Role of Fanconi DNA repair pathway in neural stem cell homeostasis

Defects in DNA repair pathways have been involved in collapse of early neurogenesis leading to brain development abnormalities and embryonic lethality. However, consequences of DNA repair defects in adult neural stem and progenitor cells and their potential contribution in ageing phenotype are poorly understood. The Fanconi anaemia (FA) pathway, which functions primarily as a DNA damage response system, has been examined in neural stem and progenitor cells during developmental and adult neurogenesis. We have shown that loss of fanca and fancg specifically provokes neural progenitor apoptosis during forebrain development, related to DNA repair defects, which persists in adulthood leading to depletion of the neural stem cell pool with ageing. In addition, neural stem cells from FA mice had a reduced capacity to self-renew in vitro. Here, we expand upon our recent work and give further data examining possible implication of oxidative stress. Therefore, FA phenotype might be interpreted as a premature ageing of stem cells, DNA damages being among the driving forces of ageing.     

Current clinical therapies for cartilage repair, their limitation and the role of stem cells

The management of osteochondral defects of articular cartilage, whether from trauma or degenerative disease, continues to be a significant challenge for Orthopaedic surgeons. Current treatment options such as abrasion arthroplasty procedures, osteochondral transplantation and autologous chondrocyte implantation fail to produce repair tissue exhibiting the same mechanical and functional properties of native articular cartilage. This results in repair tissue that inevitably fails as it is unable to deal with the mechanical demands of articular cartilage, and does not prevent further degeneration of the native cartilage. Mesenchymal stem cells have been proposed as a potential source of cells for cell-based cartilage repair due to their ability to self-renew and undergo multi-lineage differentiation. This proposed procedure has the advantage of not requiring harvesting of cells from the joint surface, and its associated donor site morbidity, as well as having multiple possible adult donor tissues such as bone marrow, adipose tissue and synovium. Mesenchymal stem cells have multi-lineage potential, but can be stimulated to undergo chondrogenesis in the appropriate culture medium. As the majority of work with mesenchymal stem cell-derived articular cartilage repair has been carried out in vitro and in animal studies, more work still has to be done before this technique can be used for clinical purposes. This includes realizing the ideal method of harvesting mesenchymal stem cells, the culture medium to stimulate proliferation and differentiation, appropriate choice of scaffold incorporating growth factors directly or with gene therapy and integration of repair tissue with native tissue.     

Sequential In vivo Imaging of Osteogenic Stem/Progenitor Cells During Fracture Repair

Bone turns over continuously and is highly regenerative following injury. Osteogenic stem/progenitor cells have long been hypothesized to exist, but in vivo demonstration of such cells has only recently been attained. Here, in vivo imaging techniques to investigate the role of endogenous osteogenic stem/progenitor cells (OSPCs) and their progeny in bone repair are provided. Using osteo-lineage cell tracing models and intravital imaging of induced microfractures in calvarial bone, OSPCs can be directly observed during the first few days after injury, in which critical events in the early repair process occur. Injury sites can be sequentially imaged revealing that OSPCs relocate to the injury, increase in number and differentiate into bone forming osteoblasts. These methods offer a means of investigating the role of stem cell-intrinsic and extrinsic molecular regulators for bone regeneration and repair.     

Human stem cells for CNS repair

Although most peripheral tissues have at least a limited ability for self-repair, the central nervous system (CNS) has long been known to be relatively resistant to regeneration. Small numbers of stem cells have been found in the adult brain but do not appear to be able to affect any significant recovery following disease or insult. In the last few decades, the idea of being able to repair the brain by introducing new cells to repair damaged areas has become an accepted potential treatment for neurodegenerative diseases. This review focuses on the suitability of various human stem cell sources for such treatments of both slowly progressing conditions, such as Parkinson’s disease, Huntington’s disease and multiple sclerosis, and acute insult, such as stroke and spinal cord injury. Despite stem cell transplantation having now moved a step closer to the clinic with the first trials of autologous mesenchymal stem cells, the effects shown are moderate and are not yet at the stage of development that can fulfil the hopes that have been placed on stem cells as a means to replace degenerating cells in the CNS. Success will depend on careful investigation in experimental models to enable us to understand not just the practicalities of stem cell use, but also the underlying biological principles.     

