Prostate Sphere-forming Stem Cells Are Derived from the P63-expressing Basal Compartment

Prostate stem cells (P-SCs) are capable of giving rise to all three lineages of prostate epithelial cells, including basal, luminal, and neuroendocrine cells. Multiple methods have been used to identify P-SCs in adult prostates. These include in vivo renal capsule implantation of a single epithelial cell with urogenital mesenchymal cells, in vitro prostasphere and organoid cultures, and lineage tracing with castration-resistant Nkx3.1 expression (CARN), in conjunction with expression of cell type-specific markers. Both organoid culture and CARN tracing show the existence of P-SCs in the luminal compartment. Although prostasphere cells predominantly express basal cell-specific cytokeratin and P63, the lineage of prostasphere-forming cells in the P-SC hierarchy remains to be determined. Using lineage tracing with P63^CreERT2, we show here that the sphere-forming P-SCs are P63-expressing cells and reside in the basal compartment. Therefore we designate them as basal P-SCs (P-bSCs). P-bSCs are capable of differentiating into AR⁺ and CK18⁺ organoid cells, but organoid cells cannot form spheres. We also report that prostaspheres contain quiescent stem cells. Therefore, the results show that P-bSCs represent stem cells that are early in the hierarchy of overall prostate tissue stem cells. Understanding the contribution of the two types of P-SCs to prostate development and prostate cancer stem cells and how to manipulate them may open new avenues for control of prostate cancer progression and relapse.     

Fibroblast Growth Factor Signaling Is Essential for Self-renewal of Dental Epithelial Stem Cells

A constant supply of epithelial cells from dental epithelial stem cell (DESC) niches in the cervical loop (CL) enables mouse incisors to grow continuously throughout life. Elucidation of the cellular and molecular mechanisms underlying this unlimited growth potential is of broad interest for tooth regenerative therapies. Fibroblast growth factor (FGF) signaling is essential for the development of mouse incisors and for maintenance of the CL during prenatal development. However, how FGF signaling in DESCs controls the self-renewal and differentiation of the cells is not well understood. Herein, we report that FGF signaling is essential for self-renewal and the prevention of cell differentiation of DESCs in the CL as well as in DESC spheres. Inhibiting the FGF signaling pathway decreased proliferation and increased apoptosis of the cells in DESC spheres. Suppressing FGFR or its downstream signal transduction pathways diminished Lgr5-expressing cells in the CL and promoted cell differentiation both in DESC spheres and the CL. Furthermore, disruption of the FGF pathway abrogated Wnt signaling to promote Lgr5 expression in DESCs both in vitro and in vivo. This study sheds new light on understanding the mechanism by which the homeostasis, expansion, and differentiation of DESCs are regulated.     

Deletion of the PH-domain and Leucine-rich Repeat Protein Phosphatase 1 (Phlpp1) Increases Fibroblast Growth Factor (Fgf) 18 Expression and Promotes Chondrocyte Proliferation

Endochondral ossification orchestrates formation of the vertebrate skeleton and is often induced during disease and repair processes of the musculoskeletal system. Here we show that the protein phosphatase Phlpp1 regulates endochondral ossification. Phlpp1 null mice exhibit decreased bone mass and notable changes in the growth plate, including increased BrdU incorporation and matrix production. Phosphorylation of known Phlpp1 substrates, Akt2, PKC, and p70 S6 kinase, were enhanced in ex vivo cultured Phlpp1^−/− chondrocytes. Furthermore, Phlpp1 deficiency diminished FoxO1 levels leading to increased expression of Fgf18, Mek/Erk activity, and chondrocyte metabolic activity. Phlpp inhibitors also increased matrix content, Fgf18 production and Erk1/2 phosphorylation. Chemical inhibition of Fgfr-signaling abrogated elevated Erk1/2 phosphorylation and metabolic activity in Phlpp1-null cultures. These results demonstrate that Phlpp1 controls chondrogenesis via multiple mechanisms and that Phlpp1 inhibition could be a strategy to promote cartilage regeneration and repair.     

