Proliferation Of Embryonic Cells Research Articles

Inhibition of T-type calcium channels disrupts Akt signaling and promotes apoptosis in glioblastoma cells

Glioblastoma multiforme (GBM) are brain tumors that are exceptionally resistant to both radio- and chemotherapy regimens and novel approaches to treatment are needed. T-type calcium channels are one type of low voltage-gated channel (LVCC) involved in embryonic cell proliferation and differentiation; however they are often over-expressed in tumors, including GBM. In this study, we found that inhibition of T-type Ca2+ channels in GBM cells significantly reduced their survival and resistance to therapy. Moreover, either T-type selective antagonists, such as mibefradil, or siRNA-mediated knockdown of the T-type channel alpha subunits not only reduced cell viability and clonogenic potential, but also induced apoptosis. In response to channel blockade or ablation, we observed reduced phosphorylation of Akt and Rictor, suggesting inhibition of the mTORC2/Akt pathway. This was followed by reduction in phosphorylation of anti-apoptotic Bad and caspases activation. The apoptotic response was specific for T-type Ca2+ channels, as inhibition of L-type Ca2+ channels did not induce similar effects. Our results implicate T-type Ca2+ channels as distinct entities for survival signaling in GBM cells and suggest that they are a novel molecular target for tumor therapy.

Functional Expression of SCL/TAL1 interrupting locus (Stil) Protects Retinal Dopaminergic Cells from Neurotoxin-induced Degeneration

We previously isolated a dominant mutation, night blindness b (nbb), which causes a late onset of retinal dopaminergic cell degeneration in zebrafish. In this study, we cloned the zebrafish nbb locus. Sequencing results revealed that nbb is a homolog of the vertebrate SCL/TAL1 interrupting locus (Stil). The Stil gene has been shown to play important roles in the regulation of vertebrate embryonic neural development and human cancer cell proliferation. In this study, we demonstrate that functional expression of Stil is also required for neural survival. In zebrafish, decreased expression of Stil resulted in increased toxic susceptibility of retinal dopaminergic cells to 6-hydroxydopamine. Increases in Stil-mediated Shh signaling transduction (i.e. by knocking down the Shh repressor Sufu) prevented dopaminergic cell death induced by neurotoxic insult. The data suggest that the oncogene Stil also plays important roles in neural protection.

The role of cannabinoidergic system in prenatal neurodevelopment.

The role of cannabinoidergic system in prenatal neurodevelopment.

Effect of tumor suppressor PTEN gene on apoptosis and cell cycle of human airway smooth muscle cells

It is well established that hyperplasia and decreased apoptosis of airway smooth muscle cells (ASMCs) play an important role in the asthmatic airway remodeling. Tumor suppressor PTEN gene with phosphatase activity plays an important regulatory role in embryonic development, cell proliferation, and apoptosis, cell cycle regulation, migration (invasion) of the cytoskeleton. We hypotheses that PTEN gene could affect the growth and viability of ASMCs through the regulation of PI3K/Akt, MAPK, and cell cycle-related gene expression. We constructed a recombinant adenovirus to transfect ASMCs. Cells were divided into the overexpression of PTEN gene group (Ad-PTEN-GFP), negative control group (Ad-GFP), and blank control group (DMEM). The cell apoptosis of ASMCs were evaluated by Hoechst-33342 staining and PE-7AAD double-labeled flow cytometry. The cell cycle distribution was observed by flow cytometry with PI staining. The expression of PTEN, p-Akt, total-Akt, p-ERK1/2, total-ERK1/2, cleaved-Caspases-3, Caspases-9, p21, and Cyclin D1 were tested by the Western blotting. Our study revealed that overexpression of PTEN gene did not induce apoptosis of human ASMCs cultured in vitro. However, overexpression of PTEN inhibited proliferation of human ASMCs cultured in vitro and was associated with downregulation of Akt phosphorylation levels, while did not affect ERK1/2 phosphorylation levels. Moreover, overexpression of PTEN could induce ASMCs arrested in the G0/G1 phase through the downregulation of Cyclin D1 and upregulation of p21 expressions.

Fibroblast Growth Factor Receptor Inhibitors

Fibroblast growth factor receptors (FGFRs) play an important role in embryonic development, angiogenesis, wound healing, cell proliferation and differentiation. The fibroblast growth factor receptor (FGFR) isoforms have been under intense scrutiny for effective anticancer drug candidates. The fibroblast growth factor (FGF) and its receptor (FGFR) provide another pathway that seems critical to monitoring angiogenesis. Recent findings suggest that FGFR mediates signaling, regulates the PKM2 activity, and plays a crucial role in cancer metabolism. The current review also covers the recent findings on the role of FGFR1 in cancer metabolism. This paper reviews the progress, mechanism, and binding modes of recently known kinase inhibitors such as PD173074, SU series and other inhibitors still under clinical development. Some of the structural classes that will be highlighted in this review include Pyrido[2,3-d]pyrimidines, Indolin- 2-one, Pyrrolo[2,1-f][1,2,4]triazine, Pyrido[2,3-d]pyrimidin-7(8H)-one, and 1,6- Naphthyridin-2(1H)-ones.

