Serum Colony-stimulating Factor Research Articles

Colony-stimulating Factor in Patients With Chronic Neutropenia

Urinary and serum colony-stimulating factor (CSF) levels were measured in 11 patients with chronic idiopathic neutropenia without infections and in 10 normal individuals. Urinary CSF output was determined using mouse marrow target cells, and serum CSF activity was assayed with human marrow target cells by the double agar layer technique. Using these methods, there was no significant difference between CSF levels of neutropenic and normal subjects. These data indicate that CSF levels are not inversely related to the blood neutrophil count in chronic idiopathic neutropenia and suggest that CSF is not a hormone regulating the blood neutrophil count in a manner analogous to the erythropoietin regulation of circulating erythrocyte levels.

Lipid A, the active part of bacterial endotoxins in inducing serum colony stimulating activity and proliferation of splenic granulocyte/macrophage progenitor cells.

An analysis of which component of lipopolysaccharides (LPS), the lipid or the polysaccharide (PS), is active in stimulating the murine granulopoietic system has been performed. LPS with different structures, isolated from different mutant strains of Salmonella and chemical degradation products of lipopolysaccharides have been used. Lipid A obtained by acid hydrolysys of the LPS and complexed to bovine serum albumin (BSA) (lipid A-BSA) was shown to be active in generating serum colony stimulating factor (CSF) and in increasing the splenic colony forming cells (CFC) levels, although it was less active than the parent LPS. The polysaccharide (PS) showed no significant activity at the concentrations used. LPS (glycolipids) from R mutants of Salmonella minnesota were active to the same extent as the LPS. The fact that even the most defective LPS from the R mutant R595 which contains lipid A and KDO only is a potent endotoxin, points unequivocally, to lipid A, as the active principle in stimulating the granulopoietic system.

Tolerance to the granulocyte-releasing and colony-stimulating factor elevating effects of endotoxin

Injection of Salmonella typhosa endotoxin into either CF1 or C57bl/6J mice leads to prompt increases in serum colony-stimulating factor (CSF). Repeated injections of endotoxin result in a dose-related hyporesponsiveness or tolerance to this effect. Tolerance is seen after either intravenous (i.v.) or intraperitoneal (i.p.) routes of administration or challenge and occurs after one to two preinjections. Cross-tolerance to heterologous endotoxin (Escherichia coli) was also shown. This cross-tolerance is complete immediately after cessation of preinjections, but partial at later time intervals. Levels of a serum inhibitor of colony growth were decreased in tolerant mice, although this decrease is not statistically significant. Tolerant mice injected with endotoxin release granulocytes from the bone marrow normally, in spite of the absence of a CSF response. This suggests that neutrophil releasing activity (NRA) and CSF are separate entities. A marked marrow granulocytic hyperplasia develops after 7 or 20 days of endotoxin injections, despite the tolerance to the CDF-elevating effect of endotoxin. This granulocytic hyperplasia could still be mediated by serum CSF increases. A negative medullary feed-back secondary to the repetitive release of marrow granulocytes, however, is an equally plausible mechanism for the stimulation of granulocyte production. It is also possible that the decrease in serum inhibitors played a role in the sustained increase in granulopoiesis seen here.

Tolerance to the Granulocyte-releasing and Colony-stimulating Factor Elevating Effects of Endotoxin

Injection of Salmonella typhosa endotoxin into either CF1 or C57bl/6J mice leads to prompt increases in serum colony-stimulating factor (CSF). Repeated injections of endotoxin result in a dose-related hyporesponsiveness or tolerance to this effect. Tolerance is seen after either intravenous (i.v.) or intraperitoneal (i.p.) routes of administration or challenge and occurs after one to two preinjections. Cross-tolerance to heterologous endotoxin (Escherichia coli) was also shown. This cross-tolerance is complete immediately after cessation of preinjections, but partial at later time intervals. Levels of a serum inhibitor of colony growth were decreased in tolerant mice, although this decrease is not statistically significant. Tolerant mice injected with endotoxin release granulocytes from the bone marrow normally, in spite of the absence of a CSF response. This suggests that neutrophil releasing activity (NRA) and CSF are separate entities. A marked marrow granulocytic hyperplasia develops after 7 or 20 days of endotoxin injections, despite the tolerance to the CDF-elevating effect of endotoxin. This granulocytic hyperplasia could still be mediated by serum CSF increases. A negative medullary feed-back secondary to the repetitive release of marrow granulocytes, however, is an equally plausible mechanism for the stimulation of granulocyte production. It is also possible that the decrease in serum inhibitors played a role in the sustained increase in granulopoiesis seen here.

