Human Langerhans' cells and dermal-type dendritic cells generated from CD34 stem cells express different toll-like receptors and secrete different cytokines in response to toll-like receptor ligands

Langerhans' cells (LC) and dermal dendritic cells (dDC) are located in the superficial and deeper layers of the skin respectively and represent the main dendritic cell (DC) populations of the skin. LC-like and dDC-like DC can be generated from CD34 stem cells and this system is widely used as a model for investigating these cells and in therapeutic vaccination. Here we report toll-like receptor (TLR) expression in human LC and dDC derived from CD34 stem cells. In vitro-generated DC expressed TLR-1, 2, 4, 6, 8 and 10. LC, but not dDC, expressed TLR-5, whereas only dDC expressed TLR-3. Maturation of LC was mediated by TLR-2, 4 and 5 ligands, but not by a TLR-3 ligand. dDC maturation was induced by TLR-3 and -4, but not with TLR-5 ligand and only weakly by a TLR-2 ligand. Stimulated LC secreted interleukin (IL)-1beta, low levels of tumour necrosis factor-alpha (TNF-alpha) and IL-8, but not IL-6 or IL-10. dDC secreted TNF-alpha, IL-6, IL-8 and IL-10, but little IL-1beta. IL-12p70 was not produced by ligand-stimulated dDC or LC, but was secreted by monocyte-derived DC (mdDC) stimulated with lipopolysaccharide (LPS). Thus, in vitro-generated LC and dDC detect different pathogen-associated molecules and show different cytokine-secretion profiles in response to TLR ligands.     

Mesenchymal stem cells decrease splenocytes apoptosis in a sepsis experimental model

Objective and design

Mesenchymal stem cells (MSCs) are potent modulators of immune responses. Sepsis is the association of a systemic inflammatory response with an infection. The aim of this study was to test the ability of MSCs derived from adipose tissue, which have immunomodulatory effects, and to inhibit the septic process in an experimental model of mice.

Methods

Three experimental groups (male C57BL/6 mice) were formed for the test: control group, untreated septic group and septic group treated with MSCs (1 × 10⁶ cells/animal).

Results

In the control group, there were no deaths; in the untreated septic group, the mortality rate was 100 % within 26 h; in the septic group treated with MSCs, the mortality rate reached 40 % within 26 h. The group treated with MSCs was able to reduce the markers of tissue damage in the liver and pancreas. The treated group had a reduction of inflammatory markers. Furthermore, the MSCs-treated group was able to inhibit the increase of apoptosis in splenocytes observed in the untreated septic group.

Conclusions

Our data showed that MSCs ameliorated the immune response with decrease of inflammatory cytokines and increase anti-inflammatory IL-10; moreover, inhibited splenocytes apoptosis and, consequently, inhibited tissue damage during sepsis.     

Mesenchymal stem cells derived from human adipose tissues favor tumor cell growth in vivo

Mesenchymal stem cells (MSCs) have generated a great deal of interest in clinical situations, due principally to their potential use in regenerative medicine and tissue engineering applications. However, the therapeutic application of MSCs remains limited, unless the favorable effects of MSCs for tumor growth in vivo and the long-term safety of the clinical applications of MSCs can be understood more thoroughly. In this study, MSCs derived from human adipose tissues (hASCs) together with tumor cells were transplanted subcutaneously or intracranially into BALB/c nude mice to observe tumor outgrowth. The results indicated that hASCs with H460 or U87MG cells promoted tumor growth in nude mice. Our histopathological analyses indicated that the co-injection of tumor cells with hASCs exerted no influence on the formation of intratumoral vessels. Co-culture of tumor cells with hASCs or the addition of conditioned medium (CM) from hASCs effected an increase in the proliferation of H460 or U87MG cells. Co-injection of hASCs with tumor cells effected an increase in tumor cell viability in vivo, and also induced a reduction in apoptotic cell death. CM from hASCs inhibited hydrogen peroxide-induced cell death in H460 or U87MG cells. These findings indicated that MSCs could favor tumor growth in vivo. Thus, it is necessary to conduct a study concerning the long-term safety of this technique before MSCs can be used as therapeutic tools in regenerative medicine and tissue engineering.     

