Macrophages as origin of factor increasing monocytopoiesis.

Earlier investigations had indicated that the factor increasing monocytopoiesis (FIM), present in the serum of mice and rabbits during the onset of an inflammatory response, is released by cells of the inflammatory exudate. The present study was performed to determine which cells produce and secrete this factor and to establish the kinetics of its production and secretion. FIM was assayed in vivo by intravenous injection of samples into untreated mice and monitoring the course of the number of blood monocytes in the recipients. FIM was assayed in vitro by adding samples to cultures of the macrophage cell line PU5 and determining the rate of proliferation of the cells. The results show that only macrophages contain and synthesize FIM. This factor is secreted upon exposure to a phagocytic stimulus, and after the release of preformed FIM, macrophages secrete newly synthesized FIM. Granulocytes and lymphocytes neither contain nor secrete FIM. The characteristics of FIM derived from macrophages are in all aspects similar to those of FIM in serum. Macrophage-derived FIM is a protein with a molecular weight between 10 and 25 X 10(3), its activity is cell-lineage specific and dose dependent, and it stimulates monocyte production in the bone marrow. Macrophage-derived FIM is not identical to either CSF-1 or IL-1, and has no chemotactic activity. Taken together, the present results show that FIM occurring in serum during an inflammatory response originates from macrophages at the site of the inflammation. In this way the macrophages themselves regulate the supply of circulating blood monocytes that can migrate to the site of injury when needed.

Method to select mice in the steady state for biological studies.

Collection of small amounts of blood from the orbital sinus was found to be a satisfactory method for repeated sampling in mice, which means that these animals can be selected for further study on the basis of the leukocyte count. In biomedical research it is often necessary to have detailed information about the effect of injected material on the numerical course of circulating leukocytes. However, the present study has shown that 2 stress-producing procedures on 1 day disturb the steady state, and that this disturbance is expressed in changes in the number of leukocytes. Such stress could be avoided by alteration of the experimental design to include only 1 stressful situation each day. When blood was sampled in the orbital sinus on 1 day to determine the animals' condition and on the next day only the tail was punctured as sham injection, the number of blood leukocytes remained constant throughout the observation period. Comparative studies on the numbers of monocytes, lymphocytes, and granulocytes in blood from the tail, heart, and orbital sinus showed a systematic difference in the mean numbers of certain types of leukocytes. Statistically significant differences were found between the mean numbers of monocytes, lymphocytes, and granulocytes in orbital blood from normal mice of 5 specific pathogen-free strains, i.e., Cpb:SE (Swiss), CBA/Cpb, BALB/cCpb, C3H/Rij, and DBA/2Rij.

Presence of the factor increasing monocytopoiesis (FIM) in rabbit peripheral blood during an acute inflammation.

An intraperitoneal injection of latex in rabbits was found to give rise to an increase in the number of macrophages at the site of inflammation and a concomitant monocytosis in the peripheral blood. The results showed that during the initial phase of the inflammatory reaction a humoral factor is present in the circulation of these animals that stimulates the monocyte production in the bone marrow in a concentration-dependent way. This factor has been called the factor increasing monocytopoiesis (FIM), in analogy with the name given to the factor previously found in mice. Rabbit FIM is cell-line specific since it has no effect on granulocyte or lymphocyte production, has an estimated molecular weight of between 10,000 and 25,000 daltons, was found to be sensitive to treatment with proteases, to be unaffected by glycosidases, and to be readily inactivated in vitro at 37 degrees C. Neither rabbit nor mouse FIM is species specific, since rabbit FIM evoked moderate monocytosis in mice and vice versa.

Properties of a factor increasing monocytopoiesis (FIM) occurring in serum during the early phase of an inflammatory reaction.

A factor increasing monocytopoiesis (FIM) has been demonstrated during the onset of an acute inflammatory reaction caused by an intraperitoneal injection of polystyrene latex particles. It is protein in nature, does not contain a carbohydrate moiety essential for its function, and is very probably not a glycoprotein. The molecular weight of FIM lies between 18,000 and 24,000 daltons (determined with both ultrafiltration membranes and gel filtration on Sephadex G100). The monocytosis induced by FIM is dose dependent. FIM is thermolabile, having a half-time of about 20 min at 37 degrees C in serum; temperature inactivation can be delayed by the addition of epsilon-aminocaproic acid, the half-time at 37 degrees C then being about 45 min. In vitro treatment of normal murine blood with the inducers of the inflammatory reaction does not result in FIM activity in the serum. FIM dose not have chemotactic activity toward macrophages, is not a clotting factor, is not a biologically active fragment of the complement system, and has no colony-stimulating or-enhancing activity in the vitro bone marrow colony assay. On the basis of these results, a mechanism is postulated for the humoral regulation of monocytopoiesis.

A serum facted by newborn calf serum.

An intraperitoneal injection of newborn calf serum (NBCS) into CRF Swiss mice causes an inflammatory reaction characterized by an increase in the number of macrophages in the peritoneal cavity and a concomitant monocytosis. The serum of such mice contains a monocytosis-inducing factor, as demonstrated by the intravenous injection of serum collected 18 (CalS18) and 24 hr (CalS24) after the intraperitoneal injection of NBCS. Serum from normal untreated mice, from mice given an intraperitoneal injection of sterile pyrogen-free saline, which does not cause an inflammatory reaction, or from mice 72 hr after an intraperitoneal injection of NBCS, when the inflammatory reaction has subsided, does not cause a monocytosis in test mice. Intravenous injection of CalS18 causes not only a monocytosis but also an increase in the number of promonocytes and bone marrow monocytes, suggesting an increased in the number of promonocytes and bone marrow monocytes, suggesting an increased production of monocytes. The effect of CalS18, CalS24 and CalS18 filtrate is specific for the mononuclear phagocytes, since only non-significant increases in the numbers of lymphocytes and granulocytes were observed. The active factor in CalS18 was shown to be different from the monocytosis-inducing factor present in NBCS. The monocytosis-inducing factor in CalS18 passes through an ultrafiltration membrane with an exclusion limit of 50,000 Daltons, so that the molecular weight must be below this value.