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Most Cited Cell Regeneration Articles

The most cited articles published since 2013, extracted from Scopus.

Role of Oct4 in the early embryo development

Volume 3, Issue 1, 四月 2014, Page
Guangming Wu | Hans R. Schöler | Hans R. Schöler

© 2014 Wu and Schöler; licensee BioMed Central Ltd. Oct4 is a key component of the pluripotency regulatory network, and its reciprocal interaction with Cdx2 has been shown to be a determinant of either the self-renewal of embryonic stem cells (ESCs) or their differentiation into trophoblast. Oct4 of maternal origin is postulat ed to play critical role in defining totipotency and inducing pluripotency during embryonic development. However, the genetic elimination of maternal Oct4 using a Cre-lox approach in mouse revealed that the establishment of totipotency in maternal Oct4-depleted embryos was not affected, and that these embryos could complete full-term development without any obvious defect. These results indicate that Oct4 is not essential for the initiation of pluripotency, in contrast to its critical role in maintaining pluripotency. This conclusion is further supported by the formation of Oct4-GFP- and Nanog- expressing inner cell masses (ICMs) in embryos with complete inactivation of both maternal and zygotic Oct4 expression and the reprogramming of fibroblasts into fully pluripotent cells by Oct4-deficient oocytes.

Generation of multi-gene knockout rabbits using the Cas9/gRNA system

Volume 3, Issue 1, 八月 2014, Page
Quanmei Yan | Quanjun Zhang | Huaqiang Yang | Qingjian Zou | Chengcheng Tang | Nana Fan | Liangxue Lai | Liangxue Lai

© 2014 Yan et al.; licensee BioMed Central Ltd. The prokaryotic clustered regularly interspaced short palindromic repeat (CRISPR)-associated system (Cas) is a simple, robust and efficient technique for gene targeting in model organisms such as zebrafish, mice and rats. In this report, we applied CRISPR technology to rabbits by microinjection of Cas9 mRNA and guided RNA (gRNA) into the cytoplasm of pronuclear-stage embryos. We achieved biallelic gene knockout (KO) rabbits by injection of 1 gene (IL2rg) or 2 gene (IL2rg and RAG1) Cas9 mRNA and gRNA with an efficiency of 100%. We also tested the efficiency of multiple gene KOs in early rabbit embryos and found that the efficiency of simultaneous gene mutation on target sites is as high as 100% for 3 genes (IL2rg, RAG1 and RAG2) and 33.3% for 5 genes (IL2rg, RAG1, RAG2, TIKI1 and ALB). Our results demonstrate that the Cas9/gRNA system is a highly efficient and fast tool not only for single-gene editing but also for multi-gene editing in rabbits.

Reprogramming barriers and enhancers: Strategies to enhance the efficiency and kinetics of induced pluripotency

Volume 4, Issue 1, 十一月 2015, Page
Behnam Ebrahimi

© 2015 Ebrahimi. Induced pluripotent stem cells are powerful tools for disease modeling, drug screening, and cell transplantation therapies. These cells can be generated directly from somatic cells by ectopic expression of defined factors through a reprogramming process. However, pluripotent reprogramming is an inefficient process because of various defined and unidentified barriers. Recent studies dissecting the molecular mechanisms of reprogramming have methodically improved the quality, ease, and efficiency of reprogramming. Different strategies have been applied for enhancing reprogramming efficiency, including depletion/inhibition of barriers (p53, p21, p57, p16 Ink4a /p19 Arf , Mbd3, etc.), overexpression of enhancing genes (e.g., FOXH1, C/EBP alpha, UTF1, and GLIS1), and administration of certain cytokines and small molecules. The current review provides an in-depth overview of the cutting-edge findings regarding distinct barriers of reprogramming to pluripotency and strategies to enhance reprogramming efficiency. By incorporating the mechanistic insights from these recent findings, a combined method of inhibition of roadblocks and application of enhancing factors may yield the most reliable and effective approach in pluripotent reprogramming.

Generation of tooth-like structures from integration-free human urine induced pluripotent stem cells

Volume 2, Issue 1, 七月 2013, Page
Jinglei Cai | Yanmei Zhang | Pengfei Liu | Pengfei Liu | Shubin Chen | Xuan Wu | Xuan Wu | Yuhua Sun | Ang Li | Ke Huang | Rongping Luo | Lihui Wang | Ying Liu | Ying Liu | Ting Zhou | Shicheng Wei | Shicheng Wei | Guangjin Pan | Duanqing Pei

© 2013 Cai et al.; licensee BioMed Central Ltd. Background: Tooth is vital not only for a good smile, but also good health. Yet, we lose tooth regularly due to accidents or diseases. An ideal solution to this problem is to regenerate tooth with patients' own cells. Here we describe the generation of tooth-like structures from integration-free human urine induced pluripotent stem cells (ifhU-iPSCs). Results: We first differentiated ifhU-iPSCs to epithelial sheets, which were then recombined with E14.5 mouse dental mesenchymes. Tooth-like structures were recovered from these recombinants in 3 weeks with success rate up to 30% for 8 different iPSC lines, comparable to H1 hESC. We further detected that ifhU-iPSC derived epithelial sheets differentiated into enamel-secreting ameloblasts in the tooth-like structures, possessing physical properties such as elastic modulus and hardness found in the regular human tooth. Conclusion: Our results demonstrate that ifhU-iPSCs can be used to regenerate patient specific dental tissues or even tooth for further drug screening or regenerative therapies.

