Therapeutic potential of circulating endothelial cells

The discovery of circulating endothelial progenitors (EPCs) in human blood has completely modified the concept of post natal angiogenesis. Many studies have further confirmed the existence of EPCs, their medullar origin and capacity to be integrated in vascular structures at the sites of neoangiogenesis. They suggest that EPCs could be naturally involved in the prevention of ischemia by participating directly to the formation of collateral vessels. These cells have a high tropism for the sites of angiogenesis. They may thus be used as a powerful tool to prevent or treat ischemic diseases and constitute an alternative to the risky local injection of pro-angiogenic growth factors. The efficiency of bone marrow cells autologous transplantation was recently proved by the first clinical trials. Bone marrow mononuclear cells locally injected to patients with limb or cardiac ischemia have been shown to improve reperfusion in ischemic tissues. These trials are still preliminary: they were performed with heterogeneous cells only on few patients and were not randomized. However they raise important questions, essentially on the necessity of cell purification prior to injection and on the nature of the bone marrow cells which are really involved in ischemic tissue regeneration. Further investigations are then required to improve the cell therapy outcome by 1) using more defined cell population, 2) clearly demonstrating the long term improvement of vascular function and 3) performing extensive analysis of the possible side effects.     

Mesenchymal stem cells in ischemic tissue regeneration

Diseases caused by ischemia are one of the leading causes of death in the world. Current therapies for treating acute myocardial infarction, ischemic stroke, and critical limb ischemia do not complete recovery. Regenerative therapies opens new therapeutic strategy in the treatment of ischemic disorders. Mesenchymal stem cells(MSCs) are the most promising option in the field of cell-based therapies, due to their secretory and immunomodulatory abilities, that contribute to ease inflammation and pr...     

Influence of temperature on ischemic brain: basic and clinical principles

In the last decades, the interest in the association between body temperature and stroke outcome has reemerged, and the use of animal models has made it possible to know the underlying pathogenic mechanisms involved, most of them with pending confirmation in human clinics. In this work, we will review the effects of hyperthermia and hypothermia and its pathogenesis on ischemic stroke, and the evidence of the efficacy and safety of anti-hyperthermic and hypothermic treatments. We will describe how treatment of hyperthermia on ischemic stroke patients, improves patient comfort and outcome, both in the short and the long term, but new clinical studies are needed in this field. Despite the theoretical and experimental bases in favor of hypothermia for the treatment of brain ischemia, there is no definitive clinical evidence that has proved its benefits, so far. With current knowledge, an objective of a body temperature between 35.5 and 36.5 °C seems an optimal therapeutic target for both hyperthermic and normothermic patients.     

AAV-mediated gene therapy in mouse models of recessive retinal degeneration

In recent years, more and more mutant genes that cause retinal diseases have been detected. At the same time, many naturally occurring mouse models of retinal degeneration have also been found, which show similar changes to human retinal diseases. These, together with improved viral vector quality allow more and more traditionally incurable inherited retinal disorders to become potential candidates for gene therapy. Currently, the most common vehicle to deliver the therapeutic gene into target retinal cells is the adenoassociated viral vector (AAV). Following delivery to the immuno-privileged subretinal space, AAV-vectors can efficiently target both retinal pigment epithelium and photoreceptor cells, the origin of most retinal degenerations. This review focuses on the AAV-based gene therapy in mouse models of recessive retinal degenerations, especially those in which delivery of the correct copy of the wild-type gene has led to significant beneficial effects on visual function, as determined by morphological, biochemical, electroretinographic and behavioral analysis. The past studies in animal models and ongoing successful LCA2 clinical trials, predict a bright future for AAV gene replacement treatment for inherited recessive retinal diseases.     

缺血性中风动物模型研究现状

利用实验动物或细胞模型完全模拟人类的中风十分困难,动物模型与临床的拟合具有重要意义。本文对目前缺血性中风动物模型研究中的实验动物选择、模型评价标准及造模方法 ,以及主要局灶缺血模型的优缺点进行述评,为缺血性中风的基础和应用研究选择合适的实验动物模型提供参考。     

Cell-replacement therapy and neural repair in the retina

Visual impairment severely affects the quality of life of patients and their families and is also associated with a deep economic impact. The most common pathologies responsible for visual impairment and legally defined blindness in developed countries include age-related macular degeneration, glaucoma and diabetic retinopathy. These conditions share common pathophysiological features: dysfunction and loss of retinal neurons. To date, two main approaches are being taken to develop putative therapeutic strategies: neuroprotection and cell replacement. Cell replacement is a novel therapeutic approach to restore visual capabilities to the degenerated adult neural retina and represents an emerging field of regenerative neurotherapy. The discovery of a population of proliferative cells in the mammalian retina has raised the possibility of harnessing endogenous retinal stem cells to elicit retinal repair. Furthermore, the development of suitable protocols for the reprogramming of differentiated somatic cells to a pluripotent state further increases the therapeutic potential of stem-cell-based technologies for the treatment of major retinal diseases. Stem-cell transplantation in animal models has been most effectively used for the replacement of photoreceptors, although this therapeutic approach is also being used for inner retinal pathologies. In this review, we discuss recent advances in the development of cell-replacement approaches for the treatment of currently incurable degenerative retinal diseases.     

