Cell Cycle Progression Of Lymphocytes Research Articles

Licorice infusion: Chemical profile and effects on the activation and the cell cycle progression of human lymphocytes

A licorice infusion (LI) and its major constituents were investigated for their capacity to stimulate the activation and the cell cycle progression of human lymphocytes, measured by the CD69 expression and DNA content, respectively. The chemical profile of LI was determined by high-performance liquid chromatography-diode array detection (HPLC-DAD). Results: Two major components of LI were identified as liquiritin (1) and glycyrrhizin (2); total flavones and flavonols were shown as its minor constituents. The LI (100-800 μg/ml) stimulated the expression of CD69 on lymphocytes in a concentration-independent manner. Values of the activation index (AI) of total lymphocytes treated with LI (100-800 μg/ml) did not differ significantly among them (P < 0.05), but were 50% lower than the AI value exhibited by cells treated with phytohemagglutinin (PHA). The LI showed a similar effect on T cells, but on a lower scale. Compounds 1 and 2 (12-100 μg/ml) did not stimulate the CD69 expression on lymphocytes. The LI, 1 and 2 showed no meaningful effect on cell cycle progression of lymphocytes. The experimental data indicates that LI stimulates the activation of lymphocytes as a result of a proliferation-independent process. This finding suggests that LI could be considered as a potential specific immune stimulator.

Degradation of cyclin D3 independent of Thr-283 phosphorylation

Cyclin D3 has been shown to play a major role in the regulation of cell cycle progression in lymphocytes. It is therefore important to understand the mechanisms involved in the regulation of this protein. We have previously shown that both basal and cAMP-induced degradation of cyclin D3 in Reh cells is dependent on Thr-283 phosphorylation by glycogen synthase kinase-3beta (GSK-3beta). We now provide evidence of an alternative mechanism being involved in the regulation of cyclin D3 degradation. Treatment of lymphoid cells with okadaic acid (OA), an inhibitor of protein phosphatases 1 and 2A (PP1 and PP2A), induces rapid phosphorylation and proteasomal degradation of cyclin D3. This degradation is not inhibited by the GSK-3beta inhibitors lithium or Kenpaullone, or by substitution of Thr-283 with Ala on cyclin D3, indicating that cyclin D3 can be degraded independently of Thr-283 phosphorylation and GSK-3beta activity. Interestingly, in vitro experiments revealed that PP1, but not PP2A, was able to dephosphorylate cyclin D3 efficiently, and PP1 was found to associate with His-tagged cyclin D3. These results support the hypothesis that PP1 constitutively keeps cyclin D3 in a stable, dephosphorylated state, and that treatment of cells with OA leads to phosphorylation and degradation of cyclin D3 through inhibition of PP1.

Enhancement of activation-induced apoptosis of lymphocytes by the cytotoxic ribonuclease onconase (Ranpirnase).

Onconase (Onc) is an amphibian ribonuclease of the pancreatic RNase family that is cytostatic and cytotoxic to several tumor lines. It also shows anti-tumor activity in mouse tumor models and is currently in phase III clinical trials. In animal tests and clinical trials Onc shows lesser toxicity and fewer side effects compared to most chemotherapeutic drugs. Intriguingly, repeated infusions of this protein do not cause apparent immunological reactions in patients. The aim of the present study was to investigate sensitivity to Onc of human lymphocytes during their mitogenic stimulation in response to the polyvalent mitogen phytohemagglutinin (PHA), and in mixed allogeneic lymphocyte cultures. Unexpectedly, we observed that frequency of cells undergoing activation-induced apoptosis was markedly increased in all cultures containing Onc. Apoptosis was measured by flow cytometry using markers that detect activation of caspases, the in situ presence of DNA strand breaks, and loss of fragmented DNA ('sub-G1' cell subpopulation). The enhancement of frequency of activation-induced apoptosis (up to 244%) was observed at 4.2-83 nM Onc concentration, which is at least an order magnitude lower than its minimal concentration reported to affect proliferation or induce apoptosis of leukemic and solid tumor cell lines. The cell cycle progression of lymphocytes that responded to PHA mitogenically was not affected at 8.3 or 83 nM Onc concentration. Because activation-induced apoptosis is the key mechanism regulating several in vivo immunological functions including induction of tolerance, the observed effects of Onc may explain the apparent lack of immune reactions to this protein in treated patients. The propensity of Onc to potentiate the activation-induced apoptosis suggests that this drug may have clinical utility as immunomodulating agent, e.g., to suppress transplant rejection or treat autoimmune diseases.