Wnt/β-catenin signaling in heart regeneration

Volume 4, Issue 1, 七月 2015, Page
Gunes Ozhan | Gunes Ozhan | Gilbert Weidinger

� 2015 Ozhan and Weidinger. The ability to repair damaged or lost tissues varies significantly among vertebrates. The regenerative ability of the heart is clinically very relevant, because adult teleost fish and amphibians can regenerate heart tissue, but we mammals cannot. Interestingly, heart regeneration is possible in neonatal mice, but this ability is lost within 7 days after birth. In zebrafish and neonatal mice, lost cardiomyocytes are regenerated via proliferation of spared, differentiated cardiomyocytes. While some cardiomyocyte turnover occurs in adult mammals, the cardiomyocyte production rate is too low in response to injury to regenerate the heart. Instead, mammalian hearts respond to injury by remodeling of spared tissue, which includes cardiomyocyte hypertrophy. Wnt/β-catenin signaling plays important roles during vertebrate heart development, and it is re-activated in response to cardiac injury. In this review, we discuss the known functions of this signaling pathway in injured hearts, its involvement in cardiac fibrosis and hypertrophy, and potential therapeutic approaches that might promote cardiac repair after injury by modifying Wnt/β-catenin signaling. Regulation of cardiac remodeling by this signaling pathway appears to vary depending on the injury model and the exact stages that have been studied. Thus, conflicting data have been published regarding a potential role of Wnt/β-catenin pathway in promotion of fibrosis and cardiomyocyte hypertrophy. In addition, the Wnt inhibitory secreted Frizzled-related proteins (sFrps) appear to have Wnt-dependent and Wnt-independent roles in the injured heart. Thus, while the exact functions of Wnt/β-catenin pathway activity in response to injury still need to be elucidated in the non-regenerating mammalian heart, but also in regenerating lower vertebrates, manipulation of the pathway is essential for creation of therapeutically useful cardiomyocytes from stem cells in culture. Hopefully, a detailed understanding of the in vivo role of Wnt/β-catenin signaling in injured mammalian and non-mammalian hearts will also contribute to the success of current efforts towards developing regenerative therapies.

The involvement of interleukin-22 in the expression of pancreatic beta cell regenerative Reg genes

Volume 2, Issue 1, 四月 2013, Page
Thomas Hill | Olga Krougly | Enayat Nikoopour | Stacey Bellemore | Edwin Lee-Chan | Lynette A. Fouser | David J. Hill | David J. Hill | Bhagirath Singh | Bhagirath Singh | Bhagirath Singh

© 2013 Hill et al.; licensee BioMed Central Ltd. Background: In Type 1 diabetes, the insulin-producing β-cells within the pancreatic islets of Langerhans are destroyed. We showed previously that immunotherapy with Bacillus Calmette-Guerin (BCG) or complete Freund's adjuvant (CFA) of non-obese diabetic (NOD) mice can prevent disease process and pancreatic β-cell loss. This was associated with increased islet Regenerating (Reg) genes expression, and elevated IL-22-producing Th17 T-cells in the pancreas. Results: We hypothesized that IL-22 was responsible for the increased Reg gene expression in the pancreas. We therefore quantified the Reg1, Reg2, and Reg 3δ (INGAP) mRNA expression in isolated pre-diabetic NOD islets treated with IL-22. We measured IL-22, and IL-22 receptor(R)- α mRNA expression in the pancreas and spleen of pre-diabetic and diabetic NOD mice. Our results showed: 1) Reg1 and Reg2 mRNA abundance to be significantly increased in IL-22-treated islets in vitro; 2) IL-22 mRNA expression in the pre-diabetic mouse pancreas increased with time following CFA treatment; 3) a reduced expression of IL-22R α following CFA treatment; 4) a down-regulation in Reg1 and Reg 2 mRNA expression in the pancreas of pre-diabetic mice injected with an IL-22 neutralizing antibody; and 5) an increased islet β-cell DNA synthesis in vitro in the presence of IL-22. Conclusions: We conclude that IL-22 may contribute to the regeneration of β-cells by up-regulating Regenerating Reg1 and Reg2 genes in the islets.

