粒细胞集落刺激因子/粒-巨噬细胞集落刺激因子联合动员异基因造血干细胞移植

背景：目前FDA已批准应用于临床外周血造血干细胞移植的动员剂只有粒细胞集落刺激因子和粒-巨噬细胞集落刺激因子,其中粒细胞集落刺激因子单药应用是目前最主要的动员方案,但可引起供者骨骼肌肉酸痛、发热等不良反应。 目的：回顾性分析以粒细胞集落刺激因子/粒-巨噬细胞集落刺激因子联合应用为动员方案的异基因造血干细胞移植临床效果。 方法：选择2004-01/2009-10河南省人民医院血液科以粒细胞集落刺激因子与粒-巨噬细胞集落刺激因子联合应用为动员方案进行血缘全相合异基因造血干细胞移植51例,分析移植物成分、造血重建及移植物抗宿主病的发生率。 结果与结论：动员96 h后 CD34+细胞占单个核细胞的比例为(0.97±0.13)%,CD34+CD38-细胞占CD34+细胞的比例为(37.49±4.03)%;移植后的快速造血重建与 CD34+细胞、CD34+CD38-细胞输入量呈负相关。Ⅰ度、Ⅱ~Ⅳ度急性移植物抗宿主病的发生率分别为25.5%,15.7%;局限性、广泛性慢性移植物抗宿主病的发生率分别为39.2%,21.2%。提示在血缘全相合异基因造血干细胞移植中,粒细胞集落刺激因子/粒-巨噬细胞集落刺激因子联合可有效实现干细胞动员,所获CD34+细胞完全可以满足快速造血重建的需要;输入较多的CD34+细胞、CD34+CD38-细胞可能利于快速造血重建。     

国产与进口粒细胞集落刺激因子动员外周血造血干细胞的效果对照

为评估国产重组人粒细胞集落刺激因子在动员外周造血干细胞的效果,选择2001/2005收治的20例自体外周血造血干细胞移植患者,男11例,女9例,其中急性髓细胞白血病11例,急性淋巴细胞白血病5例,恶性淋巴瘤3例,多发性骨髓瘤1例。根据患者对药价承受能力分为2组：国产重组人粒细胞集落刺激因子动员组、进口重组人粒细胞集落刺激因子动员组,每组各10例,除急性髓细胞白血病采用大剂量阿糖胞苷动员外,其他均应用大剂量环磷酰胺动员,当白细胞降至低谷开始回升时加国产与进口重组人粒细胞集落刺激因子,白细胞升至5×109 L-1以上开始采集外周血造血干细胞。所有患者均移植成功,两者在给药剂量、用药天数,采集干细胞质量、造血功能重建及药物毒副反应等方面差异均无显著性意义。     

粒细胞集落刺激因子与异基因造血干细胞移植中移植物抗宿主病及移植物抗白血病的免疫调节效应

文章从细胞水平、动物模型、临床应用等方面对粒细胞集落刺激因子在移植物抗宿主病和移植物抗白血病中的免疫调节作用作进一步的研究。得出结论：粒细胞集落刺激因子可从T细胞、抗原递呈细胞、调节性T细胞、自然杀伤细胞等途径发挥其免疫调节作用,诱导移植物中的T细胞产生免疫耐受,使得移植物抗宿主病和移植物抗白血病效应产生分离,从而改善异基因造血干细胞移植患者的预后,因此,输注经粒细胞集落刺激因子动员的供者淋巴细胞是其中值得关注的、有前途的一种方法。但其作用的途径机制、输注的时机、输注的细胞数等仍有待进一步的实验室和临床研究。     

