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The molecular mechanisms of the inhibitory action of fructose- 2,6-bisphosphate (F-2,6-P2) on fructose-1,6-biphosphatase (FB-Pase-1), the key enzyme of gluconeogenesis, and the those of the activating action of F-2,6-P2 on phosphofructo-1-kinase (PFK-1), the key enzyme of glycolysis, NMR spectroscopy first provided direct evidence for the fact that F-2,6-P2 was involved in the regulation of the sedoheptulose cycle of a nonoxidative stage of the pentosephosphate pathway. Procedures were developed in measuring the levels of F-2,6-P2 in the cell of experimental animal tissues and human blood lymphocytes. Naturally different emergencies substantially affected the F-2,6-P2 system by triggering these or those mechanisms controlling the activity of enzymes of this system. Vanadium-containing compounds were demonstrated to have a positive action on carbohydrate metabolism in diabetic (streptozotocin-induced) rat hepatocytes.

Content of fructose-2,6-bisphosphate was drastically decreased (about 5-fold) in hepatocyte suspension obtained from rats with streptozotocin-induced diabetes, as compared with that of control animals. At the same time, activity of fructose-1,6-bisphosphatase was increased 2-fold in these hepatocytes. Incubation mixture of the hepatocytes from the diabetes impaired animals contained only low amounts of lactate.

The study was carried out on primary coculture Kupffer cells and liver fibroblasts of newborn and mature rats. At first Kupffer cells and liver fibroblasts were taken on the equal quantity. There were observed significant decrease of Kupffer cells quantity and fibroblasts death after 4-5 days in coculture. Mitotic and functional activity liver fibroblasts, adhesion of Kupffer cells were increased by the using of conditioned medium of newborn rat hepatocytes. The rise of mitotic activity of liver fibroblasts and quantity of nucleolus in their nuclei forestalls of the increase Kupffer cells size, appearing of Kupffer cells with some nuclei.

The monolayer culture of 3-day-old rat hepatocytes was treated by antimicrotubule agent colchicine after activation with isoproterenol. Alterations in the amount of lysosomes with the size of 1.0-2.5 and 3-6 mkm per cell were studied with light microscope after staining with the vital dye neutral red and after identification of acid phosphatase by plumbum precipitation in the frozen preparations. It was shown that isoproterenol application (20 microM) within 1.5 h did not change significantly the number of lysosomes sized 1.0-2.5 microns but stimulated the increase in the amount of lysosomes 3-6 mkm in size. The addition of 2.5 mM colchicine, after the isoproterenol treatment, decreases greatly the number of lysosomes with the size of 1.0-2.5 and 3-6 mkm. A considerable part of the large lysosomes can be resistant to the colchicine action. It is concluded that the microtubules play an important role in lysosome functioning.

Content of cyclic nucleotides was studied in primary culture of newborn rat heart cells after administration of the polyether-containing antibiotic omomycin (985-I, 16-diethyl indanomycin). Level of cGMP was increased within 10 min of the cells exposition with 4 micrograms/ml of the antibiotic, while the nucleotide content elevation was more significant in presence of 1-methyl-3-isobutyl xanthine (10(-4) M). When the exposition time was prolonged up to 20 min, content of cGMP returned to control values in absence of isobutyl xanthine and remained at the elevated level in presence of the drug. Content of cAMP was unaltered under these conditions. Omomycin may be used in experiments, where distinct increase of the cellular cGMP content is required.

Effects of insulin was studied in the monolayer hepatocyte cultures of new born rats under anoxic conditions. Insulin 10(-7) M stabilized lysosomal membranes obtained after 20 min anoxia. I hr incubation of hepatocytes with insulin in normal conditions, which was followed by anoxia, caused a significant increase of acid phosphatase activity in the fraction enriched with lysosomes. Insulin caused more distinct stabilization effect on lysosomal membranes when its action was prolonged. On the other hand, insulin caused more than 2-fold increase of cAMP content in hepatocytes within 2 min of exposition as compared with control cultures. When exposition of the cells with insulin exceeded 2 min, lowering of cAMP content was observed. These data appear to indicate that stabilizing effect of insulin was secondary towards cAMP level increase. Indirect connection between cAMP level, insulin stabilizing action and the state of lysosomal membranes were noted in the rat hepatocyte culture.

The monolayer cultures of newborn rats hepatocytes were treated with substrate starvation (20 min.) anoxia accompanied with substrate starvation (1 h.), following rehabilitation (1 h.) and cooling (4 degrees C, 30 min). After the neutral red staining we have examined the alterations in lysosomes quantity and the development of phagocytosis; we also tested the hepatocytes distribution due to the amount of the second lysosomes per cell. We have shown that a short time pathological treatment caused a decrease in large lysosomes (1-3 microns) amount in significant portion of hepatocytes. There were neither increase in phagosomes amount per cell nor alterations in the quantity of cells, containing phagosomes. However a long time pathological treatment caused the increase in phagosomes and cells. We propose that initial stage of lysosomal apparatus alterations, connected with reduction of CAMP level, is accompanied by a stamping of large lysosomes.

A direct observation of primary cultures of 3-day-old rat hepatocytes was performed with the help of time-consuming cinematography. The process of monolayer formation has been examined. In addition, the continuous preservation of bile canaliculi was observed. A significant increase in hepatocyte mobility was marked in 6-7-day cultures. The structural analysis of cultured cells during the modelling of anoxia with substrate deficiency revealed a succession in hepatocyte and fibroblast alterations. The hepatocyte destruction was accompanied with changes in the cell form, that become rounded, and in the appearance of small formations protruding on the cell surface. The fibroblast destruction appeared after cell contracture.