Targeted genome engineering in human induced pluripotent stem cells by penetrating TALENs

Volume 2, Issue 1, 六月 2013, Page
Renli Ru | Yongchao Yao | Songlin Yu | Benpeng Yin | Wanwan Xu | Siting Zhao | Li Qin | Xiaoping Chen | Xiaoping Chen

� 2013 Ru et al.; licensee BioMed Central Ltd. Background: Zinc-finger nucleases (ZFNs) and transcription activator-like effector nucleases (TALENs) have been successfully used to knock out endogenous genes in stem cell research. However, the deficiencies of current gene-based delivery systems may hamper the clinical application of these nucleases. A new delivery method that can improve the utility of these nucleases is needed. Results: In this study, we utilized a cell-penetrating peptide-based system for ZFN and TALEN delivery. Functional TAT-ZFN and TAT-TALEN proteins were generated by fusing the cell-penetrating TAT peptide to ZFN and TALEN, respectively. However, TAT-ZFN was difficult to purify in quantities sufficient for analysis in cell culture. Purified TAT-TALEN was able to penetrate cells and disrupt the gene encoding endogenous human chemokine (C-C motif) receptor 5 (CCR5, a co-receptor for HIV-1 entry into cells). Hypothermic treatment greatly enhanced the TAT-TALEN-mediated gene disruption efficiency. A 5% modification rate was observed in human induced pluripotent stem cells (hiPSCs) treated with TAT-TALEN as measured by the Surveyor assay. C onclusions: TAT-TALEN protein-mediated gene disruption was applicable in hiPSCs and represents a promising technique for gene knockout in stem cells. This new technique may advance the clinical application of TALEN technology.

Defining synthetic surfaces for human pluripotent stem cell culture

Volume 2, Issue 1, 十一月 2013, Page
Jack W. Lambshead | Jack W. Lambshead | Laurence Meagher | Carmel O'Brien | Carmel O'Brien | Andrew L. Laslett | Andrew L. Laslett | Andrew L. Laslett

� 2013 Lambshead et al.; licensee BioMed Central Ltd. Human pluripotent stem cells (hPSCs) are able to self-renew indefinitely and to differentiate into all adult cell types. hPSCs therefore show potential for application to drug screening, disease modelling and cellular therapies. In order to meet this potential, culture conditions must be developed that are consistent, defined, scalable, free of animal products and that facilitate stable self-renewal of hPSCs. Several culture surfaces have recently been reported to meet many of these criteria although none of them have been widely implemented by the stem cell community due to issues with validation, reliability and expense. Most hPSC culture surfaces have been derived from extracellular matrix proteins (ECMPs) and their cell adhesion molecule (CAM) binding motifs. Elucidating the CAM-mediated cell-surface interactions that are essential for the in vitro maintenance of pluripotency will facilitate the optimisation of hPSC culture surfaces. Reports indicate that hPSC cultures can be supported by cell-surface interactions through certain CAM subtypes but not by others. This review summarises the recent reports of defined surfaces for hPSC culture and focuses on the CAMs and ECMPs involved.

Enzymatic and non-enzymatic isolation systems for adipose tissue-derived cells: Current state of the art

Volume 4, Issue 1, 九月 2015, Page
Eleni Oberbauer | Eleni Oberbauer | Carolin Steffenhagen | Carolin Steffenhagen | Christoph Wurzer | Christoph Wurzer | Christian Gabriel | Christian Gabriel | Christian Gabriel | Heinz Redl | Heinz Redl | Susanne Wolbank | Susanne Wolbank

� 2015 Oberbauer et al. In the past decade, adipose tissue became a highly interesting source of adult stem cells for plastic surgery and regenerative medicine. The isolated stromal vascular fraction (SVF) is a heterogeneous cell population including the adipose-derived stromal/stem cells (ASC), which showed regenerative potential in several clinical studies and trials. SVF should be provided in a safe and reproducible manner in accordance with current good manufacturing practices (cGMP). To ensure highest possible safety for patients, a precisely defined procedure with a high-quality control is required. Hence, an increasing number of adipose tissue-derived cell isolation systems have been developed. These systems aim for a closed, sterile, and safe isolation process limiting donor variations, risk for contaminations, and unpredictability of the cell material. To isolate SVF from adipose tissue, enzymes such as collagenase are used. Alternatively, in order to avoid enzymes, isolation systems using physical forces are available. Here, we provide an overview of known existing enzymatic and non-enzymatic adipose tissue-derived cell isolation systems, which are patented, published, or already on the market.

Glbase: A framework for combining, analyzing and displaying heterogeneous genomic and high-throughput sequencing data

Volume 3, Issue 1, 一月 2014, Page
Andrew P. Hutchins | Ralf Jauch | Ralf Jauch | Mateusz Dyla | Mateusz Dyla | Diego Miranda-Saavedra

© 2014 Hutchins et al.; licensee BioMed Central Ltd. Genomic datasets and the tools to analyze them have proliferated at an astonishing rate. However, such tools are often poorly integrated with each other: each program typically produces its own custom output in a variety of non-standard file formats. Here we present glbase, a framework that uses a flexible set of descriptors that can quickly parse non-binary data files. glbase includes many functions to intersect two lists of data, including operations on genomic interval data and support for the efficient random access to huge genomic data files. Many glbase functions can produce graphical outputs, including scatter plots, heatmaps, boxplots and other common analytical displays of high-throughput data such as RNA-seq, ChIP-seq and microarray expression data. glbase is designed to rapidly bring biological data into a Python-based analytical environment to facilitate analysis and data processing. In summary, glbase is a flexible and multifunctional toolkit that allows the combination and analysis of high-throughput data (especially next-generation sequencing and genome-wide data), and which has been instrumental in the analysis of complex data sets. glbase is freely available at http://bitbucket.org/oaxiom/glbase/.