The potential therapeutic use of stem cells in cartilage repair

As our population demographics change, osteoarthritis and cartilage defects are becoming more prevalent. The discovery of stems cells and their ability for indefinite regeneration has revolutionised the way cartilage problems are viewed. Tissue engineering has been shown to be the ideal way of repairing articular cartilage lesions, i.e. back to native tissue. Cartilage is an ideal tissue engineering target as it is avascular, aneural and alymphatic. The two main types of stem cells being investigated in chondrogenesis are embryological and mesenchymal stem cells. Research into embryological stem cells has been surrounded by controversy because of ethical, religious and social concerns. We discuss the use of embryological and mesenchymal stem cells in cartilage repair and the various factors involved in the differentiation into chondrocytes. We also discuss commonly used mesenchymal stem cell markers and their limitations.     

Placenta- versus bone-marrow-derived mesenchymal cells for the repair of segmental bone defects in a rabbit model

Tissue-engineered bones (TEBs) constructed with bone-marrow-derived mesenchymal stem cells (BMSCs) seeded on biomaterial scaffolds have achieved good results for bone defect repair in both animal experiments and clinical trials. This has been limited, however, by the source and quantity of BMSCs. We here explored TEBs constructed by placenta-derived mesenchymal stem cells (PMSCs) and compared their effect for the repair of critical-sized segmental osteoperiosteal defects with TEBs constructed with BMSCs. PMSCs were isolated from rabbit placenta by gradient centrifugation and in vitro monolayer culturing, and BMSCs were isolated from the hindlimb bone marrow of newborn rabbit. Primary cultured PMSCs and BMSCs were uniformly in a spindle shape. Immunocytochemistry indicated that both types of cells are positive for CD44 and CD105, and negative for CD34 and CD40L, confirming that they are mesenchymal stem cells. BrdU-labeled PMSCs and BMSCs were respectively co-cultured with bio-derived bone materials to construct TEBs in vitro. Critical-sized segmental osteoperiosteal defects of radii were created in 24 rabbits by surgery. The defects were repaired with TEBs constructed with PMSCs and BMSCs. The results showed that TEBs constructed by both PMSCs and BMSCs could repair the osteoperiosteal defects in a 'multipoint' manner. Measurement of radiography, histology, immunohistochemistry, alkaline phosphatase activity, osteocalcin assaying and biomechanical properties have found no significant difference between the two groups at 2, 4, 8 and 12 weeks after the transplantation (P > 0.05). Taken together, our results indicate that PMSCs have similar biological characteristics and osteogenic capacity to BMSCs and can be used as a new source of seeding cells for TEBs.     

Stem Cell Research: A Novel Boulevard towards Improved Bovine Mastitis Management

The dairy industry is a multi-billion dollar industry catering the nutritional needs of all age groups globally through the supply of milk. Clinical mastitis has a severe impact on udder tissue and is also an animal welfare issue. Moreover, it significantly reduces animal value and milk production. Mammary tissue damage reduces the number and activity of epithelial cells and consequently contributes to decreased milk production. The high incidence, low cure rate of this highly economic and sometimes deadly disease is an alarming for dairy sector as well as policy makers. Bovine mammary epithelial cells (MECs) and their stem cells are very important in milk production and bioengineering. The adult mammary epithelium consists of two main cell types; an inner layer of luminal epithelial cells, which produce the milk during lactation, and an outer layer of myoepithelial cells resting on a basement membrane, which are responsible for pushing the milk through the ductal network to the teat cistern. Inner layer of columner/luminal cells of bovine MECs, is characterized by cytokeratin18, 19 (CK18, CK19) and outer layer such as myoepithelial cells which are characterized by CK14, α-smooth muscle actin (α-SMA) and p63. Much work has been done in mouse and human, on mammary gland stem cell research, particularly in cancer therapy, but stem cell research in bovine is still in its infancy. Such stem/progenitor cell discoveries in human and mouse mammary gland bring some hope for application in bovines. These progenitors may be therapeutically adopted to correct the structural/cytological defects in the bovine udder due to mastitis. In the present review we focused on various kinds of stem/progenitor cells which can have therapeutic utility and their possibilities to use as a potential stem cell therapy in the management of bovine post-mastitis damage in orders to restore milk production. The possibilities of bovine mammary stem cell therapy offers significant potential for regeneration of tissues that can potentially replace/repair diseased and damaged tissue through differentiation into epithelial, myoepithelial and/or cuboidal/columnar cells in the udder with minimal risk of rejection and side effects.     