造血干细胞动员和采集对正常供者的影响

背景：粒细胞集落刺激因子动员是当前健康志愿供者采用的主要方法。据国内外文献报告：在粒细胞集落刺激因子动员中已发生少数严重不良反应,这引起了国人对中国非血缘健康供者安全的忧虑。粒细胞集落刺激因子作为动员剂对健康志愿供者有何影响？是否安全？ 目的：观察粒细胞集落刺激因子对正常供者的影响。 方法：选择2003-01/2008-12在海口市人民医院接受粒细胞集落刺激因子5~10 μg/(kg·d)动员剂进行外周血造血干细胞捐赠者16例,观察动员及采集过程的不良反应,检测动员前后血常规CD3、CD19、CD3+4、CD3+8细胞比值在动员前后变化,并随访所有供者。 结果与结论：10例供者在重组人粒细胞集落刺激因子动员过程中无任何不适,有3例出现低热,头痛、肌肉及骨骼疼痛、腰痛等,3例供者出现发热,其严重程度均在Ⅰ级,但无需终止动员。白细胞经重组人粒细胞集落刺激因子动员后数量较动员前升高,停止动员后3 d全部供者的白细胞恢复至动员前水平。血红蛋白、血小板、CD3、CD19、CD3+4、CD3+8细胞比值在动员前、动员后72,96 h无明显变化。提示健康供者可耐受粒细胞集落刺激因子5~10 μg/(kg·d),动员和采集过程;且对T细胞亚群无影响。     

粒细胞集落刺激因子动员对大鼠内皮祖细胞体外扩增的影响

背景：内皮祖细胞在缺血性疾病治疗应用中存在数量低的问题，许多细胞因子影响其动员效果和功能。 目的：观察粒细胞集落刺激因子动员对体外扩增骨髓源性内皮祖细胞数量及功能的影响。 设计、时间及地点：细胞学体外观察，于2007-08/2008-03在北京大学人民医院肝病研究所完成。 材料：SPF级健康成年SD雄性大鼠16只，随机分为动员组和对照组，8只/组。重组人粒细胞集落刺激因子为麒麟鲲鹏生物药业有限公司产品。 方法：实验前动员组大鼠皮下注射经生理盐水稀释的重组人粒细胞集落刺激因子10μg/kg，2次/d，共5 d。采用密度梯度离心法分别从两组大鼠骨髓获取单个核细胞，洗涤后按2×107/孔接种在包被大鼠纤连蛋白的6孔板上，采用贴壁法纯化得到内皮祖细胞。 主要观察指标：骨髓单个核细胞集落数及细胞表型，骨髓源性内皮祖细胞贴壁情况及细胞表型，通过DiI-acLDL摄取、体外血管生成进行内皮祖细胞功能学鉴定，透射电镜观察细胞超微结构。 结果：与对照组比较，动员组骨髓单个核细胞集落增多，CD45+/CD34+、CD133+和Flk-1+阳性率均明显升高（F=5.655~61.892，P < 0.05）。培养第7，9天与对照组比较，动员组骨髓源性内皮祖细胞贴壁数、CD45+/CD34+、CD133+和Flk-1+阳性率均明显升高（F=5.694~38.879，P < 0.05）。贴壁细胞F1TC-UEA-1/Dil-acLDL双荧光染色阳性率为90%，接种于Matrigel包被的培养板24 h后形成毛细血管索样结构，胞质内有Weibel-Palade小体存在。 结论：粒细胞集落刺激因子动员能增加骨髓源性内皮祖细胞的数量，并促进其分化、增殖功能。     

粒细胞集落刺激因子神经保护作用的研究进展

粒细胞集落刺激因子（Granulocyte Colony—Stimulating Factor,G—CSF）首先发现于1980年,Metcalf等在研究中发现内毒素处理过的小鼠血清或条件培养基中的一种活性成分,这种成分具有诱导小鼠骨髓单核细胞白血病细胞集落分化的功能,被命名为粒细胞-巨噬细胞分化因子（Granu—loycyte—Macrophage Differentiation Factor．GMDF）,此后研究发现它能在体外选择性地刺激造血前体细胞分化为粒细胞集落,因此将它改称为G—CSF。时至今日,G—CSF已逐渐应用于血液系统、心血管系统、神经系统等疾病的治疗,是一个非常有临床发展前景的新药。     

