The presumed atypical chemokine receptor CXCR7 signals through G(i/o) proteins in primary rodent astrocytes and human glioma cells

SDF-1/CXCL12 binds to the chemokine receptors, CXCR4 and CXCR7, and controls cell proliferation and migration during development, tumorigenesis, and inflammatory processes. It is currently assumed that CXCR7 would represent an atypical or scavenger chemokine receptor which modulates the function of CXCR4. Contrasting this view, we demonstrated recently that CXCR7 actively mediates SDF-1 signaling in primary astrocytes. Here, we provide evidence that CXCR7 affects astrocytic cell signaling and function through pertussis toxin-sensitive G(i/o) proteins. SDF-1-dependent activation of G(i/o) proteins and subsequent increases in intracellular Ca(2+) concentration persisted in primary rodent astrocytes with depleted expression of CXCR4, but were abolished in astrocytes with depleted expression of CXCR7. Moreover, CXCR7-mediated effects of SDF-1 on Erk and Akt signaling as well as on astrocytic proliferation and migration were all sensitive to pertussis toxin. Likewise, pertussis toxin abolished SDF-1-induced activation of Erk and Akt in CXCR7-only expressing human glioma cell lines. Finally, consistent with a ligand-biased function of CXCR7 in astrocytes, the alternate CXCR7 ligand, I-TAC/CXCL11, activated Erk and Akt through β-arrestin. The demonstration that SDF-1-bound CXCR7 activates G(i/o) proteins in astrocytes could help to explain some discrepancies previously observed for the function of CXCR4 and CXCR7 in other cell types.     

Enhanced expression of the CXCl12/SDF-1 chemokine receptor CXCR7 after cerebral ischemia in the rat brain

Expression patterns of the second SDF-1 receptor RDC1/CXCR7 were examined after focal ischemia in rats using in situ hybridization. CXCR7 mRNA was identified in the ventricle walls as well as neuronal, astroglial, and vascular cells. After ischemia, intact cortical regions showed a rapid, 4 days-lasting increase in neuronal CXCR7 expression. In the ischemic tissue CXCR7 expression was scarce and associated with blood vessels. Between days 2 and 10 after ischemia-onset, SDF-1 expression increased strongly in the peri-infarct and infarct region, which was accompanied by the appearance of numerous CXCR4-expressing but not CXCR7-expressing cells. These patterns suggest that SDF-1 may influence vascular, astroglial, and neuronal functions via CXCR7 and mediate cell recruitment to ischemic brain areas via CXCR4.     

The relationship between SDF-1α/CXCR4 and neural stem cells appearing in damaged area after traumatic brain injury in rats

Abstract

Objective: The actual relationship between neural stem cells and SDF-1α/CXCR4 after brain injury has not yet been elucidated, although recent studies have speculated that stromal cell-derived factor-1α (SDF-1α) and its receptor, CXCR4, could contribute to neural stem cells migration after brain injury. In the present study, the temporal relationship between neural stem cells (NSCs) and SDF-1α/CXCR4 around a damaged area was investigated using a rat traumatic brain injury (TBI) model.

Methods: We used molecular biology techniques and immunohistochemistry to investigate the relationship between SDF-1α/CXCR4 expression and NSCs existence around a damaged area after TBI in the rat brain.

Results: SDF-1α mRNA expression and SDF-1α protein synthesis did not increase after TBI. However, SDF-1α leaked from the injured area and diffused into the cortex 1–3 days after TBI. Subsequently, the levels of CXCR4 mRNA expression and CXCR4 protein synthesis increased significantly. Many small cells with a nestin-positive cytoplasm and fibers also showed immunopositivity for both CXCR4 and SOX-2, but not for GFAP, 3–7 days after TBI. Moreover, a proportion of the CXCR4-positive cells and fibers also showed immunostaining for neurofilaments.

Discussion: These results suggest that the leaked SDF-1α attracted CXCR4-positive NSCs as well as elongated nerve fibers. It is considered that the SDF-1α/CXCR4 system in the brain contributes to neural stem cells appearance and maturation after TBI. Therefore, exploitation of the SDF-1α/CXCR4 system around a damaged area may improve the brain dysfunction after TBI.     

