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Biomedical subjects

J J Sedmak

Publications and source records attributed to J J Sedmak.

10 recordsLinked to original sources
PubMed1992

Cytotoxicity of Clostridium difficile toxin A for human colonic and pancreatic carcinoma cell lines.

The use of bacterial exotoxins may constitute novel adjuncts to treatment of gastrointestinal tract malignancies. Clostridium difficile toxin A was evaluated for its cytotoxic effect in vitro on 24 human cell lines and strains including carcinomas of the colon, pancreas, prostate, lung, breast, and lymphoid malignancies, as well as nonmalignant tissues. All nine colon and five pancreas cell lines were extraordinarily sensitive to the cytotoxic effect of Clostridium difficile toxin A at very low concentrations. This effect, which occurred rapidly and was dose dependent, was observed in all cells of seven colon and two pancreas cell lines at concentrations as low as 1-5 ng/ml (10(-12) to 10(-11) M), whereas cells derived from other sites required 60 to greater than 500 ng/ml to achieve an equivalent effect. The data suggest that Clostridium difficile toxin A may have potential therapeutic value in the treatment of some gastrointestinal tract cancers.

Bacterial Toxins↗
PubMed1990

Actin and lamin comprised filaments in the nuclei of Chinese hamster ovary cells affected with Clostridium difficile enterotoxin A.

The major change in the ultrastructure of Chinese hamster ovary (CHO) cells treated with Clostridium difficile enterotoxin A was the formation of bundles of filaments in the nucleoplasm. The filaments appeared after 2.5 h and disappeared by the fourth hour of incubation with the toxin. To partially characterize these filaments, laser diffractometry and ultrastructural immunocytochemistry were employed. The bundles consist of coiled filaments of about 11 nm and 16 nm diameter and a main long axial periodicity of about 26 nm. Postembedding immunogold labelling with monoclonal antibodies demonstrated that these intranuclear filaments are comprised mainly of actin, but also contain lamin and vinculin. Between 2.5 and 3 h of incubation with enterotoxin A, the lamins were located close to the nuclear envelope, but they were also consistently found scattered in the cytoplasm and nucleoplasm. Such an unusual distribution of lamins might mediate the adverse effects of enterotoxin A on nuclear organization and function and thus cause the eventual death of CHO cells.

Actin Cytoskeleton↗
PubMed1986

Mouse beta-interferon reduces RNA efflux from isolated nuclei.

Treatments of isolated L929 cell nuclei with MuIFN-beta reduced the energy-dependent efflux of 3H-RNA from these nuclei in a dose-dependent manner. These observations provide evidence for a direct physiological effect of IFN on the cell nucleus.

Animals↗
PubMed1978

Thermal and vortical stability of purified human fibroblast interferon.

The loss of biological activity upon heating or agitation of human interferons is markedly altered by changing their aqueous environment. Low pH significantly stabilizes liquid fibroblast interferon at 68 degrees C and 37 degrees C whereas chaotropic salts stabilize at 68 degrees C but not at 37 degrees C; this anomalous result may be due to reactivation of biological activity at the higher temperature. The concentration of extraneous proteins influences the apparent thermal stability at any temperature and pH; thus, interferon was not stable even at low pH at protein concentrations less than 5 microgram/ml. Solutions of partially purified fibroblast interferon can be inactivated by mechanical stress; the addition of proteins or nonionic detergents prevents such inactivation. Freeze-dried preparations show the greatest thermal stability. The use of high-temperature, accelerated storage tests makes it possible to predict the shelf-life of freeze-dried interferon.

Drug Stability↗
PubMed1977

Effect of chaotropic salts and protein denaturants on the thermal stability of mouse fibroblast interferon.

Altering the aqueous environment, especially with agents that affect hydrogen bonds, markedly affects the stability of mouse L cell interferon. Low pH stabilizes interferon whereas high pH labilizes it; heavy water further enhances interferon thermostability at pH 2 but not at pH 9. Exposure to the protein denaturants, 4 M-guanidine hydrochloride and 6 M-urea, significantly decreases the activity of interferon at pH 2 and pH9; however, the residual interferon activity is relatively thermostable. Certain chaotropic salts protect interferon against thermal destruction, and in terms of effectiveness, their sequence is in the order SCN- greater than 1- larger than or equal to Cl- = CiO4- - Br- greater than NO3-. Interferon becomes more stable to heat as the NaSCN concentration is increased from 0-25 M to 2-0 M. Molecular sieve chromatography of interferon in the presence of 1-5 M-NaSCN at pH 7 shows a shift in its apparent mol. wt. from 25000 to 42000. Unlike most proteins, the unfolded conformation of interferon appears to be more stable to heat than the molecule with a smaller Stokes' radius.

Deuterium↗
PubMed1977

Stabilization of interferons.

It is now obvious that there are a number of factors which can influence the stability of interferons. Since human fibroblast interferon is especially unstable both to heat and mechanical stress, the use of stabilizing conditions such as low pH or thioctic acid should facilitate purification and concentration of this interferon. Leukocyte interferon, on the other hand, seems to be more stable even when highly purified; however, losses in activity can occur in very dilute solutions containing low concentrations of protein. Inactivation during storage of interferons may be avoided by using freeze-dried preparations. In order to generate large quantities of purified, stable interferons for clinical use, there is a need to establish an armamentarium of different nontoxic techniques and approaches that will enable investigators to stabilize human interferons at each step in preparation, processing, concentration, purification, shipment and storage.

Fibroblasts↗
PubMed1975

The influence of physicochemical factors on the thermal inactivation of murine interferon.

The degradation of biological activity of virus-induced murine interferon was determined in linear nonisothermal and multiple isothermal tests. The stabilizing effect of pH during heating on interferon in solution was greatest at low pH, such that pH 2 greater than pH 5 greater than pH 7 greater than or equal to pH 9; freeze-dried preparations of interferon were also more heat-stable at acid pH than at neutral pH. Heat stability was a function of the H+-ion concentration rather than the ionic composition of the buffer; interferon solutions containing monovalent cations with different ionic radii had similar heat stability. A change in the H+ ion concentration was a critical event during the cooling of heated interferon: a shift in the direction of acidity contributed to stability whereas a shift towards alkalinity led to inactivation. The rate of cooling of heated interferon significantly influenced its residual activity. Rapid cooling and sudden freezing decreased the residual activities of interferons at pH 2 and 9 more than "normal" cooling, an effect not observed at pH 7. Interferon heated to 80degree C could not be reactivated at 40degree C or 55degree C. Interferon of higher apparent molecular weight was more heat-stable than that with lower apparent molecular weight. It is postulated that the physicochemical alterations in the aqueous environment significantly affecting the stability of interferon operate by producing changes in the size and/or conformation of interferon molecules. A model is proposed that relates thermal inactivation to different possible molecular states of interferon.

Animals↗
PubMed1975

The neuraminidase yield-reduction bioassay of human other interferons.

The production of neuraminidase by the recombinant influenza virus X7(F1) in human monkey, rabbit, hamster, mouse, and chicken cell cultures is inhibited by interferon. Described is a new enzyme assay for neuraminidase that can be applied to the bioassay of interferons. The advantages of this interferon bioassay are its sensitivity, reproducibility, rapidity, and convenience.

Animals↗