巨噬细胞集落刺激因子与血管平滑肌细胞增殖

目的一些以血管平滑肌细胞（ＶＳＭＣ）增殖为特点的血管疾病，在病变部位常有巨噬细胞浸润。本研究巨噬细胞集落刺激因子（ＭＣＳＦ）在ＶＳＭＣ生长调节中的作用。方法实验采用培养大鼠主动脉ＶＳＭＣ，细胞增殖观察指标采用氚标胸腺嘧啶核苷掺入法，并用Ｎｏｒｔｈｅｒｎｂｌｏｔ技术测定原癌基因表达。结果（１）Ｌ９２９细胞上清液（富含ＭＣＳＦ）及重组ＭＣＳＦ以剂量依赖关系刺激氚标胸腺嘧啶核苷掺入；（２）ＶＳＭＣ在接受刺激后表达某些原癌基因，如ｃｆｏｓ、ｃｍｙｃ、ｅｒｇ１和ＪｕｎＢ；（３）凝血酶、ＰＤＧＦ、ｂＦＧＦ与ＭＣＳＦ在促增殖作用上具有协同作用。结论ＭＣＳＦ与其它生长因子协同作用，通过自分泌／旁分泌机制调控ＶＳＭＣ增殖，从而可能在血管病变的形成和进展中起重要作用     

Macrophage colony stimulating factor prevents the progression of atherosclerosis in Watanabe heritable hyperlipidemic rabbits.

The early atherosclerotic lesion is characterized by the presence of macrophage-derived foam cells. Macrophage colony stimulating factor (M-CSF) specifically stimulates the functions of the monocyte-macrophages. To elucidate the effects of M-CSF in the atherogenic process in vivo, we administered human recombinant M-CSF into Watanabe heritable hyperlipidemic (WHHL) rabbits, an animal model for familial hypercholesterolemia. Three hundred micrograms of M-CSF were intravenously injected into WHHL rabbits aged 2.5 months, three times a week for 8.5 months. After the M-CSF treatment, we found very retarded progression of atherosclerosis. The accumulation of cholesterol ester was remarkably decreased in the aortae of M-CSF-treated animals (0.60 +/- 0.32 mg/g tissue), as compared to those of controls (4.32 +/- 0.61 mg/g tissue). Furthermore, the percentage of the surface area of the aorta with macroscopic plaque in animals treated with M-CSF was 14.3 +/- 6.2%, much less than that in controls receiving saline injection (38.8 +/- 8.0%). Thus, M-CSF definitely prevented the progression of atherosclerosis in WHHL rabbits by influencing macrophage functions.     

The relationship between humoral stimulating activity and colony stimulating factor

Humoral stimulatory activity (HSA) is detected in the sera of neutropenic rodents and humans by its effects on in vitro marrow cell [3H] TdR incorporation. Increased levels of CSA are also found in such sera; however, the relationship between these two activities has not been established. To examine this question, sera were obtained from mice, rats and humans at the peak occurrence of HSA following cyclophosphamide administration and were incubated with rabbit antiserum directed against mouse L-cell CSF. Anti-CSF neutralized the HSA effect on species-specific marrow cell [3H] TdR incorporation relative to the effects of normal sera with an efficiency of anti-CSF produced graded neutralization of the human HSA effect on human marrow cell proliferation. This neutralizing effect of anti-CSF on human HSA was obrogated by prior incubation of anti-CSF with sheep-anti-rabbit-IgG (S-anti-R-IgG) serum. The human serum activities on marrow cell proliferation, as determined by the [3H] TdR incorporation assay, correlated closely with their effects on human granulocyte colony formation in agar culture. These data indicate that human and rodent drug-induced HSA are cross-reactive with CSF as measured by a simple proliferation assay.     

Macrophage colony stimulating factor modulates the development of hematopoiesis by stimulating the differentiation of endothelial cells in the AGM region

Definitive hematopoietic stem cells arise in the aorta-gonad-mesonephros (AGM) region from hemangioblasts, common precursors for hematopoietic and endothelial cells. Previously, we showed that multipotential hematopoietic progenitors and endothelial cells were massively produced in primary culture of the AGM region in the presence of oncostatin M. Here we describe a role for macrophage-colony-stimulating factor (M-CSF) in the development of hematopoietic and endothelial cells in AGM culture. The number of hematopoietic progenitors including multipotential cells was significantly increased in the AGM culture of op/op embryos. The addition of M-CSF to op/op AGM culture decreased colony-forming unit (CFU)-GEMM, granulocyte macrophage-CFU, and erythroid-CFU, but it increased CFU-M. On the other hand, the number of cells expressing endothelial markers, vascular endothelial-cadherin, intercellular adhesion molecule 2, and Flk-1 was reduced in op/op AGM culture. The M-CSF receptor was expressed in PCLP1(+)CD45(-) cells, the precursors of endothelial cells, and M-CSF up-regulated the expression of more mature endothelial cell markers-VCAM-1, PECAM-1, and E-selectin-in PCLP1(+)CD45(-) cells. These results suggest that M-CSF modulates the development of hematopoiesis by stimulating the differentiation of PCLP-1(+)CD45(-) cells to endothelial cells in the AGM region.     