Signalling Pathway Alterations in Pituitary Adenomas: Involvement of Gsα, cAMP and Mitogen-Activated Protein Kinases

Despite extensive research on sporadic pituitary adenomas, it is not yet possible to assign one protein alteration to one specific type of pituitary adenomas. Nevertheless, alterations of the cAMP pathway appear to be molecular hallmarks of most growth hormone (GH)-secreting adenomas. However, these alterations do not confer specific phenotypes to patients carrying these alterations. In this review, we summarise the literature regarding signalling alterations observed in GH-secreting adenomas. We focus on Gsα alterations and their possible cross-talk with the extracellular signal-related kinase (ERK)1/2 pathway. In the light of results obtained on human somatotroph adenoma cells in primary culture and on models of murine somatotroph cell lines, we postulate a crucial role for ERK1/2 in GH-secreting adenomas downstream of cAMP pathway alterations that might impact the tumoural phenotype.     

Stromal cell-derived factor 1-mediated CXCR4 signaling in rat and human cortical neural progenitor cells

Stromal cell-derived factor 1 (SDF-1) and the chemokine receptor CXCR4 are highly expressed in the nervous system. Knockout studies have suggested that both SDF-1 and CXCR4 play essential roles in cerebellar, hippocampal, and neocortical neural cell migration during embryogenesis. To extend these observations, CXCR4 signaling events in rat and human neural progenitor cells (NPCs) were examined. Our results show that CXCR4 is expressed in abundance on rat and human NPCs. Moreover, SDF-1alpha induced increased NPCs levels of inositol 1,4,5-triphosphate, extracellular signal-regulated kinases 1/2, Akt, c-Jun N-terminal kinase, and intracellular calcium whereas it diminished cyclic adenosine monophosphate. Finally, SDF-1alpha can induce human NPC chemotaxis in vitro, suggesting that CXCR4 plays a functional role in NPC migration. Both T140, a CXCR4 antagonist, and pertussis toxin (PTX), an inactivator of G protein-coupled receptors, abrogated these events. Ultimately, this study suggested that SDF-1alpha can influence NPC function through CXCR4 and that CXCR4 is functional on NPC.     

骨髓单个核细胞移植对心肌基质细胞衍生因子1-CXCR4轴的影响

背景：心脏干细胞移植后心肌基质细胞衍生因子1-CXCR4轴表达及其作用越来越受到人们的关注。 目的：观察经心外膜注骨髓单个核细胞对心衰犬心脏基质细胞衍生因子1-CXCR4轴 mRNA表达的影响。 方法：16只杂种犬随机数字表法均分为移植组和对照组,植入永久起搏器。右室快速起搏三四周后建立心衰模型。移植组犬经心外膜多点注射骨髓单个核细胞悬液,对照组注射等量生理盐水。 结果与结论：快速起搏三四周后,各项超声参数及血流动力学参数较起搏前改变明显,差异有显著性意义。定量PCR检测细胞移植组基质细胞衍生因子1 mRNA及CXCR4 mRNA表达水平高于对照组(P < 0.01)。说明经心外膜注射的骨髓单个核细胞可提高心肌基质细胞衍生因子1 mRNA及CXCR4 mRNA表达水平。     

CXCR7, CXCR4 and CXCL12: an eccentric trio?

CXCR7, formerly called RDC1 is a recently deorphanized G-protein coupled receptor which binds with high affinity the inflammatory and homing chemokines CXCL11/ITAC and CXCL12/SDF-1. Despite its phylogenetic relation and ligand binding properties CXCR7 does not mediate typical chemokine receptor responses such as leukocyte trafficking. Recent findings in zebrafish indicate that a critical activity of the receptor is scavenging of CXCL12 thereby generating guidance cues for CXCR4-dependent migration. The observations do not exclude the possibility that the receptor is capable of inducing signal transduction which is suggestive from studies of tumor growth and survival. The pronounced expression in central and peripheral nervous tissue and the absence of a brain phenotype in CXCR7(-/-) mice suggest a subtle activity of the receptor.     