Extended Synaptotagmin Interaction with the Fibroblast Growth Factor Receptor Depends on Receptor Conformation,Not Catalytic Activity

We previously demonstrated that ESyt2 interacts specifically with the activated FGF receptor and is required for a rapid phase of receptor internalization and for functional signaling via the ERK pathway in early Xenopus embryos. ESyt2 is one of the three-member family of Extended Synaptotagmins that were recently shown to be implicated in the formation of endoplasmic reticulum (ER)-plasma membrane (PM) junctions and in the Ca²⁺ dependent regulation of these junctions. Here we show that ESyt2 is directed to the ER by its putative transmembrane domain, that the ESyts hetero- and homodimerize, and that ESyt2 homodimerization in vivo requires a TM adjacent sequence but not the SMP domain. ESyt2 and ESyt3, but not ESyt1, selectively interact in vivo with activated FGFR1. In the case of ESyt2, this interaction requires a short TM adjacent sequence and is independent of receptor autophosphorylation, but dependent on receptor conformation. The data show that ESyt2 recognizes a site in the upper kinase lobe of FGFR1 that is revealed by displacement of the kinase domain activation loop during receptor activation.     

成纤维细胞生长因子与其受体相互作用及其抑制剂的研究进展

成纤维细胞生长因子（FGF）有许多重要的生理功能，并与肿瘤的形成有关.为了弄清FGF与成纤维细胞生长因子受体(FGFR)相互作用的机制，人们对FGF和FGFR的各个结合结构域进行了深入、细致的研究，定位了aFGF、bFGF的肝素结合区、bFGF的受体结合区、FGF受体的肝素结合区、配体结合区和FGF受体相互结合区，提出了两个FGF与FGFR相互作用的模型，在此基础上设计了FGF的核酸类、糖类和多肽类抑制剂，为寻找新一代抗癌药物打下了理论基础.     

A Hippo and Fibroblast Growth Factor Receptor Autocrine Pathway in Cholangiocarcinoma

Herein, we have identified cross-talk between the Hippo and fibroblast growth factor receptor (FGFR) oncogenic signaling pathways in cholangiocarcinoma (CCA). Yes-associated protein (YAP) nuclear localization and up-regulation of canonical target genes was observed in CCA cell lines and a patient-derived xenograft (PDX). Expression of FGFR1, -2, and -4 was identified in human CCA cell lines, driven, in part, by YAP coactivation of TBX5. In turn, FGFR signaling in a cell line with minimal basal YAP expression induced its cellular protein expression and nuclear localization. Treatment of YAP-positive CCA cell lines with BGJ398, a pan-FGFR inhibitor, resulted in a decrease in YAP activation. FGFR activation of YAP appears to be driven largely by FGF5 activation of FGFR2, as siRNA silencing of this ligand or receptor, respectively, inhibited YAP nuclear localization. BGJ398 treatment of YAP-expressing cells induced cell death due to Mcl-1 depletion. In a YAP-associated mouse model of CCA, expression of FGFR 1, 2, and 4 was also significantly increased. Accordingly, BGJ398 treatment was tumor-suppressive in this model and in a YAP-positive PDX model. These preclinical data suggest not only that the YAP and Hippo signaling pathways culminate in an Mcl-1-regulated tumor survival pathway but also that nuclear YAP expression may be a biomarker to employ in FGFR-directed therapy.     

RasGAP Shields Akt from Deactivating Phosphatases in Fibroblast Growth Factor Signaling but Loses This Ability Once Cleaved by Caspase-3

Fibroblast growth factor receptors (FGFRs) are involved in proliferative and differentiation physiological responses. Deregulation of FGFR-mediated signaling involving the Ras/PI3K/Akt and the Ras/Raf/ERK MAPK pathways is causally involved in the development of several cancers. The caspase-3/p120 RasGAP module is a stress sensor switch. Under mild stress conditions, RasGAP is cleaved by caspase-3 at position 455. The resulting N-terminal fragment, called fragment N, stimulates anti-death signaling. When caspase-3 activity further increases, fragment N is cleaved at position 157. This generates a fragment, called N2, that no longer protects cells. Here, we investigated in Xenopus oocytes the impact of RasGAP and its fragments on FGF1-mediated signaling during G₂/M cell cycle transition. RasGAP used its N-terminal Src homology 2 domain to bind FGFR once stimulated by FGF1, and this was necessary for the recruitment of Akt to the FGFR complex. Fragment N, which did not associate with the FGFR complex, favored FGF1-induced ERK stimulation, leading to accelerated G₂/M transition. In contrast, fragment N2 bound the FGFR, and this inhibited mTORC2-dependent Akt Ser-473 phosphorylation and ERK2 phosphorylation but not phosphorylation of Akt on Thr-308. This also blocked cell cycle progression. Inhibition of Akt Ser-473 phosphorylation and entry into G₂/M was relieved by PHLPP phosphatase inhibition. Hence, full-length RasGAP favors Akt activity by shielding it from deactivating phosphatases. This shielding was abrogated by fragment N2. These results highlight the role played by RasGAP in FGFR signaling and how graded stress intensities, by generating different RasGAP fragments, can positively or negatively impact this signaling.     