Impaired therapeutic vasculogenesis by transplantation of OxLDL-treated endothelial progenitor cells

Previous in vitro studies have revealed that oxidized low density lipoprotein (OxLDL) has negative effects on the proliferation and activity of endothelial progenitor cells (EPCs). Here, we evaluated the effect of OxLDL on the therapeutic potential of EPCs in ischemia-induced neovascularization. EPCs derived from mobilized human peripheral blood mononuclear cells were cultured without or with OxLDL before transplantation. Hindlimb ischemia models were surgically induced in athymic nude mice, which then received an intracardiac injection of 3 x 10(5) EPCs. By laser Doppler perfusion image and ischemia damage score, we found that blood perfusion and ischemia damage were less well recovered in the OxLDL-treated EPC transplantation group than in controls. Histological examination showed fewer transplanted EPCs and lower capillary density in ischemic tissue. Local delivery of Stromal cell-derived factor (SDF-1) restored this defect and improved blood perfusion by recruiting OxLDL-treated EPCs to the ischemic area and increasing host capillary density. These results provide for the first time direct evidence that OxLDL impaired the therapeutic potential of EPCs in ischemia-induced neovascularization through an inhibitory effect on the migration, adhesion, and incorporation of EPCs into vasculature and/or entrapment in the perivascular region in vivo. A therapeutic strategy based on SDF-1 administration ameliorated such defects and improved postischemic neovascularization.     

Animal models of ischemic stroke: balancing experimental aims and animal care

Animal models of ischemic stroke are examples of an induced model that can present challenges from the perspectives of protocol review and animal management. The review presented here will include a brief summary of the current state of knowledge about clinical stroke; a general synopsis of important unanswered research questions that justify use of animal stroke models; an overview of various animal models of ischemic stroke, including strengths and limitations; and a discussion of animal care issues relative to ischemic stroke models. Good communication and interactive education among primary investigators, laboratory animal veterinarians and caretakers, and institutional animal care and use committee members are critical in achieving a balance between research objectives and animal care issues when using animal stroke models.     

Transplantation of endothelial progenitor cells for therapeutic neovascularization

Endothelial progenitor cells (EPCs), which were first identified in adult peripheral blood mononuclear cells (MNCs), play an important role in postnatal neovascularization. Tissue ischemia augments mobilization of EPCs from bone marrow into the circulation and enhances incorporation of EPCs at sites of neovascularization. Two methods to obtain EPCs from bone marrow, peripheral blood or cord blood MNCs have been evaluated for therapeutic neovascularization: (1) fresh isolation using anti-CD34, anti-KDR or anti-AC133 antibody, and (2) ex vivo expansion of total MNCs. In an immunodeficient mouse model of hindlimb ischemia, systemic transplantation of human ex vivo expanded EPCs improves limb survival through the enhancement of blood flow in the ischemic tissue. A similar strategy also leads to histological and functional preservation of ischemic myocardium of nude rats. Recently, a preclinical study of catheter-based, intramyocardial transplantation ofautologous EPCs in a swine model of chronic myocardial ischemia demonstrated the therapeutic potential of cell-based therapy, with attenuation of myocardial ischemia and improvement in left ventricular function. These favorable outcomes strongly suggest a therapeutic impact of EPC transplantation in clinical settings. Further basic research, with improved understanding of the mechanisms governing homing and incorporation of EPCs, will be still necessary to optimize the methodology of the cell therapy.     

Neuroprotective Effects of Bis(7)-tacrine in a Rat Model of Pressure-Induced Retinal Ischemia

The retinal ischemia–reperfusion model has been studied extensively and is an ideal animal model for studying clinical situations such as acute glaucoma and optic neuropathy. Our previous reports showed that bis(7)-tacrine had neuroprotective effects against glutamate-induced retinal ganglion cells damage through the drug’s anti-NMDA receptor effects. Here, we investigated whether bis(7)-tacrine protects the retina from ischemic injury in a rat model. Retinal ischemia was induced by raising the intraocular pressure to 120 mmHg for 90 min. Rats received intraperitoneal injections of 0.2 mg/kg bis(7)-tacrine or saline at 30 min before ischemia, and then twice a day after retinal ischemia. Morphometric evaluation showed that bis(7)-tacrine dramatically reduced the retinal damage compared with the control group. Moreover, bis(7)-tacrine suppressed ischemia-induced reductions in a- and b-wave amplitudes of electroretinography. Protein levels of p53, the tumor suppressor gene known to induce apoptosis, were increased after ischemic injury, and treatment with bis(7)-tacrine reduced the expression of the protein. Our results suggest that bis(7)-tacrine has a neuroprotective effect against ischemic injury in the rat retina, possibly through the drug’s anti-apoptotic effects. Bis(7)-tacrine may potentially be useful as a therapeutic drug in the management of ischemic retinal diseases.     