Culture of autologous bone marrow in diffusion chambers: effect of host irradiation

Autologous bone marrow (BM) cells were cultured in diffusion chambers (DC) implanted into whole-body irradiated, non-irradiated, or sham- irradiated goats. Proliferation was apparent in DC implanted in both irradiated and nonirradiated goats. However, cells in DC cultured in irradiated hosts increased in number beginning earlier, proceeded at a faster rate, and reached higher numbers than in DC in nonirradiated hosts. Growth enhancement could not have occurred as a result of radiation-induced immunosuppression in autologous hosts. The nonirradiated “target cells” in the DC therefore constituted an indicator system for stimulatory or inhibitory substances in the host. The simultaneous increase in the number of granulocytes in peripheral blood and in DC of irradiated hosts was paralleled by an initial rise in serum colony-stimulating factor (CSF). A second, prolonged period of severe granulocytopenia following irradiation of the host correlated with high levels of serum CSF. Increased numbers of megakaryocytes were seen in DC as thrombocytopenia developed in the irradiated host. DC erythropoiesis disappeared rapidly in nonirradiated goats; however, in DC of irradiated goats, the number of erythrocytic precursors increased exponentially during ablation of host erythroid marrow. Anemia did not develop in the host during the culture period. Proliferation of mononuclear cells in DC was markedly stimulated by irradiation of the host. Proliferation of macrophages appeared independent of host treatment. These observations provide strong evidence for diffusion of specific and/or nonspecific humoral hematopoietic stimulators from the host into the DC. This stimulation appears to be elicited and/or intensified by irradiation of the host.

Culture of Autologous Bone Marrow in Diffusion Chambers: Effect of Host Irradiation

Autologous bone marrow (BM) cells were cultured in diffusion chambers (DC) implanted into whole-body irradiated, non-irradiated, or sham-irradiated goats. Proliferation was apparent in DC implanted in both irradiated and nonirradiated goats. However, cells in DC cultured in irradiated hosts increased in number beginning earlier, proceeded at a faster rate, and reached higher numbers than in DC in nonirradiated hosts. Growth enhancement could not have occurred as a result of radiation-induced immunosuppression in autologous hosts. The nonirradiated "target cells" in the DC therefore constituted an indicator system for stimulatory or inhibitory substances in the host. The simultaneous increase in the number of granulocytes in peripheral blood and in DC of irradiated hosts was paralleled by an initial rise in serum colony-stimulating factor (CSF). A second, prolonged period of severe granulocytopenia following irradiation of the host correlated with high levels of serum CSF. Increased numbers of megakaryocytes were seen in DC as thrombocytopenia developed in the irradiated host. DC erythropoiesis disappeared rapidly in nonirradiated goats; however, in DC of irradiated goats, the number of erythrocytic precursors increased exponentially during ablation of host erythroid marrow. Anemia did not develop in the host during the culture period. Proliferation of mononuclear cells in DC was markedly stimulated by irradiation of the host. Proliferation of macrophages appeared independent of host treatment. These observations provide strong evidence for diffusion of specific and/or nonspecific humoral hematopoietic stimulators from the host into the DC. This stimulation appears to be elicited and/or intensified by irradiation of the host.

The role of humoral factors in the regression of leukemia in chickens as measured by in vitro colony formation

AbstractLeukemic myeloblasts induced by avian myeloblastosis virus in the chicken formed small compact (type II) colonies in semi‐solid agar medium. Normal yolk sac cells from 12‐day old embryos formed large diffuse (type I) colonies under the same conditions. Type I colony formation (but not type II) was strictly dependent upon the presence in the medium of a colony stimulating factor (CSF) present in fresh chicken serum or conditioned medium. Serum CSF levels were determined for normal, leukemic, and birds which had spontaneously regressed from myeloblastic leukemia. When type I colony formation was used as the assay, serum CSF levels of leukemic birds were found to be significantly lower than levels in either normal or regressed birds. When the same sera were tested for their ability to induce type II colonies, leukemic birds demonstrated a significantly higher CSF level than either normal or regressed sera. Regressed chickens had serum CSF levels similar to normal birds.

The bone marrow colony stimulating factor (CSF): relation of serum CSF to urine CSF.

SummaryC57BL mouse urine CSF was found to be of lower molecular weight than normal or postendotoxin mouse serum CSF. In addition the fall in activity of postendotoxin serum CSF was found to be unaltered by bilateral nephrectomy. Because of these findings, a reevaluation of previous data was made. It was concluded that the kidney may play little or no part in the clearance of serum CSF. The origin of urinary CSF was not established.

Local and systemic control of granuloctic and macrophage progenitor cell regeneration after irradiation.