Structural Analysis of the Human Fibroblast Growth Factor Receptor 4 Kinase

The family of fibroblast growth factor receptors (FGFRs) plays an important and well-characterized role in a variety of pathological disorders. FGFR4 is involved in myogenesis and muscle regeneration. Mutations affecting the kinase domain of FGFR4 may cause cancer, for example, breast cancer or rhabdomyosarcoma. Whereas FGFR1–FGFR3 have been structurally characterized, the structure of the FGFR4 kinase domain has not yet been reported. In this study, we present four structures of the kinase domain of FGFR4, in its apo-form and in complex with different types of small-molecule inhibitors. The two apo-FGFR4 kinase domain structures show an activation segment similar in conformation to an autoinhibitory segment observed in the hepatocyte growth factor receptor kinase but different from the known structures of other FGFR kinases. The structures of FGFR4 in complex with the type I inhibitor Dovitinib and the type II inhibitor Ponatinib reveal the molecular interactions with different types of kinase inhibitors and may assist in the design and development of FGFR4 inhibitors.     

成纤维细胞生长因子的特性与分子功能

成纤维细胞生长因子(FGFs)是一类蛋白质活性肽,在有机体内外显示广谱的生物学活性和功能,在生物发育学、生理学与临床药理学方面具有重要的作用和意义．对 FGFs 家族成员肽的特征特性,FGFs 的染色体定位、基因结构与功能,FGFs 的信号分子作用与促分裂素作用,FGFs 的作用受体、主动和被动调节与拮抗机理进行了综述．     

生长分化因子15及其特异性受体GFRAL信号通路

生长分化因子 15(growth differentiation factor 15,GDF15)属于转化生长因子 β(transforming growth factor β,TGF-β)超家族的成员之一,是与转化生长因子β家族成员同源性很低的新一类二聚体多肽.GDF15最初发现于活化的巨噬细胞中,可通过2种不同的细...     

Unique Intracellular Trafficking Processes Associated With Neural Cell Adhesion Molecule and Its Intracellular Signaling

Abstract

Homophilic binding of the neural cell adhesion molecule (NCAM) results in intracellular signaling, which also involves heterophilic engagement of coreceptors such as the fibroblast growth factor receptor (FGFR) and receptor protein tyrosine phosphatase-α (RPTPα). NCAM's own cellular dynamic itinerary includes endocytosis and recycling to the plasma membrane. Recent works suggest that NCAM could influence the trafficking of other receptor molecules that it associates with, particularly the FGFR. Furthermore, it was demonstrated that NCAM could undergo proteolytic processing upon activation. A processed fragment of NCAM, together with an N-terminal fragment of focal adhesion kinase (FAK), is translocated into the nucleus. Here, the authors discuss these rather unique (though not without precedence and analogues) receptor trafficking activities that are associated with NCAM and NCAM signaling.     

Type 1 Fibroblast Growth Factor Receptor in Cranial Neural Crest Cell-derived Mesenchyme Is Required for Palatogenesis

Cleft palate is a common congenital birth defect. The fibroblast growth factor (FGF) family has been shown to be important for palatogenesis, which elicits the regulatory functions by activating the FGF receptor tyrosine kinase. Mutations in Fgf or Fgfr are associated with cleft palate. To date, most mechanistic studies on FGF signaling in palate development have focused on FGFR2 in the epithelium. Although Fgfr1 is expressed in the cranial neural crest (CNC)-derived palate mesenchyme and Fgfr1 mutations are associated with palate defects, how FGFR1 in palate mesenchyme regulates palatogenesis is not well understood. Here, we reported that by using Wnt1^Cre to delete Fgfr1 in neural crest cells led to cleft palate, cleft lip, and other severe craniofacial defects. Detailed analyses revealed that loss-of-function mutations in Fgfr1 did not abrogate patterning of CNC cells in palate shelves. However, it upset cell signaling in the frontofacial areas, delayed cell proliferation in both epithelial and mesenchymal compartments, prevented palate shelf elevation, and compromised palate shelf fusion. This is the first report revealing how FGF signaling in CNC cells regulates palatogenesis.     