CD36: A multi-modal target for acute stroke therapy

A role for CD36 in the pathogenesis of atherosclerosis, inflammation and lipid metabolism has been well-documented. However, little is known about the role of CD36 in cerebral ischemia. The intent of this review is to develop the concept that CD36, whose functions have been implicated in other pathological events, is a prototypic inflammatory receptor that contributes to the pathogenesis of cerebral ischemia. The importance of CD36 as a treatment target is indicated by the fact that many treatment strategies that are effective in experimental models of stroke exhibit little or no efficacy in clinical trials. The failure of clinical trials may be due to the use of animal models of stroke that do not reflect traditional risk factors for stroke in humans. The discussion will be focused on two risk factors, hyperlipidemia and diabetes, that modulate CD36 responses. Blocking the expression and function of CD36 by pharmacological or genetic means will provide insight not only toward identifying CD36 as a novel molecular target but also for developing effective therapeutic strategies to treat stroke victims. More importantly, coupling clinically relevant conditions with CD36-mediated ischemic injury may provide an appropriate animal model paradigm and develop a scientific understanding that could lead to clinical translational studies involving human subjects.     

Endothelial progenitor cells for postnatal vasculogenesis

In the past decade, researchers have defined committed stem or progenitor cells from various tissues, including bone marrow, peripheral blood, brain, liver, and reproductive organs, in both adult animals and humans. Whereas most cells in adult organs are composed of differentiated cells, which express a variety of specific phenotypic genes adapted to each organ's environment, quiescent stem or progenitor cells are maintained locally or in the systemic circulation and are activated by environmental stimuli for physiological and pathological tissue regeneration. Recently, endothelial progenitor cells (EPCs) were isolated from peripheral blood CD34, Flk-1, or AC133 antigen-positive cells, which are considered to include a hematopoietic stem cell population, and were shown to be incorporated into foci of neovascularization. This finding, that circulating EPCs may home to sites of neovascularization and differentiate into endothelial cells in situ, is consistent with "vasculogenesis," a critical paradigm for embryonic neovascularization, and suggests that vasculogenesis and angiogenesis may constitute complementary mechanisms for postnatal neovascularization. Previous reports demonstrating therapeutic potential of EPC transplantation in animal models of hindlimb and myocardial ischemia opened the way to the clinical application of cell therapy: the replacement of diseased or degenerating cell populations, tissues, and organs. In this review, we summarize biological features of EPCs and speculate on the utility of EPCs for vascular and general medicine. cell transplantation; ischemia; neovascularization; stem cell     

Gene modification with integrin-linked kinase improves function of endothelial progenitor cells in pre-eclampsia in vitro

Integrin-linked kinase (ILK), a multifunctional serine-threonine protein kinase, has been shown to have implications for the treatment of ischemia vascular diseases by promoting angiogenesis in various tissues. However, whether this kinase has therapeutic potential in pre-eclampsia is not well studied. In this report, we determined the changes in the production and action of ILK on endothelial progenitor cells (EPCs) isolated from patients with pre-eclampsia. The effects of ILK transfection on proliferation, migration, and angiogenesis of EPCs were investigated. We showed that EPCs transfected with the ILK gene expressed high levels of ILK protein and mRNA. Transfection with ILK also enhanced the proliferative, migratory, and angiogenic capabilities of EPCs, and promoted the production of VEGF. These results suggest that ILK gene transfection is an effective approach to augment angiogenic properties of EPCs in vitro and providing basis for clinical cell-based gene therapy in patients with pre-eclampsia.     

