肝细胞生长因子及受体传递的调节与内吞作用

肝细胞生长因子(HGF)是一种具有多重功能的细胞调控因子。HGF与其受体Met酪氨酸激酶(c-Met)的结合可激发多种生物学反应,从而调节细胞的增殖、分化、形态发生和侵袭运动等。有多种因素参与了HGF/c-Met信号传导的调控,从而防止信号的过度放大,其中Cbl1、Rab、泛素化激酶和HGF/c-Met的内吞等发挥了重要的作用。因此,对HGF/c-Met内吞过程的研究,了解内吞对于HGF/c-Met的信号传导及其调控的影响,探讨HGF/c-Met信号传导通路的调控机理和相互作用模式,可进一步阐明HGF/c-Met信号传导的调控机制,从而验证肝细胞中内吞作用直接调节HGF/c-Met信号通路的作用机制。     

神经干细胞的分化影响其对肝细胞生长因子的趋化性迁移

神经干细胞（neural stem cells,NSCs）的定向迁移对神经系统发育和损伤后修复至关重要,但NSCs的定向迁移与NSCs的分化之间的关系鲜有研究。该研究以此为切入点,以肝细胞生长因子（hepatocyte growth factor,HGF）为趋化因子,神经干细胞系C17．2为研究对象,首先,建立了不同分化阶段的NSCs（分别分化0,12,24,72h）的分化模型;其次,运用Boyden chamber和Dunn chamber研究了不同分化状态下的NSCs对HGF的趋化性迁移。Boyden chamber结果显示：下室加入HGF后,分化12,24h的NSCs迁移至膜下方的细胞数目显著高于分化0,72h的NSCs;Dunn chamber结果显示：分化12,24h的NSCs迁移效率显著高于分化0,72h的NSCs。这些结果表明,NSCs的分化影响其对HGF的趋化性迁移,为在临床上更有效地利用NSCs治疗各种神经系统退行性疾病提供了理论依据。     

细胞生长因子与创伤愈合

创伤愈合是一个复杂的过程,传统将这一过程分为血凝期、炎症期、增值期、组织重构四个相互重叠的时期。多种细胞生长因子参与了这一过程,这些细胞因子包括EGF、bFGF、VEGF、PDGF等。本文就创伤愈合过程以及其涉及的细胞生长因子的生物学特性、创伤时的表达情况、对创面愈合的促进作用、应用前景作一概述。     

Expression of HGF/SF,HGFI/MSP,and c-met suggests new functions during early chick development

We report the cloning of full-length cDNAs for a plasminogen-related growth factor, hepatocyte growth factor/scatter factor (HGF/SF), its tyrosine kinase receptor, c-met, and a close member of the same family, hepatocyte growth factor-like/macrophage stimulating protein (HGFI/MSP), from the chick. We have used these cDNAs to provide the first report of the expression of this family of growth factors and the c-met receptor at early stages of vertebrate development. RNAase protection and wholemount in situ hyb ridization were used on chick embryos between formation of the primitive streak and early organogenesis. We find patterns of expression for HGF/SF and its receptor c-met consistent with their known roles in ep ithelial-mesenchymal transformation and angiogenesis. In addition, these genes and HGFI/MSP are expressed in discrete locations within developing somites, suggesting a role in paraxial mesodermal development. Very strong and early expression of HGF/SF in the elevating limb buds suggests its involvement in limb outgrowth. HGFI/MSP is expressed in the notochord and then in the prospective floor plate region and could play a role in development of the neural tube. Interestingly, c-met is often more closely as sociated with HGFI/MSP than with its known ligand, HGF/SF, raising the possibility that c-met expression may be induced by HGFI/MSP. © 1995 Wiley-Liss, Inc.     