Characterization of a novel cell penetrating peptide derived from human Oct4

Volume 3, Issue 1, 一月 2014, Page
Eva Harreither | Hanna A. Rydberg | Helene L. Åmand | Vaibhav Jadhav | Lukas Fliedl | Christina Benda | Miguel A. Esteban | Duanqing Pei | Nicole Borth | Nicole Borth | Regina Grillari-Voglauer | Regina Grillari-Voglauer | Regina Grillari-Voglauer | Oliver Hommerding | Frank Edenhofer | Frank Edenhofer | Bengt Nordén | Johannes Grillari | Johannes Grillari | Johannes Grillari

© 2014 Harreither et al.; licensee BioMed Central Ltd. Background: Oct4 is a transcription factor that plays a major role for the preservation of the pluripotent state in embryonic stem cells as well as for efficient reprogramming of somatic cells to induced pluripotent stem cells (iPSC) or other progenitors. Protein-based reprogramming methods mainly rely on the addition of a fused cell penetrating peptide. This study describes that Oct4 inherently carries a protein transduction domain, which can translocate into human and mouse cells.Results: A 16 amino acid peptide representing the third helix of the human Oct4 homeodomain, referred to as Oct4 protein transduction domain (Oct4-PTD), can internalize in mammalian cells upon conjugation to a fluorescence moiety thereby acting as a cell penetrating peptide (CPP). The cellular distribution of Oct4-PTD shows diffuse cytosolic and nuclear staining, whereas penetratin is strictly localized to a punctuate pattern in the cytoplasm. By using a Cre/loxP-based reporter system, we show that this peptide also drives translocation of a functionally active Oct4-PTD-Cre-fusion protein. We further provide evidence for translocation of full length Oct4 into human and mouse cell lines without the addition of any kind of cationic fusion tag. Finally, physico-chemical properties of the novel CPP are characterized, showing that in contrast to penetratin a helical structure of Oct4-PTD is only observed if the FITC label is present on the N-terminus of the peptide.Conclusions: Oct4 is a key transcription factor in stem cell research and cellular reprogramming. Since it has been shown that recombinant Oct4 fused to a cationic fusion tag can drive generation of iPSCs, our finding might contribute to further development of protein-based methods to generate iPSCs.Moreover, our data support the idea that transcription factors might be part of an alternative paracrine signalling pathway, where the proteins are transferred to neighbouring cells thereby actively changing the behaviour of the recipient cell.

Generation of functional organs from stem cells

Volume 2, Issue 1, 一月 2013, Page
Yunying Liu | Yunying Liu | Ru Yang | Zuping He | Wei Qiang Gao | Wei Qiang Gao

© 2013 Liu et al.; licensee BioMed Central Ltd. We are now well entering the exciting era of stem cells. Potential stem cell therapy holds great promise for the treatment of many diseases such as stroke, traumatic brain injury, Alzheimer's disease, Parkinson's disease, amyotrophic lateral-sclerosis, myocardial infarction, muscular dystrophy, diabetes, and etc. It is generally believed that transplantation of specific stem cells into the injured tissue to replace the lost cells is an effective way to repair the tissue. In fact, organ transplantation has been successfully practiced in clinics for liver or kidney failure. However, the severe shortage of donor organs has been a major obstacle for the expansion of organ transplantation programs. Toward that direction, generation of transplantable organs using stem cells is a desirable approach for organ replacement and would be of great interest for both basic and clinical scientists. Here we review recent progress in the field of organ generation using various methods including single adult tissue stem cells, a blastocyst complementation system, tissue decellularization/recellularization and a combination of stem cells and tissue engineering.

The interplay between metabolic homeostasis and neurodegeneration: Insights into the neurometabolic nature of amyotrophic lateral sclerosis

Volume 4, Issue 1, 八月 2015, Page
S. T. Ngo | S. T. Ngo | S. T. Ngo | S. T. Ngo | F. J. Steyn | F. J. Steyn

� 2015 Ngo and Steyn. Amyotrophic lateral sclerosis (ALS) is a fatal, neurodegenerative disease that is characterized by the selective degeneration of upper motor neurons and lower spinal motor neurons, resulting in the progressive paralysis of all voluntary muscles. Approximately 10 % of ALS cases are linked to known genetic mutations, with the remaining 90 % of cases being sporadic. While the primary pathology in ALS is the selective death of upper and lower motor neurons, numerous studies indicate that an imbalance in whole body and/or cellular metabolism influences the rate of progression of disease. This review summarizes current research surrounding the impact of impaired metabolic physiology in ALS. We extend ideas to consider prospects that lie ahead in terms of how metabolic alterations may impact the selective degeneration of neurons in ALS and how targeting of adenosine triphosphate-sensitive potassium (K ATP ) channels may represent a promising approach for obtaining neuroprotection in ALS.