细胞因子对内皮祖细胞功能影响的研究进展

内皮祖细胞(EPCs)是一种具有较强增殖能力的前体细胞,血管损伤或者缺血会刺激骨髓EPCs动员,迁移、归巢于相应的靶位,然后分化为内皮细胞(ECs),从而参与血管修复和血管新生。因此,EPCs的成功发现为缺血性和血管损伤性疾病的治疗提供了新策略。但是EPCs存在动员率低、靶向性较差和功能不全等问题。大量研究显示细胞因子对EPCs的动员、归巢、增殖和分化等均起着重要的调节作用,同时,通过调控细胞因子能改善EPCs的功能活性,因此选择合适的细胞因子来提高EPCs功能变得非常重要。现总结了近年来细胞因子对EPCs功能影响的研究进展,并提出有待解决的问题和作一定的展望。     

Kallikrein-kinin in stem cell therapy

The tissue kallikrein-kinin system exerts a wide spectrum of biological activities in the cardiovascular, renal and central nervous systems. Tissue kallikrein-kinin modulates the proliferation, viability, mobility and functional activity of certain stem cell populations, namely mesenchymal stem cells(MSCs), endothelial progenitor cells(EPCs), mononuclear cell subsets and neural stem cells. Stimulation of these stem cells by tissue kallikrein-kinin may lead to protection against renal, cardiovascular and neural damage by inhibiting apoptosis, inflammation, fibrosis and oxidative stress and promoting neovascularization. Moreover, MSCs and EPCs genetically modified with tissue kallikrein are resistant to hypoxia- and oxidative stress-induced apoptosis, and offer enhanced protective actions in animal models of heart and kidney injury and hindlimb ischemia. In addition, activation of the plasma kallikrein-kinin system promotes EPC recruitment to the inflamed synovium of arthritic rats. Conversely, cleaved high molecular weight kininogen, a product of plasma kallikrein, reduces the viability and vasculogenic activity of EPCs. Therefore, kallikrein-kinin provides a new approach in enhancing the efficacy of stem cell therapy for human diseases.     

Increased oxidative stress during hyperglycemic cerebral ischemia

In this review, we summarize the role of hyperglycemia during cerebral ischemia. Hyperglycemia occurring during experimental and clinical stroke has been associated with increased cerebral damage. Increased oxidative stress resulting from hyperglycemia is believed to contribute to the exacerbated damage. More specifically, superoxide, nitric oxide and peroxynitrite are believed to play an important role in cerebral damage. This also involves increased recruitment of various blood cells to the ischemic zone that contribute to inflammation. We present data from our group and others that demonstrate that free radical production is increased during hyperglycemic stroke in rodents. Recent data suggest that inflammation is an important component of ischemic damage under both normo- and hyperglycemic conditions. We summarize numerous studies that indicate that a variety of antioxidant (inhibition of free radical production, scavenging of free radicals and increasing free radical degradation) and anti-inflammatory strategies decrease cerebral infarction. Finally, we compare the success of some of these strategies in clinical trials compared to the animal models.     

Prospects for the future: the role of free radicals in the treatment of stroke

Studies using animal models of stroke have demonstrated that free radicals are highly reactive molecules generated predominantly during cellular respiration and normal metabolism. Imbalance between cellular production of free radicals and the ability of cells to defend against them is referred to as oxidative stress. After ischemic brain damage introduced by ischemic stroke or reperfusion, production of reactive oxygen species may increase, sometimes drastically, leading to tissue damage via several different cellular molecular pathways. The damage can become more widespread due to weakened cellular antioxidant defense systems after ischemic stroke. These experimental findings have important implications for the treatment of human cerebral ischemia. Agents directed at eliminating oxygen radicals must be administered before, or in the early stages of, reperfusion after ischemia. The therapeutic window seems to be narrow and limited to, at most, a few hours. Future research may clarify the current hypothesis that the accuracy of gene expression could account for the recovery of cellular function after ischemic stroke. This may open the window to the future use of drug combinations that may be rationally administered sequentially. If the phenomenon of ischemic tolerance plays a role in this concept is still a matter of debate.     

Effects of intranasal guanosine administration on brain function in a rat model of ischemic stroke

Ischemic stroke is a major cause of morbidity and mortality worldwide and only few affected patients are able to receive treatment, especially in developing countries. Detailed pathophysiology of brain ischemia has been extensively studied in order to discover new treatments with a broad therapeutic window and that are accessible to patients worldwide. The nucleoside guanosine (Guo) has been shown to have neuroprotective effects in animal models of brain diseases, including ischemic stroke. In a rat model of focal permanent ischemia, systemic administration of Guo was effective only when administered immediately after stroke induction. In contrast, intranasal administration of Guo (In-Guo) was effective even when the first administration was 3 h after stroke induction. In order to validate the neuroprotective effect in this larger time window and to investigate In-Guo neuroprotection under global brain dysfunction induced by ischemia, we used the model of thermocoagulation of pial vessels in Wistar rats. In our study, we have found that In-Guo administered 3 h after stroke was capable of preventing ischemia-induced dysfunction, such as bilateral suppression and synchronicity of brain oscillations and ipsilateral cell death signaling, and increased permeability of the blood-brain barrier. In addition, In-Guo had a long-lasting effect on preventing ischemia-induced motor impairment. Our data reinforce In-Guo administration as a potential new treatment for brain ischemia with a more suitable therapeutic window.