Localization of Basic Fibroblast Growth Factor in Bovine Adrenal Chromaffin Cells

Abstract: We have investigated basic fibroblast growth factor (FGF-2) localization in and release from isolated bovine adrenal chromaffin cells. In contrast to previous reports, we found no evidence of fibroblast growth factor (FGF) storage in catecholamine-containing chromaffin granules. Subcellular fractionation studies did not show enrichment of FGF-2 immunoreactivity in granules, and cholinergic stimulation failed to release FGF-2 into the medium. Our results suggest that adrenal chromaffin cells resemble other FGF-2-synthesizing cell types with respect to FGF storage and secretion.     

表皮生长因子对精原干细胞的作用机理

为了探究表皮生长因子(EGF)对体外培养的精原干细胞增殖的调控作用及其作用机制.应用不连续Percoll梯度液和选择性贴壁法分离纯化精原干细胞,c-kit细胞免疫组化鉴定细胞,MTT法研究EGF对精原干细胞增殖的效应,再加入JAK-STAT信号通路特异性抑制剂AG490,探究EGF对精原干细胞增殖作用的可能机制.c-kit细胞免疫组化结果显示分离得到细胞为精原干细胞;MTT结果显示各实验组比对照组细胞数量均有显著增多(P0.01);与对照组相比,加入AG490组的活细胞数有显著下降(P0.01).实验结果表明EGF能够促进精原干细胞的增殖,并且可以通过JAK-STAT信号通路起作用.     

Fibroblast Growth Factor Inhibits Expression of Neural Markers in Cultures of Zebrafish Early Embryo Cells

Basic fibroblast growth factor (FGF) regulation of developmental markers in cell cultures derived from early zebrafish embryos was examined with the goal of in vitro culture of zebrafish embryonic stem cells and gaining an understanding of extracellular influences on early embryonic development. Markers were stem/primordial germ cell markers pou-2 and vas, neural markers zp-50, pax[zf-a], en-3, and wnt-1, and mesodermal markers gsc and myoD. Previously we had shown that FGF prevents the development of zebrafish pigment cells in vitro. In our culture system, FGF reduced expression of neural-specific markers, possibly implicating the FGF family in suppression of early neural cell development. Exposure to FGF for 24 hours at the time of seeding the cells was sufficient to suppress neural marker expression for a subsequent 4 days of culture, while absence of FGF for the first 24 hours of culture nullified the effect of FGF added subsequently. FGF predictably increased expression of gsc and myoD. Vas expression was unaffected, while pou-2 expression decreased with time in culture in the presence or absence of FGF. However, in situ hybridization identified a subpopulation of cells expressing pou-2, suggesting the possible continued existence of undifferentiated stem cells in the cultures. Received March 10, 2000; accepted May 10, 2000.     

间充质干细胞向成骨细胞分化的信号通路

间充质干细胞具有向成骨细胞分化的潜能,可体外分离、培养和扩增,是骨组织工程中理想的种子细胞。近年的研究表明间充质干细胞的成骨分化受到多种信号通路的调控,现就其中研究较为深入的MAPK和Notch通路的情况作一简要综述。     

Basic Fibroblast Growth Factor Prevents the Memory Impairment Induced by Gastrin-Releasing Peptide Receptor Antagonism in Area CA1 of the Rat Hippocampus

Increasing evidence indicates that the gastrin-releasing peptide receptor (GRPR) is implicated in regulating synaptic plasticity and memory formation in the hippocampus and other brain areas. However, the molecular mechanisms underlying the memory-impairing effects of GRPR antagonism have remained unclear. Here we report that basic fibroblast growth factor (bFGF/FGF-2) rescues the memory impairment induced by GRPR antagonism in the rat dorsal hippocampus. The GRPR antagonist [D-Tpi⁶, Leu¹³ psi(CH₂NH)-Leu¹⁴] bombesin (6–14) (RC-3095) at 1.0 μg impaired, whereas bFGF at 0.25 μg enhanced, 24 h retention of inhibitory avoidance (IA) when infused immediately after training into the CA1 hippocampal area in male rats. Coinfusion with an otherwise ineffective dose of bFGF blocked the memory-impairing effect of RC-3095. These findings suggest that the memory-impairing effects of GRPR antagonists might be partially mediated by an inhibition in the function and/or expression of neuronal bFGF or diminished activation of intracellular protein kinase pathways associated with bFGF signaling.     

