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Expression profiling of human adipose tissue derived stem cells cultured at various oxygen tensions

ABSTRACT: Adipose tissue is a plentiful and easily accessible source of mesenchymal stem cells that have been shown to be multi potent and possibly have immuno suppressive capacity(Zuk, Zhu et al. 2002; McIntosh, Zvonic et al. 2006; Prichard, Reichert et al. 2008). Several studies have identified hypoxia and the molecular effector of reduced oxygen tension, HIF1α to be essential in the differentiation processes, stem cell proliferation and survival as well as in maintenance of stem cell characteristics (Lennon, Edmison et al. 2001; Goda, Ryan et al. 2003; Wang, Fermor et al. 2005; Lin, Lee et al. 2006; Malladi, Xu et al. 2007; Ohnishi, Yasuda et al. 2007). The fact that oxygen has a central role in cell development and metabolism makes the regulation of oxygen tension a valuable tool in the attempt of exploiting stem cells to their full potential. In the current study we investigate the transcriptional changes caused by two weeks of monolayer hypoxic expansion of human adipose tissue derived stem cells (ASCs). Application of the XVivo hypoxic workstation (BioSpherix, Redfield, NY) made it possible to conduct the expansion of six ASC lines at four different hypoxic conditions in parallel covering the range from mild (15% oxygen) to full hypoxic (1% oxygen) conditions. In this comprehensive investigation the Illumina bead microarray platform was employed to give a profound and new insight in the molecular changes effectuated by hypoxic expansion. The findings of this study demonstrate the significance of biologic variation in the transcriptional response to mild hypoxic expansion and display evidence that full hypoxic growth conditions at 1% oxygen tension selects for a more prolific cell population of reduced chondrogenic potential. Keywords: hypoxia response

ORGANISM(S): Homo sapiens

PROVIDER: GSE12884 | GEO | 2009/09/22

SECONDARY ACCESSION(S): PRJNA110873

REPOSITORIES: GEO

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Project description:Adipose tissue is a plentiful and easily accessible source of mesenchymal stem cells that have been shown to be multi potent and possibly have immuno suppressive capacity(Zuk, Zhu et al. 2002; McIntosh, Zvonic et al. 2006; Prichard, Reichert et al. 2008). Several studies have identified hypoxia and the molecular effector of reduced oxygen tension, HIF1Î± to be essential in the differentiation processes, stem cell proliferation and survival as well as in maintenance of stem cell characteristics (Lennon, Edmison et al. 2001; Goda, Ryan et al. 2003; Wang, Fermor et al. 2005; Lin, Lee et al. 2006; Malladi, Xu et al. 2007; Ohnishi, Yasuda et al. 2007). The fact that oxygen has a central role in cell development and metabolism makes the regulation of oxygen tension a valuable tool in the attempt of exploiting stem cells to their full potential. In the current study we investigate the transcriptional changes caused by two weeks of monolayer hypoxic expansion of human adipose tissue derived stem cells (ASCs). Application of the XVivo hypoxic workstation (BioSpherix, Redfield, NY) made it possible to conduct the expansion of six ASC lines at four different hypoxic conditions in parallel covering the range from mild (15% oxygen) to full hypoxic (1% oxygen) conditions. In this comprehensive investigation the Illumina bead microarray platform was employed to give a profound and new insight in the molecular changes effectuated by hypoxic expansion. The findings of this study demonstrate the significance of biologic variation in the transcriptional response to mild hypoxic expansion and display evidence that full hypoxic growth conditions at 1% oxygen tension selects for a more prolific cell population of reduced chondrogenic potential. Keywords: hypoxia response To investigate the transcriptional response to hypoxia, adipose tissue derived stem cells (ASCs) from six female individuals were cultured for 14 days at different oxygen tensions (ambient, 15%, 10%, 5% and 1%). Gene expression levels at each oxygen tension were compared to ambient conditions (Ambient oxygen tension) using Illumina Human-6 v2.0 microarrays.

