Granulocyte colony-stimulating factor inhibits the endogenous leukotriene production in tumour patients

Summary. Granulocyte colony-stimulating factor (G-CSF) is virtually devoid of inflammatory side-effects when given to patients in therapeutic doses. This is in contrast to other haemopoietic cytokines such as granulocyte-macrophage colony-stimulating factor (GM-CSF) or interleukin-3 (IL-3) which may promote inflammatory reactions by increasing the number and/or activity of monocytes, eosinophils, mast cells and basophils. Inflammatory reactions to GM-CSF and IL-3 appear to be related to an increased formation of leukotrienes, known as potent mediators of allergy and inflammation. Here we report that, in contrast to GM-CSF or IL-3, G-CSF has the potential to inhibit the leukotriene production in vivo. G-CSF may thus act as an anti-inflammatory agent. The differential effects of G-CSF and other haemopoietins on endogenous leukotrienes may be of major clinical significance.     

Granulocyte colony-stimulating factor prevents progression of monocrotaline-induced pulmonary arterial hypertension in rats.

BACKGROUND: Regeneration of the lung microvasculature and replacing pulmonary artery lesions with functional endothelial cells could be a novel and effective therapeutic strategy for treating advanced pulmonary arterial hypertension (PAH). In the present study it was postulated that granulocyte colony-stimulating factor (G-CFS), which induces the proliferation of endothelial cells, would stimulate endothelial regeneration in situ at sites of impaired lung vasculature and prevent the development of PAH. METHODS AND RESULTS: Daily administration of G-CSF for 48 days did not affect the hemodynamism of normal Fischer 344 rats. PAH was induced with monocrotaline (60 mg/kg) and G-CSF was administered daily (100 microg/kg per day). Echocardiographic findings and an invasive catheter study indicated a significant decrease in the progression of PAH in rats given G-CSF. Furthermore, G-CSF increased Ki-67 positivity in the pulmonary arteries of PAH rats but did not accelerate c-kit positive cell recruitment into peripheral blood. Daily doses of G-CSF at both 2 and 100 microg/kg improved the survival and body weight gain of PAH rats. CONCLUSIONS: G-CSF improved the progression of PAH in a rat model, possibly by stimulating pulmonary endothelial cells to proliferate at sites of impaired lung vasculature. These findings show that cytokine therapy for PAH is valid based on the concept of vascular regeneration.     

Granulocyte colony-stimulating factor inhibits the mitochondria-dependent activation of caspase-3 in neutrophils.

The exact mechanism of apoptosis in neutrophils (PMNs) and the explanation for the antiapoptotic effect of granulocyte colony-stimulating factor (G-CSF) in PMNs are unclear. Using specific fluorescent mitochondrial staining, immunofluorescent confocal microscopy, Western blotting, and flow cytometry, this study found that PMNs possess an unexpectedly large number of mitochondria, which are involved in apoptosis. Spontaneous PMN apoptosis was associated with translocation of the Bcl-2-like protein Bax to the mitochondria and subsequent caspase-3 activation, but not with changes in the expression of Bax. G-CSF delayed PMN apoptosis and prevented both associated events. These G-CSF effects were inhibited by cycloheximide. The general caspase inhibitor z-Val-Ala-DL-Asp-fluoromethylketone (zVAD-fmk) prevented caspase-3 activation and apoptosis in PMNs, but not Bax redistribution. PMN-derived cytoplasts, which lack a nucleus, granules, and mitochondria, spontaneously underwent caspase-3 activation and apoptosis (phosphatidylserine exposure), without Bax redistribution. zVAD-fmk inhibited both caspase-3 activation and phosphatidylserine exposure in cultured cytoplasts. Yet, G-CSF prevented neither caspase-3 activation nor apoptosis in cytoplasts, confirming the need for protein synthesis in the G-CSF effects. These data demonstrate that (at least) 2 routes regulate PMN apoptosis: one via Bax-to-mitochondria translocation and a second mitochondria-independent pathway, both linked to caspase-3 activation. Moreover, G-CSF exerts its antiapoptotic effect in the first, that is, mitochondria-dependent, route and has no impact on the second.     