Vascularization mediated by mesenchymal stem cells from bone marrow and adipose tissue: A comparison

Volume 4, Issue 1, 十月 2015, Page
Karoline Pill | Karoline Pill | Sandra Hofmann | Sandra Hofmann | Sandra Hofmann | Heinz Redl | Heinz Redl | Wolfgang Holnthoner | Wolfgang Holnthoner

� 2015 Pill et al. Tissue-engineered constructs are promising to overcome shortage of organ donors and to reconstruct at least parts of injured or diseased tissues or organs. However, oxygen and nutrient supply are limiting factors in many tissues, especially after implantation into the host. Therefore, the development of a vascular system prior to implantation appears crucial. To develop a functional vascular system, different cell types that interact with each other need to be co-cultured to simulate a physiological environment in vitro. This review provides an overview and a comparison of the current knowledge of co-cultures of human endothelial cells (ECs) with human adipose tissue-derived stem/stromal cells (ASCs) or bone marrow-mesenchymal stem cells (BMSCs) in three dimensional (3D) hydrogel matrices. Mesenchymal stem cells (MSCs), BMSCs or ASCs, have been shown to enhance vascular tube formation of ECs and to provide a stabilizing function in addition to growth factor delivery and permeability control for ECs. Although phenotypically similar, MSCs from different tissues promote tubulogenesis through distinct mechanisms. In this report, we describe differences and similarities regarding molecular interactions in order to investigate which of these two cell types displays more favorable characteristics to be used in clinical applications. Our comparative study shows that ASCs as well as BMSCs are both promising cell types to induce vascularization with ECs in vitro and consequently are promising candidates to support in vivo vascularization.

Generation of knockout rabbits using transcription activator-like effector nucleases

Volume 3, Issue 1, 五月 2014, Page
Yu Wang | Nana Fan | Jun Song | Juan Zhong | Xiaogang Guo | Weihua Tian | Quanjun Zhang | Fenggong Cui | Li Li | Philip N. Newsome | Philip N. Newsome | Jon Frampton | Miguel A. Esteban | Liangxue Lai

© 2014 Wang et al.; licensee BioMed Central Ltd. Zinc-finger nucleases and transcription activator-like effector nucleases are novel gene-editing platforms contributing to redefine the boundaries of modern biological research. They are composed of a non-specific cleavage domain and a tailor made DNA-binding module, which enables a broad range of genetic modifications by inducing efficient DNA double-strand breaks at desired loci. Among other remarkable uses, these nucleases have been employed to produce gene knockouts in mid-size and large animals, such as rabbits and pigs, respectively. This approach is cost effective, relatively quick, and can produce invaluable models for human disease studies, biotechnology or agricultural purposes. Here we describe a protocol for the efficient generation of knockout rabbits using transcription activator-like effector nucleases, and a perspective of the field.

Pharmacological manipulation of transcription factor protein-protein interactions: Opportunities and obstacles

Volume 4, Issue 1, 三月 2015, Page
Frank Fontaine | Jeroen Overman | Mathias Fran�ois

� 2015 Fontaine et al.; licensee BioMed Central. Much research on transcription factor biology and their genetic pathways has been undertaken over the last 30 years, especially in the field of developmental biology and cancer. Yet, very little is known about the molecular modalities of highly dynamic interactions between transcription factors, genomic DNA, and protein partners. Methodological breakthroughs such as RNA-seq (RNA-sequencing), ChIP-seq (chromatin immunoprecipitation sequencing), RIME (rapid immunoprecipitation mass spectrometry of endogenous proteins), and single-molecule imaging will dramatically accelerate the discovery rate of their molecular mode of action in the next few years. From a pharmacological viewpoint, conventional methods used to target transcription factor activity with molecules mimicking endogenous ligands fail to achieve high specificity and are limited by a lack of identification of new molecular targets. Protein-protein interactions are likely to represent one of the next major classes of therapeutic targets. Transcription factors, known to act mostly via protein-protein interaction, may well be at the forefront of this type of drug development. One hurdle in this field remains the difficulty to collate structural data into meaningful information for rational drug design. Another hurdle is the lack of chemical libraries meeting the structural requirements of protein-protein interaction disruption. As more attempts at modulating transcription factor activity are undertaken, valuable knowledge will be accumulated on the modality of action required to modulate transcription and how these findings can be applied to developing transcription factor drugs. Key discoveries will spawn into new therapeutic approaches not only as anticancer targets but also for other indications, such as those with an inflammatory component including neurodegenerative disorders, diabetes, and chronic liver and kidney diseases.