PCL-Based Composite Scaffold Matrices for Tissue Engineering Applications

Biomaterial-based scaffolds are important cues in tissue engineering (TE) applications. Recent advances in TE have led to the development of suitable scaffold architecture for various tissue defects. In this narrative review on polycaprolactone (PCL), we have discussed in detail about the synthesis of PCL, various properties and most recent advances of using PCL and PCL blended with either natural or synthetic polymers and ceramic materials for TE applications. Further, various forms of PCL scaffolds such as porous, films and fibrous have been discussed along with the stem cells and their sources employed in various tissue repair strategies. Overall, the present review affords an insight into the properties and applications of PCL in various tissue engineering applications.     

Cord blood for tissue regeneration

Umbilical cord blood (CB) has become a commonly accepted source of hematopoietic stem cells for transplantation in children and adults. It is readily available and outperforms bone marrow (BM) as well as peripheral blood stem cells in terms of tolerance for HLA‐mismatches between donor and recipient and its decreased graft‐versus‐host disease. Clinical use has been expanded from hematological malignancies to various areas such as treatment of metabolic genetic disorders or to induce angiogenesis. For the last years CB has been under intense experimental investigation in in vitro differentiation models as well as in preclinical animal models. Since CB‐derived stem cells offer multiple advantages over adult stem cells from other sources like BM, CB may provide a future source of stem cells for tissue repair and regeneration. To facilitate the use of CB‐derived stem cells in clinical scenarios, the biology of these cells needs to be further explored in detail particularly with regard to the fact that different non‐hematopoietic stem cell populations occur within CB. Here we explore the most consistent and the most contradictory data referring to the differentiation potential of CB‐derived stem cells and give an outlook on their potential clinical value including and possible reprogramming into IPS cells. J. Cell. Biochem. 108: 762–768, 2009. © 2009 Wiley‐Liss, Inc.     

Adult stem cells: seing is not being

Recent unexpected observations in adult rodents that stem/progenitor cells located in the bone marrow, but also in other tissues, could, after their transplantation to an irradiated host contribute to the regeneration of damaged organs such as brain, liver, pancreas or muscle, have raised much hope for future therapeutic applications. These data have also initially been interpreted as a proof of a possible transdifferentiation or plasticity of adult stem cells located in these tissues. Additional experiments rigorously analyzed have tempered initial enthusiasm, by showing that if marrow cells do migrate in damaged muscles and liver, their contribution to organ repair is low, and in some cases, explained by cell fusion. Nevertheless, among bone marrow cells, two categories of stem cells now emerge that have a potentially tremendous interest in cell therapy, if we succeed in understanding how to purify, amplify and differentiate these more efficiently and reproducibly.     

Muscle-derived stem cells: implications for effective myoblast transfer therapy

Stem cells have been proposed as a wonder solution for tissue repair in many situations and have attracted much attention in the media for both their therapeutic potential and ethical implications. In addition to the excitement generated by embryonic stem cells, research has now identified a number of stem cells within adult tissues which pose much more realistic targets for therapeutic interventions. Myoblast transfer therapy (MTT) has long been viewed as a potential therapy for the debilitating muscle-wasting disorder Duchenne Muscular Dystrophy. This technique relies on the transplantation of committed muscle precursor cells directly into the muscle fibres but has had little success in clinical trials. The recent discovery of a population of cells within adult muscle with stem cell-like characteristics has interesting implications for the future of such putative cell transplantation therapies. This review focuses on the characterization and application of these potential muscle-derived stem cells (MDSC) to MTT.     

