Showing papers on "In vivo published in 1969"

Bioassay of Parathyroid Hormone in Vitro with a Stable Preparation of Adenyl Cyclase from Rat Kidney

Abstract: A bioassay for parathyroid hormone in vitro was developed based upon the activation of adenyl cyclase from the renal cortex of the rat. Preparation of the enzyme in 0.05M Tris-HCl, pH 7.4, containing 10% v/v dimethylsulfoxide allowed storage in liquid nitrogen for periods in excess of 30 days without significant loss of sensitivity to parathyroid hormone or fluoride. The assay was sensitive to as little as 0.14 USP U of parathyroid hormone, and showed a log-linear dose response over the range of 0.14– 1.13 USP U of hormone. The average index of precision in a series of 23 assays was 0.083. Theassay was highly specific for parathyroid hormone and showed appropriate increments in potency for hormone preparations at consecutive stages of purification. Results with the adenyl cyclase method in vitro were similar to those obtained with a standard bioassay in vivo. There was no significant interference from other hormones and proteins, including nonhormonal contaminants found in crude parathyroid extracts. The ...

A phosphorylation site in brain and the delayed neurotoxic effect of some organophosphorus compounds.

Abstract: 1. It is proposed that part of a neurotoxic dose of di-isopropyl phosphorofluoridate will be covalently bound in vivo to a specific component in the brain and spinal cord as the initial biochemical event in the genesis of the lesion. 2. A test system in vitro was devised that removes many di-isopropyl phosphorofluoridate-binding sites and indicates that the specific component may be a protein present in brain at a concentration comparable with that of the cholinesterases. 3. The site was found to be present and capable of binding di-isopropyl phosphorofluoridate in vitro in brain samples taken from either normal hens or those dosed with organophosphorus esterase inhibitors that are not neurotoxic. 4. Very little of the specific binding activity was found in brain samples from hens pre-dosed with a variety of neurotoxic organophosphorus compounds. 5. A solubilized preparation of the active brain component was obtained, suitable for further purification and study.

Lipid metabolism in tissue culture cells.

Abstract: Publisher Summary The development of methods for routine cultivation of cells in vitro has provided the biochemist with a useful system for the study of cellular lipid metabolism, making it possible to obtain large quantities of a homogeneous population of cells. The degree of biological variation encountered in a tissue culture system is less than that obtained from studies employing whole animals. In addition, experimental conditions can be easily varied at will in cell culture systems. The differences between tissue culture systems and other experimental systems must be kept in mind when evaluating metabolic data obtained from in vitro cell studies. Most of the cells maintained in tissue culture are dedifferentiated in the sense that they do not carry out many of the specialized functions found in differentiated cells. These cells are not organized into specialized tissues as are cells in vivo, limiting some aspects of cellular interactions. Cells in culture are rapidly proliferating, and most lipid studies have been conducted with mixoploid cells derived from malignant tissue. The cellular growth environment of cells cultivated in vitro differs from that of cells in vivo.

Factor XIII in Human Plasma and Platelets

Abstract: Plasma and platelet factor XIII levels were measured in normal human donors and in a patient congenitally deficient in factor XIII. The purpose of these experiments was to study the role of platelet factor XIII in blood coagulation. On polyacrylamide disc electrophoresis, factor XIII activity in extracts of washed normal platelets appeared as a single peak. This peak was missing or very low when factor XIII-deficient platelet extract was used. The patient was also studied before and after transfusion of fresh frozen plasma. Before transfusion, factor XIII activity could not be detected in the patient's plasma or platelet extracts. 24 hr after transfusion the plasma factor XIII level was still at the anticipated value, but factor XIII activity could not be detected in the platelet extracts. These experiments imply that plasma factor XIII was not transported across the platelet membrane in measurable quantities in vivo. This suggests that platelet factor XIII is a true platelet component and originates in the platelet precursor, the megakaryocyte. Thrombelastographic studies suggest that platelet factor XIII participates in the coagulation process. Thrombelastograms of factor XIII-deficient samples had decreased maximum amplitude and clot elasticity values. The abnormalities were fully corrected by the addition of washed normal platelets to factor XIII-deficient plasma; preincubation of the normal platelets in the deficient plasma had no additional effect. This indicates that platelet factor XIII is immediately available during clot formation.

The selective elimination of immunologically competent cells from bone marrow and lymphocyte cell mixtures. Iii. In vitro test for detection of immunocompetent cells in fractionated mouse spleen cell suspensions and primate bone marrow suspensions.

