Circadian rhythms in epidermal mitotic activity

Virchows Archiv B Cell Pathology - Circadian variations in mitotic index, mitotic rate (cells entering mitosis per hour) and mitotic duration were investigated in the epidermis of the back and ear...     

Studies on the control of mitotic activity

Summary A tetraploid cell population was produced in the primary root meristem of Pisum sativum by one-half hour treatments with various concentrations of colchicine. The tetraploid population so produced was found to be reasonably synchronous in its passage through successive mitotic cycles with the degree of synchrony being more or less proportional to the concentration of colchicine used. The average time between mitoses appears to be of the order of 12 hours which agrees well with previous estimates. Treatments of roots containing tetraploid populations with 2.52 × 10^–5 M. actidione for 15 minutes were used to demonstrate the possibility of using the system for studies on the differential susceptibility of cells at different stages of the mitotic cycle.This work was carried out under contract number RG-4835 of the National Institutes of Health, United States Public Health Service, and an Institutional Grant from the American Cancer Society.Contribution number 59-26 of the Department of Botany and Plant Pathology, Michigan State University, East Lansing, Michigan.Predoctoral Fellow (CF-9871) of the National Cancer Institute, United States Public Health Service     

Caspase activity is not required for the mitotic checkpoint or mitotic slippage in human cells

Biochemical studies suggest that caspase activity is required for a functional mitotic checkpoint (MC) and mitotic slippage. To test this directly, we followed nontransformed human telomerase immortalized human retinal pigment epithelia (RPE-1) cells through mitosis after inhibiting or depleting selected caspases. We found that inhibiting caspases individually, in combination, or in toto did not affect the duration or fidelity of mitosis in otherwise untreated cells. When satisfaction of the MC was prevented with 500 nM nocodazole or 2.5 μM dimethylenastron (an Eg5 inhibitor), 92-100% of RPE-1 cells slipped from mitosis in the presence of pan-caspase inhibitors or after simultaneously depleting caspase-3 and -9, and they did so with the same kinetics (~21-22 h) as after treatment with nocodazole or Eg5 inhibitors alone. Surprisingly, inhibiting or depleting caspase-9 alone doubled the number of nocodazole-treated, but not Eg5-inhibited, cells that died in mitosis. In addition, inhibiting or depleting caspase-9 and -3 together accelerated the rate of slippage ~40% (to ~13-15 h). Finally, nocodazole-treated cells that recently slipped through mitosis in the presence or absence of pan-caspase inhibitors contained numerous BubR1 foci in their nuclei. From these data, we conclude that caspase activity is not required for a functional MC or for mitotic slippage.     

Cyclical mitotic activity in the adeno-hypophysis of Rana temporaria

Summary In the pars distalis, the pars intermedia and the pars tuberalis of the hypophysis of Rana temporaria, an annual mitotic cycle is described.In the pars distalis there exists a gradient of mitotic activity, corresponding to a gradient of distribution of the PAS positive cells.In the pars distalis, the rate of cell degeneration and of cell renewal of the PAS positive cells seems to be faster than of the orange G positive cells.The replacement of degenerated PAS positive cells of the pars distalis seems to occur mainly during the months immediately following on the period of maximal activity of the PAS positive cells.     

Modeling haplotype block variation using Markov chains

Models of background variation in genomic regions form the basis of linkage disequilibrium mapping methods. In this work we analyze a background model that groups SNPs into haplotype blocks and represents the dependencies between blocks by a Markov chain. We develop an error measure to compare the performance of this model against the common model that assumes that blocks are independent. By examining data from the International Haplotype Mapping project, we show how the Markov model over haplotype blocks is most accurate when representing blocks in strong linkage disequilibrium. This contrasts with the independent model, which is rendered less accurate by linkage disequilibrium. We provide a theoretical explanation for this surprising property of the Markov model and relate its behavior to allele diversity.     

Taxol-induced mitotic block triggers rapid onset of a p53-independent apoptotic pathway.

