Ischaemia/reperfusion induced cardiac stem cell homing to the injured myocardium by stimulating stem cell factor expression via NF-κB pathway

Ischaemia/reperfusion (I/R) is a major cause of heart failure. Recently cardiac stem cells (CSCs) were proposed as the most appropriate cell type for heart disease therapy. However, it is still unclear whether I/R can stimulate the CSCs homing to the injured myocardium. Male Sprague–Dawley rats were subjected to a 30-min ischaemia followed by reperfusion of different intervals. RT-PCR, western blotting and immunohistochemistry were performed to detect stem cell factor (SCF) expression at mRNA and protein levels respectively. Activation of nuclear factor-κB (NF-κB) was determined by electrophoretic mobility shift assay. To assess the homing of CSCs in vivo , BrdU-labelled CSCs were injected into AV-groove before induction of ischaemia and examined by immunofluorescent staining in the injured myocardium after I/R. From day 3 to day 6 after reperfusion, the accumulation of CSCs was significantly elevated in the injured area, which was matched with the increased SCF expression during I/R. Pretreatment of rats with NF-κB inhibitor, N -acetyl- l -cysteine (NAC) not only suppressed NF-κB activation induced by I/R but also attenuated SCF expression. Further analysis revealed that I/R induced phosphorylation of IκBα after 15 min of reperfusion, and the raised phosphor-IκBα returned to the basal level at 2 h of reperfusion. In simulated I/R(SI/R) in vitro , it enhanced NF-κB activation and SCF expression in cultured neonatal rat cardiomyocytes, which was markedly inhibited by NF-κB decoy oligodeoxynucleotide or NAC. Taken together, our results demonstrated that I/R induced CSCs homing to the injured myocardium by stimulating myocardial SCF expression via activation of NF-κB.     

Monocyte chemotactic protein 1 increases homing of mesenchymal stem cell to injured myocardium and neovascularization following myocardial infarction

Objective:To investigate the effect of MCP-1 on mesenchymal stem cells(MSCs) homing to injured myocardium in a rat myocardial infarction(MI) model. Methods:Rat myocardial infarction model was established by permanent left anterior descending branch ligation. Mesenchymal stem cells from donor rats were cultured in IMDM and labeled with BrdU. The Rats were divided into two groups. Monocyte chemotactic protein 1(MCP-1) expression were measured by in situ hybridization and immunohistochemistry in the sham operated or infarcted hearts at 1,2, 4,7,14 and 28 days post operation in MCP-1 detection group. The rats were injected with MCP-1, anti-MCP-1 antibody or saline 4 days after myocardial infarction in intervention group. Then, a total of 5 × 106 cells in 2.5 ml of PBS were injected through the tail vein. The number of the labeled MSCs in the infarcted hearts was counted 3 days post injection. Cardiac function and blood vessel density were assessed 28 days post injection. Results:Self-generating MCP-1 expression was increased at the first day, peaked at the 7th day and decreased thereafter post MI and remained unchanged in sham operated hearts. The MSCs enrichment in the host hearts were more abundant in the MI groups than that in the non-Mi group(P = 0.000), the MSCs enrichment in the host hearts were more abundant in the MCP-1 injected group than that in the anti-MCP-1 antibody and saline injected groups (P = 0.000). Cardiac function was improved more in MCP-1 injected group than anti-MCP-1 antibody and saline injected groups(P= 0.000). Neovascularization in MCP-1 injected group significantly increased compared with that of other groups(P = 0.000). Conclusion: Myocardial MCP-1 expression was increased only in the early phase post MI. MCP-1 may enhance MSCs homing to the injured heart and improve cardiac function by promoting neovascularization.     

干细胞治疗修复心肌的机制研究进展

干细胞移植治疗心肌梗死为临床治疗心梗提供了诱人的前景.关于其治疗机制的研究不断取得进展,目前已经历了细胞分化,促血管生成等阶段.机制的研究也为治疗提供了新的线索.本文就干细胞移植治疗心梗的机制作一综述.     

