor xenografts grown on the chorioallantoic membrane of fertilized chicken eggs. Our data show that the amount of internalized nanoparticles, the uptake kinetics, and its mechanism may differ considerably between primary cells and a related tumor cell line, whether differentiated or not, and that particle uptake by these cells is critically dependent on particle opsonization by serum proteins.

Negative growth regulation in a glioblastoma tumor cell line that conditionally expresses human wild-type p53.

Abstract: To investigate the effect that human wild-type p53 (wt-p53) expression has on cell proliferation we constructed a recombinant plasmid, pM47, in which wt-p53 cDNA is under transcriptional control of the hormone-inducible mouse mammary tumor virus promoter linked to the dominant biochemical selection marker gene Eco gpt. The pM47 plasmid was introduced into T98G cells derived from a human glioblastoma multiforme tumor, and a stable clonal cell line, GM47.23, was derived that conditionally expressed wt-p53 following exposure to dexamethasone. We show that induction of wt-p53 expression in exponentially growing cells inhibits cell cycle progression and that the inhibitory effect is reversible upon removal of the inducer or infection with simian virus 40. Moreover, when growth-arrested cells are stimulated to proliferate, induction of wt-p53 expression inhibits G0/G1 progression into S phase and the cells accumulate with a DNA content equivalent to cells arrested in the G0/G1 phase of the cell cycle. Taken together, these studies suggest that wt-p53 may play a negative role in growth regulation.

Induction of terminal differentiation in human promyelocytic leukemia cells by tumor-promoting agents

Abstract: Human promyelocytic leukemia cells (HL-60) were induced to differentiate into mature cells by the tumor-promoting agent phorbol-12-myristate-13-acetate and other related phorbol diesters. Differentiation was determined by an increase in the percent of myelocytes, metamyelocytes, and other mature myeloid cells as well as by an increase in the percent of phagocytizing cells. Induction of differentiation could be determined after 2 days of treatment with phorbol-12-myristate-13-acetate at a dose as low as 6 X 10(11) M. A correlation was found between reported tumor-promoting activity of a series of phorbol esters and their ability to induce myeloid differentiation and to inhibit cell growth. It is suggested that tumor-promoting agents like chemicals that induce terminal differentiation in these cells, at extremely low concentrations, may be used as a tool in the study of the control of cell growth, cell differentiation, and malignancy in human leukemic cells.