Macrophage colony stimulating factor restores in vivo bone resorption in the op/op osteopetrotic mouse

The op/op variant of murine osteopetrosis is a recessive mutation characterized by impaired bone resorption due to lack of osteoclasts. Cultured osteoblasts and fibroblasts from this mutant do not secrete M-CSF activity and resident macrophages are absent in bone marrow. This failure has been related to a mutation within the M-CSF coding region. We report now that the administration of recombinant human M-CSF (rhM-CSF) corrects in vivo the impaired bone resorption in this animal. The treatment restores the bone marrow cavity virtually absent in the op/op animal and induces the appearance of resorbing osteoclasts and of resident bone marrow macrophages. This proves that the deficiency of M-CSF is the cause of the op/op bone disorder and that this cytokine is directly or indirectly necessary for physiological osteoclastogenesis, the resulting bone resorption and for the establishment of bone marrow hemopoiesis.     

The marrow colony forming cell and serum colony stimulating factor of the chicken.

Marrow cells of the chicken produced colonies in semisolid media. Developing colonies consisted of granulocytes, macrophages or a mixture of these two cell types. The granulocyte-macrophage CFC was nonadherent. An adherent 'CFC' was also present and it differed in several ways from the nonadherent CFC: (a) clones contained only macrophages, (b) they contained a core of nonrefractile cells, (c) their appearance was delayed 1-2 weeks, (d) they were unaffected by the presence of erythrocytes and (e) the efficiency of cloning was increased but the percentage of clones able to produce 50 or more cells was markedly decreased, i.e., the cluster/colony ratio was increased. The growth of both colony types was strictly dependent on the presence of CSF. Data obtained from dose-response studies on unfractionated marrow indicated that clusters and colonies were derived from single cells. The CSF of chicken serum yielded sigmoid dose-response curves when tested on marrow cells. Calf serum could not support cluster or colony formation when tested alone but it did have an enhancing effect on the CSF of chicken serum. Levels of serum CSF were increased by injecting chickens with bacterial endotoxin. This phenomenon occurred with five chicken lines tested, but certain chickens of the Kimber line did not respond to endotoxin with elevated levels of CSF.     

Purification and properties of bacterially synthesized human granulocyte-macrophage colony stimulating factor

Human granulocyte-macrophage colony stimulating factor (GM-CSF) has been synthesized in high yield using a temperature inducible plasmid in Escherichia coli. The human GM-CSF is readily isolated from the bacterial proteins because of its differential solubility and chromatographic properties. The bacterially synthesized form of the human GM-CSF contains an extra methionine residue at position 1, but otherwise it is identical to the polypeptide predicted from the cDNA sequence. The specific activity of 2.9 X 10(7) units/mg of protein for purified bacterially synthesized human GM-CSF indicates that despite the lack of glycosylation, the molecule is substantially in its native conformation. This molecule stimulated the same number and type of both seven- and 14-day human bone marrow colonies as the CSF alpha preparation from human placental conditioned medium. Human GM-CSF had no activity on murine bone marrow or murine leukemic cells. There was no detectable, direct stimulation of adult human erythroid burst forming units (BFU-E) by the bacterially synthesized human GM-CSF. Although impure preparations containing native human GM-CSF (eg, human placental conditioned medium) stimulated the formation of mixed colonies, even in the presence of erythropoietin, the bacterially synthesized human GM-CSF failed to stimulate the formation of mixed colonies from adult human bone marrow cells. The bacterially synthesized human GM-CSF increased N-formyl-methionyl-leucyl- phenylalanine (FMLP)-induced superoxide production and lysozyme secretion. Antibody-dependent cytotoxicity and phagocytosis by human neutrophils was stimulated by the bacterially synthesized human GM-CSF and eosinophils were also activated in the antibody-dependent cytotoxicity assay.     

Granulocyte colony stimulating factor in neutropenic patients with infective endocarditis

A well known complication in the treatment of infectious endocarditis is development of neutropenia caused by treatment with antibiotics in high concentrations over long periods. Neutropenia often necessitates discontinuation of antibiotic treatment. Three patients with infectious endocarditis who developed neutropenia are reported. The patients were treated with granulocyte colony stimulating factor (G-CSF), a haematopoietic growth factor that stimulates neutrophils. G-CSF induced an immediate increase in white blood cell count, primarily neutrophils. G-CSF may be effective in ameliorating neutropenia in patients who receive antibiotics for treatment of infectious endocarditis.