A comparison of the gene expression profiles of CRL-1807 colonocytes exposed to endogenous AAPH-generated peroxides and exogenous peroxides from heated oil

Oxidation of PUFAs in the diet has the potential to be genotoxic and hence carcinogenic. Such carcinogenic processes originate within stem cells of the colon. These cells appear to be predisposed to the carcinogenic process. In colon cells (CRL-1807) exposed to chemical reactions simulating exogenous and endogenous peroxidation reactions, we have observed that undifferentiated cells could mount an effective recombinational repair/TCR response to an endogenous peroxidative DNA damage insult, but not to an external exogenous peroxidative insult as one would encounter from a dietary source. This may suggest that defects in such specific DNA repair may play a role in tumour development in undifferentiated colonocytes exposed to a diet-derived lipid peroxides.     

In vitro organogenesis using multipotent cells

The establishment of efficient methods for promoting stem cell differentiation into target cells is important not only in regenerative medicine, but also in drug discovery. In addition to embryonic stem (ES) cells and various somatic stem cells, such as mesenchymal stem cells derived from bone marrow, adipose tissue, and umbilical cord blood, a novel dedifferentiation technology that allows the generation of induced pluripotent stem (iPS) cells has been recently developed. Although an increasing number of stem cell populations are being described, there remains a lack of protocols for driving the differentiation of these cells. Regeneration of organs from stem cells in vitro requires precise blueprints for each differentiation step. To date, studies using various model organisms, such as zebrafish, Xenopus laevis , and gene-targeted mice, have uncovered several factors that are critical for the development of organs. We have been using X. laevis , the African clawed frog, which has developmental patterns similar to those seen in humans. Moreover, Xenopus embryos are excellent research tools for the development of differentiation protocols, since they are available in high numbers and are sufficiently large and robust for culturing after simple microsurgery. In addition, Xenopus eggs are fertilized externally, and all stages of the embryo are easily accessible, making it relatively easy to study the functions of individual gene products during organogenesis using microinjection into embryonic cells. In the present review, we provide examples of methods for in vitro organ formation that use undifferentiated Xenopus cells. We also describe the application of amphibian differentiation protocols to mammalian stem cells, so as to facilitate the development of efficient methodologies for in vitro differentiation.     

CommentaryDNA Base Excision Repair Defects in Human Pathologies

DNA base excision repair (BER) is the main pathway for repair of endogenous damage in human cells. It was expected that a number of degenerative diseases could derive from BER defects. On the contrary, the link between BER defects and human pathology is elusive and the literature is full of conflicting results. The fact that most studies have investigated DNA variations but not their functional consequences has probably contributed to this confusing picture. From a functional point of view, it is likely that gross BER defects are simply not compatible with life and only limited reductions can be observed. Notwithstanding those limits, the pathological consequences of partial BER defects might be widespread and significant at the population level. This starts to emerge in particular for colorectal and lung cancer.     

Stem and progenitor cells for neurological repair: minor issues, major hurdles, and exciting opportunities for paracrine-based therapeutics

The transplantation of cultured stem and progenitor cells is a key element in the rapidly growing field of regenerative medicine. Based on their ability to rescue and/or repair injured tissue and partially restore organ function, multiple types of stem/progenitor cells have already entered into clinical trials. However, despite several decades of intense research, the goal to apply culture-expanded stem/progenitor cells in a manner that can effectively replace cells after injury has yet to be realized. Many sources of potentially useful cells are available, but something is clearly missing. In addition, recent studies suggest that paracrine effects of secreted or released factors are responsible for most of the benefits observed after cell transplantation, rather than direct cell replacement. These data call into question the need for cell transplantation for many types of therapy, in particular for acute injuries such as myocardial infarction and stroke. In this review, we examine current progress in the area of cell transplantation and minor issues and major hurdles regarding the clinical application of different cell types. We discuss the "paracrine hypothesis" for the action of transplanted stem/progenitor cells as an opportunity to identify defined combinations of biomolecules to rescue and/or repair tissues after injury. Although many of the concepts in this review will apply to multiple injury/repair systems, we will focus primarily on stem/progenitor cell-based treatments for neurological disorders and stroke.     

