Chemokine stromal cell-derived factor 1alpha activates basophils by means of CXCR4

BACKGROUND: The CXC chemokine receptor 4 (CXCR4) is predominantly expressed on inactivated naive T lymphocytes, B lymphocytes, dendritic cells, and endothelial cells. CXC chemokine stromal cell-derived factor 1alpha (SDF-1alpha) is the only known ligand for CXCR4. To date, the CXCR4 expression and function of SDF-1alpha in basophils are unknown. OBJECTIVE: The purpose of this study was to investigate the expression of CXCR4 and functions of SDF-1alpha in basophils and to characterize the role of the CXCR4-SDF-1alpha receptor ligand pair in the allergic inflammation. METHODS: Basophil purification, flow cytometry, real-time quantitative RT-PCR assay, Northern blotting, intracellular free Ca(2+) change, chemotaxis assay, and histamine release assay were used. RESULTS: CXCR4 is abundantly expressed on peripheral blood resting basophils (91%). Likewise, CXCR4 messenger (m)RNA is expressed in resting basophils (3.2 x 10(3) copies per 2 x 10(2) cells). The existence of CXCR4 mRNA was also confirmed in basophils by means of Northern blot analysis. SDF-1alpha induces an increase in intracellular free Ca(2+) in basophils. SDF-1alpha activates basophils to chemotaxis (chemotactic index = 3.8) and histamine release (36% of total content) through CXCR4 on the cells. The chemokines SDF-1alpha, eotaxin, RANTES, monocyte chemoattractant protein (MCP) 1, and macrophage inflammatory protein (MIP) 1alpha have been demonstrated at different potencies in induction of chemotaxis (eotaxin > SDF-1alpha > RANTES congruent with MCP-1 > MIP-1alpha) and histamine release (MCP-1 congruent with SDF-1alpha > eotaxin > RANTES > MIP-1alpha). The optimal concentration seen for SDF-1alpha effects (chemotaxis and histamine release) on basophils was 100 ng/mL. CONCLUSION: These results indicate that the CXCR4-SDF-1alpha receptor ligand pair may be important for the recruitment and activation of the basophils, which is a characteristic effector cell of the allergic inflammation.     

Functional expression of chemokine receptor CXCR4 on human epithelial cells

Chemokines and their receptors play an important role in the process of leucocyte recruitment at sites of inflammation. However, recent evidence suggests that these proteins can also regulate non-leucocyte cell functions such as angiogenesis, migration and proliferation. We have investigated the expression of the CXC chemokine receptor 4 (CXCR4) on primary cultures of type II alveolar epithelial cells, their transformed counterpart, the A549 cell line and also on other epithelial cell lines from various tissues. We found that all epithelial cell types tested express mRNA for CXCR4. Flow cytometric analysis and immunocytochemical staining shows that CXCR4 chemokine receptor is abundantly expressed on the surface of A549 epithelial cells. Furthermore, A549 cells responded to the CXCR4 ligand, stromal-derived factor-1alpha (SDF-1alpha) with a rapid and robust calcium mobilization and not to other CXC chemokines, suggesting that CXCR4 is functionally active and is able to couple to G-protein signalling mechanisms. A549 cells did not proliferate in response to either SDF-1alpha or interleukin-8 (IL-8) CXC chemokines. These findings may have important implications for epithelial physiology and pathology.     

Selective sequestration of X4 isolates by human genital epithelial cells: Implication for virus tropism selection process during sexual transmission of HIV

X4 and R5 HIV strains are present in the semen of men infected with HIV but R5 isolates are transmitted preferentially. The role of human epithelial cells in this selection is addressed. Three human cervical cell lines-CaSki, SiHa, and HEC1A-and normal human vaginal cells from HIV-negative donors were characterized for HIV receptor expression and incubated with X4 and R5 laboratory-adapted strains or primary isolates. The infection was assessed by detection of intracellular HIV DNA. The three cell lines were shown to express on their surface the CXCR4 and GalCer molecules, but not the CD4 and CCR5 ones. The three cell lines and normal human vaginal cells were found to be selectively permissive to X4 HIV entry; the preincubation of the cell lines with rhSDF-1 inhibited this infection. The detection of the intracellular proviral DNA in the cell lines and in normal human vaginal cells demonstrated a selective integration of X4 strains. Additional experiments showed that no extracellular RNA was detected in the supernatants of HEC1A cells infected by X4 isolates either after 18 days of culture or after incubation with PHA-stimulated PBMCs and that no transmission occurred after co-culture between infected HEC1A cells and PHA-stimulated PBMCs. These results suggest specific sequestration of X4 strains by genital epithelial cells, which could explain, at least in part, the HIV tropism selection process during sexual intercourse.     