BACKGROUND: At therapeutic concentrations, the antineoplastic agent taxol selectively perturbs mitotic spindle microtubules. Taxol has recently been shown to induce apoptosis, similar to the mechanism of cell death induced by other antineoplastic agents. However, taxol has shown efficacy against drug-refractory cancers, raising the possibility that this pharmacological agent may trigger an alternative apoptotic pathway. MATERIALS AND METHODS: The kinetics and IC50 of mitotic (M) block, aberrant mitosis, and cytotoxicity following taxol treatment were analyzed in human cell lines as well as normal mouse embryo fibroblasts (MEFs) and MEFs derived from p53-null mice. Apoptosis was followed by DNA gel electrophoresis and by in situ DNA end-labeling (TUNEL). RESULTS: Taxol induced two forms of cell cycle arrest: either directly in early M at prophase or, for those cells progressing through aberrant mitosis, arrest in G1 as multimininucleated cells. TUNEL labeling revealed that DNA nicking occurred within 30 min of the arrest in prophase. In contrast, G1-arrested, multimininucleated cells became TUNEL positive only after several days. In the subset of cells that became blocked directly in prophase, both wt p53-expressing and p53-null MEFs responded similarly to taxol, showing rapid onset of DNA nicking and apoptosis. However, p53-null MEFs progressing through aberrant mitosis failed to arrest in the subsequent G1 phase or to become TUNEL positive, and remained viable. CONCLUSIONS: Taxol induces two forms of cell cycle arrest, which in turn induce two independent apoptotic pathways. Arrest in prophase induces rapid onset of a p53-independent pathway, whereas G1-block and the resulting slow (3-5 days) apoptotic pathway are p53 dependent.     

Ionophoric activity of pluronic block copolymers

Pluronic block copolymers (triblock copolymers of poly(ethylene oxide) and poly(propylene oxide)) exhibit a chemosensitizing effect on multidrug resistant cell lines. Changes in membrane permeability are hypothesized to be responsible because inhibition of drug transport mediated by both the multidrug-resistance-associated protein and the P-glycoprotein drug efflux system has been observed. To test this hypothesis, we now have studied the ion conductivity mediated by Pluronic L61. Besides a detergent-like action, the copolymer was able to form regular channels and to exhibit carrier activity. Long living ion channels were formed by polymer oligomerization. Aggregate equilibrium was shifted toward L61 monomers and dimers, which operated as mobile carriers. Copolymer-induced membrane permeability for potassium ions (1 M KCl) was less than 10(-8) cm s(-1), whereas the permeability for uncharged doxorubicin molecules (1 mM) was equal to 5 x 10(-4) cm s(-1). The results are consistent with reports about an increased doxorubicin accumulation in cells (Venne, Li, S., Mandeville, R., Kabanov, A., and Alakhov, V. Y. (1996) Cancer Res. 56, 3626-3629). However, the increased permeability contrasts with the polymer-mediated decrease of drug efflux from cells. Preferential polymer binding to membrane proteins may mask the unspecific effect of L61 observed on lipid bilayers.     

Isolation of mitotic apparatus containing vesicles with calcium sequestration activity

We present the first report of isolated mitotic apparatus with vesicular calcium sequestration. Phase-contrast, differential interference contrast and polarized light microscopy as well as transmission and scanning electron microscopic examinations revealed structures comparable to mitotic apparatus in vivo. Numerous membrane-bound vesicles which retained their osmotic activity were present throughout. Microtubules, yolk, ribosomes and condensed chromatin were also present. The protein composition of mitotic apparatus was not dramatically altered by treatment with 0.5% Triton X-100, even though vesicles were destroyed and yolk was extracted. Calcium sequestration was demonstrated with ATP-dependent accumulation of ⁴⁵Ca by mitotic apparatus whose vesicles were left intact. Compared with controls for which no nucleotide was added, accumulation by mitotic apparatus with intact vesicles was enhanced to 184% when it was present. When ATP was supplemented with the divalent ionophore A23187, the calcium retention level was comparable to that of the control to which no nucleotide was added. Finally, the calcium accumulation by mitotic apparatus treated with either of the nonhydrolyzable ATP analogs AMPPCP or AMPPNP resulted in calcium retention levels similar to those of controls. The solubilization of vesicles with Triton X-100 abolished calcium accumulation in the presence or absence of any of the above additives. Resolution of vesicles on sucrose step gradients after ⁴⁵Ca-oxalate loading with ATP or AMPPCP indicates that a specific vesicular fraction sequesters ⁴⁵Ca.