Influence of gonadotropin (FSH + LH) and thyrotropin (TSH) on the multiplication of Chinese hamster ovary (CHO) cells. Impact of the age of the culture.

CHO cells repeatedly treated with gonadotropin showed peak division rates after their third exposure and a decrease in the mitotic rate after their fourth exposure. Thyrotropin induced a considerable decrease in the mitotic rate following the first exposure, a significant increase after the second and a further decrease following the third and fourth exposures. The pattern did not differ between the two hormones when the cells were exposed further. The age (density of the cell cultures) had an appreciable influence on hormone-provoked changes in the mitotic rate, this differing only in intensity and never in the response following the initial re-exposure.     

Pharmacological effects of vinorelbine on body temperature and locomotor activity circadian rhythms in mice

The effects of vinorelbine (VRL) on the circadian rhythms in body temperature and locomotor activity were investigated in unrestrained B6D2F₁ mice implanted with radio-telemetry transmitters. A single intravenous VRL dose (24 or 12 mg/kg) was given at 7 h after light onset (HALO), a time of high VRL toxicity, and resulted in transient suppression of temperature and activity circadian rhythms in mice kept in light-dark (LD) 12h:12h. Such suppression was dose-dependent. It occurred within 1-5 d after VRL dosing. Recovery of both rhythms was partially complete within 5 d following the high dose and within 2 or 3 d after the low dose and was not influenced by suppression of photoperiodic synchronization by housing in continuous darkness. Moreover, VRL induced a dose-dependent relative decrease in amplitude and phase shift of the temperature circadian rhythm. The mesor and amplitude of the activity rhythm were markedly reduced following the VRL administration. The relevance of VRL dosing time was studied in mice housed in LD 12h:12h. Vinorelbine was injected weekly (20 mg/kg/injection) for 3 wk at 6 or 18 HALO. Vinorelbine treatment ablated the rest-activity and temperature rhythms 3-6 d after each dose, with fewer alterations after VRL dosing at 18 HALO compared to 6 HALO, especially for the body temperature rhythm. There was at least partial recovery 1 wk after dosing, suggesting the weekly schedule of drug treatment is acceptable for therapeutic purposes. Our findings demonstrate that VRL can transiently, yet profoundly, alter circadian clock function. Vinorelbine-induced circadian dysfunction may contribute to the toxicokinetics of this and possibly other anticancer drugs.     

Different apoptotic responses and patterns in adhering and floating neoplastic cell cultures: effects of microtubule antagonists

The relationship between apoptotic progression and cell cycle perturbation induced by microtubule-destabilising (vinblastine, Colcemid) and -stabilising (taxol) drugs was studied in two mesenchyme-derived neoplastic cell lines, growing as suspension (Jurkat) and monolayer (SGS/3A) culture, by morphocytochemical and biochemical approaches. The same kind of drug induced different effects on the cell kinetics (proliferation, polyploidisation, death) of the two cell lines. In floating cells, the drugs appeared more effective during the S phase, while in adherent cells they were more effective during the G2/M phase. Moreover two distinct neoplasia-associated apoptotic phenotypes emerged: the first pattern was the typical one and was found in cells with a low transition through the S/G2 phase (Jurkat), and the second one was mainly characterised by a cell death derived from micronucleated and mitotic cells, as a consequence of a low transition through the M/G1 phase (SGS/3A). Our data show that the machinery required for the trigger and progression of apoptosis is present in every cell cycle phase, also in conditions of karyological alterations (aneugenic micronucleations). On the other hand, a different sensitivity of the two microtubular components (interphasic network and mitotic spindle) appears to be related to the anchorage-dependence or -independence during the cell growth disturbances after exposure to antimicrotubular drugs.     

Effect of a chalone-containing preparation from Ehrlich ascites tumor on DNA synthesis and cell division in the tumor in relation to the time of day

The mitosis inhibitory action of chalone-containing preparation of the Ehrlich ascite tumour was shown to depend on the time of its administration on round the clock, and on the circadian rhythm phase of the mitotic activity in this tumour. This allowed a conclusion that the chalone system of the tumour may be involved in the formation of the circadian rhythm of cell division. It was found that Ehrlich's ascite tumour chalone system regulated DNA synthesis influencing the cell passage from G1-phase of the mitotic cycle to S-phase, and the processes occurring during S-phase.     

