Two kinds of extracts of peripheral blood leukocytes induced erythroblast increase and maturation in uitro in rabbit marrow cells. One extract was a secretion or exudate into plasma from intact leukocytes; the other was the supernatant after homogenizing the cells in ethanol. The extracts were less active than the intact leukocytes. The response with the two extracts together was a little greater than with either alone, but less than additive.
Mitoses are rare in basophilic erythroblasts in stained smears of normal rabbit marrow cells, and relatively common among the acidophilic erythroblasts, usually 0.5-1 % , even without treatment which accumulates them. The difference cannot be asscribed to slow DNA synthesis in the former. On the contrary, DNA synthesis is more intense in the basophilic (1-9) and the DNA doubling time is about one-third that of the acidophilic erythroblasts (10). The initial purpose of the experiments described here was to observe the effects of colcemid treatment. This substance arrests mitoses in prophase-metaphase and thus causes them to accumulate. One might then see mitoses in basophilic erythroblasts. The long diameters of the erythroblasts were measured and these observations suggested answers to two other questions in this area. One concerns the timing of when the basophilic erythroblasts lose their cytoplasmic basophilia relative to their cell division. The other is the origin of the large acidophilic erythroblasts called Line 2 cells (1 1, 12). Their average diameter is that of the large basophilic erythroblasts; compared to polychromatic erythroblasts, their cytoplasm stains a clear yellow-orange without the hint of green; they are seen in mitosis more frequently.Materials and methods. Normal, adult white New Zealand rabbits obtained commercially were used. Colcemid was injected intravenously in doses ranging from 0 to 5 mg/kg. After 6 hr, which is the longest time before the cells escape from the effects of colcemid, the animals were killed, smears made of the marrow cells, fixed in absolute methanol, and stained with benzidine and Giemsa solutions (13). The long diameters of the erythroblasts were measured on the stained smear by means of a micrometer eyepiece. The size recorded was the nearest whole number on the scale; 1 division = 0.794 nm. Eleven animals were used. 2 1,574 erythroblasts were tallied and classified, 4,142 were measured. Table I1 gives the size distribution in five classes of erythroblasts: "large" and "small" basophilic erythroblasts, Line 2 cells, and acidophilic erythroblasts not in mitoses and in mitoses. The partition of the basophilic erythroblasts into "large" and "small" was based on the hypothesis that some of the large gave rise to the Line 2 cells, the "small" to the acidophilic erythroblasts in mitosis, in each case without change in size. The numerical details are as follows. As there were a negligible number of acidophilic erythroblasts in mitosis of size 13 (scale divisions) or larger, the percent of the total Line 2 cells of sizes 13, 14, and 15 was taken to be equal to the percent of "large" basophilic erythroblasts of the same sizes, in this case 48.7%. There were 366 basophilic erythroblasts of these sizes; the total of "large" basophilic erythroblasts was therefore 366/48.7 X 100 = 751. The total basophilic erythroblasts counted were 976; the total of "small" basophilic erythroblasts was therefore 976 -751 = 225. Their size distribution was taken to be the same as that of the aci...
Blood leukocytes incubated in vitro with rabbit-marrow cells induced a several-fold increase in basophilic erythroblasts and smaller increases in acidophils and reticulocytes. The main effect was nearly complete in one hour at 37". Erythropoietin augmented the leukocyte effect; anti-erythropoietin inhibited i t with or without erythropoietin. The erythroblast increase came entirely from the marrow cells; the precursor cell class has not been identified, except that division of pre-existing basophils appears to be excluded. Autologous and homologous leukocytes were about equally effective.A method is described of measuring on stained smears changes in relative concentrations of different classes of cells induced experimentally. A method of preparing highly concentrated peripheral blood leukocytes is described.
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