Separation of mouse spleen haematopoietic cell fractions for transplantation purposes in lethally irradiated mice.

Mouse cells were fractioned by adherence chromatography on glass beads. The fractionation of spleen cells yielded 5 different fractions, whose transplantation effect was tested by the method of Till and McCulloch. The results of these transplantation tests showed that the chosen form of adherence chromatography was not sufficiently successful, since none of the resultant fractions contained solely colony-forming units (CFU) and in none of them were CFU present in an exeptionally high concentration.     

T-cell differentiation in lethally irradiated and reconstituted mice

During the first 3 days after irradiation and reconstitution (IR) with hemopoietic stem cells a small number of cells with the capacity to form colonies (CFU-s) can be detected in the thymus. This number is increased when thymectomized mice are used as recipients for colony determination. In thymus-cell suspensions from animals 4 days after IR no CFU-s can be found either in normal or in thymectomized recipients. Tracer studies with ¹¹¹In-labeled oxine revealed that thymocytes obtained from animals 3 days after IR contain a large number of cells with a strong preference for the thymus. This number decreases in the following days after irradiation; these cells are thought to represent an intermediate cell between stem cell and T cell.     

Reconstitution of lethally irradiated mice by cells isolated from adipose tissue

It is suggested that hematopoietic stem cells (HSC) could be found in several tissues of mesodermic origin. Among these, adipose tissue can expand throughout adult life and its expansion is not only due to mature adipocyte hypertrophy but also to the presence of precursor cells in stroma-vascular fraction (SVF). Here we report that transplantation of cells isolated from mice adipose tissue can efficiently rescue lethally irradiated mice and results in a reconstitution of major hematopoietic lineages. Donor cells can be detected in blood and in hematopoietic tissues of recipient mice. Adipose tissue contains a significant percentage of CD34, CD45 positive cells, and SVF cells were able to give rise to hematopoietic colonies in methylcellulose. We demonstrate the presence of hematopoietic progenitors in adipose tissue by phenotypic and functional characteristics. Thus adipose tissue could be considered as an important and convenient source of cells able to support hematopoiesis.     

Early membrane injury in lethally irradiated salivary gland cells

The early manifestations of radiation injury in salivary glands were investigated in the rat. The animals received a single X-ray dose in the range of 200-2000 rad to their neck area. Glandular changes during the first 24 hours were studied by light and electron microscopy and by measuring serum amylase activity. The amount of cell necrosis was quantitated and expressed as necrosis index (NI), Parotid NI and serum amylase activity 24 hours following irradiation were directly proportional to the X-ray dose. The submandibular gland cells were radioresistant and so were the mucous cells of the sublingual gland. The major increase in parotid acinar cell necrosis occurred between 12 and 24 hours after irradiation. However, more than 100 per cent increase in serum amylase level was detected prior to the onset of any significant cell necrosis. As early as two hours following irradiation signs of cell membrane injury were demonstrable in the parotid by electron microscopy and consisted of intracellular oedema, sequestered degenerative cell membranes, and an accumulation of intramitochondrial particles. None of these changes was detectable in the submandibular gland. The implication of membrane injury in the lethal effects of radiation on parotid cells is discussed.     

Transplantation of bone marrow cells into lethally irradiated mice. Morphology of megakaryocytes]

We described morphological changes of megakaryocytes in the bone marrow and spleen of lethally irradiated mice, dependent on the time lapse following bone marrow transplantation. The functional active megakaryocytes of various morphological types were found to predominate in the tissues on about day 20 to 25.     

In vivo natural antitumor resistance against murine EL-4 lymphoma cells in lethally irradiated syngeneic C57Bl/6 mice

Natural resistance has been detected in lethally irradiated C57Bl/6 (B6) mice inoculated intravenously with the ascites form of a syngeneic B6 leukemia. EL-4 cells were injected into lethally irradiated (800 R) B6 mice and tumor cell proliferation was evaluated by 125IUdR uptake in different organs 4 days after the challenge. Differential growth of lymphoma cells was observed when young mice were injected as compared with older mice and when mice were treated with agents known to interfere with natural resistance (e.g., poly(I:C), FLV-P, carrageenan, cyclophosphamide, high doses of irradiated cells). Similar results were obtained by measuring rapid clearance of 125IUdR-labeled EL-4 cells from lungs of intact B6 mice. In vivo cold competition studies, employing EL-4 and several other tumor lines of the same or different haplotype, showed that only EL-4 and RBL-5 cells were capable of inhibiting syngeneic resistance against EL-4 tumor. On the contrary, YAC-1 lymphoma cells, the most susceptible target to natural killer-mediated cytotoxicity in vitro, did not compete. These results suggest that EL-4 cells express membrane determinants not detectable on normal H-2b parental bone marrow cells and are susceptible to natural resistance against hemopoietic tumor cells in lethally irradiated syngeneic B6 mice.     

Effect of erythrocyte degradation products on the migration of hematopoietic stem cells in lethally irradiated mice

In experiments on CBA mice it was shown that erythrocytes administered at the stage of prehemolysis or the stromal fraction of erythrolysate caused an additional increase in the haemopoietic stem cell migration which had been intensified by hemorrhage or hypoxic hypoxia.     

Graft-versus-host reaction-like phenomenon induced by BCG in mice lethally irradiated and transferred with syngeneic bone marrow cells.

Heat-killed BCG in paraffin exerted a lethal effect on CS7BL/6 mice irradiated lethally and transferred with syngeneic bone marrow cells. Such an effect was not detectable when mice were subjected to adult thymectomy and used as the hosts. Lymphoid cells from such nonthymectomized mice exhibited cytotoxicity to syngeneic tumor cells but not to allogeneic tumor cells in an in vivo cytotoxicity test and induced splenomegaly in sublethally irradiated syngeneic recipients after systemic transfer. The cytotoxicity of such lymphoid cells was abolished by a treatment with anti-θ serum and complement. In the bone marrow of mice irradiated and transferred with bone marrow cells, the number of nucleated cells, the ratio of myeloid to erythroid cell series, and the percentage of lymphocytes were increasd by BCG injection. These results suggest the possibility that self-tolerance may be broken by BCG stimulation in the process of reconstitution of lymphoid cells in the irradiated mice.     

T-cell differentiation in lethally irradiated and reconstituted mice: Functional recovery of PNA-fractionated subpopulations

After lethal irradiation and reconstitution (IR) there is a 6-day lag before thymus cells start to proliferate. In the first 6 days there is a relative enrichment of PNA-negative cells, which are Ly-1⁺ and radioresistant. After IR the mitogenic responses of the PNA-negative cells recover earlier than those of the PNA-positive cells; the implications of this finding for different T-cell lineages are discussed.