Abstract: SUMMARY Application of the density gradient centrifugation technique for fractionation of mouse spleen cell suspensions to primate bone marrow, as a method to separate the immunocompetent cells from the haemopoietic stem cells to be grafted, resulted in a moderate but still fatal secondary disease in the recipient. Therefore, minor changes of the separation technique for primate bone marrow were necessary. Since adequate microscopic identification of the two cell types to be separated is not possible, assays for their functional characteristics had to be developed. The impracticability of in vivo assays in primates necessitates an investigation of in vitro properties of the immunocompetent cell and haemopoietic stem cell. With regard to the immunocompetent cells, the phenomenon of transformation of lymphoid cells by phytohemagglutinin was used. The reliability of this in vitro test, called phytohemagglutinin-response test, was ascertained by quantitative comparison with the graft-versus-host assay in vivo in mice in different cell suspensions under a variety of conditions. With regard to the in vitro test for haemopoietic stem cells, a method is described which appeared to be not specifically indicative for stem cells but, nevertheless has been found to be of value as a check on the reproducibility of the separation technique

Antagonistic Action of Anti-androgens on the Formation of a Specific Dihydrotestosterone-Receptor Protein Complex in Rat Ventral Prostate

Abstract: Cyproterone (1,2α-methylene-6-chloro-Δ4,6-pregnadien-17α-ol-3,20-dione) 17α-acetate, a potent anti-androgen, suppressed the uptake of radioactive androgens in vivo by the ventral prostate of rats. This was accompanied by a decrease in the retention of 5α-dihydrotestosterone (17β-hydroxy-5α-androstan-3-one) by prostate cell nuclei . Cyproterone and its 17α-acetate (less than 0.5 µg/ml) also inhibited the formation of a specific dihydrotestosterone-protein complex in prostate cell nuclei when minced prostate was incubated with radioactive testosterone or dihydrotestosterone. Estradiol-17β, diethylstilbestrol, and progesterone, but not hydrocortisone succinate, also suppressed the retention of dihydrotestosterone by prostatic cell nuclei in vitro, but to a much lesser extent than cyproterone.

Studies on serum-mediated inhibition of cellular immunity to spontaneous mouse mammary tumors.

Abstract: Sera from mice which have either spontaneously developed a mammary tumor, or which have been immunized against such a tumor, have been shown capable of abrogating the ability of lymph-node cells from these mice to specifically inhibit colony formation by mammary tumor cells in vitro. Sera from mice sensitized to another, independently arising, syngeneic mammary tumor or to a syngeneic Moloney sarcoma virus-induced tumor are less active in this respect, indicating that this reaction is specific. It is suggested that these observations are related to the in vivo phenomenon called immunological enhancement. Inhibition Serique de l'Immunite des Tumeurs Mammaires On a observe que des serums de souris chez lesquelles une tumeur niamniaire est apparue spontanement ou qui ont ete immunisees contre ce type de tumeur ont pu empecher Ies cellules de ganglions lymphatiques de ces souris d'inhiber specifquenient la formation de colonies par des cellules de tumeurs mammuires in vitro. Les serums de souris sensibilisees a une autre tumeur mammaire syngeneique apparaissant independamment, ou a une tumeur syngeneique provoquee par le virus du sarcome de Moloney, sont moins actifs a cet egard, ce qui indique qu'il s'agit d'une reaction specifique. II est possible que ces observations soient en rdation avcc le phenomene in vivo appele „facilitation immunologique”.

In vitro evaluation of immunosuppressive drugs

Abstract: In vivo testing of immunosuppressive drugs is possible only in animals and the results cannot be extrapolated to humans. None of the in vitro tests used so far has proved satisfactory. The new in vitro method described here is sensitive enough for the testing of metabolites present in human serum after the administration of drugs. It is based on the inhibition of spontaneous rosette forming cells (RFC), the nature of formation of which may differ from that of immune rosette formation. This spontaneous phenomenon is usually supposed to result from natural antibody producing cells, and the number of spontaneous RFC in mice with sheep red blood cells (SRBC) is more than ten times smaller than in mice immunized with SRBC1.

The in vivo pH of the extravascular space of the lung

Abstract: The partition of 5,5-dimethyloxazolidine-2,4-dione (DMO) and of 11 amines between the vascular and extravascular spaces of the lung has been determined by the multiple indicator dilution technique. Four amines (nicotine, pentylamine, quinine, and benzylamine) were found to have pH-sensitive tissue to blood concentration ratios. Of these, tritiated nicotine appears to be the nost satisfactory indicator of tissue pH and values for the pH of the pulmonary extravascular space (pH(e)) have been calculated from the nicotine data. At an arterial pH (pH(art)) between 7.38 and 7.43 pH(e) averaged 6.69 +/-0.07. Changes in pH(e) usually paralleled but were consistently less than concomitant changes in pH(art). Alterations in P(CO2) at constant pH(art) regularly produced relatively small, parallel changes in extravascular hydrogen ion concentrations. Local alterations in tissue pH due to P(CO2) changes are apparently buffered quite rapidly and the pH(e) of the lung seems more closely linked to pH(art) than the cellular pH of other tissues.DMO, guanidine, methylamine, morphine, and atropine were confined to the vascular volume during the first circulation and could not be used to measure tissue pH. Histamine appeared to be bound to a pH-insensitive site. The extravascular distributions of antipyrine and aniline were unresponsive to alterations in arterial pH, presumably because they are essentially uncharged at pH levels found in the lung.

