Cell Cycle Progression Markers Research Articles

A generalized model for multi-marker analysis of cell cycle progression in synchrony experiments

Motivation: To advance understanding of eukaryotic cell division, it is important to observe the process precisely. To this end, researchers monitor changes in dividing cells as they traverse the cell cycle, with the presence or absence of morphological or genetic markers indicating a cell's position in a particular interval of the cell cycle. A wide variety of marker data is available, including information-rich cellular imaging data. However, few formal statistical methods have been developed to use these valuable data sources in estimating how a population of cells progresses through the cell cycle. Furthermore, existing methods are designed to handle only a single binary marker of cell cycle progression at a time. Consequently, they cannot facilitate comparison of experiments involving different sets of markers.Results: Here, we develop a new sampling model to accommodate an arbitrary number of different binary markers that characterize the progression of a population of dividing cells along a branching process. We engineer a strain of Saccharomyces cerevisiae with fluorescently labeled markers of cell cycle progression, and apply our new model to two image datasets we collected from the strain, as well as an independent dataset of different markers. We use our model to estimate the duration of post-cytokinetic attachment between a S.cerevisiae mother and daughter cell. The Java implementation is fast and extensible, and includes a graphical user interface. Our model provides a powerful and flexible cell cycle analysis tool, suitable to any type or combination of binary markers.Availability: The software is available from: http://www.cs.duke.edu/~amink/software/cloccs/.Contact: michael.mayhew@duke.edu; amink@cs.duke.edu

The Tumor Suppressor Merlin Is Required for Cell Cycle Exit, Terminal Differentiation, and Cell Polarity in the Developing Murine Lens

PURPOSE. Neurofibromatosis type 2 (NF2) is an autosomal-dominant CNS tumor syndrome that affects 1:25,000 children and young adults. More than 50% of NF2 patients also develop posterior subcapsular cataracts (PSCs). The authors deleted Nf2 from the lens to determine its role in fiber cell differentiation. METHODS. Nf2 was conditionally deleted from murine lenses using the LeCre transgene. Standard histology and immunohistochemical and immunofluorescent methods were used to analyze lens morphology and markers of cell cycle progression, differentiation, and cell junctions in wild-type and knockout lenses from embryonic day 10.5 through postnatal day 3. RESULTS. Fiber cells lacking Nf2 did not fully exit the cell cycle and continued to express epithelial cell markers, such as FoxE3 and E-cadherin, despite expressing the fiber cell marker Prox1. Many fiber cells lost their elongated morphology. Markers of apical-basal polarity, such as ZO-1, were mislocalized along the lateral and basal membranes of fiber cells. The lens vesicle failed to separate from the surface ectoderm, and prospective lens and corneal epithelial cells formed a multilayered mass of cells at the surface of the eye. Herniation of this membrane caused the fiber mass to erupt through the cornea. CONCLUSIONS. Nf2 is required for complete fiber cell terminal differentiation, maintenance of cell polarity, and separation of lens vesicle from corneal epithelium. Defects identified in fiber cell differentiation may explain the formation of PSCs in patients with NF2. The lens provides an assay system to identify pathways critical for fiber cell differentiation and to test therapies for the tumors that occur in patients with NF2.

Premature Cell Cycle Entry Induced by Hepatitis B Virus Regulatory HBx Protein during Compensatory Liver Regeneration

The cycles of cell death and compensatory regeneration that occur during chronic hepatitis B virus (HBV) infection are central to viral pathogenesis and are a risk factor for the development of liver cancer. The HBV genome encodes one regulatory protein, HBx, which is required for virus replication, although its precise role in replication and pathogenesis is unclear. Because HBx can induce the G(0)-G(1) transition in cultured cells, the purpose of this study was to examine the effect of HBx during liver regeneration. Transgenic mice expressing HBx (ATX) and their wild-type (WT) littermates were used in the partial hepatectomy (PH) model for compensatory regeneration. Liver tissues collected from ATX and WT mice at varying sacrifice time points after PH were examined for markers of cell cycle progression. When compared with WT liver tissues, ATX livers had evidence of premature cell cycle entry as assessed by several variables (BrdUrd incorporation, proliferating cell nuclear antigen and mitotic indices, and reduced steady-state p21 protein levels). However, HBx did not affect apoptosis, glycogen storage, or PH-induced steatosis. Together, these results show that HBx expression can induce cell cycle progression within the regenerating liver. Our data are consistent with a model in which HBx expression contributes to liver disease and cancer formation by affecting early steps in liver regeneration.