CXCR4在溃疡性结肠炎小鼠蛋白C系统变化中的作用

目的:蛋白C系统(PCS)是血管内皮功能重要的介导者,而微血管内皮细胞是参与溃疡性结肠炎(UC)最重要的非免疫细胞,本研究旨在探讨趋化因子受体CXCR4在UC病理过程中PCS变化的作用。方法:动物实验分为对照组和UC模型组,进行大体积分、组织病理学积分及溃疡指数评估;测定各组结肠组织和血浆中髓过氧化物酶(MPO)、环氧合酶-2(COX-2)、基质细胞衍生因子-1α(SDF-1α)和单核细胞趋化蛋白-1(MCP-1)的mRNA水平及活性;检测各组结肠CXCR4、β-arrestin、p-JNK、内皮细胞蛋白C受体(EPCR)和血栓调节蛋白(TM)的水平及分布;观察蛋白C(PC)和蛋白S(PS)的活性。分离、培养、鉴定结肠微血管内皮细胞,分为对照组、SDF-1α组、CXCR4沉默组和CXCR4过表达组,观察各组细胞EPCR、TM、β-arrestin和p-JNK的蛋白水平,以及PC、PS和激活的蛋白C(APC)活性。结果:与对照组比,模型组小鼠大体积分、组织病理学积分及溃疡指数升高(P0.05)。结肠组织和血浆MPO、COX-2、SDF-1α和MCP-1 mRNA水平和活性显著升高(P0.01)。结肠组织CXCR4、β-arrestin和p-JNK的蛋白水平上调,EPCR表达下调,血浆PC和PS活性明显降低(P0.05或P0.01)。CXCR4过表达进一步加重SDF-1α对结肠黏膜微血管内皮细胞PCS的抑制,同时进一步升高β-arrestin和p-JNK的蛋白水平(P0.05)。结论:UC时PCS被抑制,CXCR4参与其中,其可能的机制是CXCR4在UC病理过程中介导趋化因子通过β-arrestin-JNK信号通路进一步影响血管内皮细胞功能,从而抑制PCS。     

CXCR4 positive bone mesenchymal stem cells migrate to human endothelial cell stimulated by ox-LDL via SDF-1α/CXCR4 signaling axis

Background

Bone mesenchymal stem cells (BMSCs) are attractive candidates for cell based therapies to cardiovascular disease such as infarction and atherosclerosis; however, the mechanisms responsible for stem cell chemotaxis and homing remain unknown. Chemokine stromal cell-derived factor 1 (SDF-1α) is involved in the process of atherogenesis. This study was aimed at investigating whether the SDF-1α of human umbilical vein endothelial cells (HUVECs) plays a role in migration of BM-derived CXCR4⁺(receptor for SDF-1α) stem cells.

Methods

HUVECs were cultured from human umbilical cords and was treated with ox-LDL. The mRNA and protein expression of SDF-1α was detected in HUVECs. CXCR4⁺BMSCs from bone marrow were isolated and were tested by migration and adhesion assays.

Results

It was found that ox-LDL induced HUVECs to increase the mRNA and protein expression of SDF-1α. Ox-LDL increased the migratory and adhesion response of CXCR4⁺BMSCs. When the neutralizing SDF-1α antibody abrogated the secreted SDF-1α, the migration and adhesion response of CXCR4⁺BMSCs markedly decreased.

Conclusions

Our data indicated that the endothelial cells (ECs) stimulated by ox-LDL could increase the BMSCs migratory response via SDF-1α/CXCR4 signaling axis. These findings provide a new paradigm for biological effects of ox-LDL and have implications for novel stem cell therapeutic strategies for atherosclerosis.     