成纤维细胞生长因子9在抑郁症及其运动干预调节中的作用

成纤维细胞生长因子9（fibroblast growth factor 9, FGF9）是成纤维细胞生长因子（fibroblast growth factor,FGF）家族成员之一,属于一种自分泌或旁分泌生长因子。在脑组织中,FGF9主要表达于海马和皮质区,具有促进细胞增殖和维持细胞存活的功能。研究发现,FGF家族在抑郁症患者的多个脑区出现表达紊乱,FGF9在抑郁行为中扮演着负调控角色,但其介导抑郁行为的分子机制尚不清楚。本文综述了FGF9及其家族成员在抑郁中的作用; 围绕其受体（FGFR）信号在中枢神经系统中的功能特点,深入分析FGF9调节抑郁行为中的作用机制;结合运动抗抑郁的神经营养假说,提出经由FGFR/GSK3β/β-catenin通路的FGF信号,可能介导抑郁症的运动干预机制的假设。这些将为FGF9介导抑郁行为和运动抗抑郁的有关研究提供理论的基础和探索的思路。     

组织特异性启动子介导的反义FGF8b RNA对前列腺癌细胞生长增殖能力的影响

利用启动子的组织特异性和治疗基因组织特异表达的特点 ,设计出前列腺癌靶向基因治疗的新方案 .利用DNA重组技术将前列腺组织特异性启动子 (probasin基因启动子 )和在前列腺癌细胞中高表达成纤维细胞生长因子 (FGF) 8b反义cDNA克隆到逆转录病毒载体pSIR中构建成重组体PB 反义FGF8b pSIR .经转染包装细胞PT 6 7后将产生的复制缺陷型逆转录病毒体外感染前列腺癌细胞系PC 3M ,体外检测其生长增殖和侵袭转移能力的变化 .结果表明 ,与对照组相比 ,前列腺癌细胞感染产生反义FGF8bRNA的逆转录病毒后生长速度减慢 ,集落形成能力下降 ,体外侵袭转移能力降低 (P <0 0 1) .体外试验表明 ,前列腺组织特异性启动子介导的反义FGF8bRNA可有效降低前列腺癌细胞的体外生长增殖和转移能力 ,这为体内靶向前列腺癌基因治疗奠定了可靠的基础 .     

VEGF通过调节黏着斑促进间充质干细胞的黏附及铺展

间充质干细胞（mesenchymalstemcells,MSCs）具有多向分化潜能并能在体外趋化剂或细胞因子的作用下进行定向迁移,体内移植后可趋向迁移至脑瘤病灶区。细胞黏附是细胞迁移的首要条件,了解细胞黏附及其调控有助于细胞迁移机制的研究。细胞黏附及铺展涉及到黏着斑（f0－caladhesions,FAs）的动态变化以及细胞骨架的重排。细胞铺展面积在黏附过程中逐渐增大,黏附初期形成的小的黏着复合物逐渐成熟,聚集在一起形成较大的FAs。肌动蛋白（F—actin）聚集形成的螺线圈样微丝结构逐渐被应力纤维代替,细胞也由圆形变为具有极性的梭形或多角形。黏着斑激酶（focal adhesion kinase,FAK）和桩蛋白（paxillin）具有调节FAs聚合及骨架重排的作用,其中,Y397－FAK和Y31／Y118－paxillin的磷酸化活性在细胞铺展过程中不断变化。FAs组装时,Y397－FAK的磷酸化活性升高;FAs成熟后,Y397．FAK的磷酸化活性下降。活化的FAK能够磷酸4LY31／Y118-paxillin,激活paxillin参与调节细胞骨架的形成和排列。血管内皮生长因子（vascular endothelial growthfactor,VEGF）诱导~SMSCs黏附过程中,细胞面积变大,完全铺展的时间缩短,黏着斑及细胞骨架的形成均提前。另外,VEGF诱导的细胞铺展过程中形成的FAs形态细长,数量较多。该研究表明,VEGF通过调节黏着斑和细胞骨架促L~MSCs的黏附与铺展,提示vEGF可以通过调节黏着斑进而调控MSCs的定向迁移,为细胞迁移行为的研究提供理论基础。