[The study of molecular mechanisms of action of natural amino acids and serotonin on adenylyl and guanylyl cyclases of the ciliates].

It has been previously shown that some amino acids and their derivatives are capable of regulating the activity of adenylyl cyclase (AC) and guanylate cyclase (GC) in free-living ciliates Dileptus anser and Tetrahymena. The aim of this work was to study the molecular mechanisms of action of methionine, tyrosine, alanine and neurohormone serotonin on the activity of enzymes-cyclases and the identification of their specific receptors in D. anser and T. pyriformis. Methionine and serotonin significantly increased the basal AC activity in both ciliates, and the AC effect of serotonin in T. pyriformis was carried out with the participation of Ca2+-dependent form of AC and heterotrimetic G proteins. AC stimulating effect of tyrosine and alanine was expressed weakly and only detected in D. anser. Serotonin is both ciliates and alanine in D. anser stimulated GC activity, whereas methionine and tyrosine had no effect on GC. Methionine and serotonin bind to surface receptors of the ciliates with high affinity. K(D) for [methyl-3H] methionine binding to D. anser and T. pyriformis were 7.5 and 35.6 nM, and for [3H] serotonin binding were 2.7 and 4.7 nM, respectively. Alanine and tyrosine bind to the ciliates with low affinity. Thus, ciliates D. anser and T. pyriformis have chemosignaling systems regulated by amino acids and their derivatives and including the enzymes with cyclase activity. There is an assumption that these systems are similar to hormonal signaling systems of higher eukaryotes and are their predecessors.

[First experience with intrasurgical brachytherapy in the combined treatment for locally advanced laryngeal cancer].

Results of combined radiosurgical treatment with larynx preservation, and pre- and postoperative distant external irradiation, were evaluated. Forty-eight patients with locally advanced laryngeal cancer received single-step individually-tailored tissue application, followed by high-dose brachytherapy. Discriminative function of the larynx was restored in all patients within 2 days involving swallowing disturbances in 8%. Breathing function was restored in 98% within 2 days to 6 months after completion of treatment. Three-year relapse-free survival was 95.8%.

[Criteria for multispiral X-ray computed tomography for the choice of extent of surgery on the tumour spreading outside the larynx].

The authors summarize the experience with the treatment of 38 patients with disseminated laryngeal tumours (stages III and IV). All the patients underwent comprehensive medical examination using a variety of methods. Their advantages and drawbacks are analysed with reference to diagnostic value, potential for the detection of metastases, and possibility to use for the choice of extent of surgical intervention. Criteria for multispiral X-ray computed tomography have been developed to be applied to the identification of tumour spread outside the larynx and invasion of its cartilaginous structures.

[Up-to-date methods of tomographic diagnostics for the evaluation of effectiveness of the treatment of laryngeal and laryngopharyngeal cancer].

Currently employed methods of tomographic diagnostics for the evaluation of effectiveness of the treatment of laryngeal and laryngopharyngeal cancer are described. The authors underscore that increasingly more attention tends to be given to magnetoresonance imaging (MRI) and especially X-ray computer tomography (CT) due to their wide application and high informative value whereas nuclear medicine techniques, such as single-photon emission CT using thallium-200, 11C-methionine and (18)F-positron emission tomography, are considered to be less important despite their great diagnostic potentialities. It is emphasized that X-ray CT with intravenous administration of contrast material should be regarded as the method of choice for the evaluation of larynx and pharynx conditions following treatment of their tumours. MRI should be reserved for patients in whom X-ray CT with contrast enhancement proves impracticable.

[The role of multispiral computed tomography in combined diagnostics of laryngeal cancer for the choice of treatment strategy and extent of surgical intervention].

This paper summarizes experience with the treatment of 50 patients with laryngeal cancer at different clinical stages of the pathological process. All the patients underwent comprehensive medical examination. Advantages and disadvantages of each method were evaluated in terms of efficiency for the assessment of the extent of laryngeal tumour and identification of metastases followed by the choice of treatment strategy and extent of surgical intervention.

[Functional characteristics of calcium-sensitive adenylyl cyclase of ciliate Tetrahymena pyriformis].

