Cell Regeneration最新文献

{"title":"Combined influence of basal media and fibroblast growth factor on the expansion and differentiation capabilities of adipose-derived stem cells","authors":"Mark Ahearne ,&nbsp;Joanne Lysaght ,&nbsp;Amy P Lynch","doi":"10.1186/2045-9769-3-13","DOIUrl":"10.1186/2045-9769-3-13","url":null,"abstract":"<div><h3>Background</h3><p>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.</p></div><div><h3>Findings</h3><p>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.</p></div><div><h3>Conclusions</h3><p>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.</p></div>","PeriodicalId":9811,"journal":{"name":"Cell Regeneration","volume":"3 1","pages":"Article 3:13"},"PeriodicalIF":0.0,"publicationDate":"2014-01-01","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"https://sci-hub-pdf.com/10.1186/2045-9769-3-13","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"33145248","PeriodicalName":null,"FirstCategoryId":null,"ListUrlMain":null,"RegionNum":0,"RegionCategory":"","ArticlePicture":[],"TitleCN":null,"AbstractTextCN":null,"PMCID":"OA","EPubDate":null,"PubModel":null,"JCR":null,"JCRName":null,"Score":null,"Total":0}

{"title":"Generation of knockout rabbits using transcription activator-like effector nucleases","authors":"Yu Wang ,&nbsp;Nana Fan ,&nbsp;Jun Song ,&nbsp;Juan Zhong ,&nbsp;Xiaogang Guo ,&nbsp;Weihua Tian ,&nbsp;Quanjun Zhang ,&nbsp;Fenggong Cui ,&nbsp;Li Li ,&nbsp;Philip N Newsome ,&nbsp;Jon Frampton ,&nbsp;Miguel A Esteban ,&nbsp;Liangxue Lai","doi":"10.1186/2045-9769-3-3","DOIUrl":"10.1186/2045-9769-3-3","url":null,"abstract":"<div><p>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.</p></div>","PeriodicalId":9811,"journal":{"name":"Cell Regeneration","volume":"3 1","pages":"Article 3:3"},"PeriodicalIF":0.0,"publicationDate":"2014-01-01","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"https://sci-hub-pdf.com/10.1186/2045-9769-3-3","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"32824474","PeriodicalName":null,"FirstCategoryId":null,"ListUrlMain":null,"RegionNum":0,"RegionCategory":"","ArticlePicture":[],"TitleCN":null,"AbstractTextCN":null,"PMCID":"OA","EPubDate":null,"PubModel":null,"JCR":null,"JCRName":null,"Score":null,"Total":0}

{"title":"glbase: a framework for combining, analyzing and displaying heterogeneous genomic and high-throughput sequencing data","authors":"Andrew Paul Hutchins ,&nbsp;Ralf Jauch ,&nbsp;Mateusz Dyla ,&nbsp;Diego Miranda-Saavedra","doi":"10.1186/2045-9769-3-1","DOIUrl":"10.1186/2045-9769-3-1","url":null,"abstract":"<div><p>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 <span>http://bitbucket.org/oaxiom/glbase/</span><svg><path></path></svg>.</p></div>","PeriodicalId":9811,"journal":{"name":"Cell Regeneration","volume":"3 1","pages":"Article 3:1"},"PeriodicalIF":0.0,"publicationDate":"2014-01-01","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"https://sci-hub-pdf.com/10.1186/2045-9769-3-1","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"32824082","PeriodicalName":null,"FirstCategoryId":null,"ListUrlMain":null,"RegionNum":0,"RegionCategory":"","ArticlePicture":[],"TitleCN":null,"AbstractTextCN":null,"PMCID":"OA","EPubDate":null,"PubModel":null,"JCR":null,"JCRName":null,"Score":null,"Total":0}

{"title":"Characterization of a novel cell penetrating peptide derived from human Oct4","authors":"Eva Harreither ,&nbsp;Hanna A Rydberg ,&nbsp;Helene L Åmand ,&nbsp;Vaibhav Jadhav ,&nbsp;Lukas Fliedl ,&nbsp;Christina Benda ,&nbsp;Miguel A Esteban ,&nbsp;Duanqing Pei ,&nbsp;Nicole Borth ,&nbsp;Regina Grillari-Voglauer ,&nbsp;Oliver Hommerding ,&nbsp;Frank Edenhofer ,&nbsp;Bengt Nordén ,&nbsp;Johannes Grillari","doi":"10.1186/2045-9769-3-2","DOIUrl":"10.1186/2045-9769-3-2","url":null,"abstract":"<div><h3>Background</h3><p>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.</p></div><div><h3>Results</h3><p>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.</p></div><div><h3>Conclusions</h3><p>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.</p><p>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.</p></div>","PeriodicalId":9811,"journal":{"name":"Cell Regeneration","volume":"3 1","pages":"Article 3:2"},"PeriodicalIF":0.0,"publicationDate":"2014-01-01","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"https://sci-hub-pdf.com/10.1186/2045-9769-3-2","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"32824084","PeriodicalName":null,"FirstCategoryId":null,"ListUrlMain":null,"RegionNum":0,"RegionCategory":"","ArticlePicture":[],"TitleCN":null,"AbstractTextCN":null,"PMCID":"OA","EPubDate":null,"PubModel":null,"JCR":null,"JCRName":null,"Score":null,"Total":0}