大鼠急性心肌梗死后骨髓干细胞归巢于缺血心肌的时间窗研究

目的：初探心肌梗死后骨髓干细胞归巢于心肌组织的时间窗。 方法： 110只大鼠随机分为4组，GM-CSF干预组(n=40)皮下注射GM-CSF (50 μg/kg/d) 5 d，对照组(n=40)皮下注射同等剂量生理盐水，GM-CSF干预假手术组(n=15)，对照假手术组(n=15)，结扎左冠状动脉前降支复制急性心肌梗塞动物模型，同时复制假手术模型。各组在心梗后1 、3 、5 、10 d免疫组化法观察心肌组织中归巢的ckit+细胞数量，28 d测定血压、室内压及±dp/dt值变化，观察两组大鼠瘢痕组织修复差异。 结果： ①心梗后28 d GM-CSF干预组心功能显著高于对照组(P<0.05)；②GM-CSF干预组瘢痕组织横径显著高于对照组(P<0.05)，梗死区平均心肌面积显著高于对照组(P<0.05)；③GM-CSF干预组与对照组1，3，5 d均有ckit+细胞归巢现象，以第5 d最为密集，第10 d未再有新归巢的ckit+细胞，GM-CSF干预组各时点ckit+细胞归巢数量显著高于对照组(P<0.05)。 结论： 骨髓干细胞归巢的时间窗为心梗后10 d以内。     

Electron-microscopic autoradiography of RNA synthesis in the injured myocardium

A burn of the wall of the left ventricle was produced in newborn rats. Synthesis of RNA in the muscle cells of the heart at a distance from the site of the burn was investigated by electron-microscopic autoradiography 24 h after injury. The tissue was fixed 2 and 6 h after injection of uridine-³H. The density of distribution of silver grains above the nucleus and cytoplasm of the cardiomyocytes was lower in the experimental animals than in the controls.Department of Pathological Anatomy, A. V. Vishnevskii Institute of Surgery, Academy of Medical Sciences of the USSR, Moscow. (Presented by Academician of the Academy of Medical Sciences of the USSR A. V. Smol'yannikov.) Translated from Byulleten' Éksperimental'noi Biologii i Meditsiny, Vol. 83, No. 6, pp. 751–753, June, 1977.     

骨髓干细胞动员对大鼠缺血心肌的治疗作用

目的：了解干细胞动员剂动员的骨髓干细胞的心肌细胞分化潜能及对缺血心肌的影响。方法：用异丙肾上腺，素(ISO)制作大鼠心肌梗死模型，联用粒细胞集落刺激因子(C-CSV)和辛伐他汀动员大鼠自体骨髓干细胞释放和迁移至心肌梗死区，用ISO后24小时、4周杀死大鼠，取出心脏，通过免疫组化、HE染色和VG染色方法观察大鼠心梗灶的CD34^ 细胞的浸润及心肌血管再生、心肌纤维化的情况。结果：用ISO后24小时，动员组大鼠心梗区可见CD34^ 细胞浸润，并有CD34^ 的新生心肌细胞生长，4周时瘢痕组织少，心肌纤维化程度轻，缺血心肌基本结构得到保护。结论：急性心梗发生后，联用粒细胞集落刺激因子(G-CSF)和辛伐他汀能迅速动员自体骨髓干细胞向心梗区内迁移、存活和向心肌细胞、血管内皮细胞等分化；并能抑制缺血心肌纤维化和保护缺血心肌基本结构。     

Repair of infarcted myocardium using mesenchymal stem cell seeded small intestinal submucosa in rabbits

Myocardial infarction (MI) remains a common and deadly disease. Using tissue-engineered cardiac grafts to repair infarcted myocrdium is considered to be a therapeutic approach. This study tested the feasibility of using MSCs-seeded SIS to repair chronic myocardial infarction in a rabbit model. MI in rabbits was created by ligation of the left anterior descending artery. BrdU-labeled mesenchymal stem cells (MSCs) were seeded on the small intestinal submucosa and cultured for 5–7 days prior to implantation. Four weeks after myocardial infarction, cardiac grafts were implanted onto the epicardial surface of infarcted myocardium. Four weeks after implantation of the membranes, a serial of tests including echocardiography, hemodynamics, histology and immunohistochemistry were undertaken to evaluate the effect of the implanted grafts on recovery of the infarcted myocardium. It was shown that left ventricular contractile function and dimension, the capillary density of the infarcted region, and myocardial pathological changes were significantly improved in rabbits implanted either SIS or MSCs-seeded SIS. But the MSCs-seeded SIS was more effective. Immunofluorescence staining demonstrated the migration of Brdu-labeled MSCs from the membrane into the infarcted area and their differentiation to cardiomyocytes and smooth muscle cells. Taken together, these results suggest that MSCs-seeded SIS can be used to repair chronic myocardial infarction, which enhances myocardial regeneration.     

