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The immunological action of RNA mononucleotides was studied in animal experiments. The most pronounced activation of macrophagal glycolysis urea cycle, oxidative phosphorylation, lysosomal hydrolases was induced by uridine 5'-monophosphate (5'-UMP) and guanosine 5'-monophosphate (5'-GMP); 5'-GMP also induced the maximum increase of the expression of FC gamma receptors. 5'-UMP ensured cell activation comparable with the total action of all mononucleotides. 5'-UMP and 5'-GMP, used in combination, produced the highest stimulating effect on macrophages, and the addition of low-active adenosine 5'-monophosphate (5'-AMP) to active 5'-UMP did not decrease the stimulating potency of the latter. The stimulating activity of sodium nucleinate exceeded that of all mononucleotides and their combinations. 5'-GMP and 5'-AMP induced the maximum activation of oxygen metabolism, evaluated by chemiluminescence, while 5'-UMP and cytidine 5'-monophosphate (5'CMP) proved to be inactive. The shift of the phosphate group to the third carbon atom or the production of the oligonucleotide 5'-UMP consisting of 5-15 nucleotides resulted in the appearance of the capacity for stimulating oxygen metabolism in macrophages. Their in vitro cultivation with 5'-GMP and 5'-AMP induced the maximum increase of cell spreading in comparison with other mononucleotides, while the maximum increase of phagocytosis was ensured only by 5'-UMP and 5'-GMP.2+ 5'-UMP and 5'-GMP enhanced nonospecific resistance to Salmonella typhi infection, and sodium nucleinate, to Pseudomonas pseudomallei and Pseudomonas mallei infections.

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A method of the Poisson fitting of flow cytometry data is suggested. The estimation of distribution parameters was carried out by the steepest descent method. The phagocytosis of fluorescent latex particles by murine macrophages was used as an experimental model. The results obtained by the Poisson fitting of cytofluorograms were in good agreement with the visual estimation of macrophage distribution with regard to the amount of uptaken particles. A high value of correlation index (more than 0.99) showed the validity of the suggested fitting method. The applicability of the method to cytofluorimetric analysis of human lymphocytes was successfully demonstrated.

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The interrelation between structural changes and oxygen consumption by the phagocyting macrophage was studied. The mean number of phagocyted particles was estimated by the method of stereological transformation. It is found that the uptake of yeast particles and CN- -nonsensitive oxygen consumption is related to the concentration of yeast cells in the incubation medium. A positive correlation was established between the oxygen consumption and the mean number of phagocyted particles. The results obtained may suggest that the "respiration burst" takes place in the contact area of the macrophage and the phagocyted material, and its extent probably depends on the surface of that contact area.

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Various complexes of phagocytes with adsorbed (I) and entrapped (J) particles are formed during phagocytosis. Method of stereological reconstruction is proposed that allows to demonstrate the actual distribution of these complexes on the basis of morphometric analysis of their ultrathin sections. The principle of the method lies on the probability simulation of section distribution for a given distribution of complexes and on the solution of the reverse problem by stepwise determination of the relative quantity of each complex type (from the most complicated to the most simple one, when I = 0 and J = 0). The stereological analysis of phagocytosing murine peritoneal macrophages revealed an absolutely different and more adequate kinetical picture of phagocytosis, as compared to the morphometric data.

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Morphometry and cytofluorometry of the kinetics of phagocytosis were applied to study the quantitative mechanisms of acidification in the area of contact between the plasma membrane of macrophages and Candida albicans yeast cells conjugated with fluorescein isothiocyanate. It was found by stereological transformation of the morphometry data that the main part of macrophages phagocytose the limiting amount of particles by minute 5-10. A good agreement was established between the cytofluorometric histograms and the stereological data. This made it possible to evaluate, using the calibration curve, the pH on the surface of the phagocytosed material based on the fluorescence quenching. As an advantage of the suggested comprehensive approach to the study of acidification, the authors stress the possibility of making structural-functional analysis of the rearrangements occurring in the intact phagocytosing cell.

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Activated peritoneal macrophages exhibiting phagocytosing capacity produced an electron-dense precipitate of formazan in contact sites of macrophage plasmalemma and phagocytosed yeast cells. No production of formazan occurred, when non-opsonized latex particles were ingested by macrophages. Formazan precipitation could be prevented by anaerobiosis but not by addition of cyanide.