Two proteins were isolated from preparations of weakly bound nonhistone proteins, obtained from highly purified chromatin with 0.35 M NaCl. These two protein residues from chromatin RNP-particles have the buoyant density of 1.41 g/cm3 as shown by centrifugation in CsCl gradients and contain heterogeneous fast labelling nuclear RNA. According to SDS polyacrylamide gel electrophoresis the molecular weights of these proteins are 70000 and 43000 daltons.

Acid phosphatase activity and cAMP level were studied in primary hepatocyte culture of new born rats under conditions of anoxia and substrate deprivation (incubation of the cells in Hanks salt solution). Incubation of hepatocytes in Hanks salt solution within one hour under conditions of anoxia caused a significant increase in free (cytosolic) enzyme activity. Substitution of Hanks salt solution by normal tissue culture medium and reoxygenation after 1 hr anoxia resulted in a decrease of free acid phosphatase activity, whereas activity of the enzyme in lysosomal fraction was increased. Content of cAMP was decreased distinctly after 15 min incubation of hepatocyte culture under conditions of anoxia and substrate deprivation and was increased above control values within 5 min of reoxygenation and substitution of Hanks salt solution by normal tissue culture media. Addition of cAMP-containing liposomes to hepatocyte culture under these experimental conditions led to a decrease in free acid phosphatase activity and to an increase of the enzyme activity in lysosomal fraction. cAMP appears to modulate the lability of hepatocyte lysosomal membrane. The mechanisms involved in these processes are discussed.

Two discreet in size molecular classes of metabolically stable messenger-like RNA molecules with sedimentation coefficientes about 28S and 18S have been revealed in the nuclei of pigeon bone marrow cells. The structural pecularities of 28S RNA class were investigated more carefully. It was shown to constitute the largest fraction of nuclear messenger-like RNA and is characterized by a GC/AU ratio 1.13. As pulse-labeled nuclear RNA, 28S stable nuclear RNA hybridizes with the unique and rare repeated DNA, it does not contain poly(A)-sequences and is found in a form of RNP particles with density of 1.41 g/sm3, tightly bound to chromatin.

Acid phosphatase activity, ATP and cyclic nucleotide levels were studied in primary cardiomyocyte cultures of newborn rats under hypoxic conditions. It was shown that incubation of cardiac cells in Eagle's medium supplemented with 10% calf serum during hypoxia (5% O2, 5% CO2, 90% N2) caused an hour later a decrease in lysosomal acid phosphatase activity, whereas free enzyme activity increased. The exposure of cells in Hanks salt solution to one hour hypoxia resulted in a 3-fold decrease of ATP level and further increase in free acid phosphatase activity. It was found that cAMP and cGMP levels do not depend on the presence of hypoxia. The mechanisms involved in the regulation of lysosomal membrane state are discussed.

The primary cultures of 3-day old rats heart myocytes were used for studying hypoxia. The cells were gassed for 1 or 2 hours with 100% N2 or with the mixture of 90% N2, 5% CO2, 5% O2. The cells' morphology was tested by the light microscopy. The contractility of the cells was lost after oxygen deprivation. But it was reversible when the cells were exposed to 5% O2 for an hour and then were returned to the normal conditions. Oxygen deprivation changed the cell's morphology so that vacuolization, bubbling, contracture, exfoliation of the cell membrane from the glass surface could be observed. The number of the cells with morphological alterations increased when the content of oxygen in the gas mixture was lowered and the time of gassing was prolonged. The authors assume that the primary culture of the myocardial cells is a suitable model for studying the metabolic patterns of reversible injuries caused by one hour hypoxia (5% O2).

A primary culture of hepatocytes from new born rats was used as an experimental model suitable for studies of the cell lysosomal apparatus and of molecular mechanisms involving cyclic nucleotides for initiation of lysosomal response to extremal conditions. Substrate deficiency and anoxia of the cultivated hepatocytes were found to cause distinct alterations in the state of lysosomal apparatus accompanied by an increase in free activity of acid phosphatase.

Using isolated nuclei of salivary grand cells of Chironomus plumosus larvae, it was revealed that the integrity of nuclei was a necessary condition for realization of protein synthesis in the nuclei. The involvement of nuclear particles in the above synthesis is supposed.

This review deals with the analysis of the conflinting situations that appear during the study of the translational apparatus in cell nuclei. Experimental approaches used to unravel this problem are surveyed.

Activity of some lysosomal enzymes was studied in the primary heart cell culture of newborn rats under conditions of hypoxia and in isolated ischemic perfused heart, in liver and heart tissues of dogs after traumatic shock and after experimental myocardium infarction as well as in liver tissue of rats after 10 min experimental heart arrest and during the postresuscitation period. Decrease in cAMP and elevation in cGMP levels, impairment of the adenylate cyclase hormonal regulation, decrease in activity of cAMP-dependent protein kinases were shown to correlate distinctly with an increase of acid hydrolases free activity in liver and heart cell cytoplasm of dogs with traumatic shock and with experimental myocardium infarction. Free activity of acid phosphatase was increased in primary heart cell culture under conditions of hypoxia and in isolated ischemic perfused rat heart. Intraperitoneal administration of cAMP-containing liposomes into the rats with 10 min heart arrest led to stabilization of liver lysosomal membranes within 1 hr of 4 hrs of the post-resuscitation period. The mechanisms involved in stabilization of lysosomal membranes are discussed.