The rexinoid LG100268 and the synthetic triterpenoid CDDO-methyl amide are more potent than erlotinib for prevention of mouse lung carcinogenesis

Female A/J mice injected with the carcinogen vinyl carbamate develop atypical adenomatous hyperplasias in lungs 4 weeks after injection with the carcinogen. The number and severity of tumors then increase over time, making these mice a useful model for evaluating potential chemopreventive agents. The rexinoid LG100268 (LG268), a selective ligand for the retinoid X receptor, and the methyl amide of the synthetic triterpenoid 2-cyano-3,12-dioxooleana-1,9(11)-dien-28-oic acid (CDDO) both significantly reduced the number, size, and severity of the histopathology of lung tumors in female A/J mice when fed in diet for 14 to 20 weeks. The total tumor burden was 85% to 87% lower in mice fed LG268 and CDDO-MA than in controls, and the percentage of high-grade tumors decreased from 59% in the controls to 25% or 30% with CDDO-MA and LG268. Erlotinib, which is used to treat lung cancer patients and is an inhibitor of the epidermal growth factor receptor, was less effective in this model. Immunohistochemical staining of geminin, a marker of cell cycle progression, was higher in lung sections from control mice than in mice treated with LG268. Because rexinoids and triterpenoids signal through different biological pathways, they should be tested in combination for the prevention of lung cancer.

Differential Involvement of Cell Cycle Reactivation between Striatal and Cortical Neurons in Cell Death Induced by 3-Nitropropionic Acid

Recent evidence suggests that unscheduled cell cycle activity leads to neuronal cell death. 3-Nitropropionic acid (3-NP) is an irreversible inhibitor of succinate dehydrogenase and induces cell death in both striatum and cerebral cortex. Here we analyzed the involvement of aberrant cell cycle progression in 3-NP-induced cell death in these brain regions. 3-NP reduced the level of cyclin-dependent kinase inhibitor p27 in striatum but not in cerebral cortex. 3-NP also induced phosphorylation of retinoblastoma protein, a marker of cell cycle progression at late G(1) phase, only in striatum. Pharmacological experiments revealed that cyclin-dependent kinase activity and N-methyl-d-aspartate (NMDA) receptor were cooperatively involved in cell death by 3-NP in striatal neurons, whereas only NMDA receptor was involved in 3-NP-induced neurotoxicity in cortical neurons. Death of striatal neurons was preceded by elevation of somatic Ca(2+) and activation of calpain, a Ca(2+)-dependent protease. Both striatal p27 down-regulation and cell death provoked by 3-NP were dependent on calpain activity. Moreover, transfection of p27 small interfering RNA reduced striatal cell viability. In cortical neurons, however, there was no change in somatic Ca(2+) and calpain activity by 3-NP, and calpain inhibitors were not protective. These results suggest that 3-NP induces aberrant cell cycle progression and neuronal cell death via p27 down-regulation by calpain in striatum but not in the cerebral cortex. This is the first report for differential involvement of cell cycle reactivation in different brain regions and lightens the mechanism for region-selective vulnerability in human disease, including Huntington disease.

Trophic factors and neuronal interactions regulate the cell cycle and Pax6 expression in Müller stem cells

The finding that Müller cells have stem cell properties in the retina has led to the hypothesis that they might be a source for replacing neurons lost in neurodegenerative diseases. However, utilization of Müller cells for regenerative purposes in the mammalian eye still requires identifying those factors that regulate their multipotentiality and proliferation. In addition, because Pax6 expression is indispensable for eye development, its regulation would be required during regeneration. In the present study we investigated the regulation of cell-cycle progression and Pax6 expression in pure Müller glial cell cultures and neuroglial cocultures from rat retinas. At early times in vitro, glial cells showed high expression of Pax6 and of nestin, a stem cell marker, and of markers of cell-cycle progression; expression of these markers decreased during development in parallel with increased glial differentiation. The addition of glial-derived neurotrophic factor, basic fibroblast growth factor, and insulin restored proliferation and also Pax6 and nestin expression in glial cells. Noteworthy, in neuroglial cocultures Müller cells retained Pax6 expression for longer periods, and, in turn, neuronal progenitors preserved their proliferative potential for several days in vitro. This suggests that neuroglial interactions mutually regulate their mitogenic capacity. In addition, in glial secondary cultures incubated with insulin, many neuroblast-like cells expressed the neuronal marker NeuN. Our results suggest that the proliferative capacity and the features of eye stem cells of Müller glial cells are regulated by molecular and cellular factors, which might then provide potential tools for manipulating retinal regeneration.