Stromal cell-derived factor-1 from biliary epithelial cells recruits CXCR4-positive cells: implications for inflammatory liver diseases

Although stromal cell-derived factor-1 (SDF-1) plays an important role in hematopoiesis in the fetal liver, the role after birth remains to be clarified. We investigated the role of SDF-1 and its receptor, CXCR4, in 75 patients; this included controls and patients with viral hepatitis, liver cirrhosis, primary biliary cirrhosis, primary sclerosing cholangitis, and autoimmune hepatitis. Interestingly, SDF-1 appeared up-regulated in biliary epithelial cells (BEC) of inflammatory liver disease. Furthermore, in inflammatory liver diseases, SDF-1 was expressed by BEC of interlobular and septal bile ducts and by proliferated bile ductules. The message expression of SDF-1 in BEC was confirmed at a single-cell level by RT-PCR and laser capture microdissection. The plasma levels of SDF-1 were significantly higher in patients with liver diseases than in normal controls. Flow cytometric analysis of the surface expression of CXCR4 showed that most liver-infiltrating lymphocytes express CXCR4 and the intensity was up-regulated more significantly in liver-infiltrating lymphocytes than in peripheral blood lymphocytes. These results suggest that increased SDF-1 production by BEC may play an important role in the recruitment of CXCR4-positive inflammatory cells into the diseased livers. These data are significant because modulation of the SDF-1/CXCR4 interaction has therapeutic implications for inflammatory liver diseases.     

Syngenic bone marrow cells restore hepatic function in carbon tetrachloride-induced mouse liver injury

Progenitor cells in bone marrow have been explored for the treatment of liver injury. Stem cell homing to the injured tissue is regulated through stromal cell derived factor-1 (SDF-1) and its receptor CXCR4. We hypothesized that syngenic bone marrow cells (BMCs) would restore hepatic function in the injured liver through the regulation by SDF-1/CXCR4 system. After injecting carbon tetrachloride (CCl(4)), the mice were injected with syngenic BMCs or normal saline. Morphological and functional analysis of the liver was performed. Flow cytometry for the stem cell markers and CXCR4 was done with the liver, BM, and spleen cells from each group. Carboxyfluorescein diacetate succinimidyl ester was used to trace the homing of transplanted BMCs. The SDF-1 expression of the liver was assessed by immunohistochemistry. Hepatosplenomegaly and necrosis of the CCl(4)-injected mouse liver were improved after BMCs transplantation The hepatic enzymes were increased after injury and then decreased after BMCs transplantation. The expression of stem cell markers and CXCR4 was exclusively increased in the damaged liver compared to the BM and spleen, and even more elevated after BMCs transplantation. SDF-1 expression in the liver was observed after CCl(4) injection and it was elevated after BMCs transplantation. The intrinsic and extrinsic BMCs migrate specifically to the injured liver rather than BM or spleen, and the transplanted BMCs contribute to the repair of the damaged liver. SDF-1/CXCR-4 interaction plays a role in stem cell homing toward the damaged organ, and transplanted BMCs are involved in the up-regulated SDF-1 expression seen in the injured liver.     

The chemokine SDF-1α triggers a chemotactic response and induces cell polarization in human B lymphocytes

We studied the expression and possible functional role of chemokine receptors CXCR3, CXCR4 and CCR5 in normal human B lymphocytes. B cells from both peripheral blood and tonsils expressed high levels of CXCR4 but not the other chemokine receptors tested. CXCR4 ligand, stromal cell-derived factor (SDF)-1α, elicited a potent chemotactic response and induced a polarized motile phenotype in B cells, resulting in redistribution of the adhesion molecule ICAM-3 to a posterior appendage of the cell, termed uropod, and of CXCR4 receptor to the leading edge of migrating B cells. Time-lapse videomicroscopy studies revealed that SDF-1α-treated cells recruited additional bystander B cells through the uropod. SDF-1α induced levels of cellular recruitment comparable to those elicited by polarization-inducing anti-ICAM-3 monoclonal antibody, in an LFA-1/ICAM-1, −3-dependent fashion. Moreover, this chemokine increased intracellular Ca²⁺ levels in B lymphocytes, and induced a rapid CXCR4 receptor down-regulation on the cell surface membrane. These results provide new insight into the important biological role of SDF-1α in physiological processes in which B cells participate, and suggest a key role for chemokines in normal B cell trafficking and recirculation.     