In vitro Cell Migration and Invasion Assays

Migration is a key property of live cells and critical for normal development, immune response, and disease processes such as cancer metastasis and inflammation. Methods to examine cell migration are very useful and important for a wide range of biomedical research such as cancer biology, immunology, vascular biology, cell biology and developmental biology. Here we use tumor cell migration and invasion as an example and describe two related assays to illustrate the commonly used, easily accessible methods to measure these processes. The first method is the cell culture wound closure assay in which a scratch is generated on a confluent cell monolayer. The speed of wound closure and cell migration can be quantified by taking snapshot pictures with a regular inverted microscope at several time intervals. More detailed cell migratory behavior can be documented using the time-lapse microscopy system. The second method described in this paper is the transwell cell migration and invasion assay that measures the capacity of cell motility and invasiveness toward a chemo-attractant gradient. It is our goal to describe these methods in a highly accessible manner so that the procedures can be successfully performed in research laboratories even just with basic cell biology setup.     

血小板来源的生长因子通过ERK/p38/PI3K促进人视网膜色素上皮细胞的增殖、迁移

目的:探讨血小板来源的生长因子(PDGF)对体外培养的人视网膜色素上皮细胞(RPE)增殖和迁移的影响,并对参与其中的信号通路做初步研究.方法:体外培养的人视网膜色素上皮细胞与含有重组人血小板来源的生长因子的培养基(含有或不含2％(v/v)胎牛血清)共培养,用MTT法检测PDGF对RPE细胞增殖的影响,利用细胞爬片和免疫荧光技术检测PDGF对RPE细胞迁移等影响;另外分别向细胞培养物中添加PD98059,SB203580和PI3K等不同的信号通路分子抑制剂,判断参与PDGF激活的细胞活动相关的信号通路.结果:外源性PDGF能促进体外培养的人RPE的增殖和迁移.ERK1/2选择性抑制剂PD98059和PI3K抑制剂LY294002能显著的降低PDGF-BB诱导的人RPE细胞的增殖(P＜0.05),p38抑制剂SB203580没有明显的抑制作用.而对PDGF-BB诱导的RPE细胞的迁移,SB203580和LY294002有显著的抑制作用(P＜0.05),PD98059抑制作用不显著.结论:PDGF对RPE细胞的影响提示其在增生性玻璃体视网膜病变(PVR)的发展中有重要的作用,其可能为PVR提供一种新的毒副作用小的治疗手段.     

HGF/SF and menthol increase human glioblastoma cell calcium and migration

This study explored the role of transient receptor potential melastatin 8 ion channels (TRPM8) in mechanisms of human glioblastoma (DBTRG) cell migration. Menthol stimulated influx of Ca²⁺, membrane current, and migration of DBTRG cells. Effects on Ca²⁺ and migration were enhanced by pre-treatment with hepatocyte growth factor/scatter factor (HGF/SF). Effects on Ca²⁺ also were greater in migrating cells compared with non-migrating cells. 2-Aminoethoxydiphenyl borate (2-APB) inhibited all menthol stimulations. RT-PCR and immunoblot analysis showed that DBTRG cells expressed both mRNA and protein for TRPM8 ion channels. Two proteins were evident: one (130-140 kDa) in a plasma membrane-enriched fraction, and a variant (95-100 kDa) in microsome- and plasma membrane-enriched fractions. Thus, TRPM8 plays a role in mechanisms that increase [Ca²⁺]_i needed for DBTRG cell migration.     

Endothelial Injury Induces Vascular Smooth Muscle Cell Proliferation in Highly Localized Regions of a Direct Contact Co-culture System

Though previous studies have indicated a relationship between the proliferation of endothelial cells and vascular smooth muscle cells (VSMCs) in co-culture, the results have been contradictory and the signaling mechanism poorly understood. In this transmembrane co-culture study, VSMCs and endothelial cells were grown to confluence on opposite sides of a microporous membrane to mimic the intima/media border of vessels. The endothelial layer was injured, and then cultured for 3 days, resulting in partial re-endothelialization. VSMC proliferation across from the injured/partially recovered endothelial region was significantly higher than across from the de-endothelialized region (a sevenfold increase) and the uninjured region (a threefold increase). ELISA indicated that PDGF, which was undetectable in uninjured co-culture and homotypic controls, increased after injury and the addition of a piperazinyl-quinazoline carboxamide PDGF receptor inhibitor blocked VSMC proliferation across from the injured/partially recovered region. We conclude that co-culture signaling initiated by endothelial cell injury locally stimulates VSMC proliferation and that this signaling could be mediated by PDGF-BB.     