Lymphocyte beta-adrenergic refractoriness induced by theophylline or metaproterenol in healthy and asthmatic subjects

Beta-adrenergic refractoriness was assessed in human lymphocytes following in vivo administration of the beta-adrenergic agonist, metaproterenol, the phosphodiesterase inhibitor, theophylline, or both concomitantly, to normal and asthmatic subjects. In normal subjects both beta-adrenergic receptor number and isoproterenol stimulated cAMP response decreases during therapy with metaproterenol (59±3; 51±16% of control, respectively), theophylline (76±6; 78±16), or concommitant metaproterenol and theophylline (47±4; 69±13). The asthmatic subjects were of two types; one type responding to metaproterenol or theophylline therapy by down regulation of receptor number to zero or near zero values, and a second group of asthmatics insensitive to down regulation of receptor number. The results suggest that the induction of the refractory state is different between asthmatics and non-asthmatics, and that there may be a role for cAMP in the development of beta-adrenergic refractoriness, in vivo.     

Combined effect of epinephrine and chalone extracted from Ehrlich ascites carcinoma administered during different time of day on cell division in the tumor

The biphasic circadian rhythm of mitotic activity has been demonstrated in a 5-day Ehrlich's ascites carcinoma (EAC) in mice. Adrenaline injected intraperitoneally in a dose of 1.5 micrograms/g bw produced an inhibitory effect on cell division that lasted over 4 hours and reached maximum at injection to mice during light time of the day. EAC extract in a dose of 1 ml also inhibited the mitosis during 4 hours, but the greatest fall in the mitotic activity was observed during the minimum mitotic activity in the control animals. Combined administration of adrenaline and the extract resulted in the phenomenon of prolonged inhibition of cell division, that persisted for maximum 6-8 hours, if the preparations were injected in the middle of the day light time. Of definite importance was the rhythm of changes in the sensitivity of proliferating tumor cells.     

Action of the chalone from Ehrlich's ascites, tumor in mice on the mitotic activity in this tumor after single and double administrations

Chalone from Ehrlich's ascites tumour exerts a short-lived and reversible inhibitory effect on cell proliferation in the tumour both after a single and two-fold administration. 10 hours following single and two-fold injection of chalone (second injection was given at 6 p.m.), the mitotic index in tumour cells rises as compared to controls an evidence of chalone action on G(2) cell population of the mitotic cycle and synchronization of cell division. Repeated injection of chalone at 9 p.m. results in a more prolonged effect on the cells and in a more pronounced synchronization wave of G(2) cell population comparatively to its injection at 6 p.m. Thus the duration of cell inhibition in G(2) phase of the mitotic cycle depends with repeated administration of chalone, on the condition of cell population affected by chalone.     

Time-patterned drug administration: insights from a modeling approach

The physiological effects of a drug depend not only on its molecular structure but also on the time-pattern of its administration. One of the main reasons for the importance of temporal patterns in drug action is biological rhythms—particularly those of circadian period. These rhythms affect most physiological functions as well as drug metabolism, clearance, and dynamic processes that may alter drug availability and target cell responsiveness with reference to biological time. We present an overview of the importance of time-patterned signals in physiology focused on the insights provided by a modeling approach. We first discuss examples of pulsatile intercellular communication by hormones such as gonadotropin-releasing hormone, and by cyclic adenosine monophosphate (cAMP) signals in Dictyostelium amoebae. Models based on reversible receptor desensitization account in both cases for the existence of optimal patterns of pulsatile signaling. Turning to circadian rhythms, we examine how models can be used to account for the response of 24h patterns to external stimuli such as light pulses or gene expression, and to predict how to restore the physiological characteristics of altered rhythms. Time-patterned treatments of cancer involve two distinct lines of research. The first, currently evaluated in clinical trials, relies on circadian chronomodulation of anticancer drugs, while the second, mostly based on theoretical studies, involves a resonance phenomenon with the cell-cycle length. We discuss the implications of modeling studies to improve the temporal patterning of drug administration.     

Effect in vivo of beta-adrenergic stimulation, angiotensin II, dibutyryl cyclic AMP, and theophylline on tonin concentration in rat saliva and submaxillary gland.

Tonin (an enzyme present in rat submaxillary gland and saliva) has previously been shown to be able, unlike renin and reninlike substances, to release angiotensin II either directly by acting on an appropriate substrate or from angiotensin I. The administration of a beta-adrenergic drug, isoproterenol, produces a rise of tonin concentration in saliva without affecting its concentration in the submaxillary gland. Prior administration of a beta blocker, propranolol, partially prevents this effect. The administration of theophylline increases the tonin concentration in both saliva and the submaxillary gland, whereas dibutyryl cyclic AMP increases tonin concentration in the former. These results suggest that beta-adrenergic stimulation enhances both tonin release into the saliva and tonin synthesis in the submaxillary gland, and that these effects might be mediated by cyclic AMP. Infusion of angiotensin II blocked the stimulatory effect of isoproterenol on salivary tonin. 1Sar-8Ile-angiotensin II is both a weak antagonist of angiotensin II in this respect and a strong agonist in terms of blocking the effect of isoproterenol another role mirrored in other physiological mechanisms of derivatives of angiotensin II.     