间质细胞衍生因子1及其受体CXCR4在浸润性乳腺癌中的表达及其临床意义

目的 研究间质细胞衍生因子1(SDF-1)及其受体CXCR4在浸润性乳腺癌中的表达,并分析其与浸润性乳腺癌相关临床病理指标及淋巴结转移之间的关系.方法 采用免疫组织化学LSAB方法检测SDF-1/CXCR4在120例浸润性乳腺癌中的表达情况;采用地高辛标记的寡核苷酸探针进行原位杂交以检测趋化因子SDF-1在肿瘤环境中表达的部位及来源.结果 (1)SDF-1主要表达于肿瘤细胞的胞质和胞膜;SDF-1的胞质表达在淋巴结阳性组高于阴性组(P=0.033),且其表达程度与淋巴结受累数目、病理学分期、组织学分级、肿瘤大小及ER表达等指标呈正相关(P<0.05);(2)罕见SDF-1 mRNA表达的脉管内皮却可见SDF-1蛋白的表达,其表达程度与肿瘤胞质SDF-1着色正相关(P<0.01);且淋巴管内皮SDF-1的着色与淋巴结转移程度为正相关(P=0.005);血管内皮SDF-1的着色与肿瘤环境中的淋巴细胞浸润正相关(P<0.01),且同时伴有较多淋巴细胞浸润及SDF-1血管内皮着色阳性的病例,其淋巴结的转移程度分别高于仅有上述条件之一或二者均不具备的各组病例(P<0.05);(3)CXCR4也主要表达于肿瘤细胞的胞质和胞核;CXCR4的胞质表达在淋巴结阳性组高于阴性组(P<0.05),且其表达程度与淋巴结受累数目、病理学分期、组织学分级、肿瘤大小及HER2表达等呈正相关(P=0.005),而胞核的表达仅与PR的表达情况呈正相关(P<0.01);(4)瘤细胞质CXCR4与SDF-1的表达呈正相关(P=0.001).结论 浸润性乳腺癌肿瘤细胞SDF-1和CXCR4的表达与多项临床病理指标,尤其是淋巴结转移率及转移程度有关,可作为预测乳腺癌淋巴结转移及预后的免疫病理学指标,同时应注意肿瘤微环境中SDF-1的多个来源及定位的不同意义.     

Increased expression of SDF-1/CXCR4 is associated with lymph node metastasis of invasive micropapillary carcinoma of the breast

Aims:  Stromal cell-derived factor-1 (SDF-1) and its receptor CXCR4 are implicated in tumour chemotaxis and metastasis. The aim was to examine their roles in the metastasis of invasive micropapillary carcinoma (IMPC) of the breast, a tumour with a high propensity for nodal spread.
Methods and results:  We compared the expression of SDF-1 and CXCR4 in 103 cases of breast cancer containing IMPC components with a control group of 96 cases of invasive ductal carcinoma (IDC), not otherwise specified type by immunohistochemistry and chemical in situ hybridization (CISH). The results showed that the predominant cytoplasmic expression of both SDF-1 and CXCR4 was greater in tumour cells of the IMPC components than in those of the non-IMPC components and the control IDC cases, and was correlated significantly with the number of positive lymph nodes ( P  < 0.05). SDF-1 expression on cell membranes was less frequently identified in IMPC than IDC ( P  = 0.021). Immunohistochemical detection of SDF-1 in endothelial cells of lymphatic vessels was more common in IMPC ( P   =  0.007) and correlated significantly with lymph node status ( P  = 0.002), although SDF-1 mRNA was rarely detected by CISH.
Conclusions:  This study suggests that up-regulation of cytoplasmic expression of SDF-1/CXCR4 might be one of the molecular mechanisms facilitating lymph node metastasis of IMPC.     