尾静脉注射骨髓基质细胞和腹腔注射粒细胞集落刺激因子治疗大鼠脑梗死*☆

背景：骨髓间充质干细胞具有成神经分化特性,有很多试验也证实粒细胞集落刺激因子可以用于改善脑梗死后的神经功能。 目的：比较静脉移植骨髓间充质干细胞和腹腔注射粒细胞集落刺激因子动员干细胞来治疗大脑中动脉闭塞模型大鼠疗效。 方法：实验以改良的Zea-longa线栓法阻断大脑中动脉建立SD大鼠脑梗死模型,造模24 h后分别通过尾静脉注射骨髓间充质干细胞或腹腔注射粒细胞集落刺激因子。 结果与结论：两种治疗方法均可改善脑梗死模型大鼠的运动和认知功能,且粒细胞集落刺激因子对脑梗死模型大鼠的运动和认知功能的改善比尾静脉注射骨髓间充质干细胞明显,移植后第7,14天,粒细胞集落刺激因子组梗死面积小于骨髓间充质干细胞组(P < 0.05),粒细胞集落刺激因子组BrdU阳性细胞数多于骨髓间充质干细胞组(P < 0.05)。提示粒细胞集落刺激因子动员骨髓干细胞治疗脑梗死的疗效可能优于骨髓间充质干细胞静脉注射的移植方法。     

银屑病患者骨髓基质细胞分泌干细胞因子和粒细胞集落刺激因子的水平

背景：课题组在前期研究中发现银屑病患者骨髓造血细胞活性存在异常。 目的：观察银屑病患者骨髓基质细胞分泌干细胞因子和粒细胞集落刺激因子的水平。 设计、时间及地点：病例-对照观察,于2007-10/2008-08在太原市中心医院皮肤科实验室完成。 对象：选择太原市中心医院皮肤科临床及病理确诊为寻常型银屑病的门诊患者24例,男16例,女8例;年龄16~59岁;另取血液科骨穿后经筛选的正常骨髓20例作为对照组,性别、年龄与银屑病患者匹配。 方法：采用密度梯度离心法分离银屑病患者与对照组骨髓单一核细胞,通过贴壁法培养骨髓基质细胞,收集传了3代后又培养72 h的骨髓基质细胞及培养上清。 主要观察指标：流式细胞仪检测细胞表面标志并用ELISA法检测上清液中干细胞因子和粒细胞集落刺激因子的水平。 结果：流式细胞仪检测结果显示,90%以上细胞表面抗原高表达CD29,而CD34、CD45及HLA-DR表达阴性,即骨髓基质细胞纯度在90%以上。银屑病组骨髓基质细胞培养上清液的干细胞因子和粒细胞集落刺激因子表达均显著高于对照组 (P < 0.01)。 结论：银屑病患者骨髓基质细胞分泌干细胞因子和粒细胞集落刺激因子存在异常,提示银屑病患者骨髓造血微环境发生了改变。     

膀胱内灌注粒细胞-巨噬细胞集落刺激因子预防造血干细胞移植后出血性膀胱炎发生

背景：出血性膀胱炎是造血干细胞移植后常见并发症之一。粒细胞-巨噬细胞集落刺激因子除影响造血干/祖细胞的增殖和分化外,还能调节炎性反应中单核细胞、粒细胞、淋巴细胞以及内皮细胞的功能。 目的：探讨膀胱内灌注粒细胞-巨噬细胞集落刺激因子对造血干细胞移植后出血性膀胱炎的预防作用。 设计：病例分析。 对象：中山大学附属中山医院2004-01/2006-08接受异基因造血干细胞移植的血液病患者15例作为常规治疗组,2006-09/2008-12接受异基因造血干细胞移植的血液病患者16例作为粒细胞-巨噬细胞集落刺激因子组。 方法：常规治疗组采用常规方法即美司钠、水化、碱化尿液预防出血性膀胱炎,粒细胞-巨噬细胞集落刺激因子组在其基础上,在应用环磷酰胺前24 h开始向膀胱内灌注粒细胞-巨噬细胞集落刺激因子,至环磷酰胺停用3 d后拔除导尿管,用生理盐水冲洗膀胱后排空膀胱,然后向300 μg粒细胞-巨噬细胞集落刺激因子中加入生理盐水10 mL、利多卡因5 mL注入膀胱,保留60~120 min。 主要观察指标：出血性膀胱炎的发生及其与移植物抗宿主病的相关性,巨细胞病毒感染及泌尿系统感染的发生情况。 结果：与常规治疗组比较,粒细胞-巨噬细胞集落刺激因子组出血性膀胱炎发生率明显降低(χ2=4.39,P < 0.05),出血性膀胱炎平均持续时间、患者平均住院时间均明显缩短(t＝3.97,P < 0.05;t＝3.13,P < 0.05),Ⅲ度以上出血性膀胱炎发生率明显降低(χ2=5.04,P < 0.05)。出血性膀胱炎严重程度与移植物抗宿主病严重程度、持续时间有关(r=0.76)。与常规治疗组比较,粒细胞-巨噬细胞集落刺激因子组巨细胞病毒感染率略微下降但无明显差异(χ2=0.28,P > 0.05),巨细胞病毒感染阳性患者Ⅲ度以上出血性膀胱炎发生率升高。与常规治疗组比较,粒细胞-巨噬细胞集落刺激因子组泌尿系统感染发生率略微下降但无明显差异(χ2=0.28,P > 0.05)。 结论：粒细胞-巨噬细胞集落刺激因子膀胱内灌注耐受性好,临床取得了较好疗效,是预防造血干细胞移植后出血性膀胱炎的有效措施。     