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A new method is elaborated for detecting acidification in phagocytes on the ultrastructural level. The method is based on the reaction between Cu2+ and [Fe(CN)6]4- which form a pellet of cupric ferrocyanide in the neutral medium. It is possible to induce pellet formation under definitely determined pH values on adding different amounts of chelating agent (citrate) to the reaction mixture. The fine-grained electron dense pellet of cupric ferrocyanide persists throughout the whole subsequent procedure of fixation, dehydration and embedding of the biological material for electron microscopy. Data are presented on the degree of acidification and its localization in subcellular structures of the phagocyte during phagocytosis.

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A new electron-cytochemical method for detection of phagolysosomal structures in intact cells is proposed, which is more susceptible and simple in comparison with methods described elsewhere. The new method involves the addition of sodium beta-glycerophosphate and Pb(NO3)2 to the medium where phagocytes are incubated with phagocytic material. After the fusion of a phagosome containing the phagocytic material, sodium beta-glycerophosphate (the substrate for a marker lysosomal enzyme--acid phosphatase) and Pb2+ with primary lysosomes, hydrolysis of this substrate occurs with the eventual appearance of a deposit of Pb3(PO4)2.

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Using flow cytofluorimetry, acidification in macrophages was studied during phagocytosis of yeast cells Candida albicans conjugated with FITC. Two phases of acidification were found: one corresponding presumably to yeast cell absorption on the macrophage surface, and the other one being due to cell incorporation into phagolysosomes. The phases are characterized by pH values of 6.3 +/- 0.5 and 4.5 +/- 0.5, respectively. Addition of 10 mM NH4Cl cancelled the observed acidification, whereas the presence of cytochalasin B (2 . 10(-5) M) prevented the very process of acidification.

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The authors studied the effect of high-fat and high-protein diets on the activity of some enzymes of rat gastric mucosa parietal cells. It was found that the high-protein diet (40% of protein in terms of caloricity) causes an increase in the activity of both the mitochondrial enzymes of Mg2+-ATPase and HCO-3-ATPase and enzymes of the plasma membrane (K+-ATPase and cytoplasm (carboanhydrase). The high-fat diet (60%) of fat in terms of caloricity) produces different effects on the above-indicated enzymes: the activity of Mg2+-ATPase decreases, while that of HCO-3-ATPase rises; at the same time the activity of K+-ATPase and carboanhydrase diminishes. It was shown that the enzymatic systems responsible for hydrochloric acid secretion in the stomach show a delicate response to a qualitatively different nutrition.

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The effect of the thiamine deficient diet on the ultrastructure and secretory response of parietal cells of the rat gastric mucosa was studied. The control group consisting of fasting animals showed 60% of the parietal cells to be in a presecretory state. Subcutaneous administration of histamine to the control animals was accompanied by secretion of 512 +/- 128 mkekv H+ over 3 hours, with only 18% of the parietal cells remaining not involved in the secretory process. In thiamine-free diets the number of secreting cells in fasting animals amounted to 23%, rising to 35% after histamine administration. At the same time a three-fold decrease in the secretory response (170 +/- 21 mkekv H+ over 3 h) was observed. Thiamine deficiency in the diet is also accompanied by inhibition of the transketolase activity in the gastric mucosa homogenate, which varied from 0.706 +/- 0.032 mcmole (control animals) to 0.344 +/- 0.034 mcmole of sedoheptuloso-7-phsophate/h per 1 mg protein (test animals). Potentialities of the pentose cycle participation in secretion are discussed.

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The distribution of carbonic anhydrase, K+-ATPase and K+-phosphatase in the subcellular fractions of gastric mucosa was studied. It was found that 90% of carbonic anhydrase are localized in the hyaloplasm, whereas K+-ATPase and K+-phosphatase are predominantly localized in the microsomal fraction. Subfractionation of the microsomal fraction in a sucrose density gradient showed that the membrane-bound carbonic anhydrase (5% of total content) and K+-ATPase are bound to various cell organelles. It is concluded that carbonic anhydrase functions as an intracellular pH-stat and is not directly involved in proton generation by the cell.

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Electron--microscope and biochemical studies in rats and dogs showed that secretory membranes of parietal cells were in dynamic equilibrium with plasmalemma in activated secretion. During secretion, a transposition of secretory membranes occurs: 53.2% of tubulovesicular membranes transform into membranes of intracellular canaliculi and apical surface. K+--ATPace and carbonic anhydrase activities increased (55.5% and 62.9%, resp.). The system of secretory membranes and enzyme systems of parietal cells seem to take part synchronously in the process of gastric acid secretion.

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