Project description:Several studies indicate that plant MIR genes are transcribed by RNA Pol II, similar to what is found in animals (Cai et al., 2004; Lee et al., 2004). By sequencing the 5' transcript ends Xie et al. (2005) mapped the transcription start sites (TSSs) for 52 MIR genes. This list was expanded upon through computational prediction of the core promoters (Zhou et al., 2007). In addition to the TATA box sequence motifs located upstream of the TSSs (Xie et al. 2005), Megraw et al. (2006) identified other transcription factor binding motifs in the promoter of Arabidopsis MIR genes. They showed that within the 800 nucleotides region upstream of TSSs, sequences resembling the binding sites for the transcription factors AtMYC2, ARF, SORLREP3, and LFY were overrepresented relative to protein-coding gene promoters and randomly sampled genomic sequences (Megraw et al. 2006). While these previous studies are instrumental in establishing our working understanding of MIR gene transcription in plants, there are several major knowledge gaps that urgently need to be addressed. Binding of RNA Pol II to the identified MIR promoter regions has not been tested. Because expression of MIR genes does not involve translation, transcriptional control by RNA Pol II might be different from protein-coding genes. In addition, few of the predicted cis elements have been functionally tested. Futher, the number of annotated miRNA genes has since increased from 199 to 232 (miRBase release 17; Kozomara and Griffiths-Jones, 2011). None of the newly identified genes have been subject to examination for promoter and regulatory sequences. Compared to the previously known miRNAs, these genes have narrower phylogenetic distribution and exhibit weaker expression level and more prominent tissue-specific expression (Rajagopalan et al., 2006; Fahlgren et al. 2007; Yang et al., 2011). Based on these observations, it has been argued that continuous gene birth and death allows beneficial miRNAs to be maintained while deleterious ones avoided (Rajagopalan et al., 2006; Fahlgren et al. 2007; Chen and Rajewsky, 2007; Axtell and Bowman, 2008). Identification of the promoter regions of these MIR genes and their comparison to those of the conserved are highly desirable to fully elucidate miRNA based gene regulation. In the current study, we performed chromatin immunoprecipitation (ChIP)-chip for the Pol II complex using the Affymetrix 'At35b_MR_v04' array. We designed a computational approach using genome-wide RPol II binding patterns to identify the promoter region and transcription start site of pri-miRNAs that are actively transcribed.

Project description:Male factors account for approximately 40% of all infertility. Conventional semen analysis focus on seminal physicochemical property and spermatozoa morphology. They yield no information concerning the functional competence of the spermatozoa(Petrunkina et al., 2007). Owning to the essential role of proteins in the reproductive process, comprehensive and systematic identification of proteome is critical to gain new insights into spermatogenesis and infertility. Recent advances in mass spectrometry have allowed the identification of hundreds to thousands of protein in spermatozoa(Oliva et al., 2008; Amaral et al., 2014; Codina et al., 2015). In major domestic mammalian species, global spermatozoa proteomic profiles have been described in rodents(Baker et al., 2008a; Baker et al., 2008b) and bovine(Peddinti et al., 2008). Meanwhile, lacking of new protein synthesis in spermatozoa supports the current view that the regulation of sperm maturation is controlled by exogenous proteins(O'Rand et al., 2011) (e.g., during epididymal transit). Among the seminal plasma proteins, The human seminal plasma has been comprehensively described (Pilch and Mann, 2006; Batruch et al., 2011; Milardi et al., 2012; Milardi et al., 2013; Sharma et al., 2013). In domestic animal species, some studies performing a systematic analysis on using high throughput proteomics have been performed(Kelly et al., 2006; Moura et al., 2007; Souza et al., 2012; Druart et al., 2013; Soleilhavoup et al., 2014). Buffalos (Bubalus bubalis) are adapted to hot–humid tropical climatic conditions, but have low reproductive efficiency. The low sperm motility maybe contributed to protein components variation of spermatozoa and seminal plasma. The composition of buffalo spermatozoa and buffalo seminal plasma (BSP) remains unknown. Proteomic study would be beneficial to the elucidation of the roles of sperm and BSP proteins in regulation of maturation, motility and fertilization. As such, the aim of the current study was to extensively characterize the differentially expressed protein of buffalo spermatozoa and seminal plasma using a comparative proteomics.

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Expression profiling of human adipose tissue derived stem cells cultured at various oxygen tensions

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