Autologous stem cell transplantation in acute lymphocytic leukemia.

Autologous stem cell transplantation (ASCT) as well as allogeneic stem cell transplantation and conventional chemotherapy (CT) are less effective at treating acute lymphocytic leukemia (ALL) than acute myelocytic leukemia (AML). Chemoresistance and late relapses are hallmarks of ALL. In this context, the question of whether ASCT is superior to CT remains unanswered. In vitro marrow purging using monoclonal antibodies is not routinely used. This review summarizes the results of ASCT for adult and childhood ALL. Statistics from the European Group for Blood and Marrow Transplantation reveal a transplant-related mortality at 5 years of 11% +/- 1%, a relapse incidence of 60% +/- 2%, and a leukemia-free survival (LFS) and overall survival (OS) of 36% +/- 2% and 42% +/- 2%, respectively in 1,366 adults autografted in first remission (CR1). In 269 children, the LFS and OS were 50% +/- 3% and 54% +/- 3%, respectively. There was no evidence in favor of purging the autograft in vitro. In contrast, multicentric and single-institution studies have found better results in adults autografted in CR1, with LFS at 5 years from 46% to 64%, possible efficacy of marrow in vitro purging with mafosfamide (LFS 52%), and improvement in outcome with additional measures post-ASCT, such as maintenance chemotherapy (LFS 57%). Further, as already observed for AML, analyses by risk groups suggest that ASCT may essentially benefit good- but not poor-risk patients. For patients with the Ph1/bcr-abl translocation, the role of STI571 anti-tyrosine kinase for in vivo purging before stem cell harvesting is being investigated.     

Baculovirus-transduced,VEGF-expressing adipose-derived stem cell sheet for the treatment of myocardium infarction

Cell sheet technology has been widely employed for the treatment of myocardial infarction (MI), but cell sheet fabrication generally requires the use of thermo-responsive dishes. Here we developed a method for the preparation of adipose-derived stem cell (ASC) sheet that obviated the need of thermo-responsive dishes. This method only required the seeding of rabbit ASC onto 6-well plates at an appropriate cell density and culture in appropriate medium, and the cells were able to develop into ASC sheet in 2 days. The ASC sheet allowed for transduction with the hybrid baculovirus at efficiencies >97%, conferring robust and prolonged (>35 days) overexpression of vascular endothelial growth factor (VEGF). The ASC sheet was easily detached by brief (10 s) trypsinization and saline wash, while retaining the extracellular matrix and desired physical properties. The ASC sheet formation and VEGF expression promoted cell survival under hypoxia in vitro. Epicardial implantation of the VEGF-expressing ASC sheet to rabbit MI models reduced the infarct size and improved cardiac functions to non-diseased levels, as judged from the left ventrical ejection fraction/myocardial perfusion. The VEGF-expressing ASC sheet also effectively prevented myocardial wall thinning, suppressed myocardium fibrosis and enhanced blood vessel formation. These data implicated the potential of this method for the preparation of genetically engineered ASC sheet and future MI treatment.     

Mesenchymal stem cell transplantation accelerates hearing recovery through the repair of injured cochlear fibrocytes