Effects of chemical manipulation of mitotic arrest and slippage on cancer cell survival and proliferation

Microtubule-targeting cancer therapies interfere with mitotic spindle dynamics and block cells in mitosis by activating the mitotic checkpoint. Cells arrested in mitosis may remain arrested for extended periods of time or undergo mitotic slippage and enter interphase without having separated their chromosomes. How extended mitotic arrest and mitotic slippage contribute to subsequent cell death or survival is incompletely understood. To address this question, automated fluorescence microscopy assays were designed and used to screen chemical libraries for modulators of mitotic slippage. Chlorpromazine and triflupromazine were identified as drugs that inhibit mitotic slippage and SU6656 and geraldol as chemicals that stimulate mitotic slippage. Using the drugs to extend mitotic arrest imposed by low concentrations of paclitaxel led to increased cell survival and proliferation after drug removal. Cells arrested at mitosis with paclitaxel or vinblastine and chemically induced to undergo mitotic slippage underwent several rounds of DNA replication without cell division and exhibited signs of senescence but eventually all died. By contrast, cells arrested at mitosis with the KSP/Eg5 inhibitor S-trityl-L-cysteine and induced to undergo mitotic slippage were able to successfully divide and continued to proliferate after drug removal. These results show that reinforcing mitotic arrest with drugs that inhibit mitotic slippage can lead to increased cell survival and proliferation, while inducing mitotic slippage in cells treated with microtubule-targeting drugs seems to invariably lead to protracted cell death.     

Effect of opioid receptor ligands on cell division processes in the corneal epithelium of the white rat

The effects of the application of synthetic leu-enkephalin analogue--dalargin and naloxone on DNA synthesis and cell division in corneal epithelium were studied with the drugs injected once or five times. The influence of one and five intraperitoneal injections of the drugs in question at a dose of 10 micrograms/kg body weight was also investigated. In both cases the opioid receptor ligands examined activated DNA synthesis with the following adequate increase in mitotic index.     

Identification of CDK- and cyclin-like proteins in the eye of Bulla gouldiana

The ocular circadian rhythm in the eye of Bulla gouldiana is generated by a rhythm in membrane potential of retinal neurons that is driven by alterations in potassium conductance. Since potassium conductance may be modulated by the phosphorylation of potassium channels, the circadian rhythm may reflect rhythmic changes in protein kinase activity. Furthermore, the circadian rhythm recorded from the Bulla eye can be phase shifted by agents that affect protein synthesis and protein phosphorylation on tyrosine residues. Interestingly, the eukaryotic cell division residues. Interestingly, the eukaryotic cell division cycle is generated by similar processes. Rhythmic cell division is regulated by periodic synthesis and degradation of a protein, cyclin, and periodic tyrosine phosphorylation of a cyclin-dependent kinase (cdk), p34^cdc2. The interaction between these two proteins results in rhythmic kinase activity of p34^cdc2. Both cyclin and p34^cdc2 are pat of two diverse gene families, some of whose members have been localized to postmitotic cell types with no function yet determined. In the current work, we identify proteins similar to the cdks and cyclin in the eye of Bulla. Neither of these ocular proteins are found in mitotic cells in Bulla, and the cdk-like protein (p40) is specific to the eye. Furthermore, the concentration of the cyclin-like protein (p66) is affected by treatments that phase shift the circadain rhythm. The identification of cdk and cyclin-like proteins in the Bulla eye is consistent with the hypothesis that the biochemical mechanism responsible for generating the ocular circadian rhythm in Bulla is related to the biochemical mechnism that regulates the eukaryotic cell division cycle. 1994 John Wiley & Sons, Inc.     

Chronopharmacology and Chronotherapeutics: Definitions and Concepts

Most knowledge of medications has been derived from single- and multiple-dose investigations in which pharmacokinetic and pharmacodynamic phenomena have been evaluated following one, usually, daytime drug administration. Chronopharmacologic studies involving the evaluation of such phenomena after each of several different clock-hour treatments during the day- and nighttime reveal that biological rhythmic processes, such as those of 24 hr, can profoundly affect the kinetics and effects of various medications. Several new concepts have arisen based on findings from chronopharmacologic investigations, such as chronokinetics, chronesthesy and chronergy. These are defined and discussed herein using illustrative examples. A major goal of chronopharmacologic research is to devise chronotherapeutic interventions. Chronotherapeutics is the optimization of drug effects and/or minimization of toxicity by timing medications with regard to biological rhythms. Chronotherapeutics takes into account predictable administration-time-dependent variation in the pharmacokinetics of drugs as well as the susceptibility of target tissues due to temporal organization of physiochemical processes and functions of the body as circadian and other rhythms. The unequally divided and once-daily theophylline treatment schedules for the clinical management of nocturnal asthma, which are discussed in this issue, represent steps toward a chronotherapy.     