粒细胞集落刺激因子动员外周血干细胞富集致肺损伤1例

背景：使用粒细胞集落刺激因子动员外周血干细胞后导致急性肺损伤仅有零星报道，机制尚不清楚。 目的：探讨应用重组人粒细胞集落刺激因子动员外周血干细胞富集导致肺损伤的临床表现、机制。 方法：报告1例粒细胞集落刺激因子动员外周血干细胞时导致咯血和影像学肺部结节状、片状阴影，并复习相关文献。 结果与结论：粒细胞集落刺激因子可能会导致肺损伤，机制可能和中性粒细胞在肺部聚集、黏附、释放炎症递质有关。     

黄精含药血清促进骨髓间充质干细胞增殖的效应及机制

背景：近年来,对传统单味中药有效成分、复方中药及含药血清调控骨髓间充质干细胞增殖与分化作用的研究已有报道,但鲜有涉及到机制的研究。 目的：观察黄精含药血清体外对骨髓间充质干细胞增殖的影响,并分析其作用途径与机制。 方法：雄性SD大鼠用于骨髓间充质干细胞的提取;雌性SD大鼠以中药水溶液灌胃后定时取血制备含药血清。全骨髓贴壁法分离培养大鼠骨髓间充质干细胞,将传至P3代的细胞,制备成单细胞悬液接种于96孔培养板后,分别加入黄精含药血清及空白血清,各组血清浓度分别为培养液体积的5%,10%,15%,20%,25%及30%。MTT法检测黄精含药血清对骨髓间充质干细胞增殖的影响;RT-PCR检测各细胞因子mRNA表达;实时PCR检测粒细胞集落刺激因子mRNA表达。 结果与结论：与空白血清组相比,体积分数10%的黄精含药血清具有明显促进骨髓间充质干细胞增殖的作用(P < 0.05),并显著促进粒细胞集落刺激因子mRNA的表达(P < 0.05)。提示体积分数10%的黄精含药血清具有促进骨髓间充质干细胞增殖的作用,可能与其促进粒细胞集落刺激因子mRNA表达有关。     

Increased platelet aggregation and in vivo platelet activation after granulocyte colony-stimulating factor administration. A randomised controlled trial