Stromal-derived factor 1 signalling regulates radial and tangential migration in the developing cerebral cortex

Stromal-derived factor 1 (SDF-1), a known chemoattractant, and its receptor CXCR4 are widely expressed in the developing and adult cerebral cortex. Recent studies have highlighted potential roles for SDF-1 during early cortical development. In view of the current findings, our histological analysis has revealed a distinct pattern of SDF-1 expression in the developing cerebral cortex at a time when cell proliferation and migration are at peak. To determine the role of chemokine signalling during early cortical development, embryonic rat brain slices were exposed to a medium containing secreted SDF-1 to perturb the endogenous levels of chemokine. Alternatively, brain slices were treated with 40 muM of T140 or AMD3100, known antagonists of CXCR4. Using these experimental approaches, we demonstrate that chemokine signalling is imperative for the maintenance of the early cortical plate. In addition, we provide evidence that both neurogenesis and radial migration are concomitantly regulated by this signalling system. Conversely, interneurons, although not dependent on SDF-1 signalling to transgress the telencephalic boundary, require the chemokine to maintain their tangential migration. Collectively, our results demonstrate that SDF-1 with its distinct pattern of expression is essential and uniquely positioned to regulate key developmental events that underlie the formation of the cerebral cortex.     

Regional and cellular localization of the CXCl12/SDF-1 chemokine receptor CXCR7 in the developing and adult rat brain

The chemokine stromal cell-derived factor-1 (SDF-1) regulates neuronal development via the chemokine receptor CXCR4. In the adult brain the SDF-1/CXCR4 system was implicated in neurogenesis, neuromodulation, brain inflammation, tumor growth, and HIV encephalopathy. Until the recent identification of RDC1/CXCR7 as the second SDF-1 receptor, CXCR4 was considered to be the only receptor for SDF-1. Here we provide the first map of CXCR7 mRNA expression in the embryonic and adult rat brain. At embryonic stages, CXCR7 and CXCR4 were codistributed in the germinative zone of the ganglionic eminences, caudate putamen, and along the routes of GABAergic precursors migrating toward the cortex. In the cortex, CXCR7 was identified in GABAergic precursors and in some reelin-expressing Cajal-Retzius cells. Unlike CXCR4, CXCR7 was abundant in neurons forming the cortical plate and sparse in the developing dentate gyrus and cerebellar external germinal layer. In the adult brain, CXCR7 was expressed by blood vessels, pyramidal cells in CA3, and mature dentate gyrus granule cells, which is reminiscent of the SDF-1 pattern. CXCR7 and CXCR4 overlapped in the wall of the four ventricles. Further neuronal structures expressing CXCR7 comprised the olfactory bulb, accumbens shell, supraoptic and ventromedial hypothalamic nuclei, medial thalamus, and brain stem motor nuclei. Also, GLAST-expressing astrocytes showed signals for CXCR7. Thus, CXCR4 and CXCR7 may cooperate or act independently in SDF-1-dependent neuronal development. In mature neurons and blood vessels CXCR7 appears to be the preponderant SDF-1-receptor.     

诺帝对CXCR4介导的人恶性胶质瘤细胞钙流和白细胞介素8分泌的抑制作用

目的 观测诺帝(Nordy)对人恶性胶质瘤细胞系U87细胞趋化因子受体CXCR4活化后钙流变化及白细胞介素8(IL-8)产生的影响.方法 采用Fluo-4/AM标记钙离子、激光共聚焦显微术观测CXCR4被其配体间质细胞衍生因子-1α(SDF-1α)活化后U87细胞钙流的变化,并通过酶联免疫吸附试验(ELISA)检测细胞培养上清中IL-8的含量;以CXCR4抑制剂AMD3100为对照,用自行合成的化合物诺帝顸处理U87细胞,再用SDF-1α刺激,观测二者对CXCR4活化后引起的钙流变化和IL-8分泌的影响.结果 25～100ng/ml SDF-1α可不同程度地引起U87细胞胞内钙流增加和IL-8分泌量增多,与对照组(无SDF-1α刺激)相比均有显著统计学意义(P<0.05);用1～10μmol/LAMD3100或25～100μmol/L诺帝预处理细胞,再用SDF-1α刺激,上述效应受到抑制.结论 诺帝可抑制CXCR4活化引起的钙流和IL-8产生,这可能是其抗胶质瘤血管生成机制之一.     

