THE MECHANISM OF COLCHICINE INHIBITION OF MITOSIS

H3-colchicine of high specific activity (2.5 curies per mM) was prepared in order to study the mechanism of colchicine inhibition of mitosis in cultures of human cells, strain K.B. No direct effects on the duration of the cell cycle or macromolecular synthesis were demonstrable at a concentration of colchicine which completely inhibited mitosis. The radioactive compound was bound to the cells at a rate proportional to colchicine concentration. The binding appeared to be reversible since the radioactivity of the cells reached a maximum value for a given concentration and was slowly lost after resuspension of the cells in fresh medium. A suitable exposure to colchicine produced accumulation of metaphase-blocked mitoses after the colchicine was removed from the medium. An exposure of 6 to 8 hours at 10-7 M was sufficient to block essentially all the cells in metaphase, thus indicating that colchicine is bound to the majority of interphase cells. The data are in quantitative agreement with a mechanism involving reversible binding of colchicine to a set of cellular sites. Based on the correlation between the time of first appearance of blocked mitoses and the radioactivity per cell, it is suggested that if a critical fraction (3 to 5 per cent) of the sites are complexed, the cell is unable to form a functional mitotic spindle.

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Compartmented pyruvate in perfused working heart

AJP Heart and Circulatory Physiology ◽

10.1152/ajpheart.1985.249.3.h439 ◽

1985 ◽

Vol 249 (3) ◽

pp. H439-H449 ◽

Cited By ~ 11

Author(s):

R. Bunger

Keyword(s):

Specific Activity ◽

High Specific Activity ◽

Inhibitory Effect ◽

Quantitative Agreement ◽

Myocardial Substrate ◽

14Co2 Production ◽

The Mean ◽

Glycerol 3 Phosphate Dehydrogenase ◽

Perfused Hearts ◽

Working Heart

Pyruvate compartmentation and lactate dehydrogenase (LDH) were studied in isolated perfused working guinea pig hearts. The mean intracellular pyruvate (Pyr) contents increased with perfusate Pyr (0-2 mM) but varied only slightly with glucose (0-10 mM) and additional insulin (0.04-5 U/l), respectively. With 5-10 mM glucose plus 5 U/l insulin, but not with Pyr or lactate (Lac) as substrates, a near equilibrium between the LDH and the glycerol-3-phosphate dehydrogenase seemed to exist. Evidence for an inhibitory effect of Pyr on the activity of the LDH system of the perfused hearts was not obtained. With [U-14C]glucose as sole substrate, the specific activity of coronary venous Lac was near half that of precursor glucose. 14CO2 production was thus in quantitative agreement with rates of pyruvate oxidation that were determined as glucose uptake minus (Pyr + Lac) release. In contrast, with 0.2 mM [1-14C]Pyr plus 5 mM glucose, the ratio of 14CO2 production to specific activity of Lac overestimated Pyr oxidation judged from myocardial substrate balances and O2 uptake, respectively; here, at least three pools of [14C]HCO-3 and [14C]lac, respectively, were kinetically demonstrable during washout of trace amounts of 14C-labeled Pyr. Evidently, the specific activity of Lac was equivalent to that of mitochondrial oxidized Pyr provided [14C]glucose was the sole or major precursor of cellular pyruvate. However, exogenously applied [1-14C]Pyr of high specific activity seemed to induce intracellular formation of both a highly and lowly labeled Pyr; the latter Pyr compartment did not seem in ready equilibrium with the cell physiologically prevailing highly labeled Pyr pool.

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Accumulation of adrenocorticotropin secretory granules in the midbody of telophase AtT20 cells: evidence that secretory granules move anterogradely along microtubules

The Journal of Cell Biology ◽

10.1083/jcb.104.4.1047 ◽

1987 ◽

Vol 104 (4) ◽

pp. 1047-1057 ◽

Cited By ~ 29

Author(s):

J Tooze ◽

B Burke

Keyword(s):