Calcium-sensitive forms of adenylyl cyclase (AC) were revealed in most vertebrates and invertebrates and also in some unicellular organisms, in particular ciliates. We have shown for the first time that calcium cations influence the AC activity of ciliate Tetrahymena pyriformis. These cations at the concentrations of 0.2-20 microM stimulated the enzyme activity, and maximum of catalytic effect was observed at 2 microM Ca2+. Calcium cations at a concentrations of 100 microM or higher inhibited the AC activity. Calmodulin antagonists W-5 and W-7 at the concentrations of 20-100 microM inhibited the catalytic effect induced by 5 microM Ca2+ and blocked the effect at higher concentrations of Ca2+. Chloropromazine, another calmodulin antagonist, reduced Ca2+-stimulated AC activity only at the concentrations of 200-1000 microM. AC stimulating effects of serotonin, EGF and cAMP increased in the presence of 5 microM Ca2+. AC stimulating effects of EGF, cAMP and insulin decreased in the presence of 100 microM Ca2+, and AC stimulating effect of cAMP decreased also in the presence of calmodulin antagonists (1 mM). At the same time, stimulating effect of D-glucose in the presence of Ca2+ and calmodulin antagonists did not change essentially. The data obtained speak in favor of the presence of calcium-sensitive forms of AC in ciliate T. pyriformis which mediate enzyme stimulation by EGF, cAMP, insulin, and serotonin.

[The role of X-ray computed tomography in the detection of metastatic lesions in the regional lymphatic nodes in patients presenting with primary diagnosis of laryngeal and laryngo-pharyngeal cancer].

The objective of this study was to evaluate the clinical significance of X-ray multispiral computed tomography with the use of internal contrast for the clinical diagnostics of metastatic lesions in the regional lymphatic nodes of patients presenting with primary diagnosis of squamous cell cancer of the larynx and laryngopharynx. A total of 78 patients with laryngeal cancer and 22 with the primary diagnosis of laryngo-pharyngeal cancer were examined by X-ray computed tomography that revealed metastatic lesions in the regional lymphatic nodes. A detailed description of the most characteristic metastatic lesions is presented, viz. (1) active accumulation of the contrast agent in the peripheral parts of the lymph nodes, (2) the presence of a central necrotic zone, (3) the indistinct outer contour of the affected lymphatic nodes, and (4) compaction of the fatty tissue surrounding them. The use of computed tomography allowed localization of the affected nodes to be correctly determine in all the examined cases. Moreover, the method made it possible to evaluate the extent of the carcinogenic process and its clinical stage.

[Change of mating type induced by actinomycin D in the ciliate Dileptus anser].

Effect of actinomycin D on the expression and inheritance of mating types (MTs) in mature laboratory clones of the ciliate Dileptus anser was studied. Each mature clone of these ciliates isolated from natural population appears to belong to one of the three complementary MTs - MT I, MT II or MT III. Its MT does not vary in the course of further laboratory cultivation of the clone in a series of vegetative generations. However, after treatment with actinomycin D (15 microg/ml, 3 days), such clones became hereditarily unstable for their MTs. At weekly testing for MT over the course of at least 15 weeks after treatment (which corresponds to more than 100 cellular divisions), many subclones of the treated clone reversibly changed their MT for another, and (or) showed temporary state of immaturity or partial maturity. These data testify in favour of our hypothesis of epigenetic MT determination in D. anser. Considering that actinomycin D induces heritable destabilization of some characters in amebas Amoeba proteus which is obviously of epigenetic nature, this antibiotic can possibly be regarded as "epimutagen".

[Multispiral computed tomographic semiotics of laryngeal cancer].

Multispiral computed tomography (MSCT) with intravenous bolus contrasting is a currently available method for radiodiagnosis of laryngeal cancer. MSCT is of much higher informative value in estimating the extent of a tumorous lesion than the traditional radiodiagnostic techniques: linear tomography, lateral X-ray study, roentgenoscopy and roentgenography of the laryngopharynx and esophagus with barium meal.

[Mating types in the ciliate Dileptus anser. Inheritance and genetic determination].