Priming of mononuclear cells with a combination of growth factors enhances wound healing via high angiogenic and engraftment capabilities

Recently, we demonstrated that a specific combination of growth factors enhances the survival, adhesion and angiogenic potential of mononuclear cells (MNCs). In this study, we sought to investigate the changes of the angiogenic potential of MNCs after short‐time priming with a specific combination of growth factors. MNCs were isolated using density gradient centrifugation and incubated with a priming cocktail containing epidermal growth factor (EGF), insulin‐like growth factor (IGF)‐1, fibroblast growth factor (FGF)‐2, FMS‐like tyrosine kinase (Flt)‐3L , Angiopoietin (Ang)‐1, granulocyte chemotactic protein (GCP)‐2 and thrombopoietin (TPO) (all 400 ng/ml) for 15, 30 and 60 min. Wounds in nonobese diabetic‐severe combined immune deficiency (NOD‐SCID) mice were created by skin excision followed by cell transplantation. We performed a qRT‐PCR analysis on the growth factor–primed cells. The angiogenic factors vascular endothelial growth factor (VEGF)‐A, FGF‐2, hepatocyte growth factor (HGF), platelet‐derived growth factor (PDGF) and interleukin (IL)‐8 and the anti‐apoptotic factors IGF‐1 and transforming growth factor‐β1 were significantly elevated in the MNCs primed for 30 min. (T30) compared with the non‐primed MNCs (T0). The scratch wound assay revealed that T30‐ conditioned media (CM) significantly increased the rate of fibroblast‐mediated wound closure compared with the rates from T0‐CM and human umbilical vein endothelial cells (HUVEC)‐CM at 20 hrs. In vivo wound healing results revealed that the T30‐treated wounds demonstrated accelerated wound healing at days 7 and 14 compared with those treated with T0. The histological analyses demonstrated that the number of engrafted cells and transdifferentiated keratinocytes in the wounds were significantly higher in the T30‐transplanted group than in the T0‐transplanted group. In conclusion, this study suggests that short‐term priming of MNCs with growth factors might be alternative therapeutic option for cell‐based therapies.     

Permeable FGF-1 Nuclear Localization Signal Peptide Stimulates DNA Synthesis in Various Cell Types but Is Cell-Density Sensitive and Unable to Support Cell Proliferation

An earlier report indicated that a 26-amino-acid peptide (SA), comprised of the nuclear localization signal (NLS) of fibroblast growth factor-1 (FGF-1) and a membrane-permeable peptide, was able to stimulate DNA synthesis after it was taken up by NIH3T3 fibroblasts. Here, we report that SA, but not a mutant with the NLS motif destroyed, induced DNA synthesis in BALB/c3T3 murine fibroblasts, human vascular endothelial (HUVE) cells, and primary cultured hepatocytes, although the activity was weaker than that of FGF-1. The kinetics of SA-induced DNA synthesis and G₁cyclin expression were similar to those elicited by FGF-1, indicating that SA induces cell cycle progression. Kinetic analysis also suggested that SA stimulates only a fraction of the DNA replication in BALB/c3T3 cells. At high cell densities, SA-induced G₁cyclin expression and DNA synthesis were more strongly inhibited than those induced by FGF-1. SA did not induce cell division in HUVE and BALB/c3T3 cells and did not interfere with FGF-1-stimulated proliferation of HUVE cells. These results indicate that SA is able to partially induce cell cycle progression through a contact-inhibition sensitive signaling pathway, but it is insufficient to support cell mitosis. We also suggest that signaling by SA does not interfere with that of FGF-1.     