ABSTRACTAgar cultures of C57BL bone marrow cells were used to determine colony stimulating factor (CSF) and serum CSF‐inhibitor levels in C57BL and BALB/c mice following irradiation. Whole‐body irradiation caused an acute, dose‐dependent, rise in serum CSF levels and fall in CSF‐inhibitor levels. The regeneration of granulocytic and macrophage progenitor cells (in vitro CFCs) in the femur after 250 rads whole‐body irradiation was preceded or paralleled by a fall in serum CSF‐inhibitors and a dramatic rise in the capacity of bone‐adherent cells in the marrow (‘stromal cells’) to produce material with colony‐stimulating activity. No comparable changes were observed in the activity of marrow haemopoietic cells during regeneration or in the lungs or spleen. A similar rise in the activity of bone‐adherent cells was observed in shielded femurs during regeneration of in vitro CFCs.Regeneration of granulocytic and macrophage progenitor cells following irradiation may be regulated by fluctuations in circulating CSF‐inhibitor levels and local production of CSF within the marrow cavity.

Effect of Endotoxin on Granulopoiesis and the In Vitro Colony-forming Cell

Abstract The effect of endotoxin on the tibial in vitro colony-forming cell (CFC) and the differentiated granulocyte compartment was evaluated. The injection of 5 µg of Salmonella typhosa endotoxin into CF1 mice leads to elevated serum colony-stimulating factor (CSF) levels along with a transient granulocytopenia followed by the release of granulocytes from the marrow. The present study demonstrates that the number of tibial CFC, as assayed on soft agar, decreases within 20 min after endotoxin, reaches a nadir at 6 hr (65.8 ± 4.9% of control), and then returns to control values by 48 hr. Concomitantly, over the 48 hr after endotoxin, there are sequential increases in the marrow myeloblast-promyelocyte compartment (MPC), myelocyte compartment (MC), and the PMN compartment. Radioautographic studies utilizing 3HTdR showed increased labeling indices and grain counts in the MPC and MC at 24 or 48 hr after endotoxin, indicating a shortening of generation time and perhaps of the DNA synthetic period in these compartments. The present data suggest that elevated CSF levels seen after endotoxin administration may lead to differentiation of the marrow CFC into the granulocyte pathway. There are also changes in the differentiated granulocyte compartment, suggesting a decreased generation time that may be due to CSF or a separate regulator such as antichalone.

Humoral Stimulators of Granulocyte Production

Abstract These studies were designed to determine whether colony stimulating factor (CSF) and diffusible granulocytopoietic stimulator (DGS) represented a single substance which stimulates the production of granulocytes in vitro and in vivo. The effect of endogenous CSF upon DGS activity was studied after injection of 40 µg endotoxin. This resuited in both DGS activity and high serum CSF levels, but the time of appearance and duration of the two activities followed different patterns. In particular, CSF peaked and was declining 24 hr following endotoxin at which time DGS was not yet detectable. Then, as serum CSF fell, the DGS activity rose. These data support the concept that the DGS activity detectable with short term Millipore chamber cultures is not due to CSF and suggests the effects of CSF in vitro and DGS in vivo are due to different factors.

Tissue culture of calf bone marrow. Requirements of serum and conditioned media.

The agar culture method for growing colonies of granulocytes and macrophages from bone marrow cells has provided a technique for assaying a humoral factor, the colony-stimulating factor (CSF), which is found in plasma (1) and urine (2). This factor appears to be similar to CSF produced by feeder layers of bone marrow (3), peripheral leukocytes (4) or kidney cells (5). In general, colonies of bone marrow do not form unless this stimulant is continually present. Shad-duck and Nunna (1) found elevation of CSF in serum only during experimentally induced neutropenia, favoring the view that CSF is a humoral regulator of myelopoiesis.Although serum does not usually stimulate bone marrow colonies, Bradley, Metcalf and Robinson (6) found CSF in serum of AKR mice when leukemia developed. Bradley and Siemienowicz (7) also reported stimulation of rat bone marrow by normal rat serum. In both cases, however, fetal bovine serum was used in the culture. The question of whether normal serum can stimulate bone marrow witho...

Studies on the bone marrow colony stimulating factor (CSF): relation of tissue CSF to serum CSF.

AbstractFluctuations in the colony stimulating factor (CSF) content of submaxillary salivary gland, lung, kidney and spleen were studied in male C57BL mice which had been subjected to a variety of stimuli (endotoxin, x‐irradiation or polyadenylic‐polyuridylic acid complex) that caused elevations of the serum CSF. The temporal relationships and magnitudes of the rises in CSF were complex and differed from tissue to tissue according to the stimulus used. In the tissues examined not only was the measurable CSF content in each case found to equal or exceed the prestimulatory levels, but in some cases distinctly different forms of CSF were observed as shown by differences in the zone sedimentation, electrophoretic, and calcium phosphate binding characteristics of the material. The patterns of response to the stimuli investigated suggested that tissue injury either directly, or indirectly via the release of various cellular constituents, might mediate the release of CSF by various widely disseminated and/or differing cell types.