Commentary: DNA base excision repair defects in human pathologies

DNA base excision repair (BER) is the main pathway for repair of endogenous damage in human cells. It was expected that a number of degenerative diseases could derive from BER defects. On the contrary, the link between BER defects and human pathology is elusive and the literature is full of conflicting results. The fact that most studies have investigated DNA variations but not their functional consequences has probably contributed to this confusing picture. From a functional point of view, it is likely that gross BER defects are simply not compatible with life and only limited reductions can be observed. Notwithstanding those limits, the pathological consequences of partial BER defects might be widespread and significant at the population level. This starts to emerge in particular for colorectal and lung cancer.     

Bone regeneration in a rabbit ulna defect model: use of allogeneic adipose-derivedstem cells with low immunogenicity

Tissue engineering provides new potential treatments for the repair of bone defects. Bone-marrow-derived mesenchymal stem cells (BMSCs) represent an attractive cell source for therapeutic applications involving tissue engineering, although disadvantages, such as pain of harvest and low proliferation efficiency, are major limitations to the application of BMSCs in the clinic. Adipose-derived stem cells (ASCs) with their multilineage potential and satisfactory proliferation potential can be induced into the osteogenic lineage in vitro and can be anchored onto suitable scaffolds as seed cells to repair bone defects successfully in an autologous setting. Previous studies have indicated that both undifferentiated BMSCs and ASCs exhibit immunosuppression and immunoprivilege properties. We compare the immuno-function of undifferentiated and osteo-differentiated ASCs in vitro and explore the feasibility of applying allogeneic ASCs to the repair of ulnar bone defects in the rabbit model. Our study demonstrates that allogeneic osteogenic differentiated ASCs maintain low immunogenicity and negative immunomodulation. The allogeneic osteogenic differentiated ASCs combined with demineralized bone matrix successfully regenerate ulnar bone defects in rabbits without immunosuppressive therapies.     

Different roles of TGF-β in the multi-lineage differentiation of stem cells

Stem cells are a population of cells that has infinite or long-term self-renewal ability and can produce various kinds of descendent cells.Transforming growth factor β(TGF-β) family is a superfamily of growth factors,including TGF-β1,TGF-β2 and TGF-β3,bone morphogenetic proteins,activin/inhibin,and some other cytokines such as nodal,which plays very important roles in regulating a wide variety of biological processes,such as cell growth,differentiation,cell death.TGF-β,a pleiotropic cytokine,has been proved to be differentially involved in the regulation of multi-lineage differentiation of stem cells,through the Smad pathway,non-Smad pathways including mitogen-activated protein kinase pathways,phosphatidylinositol-3-kinase/AKT pathways and Rholike GTPase signaling pathways,and their cross-talks.For instance,it is generally known that TGF-β promotes the differentiation of stem cells into smooth muscle cells,immature cardiomyocytes,chondrocytes,neurocytes,hepatic stellate cells,Th17 cells,and dendritic cells.However,TGF-β inhibits the differentiation of stem cells into myotubes,adipocytes,endothelial cells,and natural killer cells.Additionally,TGF-β can provide competence for early stages of osteoblastic differentiation,but at late stages TGF-β acts as an inhibitor.The three mammalian isoforms(TGF-β1,2 and 3) have distinct but overlapping effects on hematopoiesis.Understanding the mechanisms underlying the regulatory effect of TGF-β in the stem cell multi-lineage differentiation is of importance in stem cell biology,and will facilitate both basic research and clinical applications of stem cells.In this article,we discuss the current status and progress in our understanding of different mechanisms by which TGF-β controls multi-lineage differentiation of stem cells.