Cross‐talk between the VEGF‐A and HGF signalling pathways in endothelial cells

Background information. Endothelial cells play a major role in angiogenesis, the process by which new blood vessels arise from a pre‐existing vascular bed. VEGF‐A (vascular endothelial growth factor‐A) is a key regulator of angiogenesis during both development and in adults. HGF (hepatocyte growth factor) is a pleiotropic cytokine that may promote VEGF‐A‐driven angiogenesis, although the signalling mechanisms underlying this co‐operation are not completely understood. Results. We analysed the effects of the combination of VEGF‐A and HGF on the activation of VEGFR‐2 (VEGF receptor‐2) and c‐met receptors, and on the stimulation of downstream signalling pathways in endothelial cells. We found that VEGFR‐2 and c‐met do not physically associate and do not transphosphorylate each other, suggesting that co‐operation involves signalling events more distal from receptor activation. We demonstrate that the VEGF isoform VEGF‐A₁₆₅ and HGF stimulate a similar set of MAPKs (mitogen‐activated protein kinases), although the kinetics and strengths of the activation differ depending on the growth factor and pathway. An enhanced activation of the signalling was observed when endothelial cells were stimulated by the combination of VEGF‐A₁₆₅ and HGF. Moreover, the combination of VEGF‐A and HGF results in a statistically significant synergistic activation of ERK1/2 (extracellular‐signal‐regulated kinase 1/2) and p38 kinases. We demonstrated that VEGF‐A₁₆₅ and HGF activate FAK (focal adhesion kinase) with different kinetics and stimulate the recruitment of phosphorylated FAK to different subsets of focal adhesions. VEGF‐A₁₆₅ and HGF regulate distinct morphogenic aspects of the cytoskeletal remodelling that are associated with the preferential activation of Rho or Rac respectively, and induce structurally distinct vascular‐like patterns in vitro in a Rho‐ or Rac‐dependent manner. Conclusions. Under angiogenic conditions, combining VEGF‐A with HGF can promote neovascularization by enhancing intracellular signalling and allowing more finely regulated control of the signalling molecules involved in the regulation of the cytoskeleton and cellular migration and morphogenesis.     

Age-related nitration/dysfunction of myogenic stem cell activator HGF

Mechanical perturbation triggers activation of resident myogenic stem cells to enter the cell cycle through a cascade of events including hepatocyte growth factor (HGF) release from its extracellular tethering and the subsequent presentation to signaling-receptor c-met. Here, we show that with aging, extracellular HGF undergoes tyrosine-residue (Y) nitration and loses c-met binding, thereby disturbing muscle homeostasis. Biochemical studies demonstrated that nitration/dysfunction is specific to HGF among other major growth factors and is characterized by its locations at Y198 and Y250 in c-met-binding domains. Direct-immunofluorescence microscopy of lower hind limb muscles from three age groups of rat, provided direct in vivo evidence for age-related increases in nitration of ECM-bound HGF, preferentially stained for anti-nitrated Y198 and Y250-HGF mAbs (raised in-house) in fast IIa and IIx myofibers. Overall, findings highlight inhibitory impacts of HGF nitration on myogenic stem cell dynamics, pioneering a cogent discussion for better understanding age-related muscle atrophy and impaired regeneration with fibrosis (including sarcopenia and frailty).     