Mesenchymal stem cells in tissue engineering

The repair of diseased or damaged cartilage remains one of the most challenging problems of musculoskeletal medicine. Tissue engineering advances in cartilage repair have utilized autologous and allogenic chondrocyte and cartilage grafts, biomaterial scaffolds, growth factors, stem cells, and genetic engineering. The mesenchymal stem cell has specifically attracted much attention because of its accessibility, potential for differentiation, and manipulability to modern molecular, tissue and genetic engineering techniques. Mesenchymal stem cells provide invaluable tools for the study of tissue repair when combined with a carrier vehicle/matrix scaffold, and/or bioactive growth factors. However, an underappreciated source of knowledge lies in the relationship between fetal development and adult tissue repair. The multitude of events that take place during fetal development which lead from stem cell to functional tissue are poorly understood. A more thorough understanding of the events of development as they pertain to cartilage organogenesis may help elucidate some of the unknowns of adult tissue repair.     

Mesenchymal stem cells in autoimmune disease

Autoimmune diseases afflict more than 3% of the U.S. population. Current therapy for mild to moderate cases is symptomatic, however advanced cases suffer high morbidity and mortality. Advanced patients have benefited from stem cell therapy in the form of bone marrow transplantation in conjunction with high-dose cytotoxic therapy. Broader application of stem cell therapy requires better understanding of how adult stem cells affect development and foster treatment of autoimmune pathologies, and of better ways to manipulate the host immune responses. While extensive research documents the role of hematopoietic stem cells (HSCs) in autoimmune disease, few studies have addressed if and how mesenchymal stem cells (MSCs) contribute to their etiopathology. Recent characterization of MSCs and their role in hematopoiesis and immune modulation suggest that their potential for cell therapy extends beyond their traditional accessory function in HSC engraftment. MSCs contribute significantly to tissue restructuring and immune functioning, in addition to facilitating durable, long-lasting stem cell engraftment. MSCs are relatively easy to obtain and expand in in vitro cultures, rendering them a prime candidate for genetic manipulations for stem cell therapy. They have the potential to differentiate into multiple lineages such as osteoblasts, adipose tissue, cartilage, tendon, and stromal cells. The role of MSCs for autoimmune disease therapy could thus be based both on immune function modulation and contribution to hematopoiesis. In this review, we examine the biology of MSCs, and their potential for cell therapy of autoimmune disease.     

树突状细胞Toll样受体介导的信号传导通路

树突状细胞(DC)是体内功能最强的抗原提呈细胞,是机体联系固有免疫应答和适应性免疫应答的桥梁.DC表面的Toll样受体(TLRs)在接受外界刺激信号和诱导机体产生免疫应答方面具有核心作用.TLRs介导的胞内信号传导通路主要有两条:髓样分化蛋白88(MyD88)依赖途径与MyD88非依赖途径.这两条传导通路中的大部分接头蛋白分子是一致的,但在某些关键点上又有所不同,因此决定了它们的功能既相互交叉又彼此独立.     

Mesenchymal stem cells enhance angiogenesis in mechanically viable prevascularized tissues via early matrix metalloproteinase upregulation

Angiogenesis, the sprouting of new blood vessels from existing vasculature, is a complex biological process of interest to both the treatment of numerous pathologies and the creation of thick engineered tissues. In the context of tissue engineering, one potential solution to the diffusion limitation is to create a vascular network in vitro that can subsequently anastomose with the host after implantation, allowing the implantation of thicker, more complex tissues. In this study, the ability of endothelial cells to sprout and form stable vascular networks in 3-dimensional (3D) fibrin matrices was investigated as a function of matrix density in a prevascularized tissue model. The results demonstrate that while increasing matrix density leads to a nearly 7-fold increase in compressive stiffness, vascular sprouting is virtually eliminated in the most dense matrix condition. However, the addition of human mesenchymal stem cells (HMSCs) to the denser matrices reverses this effect, resulting in an up to a 7-fold increase in network formation. Although the matrix metalloproteinases (MMPs) MMP-2, MMP-9, and MT1-MMP are all upregulated early on with the addition of HMSCs, MT1-MMP appears to play a particularly important role in the observed angiogenic response among these proteases. This study provides a means to design stiffer prevascularized tissues utilizing naturally derived substrates, and its results may yield new mechanistic insights into stem cell-based angiogenic therapies.     