Hybridological analysis of mating types (MTs) has been first made for the lower ciliate Dileptus anser. Clones of an initially unknown genotype belonging to three MTs (MT I, MT II and MT III), characteristic of D. anser, were isolated from natural reservoirs and further used for crosses. In one group crosses, synclonal inheritance and typical Mendelian behaviour of the character were observed over sexual generations of ciliates. The results suggest that MTs in D. anser may be directly controlled by a single mat locus with three alleles showing peck-order dominance (mat1 > mat2 > mat3). In other words, cells with mat1/mat1, mat1/mat2 and mat1/mat3 genotypes belong to MT I, those with mat2/mat2 and mat2/mat3, and the mat3/mat3 belong to MT II and MT III, respectively. Sexually mature exconjugant clones stably retain their MTs corresponding to their genotypes on vegetative reproduction. The progeny of other group crosses showed various deviations from typical Mendelian behaviour of the character. In some cases, standard Mendelian ratios were more or less violated. Most typical was instability of differentiation for MT in maturing exconjugant clones. Shortly after their maturation, the majority of clones change their MT, rather frequently more than once, although the finally established MT is stably inherited afterwards, during vegetative reproduction. When unstable, exconjugant clones can successively express two or even three MTs characteristic of this species, including MTs that should not have been expected on the basis of parental genotypes available in a given cross. It looks likely that the mat locus in D. anser is complex and multipotential; it is inherited as a whole providing for expression of any MT characteristic of the species (in this respect bearing similarity with Tetrahymena thermophila). Other mechanisms, epigenetic in particular (Nanney, 1958), determine the final expression of one of the three MT potentialities by a given exconjugant clone. Stable, persistent functioning of these mechanisms ensures a stable differentiation for MT and Mendelian behaviour of the character in sexual generations and in crosses. Any disturbances in differentiation control may trigger MT instability in maturing exconjugant clones and violation of regular Mendelian behaviour.

[Mating types in the ciliate Dileptus anser. Shortening of maturation period after micrurgical bisection of exconjugant cells]. / Tipy sparivaniia u infuzorii Dileptus anser. Uskorenie polovogo sozrevaniia posle mikrurgicheskoi fragmentatsii ékskon"iugantnykh kletok.

In many ciliates, young exconjugant clones demonstrate sexual immaturity: they are not able to conjugate with mature cells of complementary mating types (MTs). After several scores of cell divisions, a short period of adolescence (partial maturity) commonly occurs followed by maturation, after which these cells are able to conjugate with cells of other, i.e. complementary, MTs again. Tavrovskaja (1981) reported a significant reduction in the maturity period in Dileptus anser clones, grown from exconjugant ciliates regenerated from small cell fragments. To verify this, in the present study exconjugant D. anser cells were bisected with glass needle after 3 to 4 cell divisions following conjugation. The same procedure was performed with survived and regenerated cells on the 2nd and 3rd days. The clones thus obtained were cultivated, and their ability to mate with each of the three standard clones of MT I, II and III was tested week by week. Indeed, in 22 F1 clones from cross N 7C (MT I) x N 2 (MT II) the immaturity period was reduced 1.4-3.4-fold (2.18 in average) after a threefold bisection, as compared with that in intact subclones. Similarly, in 27 F1 clones from cross N 20 (MT I) x N 14 (MT II) this period was shortened 1.6-3.0-fold (2.19 in average). 12 of these clones showed a 0.9-2.4-fold (1.53 in average) reduction after a single bisection, and 1.6-2.8 (2.12 in average) after a threefold operation. Thus, micrurgical fragmentation of young exconjugant cells can be used to accelerate maturation in D. anser.

[Mating types in the ciliate Dileptus anser. Genetical instability in the mating type system]. / Tipy sparivaniia u infuzorii Dileptus anser. Geneticheskaia nestabil'nost' v sisteme tipov sparivaniia.

In F1 and F2 from a cross between two clones of Dileptus anser isolated from natural sources (MT 1 x x MT III; MT = mating type), along with "normal" clones, many clones were observed demonstrating abnormal phenotype with respect to the MT-character. Irregular features of the latter were as follows: a) a delay in maturation; b) temporary reversion to immature or adolescent state, which means instability of maturity state; c) expression of MT I and MT III, rather than MT II as in properly matured clones; d) changes in MT (i.e., MT instability); e) appearance of totally unexpected MTs in terms of the scheme of genetic control of MTs in D. anser previously suggested by Afon'kin and Yudin (1987)--e.g., of all three MTs in F1 from the initial (analysing!) cross. Amazingly, these abnormal D. anser clones closely resembled some selfer-clones of Tetrahymena pigmentosa, previously reported as an example of genetic instability in the ciliate MT system (Simon, 1980; Simon, Orias, 1987).

[Re-expression of various i-antigens in Dileptus anser after temporary transformation of serotype]. / Vozobnovlenie sovmestnoi ékspressii raznykh i-antigenov infuzoriiami Dileptus anser posle vremennoi transformatsii serotipa.

RNP particles containing 20S prosomes (alpha RNP) isolated from human epidermoid carcinoma cell line A-431 are shown to posses strong and regulated endonuclease activity specific for high-molecular-weight RNA, particularly, specific mRNAs. Furthermore, alpha-RNP destabilize the 3'-untranslated regions of c-myc mRNA, creating a specific cleavage pattern. Cleavage point within Alu sequence in high-molecular-weight RNA has been localized by primer-extension method. This RNase activity is induced under the action of EGF. alpha-RNP involvement in the coordinated control of processing and stability of specific messenger RNA molecules is suggested. The endoribonuclease activity of alpha-RNP can represent a link between EGF signalling pathway and RNA processing and degradation.

