POU5F1 Gene Expression Research Articles

Comparison of SOX2 and POU5F1 Gene Expression in Leukapheresis-Derived CD34+ Cells before and during Cell Culture

Bone marrow is an abundant source of both hematopoietic as well as non-hematopoietic stem cells. Embryonic, fetal and stem cells located in tissues (adipose tissue, skin, myocardium and dental pulp) express core transcription factors, including the SOX2, POU5F1 and NANOG gene responsible for regeneration, proliferation and differentiation into daughter cells. The aim of the study was to examine the expression of SOX2 and POU5F1 genes in CD34-positive peripheral blood stem cells (CD34+ PBSCs) and to analyze the influence of cell culture on the expression of SOX2 and POU5F1 genes. The study material consisted of bone marrow-derived stem cells isolated by using leukapheresis from 40 hematooncology patients. Cells obtained in this process were subject to cytometric analysis to determine the content of CD34+ cells. CD34-positive cell separation was conducted using MACS separation. Cell cultures were set, and RNA was isolated. Real-time PCR was conducted in order to evaluate the expression of SOX2 and POU5F1 genes and the obtained data were subject to statistical analysis. We identified the expression of SOX2 and POU5F1 genes in the examined cells and demonstrated a statistically significant (p < 0.05) change in their expression in cell cultures. Short-term cell cultures (<6 days) were associated with an increase in the expression of SOX2 and POU5F1 genes. Thus, short-term cultivation of transplanted stem cells could be used to induce pluripotency, leading to better therapeutic effects.

Comparison of POU5F1 gene expression and protein localization in two differentiated and undifferentiated spermatogonial stem cells.

The POU domain, class 5, transcription factor 1 (POU5F1), plays a vital role in creating pluripotency and maintaining self-renewal of the spermatogonial stem cells (SSCs). In this experimental research, the gene and protein expression of POU5F1 in two populations of differentiated and undifferentiated spermatogonia were examined, by immunohistochemistry (IMH), immunocytochemistry (ICC) and Fluidigm real-time RT-PCR. Our study was extended with online databases and the creation of PPI networks. The results indicated that the POU5F1 protein was localized in the basal compartment of seminiferous tubules. Under in vitro conditions, isolated SSC colonies were ICC-positive for the POU5F1, but the protein expression level of POU5F1 in the undifferentiated populations was higher than that in differentiated. A significant POU5F1 mRNA expression was seen in passage 4 compared to passage 0 for both populations. POU5F1 has a significantly higher mRNA expression in undifferentiated SSCs than that in differentiated SSCs, also in mESCs than in SSC-like cells. Bioinformatic analysis on POU5F1 shows its impressive connection with other genes involved in spermatogonia differentiation. These results support the advanced investigations of spermatogonia differentiation, both in vitro and in vivo. A better understanding of the POU5F1 gene and its function during differentiation will give the scientific community an open perspective for the development of direct differentiation of SSC to other male germline cells which is very important in infertility treatment.

Influence of the Type of Delivery, Use of Oxytocin, and Maternal Age on POU5F1 Gene Expression in Stem Cells Derived from Wharton’s Jelly within the Umbilical Cord

The paper presents an evaluation of the POU5F1 gene expression in mesenchymal stem cells derived from Wharton's jelly within the umbilical cord, collected from 36 patients during labor. The study is the first one to show that the expression of POU5F1 in mesenchymal stem cells has been dependent on maternal age, birth order, route of delivery, and use of oxytocin. Our research proves that the POU5F1 gene expression in mesenchymal stem cells decreases with each subsequent pregnancy and delivery. Wharton's jelly stem cells obtained from younger women and during their first delivery, as well as patients treated with oxytocin, show higher POU5F1 gene expression when compared with the subsequent deliveries. This leads to a conclusion that they are characterized by a lower level of differentiation, which in turn results in their greater plasticity and greater proliferative potential. Probably, they are also clinically more useful.

Abstract 3687: Colorectal cancer stem cell expressing POU5F1 promotes liver metastasis

