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Results for “Diffusion Chambers, Culture”

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PubMed1976

Growth of in vitro colony-forming cells from normal human peripheral blood leukocytes cultured in diffusion chambers.

The growth of granulopoietic progenitor cells (CFU-C) in diffusion chambers during culture of peripheral blood leukocytes from 10 normal subjects has been studied. At various times after initiation of diffusion chamber culture, cells harvested from the chambers were transferred to agar culture for measurement of CFU-C concentration. Under these conditions colonies could be grown successfully in agar culture provided pronase, necessary for the chamber harvesting procedure, was first removed by careful washing. A marked increase in the number of CFU-C, up to 25-fold the initial value, was observed in 8 out of 10 subjects. Here the growth pattern was similar, independent of the initial CFU-C throughout the diffusion chamber culture period was very poor. The growth of CFU-C from a given individual's blood was shown to be reproducible in repeated studies in 2 subjects, one of whom showed a proliferative and the other a non-proliferative pattern. Evidence suggests that the increase in CFU-C in diffusion chambers is the result of both self-renewal of these cells and influx from a more primitive compartment, although the present data do not allow an estimate of the relative magnitude of each.

Cell Division↗

PubMed1983

[Neurons of the enteral portion of the metasympathetic system cultured in diffusion chambers].

A culture of the neurons of metasympathetic system's enteral portion of 3-5-day old rats was obtained and fragments of the mienteral plexus were implanted to adult rats-recipients. A technique of cultivation was developed for in vivo microscopy of culture's neurons. Migration of neuroblasts from the deplantate area was revealed as well as the specific features of neurons differentiation. The neurons were shown to acquire characteristic morphological features corresponding to Dogel's I and II types of neurons; growth of neurites was shown to aid to formation of neuropil and microglia. Histochemical analysis revealed a product of response to cholinesterase which is the index of formation of the acetylcholine hydrolysis system in the culture's I type neurons. The technique of electron-microscopic scanning enables to watch in detail the process of formation of interneuronal connections and formation of a synapse.

Animals↗

PubMedDate not supplied

[Neutrophils do not affect clonal growth of granulocyte progenitor cells cultured in vivo in diffuse chambers].

The effect of polymorphonuclear leukocytes on the growth of granulocytic progenitors in the diffusion chamber culture (CFU-dG) has been studied. It has been shown that peripheral blood neutrophils do not affect the myeloid colony formation in this culture system. The inhibition of CFU-dG growth, observed in the presence of peripheral blood leukocytes was dependent on prostaglandins release from monocytes.

Animals↗

PubMed1995

[Changes in the myocardium of adult rats when cultured in diffusion chambers in vivo].

Morpho-functional changes of left ventricle myocardium were studied in 30 adult male rats by the method of subcutaneous implantation of diffusion chambers. The transplants were examined in 1, 3, 6, 8 and 10 days after the beginning of cultivation. The method of light microscopy at the 1-2 microm sections was used after histological processing of specimens for the identification of muscle and non-muscle cells and studying of their particular changes. In the muscular fibres there were cardiomyocytes of the destructive, survived and reconstructed forms. It was shown that smooth muscle cells and periocytes are separated from blood microvessels and migrated into the interstitial space. Multinuclear drawing rods formated from activational endotheliocytes were detected between the muscular fibers. Furthermore the cells containing large amount of lipid granules-¿lypophags¿ and the two types of macrophage-like cells were demonstrated. The area of growth including fibroblast-like and spindle-shaped cells were observed around the margins of transplants on the late stages of implantation.

Animals↗

PubMed1978

Sister chromatid exchanges induced by cyclophosphamide in V79 cells cultured in diffusion chambers in mice.

Chinese hamster cells V79 were cultured in diffusion chambers (DC) and implanted into mice. An exponential growth was observed from the 2nd to 4th day after implantation. The maximum growth was reached on the 6th day. After that, cell growth and viable cell counts decreased. Three days after implantation of DC with V79 cells, the hosts received 6 hourly injections of 0.2 ml of 5-bromodeoxyuridine (BUdR) solution at concentrations of 0.125 to 1.0 x 10(-2) M. DC were removed for chromosome and sister-chromatid exchanges (SCE) analyses 24 h after the first BUdR injection. The frequency of metaphases with differentially stained chromatids, with aberrations, and the number of SCE per cell increased with BUdR dose. The frequency of metaphases with differentially stained chromatids was also positively correlated with the duration of BUdR exposure or the number of hourly injections of BUdR-solution. The effects of cyclophosphamide (CY) in V79 cells in DC in mice were studied. Injections of CY at 2.5, 5, 10 and 15 microgram per gram of body weight to the hosts caused an increase in the number of SCE per cell in a linear manner. The results from this study indicate that V79 cells cultured in DC in mice may provide a potential test system for mutagenicity.