Although demyelination is a cardinal feature in multiple sclerosis, axonal injury also occurs. We tested whether a delay in axonal degeneration could affect the disease severity in two models for multiple sclerosis: experimental autoimmune encephalomyelitis (EAE) and Theiler's murine encephalomyelitis virus (TMEV) infection. We compared wild-type C57BL/6 (B6) mice with C57BL/Wld(s) (Wld) mice, which carry a mutation that delays axonal degeneration. In EAE, both mouse strains were sensitized with myelin oligodendrocyte glycoprotein (MOG)(35-55) peptide and showed a similar disease onset, MOG-specific lymphoproliferative responses, and inflammation during the acute stage of EAE. However, during the chronic stage, B6 mice continued to show paralysis with a greater extent of axonal damage, demyelination, and MOG-specific lymphoproliferative responses compared with Wld mice, which showed complete recovery. In TMEV infection, only Wld mice were paralyzed and had increased inflammation, virus antigen-positive cells, and TMEV-specific lymphoproliferative responses versus infected B6 mice. Because TMEV can use axons to disseminate in the brain, axonal degeneration in B6 mice might be a beneficial mechanism that limits the virus spread, whereas slow axonal degeneration in Wld mice could favor virus spread. Therefore, axonal degeneration plays contrasting roles (beneficial versus detrimental) depending on the initiator driving the disease.     

The cell of origin and the leukemia stem cell in acute myeloid leukemia

There is experimental and observational evidence that the cells of the leukemic clone in acute myeloid leukemia (AML) have different phenotypes even though they share the same somatic mutations. The organization of the malignant clone in AML has many similarities to normal hematopoiesis, with leukemia stem cells (LSCs) that sustain leukemia and give rise to more differentiated cells. LSCs, similar to normal hematopoietic stem cells (HSCs), are those cells that are able to give rise to a new leukemic clone when transplanted into a recipient. The cell of origin of leukemia (COL) is defined as the normal cell that is able to transform into a leukemia cell. Current evidence suggests that the COL is distinct from the LSC. Here, we will review the current knowledge about LSCs and the COL in AML.     

Immunogenetics of hematopoietic stem cell transplantation: the contribution of microsatellite polymorphism studies

Polymorphisms of short tandem repeats of <10 nucleotides, or microsatellites (Msat), are largely used for post-transplant chimerism analyses in clinical hematopoietic stem cell transplantation (HSCT). Compared to single nucleotide polymorphisms (SNP), they have the advantage of a higher degree of allelic polymorphism and thus a potentially larger degree of informativity. Msat markers contribute to approximately 3% of the human genome and have been highly informative in disease association studies, population genetics, forensic medicine and organ and HSC transplantation. They allowed to expand our knowledge of the haplotypic structure of the HLA complex, including the noncoding sequences in the MHC, and to reach a better characterization of immunological phenotypes. Among the different immunogenetic studies in HSCT patients reviewed here, four Msat loci linked to cytokine genes have been analysed by a number of laboratories as potential candidates markers for HSCT outcome: IFNG, TNFd, IL-10(-1064) and IL-1RN. The low patient numbers and high diversity of clinical parameters account for some heterogeneity of the results. Among the trends starting to emerge from these studies, specific TNFd Msat alleles seem to be associated with acute graft-versus-host disease and mortality. Patient/donor Msat incompatibilities have also been used as surrogate markers to map biologically relevant polymorphisms, with a main focus on MHC-resident genetic variation. High throughput SNP typing and next-generation sequencing technologies will allow acquisition of large-scale genomic data and should allow refined analyses of clinically relevant genotypes in the transplantation settting, although the heterogeneity of the study cohorts will remain an issue. The analysis of Msat polymorphisms may still have a place in functional studies on the impact of Msat diversity in the control of immune response gene expression.     

Myocyte reactions at the borders of injured and healing rat myocardium

To better understand the apparent tendency of myocardium to heal by scarring rather than by restoration of normal structure, we have examined by light and electron microscopy the basic reactions of rat ventricular myocytes and their interactions with the extracellular matrix. Using three different types of necrotizing injuries, we found that necrotic myocytes separated from viable myocytes at intercalated discs leaving blunt-ended stumps at the edge of each lesion; the basal lamina of necrotic myocytes remained largely intact and spanned the gap between viable myocytes on opposite sides of each lesion. A small number of stumps were capped off by a new layer of basal lamina and showed no evidence of proliferative activity. The majority of the stumps developed cell processes that extended along the acellular myocyte basal lamina sheaths. These processes had one of two different fates. Some became apposed to similar processes, formed intercalated disc attachments, increased myofibrillar mass, and appeared to be associated with muscle reconstruction. Others developed elongate tapered ends, which terminated in myotendinous connections to scar tissue. The outcome of healing necrotic myocardium, like the healing of noncardiac necrotic tissue injuries, appears to be a function of cell growth and extracellular framework guidance; however, unlike healing of noncardiac tissues, healing of myocardium is uniquely complicated by continuing muscle contractions. We think the rhythmic pulling at the edges of necrotic lesions induces formation of myotendinous attachments, which anchor myocytes to scar tissue and probably prevent further growth.     