Tritiated thymidine labelling of normal human epidermal cell nuclei. A comparison, in the same subjects, of in vivo and in vitro techniques.

Abstract: SUMMARY.— Only 2 methods for the incorporation of radioactive tracers into skin i.e. the local in vivo method and the in vitro method can be seriously considered for routine clinical use in man. Both methods-and especially the in vitro method as modified by us—are described in detail. The patterns of epidermal labelling and the labelling indices seen in the same skin fields, from the same 6 healthy human male volunteers, with the in vivo and in vitro techniques are directly compared. These two methods were found to yield very similar results. Thus, the average percentage of labelled basal cell nuclei was found to be 2·2% (range 1·9%, to 2·6%) with the local in vivo method and 2·3% (range 2·0% to 2·6%) with the in vitro method. The average percentage of labelled epidermal cell nuclei was found to be 2·1% (1·7% to 2·5%) with the in vivo method and 2·2% (range 1·4% to 3·0%) with the in vitro method for the same skin field. It is concluded that the local in vivo labelling method will probably provide invaluable data on cell and metabolite turnover rates, and its use should perhaps be limited to such studies. For all other purposes, however, the in vitro method, here detailed, seems to be preferable for routine clinical studies because of its safety, simplicity and reproducibility.

In vivo reactions of nitroso compounds

Abstract: A number of N-alkylnitroso compounds have very potent biological actions, including acute cellular injury in the liver and other organs, carcinogenesis, mutagenesis, and teratogenesis. Aspects of these biological actions have been discussed in several recent The mechanism of toxic liver injury by nitrosamines has been discussed by Magee and Lee! The biological actions of the N-alkylnitroso compounds are determined to some extent by their chemical properties. The dialkylnitrosamines, of which dimethylnitrosamine is the simplest example (FIGURE l), are relatively stable chemically, while some N-nitrosamides, for example, N-nitrosomethylurea (FIGURE l), are relatively unstable in aqueous solution, the rate of decomposition being dependent on pH. According to Druckrey and co-workersz the half-life of N-nitrosomethylurea is 125 hours at pH 4.0, 24 hours at pH 6.5, 1.2 hours at pH 7, and 0.1 hours at pH 8.

The assessment of the nutritive value of silage by determination of in vitro digestibility on homogenates prepared from fresh undried silage

Abstract: A report is presented on studies on the relationship between in vitro digestibility of 18 silage samples and their in vivo digestibilities. Various analytical techniques are compared and the method adopted, on grounds of suitability for routine operation and accuracy of prediction of in vivo data, is based on analysis of homogenates of fresh silage. The homogenates, sufficiently uniform to permit of volumetric subsampling at acceptable error levels, are prepared in a new design of homogenizer. The impact of the % dry matter of the silage, as fed, on the accuracy of in vitrol in vivo relationships is considered and a bivariate regression plane is proposed.

Metabolism of some phenylethylamines and their beta-hydroxylated analogs in brain.

Abstract: In rat brain slices, phenylethylamine derivatives were converted predominantly to acidic metabolites, whereas the β-hydroxylated derivatives (norepinephrine, normetanephrine and octopamine) were converted predominantly to neutral metabolites. Normetanephrine-H3 incubated with guinea-pig brain slices was converted mainly to 3-methoxy-4-hydroxyphenylglycol, whereas normetanephrine incubated with liver slices was metabolized primarily to 3-methoxy-4-hydroxymandelic acid. Studies comparing metabolism of i.v. and intracisternally injected normetanephrine indicate that reduction of the intermediate aldehyde is the predominant route of metabolism of β-hydroxylated phenylethylamine derivatives in vivo as well as in vitro in brain tissue.

Interactions of 6,7-3H-17beta-estradiol with mammary gland and other organs of the C3H mouse in vivo.

Abstract: After injection of a physiological amount of 6,7-3H-17β-estradiol into ovariectomized C3H mice,the nonvolatile radioactivity in the mammary gland consists almost exclusively of “free” 17β-estradiol. The mammary gland is able to concentrate and retain 17β-estradiol against a large concentration gradient with the blood. On a dry weight basis, the amount of hormone retained by the mammary tissue is about 1/3 that of uterus, 2/3 that of vagina, 11 times that of omental adipose tissue, 17 times that of lung and 42 times that of muscle. Estrogenic or antiestrogenic substances but not steroids without estrogenic activity are able to compete with 17β-estradiol for the process of accumulation and retention, an indication that this process is highly specific. In all tissues the pool of hormone specifically retained shows a half-life (turnover ti.ne) of about 7 hr. These findings suggest that the mammary gland is able to bind 17β-estradiol in a highly specific manner, and furthermore that this ability is not a pecul...