Antiproliferative potential of sarcophine and its semisynthetic sulfur-containing derivatives against human mammary carcinoma cell lines

The marine-derived cembranoid sarcophine (1) and its sulfur-containing semisynthetic derivatives (2–6) were evaluated for anticancer potential using cell cycle progression markers. No effect on MCF-7 cell viability or apoptosis was seen with these derivatives at concentrations of up to 100 μM after 72 h of incubation. At 100 μM, sarcophine and its derivatives 2–5 arrested the MCF-7 cells in G0/G1 phase, with concomitant decrease in the cell populations at S and G2+M phases. MDA-MB-231 cells were not responsive to any of the derivatives. Our preliminary results suggest that the sulfur-containing derivatives of sarcophine, especially 2 and 3, show potent and cell-specific antiproliferative activity.

The liver that caveolin built

Regeneration after liver damage requires caveolin and lipid droplets, according to Manuel Fernandez, Cecilia Albor (Universitat de Barcelona, Spain), Albert Pol, Robert Parton (University of Queensland, Brisbane, Australia), and colleagues. The results suggest that caveolin helps process or package storage lipids, and that the system somehow communicates with the cell cycle machinery. Figure 1 Lipid drops induced after partial hepatectomy (left) are missing in mice lacking caveolin (right). Caveolin is best known as an essential component of ordered plasma membrane domains called caveolae. Besides overseeing signaling and sorting in caveolae, caveolin has also been seen around lipid droplets after partial hepatectomy. These lipid droplets have a core of storage lipids surrounded by a single monolayer of phospholipids. They are thought to bud from the ER membrane after forming between its two leaflets. In partially hepatectomized mice lacking caveolin, the group found that initial events were intact: the two main signaling pathways that initiate recovery were activated, and levels of fatty acids increased in the serum and inside cells. But then things deteriorated. The signaling pathways persisted longer than normal, and lipid droplet accumulation and a marker of cell cycle progression were greatly reduced. The outcome was approximately half as much liver regeneration, and eightfold more death. Glucose feeding rescued these cell cycle and survival defects. The glucose may be acting by providing an alternate source of energy, membrane lipids, or both. Lipid droplet formation and caveolin's involvement in it remain a mystery. Perhaps caveolin helps package lipids in a way that allows them to be used properly. The group is now interested in seeing which stages of lipid droplet formation and lipid metabolism are blocked in the mutants, and in teasing apart the link from lipid metabolism to cell cycle. Reference: Fernandez, M.A., et al. 2006. Science. 313:1628–1632. [PubMed]

P27 small interfering RNA induces cell death through elevating cell cycle activity in cultured cortical neurons: a proof-of-concept study.

Recent research has demonstrated that cell cycle-associated molecules are activated in multiple forms of cell death in mature neurons, and raised a hypothesis that unscheduled cell cycle activity leads to neuronal cell death. But there is little evidence that changes in endogenous level of these molecules are causally associated with neuronal cell death. Here we transfected small interfering RNA (siRNA) targeting cyclin-dependent kinase (CDK) inhibitor p27, which plays an important role in cell cycle arrest at G1-S phase, into cultured cortical neurons. Transfection of p27 siRNA reduced neuronal viability in a time-dependent manner. p27 siRNA induced phosphorylation of retinoblastoma protein (Rb), a marker of cell cycle progression at late G1 phase. Moreover, phosphorylation of Rb and neuronal cell death provoked by p27 siRNA were abrogated by pharmacological CDK inhibitors, olomoucine and purvalanol A. Our data demonstrate that a decrease in endogenous p27 induces neuronal cell death through elevating cell cycle activity.

Cell Cycle Defects Contribute to a Block in Hormone-induced Mammary Gland Proliferation in CCAAT/Enhancer-binding Protein (C/EBPβ)-null Mice