雌激素受体在人类垂体腺瘤中的表达

目的研究雌激素受体(ER)在人类垂体腺瘤中的表达及其与垂体腺瘤临床特征及所分泌激素类型的关系.方法应用免疫组化S-P法和原位杂交法检测55例垂体腺瘤中ER蛋白及ERmRNA的表达,并采用S-P法对其中53例标本进行激素分型.结果在不同性别及不同病程垂体腺瘤患者中,ER蛋白及ERmRNA在肿瘤组织中的表达差异无显著性.不同体积垂体腺瘤中,ER蛋白及ERmRNA表达阳性率有随肿瘤体积增大而增高的趋势,但ER蛋白表达仅在大腺瘤组和巨大腺瘤组具有显著差异(P＜0.05).53例大腺瘤及巨大腺瘤中,部分PRL、FSH、LH单激素腺瘤及部分多激素腺瘤有ER蛋白及ERmRNA表达,而全部GH、ACTH、TSH单激素腺瘤均无ER蛋白及ERmRNA表达,4例无功能腺瘤均无ER蛋白表达,仅1例有ERmRNA表达.结论垂体腺瘤患者的性别和病程不影响肿瘤组织中ER的表达.推测ER蛋白及ERmRNA表达与垂体腺瘤的高增殖性及垂体腺瘤的激素分泌类型有关.     

基质细胞衍生因子-1对间质干细胞迁移的影响

目的:观察基质细胞衍生因子-1(SDF-1)在体内外对大鼠骨髓间质干细胞(rMSCs)的趋化诱导作用,探讨SDF-1对rMSCs迁移影响的可能机制。方法:应用体外细胞迁移实验及大鼠脑梗死模型体内移植,观察SDF-1对rMSCs的迁移影响。流式细胞术与RT-PCR检测rMSCs的CXC趋化因子受体4(CXCchemokinereceptor4,CXCR4)表达。结果:在SDF-1存在时,rMSCs迁移活跃,应用抗体封闭CXCR4后,这种迁移显著减弱。体内移植的rMSCs主要聚集在脑梗死灶周围,但在封闭CXCR4后,这种聚集现象大大减弱。流式细胞术示仅小部分rMSCs表面表达CXCR4,但经TritonX-100处理后,表达CXCR4的rMSCs增加。结论:SDF-1可通过CXCR4对rMSCs起趋化作用,针对这种作用可望调控干细胞向靶组织的趋化聚集量,达到治疗目的。     

Estrogen receptors in brain tumors

We examined the cytosolic estrogen receptor (ER) level in tumor tissue from 77 patients: 36 meningiomas, 20 gliomas (12 glioblastomas, 2 cerebellar astrocytomas, 2 ependymomas, and 4 medulloblastomas), 8 neurinomas, 7 pituitary adenomas (2 prolactin-producing adenomas, 1 growth hormone-producing adenoma, and 4 nonfunctioning adenomas), and 6 metastatic brain tumors (1 from breast cancer, 4 from lung cancers, and 1 from colon cancer). Nuclear ER levels were assayed in 11 meningiomas and 2 glioblastomas. ER was determined by the dextran-coated charcoal method and calculated by Scatchard analysis. Cytosolic ER was detected in 100% of the pituitary adenomas, 50% of the meningiomas, 50% of the metastatic brain tumors, 25% of the neurinomas, and 15% of the gliomas. In gliomas, only medulloblastomas had ER activity. Nuclear ER was found in three premenopausal women with meningioma. The dissociation constant of the ER complex was, in each case, less than 10(-9) M. These observations suggest that some brain tumors may be responsive to estrogen via the cellular ER.     