Dissecting microRNA-mediated regulation of stemness, reprogramming, and pluripotency

Volume 5, Issue 1, 三月 2016, Page
Young Jin Lee | Suresh Ramakrishna | Suresh Ramakrishna | Himanshu Chauhan | Won Sun Park | Seok Ho Hong | Seok Ho Hong | Kye Seong Kim | Kye Seong Kim

� 2016 Lee et al. Increasing evidence indicates that microRNAs (miRNAs), endogenous short non-coding RNAs 19-24 nucleotides in length, play key regulatory roles in various biological events at the post-transcriptional level. Embryonic stem cells (ESCs) represent a valuable tool for disease modeling, drug discovery, developmental studies, and potential cell-based therapies in regenerative medicine due to their unlimited self-renewal and pluripotency. Therefore, remarkable progress has been made in recent decades toward understanding the expression and functions of specific miRNAs in the establishment and maintenance of pluripotency. Here, we summarize the recent knowledge regarding the regulatory roles of miRNAs in self-renewal of pluripotent ESCs and during cellular reprogramming, as well as the potential role of miRNAs in two distinct pluripotent states (na�ve and primed).

Aberrant DNA methylation reprogramming during induced pluripotent stem cell generation is dependent on the choice of reprogramming factors

Volume 3, Issue 1, 二月 2014, Page
Aline C. Planello | Aline C. Planello | Junfeng Ji | Junfeng Ji | Vivek Sharma | Vivek Sharma | Rajat Singhania | Faridah Mbabaali | Fabian Müller | Javier A. Alfaro | Javier A. Alfaro | Christoph Bock | Christoph Bock | Christoph Bock | Daniel D. De Carvalho | Daniel D. De Carvalho | Nizar N. Batada | Nizar N. Batada

© 2014 Planello et al.; licensee BioMed Central Ltd. The conversion of somatic cells into pluripotent stem cells via overexpression of reprogramming factors involves epigenetic remodeling. DNA methylation at a significant proportion of CpG sites in induced pluripotent stem cells (iPSCs) differs from that of embryonic stem cells (ESCs). Whether different sets of reprogramming factors influence the type and extent of aberrant DNA methylation in iPSCs differently remains unknown. In order to help resolve this critical question, we generated human iPSCs from a common fibroblast cell source using either the Yamanaka factors (OCT4, SOX2, KLF4 and cMYC) or the Thomson factors (OCT4, SOX2, NANOG and LIN28), and determined their genome-wide DNA methylation profiles. In addition to shared DNA methylation aberrations present in all our iPSCs, we identified Yamanaka-iPSC (Y-iPSC)-specific and Thomson-iPSC (T-iPSC)-specific recurrent aberrations. Strikingly, not only were the genomic locations of the aberrations different but also their types: reprogramming with Yamanaka factors mainly resulted in failure to demethylate CpGs, whereas reprogramming with Thomson factors mainly resulted in failure to methylate CpGs. Differences in the level of transcripts encoding DNMT3b and TET3 between Y-iPSCs and T-iPSCs may contribute partially to the distinct types of aberrations. Finally, de novo aberrantly methylated genes in Y-iPSCs were enriched for NANOG targets that are also aberrantly methylated in some cancers. Our study thus reveals that the choice of reprogramming factors influences the amount, location, and class of DNA methylation aberrations in iPSCs. These findings may provide clues into how to produce human iPSCs with fewer DNA methylation abnormalities.

Stem cell therapy for amyotrophic lateral sclerosis

Volume 4, Issue 1, 十一月 2015, Page
Zhijuan Mao | Suming Zhang | Hong Chen

� 2015 Mao et al. Amyotrophic lateral sclerosis (ALS) is a fatal neurodegenerative disorder characterized by the loss of motor neurons. Currently, no effective the rapy is available to treat ALS, except for Riluzole, which has only limited clinical benefits. Stem-cell-based therapy has been intensively and extensively studied as a potential novel treatment strategy for ALS and has been shown to be effective, at least to some extent. In this article, we will review the current state of research on the use of stem cell therapy in the treatment of ALS and discuss the most promising stem cells for the treatment of ALS.

GATA2-/-human ESCs undergo attenuated endothelial to hematopoietic transition and thereafter granulocyte commitment

Volume 4, Issue 1, 八月 2015, Page
Ke Huang | Ke Huang | Juan Du | Juan Du | Ning Ma | Ning Ma | Jiajun Liu | Pengfei Wu | Pengfei Wu | Xiaoya Dong | Minghui Meng | Minghui Meng | Wenqian Wang | Xin Chen | Xin Chen | Xi Shi | Xi Shi | Qianyu Chen | Qianyu Chen | Zhongzhou Yang | Zhongzhou Yang | Shubin Chen | Shubin Chen | Jian Zhang | Jian Zhang | Yuhang Li | Wei Li | Wei Li | Yi Zheng | Yi Zheng | Jinglei Cai | Jinglei Cai | Peng Li | Peng Li | Xiaofang Sun | Xiaofang Sun | Jinyong Wang | Jinyong Wang | Duanqing Pei | Duanqing Pei | Guangjin Pan | Guangjin Pan