Monoclonal antibody 425 inhibits growth stimulation of carcinoma cells by exogenous EGF and tumor-derived EGF/TGF-alpha

Carcinoma cells frequently coexpress transforming growth factor (TGF)-alpha and its receptor, the epidermal growth factor (EGF) receptor, implicating an autocrine function of carcinoma-derived TGF-alpha. Using a monoclonal antibody (425) to the EGF-receptor, we investigated the role of exogenous and tumor cell-derived EGF/TGF-alpha mitogenic activities in proliferation of cell lines derived from solid tumors. Monoclonal antibody 425 was chosen for these studies because it inhibits binding of EGF/TGF-alpha to the EGF-receptor and effectively blocks activation of the EGF-receptor by EGF/TGF-alpha. Seven malignant cell lines originating from carcinomas of colon, pancreas, breast, squamous epithelia, and bladder expressed surface EGF-receptor and secreted EGF/TGF-alpha-like mitogenic activities into their tissue culture media. All cell lines were maintained in a defined medium free of exogenous EGF/TGF-alpha. EGF and TGF-alpha added to the culture medium stimulated proliferation of five cell lines to comparable levels. EGF/TGF-alpha-dependent proliferation was significantly reduced by addition of MAb 425 to culture media. In addition, monoclonal antibody 425 reduced proliferation of the five EGF/TGF-alpha responsive cell lines in the absence of exogenous EGF/TGF-alpha. Antiproliferative effects induced by monoclonal antibody 425 were reversible and could be overcome by addition of EGF to culture media. Our results indicate that tumor-derived EGF-receptor-reactive mitogens can promote proliferation of carcinoma cells in an autocrine fashion.     

Structural basis of MET receptor dimerization by the bacterial invasion protein InlB and the HGF/SF splice variant NK1

The structural basis of ligand-induced dimerization of the receptor tyrosine kinase MET by its natural ligand hepatocyte growth factor/scatter factor (HGF/SF) is not well understood. However, interesting insight into the molecular mechanism of MET dimerization has emerged from crystal structures of MET in complex with a bacterial agonist, the invasion protein internalin B (InlB) from pathogenic Listeria monocytogenes. MET activation by InlB promotes uptake of bacteria into host cells. Structural and biophysical data suggest that InlB is monomeric on its own but dimerizes upon binding to the membrane-anchored MET receptor promoting the formation of a signaling active 2:2 complex. The dimerization interface is small and unusually located on the convex side of the curved InlB leucine-rich repeat (LRR) domain. As InlB does not dimerize in solution, the dimerization site could only be identified by studying packing contacts of InlB in various crystal forms and had to be proven by scrutinizing its biological relevance in cellular assays. InlB dimerization is thus an example of a low-affinity contact that appears irrelevant in solution but becomes physiologically significant in the context of 2-dimensional diffusion restricted to the membrane plane. The resulting 2:2 InlB:MET complex has an InlB dimer at its center with one MET molecule bound peripherally to each InlB. This model of ligand-mediated MET dimerization may serve as a blue-print to understand MET activation by NK1, a naturally occurring HGF/SF splice variant and MET agonist. Crystal structures of NK1 repeatedly show a NK1 dimer, in which residues implicated in MET-binding are located on the outside. Thus, MET dimerization by NK1 may also be ligand-mediated with a NK1 dimer at the center of the 2:2 complex with one MET molecule bound peripherally to each NK1. This article is part of a Special Issue entitled: Emerging recognition and activation mechanisms of receptor tyrosine kinases.     

Regulation of Adhesion and Migration in the Germinal Center Microenvironment

T cell dependent humoral immune responses are initiated by the activation of naive B cells in the T cell areas of the secondary lymphoid tissues. This primary B cell activation leads to migration of germinal center (GC) cell precursors into B cell follicles where they engage follicular dendritic cells (FDC) and T cells, and differentiate into memory B cells or plasma cells. Both B cell homing to the GC and interaction with FDC critically depend on integrin-mediated adhesion. We have recently indentified the c-met-encoded receptor tyrosine kinase and its ligand, the growth and motility factor hepatocyte growth factor/scatter factor (HGF/SF), as a novel paracrine signalling pathway regulating B cell adhesion (van der Voort et al., 1997, J. Exp. Med. 185, 2121–2131). The c-Met protein is expressed on B cells localized in the dark zone of the GC (centroblasts) and is induced by CD40 plus BCR ligation. Stimulation of c-Met with HGF/SF. which is produced at high levels by tonsillar stromal cells and FDC, leads to receptor phosphorylation and to enhanced integrin-mediated adhesion of B cells to both VCAM-l and fibronectin. Interestingly, these responses to HGF/SF are promoted by heparan-sulfate proteoglycan forms of CD44 (CD44-HS). Like c-Met, CD44-HS is induced on B cells by CD40 ligation. It efficiently binds HGF/SF and strongly promotes signalling through c-Met. We conclude that integrin regulation during antigen specific B cell differentiation involves cross-talk between the HGF/SF-c-Met pathway and CD44-HS.     