脑梗死患者外周血SDF-1/CXCR7表达水平与颈总动脉内膜中层厚度及斑块积分的相关性分析

目的 探讨基质细胞衍生因子-1(Stromal cell-derived factor-1,SDF-1)/趋化因子受体7(Chemokine receptor 7,CXCR7)在脑梗死患者血清中的表达水平变化及其与颈动脉内膜中层厚度(Intima-media thickness,IMT)、斑块积分的相关性。方法 选取2017年6月-2019年4月本院收治的186例脑梗死患者进行研究,称脑梗死组; 并选取同期127例体检健康者进行对照研究,称对照组; 比较2组一般资料; 采用实时荧光定量PCR(Real-time fluorescent quantitative PCR,qRT-PCR)法检测2组血清SDF-1,CXCR7表达水平; 利用彩色多普勒超声仪检测2组IMT,记录斑块数量和大小,并进行斑块积分; Pearson法分析脑梗死患者血清SDF-1,CXCR7表达水平与IMT、斑块积分,SDF-1与CXCR7的关系; Logistic法分析脑梗死的影响因素。结果 脑梗死组甘油三酯(Triacylglycerol,TG)、总胆固醇(Total cholesterol,TC)、低密度脂蛋白-胆固醇(Low density lipoprotein cholesterol,LDL-C)、血清SDF-1、CXCR7水平、IMT、斑块积分均明显高于对照组(P<0.05),高密度脂蛋白-胆固醇(High density lipoprotein cholesterol,HDL-C)水平明显低于对照组(P<0.05); 脑梗死血清SDF-1表达水平与IMT、斑块积分,CXCR7表达水平与IMT、斑块积分,SDF-1表达水平与CXCR7表达水平均呈正相关(0.310<r<0.491,P<0.05); LDL-C,SDF-1,CXCR7表达水平、IMT、斑块积分为脑梗死发生的危险因素(P<0.05); 高密度脂蛋白胆固醇水平为脑梗死发生的保护因素(P<0.05)。结论 脑梗死患者血清SDF-1,CXCR7表达水平上调,两者与IMT、斑块积分均呈正相关,SDF-1,CXCR7可能与IMT、斑块数量、大小相互影响,从而共同在脑梗死发病过程中发挥作用。     

Characterization and visualization of [125I] stromal cell-derived factor-1alpha binding to CXCR4 receptors in rat brain and human neuroblastoma cells

Stromal cell-Derived Factor-1 (SDF-1alpha), binds to the seven-transmembrane G protein-coupled CXCR4 receptor and modulates cell migration, differentiation, and proliferation. CXCR4 has been reported to be expressed in various tissues including brain. Moreover, CXCR4 has recently been shown to be one of the coreceptors for HIV-1 infection which could be implicated in HIV encephalitis. In the present study, the binding properties and autoradiographic distribution of [125I]SDF-1alpha binding to CXCR4 were characterized in the adult rat brain. SDF-1alpha binding and CXCR4 coupling system were also studied in human neuroblastoma cell line SK-N-SH. The binding of [125I]SDF-1alpha on rat brain sections was specific, time-dependent and reversible. The highest densities of CXCR4 were detected in the choroid plexus of the lateral and the dorsal third ventricle. Lower densities of [125I]SDF-1alpha binding sites were observed in various brain regions including cerebral cortex, anterior olfactory nuclei, hippocampal formation, thalamic nuclei, blood vessels and pituitary gland. In the choroid plexus, the IC(50) and K(d) of [125I]SDF-1alpha binding were respectively 0.6 nM and 0. 36 nM. Similar IC(50) values were obtained in other brain structures. A CXCR4 antagonist, bicyclam, competed with SDF-1alpha binding (30% inhibition at 10(-6) M). In SK-N-SH cells, [125I]SDF-1alpha bound to CXCR4 with a K(d) of 5.0 nM and a maximal binding capacity of 460 fmol/mg of protein. SDF-1alpha induced a rapid and transient intracellular calcium increase in SK-N-SH cells. These findings suggest that CXCR4 is highly expressed in some brain structures and have a regulatory role in the nervous system. The significance of this expression in the brain parenchyma and more specifically in the choroid plexus remains to be clarified in the normal as well as in the infected brain.     

Expression of SDF-1 and CXCR4 during reorganization of the postnatal dentate gyrus

Previous studies have demonstrated that stromal cell-derived factor 1 (SDF-1) is crucial for early dentate development; however, the mouse mutants for this chemokine and its only receptor, CXCR4, are neonatally lethal, making conclusions about the role of these molecules in postnatal development difficult to sustain. Previous expression analyses have used single labeling, but the distribution of CXCR4 is complex and to determine the cell types expressing CXCR4 requires multiple marker labeling. In this study, we examined the distribution of SDF-1 and CXCR4 mRNAs during the first postnatal weeks, combining these markers with several other cell-type-specific markers. We found that SDF-1 has three sites of expression: (1) continuation of prenatal expression in the meninges; (2) expression in Cajal-Retzius cells occupying the molecular layer of the upper and lower blades of the dentate, and (3) the maturing dentate granule neurons themselves. The timing of expression in these three sites corresponds to alterations in the distribution of the primary cell types expressing CXCR4 during the same periods, notably the expression of CXCR4 in radial-glial-like GFAP-expressing dentate precursors and immature dentate granule neurons. Taken together, our data suggest potential ongoing roles for SDF-1/CXCR4 signaling in the dentate gyrus during the early postnatal period that will be tested in the future with more precise genetic approaches.