粒细胞集落刺激因子通过PI3K-Akt通路抑制缺血再灌注损伤

目的:研究经冠状动脉(冠脉)内注入粒细胞集落刺激因子(G-CSF)对大鼠心肌缺血再灌注损伤的预防作用,及其与PI3K-Akt信号转导途径的关系。方法:制备大鼠心肌缺血再灌注模型。再灌注同时经冠脉内给予G-CSF或G-CSF+PI3K激酶抑制剂(LY294002),灌注120min后应用TUNEL法检测细胞凋亡,免疫组织化学法观察凋亡相关蛋白Bax及Bcl-2及caspase-3表达情况,免疫印迹检测总-Akt(t-Akt)及磷酸化Akt(p-Akt)表达。结果:缺血再灌注同时给予冠脉内G-CSF治疗能够显著减轻缺血区心肌细胞凋亡,降低Bax、caspase-3的表达,增高心肌细胞内Bcl-2表达,同时p-Akt的蛋白水平显著增高(P<0.01)。LY294002阻断Akt活化后,抑制了G-CSF诱导的抗心肌细胞凋亡作用。结论:缺血再灌注同时冠脉内给予G-CSF可保护梗死区残存的心肌细胞,减少缺血再灌注所诱导的心肌细胞凋亡,其保护机制与PI3K-Akt信号通路有关。     

Granulocyte colony-stimulating factor producing lung large cell carcinoma with sarcomatous transformation]

We report a case of granulocyte colony-stimulating factor (G-CSF) producing lung large cell carcinoma with sarcomatous transformation. A 57-year-old man was admitted for evaluation of an abnormal shadow near the right pulmonary hilus on chest X-ray film. Brush cytology specimen from right B6 disclosed large cell carcinoma. His clinical stage was already IIIB (T3N3M0) on admission, so we immediately administered chemotherapy and radiation therapy. However, the patient's condition gradually deteriorated and he died of respiratory failure. Although there was no evidence of infection throughout his clinical course, his peripheral white blood cell count gradually increased and reached 47,000/mm3 (neutrophils 96%) before death. Histological study of the autopsy specimen revealed that the primary tumor was composed of two different elements (large cell carcinoma and spindle cell sarcomatoid element). The spindle cell sarcomatoid element appeared to have arisen from sarcomatous transformation of carcinoma cells, because the transition margin of the carcinoma to the sarcomatoid element was smooth, and special and immunohistochemical staining of both elements showed the same properties. Moreover, immunohistochemical study with monoclonal antibody to human G-CSF, 4A6 clearly demonstrated granular staining of G-CSF in the cytoplasm of tumor cells.     

Granulocyte colony-stimulating factor inhibits spontaneous cytochrome c release and mitochondria-dependent apoptosis of myelodysplastic syndrome hematopoietic progenitors

Low-risk myelodysplastic syndromes (MDS), including refractory anemia and sideroblastic anemia, are characterized by increased apoptotic death of erythroid progenitors. The signaling pathways that elicit this pathologic cell death in MDS have, however, remained unclear. Treatment with erythropoietin in combination with granulocyte colony-stimulating factor (G-CSF) may synergistically improve the anemia in patients with MDS, with a concomitant decrease in the number of apoptotic bone marrow precursors. Moreover, we have previously reported that G-CSF inhibits Fas-induced caspase activation in sideroblastic anemia (RARS). The present data demonstrate that almost 50% of erythroid progenitor cells derived from patients with MDS exhibit spontaneous release of cytochrome c from mitochondria with ensuing activation of caspase-9, whereas normal erythroid progenitors display neither of these features. G-CSF significantly inhibited cytochrome c release and suppressed apoptosis, most noticeably in cells from patients with sideroblastic anemia. Furthermore, inhibition of caspase-9 suppressed both spontaneous and Fas-mediated apoptosis of erythroid progenitors in all low-risk MDS cases studied. We propose that the increased sensitivity of MDS progenitor cells to death receptor stimulation is due to a constitutive activation of the mitochondrial axis of the apoptotic signaling pathway in these cells. These studies yield a mechanistic explanation for the beneficial clinical effects of growth factor administration in patients with MDS, and provide a model for the study of growth factor-mediated suppression of apoptosis in other bone marrow disorders.     