三种来源于不同围产期组织间充质干细胞的比较

背景：围产期组织作为间充质干细胞的良好来源已经受到广泛关注。 目的：比较3种来源于不同围产期组织的间充质干细胞的特性。 方法：以“mesenchymal stem cells, fetal blood, umbilical cord, placenta”为检索词,检索PudMed数据库中2002年1月至2012年4月的文献,纳入与脐血、脐带、胎盘来源的间充质干细胞相关的权威性且具有代表性的文献。 结果与结论：计算机初检得到248篇文献,对其中16 篇文献进行综述。主要阐述3种来源于不同围产期组织的间充质干细胞的分离培养方法以及生物学特性等方面内容,相对于脐血、胎盘来源的间充质干细胞,来源于脐带华通胶质的间充质干细胞由于其分离方法简单,分离成功率高,扩增潜能较高,分化潜能较高以及致瘤性低而成为相对优越的间充质干细胞来源。     

不同组织来源间充质干细胞向血管内皮细胞诱导分化的研究与进展

文章快速阅读：文题释义： 血管内皮细胞：亦称内皮细胞,是胚胎发育最早分化的组织,且属于中胚层,通常指在心、血管和淋巴管内表面的单层扁平上皮,其形成血管的内壁。血管内皮细胞具有吞噬异物、细菌和衰老及坏死组织的功能,还参与免疫活动。理论上间充质干细胞可以分化为血管内皮细胞。 诱导间充质干细胞分化为内皮细胞的主要因素：细胞分化是由许多分子信号通路调控的复杂过程,在细胞分化早期干细胞通过旁分泌作用释放信号蛋白分子,如血小板衍生生长因子、血管内皮生长因子A、转化生长因子β和Wnts等,刺激距离较远的细胞内的信号级联,使细胞定向分化。 摘要 背景：间充质干细胞可诱导分化为血管内皮细胞,在体外可形成血管移植物用于临床治疗缺血性疾病、血管组织工程及再生医学等领域。 目的：总结间充质干细胞的生物学特征,不同组织来源间充质干细胞诱导分化为血管内皮细胞的相关研究进展。 方法：文章由第一作者于2015年9至11月期间检索PubMed、Sciencedirect、Medline数据库2000年1月至2015年6月的相关文章,限定文章语言种类为English,检索词为“mesenchymal stem cells,vascular endothelial cell,cell differentiation”,初检文章156篇,筛选后对51篇文章进行分类总结。 结果与结论：①间充质干细胞来源广泛,其中骨髓来源研究最早最多,但随供者年龄增大,其增殖及分化能力随之减弱;②脐带、胎盘、羊膜都是产妇自体产物且分娩后会遗弃,故获取间充质干细胞比较容易,同时对新生儿和产妇不会造成任何痛苦和心理负担,受者更易于接受,不会引起道德伦理及法律方面的纠纷;③脐血来源充足,免疫原性较弱,且受胎盘屏障保护使其成分被病毒、细菌污染的概率低;④羊水因取材方式等限制其在临床应用。 中国组织工程研究杂志出版内容重点：干细胞;骨髓干细胞;造血干细胞;脂肪干细胞;肿瘤干细胞;胚胎干细胞;脐带脐血干细胞;干细胞诱导;干细胞分化;组织工程 ORCID: 0000-0002-2983-8929(陈国华)     

Maternal neutrophil toll-like receptor mRNA expression is down-regulated in preeclampsia

Citation
Nitsche JF, Jiang S‐W, Brost BC. Maternal neutrophil Toll‐like receptor mRNA expression is down‐regulated in preeclampsia. Am J Reprod Immunol 2011; 66: 242–248 Problem There are many immunological changes in preeclampsia. For example, TLR‐4 expression is increased in the placenta during preeclampsia. However, data on TLR expression in other tissues during preeclampsia are lacking. This study aimed to determine whether TLR mRNA expression in maternal neutrophils is altered in preeclampsia. Method of Study A case–control study using standard quantitative real‐time PCR techniques was performed to assess TLR‐2 and TLR‐4 mRNA expression in 12 patients with mild preeclampsia and 18 normal pregnant controls at similar gestational ages. Results Compared to normal pregnant controls, there was a significant decrease in TLR‐2 and TLR‐4 expression in women with mild preeclampsia. Conclusion TLR‐2 and TLR‐4 expression in maternal neutrophils is decreased in preeclampsia. Given the many immunological changes in preeclampsia, this may represent an adaptation to the increased inflammatory signals present in preeclampsia. Further study is needed to clarify the role of the TLR in preeclampsia.     