Abstract Background: The gene POU5F1, encoding the POU domain, class 5, is a transcription factor and the expression is found in embryonic stem cells. POU5F1 gene controls self-renewal and differentiation, and regulates pluripotency and proliferation in embryonic stem cells. However, the role of POU5F1 in metastatic CRC (mCRC) is not well known. We focused on the relationship between POU5F1 gene expression and liver metastasis of CRC and clarify the role and abilities of POU5F1-positive CRC cells. Methods: The study included 158 patients with CRC who underwent surgery from 2009 to 2011. The correlations between the POU5F1 gene expression and the clinical parameters were assessed, and liver-metastasis-free survival (LMFS) was evaluated in these patients. POU5F1-EGFP-positive cells were established by the construct with lentiviral vector, and we examine their sub-population and ability. The capacity to form liver metastasis in vivo was examined using CRC cell lines and primary cultured CRC cells. Results: In the survival analysis, LMFS was significantly poor in the high POU5F1 expression group compared to the low-expression group (P=0.008). Multivariate analyses showed that POU5F1 expression (P=0.015) and TNM stage (p&amp;lt;0.001) significantly correlated with LMFS. POU5F1-EGFP-positive cells highly expressed stem-cell-associated markers and they had self-renewal and differentiation abilities. POU5F1-high cells actively formed liver metastasis. Conclusion: The POU5F1expression correlated with liver metastasis in CRC patients. POU5F1-positive cells have the self-renewal and differentiation abilities as cancer stem cells. POU5F1 contributes to the ability forming liver metastasis in CRC. Citation Format: Norikatsu Miyoshi, Shiki Fujino, Masaru Sasaki, Kazuhiro Saso, Hidekazu Takahashi, Naotsugu Haraguchi, Taishi Hata, Chu Matsuda, Tsunekazu Mizushima, Masaki Mori, Yuichiro Doki. Colorectal cancer stem cell expressing POU5F1 promotes liver metastasis [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2019; 2019 Mar 29-Apr 3; Atlanta, GA. Philadelphia (PA): AACR; Cancer Res 2019;79(13 Suppl):Abstract nr 3687.

The POU5F1 gene expression in colorectal cancer: a novel prognostic marker.

The POU5F1 gene, which encodes the POU domain, class 5, transcription factor 1 (also known as Oct-4), is expressed in embryonic stem cells where it regulates pluripotency and proliferation. Few studies have examined the expression and significance of POU5F1 in cancer tissues. The aim of this study was to clarify its significance in colorectal cancer (CRC). The study included 95 patients who underwent surgery for CRC from 2009 to 2011. The correlations between the POU5F1 gene expression and the clinical parameters were assessed in these patients. The serum TP53 expression levels were also examined by an enzyme-linked immunosorbent assay. Patients with a high POU5F1 expression were significantly more likely to have a poor prognosis than those with a low expression. In contrast, patients with a low POU5F1 expression had a better disease-free survival after curative surgical resection than those with a high expression (P = 0.007). Multivariate analyses showed that the POU5F1 expression (P = 0.003) and lymph node metastasis (P < 0.001) were significantly correlated with the disease-free survival. The prognosis was significantly related to the serum TP53 levels, according to the POU5F1 expression. The POU5F1 expression was suggested to be a prognostic factor in patients with CRC.

Delayed blastocyst formation or an extra day culture increases apoptosis in pig blastocysts

In the present study, the timing was examined of blastocyst collection/formation or of how the duration of post-blastulation culture affected the quality and developmental competence of in vitro-produced pig parthenogenetic embryos. The earliest apoptotic signals were observed at the morula stage while the earliest cytoplasmic fragmentation was observed before the 4- to 8-cell stage of embryo development. Nuclear condensation was detected in morulae and blastocysts, but not all condensed nuclei were positive for the apoptotic signal (TUNEL staining). The mean blastocyst diameter increased with delayed blastocyst collection or extended post-blastulation culture, but decreased with delayed blastocyst formation. Delayed blastocyst collection/formation or an additional day of post-blastulation culture increased the frequencies of apoptosis, condensed nuclei, and low quality blastocysts (those showing a nuclear destruction that negated counting of the nuclei); increased the expression of the pro-apoptotic BAX gene; and reduced the ratio of ICM (inner cell mass) cells to TE (trophectoderm) cells. In addition, delayed blastocyst formation decreased POU5F1 gene expression. These results suggest that a delay in blastocyst collection/formation or an additional day of culture could increase the incidence of apoptosis, decrease the ICM:TE cell ratio, and influence the gene expression and diameter of blastocysts derived from in vitro-produced pig embryos. These findings provide a useful reference for improving the quality of in vitro-produced embryos.

201P POU5F1 gene expression in colorectal cancer: a novel prognostic marker after curative surgical resection

201P POU5F1 gene expression in colorectal cancer: a novel prognostic marker after curative surgical resection

201P POU5F1 gene expression in colorectal cancer: a novel prognostic marker after curative surgical resection

201P POU5F1 gene expression in colorectal cancer: a novel prognostic marker after curative surgical resection

Supplementation of bovine embryo culture medium with L-arginine improves embryo quality via nitric oxide production.

Nitric oxide (NO) is a cell-signaling molecule that regulates a variety of molecular pathways. We investigated the role of NO during preimplantation embryonic development by blocking its production with an inhibitor or supplementing in vitro bovine embryo cultures with its natural precursor, L-arginine, over different periods. Endpoints evaluated included blastocyst rates, development kinetics, and embryo quality. Supplementation with the NO synthase inhibitor N-Nitro-L-arginine-methyl ester (L-NAME) from Days 1 to 8 of culture decreased blastocyst (P < 0.05) and hatching (P < 0.05) rates. When added from Days 1 to 8, 50 mM L-arginine decreased blastocyst rates (P < 0.001); in contrast, when added from Days 5 to 8, 1 mM L-arginine improved embryo hatching rates (P < 0.05) and quality (P < 0.05) as well as increased POU5F1 gene expression (P < 0.05) as compared to the untreated control. Moreover, NO levels in the medium during this culture period positively correlated with the increased embryo hatching rates and quality (P < 0.05). These data suggest exerts its positive effects during the transition from morula to blastocyst stage, and that supplementing the embryo culture medium with L-arginine favors preimplantation development of bovine embryos.