Animals↗

PubMed1979

Culture of human haemopoietic cells in diffusion chambers. I. Growth of normal peripheral blood cells.

Growth of mononuclear cells from human peripheral blood from 10 normal individuals was tested in diffusion chamber culture over a period up to 17 or 21 days. After an initial decrease during the first few days an increase of the total cell number was observed with maximal values on day 13. In all individuals growth of undifferentiated blast cells, lymphocytes, plasma cells, immature and mature granulopoietic cells, macrophages, and megakaryocytes occurred. In all individuals the different cell types had similar growth patterns in diffusion chamber culture. The considerable numerical variations which were seen in the granulopoietic cells were probably due to different stem cell concentrations in the peripheral blood of the investigated individuals. The results indicate that the diffusion chamber technique is a valuable method for the detection of haemopoietic stem cells and the culture of lymphocytic cells in man.

Adult↗

PubMed1977

Effect of human leucocyte interferon on DNA synthesis in human psoriatic skin cultured in diffusion chambers.

Pieces of human psoriatic skin were cultured for five days in diffusion chambers implanted intraperitoneally in mice. Partially purified human leucocyte interferon was injected into the experimental animals daily during the culture period. Phosphate buffered saline and, in one experiment, also mock interferon were used as control materials. On the fifth day, incorporation of 3H-thymidine into the epidermal cells was studied autoradiographically. The results showed that human leucocyte interferon does not depress DNA synthesis in human psoriatic epidermal cells cultured by this method. In fact, DNA synthesis seemed to be stimulated in two of the three experiments but the effect was not statistically significant.

Animals↗

PubMed1988

[Hormone-dependent characteristics of the myocyte cell surface in the human myoma and myometrium cultured in diffusion chambers].

The normal and the pathologically changed human myometrium was cultured in the diffusion chamber implanted in the subcutaneous cellular tissue of the rat. In the absence of hormonal influence the growth of myometrium culture is only insignificant, while no myoma cell growth was found at all. Estradiol stimulates the growth and development of the myometrium culture and the myoma. A combined action of estradiol and progesterone stimulates the collagen formation.

Animals↗

PubMed1977

Growth and differentiation of normal and leukemic human bone marrow cells cultured in diffusion chambers.

Bone marrow from healthy, normal volunteers and patients with acute myelocytic leukemia was cultured in diffusion chambers implanted into cyclophosphamide pretreated mice. Chambers were removed at regularly scheduled intervals over a period of 28 days. Total and differential cell counts were then done on the contents of each chamber. Normal human bone marrow showed an orderly pattern of growth and differentiation which was not found with leukemic bone marrow. Monocytes and macrophages were the predominant cell types in the diffusion chambers filled with normal marrow after day 10 of culture. Although leukemic specimens showed predominantly leukemic cells, a few mature polymorphonuclear leukocytes could be found throughout the entire culture period. Questions about the nature of the defect in acute myelocytic leukemia and the significance of the in vivo culture system are discussed. The results of these studies are compared and contrasted with studies of a similar type.

Animals↗

PubMed1986

[Thermal sensitivity of Ehrlich ascites cancer cells forming colonies in agar cultures in diffusion chambers].

Thermosensitivity of clonogenic ascites Ehrlich carcinoma cells (AEC), forming colonies in agar cultures in diffusion chambers was studied when heating them in vitro at 41-44 degrees C. The clonogenic cells of AEC are shown not to essentially differ both in the colony-forming efficiency (CFE) and in the proliferative status on the 3rd and 7th days. No essential differences are revealed in thermosensitivity of such cells. The heating time-survival curves are exponential or S-shaped with a small "shoulder". The temperature increase by 1 degree C resulted in a 2-2.5-fold decrease in the time of effective hyperthermia treatment. The values of D0 for cells treated with 42 degrees C in different experiments varied from 20.1 min to 24.6 min. Only one type of thermal damage reaction with the energy of inactivation about 150 kcal/M was observed in AEC cells in the temperature range studied.

Agar↗

PubMed1976

Chalone-induced inhibition in DNA synthesis of human psoriatic epidermal cells cultured in diffusion chambers in mice.