HIV-1协同受体CXCR4、CCR5及相关趋化因子SDF-1在胎盘的表达

目的：研究HIV-1协同受体CXCR4、CCR5及CXCR4的特异性配体SDF-1在人胎盘组织的表达，探索HIV-1子宫内垂直传播的分子机制。方法：半定量RT-PCR检测早、中、晚孕期胎盘及早孕滋养细胞CXCR4、CCR5 mRNA水平；免疫组化和免疫细胞化学检测早孕胎盘及原代培养滋养细胞CXCR4、CCR5蛋白表达；原位杂交及免疫组化分析SDF-1在早孕胎盘的表达；ELISA测定滋养细胞SDF-1的动态分泌水平。结果：各孕期胎盘表达CXCR4及CCR5 mRNA；CXCR4蛋白定位于滋养细胞，而CCR5蛋白定位于绒毛基质中。滋养细胞可转录并翻译SDF-1，且能分泌可溶性SDF-1。结论：滋养细胞同时表达CXCR4及SDF-1，SDF-1可能通过降调CXCR4而拮抗X4-HIV-1感染胎儿细胞；R5-HIV-1或许能通过滋养层裂隙感染CCR5^#基质细胞和/或Hotbauer细胞，从而发生子宫内垂直传播。     

Hypoxia- and vascular endothelial growth factor-induced stromal cell-derived factor-1alpha/CXCR4 expression in glioblastomas: one plausible explanation of Scherer's structures

The morphological patterns of glioma cell invasion are known as the secondary structures of Scherer. In this report, we propose a biologically based mechanism for the nonrandom formation of Scherer's secondary structures based on the differential expression of stromal cell-derived factor (SDF)-1alpha and CXCR4 at the invading edge of glioblastomas. The chemokine SDF-1alpha was highly expressed in neurons, blood vessels, subpial regions, and white matter tracts that form the basis of Scherer's secondary structures. In contrast, the SDF-1alpha receptor, CXCR4, was highly expressed in invading glioma cells organized around neurons and blood vessels, in subpial regions, and along white matter tracts. Neuronal and endothelial cells exposed to vascular endothelial growth factor up-regulated the expression of SDF-1alpha. CXCR4-positive tumor cells migrated toward a SDF-1alpha gradient in vitro, whereas inhibition of CXCR4 expression decreased their migration. Similarly, inhibition of CXCR4 decreased levels of SDF-1alpha-induced phosphorylation of FAK, AKT, and ERK1/2, suggesting CXCR4 involvement in glioma invasion signaling. These studies offer one plausible molecular basis and explanation of the formation of Scherer's structures in glioma patients.     

The SDF-1-CXCR4 axis stimulates VEGF secretion and activates integrins but does not affect proliferation and survival in lymphohematopoietic cells

To better define the role HIV-related chemokine receptor-chemokine axes play in human hematopoiesis, we investigated the function of the CXCR4 and CCR5 receptors in human myeloid, T- and B-lymphoid cell lines selected for the expression of these receptors (CXCR4(+), CXCR4(+) CCR5(+), and CCR5(+) cell lines). We evaluated the phosphorylation of MAPK p42/44, AKT, and STAT proteins and examined the ability of the ligands for these receptors (stromal-derived factor-1 [SDF-1] and macrophage inflammatory protein-1beta [MIP-1beta]) to influence cell growth, apoptosis, adhesion, and production of vascular endothelial growth factors (VEGF), matrix metalloproteinases (MMPs) and their tissue inhibitors (TIMPs) in these cell lines. We found that A) SDF-1, after binding to CXCR4, activates multiple signaling pathways and that in comparison with the MIP-1beta-CCR5 axis, plays a privileged role in hematopoiesis; B) SDF-1 activation of the MAPK p42/44 pathway and the PI-3K-AKT axis does not affect proliferation and apoptosis but modulates integrin-mediated adhesion to fibronectin, and C) SDF-1 induces secretion of VEGF, but not of MMPs or TIMPs. Thus the role of SDF-1 relates primarily to the interaction of lymphohematopoietic cells with their microenvironment and does not directly influence their proliferation or survival. We conclude that perturbation of the SDF-1-CXCR4 axis during HIV infection may affect interactions of hematopoietic cells with the hematopoietic microenvironment.     