Cytogenetic Effects of Tranquilizing Drugs in Vivo and in Vitro

Abstract: During a recent investigation of the effects of lysergic acid diethylamide (LSD) on human chromosomes, three individuals, included as controls, showed elevated frequencies of chromosomal breakage while being treated with chlorpromazine. 1 Subsequently, invitro observations on human leukocytes and fibroblasts suggested that diazepam is also capable of inducing chromosome damage. 2 Due to the widespread clinical use of such drugs in our population, an in-vivo and invitro cytogenetic study of several tranquilizing agents was carried out. Materials and Methods: In Vitro Studies .—Peripheral leukocyte cultures were initiated from two normal, healthy donors (one male and one female) with no known history of prior drug ingestion, radiation exposure, or recent viral infection. Chromosomes were obtained from cells of whole-blood inoculum (3 to 4 drops), grown in commercially available microculture kits, and incubated at 37 C for 72 hours. Two hours before harvest, colcemide was added at a final concentration of 0.05μg/ml

Delayed hypersensitivity to chemically induced tumors in mice and correlation with an in vitro test

Abstract: Tumors induced in CBA mice by 3-methylcholanthrene were used to confer delayed hypersensitivity on other mice of the same strain. Two methods of sensitization were successful: 1) surgical removal of a transplanted growing tumor and 2) administration of a disrupted tumor vaccine with Freund's adjuvant. Living tumor cells prepared in various ways (but not disrupted cells or extracts) elicited typical delayed cutaneous reactions when injected into the foot pads of sensitized mice; these reactions were tumor specific. Hypersensitivity in vivo was correlated with specific susceptibility to the inhibition of peritoneal cell migration in vitro.

Stimulatory and inhibitory effects of purified hypothalamic extracts on growth hormone release from rat pituitary in vitro

Abstract: Male rat anterior pituitaries were incubated in vitro for 5 hr in Krebs-Ringer bicarbonate medium and the growth hormone (GH) released from the glands was estimated by the tibial epiphyseal cartilage assay. Addition of crude sheep or rat hypothalamic extract to the pituitaries increased the GH concentration in the medium on comparison to either diluent or cortical extract-treated controls. The ovine extracts also depleted pituitary GH concentration when injected into male rats in vivo. Gel filtration of either sheep or rat hypothalamic extract on a column of Sephadex G-25 resulted in the elution from the column of 2 zones which influenced the release of GH in vitro on addition to the incubated pituitaries. The first zone of activity to be eluted increased the release of GH from the glands severalfold, whereas the second zone inhibited the release of GH to levels about one half of that released by control glands incubated in the presence of the eluting buffer. Increasing the dose in the inhibitory zone res...

Gaffkemia, a bacterial disease of the lobster, Homarus americanus: effects of the pathogen, Gaffkya homari, on the physiology of the host.

Abstract: Gaffkya homari, the pathogen causing gaffkemia, was observed in vivo to reach its stationary growth phase (approximately 108 bacteria/ml of lobster hemolymph) in 4 days at 15 C. The bacterial totals at death ranged between 5 × 108 and 1 × 1010/ml of hemolymph. Among the main effects of the infection was the great increase in hemolymph clotting times followed by the essential elimination of clotting. The impaired clotting mechanism was a result of the drastic reduction in circulating hemocyte numbers rather than a reduction in hemolymph fibrinogen levels. Hepatopancreatic glycogen levels and hemolymph non-protein nitrogen concentrations dropped to minimal levels but hepatopancreatic lipid levels, hemolymph lactic acid, carbohydrate, and pH values were not significantly affected by the infection. An extreme hyperglycemic effect, attributed to stress, was observed in both control and infected lobsters. Death from gaffkemia, other than that stemming from a wounded animal bleeding to death because of the impai...

ULTRAVIOLET LIGHT ALTERATION OF CELLULAR DEOXYRIBONUCLEIC ACID in vivo

Abstract: DNA after irradiation with ultraviolet light was immunogenic in rabbits and elicited serum antibodies reacting specifically with UV-irradiated DNA. The serological reactions were demonstrated by immunodiffusion, complement fixation, and immunofluorescence. By immunofluorescence, antisera reacted with cell nuclei of irradiated tissue sections but not with unirradiated tissue. This method was employed to show the presence of UV lesions in tissues of mice exposed to UV light. UV lesions in DNA were present in nuclei of epidermal cells, and in heavily irradiated animals they were also detected in the corium immediately below the epidermis. The method is useful not only for directly demonstrating UV lesions of DNA but also for localizing such lesions in tissues.