Pieces of human psoriatic skin and suspensions of mouse bone marrow cells as non-epidermal control tissue were cultured for four days in diffusion chambers implanted intraperitoneally in 40 mice. The mice were divided into three groups. On the second and third day of the culture partially purified epidermal extract containing epidermal chalone, granulocyte extract containing granulocyte chalone, and Hanks' solution were injected intraperitoneally into the mice of the three groups, respectively. On the fourth day, incorporation of 3H-TdR into the diffusion chamber cultures and various epithelia and several non-epidermal tissues of the host animal was studied autoradiographically and radiochemically. The results showed that the epidermal chalone induced a marked, statistically significant, depression in the labelling index of the cultured psoriatic epidermal cells and in the number of labelled cells in the host ear epidermis (31% and 62%, respectively); the other host epithelia were not significantly inhibited. A crude granulocyte extract used in most experiments was clearly toxic; the semipurified granulocyte chalone used in one experiment had no effect on the psoriatic and cultures, but affected weakly the cultured bone marrow cells. Neither the epidermal nor the granulocyte extract had any inhibitory effect on the non-epidermal tissues studied.

Animals↗

PubMed1978

The fine structure of monoclonal Hodgkin cells cultured in diffusion chambers.

Pleural effusion cells from two patients with stage IV Hodgkin's disease have been cultured continuously in diffusion chambers in mice and studied by electron microscopy after a culture period exceeding 100 days. Cell identity and monoclonal growth in culture has been documented by marker chromosomes (Hossfeld and Schmidt, 1978). These cultured cells grow in close connection, projecting pseudopode-like processes into the intercellular spaces. Most nuclei are lobulated. They always are of low electron density with a norrow rim of condensed chromatin confined to the nuclear membrane. One large prominent nucleolus and up to four smaller nucleoli are found. Nuclear pockets in case 1 and deep cytoplasmic invaginations into the nuclear area in both cases frequently occur. In the cytoplasm, besides microtubuli and fibrils, the Golgi apparatus and mitochondria are the predominant organelles. Most mitochondria appear to be dilated containing fragmented cristae. Free ribosomes and polysomal aggregates are randomly distributed. The ratio nucleoplasm:cytoplasm, on the average, is 0.7 in both cases and the cell diameters lie distinctly above those of lymphocytes. At the electron microscope level these cultured monoclonal cells of Hodgkin's disease are not distinguishable from those described in genuine Hodgkin material. Their probable origin and apparent relation to true histiocytic lymphoma cells will be discussed.

Cells, Cultured↗

PubMed1993

Transplant of bone marrow and muscle-derived connective tissue cultures in diffusion chambers for bioassay of bone morphogenetic protein.

To promote a high concentration gradient, syngeneic bone marrow and muscle connective tissue cell cultures and subcultures were exposed to bone morphogenetic protein and associated noncollagenous bone matrix proteins (BMP/NCP) in vitro, transferred to diffusion chambers, and then transplanted into the anterior abdominal wall of isogeneic rats. Both muscle- and marrow-derived cells differentiated into cartilage and chondroosteoid on the inside of diffusion chambers. New bone developed on the vascularized outside surfaces in juxtaposition to avascular tissue on the inside. Levels of glycosaminoglycans (GAGs) and DNA synthesis in tissues inside the chambers were sharply elevated. These data, correlated with histologic observations, demonstrate that the BMP/NCP recruits mesenchymal-type cells for a skeletal tissue pathway of development. Muscle-derived cells, as distinguished from marrow-derived cells, originate in a nonosseous environment but differentiate into osteoprogenitor (osteogenic stem) cells and display levels of DNA synthesis almost as high as marrow stroma-derived cells. The rise in DNA synthesis was maximal in the interval from two to three weeks after transplantation. Whether the bone marrow stroma mesenchymal cell target for BMP is only the osteogenic stem cell population or also includes colony-forming units fibroblasts (CFU-F) remains to be investigated by experiments on established cell lines.

Animals↗

PubMedDate not supplied

[Survival of clonogenic cells of Lewis lung carcinoma cells forming colonies in agar cultures in diffusion chambers during gamma-irradiation and exposure to gamma-neutron californium-252 irradiation].

Clonogenic cells forming colonies in agar cultures in diffusion chambers and those isolated from subcutaneously transplanted Lewis lung carcinoma do not differ in their sensitivity to 60Co gamma-rays with respect to tumor growth stages. The dose-survival curves for all studied cells are S-shaped with a small shoulder. A cumulative dose-survival curve for malignant clonogenic cells is characterized by the average value of mean lethal dose D0 = 2.24 Gy and extrapolation number n = 2.0. When exposed to gamma-neutron-radiation (252Cf) malignant clonogenic cells exhibit a nearly exponential dose-survival curve with D0 = 0.56 Gy (with respect to a neutron component). The RBE of gamma-neutron radiation (252Cf) is 2.5.