In contrast to hormone-dependent breast cancer, steroid hormone-induced proliferation in the normal mammary gland does not occur in the steroid-receptor positive cells but rather in adjacent cells via paracrine signaling involving several local growth factors. To help elucidate the mechanisms involved in the block in proliferation in hormone-receptor positive cells, we have utilized a CCAAT/enhancer binding protein (C/EBPbeta)-null mouse model. Loss of this transcription factor results in increased steroid and prolactin receptor expression concomitant with a 10-fold decrease in proliferation in response to pregnancy hormones. To determine the basis for this decrease, several markers of cell cycle progression were analyzed in wild type and C/EBPbeta-null mammary epithelial cells (MECs). These studies indicated that cell cycle progression in C/EBPbeta-null MECs is blocked at the G1/S transition. C/EBPbeta-null mammary glands display substantially increased levels of the activated form of transforming growth factor beta, a potent inhibitor of epithelial cell proliferation, as well as increased downstream Smad2 expression and signaling. While cyclin D1 levels were equivalent, cyclin E expression was markedly reduced in C/EBPbeta-null as compared with wildtype MECs. In addition, increased p27 stability and retention in the nucleus and decreased levels of the cdc25a phosphatase contributed to a significant loss of cdk2 kinase activity. Collectively, these changes prevent C/EBPbeta-null mammary epithelial cells from responding to hormone-induced proliferative signals.

Protein Kinase C δ Stimulates Apoptosis by Initiating G1 Phase Cell Cycle Progression and S Phase Arrest

Overexpression of protein kinase C delta (PKCdelta) stimulates apoptosis in a wide variety of cell types through a mechanism that is incompletely understood. PKCdelta-deficient cells are impaired in their response to DNA damage-induced apoptosis, suggesting that PKCdelta is required to mount an appropriate apoptotic response under conditions of stress. The mechanism through which it does so remains elusive. In addition to effects on cell survival, PKCdelta elicits pleiotropic effects on cellular proliferation. We now provide the first evidence that the ability of PKCdelta to stimulate apoptosis is intimately linked to its ability to stimulate G(1) phase cell cycle progression. Using an adenoviral-based expression system to express PKCalpha,-delta, and -epsilon in epithelial cells, we demonstrate that a modest increase in PKCdelta activity selectively stimulates quiescent cells to initiate G(1) phase cell cycle progression. Rather than completing the cell cycle, PKCdelta-infected cells arrest in S phase, an event that triggers caspase-dependent apoptotic cell death. Apoptosis was preceded by the activation of cell cycle checkpoints, culminating in the phosphorylation of Chk-1 and p53. Strikingly, blockade of S phase entry using the phosphatidylinositol 3-kinase inhibitor LY294002 prevented checkpoint activation and apoptosis. In contrast, inhibitors of mitogen-activated protein kinase cascades failed to prevent apoptosis. These findings demonstrate that the biological effects of PKCdelta can be extended to include positive regulation of G(1) phase cell cycle progression. Importantly, they reveal the existence of a novel, cell cycle-dependent mechanism through which PKCdelta stimulates cell death.

Replication‐independent chromatin loading of Dnmt1 during G2 and M phases

The major DNA methyltransferase, Dnmt1, associates with DNA replication sites in S phase maintaining the methylation pattern in the newly synthesized strand. In view of the slow kinetics of Dnmt1 in vitro versus the fast progression of the replication fork, we have tested whether Dnmt1 associates with chromatin beyond S phase. Using time-lapse microscopy of mammalian cells expressing green-fluorescent-protein-tagged Dnmt1 and DsRed-tagged DNA Ligase I as a cell cycle progression marker, we have found that Dnmt1 associates with chromatin during G2 and M. This association is mediated by a specific targeting sequence, shows strong preference for constitutive but not facultative heterochromatin and is independent of heterochromatin-specific histone H3 Lys 9 trimethylation, SUV39H and HP1. Moreover, photobleaching analyses showed that Dnmt1 is continuously loaded onto chromatin throughout G2 and M, indicating a replication-independent role of Dnmt1 that could represent a novel and separate pathway to maintain DNA methylation.

TNFα-induced hepatocyte apoptosis is associated with alterations of the cell cycle and decreased stem loop binding protein