Granulocyte colony stimulating factor. 3. Effect of alkylating agent induced granulocytopenia.

SummaryAt intervals following a single injection of cyclophosphamide, groups of rats were tested for the presence of an in vitro granulocyte colony stimulating factor (CSF). Increased serum CSF activity was demonstrated on days 5 through 7 after injection, corresponding to a phase of marked peripheral granulocytopenia. The CSF returned to base line levels as peripheral blood granulocytes returned to normal. Increased elaboration of CSF in conjunction with granulocytopenia is in accord with a possible role of this material as a regulator of granulopoiesis.

Colony-stimulating Factor and Inhibitor Levels in Acute Granulocytic Leukemia

Abstract Assays were performed on bone marrow colony-stimulating-factor (CSF) levels in the serum and urine of 33 patients with acute granulocytic and myelomonocytic leukemia. Of 251 serum samples, 30% showed elevated CSF levels as did 53% of 1422 urines tested. Levels of serum inhibitors of colony formation were abnormally low or undetectable in 57% of sera from leukemic patients. Patients in relapse were unable to respond to infections by developing high serum CSF levels and low serum-inhibitor levels were more common in patients with a short survival time and in some types of patient during relapses.

Serum Half Life in Mice of Colony Stimulating Factor Prepared from Human Urine

Summary. Partially purified preparations of the colony stimulating factor (CSF) from human urine were given as a single intravenous injection to adult C57BL mice. Assays for human CSF in the sera of these mice were performed in cultures of C57BL bone marrow cells using a discriminatory rabbit anti‐human CSF antibody preparation, incapable of blocking the action of mouse serum CSF. The half life of injected CSF varied from 1.7 to 3.7 hr (mean 2.9 hr). The injection of these human urine preparations caused a temporary elevation of mouse CSF levels in the serum of recipient mice, which prolonged the effective period of elevated CSF levels in the serum following injection.

Inhibition of bone marrow colony formation by normal and leukaemic human serum.

A FACTOR is present in the serum and urine of mice and humans which can stimulate the formation of granulocytic and macrophage colonies by mouse bone marrow cells in agar culture1–3. Levels of this colony stimulating factor (CSF) can be assayed in bone marrow cultures because of the sigmoid relationship between CSF concentration and colony numbers3. Previous studies showed that serum CSF levels were elevated in mice with various types of leukaemia1,4. Although serum colony stimulating activity seemed to be high in some patients with leukaemia5, a direct comparison of CSF levels in normal and leukaemic human sera was prevented by an inability to detect CSF in normal human serum.

Studies on colony-stimulating factor (CSF): role of the kidney in clearing serum CSF.

SummarySerum CSF levels in C57BL mice rose after bilateral nephrectomy and bilateral ureter tie. These procedures prevented the fall of serum CSF levels caused by cortisone. The urine of cortisone-injected mice contained more CSF than that of saline-injected mice. In mice, most of the CSF in the urine appears to be CSF cleared by the kidney from the serum.

Bone Marrow Colony-Stimulating Factor Following Ureteral Ligation in Germfree Mice

SummarySix days after unilateral ureteral ligation, serum colony-stimulating factor (CSF) is four times as high as in nonoperative germfree mice. The elevation in serum CSF is associated with a significant increase in peripheral blood granulocytes, but not lymphocytes, monocytes, or eosinophils. This finding supports the view that CSF not only is capable of stimulating proliferation of granulocytes and mononuclear cells in vitro, but also promotes granulopoiesis in vivo, and can be regarded as a leukopoietin.

The effects of whole body irradiation on serum colony stimulating factor and in vitro colony-forming cells in the bone marrow.

Summary. Using in vitro cultures of mouse bone marrow cells, levels of colony stimulating factor were assayed in serum from mice following whole body irradiation. No significant differences from control levels were noted in the period 1–32 days following 50, 150, 250 or 450 rads. A dose of 250 rads caused a sharp fall in the level of colony‐forming cells in the bone marrow followed by regeneration between 6 and 16 days following irradiation.Six to 8 hours following whole body irradiation, blood polymorphs and serum colony stimulating factor levels were consistently elevated. Splenectomy did not affect this response.The results suggest that serum colony stimulating factor is unlikely to be the major regulator determining regeneration of in vitro colony‐forming cells in the bone marrow following irradiation.