Stromal-derived factor-1 and its receptor, CXCR4, are constitutively expressed by mouse liver sinusoidal endothelial cells: implications for the regulation of hematopoietic cell migration to the liver during extramedullary hematopoiesis

Stromal-derived factor (SDF)-1 is the main regulating factor for trafficking/homing of hematopoietic stem cells (HSC) to the bone marrow (BM). It is possible that this chemokine may also play a fundamental role in regulating the migration of HSC to several organs during extramedullary hematopoiesis. Because liver sinusoidal endothelial cells (LSEC) constitute an extramedullary niche for HSC, it is possible that these cells represent one of the main cellular sources of SDF-1 at the liver. Here, we show that LSEC express SDF-1 at the mRNA and protein level. Biological assays showed that conditioned medium from LSEC (LSEC-CM) stimulated the migration of BM progenitor lineage-negative (BM/Lin?) cells. This effect was significantly reduced by AMD3100, indicating that the SDF-1/CXCR4 axis is involved in the stimulatory migrating effect induced by LSEC-CM. Early localization of HSC in SDF-1-expressing LSEC microenvironment together with increased levels of this chemokine in hepatic homogenates was found in an experimental model of liver extramedullary hematopoiesis. Flow cytometry studies showed that LSEC express the CXCR4 receptor. Functional assays showed that activation of this receptor by SDF-1 stimulated the migration of LSEC and increased the expression of PECAM-1. Our findings suggest that LSEC through the production of SDF-1 may constitute a fundamental niche for regulation of HSC migration to the liver. To our knowledge, this is the first report showing that LSEC not only express and secrete SDF-1, but also its receptor CXCR4.     

Trafficking of normal stem cells and metastasis of cancer stem cells involve similar mechanisms: pivotal role of the SDF-1-CXCR4 axis

The alpha-chemokine stromal-derived factor (SDF)-1 and the G-protein-coupled seven-span transmembrane receptor CXCR4 axis regulates the trafficking of various cell types. In this review, we present the concept that the SDF-1-CXCR4 axis is a master regulator of trafficking of both normal and cancer stem cells. Supporting this is growing evidence that SDF-1 plays a pivotal role in the regulation of trafficking of normal hematopoietic stem cells (HSCs) and their homing/retention in bone marrow. Moreover, functional CXCR4 is also expressed on nonhematopoietic tissue-committed stem/progenitor cells (TCSCs); hence, the SDF-1-CXCR4 axis emerges as a pivotal regulator of trafficking of various types of stem cells in the body. Furthermore, because most if not all malignancies originate in the stem/progenitor cell compartment, cancer stem cells also express CXCR4 on their surface and, as a result, the SDF-1-CXCR4 axis is also involved in directing their trafficking/metastasis to organs that highly express SDF-1 (e.g., lymph nodes, lungs, liver, and bones). Hence, we postulate that the metastasis of cancer stem cells and trafficking of normal stem cells involve similar mechanisms, and we discuss here the common molecular mechanisms involved in these processes. Finally, the responsiveness of CXCR4+ normal and malignant stem cells to an SDF-1 gradient may be regulated positively/primed by several small molecules related to inflammation which enhance incorporation of CXCR4 into membrane lipid rafts, or may be inhibited/blocked by small CXCR4 antagonist peptides. Consequently, strategies aimed at modulating the SDF-1-CXCR4 axis could have important clinical applications both in regenerative medicine to deliver normal stem cells to the tissues/organs and in clinical hematology/oncology to inhibit metastasis of cancer stem cells.     

Stromal Cell-derived Factor-1 and its Receptor CXCR4 are Upregulated Expression in Degenerated Intervertebral Discs