Background Inhibition of nuclear factor kappa B (NFκB) during liver regeneration induces hepatocyte apoptosis associated with normal DNA synthesis but decreased mitosis, suggesting that inhibition of NFκB impairs progression from S-phase through the G 2/M phase of the cell cycle. Our aim was to determine if inhibition of NFκB alters cell cycle characteristics in hepatocytes treated with tumor necrosis factor alpha (TNFα). Methods Primary hepatocytes from BALB/c mice were infected with adenoviruses expressing luciferase (control; AdLuc) or the IκB super-repressor (AdIκB) and treated with or without TNFα (30 ng/ml). Flow cytometry was performed (0 to 40 hours) to determine apoptosis and cell cycle progression. Reverse transcriptase-polymerase chain reaction and immunoblots assessed changes in cell cycle mediators and antiapoptotic factors. Results Primary hepatocytes treated with AdIκB and TNFα demonstrated significantly more S-phase cells (14% ± 3% vs 6% ± 2%, P<.05) at 14 hours compared with controls. Inhibition of NFκB with or without TNFα was associated with decreased expression of stem loop bind protein, a marker of cell cycle progression through S-phase. The NFκB-induced antiapoptotic proteins, iNOS and TRAF2, had decreased message at 9 and 12 hours, respectively, in TNFα- and AdIκB-treated cells. Conclusion Inhibition of NFκB in TNFα-treated primary mouse hepatocytes is associated with increased S-phase cell cycle retention and decreased stem loop bind protein.

Transient Cerebral Ischemia Induces Aberrant Neuronal Cell Cycle Re-entry and Alzheimer's Disease-like Tauopathy in Female Rats

Aberrant mitosis occurs in many tauopathy-related neurodegenerative diseases and is believed to precede the formation of neurofibrillary tangles. In this study, we report for the first time that transient cerebral ischemia induces aberrant mitotic proteins and hyperphosphorylation of tau protein with neurofibrillary tangle-like conformational epitopes in adult female rat cortex. Following transient cerebral ischemia in rats, initiation of apoptosis precedes and is potentially integrated with subsequent aberrant mitosis and tau hyperphosphorylation. Furthermore, inhibition of mitosis-related cyclin-dependent kinases (Cdks) by roscovitine significantly reduced the hyperphosphorylation of tau. Administration of the female sex steroid and potent neuroprotective agent, 17beta-estradiol, reduced ischemia-reperfusion-induced cerebral damage and the subsequent aberrant mitosis and tauopathies. These results provide a neuropathological basis for the higher prevalence of dementia in stroke patients and support the hypothesis that apoptosis and aberrant mitosis are integrated pathological events in neurons that may play a critical role in the development of Alzheimer's disease and other tauopathy-related neuropathology.

Cell cycle regulation in retinal progenitors by glia-derived neurotrophic factor and docosahexaenoic acid.

A recent study has shown that glia-derived neurotrophic factor (GDNF) and docosahexaenoic acid (DHA) promote the survival and differentiation of retina photoreceptors. The current study was undertaken to investigate whether these molecules participate in cell cycle regulation in retinal progenitors in vitro. Developmental changes in the expression of the stem cell marker nestin and of cell cycle and differentiated neuron markers were analyzed in neuroblasts obtained from 1-day-old rat retinas. The effects of GDNF and DHA on those changes were then determined. Expression of nestin, found in more than one third of neuroblasts at day 1, rapidly decreased during development, with most neuroblasts acquiring the photoreceptor phenotype. GDNF increased the percentage of photoreceptor progenitors expressing nestin, whereas DHA reduced it, simultaneously enhancing photoreceptor differentiation. Several markers of cell cycle progression indicated that photoreceptor progenitors maintained an active cell cycle during the first 2 days in vitro. GDNF stimulated the cell cycle, increasing the number of dividing cells and generating more photoreceptor progenitors, whereas DHA induced cell cycle exit and photoreceptor differentiation. Analysis of the expression of the cyclin-Cdk inhibitor p27(Kip1) confirmed these results. GDNF and DHA acted as molecular cues, counterbalancing the decision of photoreceptors to remain in or exit the cell cycle. The results strongly suggest that both factors participate in determining the number of photoreceptors in vitro, regulating the cell cycle and survival at early and late stages of development, respectively. Hence, GDNF and DHA may coordinately control the histogenesis of photoreceptors in the retina by modulating both neurogenesis and apoptosis.

Critical Proliferation-independent Window for Basic Fibroblast Growth Factor Repression of Myogenesis via the p42/p44 MAPK Signaling Pathway

In many cell types including myoblasts, growth factors control proliferation and differentiation, in part, via the mitogen-activated protein kinase (MAPK) pathway (also known as the extracellular regulated kinase (Erk) pathway). In C2C12 myoblast cells, insulin-like growth factor-1 and basic fibroblast growth factor (bFGF) activate MAPK/Erk, and both growth factors promote myoblast proliferation. However, these factors have opposing roles with respect to differentiation; insulin-like growth factor-1 enhances muscle cell differentiation, whereas bFGF inhibits the expression of the muscle-specific transcription factors MyoD and myogenin. Cells treated with bFGF and PD98059, a specific inhibitor of the MAPK pathway, show enhanced expression of the muscle-specific transcription factors MyoD and myogenin as compared with cells not exposed to this inhibitor. Inhibiting MAPK activity also enhances myoblast fusion and the expression of the late differentiation marker myosin heavy chain. Basic FGF mediated repression of muscle-specific genes does not result from continued cell proliferation, since bFGF-treated cells progress through only one round of cell division. We have identified a critical boundary 16 to 20 h after plating during which bFGF induced MAPK activity is able to repress myogenic gene expression and differentiation. Thus, the targets of MAPK that regulate myogenesis are functional at this time and their identification is in progress.