{"title":"Generation of multi-gene knockout rabbits using the Cas9/gRNA system","authors":"Quanmei Yan ,&nbsp;Quanjun Zhang ,&nbsp;Huaqiang Yang ,&nbsp;Qingjian Zou ,&nbsp;Chengcheng Tang ,&nbsp;Nana Fan ,&nbsp;Liangxue Lai","doi":"10.1186/2045-9769-3-12","DOIUrl":"10.1186/2045-9769-3-12","url":null,"abstract":"<div><p>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.</p></div>","PeriodicalId":9811,"journal":{"name":"Cell Regeneration","volume":"3 1","pages":"Article 3:12"},"PeriodicalIF":0.0,"publicationDate":"2014-01-01","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"https://sci-hub-pdf.com/10.1186/2045-9769-3-12","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"32824482","PeriodicalName":null,"FirstCategoryId":null,"ListUrlMain":null,"RegionNum":0,"RegionCategory":"","ArticlePicture":[],"TitleCN":null,"AbstractTextCN":null,"PMCID":"OA","EPubDate":null,"PubModel":null,"JCR":null,"JCRName":null,"Score":null,"Total":0}

{"title":"Genetic reporter analysis reveals an expandable reservoir of OCT4+ cells in adult skin","authors":"Anne Limbourg ,&nbsp;Sabine Schnabel ,&nbsp;Vladimir J Lozanovski ,&nbsp;L Christian Napp ,&nbsp;Teng-Cheong Ha ,&nbsp;Tobias Maetzig ,&nbsp;Johann Bauersachs ,&nbsp;Hassan Y Naim ,&nbsp;Axel Schambach ,&nbsp;Florian P Limbourg","doi":"10.1186/2045-9769-3-9","DOIUrl":"10.1186/2045-9769-3-9","url":null,"abstract":"<div><p>The transcription factor <em>Oct4</em> (<em>Pou5f1</em>) is a critical regulator of pluripotency in embryonic and induced pluripotent stem cells. Therefore, <em>Oct4</em> expression might identify somatic stem cell populations with inherent multipotent potential or a propensity for facilitated reprogramming. However, analysis of <em>Oct4</em> expression is confounded by <em>Oct4</em> pseudogenes or non-pluripotency-related isoforms. Systematic analysis of a transgenic <em>Oct4-EGFP</em> reporter mouse identified testis and skin as two principle sources of <em>Oct4</em>⁺ cells in postnatal mice. While the prevalence of GFP⁺ cells in testis rapidly declined with age, the skin-resident GFP⁺ population expanded in a cyclical fashion. These cells were identified as epidermal stem cells dwelling in the stem cell niche of the hair follicle, which endogenously expressed all principle reprogramming factors at low levels. Interestingly, skin wounding or non-traumatic hair removal robustly expanded the GFP⁺ epidermal cell pool not only locally, but also in uninjured skin areas, demonstrating the existence of a systemic response. Thus, the epithelial stem cell niche of the hair follicle harbors an expandable pool of Oct4+ stem cells, which might be useful for therapeutic cell transfer or facilitated reprogramming.</p></div>","PeriodicalId":9811,"journal":{"name":"Cell Regeneration","volume":"3 1","pages":"Article 3:9"},"PeriodicalIF":0.0,"publicationDate":"2014-01-01","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"https://sci-hub-pdf.com/10.1186/2045-9769-3-9","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"32824480","PeriodicalName":null,"FirstCategoryId":null,"ListUrlMain":null,"RegionNum":0,"RegionCategory":"","ArticlePicture":[],"TitleCN":null,"AbstractTextCN":null,"PMCID":"OA","EPubDate":null,"PubModel":null,"JCR":null,"JCRName":null,"Score":null,"Total":0}