Background: Although chemokine stromal cell-derived factor 1 (SDF-1) and its receptor CXCR4 induce degradation of articular cartilage in rheumatoid arthritis (RA) and osteoarthritis (OA), the association between the SDF-1/CXCR4 pathway and degradation of the cartilaginous endplate and nucleus pulposus has not been thoroughly clarified. We investigated the expression of SDF-1 and CXCR4 in intervertebral discs (IVDs).Methods: SDF-1 and CXCR4 levels in human IVDs and the rat L5/6 motion segment were quantified by enzyme-linked immunosorbent assay. SDF-1 staining was quantified using a microscope and Image-Pro Plus software. Integrated optical density (IOD) served as the measurement parameter. The number of CXCR4 immunoreactive cells was expressed as a percentage of the total number of cells.Results: SDF-1 and CXCR4 were both expressed in IVDs, and the levels of SDF-1 and CXCR4 were both significantly higher in the degeneration group than in the normal group of human (or rat) discs. Both nucleus pulposus cells and cartilaginous endplate cells expressed the CXCR4 protein. Furthermore, a positive correlation was observed between the SDF-1 IOD value and the percentage of CXCR4-positive disc cells in the nucleus pulposus and cartilaginous endplate. The SDF-1 IOD values were significantly higher in the outer annular fibrosus and bone/endplate junction region than in the nucleus pulposus and cartilaginous endplate in the rat specimens.Conclusions: Our findings suggest upregulated expression of SDF-1 and its receptor CXCR4 in degenerated IVD.     

Endothelial cells and normal breast epithelial cells enhance invasion of breast carcinoma cells by CXCR-4-dependent up-regulation of urokinase-type plasminogen activator receptor (uPAR, CD87) expression

Here we show the increase of invasion of three breast cancer cell lines (8701-BC, MDA-MB-231 and SKBR3) upon long-term co-incubation with culture medium of normal microvascular endothelial cells (MVEC) and normal breast epithelial cells (HB2). The enhancement of invasion relied on the interaction of microvascular endothelial cell and normal breast epithelial cell CXCL12 (SDF1) chemokine, whose expression by breast cancer cells was very low, with the cognate CXCR4 receptor of malignant cells, which resulted in over-expression of the urokinase-type plasminogen activator receptor (uPAR) on their surfaces. uPAR over-expression, showed by RT-PCR and Western blotting, was paralleled by increased urokinase-type plasminogen activator (uPA) partitioning on the cell surface with respect to the fluid phase, as demonstrated by zymography. Long-term interaction of SDF1 with CXCR4 stimulated sustained activation of JNK phosphorylation. Blocking antibodies to CXCR4 were able to block the endothelial/epithelial cell-dependent enhancement of invasion, as well as to inhibit SDF1-CXCR4-dependent JNK phosphorylation and uPAR over-expression of malignant cells. We suggest that acquisition of the angiogenic phenotype by breast cancer cells triggers an amplification loop, in which endothelial cells and normal breast epithelial cells of the tumour cooperate to provide facilitated routes to cell invasion and metastasis and to enhance the aggressive phenotype of cancer cells.     

SDF-1/CXCR4轴通过调控间充质干细胞定向分化修复缺氧缺血性脑损伤

目的:探讨基质细胞衍生因子-1(stromalcell-derivedfactor-1,SDF-1)/CXC趋化因子受体4(CXCchemokinereceptor4,CXCR4)轴调控间充质干细胞定向分化、修复缺氧缺血性脑损伤的作用。方法:大鼠间充质干细胞(ratmesenchymalstemcells,rMSCs)经缺氧培养不同时点(0h、6h、12h、24h、48h、72h)或SDF-1α(10μg/L)孵育后,采用RT-PCR、Westernblotting和流式细胞术检测其表面CXCR4表达的变化;建立大鼠缺氧缺血性脑损伤模型,运用RT-PCR和Westernblotting检测造模后不同时点(1d、3d、5d、7d、14d、21d)大鼠脑部海马组织中SDF-1αmRNA转录和蛋白表达的改变情况;用AMD3100(CXCR4拮抗剂)拮抗rMSCs表面CXCR4后,免疫细胞化学和Westernblotting检测rMSCs诱导向神经细胞分化中神经元特异性烯醇化酶(neuron-specificenolase,NSE)和胶质纤维酸性蛋白(glialfibrillaryacidicprotein,GFAP)等神经细胞特异性标志物的阳性率及表达变化情况。结果:低氧培养6h及12h的rMSCsCXCR4mRNA及蛋白表达水平均较常氧培养组明显增加(P<0.01),10μg/LSDF-1α孵育后rMSCs的CXCR4表达水平明显增加(P<0.01);缺氧缺血性脑损伤模型的大鼠脑内SDF-1α蛋白表达显著增加(P<0.01);5mg/LAMD3100处理后的rMSCs在向神经细胞诱导分化中NSE和GFAP蛋白的表达明显减少。结论:微小剂量的SDF-1α可诱导低氧培养的rMSCs表面CXCR4的表达,而在缺氧缺血性脑损伤模型的大鼠脑内SDF-1α表达增加,从而使SDF-1/CXCR4轴的生物学效应得以增强;CXCR4拮抗的rMSCs在分化中神经细胞特异性标志物NSE和GFAP的表达降低,表明SDF-1/CXCR4轴在rMSCs定向神经分化修复缺血缺氧脑损伤中具有重要的调控作用。     