The lack of a functional p21 WAF1/CIP1 gene ameliorates progression to chronic renal failure

Partial renal ablation leads to progressive renal insufficiency and is a model of chronic renal failure from diverse causes. We find that mice develop functional and morphologic characteristics of chronic renal failure after partial renal ablation, including glomerular sclerosis, systemic hypertension, and reduced glomerular filtration. However, we now report that littermates with a homozygous deletion of the gene for the cyclin-dependent kinase inhibitor, p21(WAF1/CIP1), do not develop chronic renal failure after ablation. The markedly different reactions of the p21(+/+) and p21(-/-) animals was not because of differences in glomerular number or degree of renal growth but rather because of the presence or absence of a normal p21 gene. Although the reaction to the stress of renal ablation is both hyperplastic and hypertrophic in the presence of a functional p21 gene, it would appear that the absence of the p21 gene may induce a more hyperplastic reaction because proliferating-cell nuclear antigen expression, a marker of cell-cycle progression, in the renal epithelium of the remnant kidney was more than five times greater in the p21(-/-) mice than in the p21(+/+) animals. Because p21 is a potent inhibitor of the cell cycle, we speculate that p21 regulates the balance between hyperplasia and hypertrophy after renal ablation. We propose that this change in response inhibits the development of chronic renal failure. These studies suggest that controlling p21 function may ameliorate or even prevent progressive end-stage renal disease.

Flow cytometry study of cell cycle, apoptosis and drug resistance in acute leukemia.

The response to therapy of leukemic cells is largely determined by their capacity of proliferation and apoptosis in presence of the administered drugs. We describe here the main markers used in flow cytometry (FCM) and involved in the assessment of cell cycle parameters: single labeling by Propidium Iodide (PI) and double labeling anti-Bromodeoxyuridine (BrdUrd)/PI which, both in vitro and in vivo, gives cell percentages in the different cell cycle phases. The markers of cell cycle progression can be divided into proliferation markers such as PCNA (proliferating cell nuclear antigen) or Ki-67 and cell cycle progression markers. The latter, which are the core of the cell cycle machinery, are molecules recently characterized (Cyclins, CDKs (cell dependent kinases), CDIs (cyclin-dependent kinase inhibitors)) and their cell expression can be analyzed using FCM. FCM is also one of the best means to detect and quantitate apoptotic cells. Several techniques are described: Nuclear labeling using Hoechst 33342: mitochondrial labeling using DiOC6(3): detection of DNA fragmentation using 1) labeling of fixed and permeabilized cells with a DNA marker or 2) labeling of the free 3' DNA ends using incorporation of labeled deoxynucleotides; detection in apoptotic cells (Bcl-2, Fas, phospholipids...). At last, we analyzed flow cytometry methods to study the cell resistance to Ara-C and anthracyclins. In combination with cell kinetic studies and detection of apoptotic cells, they should increase the efficiency of the acute leukemia treatment.

Expression of cell-cycle-dependent genes in phytohemagglutinin-stimulated human lymphocytes.

We have investigated the expression of certain cell-cycle-dependent genes in human peripheral blood mononuclear cells (PBMC) stimulated by phytohemagglutinin (PHA). The genes studied had been previously identified as cell-cycle dependent in other cell types from different species and were induced by different mitogens. One of these genes (2F1) and the gene for the interleukin 2 receptor were induced by PHA even in cultures partially depleted of accessory cells where the lymphocytes grew in size but failed to enter S phase. The other genes (c-myc, 4F1, JE-3, and KC-1) were induced only in complete cultures of PBMC stimulated by PHA. These results confirm the dissociation between growth in size and cell DNA replication that can occur during cell-cycle progression. Moreover, the time course of appearance of detectable levels of RNA for these genes suggests that they may be used as markers of cell-cycle progression in the transition of lymphocytes from G0 to S phase.