Time-patterned drug administration: insights from a modeling approach

The physiological effects of a drug depend not only on its molecular structure but also on the time-pattern of its administration. One of the main reasons for the importance of temporal patterns in drug action is biological rhythms—particularly those of circadian period. These rhythms affect most physiological functions as well as drug metabolism, clearance, and dynamic processes that may alter drug availability and target cell responsiveness with reference to biological time. We present an overview of the importance of time-patterned signals in physiology focused on the insights provided by a modeling approach. We first discuss examples of pulsatile intercellular communication by hormones such as gonadotropin-releasing hormone, and by cyclic adenosine monophosphate (cAMP) signals in Dictyostelium amoebae. Models based on reversible receptor desensitization account in both cases for the existence of optimal patterns of pulsatile signaling. Turning to circadian rhythms, we examine how models can be used to account for the response of 24h patterns to external stimuli such as light pulses or gene expression, and to predict how to restore the physiological characteristics of altered rhythms. Time-patterned treatments of cancer involve two distinct lines of research. The first, currently evaluated in clinical trials, relies on circadian chronomodulation of anticancer drugs, while the second, mostly based on theoretical studies, involves a resonance phenomenon with the cell-cycle length. We discuss the implications of modeling studies to improve the temporal patterning of drug administration.     

Changing the dosing schedule minimizes the disruptive effects of interferon on clock function

The effectiveness and toxicity of many drugs vary depending on the relationship between the dosing schedule and the 24-hour rhythms of biochemical, physiological and behavioral processes. In addition, several drugs can cause alterations to the 24-hour rhythms leading to illness and altered homeostatic regulation. However, the mechanisms of this drug-based disruption of circadian 'clock' genes remain unclear. Here, we show the disruptive effect of interferon-alpha on the rhythm of locomotor activity, body temperature and clock-gene mRNA expression in the periphery and suprachiasmatic nuclei, a primary circadian pacemaker. The rhythmicity of clock genes and the photic induction of the Per gene in suprachiasmatic nuclei were disturbed by the repetitive administration of interferon-alpha. Moreover, alteration of clock function, a new concept of adverse effects, can be overcome by optimizing the dosing schedule to minimize adverse drug effects.     

Effect of dibutyryl cAMP on cell cycle progression of rat brain tumor cells in vitro.

Autoradiographic and flow microfluorometry analyses have been applied to a study of perturbed cell kinetics in 9L rat brain tumor cells treated with dibutyryl cyclic AMP and theophylline alone and in combination in vitro. At a concentration of 1 mM each, cell growth ceased shortly after the administration of these drugs. The results indicate that cells in S and G2 phase at the time of drug administration can undergo mitosis even though a considerable prolongation of G2 phase was apparent. However, cells in G1 at the time of drug administration was arrested in that phase, whereas those cells in S or G2 were able to complete one mitosis before becoming arrested in the G1 phase. This blocking effect was reversible, and cells resumed proliferation at a normal rate shortly after the removal of these drugs.     

The cell cycle and its relationship to development in Dictyostelium discoideum.

When deprived of exogenous nutrients some amoebas of Dictyostelium discoideum do continue to progress through the cell cycle. There are two distinct periods when mitotic cell division occurs. Labeling studies show that during the first period, which begins at the onset of development and ceases at the first visible signs of aggregation (rippling), only those cells which are beyond a certain point in G2 at the initiation of development divide. The second period of mitotic activity begins at tip formation, reaches maximum activity at the grex stage, and ceases during early culmination. Significantly, examination of the development of amoebas harvested when in the stationary phase of growth (and thus arrested in G2) shows that these cells still undergo mitotic cell division during the second period but do not show any such division during the preaggregation phase. The extent to which increases in cell number can be taken to be indicative of mitotic cell division varies from one culture to another due to the presence of variable numbers of multinucleate cells which become mononucleate during the first 10 hr of development. However, when due allowance has been made for the existence of these cells in axenically growing amoebal populations, our data show that by completion of fruiting body construction there has been a doubling in cell number as a direct result of mitotic cell division. Nuclear DNA synthesis also occurs at two distinct periods during development, these coinciding with the periods of mitotic activity. However, since no more than 35% of the cells have undergone nuclear DNA synthesis by the end of the developmental phase, our results are inconsistent with the conclusion that all cells accumulate at a position in G2 at the time of aggregation. Our results do suggest, however, that mitotic cell division of a fraction of the cells may be an integral part of the developmental phase.