{"title":"A mitochondrial strategy for safeguarding the reprogrammed genome","authors":"Alessandro Prigione ,&nbsp;James Adjaye","doi":"10.1186/2045-9769-3-5","DOIUrl":"10.1186/2045-9769-3-5","url":null,"abstract":"<div><p>Genomic aberrations induced by somatic cell reprogramming are a major drawback for future applications of this technology in regenerative medicine. A new study by Ji et al. published in Stem Cell Reports suggests a counteracting strategy based on balancing the mitochondrial/oxidative stress pathway through antioxidant supplementation.</p></div>","PeriodicalId":9811,"journal":{"name":"Cell Regeneration","volume":"3 1","pages":"Article 3:5"},"PeriodicalIF":0.0,"publicationDate":"2014-01-01","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"https://sci-hub-pdf.com/10.1186/2045-9769-3-5","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"32824476","PeriodicalName":null,"FirstCategoryId":null,"ListUrlMain":null,"RegionNum":0,"RegionCategory":"","ArticlePicture":[],"TitleCN":null,"AbstractTextCN":null,"PMCID":"OA","EPubDate":null,"PubModel":null,"JCR":null,"JCRName":null,"Score":null,"Total":0}

{"title":"Aberrant DNA methylation reprogramming during induced pluripotent stem cell generation is dependent on the choice of reprogramming factors","authors":"Aline C Planello ,&nbsp;Junfeng Ji ,&nbsp;Vivek Sharma ,&nbsp;Rajat Singhania ,&nbsp;Faridah Mbabaali ,&nbsp;Fabian Müller ,&nbsp;Javier A Alfaro ,&nbsp;Christoph Bock ,&nbsp;Daniel D De Carvalho ,&nbsp;Nizar N Batada","doi":"10.1186/2045-9769-3-4","DOIUrl":"10.1186/2045-9769-3-4","url":null,"abstract":"<div><p>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, <em>de novo</em> 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.</p></div>","PeriodicalId":9811,"journal":{"name":"Cell Regeneration","volume":"3 1","pages":"Article 3:4"},"PeriodicalIF":0.0,"publicationDate":"2014-01-01","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"https://sci-hub-pdf.com/10.1186/2045-9769-3-4","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"32824475","PeriodicalName":null,"FirstCategoryId":null,"ListUrlMain":null,"RegionNum":0,"RegionCategory":"","ArticlePicture":[],"TitleCN":null,"AbstractTextCN":null,"PMCID":"OA","EPubDate":null,"PubModel":null,"JCR":null,"JCRName":null,"Score":null,"Total":0}

{"title":"Exploring the utility of organo-polyoxometalate hybrids to inhibit SOX transcription factors","authors":"Kamesh Narasimhan ,&nbsp;Kevin Micoine ,&nbsp;Emmanuel Lacôte ,&nbsp;Serge Thorimbert ,&nbsp;Edwin Cheung ,&nbsp;Bernold Hasenknopf ,&nbsp;Ralf Jauch","doi":"10.1186/2045-9769-3-10","DOIUrl":"10.1186/2045-9769-3-10","url":null,"abstract":"<div><h3>Background</h3><p>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.</p></div><div><h3>Results</h3><p>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.</p></div><div><h3>Conclusion</h3><p>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.</p></div>","PeriodicalId":9811,"journal":{"name":"Cell Regeneration","volume":"3 1","pages":"Article 3:10"},"PeriodicalIF":0.0,"publicationDate":"2014-01-01","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"https://sci-hub-pdf.com/10.1186/2045-9769-3-10","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"33377331","PeriodicalName":null,"FirstCategoryId":null,"ListUrlMain":null,"RegionNum":0,"RegionCategory":"","ArticlePicture":[],"TitleCN":null,"AbstractTextCN":null,"PMCID":"OA","EPubDate":null,"PubModel":null,"JCR":null,"JCRName":null,"Score":null,"Total":0}

{"title":"OCT4: A penetrant pluripotency inducer","authors":"Xuecong Wang ,&nbsp;Ralf Jauch","doi":"10.1186/2045-9769-3-6","DOIUrl":"10.1186/2045-9769-3-6","url":null,"abstract":"<div><p>Native OCT4 protein has the intrinsic ability of crossing cellular membranes to enter cells. This finding could revive efforts to induce pluripotency with proteins replacing nucleic acid-based approaches, and raises the intriguing question as to whether OCT4 can act non-cell-autonomously.</p></div>","PeriodicalId":9811,"journal":{"name":"Cell Regeneration","volume":"3 1","pages":"Article 3:6"},"PeriodicalIF":0.0,"publicationDate":"2014-01-01","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"https://sci-hub-pdf.com/10.1186/2045-9769-3-6","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"32824477","PeriodicalName":null,"FirstCategoryId":null,"ListUrlMain":null,"RegionNum":0,"RegionCategory":"","ArticlePicture":[],"TitleCN":null,"AbstractTextCN":null,"PMCID":"OA","EPubDate":null,"PubModel":null,"JCR":null,"JCRName":null,"Score":null,"Total":0}