Three-dimensional scaffolding to investigate neuronal derivatives of human embryonic stem cells

Access to unlimited numbers of live human neurons derived from stem cells offers unique opportunities for in vitro modeling of neural development, disease-related cellular phenotypes, and drug testing and discovery. However, to develop informative cellular in vitro assays, it is important to consider the relevant in vivo environment of neural tissues. Biomimetic 3D scaffolds are tools to culture human neurons under defined mechanical and physico-chemical properties providing an interconnected porous structure that may potentially enable a higher or more complex organization than traditional two-dimensional monolayer conditions. It is known that even minor variations in the internal geometry and mechanical properties of 3D scaffolds can impact cell behavior including survival, growth, and cell fate choice. In this report, we describe the design and engineering of 3D synthetic polyethylene glycol (PEG)-based and biodegradable gelatin-based scaffolds generated by a free form fabrication technique with precise internal geometry and elastic stiffnesses. We show that human neurons, derived from human embryonic stem (hESC) cells, are able to adhere to these scaffolds and form organoid structures that extend in three dimensions as demonstrated by confocal and electron microscopy. Future refinements of scaffold structure, size and surface chemistries may facilitate long term experiments and designing clinically applicable bioassays.     

Calpains and cathepsin D of the injured myocardium: Role of iodothyronines

Department of Biological Chemistry, Academician I. P. Pavlov Ryazan Medical Institute (Presented by Academician of the Russian Academy of Medical Sciences P. V. Sergeev.) Translated from Byulleten' Éksperimental'noi Biologii i Meditsiny, Vol. 114, No. 12, pp. 596–598, December, 1992.     

干细胞移植在急性肾损伤中的应用及研究进展

背景：干细胞移植治疗急性肾损伤是近年来研究的热点,不同来源的干细胞在治疗急性肾损伤方面都取得了很大的进展。 目的：对干细胞生物学特性、干细胞的临床研究、不同来源的干细胞治疗急性肾损伤的实验性研究、存在问题及前景进行综述。 方法：应用计算机检索中国学术期刊全文数据库(CNKI)和Pubmed 数据库2001-01/2012-02 关于干细胞移植治疗急性肾损伤的文章,检索主题词“干细胞,移植,肾脏疾病,急性肾损伤”或“stem cell,transplantation,kidney disease,acute kidney injury”。初检索到205 篇文献,据纳入标准保留41篇进行分析、综述。 结果与结论：干细胞移植是一种尝试用于急性肾损伤治疗的新方法,可以改善肾功能的损伤,加快肾脏修复。虽然仍存在不少有待解决的问题,但干细胞移植仍以其传统方法无法比拟的优势在急性肾损伤领域展现了诱人的应用前景。     

急性冬眠心肌的能量代谢特点

目的：探索冬眠心肌的能量代谢特点。方法：用悬浮人红血球的ＫＨ液灌流大鼠离体等容收缩心脏，建立急性冬眠心肌模型，以心肌三磷酸腺苷、磷酸肌酸、糖原含量，以及心肌摄取乳酸的量作为代谢指标。结果：（１）冬眠３０ｍｉｎ组心肌三磷酸腺苷（ＡＴＰ）、磷酸肌酸及糖原含量均明显低于平衡末组（Ｐ＜００５），心肌由正常情况下摄取乳酸转为释放乳酸（Ｐ＜００５），在冬眠９０ｍｉｎ组，以上指标与冬眠３０ｍｉｎ组无显著差异；（２）再灌注３０ｍｉｎ，心肌磷酸肌酸和糖原含量及心肌摄取乳酸的量与平衡末组相似（Ｐ＞００５），但心肌三磷酸腺苷含量仍较平衡末组为低（Ｐ＜００５）。结论：急性冬眠期间，心肌对氧及代谢底物的供／均需比例在低水平上保持平衡，保证了心肌的存活