Bacterial beta-galactosidase and human dystrophin genes are expressed in mouse skeletal muscle fibers after ballistic transfection.

Ballistic transfection, based on cell and tissue bombardment by the tungsten and gold microparticles covered with the gene DNA, was used for the delivery of a bacterial beta-galactosidase and a full-length cDNA copy of the human dystrophin genes into mouse skeletal muscles. CMV-lacZ, SV40-lacZ, LTR-lacZneo and full-length cDNA dystrophin (pDMD-1, approximately 16 kb) in eukaryotic expression vector pJ OMEGA driven by mouse leukaemia virus promotor (pMLVDy) were used throughout the studies. Musculus glutaeus superficialis of C57BL/6J and quadriceps femoris of mdx male mice were opened surgically under anesthesia and bombarded by means of the gene-gun technique originally developed by us. Different mixtures of gold and tungsten particles at ratios of 4:1, 1:1, 1:4 were applied. X-gal assay revealed marked beta-gal activity, both in total muscles and whole muscle fibers on histological sections, up to three months after transfection. The most intensive staining was observed after SV40-lacZ delivery. No staining was detected with LTR-lacZneo DNA as well as in untreated muscles. The higher tungsten particle concentration in the bombardment mixture correlated with more intense X-gal staining. At the gold/tungsten ratio of 1:4 the microparticles penetrated the musculus glutaeus superficialis and transfected the underlying musculus glutaeus medius as well. Immuno-cytochemical assay for human dystrophin revealed dystrophin positive myofibers (DPM) in the bombarded area up to two months after transfection. The proportion of DMP varied from 2.5% on day 17 up two 5% on day 60 after bombardment compared to only 0.5% in the control mdx mice. These results suggest the applicability of particle bombardment for gene delivery into muscle fibers.

[The characteristics of the biological action of low doses of irradiation]. / Osobennosti biologicheskogo deistviia malykh doz oblucheniia.

Biochemical, biophysical and functional properties of the genetic and membrane apparatus of the cell were considered activity and regulatory properties of the membrane and cytosolic enzymes of organs and tissues of mice exposed to radiation in the wide range of doses of 6 to 1800 mGy with the radiation intensity of 4.1 x 10(-3) and 41 x 10(-3) mGy/min. It was shown that the dose-dependence of changes in the investigated properties is of non-linear polymodal (bimodal) nature. The value of the maximum and the dose at which the latter was observed depend on the object's nature, radiation intensity and time passed after irradiation. An essential factor is that sensitivity of molecules, cells, organs and animals exposed to low-dose radiation to other damaging effects changes. The explanation is given in terms of the changes in the relation between the quantity of damages and the activity of reparation systems induced by low-dose irradiation.

[Differentiation of muscle fibers in mdx mice after ballistic transfection of cDNA of the human dystrophin gene]. / Differentsirovka myshechnykh volokon myshei MDX posle ballisticheskoi transfektsii kDNK gena distrofina cheloveka.

Changes in morphological dimensions of MDX mouse myofibres in M. rectus femoris were recorded after ballistic transfection (BT) with pHSADys and pVMMDys plasmids containing cDNA of the full-length human dystrophin gene. The dystrophin expression was observed by an immunomorphological procedure with P6 antibody and PAP method. Dystrophin positive (dyst+) myofibres were divided into two types, with a typical dystrophin expression under sarcoplasma membrane and an atypical expression through the whole sarcoplasm, respectively. The share of atypical dyst+ myofibres was seen to rise during the experiment from 27%, at 2-3 weeks after BT, up 84% by 2 months after BT. The atypical dyst+ myofibres usually underwent destruction. At the same time, the share of entire dyst+ myofibres decreased from 17 to 2-5% by 2 months. Morphological dimensions of the myofibres (square in mkm2, perimeter, smallest and largest diameters) were calculated with the help of computer analyser. The middle square of both types of dyst+ myofibres was larger than that of dyst- myofibres, both in BT target M. rectus femoris and in the same contralateral muscle, but never exceeded the value of middle square of C57B1 mouse myofibres in the same muscle. The form of dyst+ myofibres was not modified by the dystrophin expression. The nuclei of dyst+ myofibres remained in the central region of sarcoplasm. A conclusion is made that BT of human dystrophin gene inside MDX mouse myofibres allows dystrophin gene expression and enlargement of the dyst+ myofibres. Dystrophin expression is not able to induce a complete and stable differentiation of striated muscle of adult MDX mice.