Electron Microscopy ◽

Cell Cycle ◽

Mitotic Spindle ◽

Secretory Granules ◽

Thin Sections ◽

Large Numbers ◽

Golgi Region ◽

Interphase Cells ◽

Immunofluorescence Labeling ◽

Mitotic Cells

During the cell cycle the distribution of the ACTH-containing secretory granules in AtT20 cells, as revealed by immunofluorescence labeling and electron microscopy of thin sections, undergoes a cycle of changes. In interphase cells the granules are concentrated in the Golgi region, where they form, and also at the tips of projections from the cells, where they accumulate. These projections contain many microtubules extending to their tips. During metaphase and anaphase the granules are randomly distributed in the cytoplasm of the rounded-up mitotic cells. On entry into telophase there is a rapid and striking redistribution of the granules, which accumulate in large numbers in the midbody as it develops during cytokinesis. This accumulation of secretory granules in the midbody is dependent upon the presence of microtubules. The changing pattern of distribution of the secretory granules during the cell cycle fulfills the predictions of a model envisaging first that secretory granules associate with and move along interphase microtubules in a net anterograde direction away from the centrioles, and secondly that they do not associate with microtubules of the mitotic spindle during metaphase and anaphase.

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Effects of Heparin Oligosaccharides with High Affinity for Antithrombin III in Experimental Venous Thrombosis

Thrombosis and Haemostasis ◽

10.1055/s-0038-1657178 ◽

1982 ◽

Vol 47 (03) ◽

pp. 244-248 ◽

Cited By ~ 47

Author(s):

D P Thomas ◽

Rosemary E Merton ◽

T W Barrowcliffe ◽

L Thunberg ◽

U Lindahl

Keyword(s):

Antithrombin Iii ◽

Specific Activity ◽

High Specific Activity ◽

Factor Xa ◽

Blood Levels ◽

Thrombin Time ◽

High Affinity ◽

Very High

SummaryThe in vitro and in vivo characteristics of two oligosaccharide heparin fragments have been compared to those of unfractionated mucosal heparin. A decasaccharide fragment had essentially no activity by APTT or calcium thrombin time assays in vitro, but possessed very high specific activity by anti-Factor Xa assays. When injected into rabbits at doses of up to 80 ¼g/kg, this fragment was relatively ineffective in impairing stasis thrombosis despite producing high blood levels by anti-Xa assays. A 16-18 monosaccharide fragment had even higher specific activity (almost 2000 iu/mg) by chromogenic substrate anti-Xa assay, with minimal activity by APTT. When injected in vivo, this fragment gave low blood levels by APTT, very high anti-Xa levels, and was more effective in preventing thrombosis than the decasaccharide fragment. However, in comparison with unfractionated heparin, the 16-18 monosaccharide fragment was only partially effective in preventing thrombosis, despite producing much higher blood levels by anti-Xa assays.It is concluded that the high-affinity binding of a heparin fragment to antithrombin III does not by itself impair venous thrombogenesis, and that the anti-Factor Xa activity of heparin is only a partial expression of its therapeutic potential.

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Autoprothrombin C Activity from Prothrombin with Snake Venom or Trypsin

Thrombosis and Haemostasis ◽

10.1055/s-0038-1655440 ◽

1962 ◽

Vol 08 (03) ◽

pp. 425-433 ◽

Cited By ~ 1

Author(s):

Ewa Marciniak ◽

Edmond R Cole ◽

Walter H Seegers

Keyword(s):

Snake Venom ◽

Thrombolytic Agent ◽

Sodium Citrate ◽

Specific Activity ◽

High Specific Activity ◽

Citrate Solution ◽

Clotting Factors ◽

Activation Products ◽

Russell’S Viper ◽

Autoprothrombin C

SummarySuitable conditions were found for the generation of autoprothrombin C from purified prothrombin with the use of Russell’s viper venom or trypsin. DEAE chromatographed prothrombin is structurally altered and has never been found to yield autoprothrombin C and also did not yield it when Russell’s viper venom or trypsin were used. Autoprothrombin C is derived from prothrombin with tissue extract thromboplastin, but not in large amounts with the intrinsic clotting factors. With the latter thrombin and autoprothrombin III are the chief activation products. Autoprothrombin III concentrates were prepared from serum and upon activation with 25% sodium citrate solution or with Russell’s viper venom large amounts of autoprothrombin C were obtained, and this was of high specific activity. Theoretically trypsin is not a thrombolytic agent, but on the contrary should lead to intravascular clotting.

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