Granulocyte colony-stimulating factor (G-CSF) stimulates the bone marrow to produce granulocytes and stem cells and is widely used to accelerate neutrophil recovery after chemotherapy. Interestingly, specific G-CSF receptors have been demonstrated not only on myeloid cells, but also on platelets. Data on the effects of G-CSF on platelet function are limited and partly conflicting. The objective of this study was to determine the effect of G-CSF on platelet aggregation and in vivo platelet activation. Seventy-eight, healthy volunteers were enrolled into this randomised, placebo-controlled trial. Subjects received 5 μg/kg methionyl human granulocyte colony-stimulating factor (r-metHuG-CSF, filgrastim) or placebo subcutaneously for four days. We determined platelet aggregation with a whole blood impedance aggregometer with various, clinically relevant platelet agonists (adenosine diphosphate [ADP], collagen, arachidonic acid [AA], ristocetin and thrombin receptor activating peptide 6 [TRAP]). Filgrastim injection significantly enhanced ADP (+40%), collagen (+60%) and AA (+75%)-induced platelet aggregation (all p<0.01 as compared to placebo and p<0.001 as compared to baseline). In addition, G-CSF enhanced ristocetin-induced platelet aggregation (+18%) whereas TRAP-induced platelet aggregation decreased slightly (-14%) in response to filgrastim. While baseline aggregation with all agonists was only slightly but insignificantly higher in women than in men, this sex difference was enhanced by G-CSF treatment, and became most pronounced for ADP after five days (p<0.001). Enhanced platelet aggregation translated into a 75% increase in platelet activation as measured by circulating soluble P-selectin. G-CSF enhances platelet aggregation and activation in humans. This may put patients suffering from cardiovascular disease and cancer at risk for thrombotic events.     

G-CSF reduces infarct volume and improves functional outcome after transient focal cerebral ischemia in mice.

Growth factors possess neuroprotective and neurotrophic properties in vitro, but few have been extensively studied in vivo after stroke. In the present study, we investigated the potential functional benefits of granulocyte colony-stimulating factor (G-CSF) administration after focal cerebral ischemia. Male mice underwent 60-minute middle cerebral artery occlusion (MCAO) and received G-CSF (50 microg/kg, subcutaneously) or vehicle (saline) at the onset of reperfusion. Granulocyte colony-stimulating factor-treated mice killed at 48 hours after MCAO revealed a >45% reduction (P<0.05) in lesion volume. In terms of body weight recovery, and in tests of motor (grid test and rotarod) and cognitive ability (water maze), MCAO significantly worsened the outcome in vehicle-treated mice as compared with shams (P<0.05). However, G-CSF treatment was beneficial as, compared with vehicle, this significantly improved weight recovery and motor ability. This effect was most apparent on the water maze where G-CSF-treated mice were indistinguishable from shams in terms of acquiring the task. These results indicate long-term beneficial effects of a single dose of G-CSF administered on reperfusion, and illustrate the need to further investigate the mechanisms of G-CSF action.     

Granulocyte colony-stimulating factor attenuates neuronal death and promotes functional recovery after spinal cord injury in mice

Granulocyte colony-stimulating factor (G-CSF) is a protein that stimulates differentiation, proliferation, and survival of granulocytic lineage cells. Recently, a neuroprotective effect of G-CSF was reported in a model of cerebral infarction. The aim of the present study was to elucidate the potential therapeutic effect of G-CSF for spinal cord injury (SCI) in mice. We found that G-CSF is neuroprotective against glutamate-induced cell death of cerebellar granule neurons in vitro. Moreover, we used a mouse model of compressive SCI to examine the neuroprotective potential of G-CSF in vivo. Histologic assessment with cresyl violet staining revealed that the number of surviving neurons in the injured spinal cord was significantly increased in G-CSF-treated mice. Immunohistochemistry for neuronal apoptosis revealed that G-CSF suppressed neuronal apoptosis after SCI. Moreover, administration of G-CSF promoted hindlimb functional recovery. Examination of signaling pathways downstream of the G-CSF receptor suggests that G-CSF might promote functional recovery by inhibiting neuronal apoptosis after SCI. G-CSF is currently used in the clinic for hematopoietic stimulation, and its ongoing clinical trial for brain infarction makes it an appealing molecule that could be rapidly placed into trials for patients with acute SCI.     

Granulocyte colony-stimulating factor has a negative effect on stroke outcome in a murine model

The administration of CD34-positive cells after stroke has been shown to have a beneficial effect on functional recovery by accelerating angiogenesis and neurogenesis in rodent models. Granulocyte colony-stimulating factor (G-CSF) is known to mobilize CD34-positive cells from bone marrow and has displayed neuroprotective properties after transient ischemic stress. This led us to investigate the effects of G-CSF administration after stroke in mouse. We utilized permanent ligation of the M1 distal portion of the left middle cerebral artery to develop a reproducible focal cerebral ischemia model in CB-17 mice. Animals treated with G-CSF displayed cortical atrophy and impaired behavioral function compared with controls. The negative effect of G-CSF on outcome was associated with G-CSF induction of an exaggerated inflammatory response, based on infiltration of the peri-infarction area with CD11b-positive and F4/80-positive cells. Although clinical trials with G-CSF have been started for the treatment of myocardial and limb ischemia, our results indicate that caution should be exercised in applying these results to cerebral ischemia.     