The Pou5f1 distal enhancer is sufficient to drive Pou5f1 promoter-EGFP expression in embryonic stem cells

The POU5F1 transcription factor is the gatekeeper of the pluripotent state in mammals. It is essential for epigenetic reprogramming events and also for germ cell viability. Pou5f1 gene expression is tightly controlled during embryogenesis, but its regulatory regions are not fully deciphered. The GOF18ΔPE-EGFP transgene, harboring the enhanced green fluorescence protein reporter gene inserted into a 17- kilobase long mouse Pou5f1 genomic sequence, has been widely used to visualize pluripotent embryonic cells and primordial germ cells in the mouse and other mammalian species. This construct includes the Pou5f1 promoter under the control of the distal enhancer and also includes the Pou5f1 gene body and flanking sequences. In search of the essential regulatory regions of Pou5f1, we generated four shorter forms of this construct. We found that the shortest form, containing the Pou5f1 promoter and distal enhancer but lacking the gene body and upstream flanking sequences, correctly expressed EGFP in transiently transformed undifferentiated ES cells, correctly switched it off upon ES cell differentiation, and correctly kept it silenced in differentiated Hep3B cells. Similarly to the original GOF18ΔPE-EGFP, this shortest form was expressed in the fetal mouse gonad. Our data suggest that the Pou5f1 distal enhancer and proximal promoter may be sufficient to specify transgene expression in pluripotent cells.

Probiotics Can Induce Follicle Maturational Competence: The Danio rerioCase1

In the present study, the effects of the probiotic Lactobacillus rhamnosus IMC 501 on the acquisition of oocyte maturational competence was examined in zebrafish (Danio rerio). L. rhamnosus administration induced the responsiveness of incompetent follicles (stage IIIa) to 17,20-dihydroxy-4-pregnen-3-one and their in vitro maturation. Acquisition of competence by the stage IIIa follicles was further validated by changes of lhr, mprb, inhbaa (activin betaA1), tgfb1, and gdf9 gene expression, which have recently emerged as key regulators of oocyte acquisition of maturational competence, and pou5f1 gene expression, which in other models has been shown to govern the establishment of developmental competence of oocytes. In addition, a DNA microarray experiment was conducted using the same follicles, and with relative gene ontology (GO) data analysis, the molecular effects of probiotic administration emerged. Molecular analysis using PCR-DGGE (denaturing gradient gel electrophoresis) approach, providing information about only the most abundant bacterial members of the microbial community, revealed that the probiotic was able to populate the gastrointestinal tract and modulate the microbial communities, causing a clear shift in them and specifically enhancing the presence of the lactic acid bacteria Streptococcus thermophilus. At the same time, PCR-DGGE analysis revealed that the probiotic was not directly associated with the ovaries. Finally, the effects of probiotic treatment on zebrafish follicle development were also analyzed by FPA (focal plane array) Fourier transform-infrared (FT-IR) imaging, a technique that provides the overall biochemical composition of samples. Changes were found above all in stage IIIa follicles from probiotic-exposed females; the modifications, observed in protein secondary structures as well as in hydration and in bands related to phosphate moieties, allowed us to hypothesize that probiotics act at this follicle stage, affecting the maturation phase.

B-MYB Is Essential for Normal Cell Cycle Progression and Chromosomal Stability of Embryonic Stem Cells

BackgroundThe transcription factor B-Myb is present in all proliferating cells, and in mice engineered to remove this gene, embryos die in utero just after implantation due to inner cell mass defects. This lethal phenotype has generally been attributed to a proliferation defect in the cell cycle phase of G1.Methodology/Principal FindingsIn the present study, we show that the major cell cycle defect in murine embryonic stem (mES) cells occurs in G2/M. Specifically, knockdown of B-Myb by short-hairpin RNAs results in delayed transit through G2/M, severe mitotic spindle and centrosome defects, and in polyploidy. Moreover, many euploid mES cells that are transiently deficient in B-Myb become aneuploid and can no longer be considered viable. Knockdown of B-Myb in mES cells also decreases Oct4 RNA and protein abundance, while over-expression of B-MYB modestly up-regulates pou5f1 gene expression. The coordinated changes in B-Myb and Oct4 expression are due, at least partly, to the ability of B-Myb to directly modulate pou5f1 gene promoter activity in vitro. Ultimately, the loss of B-Myb and associated loss of Oct4 lead to an increase in early markers of differentiation prior to the activation of caspase-mediated programmed cell death.Conclusions/SignificanceAppropriate B-Myb expression is critical to the maintenance of chromosomally stable and pluripotent ES cells, but its absence promotes chromosomal instability that results in either aneuploidy or differentiation-associated cell death.