[Mode of serotype inheritance in exoconjugant progeny of the ciliate Dileptus anser]. / Nasledovanie serotipov v ékskon''iugantnom potomstve infuzorii Dileptus anser.

Two clones of Dileptus anser, originally isolated from natural reservoirs and referred to below as B and D clones, were found to display different serotypes, when cultured under identical laboratory conditions. On being tested with two different polyclonal rabbit immune sera against each particular clone (the classic immobilization test) these clones showed no cross-reaction. At a standard dilution (1:50) and at a standard exposure time (4 h), either of the two immune sera immobilized 100% or commonly 0% of homologous and heterologous clone cells, respectively. In addition, the difference in serotypes was confirmed by the immunofluorescence analysis. By crossing (conjugation) between B (mating type I) and D (mating type III) cells, exconjugant F1 clones were obtained. Their serotypes were then tested (the same immobilization test) with antisera against both the "parental" clones: some clones were tested before their sexual maturation in ca. one month after conjugation, while others were examined in approximately 4 months after conjugation, i.e. after reaching maturity. Each of the F1 clones could react with both immune sera, which means that they possessed the intermediate, "hybrid" phenotype. Five different F1 clones were selected, and each of them was back-crossed to both "parental" clones, B and D. We succeeded in raising 25 exconjugant F2 (B1, to be more exact) clones from F1 x B crosses and 26 clones from F1 x D crosses. The conventional testing of these clones in 5-10 weeks after conjugation provided quite unexpected results, since among them no segregation for "parental" serotypes was observed. Each of the 51 tested clones demonstrated the "hybrid" serotype--seemingly the same as that of F1 clones. Such a non-Mendelian inheritance of the character is hardly to explain from the standard, canonical assumptions on the genetic control of serotype difference between original "parental" clones (different alleles in one locus? different loci?). Also it does not seem likely that the absence of segregation could result from differential survival of various phenotypes in F2 (although the total viability of exconjugant clones appeared rather low). The above data obviously need further confirmations and experimental analyses. We attempt to discuss the obtained results in terms of the epigene hypothesis (Tchuraev, 1975) and in relation to the epigenetic control of serotype expression in species of the Paramecium aurelia complex and in Tetrahymena thermophila, which are "the chosen few" subjects in ciliate genetics.

[Role of computerized tomography in stenoses and other diseases of the trachea and main bronchi]. / Rol' komp'iuternoi tomografii pri stenozakh i drugikh zabolevaniiakh trakhei i glavnykh bronkhov.

Fifty-one patients with stenosis of the upper portion of the tracheobronchial tree of various origins were examined; the sensitivity of computer-aided tomography (CAT) in the diagnosis of stenoses of the trachea and the main bronchi was found to be 95.83%, specificity 98.31%, and accuracy 93.87%. CAT is superior to other methods of investigation (common tomography, tracheobronchoscopy) in the detection of changes in the tracheobronchial walls and the tissues adjacent to the trachea and the main bronchi. CAT is particularly valuable for the diagnosis of the causes of compression stenosis. The method permits specification and recognition of the type of involvement in traumatic injuries (6 cases), tracheoesophageal fistulae (6 cases), post-tuberculous processes (3 cases). CAT examinations are truly valuable in the diagnosis of osteoplastic tracheobronchopathy (3 cases), some other rare diseases (tracheobronchomegaly--1 case, scleroma--1 case, amyloidosis--2 cases). The authors recommend the optimal algorithm of examinations of patients with diseases of the upper portions of the tracheobronchial tree: roentgenoscopy--roentgenography--CAT--tracheobronchoscopy.

[Diagnosis of tumors of the trachea and main bronchi by computerized tomography]. / Diagnostika opukholei trakhei i glavnykh bronkhov c pomoshch'iu komp'iuternoi tomografii.

A total of 104 patients with tumors of the upper portions of the tracheobronchial tree were examined. The tumors were malignant in 66 cases (53 primary and 13 secondary ones) and benign in 38 cases. The sensitivity of CT in the diagnosis of blastomas of the trachea and the main bronchi made up 96.12%, specificity 86.15%, and accuracy 91.6%. CT permitted the detection of the tumor, specification of its site and size, definition of its contour, imaging of a non-transparent portion and of relationships with the adjacent organs and tissues, assessment of the status of the walls (by detecting dense, thickened, or deformed sites) and regional lymph nodes.