© 2015 Huang et al. Background: Hematopoiesis is a progressive process collectively controlled by an elaborate network of transcription factors (TFs). Among these TFs, GATA2 has been implicated to be critical for regulating multiple steps of hematopoiesis in mouse models. However, whether similar function of GATA2 is conserved in human hematopoiesis, especially during early embryonic development stage, is largely unknown. Results: To examine the role of GATA2 in human background, we generated homozygous GATA2 knockout human embryonic stem cells (GATA2 -/- hESCs) and analyzed their blood differentiation potential. Our results demonstrated that GATA2 -/- hESCs displayed attenuated generation of CD34 + CD43 + hematopoietic progenitor cells (HPCs), due to the impairment of endothelial to hematopoietic transition (EHT). Interestingly, GATA2 -/- hESCs retained the potential to generate erythroblasts and macrophages, but never granulocytes. We further identified that SPI1 downregulation was partially responsible for the defects of GATA2 -/- hESCs in generation of CD34 + CD43 + HPCs and granulocytes. Furthermore, we found that GATA2 -/- hESCs restored the granulocyte potential in the presence of Notch signaling. Conclusion: Our findings revealed the essential roles of GATA2 in EHT and granulocyte development through regulating SPI1, and uncovered a role of Notch signaling in granulocyte generation during hematopoiesis modeled by human ESCs.

Production of transgenic pigs over-expressing the antiviral gene Mx1

Volume 3, Issue 1, 五月 2014, Page
Quanmei Yan | Huaqiang Yang | Dongshan Yang | Bentian Zhao | Zhen Ouyang | Zhaoming Liu | Nana Fan | Hongsheng Ouyang | Weiwang Gu | Liangxue Lai | Liangxue Lai

© 2014 Yan et al.; licensee BioMed Central Ltd. The myxovirus resistance gene (Mx1) has a broad spectrum of antiviral activities. It is therefore an interesting candidate gene to improve disease resistance in farm animals. In this study, we report the use of somatic cell nuclear transfer (SCNT) to produce transgenic pigs over-expressing the Mx1 gene. These transgenic pigs express approximately 15-25 times more Mx1 mRNA than non-transgenic pigs, and the protein level of Mx1 was also markedly enhanced. We challenged fibroblast cells isolated from the ear skin of transgenic and control pigs with influenza A virus and classical swine fever virus (CFSV). Indirect immunofluorescence assay (IFA) revealed a profound decrease of influenza A proliferation in Mx1 transgenic cells. Growth kinetics showed an approximately 10-fold reduction of viral copies in the transgenic cells compared to non-transgenic controls. Additionally, we found that the Mx1 transgenic cells were more resistant to CSFV infection in comparison to non-transgenic cells. These results demonstrate that the Mx1 transgene can protect against viral infection in cells of transgenic pigs and indicate that the Mx1 transgene can be harnessed to develop disease-resistant pigs.

Combined influence of basal media and fibroblast growth factor on the expansion and differentiation capabilities of adipose-derived stem cells

Volume 3, Issue 1, 十一月 2014, Page
Mark Ahearne | Mark Ahearne | Joanne Lysaght | Amy P. Lynch | Amy P. Lynch

© 2014 Ahearne et al.; licensee BioMed Central Ltd. Background: Interest in adipose-derived stem cells (ASCs) has increased in recent years due to their multi-linage differentiation capabilities. While much work has been done to optimize the differentiation media, few studies have focused on examining the influence of different expansion media on cell behavior. In this study, three basal media (low glucose Dulbecco's modified Eagle's medium (DMEM), high glucose DMEM and DMEM-F12) supplemented with or without fibroblast growth factor 2 (FGF) were examined to assess their suitability for expanding ASCs. Findings: Flow cytometry, colony-forming unit assays (CFU-Fs) and differentiation assays were utilized to study cell behavior. High glucose media CFU-Fs produced fewest colonies while the addition of FGF increased colony size. By passage 2, the majority of cells were positive for CD44, 45, 73, 90 and 105 and negative for CD14, 31 and 45, indicating a mesenchymal phenotype. A sub-population of CD34 positive cells was present among passage 2 cells; however, by passage 4 the cells were negative for CD34. FGF has a negative effective on passage 4 ASC adipogenesis and high glucose media plus FGF-enhanced osteogenic capacity of passage 4 ASCs. FGF supplemented basal media were most suitable for chondrogenesis. High glucose media plus FGF appeared to be the most beneficial for priming ASCs to induce a keratocyte phenotype. Conclusions: These findings demonstrate the reciprocal effect FGF and basal media have on ASCs. This research has implications for those interested regenerating bone, cartilage, cornea or adipose tissues.