Vascular endothelial growth factor and basic fibroblast growth factor induce expression of CXCR4 on human endothelial cells: In vivo neovascularization induced by stromal-derived factor-1alpha

The contribution of chemokines toward angiogenesis is currently a focus of intensive investigation. Certain members of the CXC chemokine family can induce bovine capillary endothelial cell migration in vitro and corneal angiogenesis in vivo, and apparently act via binding to their receptors CXCR1 and CXCR2. We used an RNAse protection assay that permitted the simultaneous detection of mRNA for various CXC chemokine receptors in resting human umbilical vein endothelial cells (HUVECs) and detected low levels of only CXCR4 mRNA. Stimulation of HUVECs with vascular endothelial growth factor (VEGF) or basic fibroblast growth factor (bFGF) up-regulated levels of only CXCR4 mRNA. CXCR4 specifically binds the chemokine stromal-derived factor-1alpha (SDF-1alpha). Competitive binding studies using 125I-labeled SDF-1alpha with Scatchard analysis indicated that VEGF or bFGF induced an average number of approximately 16,600 CXCR4 molecules per endothelial cell, with a Kd = 1.23 x 10(-9) mol/L. These receptors were functional as HUVECs and human aorta endothelial cells (HAECs) migrated toward SDF-1alpha. Although SDF-1alpha-induced chemotaxis was inhibited by the addition of a neutralizing monoclonal CXCR4 antibody, endothelial chemotaxis toward VEGF was not altered; therefore, the angiogenic effect of VEGF is independent of SDF-1alpha. Furthermore, subcutaneous SDF-1alpha injections into mice induced formation of local small blood vessels that was accompanied by leukocytic infiltrates. To test whether these effects were dependent on circulating leukocytes, we successfully obtained SDF-1alpha-induced neovascularization from cross sections of leukocyte-free rat aorta. Taken together, our data indicate that SDF-1alpha acts as a potent chemoattractant for endothelial cells of different origins bearing CXCR4 and is a participant in angiogenesis that is regulated at the receptor level by VEGF and bFGF.     

SDF-1α/CXCR4轴通过诱导胰腺癌上皮-间充质转化促进肿瘤迁移和侵袭

目的:探讨SDF-1α/CXCR4轴对胰腺癌细胞迁移和侵袭能力的影响及其作用机制。方法:应用RT-qPCR检测4种胰腺癌细胞株CXCR4 mRNA的表达。Transwell实验检测外源性SDF-1α及其受体CXCR4靶向抑制剂AMD3100对胰腺癌细胞迁移和侵袭能力的影响。MTS法检测外源性SDF-1α及AMD3100对胰腺癌细胞活力的影响。Western blot法检测外源性SDF-1α及AMD3100对胰腺癌细胞上皮-间充质转化(EMT)相关标志物表达的影响。结果:(1) 4种胰腺癌细胞株均不同程度地表达CXCR4 mRNA,其中PANC-1细胞株表达量最高。(2)外源性SDF-1α可增强PANC-1细胞的迁移和侵袭能力,该作用可被AMD3100所阻断。(3)外源性SDF-1α处理PANC-1细胞72 h可增强细胞活力,该作用可被AMD3100阻断。(4)外源性SDF-1α通过上调SNAIL和TWIST促使PANC-1细胞发生EMT,该作用可被AMD3100所阻断。结论:SDF-1/CXCR4轴通过促进胰腺癌细胞发生EMT而促进肿瘤迁移和侵袭。