Delphinidin Reduces Cell Proliferation and Induces Apoptosis of Non-Small-Cell Lung Cancer Cells by Targeting EGFR/VEGFR2 Signaling Pathways

Epidermal growth factor receptor (EGFR) and vascular endothelial growth factor receptor 2 (VEGFR2) have emerged as two effective clinical targets for non-small-cell lung cancer (NSCLC). In the present study, we found that delphinidin, an anthocyanidin, present in pigmented fruits and vegetables, is a potent inhibitor of both EGFR and VEGFR2 in NSCLC cells that overexpress EGFR/VEGFR2. Using these cells, we next determined the effects of delphinidin on cell growth and apoptosis in vitro and on tumor growth and angiogenesis in vivo. Delphinidin (5-60 µM) treatment of NSCLC cells inhibited the activation of PI3K, and phosphorylation of AKT and MAPKs. Additionally, treatment of NSCLC cells with delphinidin resulted in inhibition of cell growth without having significant toxic effects on normal human bronchial epithelial cells. Specifically, treatment of NCI-H441 and SK-MES-1 cells with delphindin (5-60 µM) resulted in (i) cleavage of PARP protein, (ii) activation of caspase-3 and -9, (iii) downregulation of anti-apoptotic proteins (Bcl2, Bcl-xL and Mcl-1), (iv) upregulation of pro-apoptotic proteins (Bax and Bak), and (v) decreased expression of PCNA and cyclin D1. Furthermore, in athymic nude mice subcutaneously implanted with human NSCLC cells, delphinidin treatment caused a (i) significant inhibition of tumor growth, (ii) decrease in the expression of markers for cell proliferation (Ki67 and PCNA) and angiogenesis (CD31 and VEGF), and (iii) induction of apoptosis, when compared with control mice. Based on these observations, we suggest that delphinidin, alone or as an adjuvant to current therapies, could be used for the management of NSCLC, especially those that overexpress EGFR and VEGFR2.     

Proliferation and tube formation of periodontal endothelial cells

Angiogenesis is indispensable to guide a regeneration of good periodontal tissue in the wound healing after periodontal surgery. Hepatocyte growth factor is well known for a strong angiogenic factor and it may play important roles in the periodontal tissue during periodontal wound healing. In exploring the promotion of angiogenesis in the periodontal ligament, proliferative and tubulogenic responses of endothelial cells to hepatocyte growth factor and to soluble factors secreted by fibroblasts were investigated. Pavement-shaped cells isolated from a human periodontal ligament were identified as the endothelial cell by their granular immunoreactivity for factor VIII. The proliferation of the endothelial cells was accelerated by the addition of hepatocyte growth factor or fibroblast-conditioned medium, and far more by adding both than either. The endothelial cells seeded on the agar containing both hepatocyte growth factor and fibroblast products formed a dense network in a shorter time than on the agar containing either. The endothelial cells in the dense network took a tube-like structure with lumen and were covered with laminin. These results suggest that hepatocyte growth factor administered into the regenerating periodontal tissue may promote, synergistically with local factors produced by the activated fibroblast, the proliferation and tubulogenesis of the remaining endothelial cells.     