Dietary Zinc Deficiency and Protein-Energy Malnutrition Decrease in Vitro Murine T Lymphocyte Cell Cycle Progression

Abstract The objective of this study was to investigate the effects of dietary zinc deficiency and protein-energy malnutrition (PEM), alone and in combination, on murine splenic T-lymphocyte subpopulations and cell cycle progression using flow cytometry. Four month old C56BL/6 female mice were fed Zn-deficient (ZnDF), low protein (LP), combined Zn-deficient and low protein (ZnDF&LP), energy-restricted (ER) or control (CTRL) diet for 4 weeks. The CD4+/CD8+ ratio of T lymphocytes was not altered by the deficient diets, however, the total percentage of CD4+ plus CD8+ cells within the CD3+ population was less in all deficient groups compared to the CTRL group. When spleen cells were stimulated with concanavalin A (ConA) in the presence of 2-mercaptoethanol, the ZnDF&LP, ZnDF and LP groups had significantly fewer cells in both early and late S phase and significantly more cells in the G0 or resting state than the CTRL group. The ER group had significantly fewer cells in total S phase than the CTRL group. These...

Tumor necrosis factor-related apoptosis-inducing ligand (TRAIL) is an inhibitor of autoimmune inflammation and cell cycle progression.

The tumor necrosis factor-related apoptosis-inducing ligand (TRAIL) induces apoptosis of tumor cells but not normal cells; its role in normal nontransformed tissues is unknown. We report here that chronic blockade of TRAIL in mice exacerbated autoimmune arthritis, and that intraarticular TRAIL gene transfer ameliorated the disease. In vivo, TRAIL blockade led to profound hyperproliferation of synovial cells and arthritogenic lymphocytes and heightened the production of cytokines and autoantibodies. In vitro, TRAIL inhibited DNA synthesis and prevented cell cycle progression of lymphocytes. Interestingly, TRAIL had no effect on apoptosis of inflammatory cells either in vivo or in vitro. Thus, unlike other members of the tumor necrosis factor superfamily, TRAIL is a prototype inhibitor protein that inhibits autoimmune inflammation by blocking cell cycle progression.

Xenobiotic immunosuppressive agents: therapeutic effects in animal models of autoimmune diseases.

An unprecedented arsenal of new xenobiotic immunosuppressive agents has been developed recently. Most of the new immunosuppressants have been tested primarily in the treatment of allograft rejection in experimental models of transplantation, and some of the new drugs have already proven their safety and efficiency in extensive clinical trials on transplant patients. Another field for their potential application is the treatment of autoimmune diseases. This review will give an overview of the therapeutic potential of the new xenobiotic drugs in different animal models of rheumatoid arthritis, systemic lupus erythematosus, myasthenia gravis, multiple sclerosis, diabetes mellitus, thyroiditis and uveoretinitis. The new xenobiotics are either inhibitors of the de novo synthesis of nucleotides, for example mycophenolate mofetil, mizoribine, leflunomide, and brequinar, or are immunophilin-binding agents (cyclosporin, FK506 and rapamycin) that inhibit signal transduction and cell cycle progression in lymphocytes. A different mode of action is likely to account for the immunosuppressive effects of deoxyspergualin, which may interfere with intracellular chaperoning by the heat shock protein HSP70 and the activation of transcription factor NF-kappa B.

Spontaneous genomic fragility and cell cycle progression in lymphocytes of patients with cervical carcinoma

Thirty patients with invasive carcinoma of the cervix, along with 15 age-matched healthy females as controls, were studied to examine the frequency of spontaneous SCEs, chromosomal aberrations (CAs), and cell cycle progression in lymphocytes. The frequency of SCEs and TCAs was statistically significant in the patients over the control group. The correlation of SCEs and TCAs with the stage of cancer, age, and number of pregnancies was also made. The cell cycle of lymphocytes showed its prolongation in the patients, as is evident from the higher proportion of cells at M1 (metaphase of first cell division after 72 hours).