Reprogramming somatic cells to cells with neuronal characteristics by defined medium both in vitro and in vivo

Volume 4, Issue 1, 十二月 2015, Page
Songwei He | Yiping Guo | Yixin Zhang | Yuan Li | Chengqian Feng | Xiang Li | Lilong Lin | Lin Guo | Haitao Wang | Chunhua Liu | Yi Zheng | Chuanming Luo | Qiang Liu | Fuhui Wang | Hao Sun | Lining Liang | Lingyu Li | Lingyu Li | Huanxing Su | Jiekai Chen | Duanqing Pei | Hui Zheng

© 2015 He et al. Background: Currently, direct conversion from somatic cells to neurons requires virus-mediated delivery of at least one transcriptional factor or a combination of several small-molecule compounds. Delivery of transcriptional factors may affect genome stability, while small-molecule compounds may require more evaluations when applied in vivo. Thus, a defined medium with only conventional growth factors or additives for cell culture is desirable for inducing neuronal trans-differentiation. Results: Here, we report that a defined medium (5C) consisting of basic fibroblast growth factor (bFGF), N2 supplement, leukemia inhibitory factor, vitamin C (Vc), and β-mercaptoethanol (βMe) induces the direct conversion of somatic cells to cells with neuronal characteristics. Application of 5C medium converted mouse embryonic fibroblasts (MEFs) into TuJ+ neuronal-like cells, which were capable of survival after being transplanted into the mouse brain. The same 5C medium could convert primary rat astrocytes into neuronal-like cells with mature electrophysiology characteristics in vitro and facilitated the recovery of brain injury, possibly by inducing similar conversions, when infused into the mouse brain in vivo. Crucially, 5C medium could also induce neuronal chara cteristics in several human cell types. Conclusions: In summary, this 5C medium not only provides a means to derive cells with neuronal characteristics without viral transfection in vitro but might also be useful to produce neurons in vivo for neurodegenerative disease treatment.

Exploring the utility of organo-polyoxometalate hybrids to inhibit SOX transcription factors

Volume 3, Issue 1, 七月 2014, Page
Kamesh Narasimhan | Kamesh Narasimhan | Kevin Micoine | Emmanuel Lacôte | Serge Thorimbert | Edwin Cheung | Edwin Cheung | Bernold Hasenknopf | Bernold Hasenknopf | Ralf Jauch | Ralf Jauch

© 2014 Narasimhan et al.; licensee BioMed Central Ltd. Background: SOX transcription factors constitute an attractive target class for intervention with small molecules as they play a prominent role in the field of regenerative biomedicine and cancer biology. However, rationally engineering specific inhibitors that interfere with transcription factor DNA interfaces continues to be a monumental challenge in the field of transcription factor chemical biology. Polyoxometalates (POMs) are inorganic compounds that were previously shown to target the high-mobility group (HMG) of SOX proteins at nanomolar concentrations. In continuation of this work, we carried out an assessment of the selectivity of a panel of newly synthesized organo-polyoxometalate hybrids in targeting different transcription factor families to enable the usage of polyoxometalates as specific SOX transcription factor drugs.Results: The residual DNA-binding activities of 15 different transcription factors were measured after treatment with a panel of diverse polyoxometalates. Polyoxometalates belonging to the Dawson structural class were found to be more potent inhibitors than the Keggin class. Further, organically modified Dawson polyoxometalates were found to be the most potent in inhibiting transcription factor DNA binding activity. The size of the polyoxometalates and its derivitization were found to be the key determinants of their potency.Conclusion: Polyoxometalates are highly potent, nanomolar range inhibitors of the DNA binding activity of the Sox-HMG family. However, binding assays involving a limited subset of structurally diverse polyoxometalates revealed a low selectivity profile against different transcription factor families. Further progress in achieving selectivity and deciphering structure-activity relationship of POMs require the identification of POM binding sites on transcription factors using elaborate approaches like X-ray crystallography and multidimensional NMR. In summary, our report reaffirms that transcription factors are challenging molecular architectures and that future polyoxometalate chemistry must consider further modification strategies, to address the substantial challenges involved in achieving target selectivity.

Cohesin complexes with a potential to link mammalian meiosis to cancer

Volume 2, Issue 1, 六月 2013, Page
Alexander Strunnikov

� 2013 Strunnikov; licensee BioMed Central Ltd. Among multiple genes aberrantly activated in cancers, invariably, there is a group related to the capacity of cell to self-renewal. Some of these genes are related to the normal process of development, including the establishment of a germline. This group, a part of growing family of Cancer/Testis (CT) genes, now includes the meiosis specific subunits of cohesin complex. The first reports characterizing the SMC1 and RAD21 genes, encoding subunits of cohesin, were published 20 years ago; however the exact molecular mechanics of cohesin molecular machine in vivo remains rather obscure notwithstanding ample elegant experiments. The matters are complicated by the fact that the evolution of cohesin function, which is served by just two basic types of protein complexes in budding yeast, took an explosive turn in Metazoa. The recent characterization of a new set of genes encoding cohesin subunits specific for meiosis in vertebrates adds several levels of complexity to the task of structure-function analysis of specific cohesin pathways, even more so in relation to their aberrant functionality in cancers. These three proteins, SMC1β, RAD21L and STAG3 are likely involved in a specific function in the first meiotic prophase, genetic recombination, and segregation of homologues. However, at present, it is rather challenging to pinpoint the molecular role of these proteins, particularly in synaptonemal complex or centromere function, due to the multiplicity of different cohesins in meiosis. The roles of these proteins in cancer cell physiology, upon their aberrant activation in tumors, also remain to be elucidated. Nevertheless, as the existence of Cancer/Testis cohesin complexes in tumor cells appears to be all but certain, this brings a promise of a new target for cancer therapy and/or diagnostics.

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