Effect of hepatocyte growth factor/scatter factor and other growth factors on motility and morphology of non-tumorigenic and tumor cells

Summary Using an automated cell analyzer system, the effect of hepatocyte growth factor/scatter factor (HGF/SF), epidermal growth factor (EGF), basic fibroblast growth factor (bFGF), endothelial acidic fibroblast growth factor (a-FGF), platelet derived growth factor (PDGF), and recombinant human insulinlike growth factor (IGF) on the motility and morphology of Madin-Darby canine kidney (MDCK), rat hepatomas, C₂, and H_5–6 and murine mammary carcinoma (EMT-6) cells was investigated. Treatment of MDCK cells with HGF/SF, bFGF, EGF, and a-FGF resulted in an increase in average cell velocity and in the fraction of moving cells. Cells treated with the PDGF and IGF did not show significant alterations in velocity. MDCK cells treated with each growth factor were classified into groups of “fast” and “slow” moving cells based on their average velocities, and the average morphologic features of the two groups were quantitated. Fast-moving cells had larger average area, circularity, and flatness as compared to slow-moving cells. Factors that stimulated cell movement also induced alterations in cell morphologic parameters including spreading, flatness, area, and circularity. HGF/SF also scattered and stimulated motility of C₂ and H_5–6 hepatoma cells. In contrast to MDCK cells, there was no significant difference between the morphology of the fast moving and slow moving C₂ and H_5–6 cells. These studies suggest that growth factor cytokines have specific effects on motility of normal and tumor cells.     

Stimulation of human vascular endothelial cell growth by a platelet-derived growth factor and thrombin

Repair of a vascular wound is mediated by migration and subsequent replication of the endothelial cells that form the inner lining of blood vessels. We have measured the growth response of human umbilical vein endothelial cells (HuE) to two polypeptides that are transiently produced in high concentrations at the site of a wound; the platelet-derived growth factor (PDGF) and the protease thrombin. When 10⁴ HuE cells are seeded as a dense island (2-mm diameter) in the center of a 16-mm tissue culture well in medium containing 20% human serum derived from platelet-poor plasma (PDS), no increase in cell number or colony size is observed. With the addition of 0.5 ng/ml partially purified PDGF, colony size increases and the number of cells after 8 days is 4.8 × 10⁴. When human thrombin (1 μg/ml) is added along with the PDGF, the cell number rises to 9.2 × 10⁴. Thrombin alone stimulates no increase in cell number. Although partially purified PDGF stimulates endothelial cells maintained in PDS as well as those maintained in whole blood serum (WBS), pure PDGF is active only when assayed in medium that contains WBS and is supplemented with thrombin. These results suggest the existence of a second class of platelet-derived factors that enable HuE cells to respond to the mitogenic activity of the purified platelet mitogen and thrombin.     

秦皮素通过抑制EGFR及其下游AKT信号通路抑制MCF-7细胞增殖及迁移

表皮生长因子受体(EGFR)是细胞内多种信号调节通路的交汇点,其介导的信号通路与乳腺癌的发生、发展、转移和侵袭等密切相关,已成为乳腺癌治疗的新靶点之一。但目前关于秦皮素的抗乳腺癌作用与EGFR通路的关系,国内外尚未见相关报道。本研究结果表明,秦皮素能够通过抑制EGFR及其下游的AKT信号通路来发挥其抗乳腺癌作用。秦皮素在体外可促进T、B 淋巴细胞增殖及巨噬细胞吞噬能力,提示秦皮素可能促进小鼠免疫功能。Western印迹结果表明,秦皮素能显著抑制EGFR蛋白及其下游的AKT蛋白磷酸化水平。划痕实验结果表明,秦皮素能抑制MCF-7细胞的迁移。此外,秦皮素还能促进小鼠巨噬细胞的吞噬能力和代谢活力,促进T、B淋巴细胞的增殖,提高NK细胞的杀伤活力。本研究结果提示,秦皮素的抗乳腺癌作用是通过抑制EGFR信号通路,抑制MCF-7细胞迁移和促进小鼠的免疫功能等多种途径来实现的。