Inhibition of Human B Lymphocyte Cell Cycle Progression and Differentiation by Rapamycin

In this study, we have analyzed the effects of the immunosuppressive agent rapamycin on the activation of highly purified normal human B lymphocytes. When the polyclonal activators Staphylococcus aureus (SA) and soluble CD40 ligand (CD40L) were used to stimulate B cells, rapamycin inhibited both interleukin 2 (IL2)-dependent and -independent proliferation, as well as IL2-dependent differentiation into antibody-secreting cells. Cell cycle analysis indicated that rapamycin inhibited the progression of SA+IL2-stimulated B cells past the mid-G1 phase of the cell cycle. To begin to identify rapamycin-sensitive signaling events essential for B cell activation, we examined the effects of rapamycin on p34cdc2 and p33cdk2 kinase activities. SA+IL2 stimulation induced the activation of both cyclin-dependent kinases. Of interest, rapamycin abrogated the activation of both p34cdc2 and p33cdk2. Our results indicate therefore that rapamycin inhibits a number of SA- and CD40L-inducible events that may be necessary for both entry into S phase and for permitting subsequent B cell differentiation. These studies emphasize the utility of this drug as a tool to begin to dissect the activation pathways utilized by human B cells, as well as to provide implications for the therapeutic use of rapamycin in vivo.

Pig plasma modulates cell cycle kinetics but not the baseline frequency of sister-chromatid exhanges in human lymphocytes

The effect of human and pig plasma on the baseline frequency of sister-chromatid exchanges (SECs) in human and porcine plasma leukocyte cultures was studied. Human plasma leukocyte cultures, but not porcine plasma leukocyte cultures, showed at least a twofold increase in SCE frequency over whole blood culture values. Addition of pig plasma to human plasma leukocyte cultures and human plasma to pig plasma leukocyte cultures did not modify the baseline SCE frequencies. In both human and porcine cultures, cell proliferation was slower in plasma leukocyte cultures than in whole blood cultures. The addition of pig plasma to human plasma leukocyte cultures, but not the incorporation of human plasma in pig plasma leukocyte cultures, accelerated the cell cycle progression of lymphocytes. With 10% pig plasma in the plasma leukocyte culture medium, lymphocyte proliferation was similar to that in whole blood cultures. Smaller concentrations of pig plasma rendered cell cycle progression intermediate between the basal plasma leukocyte culture and whole blood culture values. Exchanging fetal calf serum for human AB serum in human plasma leukocyte cultures did not affect the cell cycle kinetics of lymphocytes but it did decrease their baseline SCE frequency.

Expression of polyamine transporter activity during B lymphocyte cell cycle progression.

The movement of resting B cells into the early stages of the cell cycle is accompanied by elevations in cellular polyamine levels. Our previous results indicate that part of this increase is a consequence of enhanced levels of ornithine decarboxylase activity in activated B cells. However, the levels of ornithine decarboxylase wane within 30 h post-stimulation, suggesting that B cells may acquire polyamines from an alternative source. Results in the present communication indicate that B cells initiate the transport of extracellular polyamines as they enter the G1 stage of the cell cycle. This induction of polyamine transport activity is seen after the stimulation of B cell cycle progression by both PMA and ionomycin, as well as cognate help from activated Th cells. The transport activities are energy and temperature sensitive, and can be competitively inhibited by unlabeled ligands. The results of the competition studies are compatible with the presence of at least two types of transporters, a low affinity transporter for both putrescine and spermidine, and a higher affinity transporter selective for spermidine. The results are discussed in terms of biochemical events associated with Th cell-mediated, B cell activation.

Spontaneous and induced sister chromatid exchange frequencies and cell cycle progression in lymphocytes of patients with carcinoma of the uterine cervix

Thirteen healthy females and thirteen untreated patients with carcinoma of the uterine cervix were studied for spontaneous and mitomycin C (MMC)-induced rates of sister chromatid exchange (SCE) and cell cycle progression. The mean values of spontaneous as well as MMC-induced SCE rates showed no statistically significant difference between groups. For studying cell cycle progression, cells in the M1, M2, and M3 stages were scored from the same samples. The percent values of cells in these stages, identified by the nature of differential sister chromatid staining, were found to be almost identical in normal as well as MMC-treated cultures in controls and patients. It was concluded that the presence of carcinoma of the uterine cervix in human females has no bearing either on spontaneous and MMC-induced SCE rates or on cell cycle progression in PHA-stimulated cultures of peripheral blood lymphocytes.