Keywords: granulocyte colony stimulating factor;  neutropenia;  endocarditis     

Binding of colony stimulating factor by sterile filtration membranes.

Loss of serum-free L-cell colony stimulating factor (CSF) activity was noted when non-sterile or ethylene oxide sterilized Millipore filters were employed; autoclaved membranes showed no inactivation. The reduction in CSF did not appear to be due to release of inhibitory or inactivating substances as extensive rinsing with various solvents did not prevent loss of CSF activity. In addition, these filtrates did not inhibit the activity of standard CSF. Less pure sources of CSF, including standard serum containing L-cell CSF and human urinary concentrate, did not lose activity upon filtration. Moreover, the addition of bovine serum albumin to serum-free CSF completely prevented membrane filtration loss. The CSF loss could be prevented by treatment of the membranes with buffers contraining 0.05% polyethylene glycol (PEG). These findings show that Millipore membranes can bind significant quantities of partially purified CSF. It is important in purification studies to recognize that CSF loss can be prevented by addition of PEG to all buffer systems.     

Granulocyte-macrophage colony stimulating factor exacerbates collagen induced arthritis in mice

OBJECTIVE—To examine the effect of granulocyte-macrophage colony stimulating factor (GM-CSF) on disease progression in the collagen induced arthritis (CIA) model in mice.
METHODS—DBA/1 mice were primed for a suboptimal CIA response by intradermal injection of chick type II collagen without a secondary immunisation. Three weeks after immunisation the mice were given four to five consecutive daily intraperitoneal injections of recombinant murine GM-CSF (15 µg; 5 × 10⁵ U), or vehicle, and arthritis development was monitored by clinical scoring of paws and calliper measurements of footpad swelling. At approximately six to eight weeks after immunisation mice were killed, their limbs removed and processed for histological analyses of joint pathology.
RESULTS—Control animals receiving a single immunisation with collagen exhibited a varied CIA response both in terms of incidence and severity. Mice treated with GM-CSF at 20 to 25 days after immunisation with collagen had a consistently greater incidence and more rapid onset of disease than the vehicle treated control mice, based on clinical assessment. GM-CSF treated mice showed higher average clinical scores and greater paw swelling than controls. Histological analyses of joints reflected the clinical scores with GM-CSF treated mice displaying more pronounced pathology (synovitis, pannus formation, cartilage and bone damage) than control mice.
CONCLUSION—GM-CSF is a potent accelerator of the pathological events leading to chronic inflammatory polyarthritis in murine CIA supporting the notion that GM-CSF may play a part in inflammatory polyarthritis, such as rheumatoid arthritis.

     

Granulocyte-macrophage colony stimulating factor: an adjuvant for cancer vaccines

Granulocyte-macrophage colony stimulating factor (GM-CSF) enhances immune responses by inducing the proliferation, maturation, and migration of dendritic cells, and the expansion and differentiation of B and T lymphocytes. There is significant data in pre-clinical animal models demonstrating the adjuvant effects of GM-CSF in a variety of cancer vaccine approaches, including cellular vaccines, viral vaccines, peptide and protein vaccines, and DNA vaccines. GM-CSF is an attractive vaccine adjuvant because of its immune modulation effects and low toxicity profile. The results in animal models have been confirmed in pilot clinical trials and several clinical trials are currently ongoing.     

Sulfasalazine induced agranulocytosis treated with granulocyte-macrophage colony stimulating factor.

We report the use of granulocyte-macrophage colony stimulating factor (GM-CSF) in a case of rheumatoid arthritis with sulfasalazine induced agranulocytosis, leading to a rapid bone marrow recovery within 7 days. This case and 2 others reported in the literature emphasize the need for further research into the possible role of GM-CSF in the treatment of drug induced agranulocytosis.     

Circadian variation of granulocyte colony stimulating factor levels in man

Glucocorticoids dose-dependently increase plasma levels of granulocyte colony stimulating factor (G-CSF). Based on the marked circadian rhythm of cortisol levels, we hypothesized that plasma levels of G-CSF may also show a diurnal rhythm. A prospective study was conducted in 12 healthy young volunteers. Blood samples were obtained every 2 h over 24 h. G-CSF levels averaged 18.0 ng/l (CI 13. 1-22.9) at 8.00 am, increased continuously and reached peak values at 10.00 p.m. Individual harmonic regression analysis showed a clear circadian rhythm. The individual differences between nadir and peak levels averaged 54% (CI 43-65%). This pronounced diurnal rhythm of G-CSF levels may help understand the circadian changes in circulating stem cells, bone marrow DNA synthesis, or bone marrow toxicity induced by some chemotherapeutic agents.