Animals↗

PubMed1982

Characterization of normal human mononuclear blood cells during diffusion chamber (DC) culture with respect to proliferation and expression of T-cell antigen.

Normal human mononuclear blood cells were cultured for 13 d in diffusion chambers and the cell harvest on days 3, 6, 9 and 13 was characterized according to morphology, growth kinetics and surface antigen expression. The greatest part of the cell harvest consisted on all days of lymphoid cells. It could be shown that most of the cells carried the T-cell marker, the main growing population thus being T-cells. The mean anti-T-binding capacity per cell increased during the initial lag phase and reached its peak when the cell count was lowest. It dropped again during the following exponential growth phase to preculture values. These changes indicate an initial increase in surface antigen during the transition of cells from resting to a more activated state before the onset of DNA-synthesis, and a subsequent decrease which may indicate functional maturation.

Adult↗

PubMed1982

Diffusion chamber (DC) culturing of haemopoietic cells: a reliable assay system for regulators of proliferation?

If i.p. DC culturing of haemopoietic cells shall serve as a reliable assay for systemic and/or local peritoneal regulators of proliferation, cell growth must not be restricted by the diffusion capacity through the chamber membranes. Diffusion was assessed by measuring uptake in DC or 51Cr-EDTA of 3H-thymidine, given intravenously to mouse hosts or to chambers incubated in vitro. Fractional incorporation of 3H-thymidine, numbers of mouse spleen colony-forming stem cells, agar colony-forming progenitors (of granulocytes and macrophages), and total cell numbers were taken to reflect cell growth. Changing the diffusion capacity did not lead to marked changes in cell growth; and vice versa, culturing cells under host or chamber conditions that increased cell growth was not associated with an appreciably increased diffusion capacity. Diffusion was increased by replacing Acropor with Millipore DC or by repeatedly cleaning the DC during the culture period. Cell growth was increased by using irradiated rather than normal host mice, and Nucleopore rather than Millipore DC in irradiated mice. Increasing the oxygen delivery to the cultured cells by using polycythaemic and hyperoxic hosts did not enhance cell growth either.

Animals↗

PubMed1979

Induction of 8-azaguanine- and ouabain-resistant mutants by cyclophosphamide and 1-(pyridyl-3)-3,3-dimethyltriazene in Chinese hamster cells cultured in diffusion chambers in mice.

A host-mediated assay is described for induction of 8-azaguanine-resistant (azgr) and ouabain-resistant (ouar) mutants in Chinese hamster V79 cells cultured in diffusion chambers (DC) in C3H mice. Injection of the hosts with the indirect mutagen/carcinogen cyclophosphamide (CPP) or 1-(pyridyl-3)-3,3-dimethyltriazene (PyDT) caused a dose-dependent increase in mutation frequency at the loci of azgr and ouar in the V79 target cells. Plating efficiency of V79 cells in DC in mice was decreased depending upon the dose of CPP or PyDT given to the hosts. In addition, the relationship between expression time and mutation frequency was examined and discussed. The data support the use of this system as an effective screening procedure for suspected environmental mutagens or carcinogens, especially those that need to be metabolically activated in vivo.

Animals↗

PubMed1978

Endotoxin-induced alterations in canine granulopoiesis: colony-stimulating factor, colony-forming cells in culture, and growth of cells in diffusion chambers.

Salmonella typhosa endotoxin injected into dogs produced elevated plasma CSF levels, transient leukopenia followed by leukocytosis, and stimulation of marrow granulopoiesis and mobilization of granulocyte-macrophage progenitors cells into the the peripheral circulation. The number of marrow CFU-c decreased to 65% of the control number within 6 h, returned to control levels by 24 h, and increased to 370% of the control number by 48h after endotoxin. The granulopoietic response was supported by a concomitant increase in the M:E ratio, an increased fraction of marrow-derived CFU-c susceptible to 3H-TdR suicide, and increased granulo-monocytopoietic activity of marrow- and peripheral blood-derived cells grown in diffusion chamber cultures. These results are consistent with the concept that endotoxin-induced CSF is a physiologic regulator of canine granulopoiesis, and that canine marrow responds to endotoxin with a significant increase in the concentration of marrow-derived granulocytic progenitors and with mobilization of granulocyte-macrophage progenitors into the peripheral circulation.

Animals↗