Trophic effect of erythropoietin and other hematopoietic factors on central cholinergic neurons in vitro and in vivo

In vitro granulocyte colony-stimulating factor (G-CSF), macrophage colony-stimulating factor (M-CSF), erythropoietin (EPO), and erythroid differentiation factor (EDF) augmented choline acetyltransferase (ChAT) activity in mouse embryonic primary septal neurons and in cholinergic hybridoma cell line, SN6.10.2.2. This is similar to the effects seen with interleukin-3 (IL-3) or granulocyte-macrophage colony-stimulating factor (GM-CSF). Moreover, in vivo GM-CSF and EPO promoted survival of septal cholinergic neurons in adult rats which had undergone fimbria-fornix transections. These results suggest that some of the hematopoietic factors act on cholinergic neurons as ‘neurotrophic factors’ to influence the differentiation, maintenance and regeneration of these neurons.     

Granulocyte colony-stimulating factor promotes growth of processes,growth associated protein 43 and microtubule-associated protein 2 expression in cultured rat retinal ganglion cells in vitro

Following granulocyte colony-stimulating factor (G-CSF) treatment,the growth of processes in cul-tured rat retinal ganglion cells (RGCs) in vitro,expression of growth associated protein 43,and expression of microtubule-associated protein 2 mRNA expression were significantly increased.In contrast,RhoA/Rock protein content was significantly reduced by G-CSF treatment.These results indicate that G-CSF promotes the growth of processes in RGCs and increases the expression of growth-associated protein 43 and micr...     

粒细胞集落刺激因子可促进纯化培养大鼠视网膜神经节细胞突起生成和生长相关蛋白43及微管相关蛋白2的表达

发现粒细胞集落刺激因子干预的体外纯化培养大鼠视网膜神经节细胞突起增多,生长相关蛋白43和微管相关蛋白2 mRNA表达明显增加,RhoA/Rock蛋白含量显著下降。显示粒细胞集落刺激因子可以促进视网膜神经节细胞突起的生成,通过抑制Rho/Rock途径,促进轴突生成相关蛋白生长相关蛋白43及微管相关蛋白2表达增加,从而促进缺氧损伤视网膜神经节细胞的轴突修复。     

G-CSF administration is neuroprotective following transient cerebral ischemia even in the absence of a functional NOS-2 gene

Granulocyte colony-stimulating factor (G-CSF) is a candidate neuroprotective factor following cerebral ischemia. To determine whether G-CSF acts partly through the inhibition of nitric oxide synthase (NOS)-2 expression, we administered G-CSF to male NOS-2−/− mice after cerebral ischemia. Although male NOS-2−/− mice exhibit resistance to the gross effects of cerebral ischemia, they display neuronal loss and skilled motor deficits following cerebral ischemia. Administration of G-CSF during reperfusion reduced motor deficit and neuronal loss. Thus, G-CSF is still effective in NOS-2 gene-deficient mice, suggesting that part of the mechanism of action is independent of NOS-2.     

The cancer patient with severe mucositis

Oropharyngeal mucositis is a painful, often dose-limiting side effect of both radiotherapy and chemotherapy. To reduce the intensity of pain and prevent systemic infection via the compromised mucosa, agents such as antiseptic mouthwashes, anti-ulcer compounds, sodium bicarbonate, saline, and allopurinol have been traditionally used with limited success. The new agents that show promise are granulocyte macrophage colony-stimulating factor (GM-CSF) and granulocyte colony-stimulating factor (G-CSF). However, best results, dosage, and means of administration still have to be determined. Other agents such as sucralfate, tretinoin, glutamine, and misoprostol are also being tested. The results reported from different testing centers are often contradictory and confusing. Basic requirements in prevention and control of mucositis are good oral hygiene, mechanical débridement of the oral tissues, and hydration.