间充质干细胞在炎症免疫调节中的作用及应用进展

背景：研究表明,间充质干细胞的主要功能有直接参与损伤修复,分泌生长因子,调节免疫和炎症,抗氧化应激等,可用于治疗多种急、慢性疾病。 目的：综述间充质干细胞在炎症免疫调节中的研究进展。 方法：以“干细胞,间充质干细胞,免疫调节,炎症,免疫细胞,炎症因子,治疗”为中文检索词,以“stem cells,mesenchymal stem cells,immune regulation,inflammation,immune cells,inflammatory factors,treatment”为英文检索词,在万方、中国知网期刊全文数据库和PubMed数据库检索2005年1月至2015年8月有关干细胞参与炎症免疫调节的文献。排除陈旧性和重复性研究,最终纳入40篇文献进行综述。 结果与结论：间充质干细胞因其具有免疫调节及多向分化的特性,越来越受到临床的重视,同时间充质干细胞易于从不同组织中获取并且具有良好的体外扩增能力,使得其在组织损伤炎症与修复的临床运用中具有广阔的前景。作为最有希望进入临床应用阶段的种子细胞,间充质干细胞在许多疾病的治疗中表现出了它的优越性,特别是对于免疫调节失衡导致的炎症性相关疾病的治疗中表现出良好的效果。相信在未来的细胞生物治疗领域,间充质干细胞将占有重要的地位。 中国组织工程研究杂志出版内容重点：干细胞;骨髓干细胞;造血干细胞;脂肪干细胞;肿瘤干细胞;胚胎干细胞;脐带脐血干细胞;干细胞诱导;干细胞分化;组织工程     

Rotavirus and coxsackievirus infection activated different profiles of toll-like receptors and chemokines in intestinal epithelial cells

Objective  To understand the inflammatory-immune response in intestinal epithelial cells after infection of rotavirus and coxsackievirus B3. Methods  We examined by quantitative PCR the expression profiles of genes encoding five toll-like receptors (TLR) and levels of three chemokines in response to rotavirus and coxsackievirus B3 infection in a human intestinal epithelial cell line (HT-29 cells). Results  We demonstrated that rotavirus induced significantly increased levels of mRNA expression for TLR2, TLR3, TLR7 and TLR8 in HT-29 cells in a time-dependent manner. In contrast, coxsackievirus B3 did not stimulate mRNA expression for TLR3. Rotavirus and coxsackievirus B3 also induced higher levels of mRNA expression for RANTES, IP-10 and IL-8 during the period of infection in a different manner. Finally, significantly elevated levels of RANTES, IP-10 and IL-8 were detected by ELISA in rotavirus-infected cells from 24 to 48 h. Conclusion  Our findings suggest that different patterns of TLRs and chemokines were induced in the initiation and modulation of immune response to rotavirus and coxsackievirus B3 infection.     

慢性乙型肝炎患者外周血单个核细胞Toll样受体3的表达降低

目的 探讨慢性乙型肝炎患者外周血单个核细胞Toll样受体3(TLR3)的表达及其临床意义.方法 分别采集慢性乙型肝炎患者和健康志愿者外周血,荧光定量PCR法检测血清HBV DNA复制水平;使用RT-PCR、流式细胞术以及免疫印迹技术分别检测外周血单个核细胞TLR3的mRNA、蛋白的表达;使用ELISA法检测血清中肿瘤坏死因子α(TNF-α)和干扰素β(IFN-p)水平.结果 慢性乙型肝炎患者外周血单个核细胞中的TLR3表达显著低于健康志愿者,且降低水平与血清HBV DNA复制水平相关;慢性乙型肝炎患者外周血TNF-α、IFN-β浓度显著低于健康志愿者,且降低的水平与血清HBV DNA复制水平相关.结论 慢性乙型肝炎患者外周血单个核细胞TLR3的表达与乙肝病毒的复制水平相关.     