[The role of ethanol during gamma-irradiation of water-salt DNA solutions]. / Rol' étanola v protsesse gamma-oblucheniia vodno-solevykh rastvorov DNK.

In this work we had assumed to clarify the role of the water structure in the process of the radiation damage of DNA. It is known that the aliphatic alcohols stabilize the water structure until the critical concentration. In this connection we have analyzed the changes of the long- and short-distance interactions in the DNA irradiated in the water-ethanol solutions with various concentration of the ethanol and ions of metal. It was shown that as the water structure becomes more stable the conformational damages in the DNA are decreasing and finally at the some concentration of the alcohol in the irradiated solution the damages disappear. By the achieving of the alcohol concentration which lead to the destruction of the water structure the radiation results in the same changes of the considered parameters as in the case of the DNA irradiated in the water-salt solution with ethanol. The analyses of the experimental data allow us to conclude that the radiation destroys the structure of the water and thus helps the positive ions from the solution to come nearer to the DNA, to say, the radiation reduce intramolecular electrostatic interactions. This concept allows us to explain the observed changes of intrinsic viscosity and the difference in the polarizabilities of the DNA in the process of the radiation damage.

[Comparative study of the action of Vibrio cholerae enterotoxin, thermolabile Escherichia coli toxin and a Salmonella typhimurium filtrate on the small intestine of laboratory animals]. / Sravnitel'noe izuchenie deistviia énterotoksina Vibrio cholerae, termolabil'nogo toksina Escherichia coli i fil'trata Salmonella typhimurium na tonkuiu kishku laboratornykh zhivotnykh.

The results of the comparative study of the effects produced by V. cholerae enterotoxin E. coli thermolabile toxin (TL-toxin) and S. typhimurum 415 filtrate on the loops of the small intestine of rabbits and C57BL/6 mice are presented. The results obtained in mice were considerably more uniform than those obtained in rabbits. For the first time the dose-effect curve for V. cholerae enterotoxin and TL-toxin, obtained on both animal species, was found to be bell-shaped. The concentration relationship for dibutyryl-cAMP proved to be similar in character. The authors suggest that these effects are based on the specific relationship between the cellular processes, controlling the transport of electrolytes and water, and the level of cAMP.

[The efficacy of exogenous and endogenous interferonization in preventing influenza]. / Effektivnost' ékzogennoi i éndogennoi interferonizatsii pri profilaktike grippa.

In this work the results of using interferon (IFN), Dibasol and the combination of these preparations for the urgent prophylaxis of influenza and acute respiratory diseases (ARD) among the employees of the Gamaleia Research Institute of Epidemiology and Microbiology (USSR Acad. Med. Sci.) are summarized. Reaferon and Dibasol decrease ARD morbidity 2 times and leukocytic IFN decreases it 1.4 times, while the combined administration of Dibasol and IFN has proved to be ineffective. Reaferon is mainly a prophylactic remedy; it has been found to bring about almost no decrease in the number of patients at the peak of morbidity, while pronouncedly decreasing it in two weeks after the administration of the preparation. Dibasol has a curative effect, sharply interrupting the beginning rise of morbidity. Reaferon normalizes the characteristics of the IFN status, decreasing the amount of circulating IFN and enhancing the capacity of leukocytes for producing alpha-IFN and gamma-IFN. For the prophylaxis of respiratory infections the use of Reaferon is advisable 3-4 weeks prior to the beginning of the epidemic and then, when the first cases of infection are registered, the course of prophylaxis with Dibasol should be carried out.