Granulocyte colony-stimulating factor improves function of neutrophils from patients with acute liver failure

OBJECTIVE: To evaluate the in vitro effects of granulocyte colony-stimulating factor (G-CSF) on function of neutrophils in acute liver failure (ALF). METHODS: Neutrophil functions (superoxide and hydrogen peroxide production; phagocytosis and killing; complement receptor expression) were determined simultaneously in 23 patients with ALF due to paracetamol overdose and compared with 23 healthy control subjects. RESULTS: Phagocytosis was reduced in neutrophils from ALF patients compared to controls (P< 0.005) and was significantly increased by incubation with 1,000 or 5,000 IU/ ml G-CSF (P< 0.05). This correlated with increased expression of CD11b (r= 0.93) and CD18 (r= 0.98) after incubation with 5,000 IU/ml G-CSF (P< 0.05). Killing was reduced in ALF neutrophils compared to controls (P< 0.005) and was similarly restored by G-CSF (P< 0.005). An increase in killing correlated with increases in production of superoxide (r = 0.96) and hydrogen peroxide (r= 0.97) by ALF neutrophils after incubation with 1,000 and 5,000 IU/ml of G-CSF when formylmethionylleucylphenylalanine (fMLP) was the stimulant. G-CSF at 5,000 IU/ml increased the production of hydrogen peroxide (P< 0.01) when zymosan was the stimulant. CONCLUSIONS: G-CSF improves the neutrophil dysfunction of ALF.     

Granulocyte colony-stimulating factor after allogeneic bone marrow transplantation

Hematopoietic growth factors have been shown to be effective in reducing the period of neutropenia after autologous bone marrow transplantation (BMT). Initial concerns over potential aggravation of graft-versus-host disease (GVHD) and increase in the incidence of relapse in patients with myeloid leukemias influenced the number of studies using hematopoietic growth factors after allogeneic BMT. We report the experience with 50 patients treated at a single institution using granulocyte colony-stimulating factor (G-CSF) after allogeneic sibling (n = 30) and matched unrelated (n = 20) BMT. The time to an absolute neutrophil count > or = 500/microL was significantly faster in patients who received G-CSF and cyclosporine and prednisone for GVHD prophylaxis when compared with historical control patients receiving the same GVHD prophylaxis (10 v 13 days, P < .01). A similar accelerated myeloid engraftment was observed for those patients who received the addition of methotrexate for GVHD prophylaxis when compared with historical control patients receiving the same GVHD prophylaxis regimen (16 v 19 days, P < .05). The median time to engraftment for patients receiving a matched unrelated BMT and G-CSF was 17 days (range 13 to 26). We did not observe any increase in GVHD or early mortality in the matched related sibling BMT. The incidence of acute GVHD in the matched unrelated BMT recipients was also low at 21%; however, 9 patients (45%) died within 100 days of the date of BMT, similar to the experience reported with granulocyte-macrophage CSF. This study confirms the efficacy of G-CSF in accelerating myeloid engraftment after allogeneic matched sibling BMT. The higher early mortality associated with patients receiving matched unrelated BMT suggests that randomized controlled trials using G-CSF after allogeneic BMT should be performed.     

粒细胞集落刺激因子促进小鼠肝再生的作用

目的:探讨粒细胞集落刺激因子(granulocyte colony-stimulating factor,G-CSF)促进肝大部切除小鼠肝再生的作用.方法:建立小鼠肝大部分切除(约70%)模型,将肝切除小鼠随机分为3组.单纯肝切除组:肝切除后24 h,腹腔注射生理盐水5 d:G-CSF+肝切除组:rhG-CSF 150 μg/(kg·d)腹腔注射5 d,24 h后进行肝切除:肝切除+G-CSF组:肝切除术后24 h,rhG-CSF 150 μg/(kg·d)腹腔注射5 d.于术后7 d取血清和肝组织,用全自动生化分析仪检测血清肝功能指标,并采用免疫组织化学方法观察肝内的增殖细胞核抗原(proliferating cell nuclear antigen, PCNA)的表达及BrdU阳性细胞.结果:肝切除+G-CSF组和G-CSF+肝切除组Brdu及PCNA表达及BrdU阳性细胞明显升高,与对照组比较差异均具有统计学意义(PCNA:74.08%±8.86%,68.91%±9.64% vs 57.36%±13.37%,均P<0.05);肝组织病理检查发现G-CSF+肝切除组27%(3/11)小鼠肝组织出现了不同程度的炎症反应,肝功生化学也显示血清ALT及AST升高,且其差异有统计学意义(均P<0.05).结论:G-CSF具有明显促进肝大部切除小鼠肝再生的作用,但也容易诱发肝脏炎症反应.     

