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The existence of a possible local control of CFU turnover was studied in mice in which one tibia only was irradiated (LR mice) and in mice in which one tibia was shielded during whole‐body irradiation (‘TBR’mice). In both the LR and ‘TBR’mice, the increased CFU turnover continued in the irradiated tibiae even after the time when in the unirradiated (shielded) tibiae it returned to normal levels. The early temporary decrease in the CFU numbers in the shielded tibiae of ‘TBR’mice is attributed to a temporary demand for increased differentiation rather than to migration of CFU. The direct control of CFU turnover appears to be local in nature, in contrast to the stimulus for CFU differentiation.

Measurements to determine the kinetic status of the morphologically unrecognizable haemopoietic precursor cells in the bone marrow are frequently carried out using techniques which inhibit or destroy cells in the DNA‐synthetic (S) phase of the cell cycle. For example, tritiated thymidine (3H‐TdR) has for many years been recognized as a highly specific label for DNA synthesis and, as such, administration of large doses of 3H‐TdR has often been used, both in vitro and in vivo, to kill cells in S. Assay of the surviving cells has then given a measure of the proportion of the total cells which are in the S‐phase of the generation cycle. Other compounds which have been used for the same purpose are: 125Iodo‐deoxyuridine (125I‐UdR), another S‐phase specific label, or hydroxyurea (HU) which prevents entry of cells into S and inhibits or kills cells already in S (Sinclair, 1965). For a variety of reasons, different laboratories tend to make different choices of the agent to be used for this purpose. As a result, it has sometimes proved difficult to marry data obtained from different sources. In the course of using 3H‐TdR, tritiated uridine (3H‐Ur), 125I‐UdR and HU in attempts toevaluate the kinetic status of bone marrow stem cells, it has become clear that their use is not straightforward and this paper presents data which illustrate some of the pitfalls associated with their use.

The ability of protaglandins E1 and E2 to stimulate the proliferation of haemopoietic stem cells (CFUs) was studied in vivo. PGE2, in a dose range of 10‐4 to 10‐1μg/g body weight and PGE1 in a dose range of 10‐5 to 10‐1μg/g body weight, produced a rapid cycling wave of CFUs. The increase in the number of CFUs in S phase was not followed by a rise in the femoral CFUs content, and except for a transient increase in femoral CFUc level, no increase in differentiation was found either. Therefore, it is proposed that haemopoiesis after PG‐induced CFU stimulation is ineffective. PGE2 did not stimulate regeneration of CFUs in a perturbed state (after sublethal irradiation). All these findings support the idea that PGEs might represent potent stimulators of the haemopoietic stem cells acting in physiological doses. However, if acting concurrently with physiological control systems PGs lead to ineffective haemopoiesis (under normal conditions) or do not exert any measurable effect (after sublethal irradiation).

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The effect of mouse serum interferon (IF) in vitro and an inducer in vivo on the proliferation of a pluripotent stem cell population with high turnover rate was studied. Proliferation rate was characterized by the number of CFUs in the S phase of the cell cycle. Increased proliferation of bone marrow stem cell populations was produced either by irradiating the donor mice with 3·36 Gy (336 rad) 60Co‐gamma rays 7 days before the experiment or by incubating normal bone marrow cells with 10–11 M concentration of isoproterenol. IF considerably reduced the number of CFUs in S phase in both cases without reducing the CFUs content of the samples. Injection of IF inducer (4 mg/kg poly I:C) into regenerating mice also inhibited the proliferation of CFUs without decreasing the femoral CFUs level. Regeneration kinetics of CFUs from irradiated poly I:C‐treated mice ran parallel with that of irradiated untreated animals but showed a characteristic delay corresponding to approximately one CFUs doubling. A transient, non‐cytotoxic proliferation inhibitory effect of IF or IF inducer is, therefore, proposed.

Increasing number of data suggests that locally produced histamine is involved in regulation of hematopoiesis. In this study the granulocyte/macrophage (CFU-GM) colony formation by normal murine or human bone marrow cells, leukaemic colony formation (CFU-L) by a murine leukemia cell line (WEHI 3B), and colony formation by bone marrow cells from patients with chronic myeloid leukemia (CML) have been examined. We detected mRNA and protein expression of histidine decarboxylase (HDC), the only enzyme responsible for histamine synthesis both in normal bone marrow progenitor cells and in leukaemic progenitors. The significance of in situ generated histamine was shown on colony formation by inhibitory action of alphaFMH (blocking HDC activity, i.e. de novo histamine formation) and by N,N-diethyl-2-[4-(phenylmethyl)phenoxy]-ethanamine-HCl (DPPE) disturbing the interference of histamine with intracellular binding sites. These data provide further confirmation of the role of histamine in development and colony formation of bone marrow derived cells.

IntroductionHematopoietic colony formation, the assembly of cycling and differentiating descendants of pluripotent progenitor cells, is regulated by both external signals and intracellular promoting factors. Among the external signals, interleukin (IL)-3 exerts growth stimulatory effects in colony formation of murine myeloid progenitor CFU-GM (granulocyte-macrophage) cells [1]. IL-3 further proved to enhance the synthesis of endogenous histamine in low density, progenitor-enriched cells of murine bone marrow [2]. Besides the intracellular synthesis of histamine, hematopoietic progenitors are known to take up exogenous histamine by a mechanism indepenedent of surface receptors [3]. Furthermore, exogenous histamine stimulates the division of progenitor cells through H2 receptors [4]. In this study, the clonogenic potencies of murine bone marrow CFU-GM (normal) and of CFU-L (leukemic) progenitor cells were studied under the influence of histamine receptor antagonists and an irreversible blocker of histidine decarboxylase (HDC). Materials and methods Clonogenic cells in cultureFor plating, bone marrow cells were suspended in McCoy5A medium supplemented with horse serum (10%) and the supernatant of a WEHI 3B myelomonocytic leukemia cell line. The latter contains IL-3, the cytokine responsible for increasing histamine synthesis, thereby keeping the cells in cycle. Cell viability was estimated with trypan blue (1 %) exclusion, and in the given concentration range more than 95 % of cells were viable.Semisolid methylcellulose culturing conditions were used for following the clonogenic capacity of murine femoral bone marrow CFU-GM (normal) cells, and of murine CFU-L (L: leukemic = WEHI 3B myelomonocytic leukemia) cells. The colony number was counted by day 7, when a colony was represented by a minimum of 50 cells in assembly. Experimental changes in the number of colonies were recorded as percentages of the controls (= 100 %) with no treatment. Influencing the activity of HDCThe activity of the HDC was irreversibly inhibited by alpha-fluoromethyl-histidine (FMH) [5]. The concentration of FMH applied was 10 -3 -10 -7 M. The effective range which produced a minimum of 30 % inhibition in colony formation was 10 -3 -10 -5 M.Triprolidine and cimetidine, (H 1 and H 2 receptor antagonists, respectively) were applied in concentrations of 10 -5 -10 -8 M. The effective range which produced a minimum of 30 % inhibition in colony formation was 10 -5 M-10 -7 M. Results and discussionThe effects of histamine antagonists on the colony number of murine CFU-GM and CFU-L cells are shown on Table 1. Cimetidine in murine bone marrow cells of other studies [3] did not inhibit the uptake of histamine but did block IL-3 induced DNA synthesis [4]. In our experiments, cimetidine produced a 20-30% inhibition of colony formation in murine CFU-GM, and a 40-50% inhibition in the CFU-L cells. Regarding the dual signal transduction of histamine H 2 receptors [6], murine CFU-L cells seem to utilize more intensely both the cyclic-AMP and the phospholipase...