含不同启动子的GFP载体在不同种属的胚胎干细胞系中表达效率的研究

目的 对比含不同启动子的GFP表达载体在hESC及小鼠胚胎干细胞(mouse embryonic stem cell, mESC)的表达效率,为探索ESC及其衍生细胞移植在体内的存活、迁移、分化及整合提供细胞模型.方法 阳离子脂质体转染hESC及mESC ES-D3,荧光显微镜下观察阳性克隆,流式细胞术(flow cytometry, FCM)计算不同载体pCX-hrGFP、pIRES-hrGFP在不同种属细胞中的表达效率.结果 两种载体在mESC的表达效率分别为pCX-hrGFP 90±2.5%, pIRES-hrGFP 0.67±0.02%, 两组间比较差异有统计学意义(P＜0.05).在hESC的表达效率分别为pCX-hrGFP 0.8±0.1%, pIRES-hrGFP 0.62±0.08%,两组间比较有统计学差异(P＜0.05).pCX-hrGFP在mESC及hESC间的表达效率差异有统计学意义 (P＜0.01),pIRES-hrGFP在mESC及hESC间的表达效率差异无统计学意义 (P＞0.05).结论 (1)CBA启动子引导的GFP表达效率高于CMV启动子.(2)同一载体在不同种属细胞内表达效率不同.     

Mesenchymal stem cells from different organs are characterized by distinct topographic Hox codes

Mesenchymal stem cells (MSC) are multipotent cells found as part of the stromal compartment of the bone marrow and in many other organs. They can be identified in vitro as CFU-F (colony forming unit-fibroblast) based on their ability to form adherent colonies of fibroblast-like cells in culture. MSC expanded in vitro retain characteristics appropriate to their tissue of origin. This is reflected in their propensity for differentiating towards specific lineages, and their capacity to generate, upon retransplantation in vivo, a stroma supporting typical lineages of hematopoietic cells. Hox genes encode master regulators of regional specification and organ development in the embryo and are widely expressed in the adult. We investigated whether they could be involved in determining tissue-specific properties of MSC. Hox gene expression profiles of individual CFU-F colonies derived from various organs and anatomical locations were generated, and the relatedness between these profiles was determined using hierarchical cluster analysis. This revealed that CFU-F have characteristic Hox expression signatures that are heterogeneous but highly specific for their anatomical origin. The topographic specificity of these Hox codes is maintained during differentiation, suggesting that they are an intrinsic property of MSC. Analysis of Hox codes of CFU-F from vertebral bone marrow suggests that MSC originate over a large part of the anterioposterior axis, but may not originate from prevertebral mesenchyme. These data are consistent with a role for Hox proteins in specifying cellular identity of MSC.     

GPI-specific phospholipase D mRNA expression in tumor cells of different malignancy

Glycosylphosphatidylinositol-specific phospholipase D (GPI-PLD) is a highly specific enzyme whose only known substrate is the GPI anchor of cell surface proteins. GPI-PLD measurements, however, are technically difficult since the enzyme is expressed at low levels in cells and tissues, and serum contains large amounts of inactive, latent GPI-PLD interfering with protein-based assays. We have therefore developed a semi-quantitative RT-PCR method to measure mRNA expression of all known GPI-PLD isoforms in cells and tissues. In human ovarian cancer cell lines, GPI-PLD mRNA expression correlated with GPI-PLD enzyme activity and with the shedding of the GPI-anchored tumor and prognostic markers, urokinase receptor and CA125, from the cell surface. This supports a potential role for this enzyme in the generation of circulating prognostic markers in malignant tumors. Similarly, in human epithelial cells of the skin, GPI-PLD mRNA expression increased with tumor progression. Whereas normal keratinocytes did not express significant amounts of GPI-PLD mRNA, expression was dramatically induced by serum in immortalized HaCaT keratinocytes and constitutively high and independent of serum in tumorigenic A431 epidermoid carcinoma cells. In addition, GPI-PLD expression was significantly increased in highly malignant, H-ras-transfected murine bladder carcinoma cells as compared to the low malignant, non-transfected parental cells. The competitive RT-PCR described here represents the first quantitative assay specific for cellular GPI-PLD isoforms, and our in vitro analyses suggest that GPI-PLD expression might be associated with tumor malignancy. This revised version was published online in July 2006 with corrections to the Cover Date.