Granulocyte colony-stimulating factor and neutrophils--forgotten mediators of inflammatory disease

Recent studies have highlighted the functional capacity of neutrophils as powerful mediators of tissue inflammation. Granule-packaged proteases and reactive oxygen intermediates, which are important for intracellular digestion during phagocytosis, are released from neutrophils during inflammation. In the extracellular environment, neutrophil-derived proteases can cause local tissue damage, but also regulate the activity of cytokines, cytokine receptors and chemokines. Neutrophils can themselves produce an array of inflammatory mediators, including cytokines, chemokines and complement; these cells also express Fc receptors, which can bind and possibly transport immune complexes into the extravascular compartment, as well as activating neutrophils at opsonised surfaces. Blood-borne neutrophils interact with, and then exit through, the endothelium of blood vessels, after which these cells die and must be removed safely. The balance between neutrophil survival and clearance is crucial to the resolution of inflammation. A major regulator of neutrophil production and survival is the cytokine granulocyte colony-stimulating factor (G-CSF). Treatment with G-CSF can exacerbate underlying inflammatory diseases in humans and mice, and G-CSF deficiency is profoundly protective against collagen-induced arthritis in mice. These findings implicate G-CSF as an important proinflammatory cytokine. This article discusses the roles of neutrophils and G-CSF during chronic inflammatory diseases.     

Granulocyte colony-stimulating factor receptors in human acute myelocytic leukemia

The binding of granulocyte colony-stimulating factor (G-CSF) to normal and human acute myeloid leukemia (AML) cells was investigated with radiolabeled recombinant human G-CSF (rhG-CSF). In all 14 cases of primary AML specific receptors for G-CSF were demonstrated on purified blast cells. The average numbers of G-CSF receptors ranged from very low to 428 receptors per cell (mean). Normal granulocytes showed G-CSF binding sites on their surface at higher densities (703 to 1,296 sites per cell). G-CSF receptors appeared to be of a single affinity type with a dissociation constant (kd) ranging between 214 and 378 pmol/L for AML blasts and 405 to 648 pmol/L for granulocytes. In 12 of 14 cases, including those with relatively low specific binding, G-CSF was a potent inducer of DNA synthesis of blasts in vitro; therefore, apparently relatively few receptors are required to permit activation of AML cell growth. However, in two cases cell cycling was not activated in response to G-CSF despite G-CSF receptor availability. The results show that G-CSF receptors of high affinity are frequently expressed on the blasts of human AML, but their presence may not be a strict indicator of the proliferative responsiveness of the cells to G- CSF.     

Granulocyte colony-stimulating factor production by adult T-cell leukaemia cells

We previously demonstrated that, in about 30% of primary adult T-cell leukaemia (ATL) cases, the leukaemic cells proliferated in response to granulocyte colony-stimulating factor (G-CSF). In the present report, we describe five patients with the acute leukaemia type of ATL who showed marked neutrophilia and elevated serum G-CSF concentrations in the absence of infection. We further examined two of these patients for detailed clinical features and cellular characteristics of the tumour cells. The white blood cell counts of both patients were 62 x 10(9)/l, consisting of approximately 90% neutrophils and 10% ATL cells. Serum concentrations of G-CSF in the two patients were 138 pg/ml and 93 pg/ml. The G-CSF concentrations in supernatants of short-term cultures of the patients' peripheral blood T-cells were 2 5 pg/ml and 13 pg/ml respectively. Immunostaining with anti-G-CSF antibody demonstrated G-CSF production by primary ATL cells in both cases. The neutrophil count fluctuated simultaneously with activity of ATL. Primary ATL cells from one patient were shown to proliferate in response to G-CSF in vitro. These results suggest autocrine growth stimulation of primary ATL cells in a subgroup of patients.