Decreased contact inhibition in mouse adipose mesenchymal stem cells.

The proliferation of embryonic cells or adult stem cells in tissue is critically regulated during development and repair. How limited the proliferation of cells, so far, is not much explored. Cell-cell contact proliferation inhibition is known as a crucial mechanism regulating cell proliferation in vitro and in vivo. In this study we examined the characters of mouse subcutaneous adipose derived stem cells (msADSC) whether they lost or get contact inhibition during in vitro culture. The characters of msADSC growth after confluence were analyzed using confocal microscope and the expression profiles of contact inhibition related genes were analyzed according to the morphological changes using real-time PCR method. msADSC showed overlapping growth between them but not after passage 14. The cell shapes were also changed after passage 14. The expression profiles of genes which are involved in contact inhibition were modified in the msADSC after passage 14. The differentiation ability of msADSCs to adipocyte, chondrocyte and osteocyte was not changed by such changes of gene expression profiles. Based on these results, it is revealed that smADSC were characterized by getting of strong cell-cell contact inhibition after passage 14 but the proliferation and developmental ability were not blocked by the change of cell-cell contact proliferation inhibition. These finding will help to understand the growth of adipose tissue, although further studies are needed to evaluate the physiological meaning of the cell-cell contact proliferation inhibition during in vitro culture of msADSC.

Fibroblast Growth Factor Receptor Inhibitors

Fibroblast growth factor receptors (FGFRs) play an important role in embryonic development, angiogenesis, wound healing, cell proliferation and differentiation. The fibroblast growth factor receptor (FGFR) isoforms have been under intense scrutiny for effective anticancer drug candidates. The fibroblast growth factor (FGF) and its receptor (FGFR) provide another pathway that seems critical to monitoring angiogenesis. Recent findings suggest that FGFR mediates signaling, regulates the PKM2 activity, and plays a crucial role in cancer metabolism. The current review also covers the recent findings on the role of FGFR1 in cancer metabolism. This paper reviews the progress, mechanism, and binding modes of recently known kinase inhibitors such as PD173074, SU series and other inhibitors still under clinical development. Some of the structural classes that will be highlighted in this review include Pyrido[2,3-d]pyrimidines, Indolin- 2-one, Pyrrolo[2,1-f][1,2,4]triazine, Pyrido[2,3-d]pyrimidin-7(8H)-one, and 1,6- Naphthyridin-2(1H)-ones. Keywords: FGFR receptor, Cancer metabolism, FGFR1 inhibitors, FGFR mediated signaling, cancer

Fibroblast Growth Factor Receptor-Mediated Activation of AKT-β-Catenin-CBP Pathway Regulates Survival and Proliferation of Murine Hepatoblasts and Hepatic Tumor Initiating Stem Cells

Fibroblast Growth Factor (FGF)-10 promotes the proliferation and survival of murine hepatoblasts during early stages of hepatogenesis through a Wnt-β-catenin dependent pathway. To determine the mechanism by which this occurs, we expanded primary culture of hepatoblasts enriched for progenitor markers CD133 and CD49f from embryonic day (E) 12.5 fetal liver and an established tumor initiating stem cell line from Mat1a−/− livers in media conditioned with recombinant (r) FGF10 or rFGF7. FGF Receptor (R) activation resulted in the downstream activation of MAPK, PI3K-AKT, and β-catenin pathways, as well as cellular proliferation. Additionally, increased levels of nuclear β-catenin phosphorylated at Serine-552 in cultured primary hepatoblasts, Mat1a−/− cells, and also in ex vivo embryonic liver explants indicate AKT-dependent activation of β-catenin downstream of FGFR activation; conversely, the addition of AKT inhibitor Ly294002 completely abrogated β-catenin activation. FGFR activation-induced cell proliferation and survival were also inhibited by the compound ICG-001, a small molecule inhibitor of β-catenin-CREB Binding Protein (CBP) in hepatoblasts, further indicating a CBP-dependent regulatory mechanism of β-catenin activity. Conclusion: FGF signaling regulates the proliferation and survival of embryonic and transformed progenitor cells in part through AKT-mediated activation of β-catenin and downstream interaction with the transcriptional co-activator CBP.

The integration of microRNA target data by biclustering techniques opens new roads for signaling networks analysis

Motivation and Objectives MicroRNAs (miRNAs) are key modulators of gene expression. In addition to their recognised role in embryonic and adult cell proliferation and differentiation (Ren et al., 2009), many recent studies on diverse types of human cancer have demonstrated that miRNAs are functionally integrated into those oncogenic pathways that are central to tumorogenesis (Olive et al., 2010). Although microarray profiling and next generation sequencing technologies have allowed researchers to discover much of their structural and functional features as well as many new miRNAs, the current challenge is to understand their specific biological functions and mechanisms through which they are able to ensure cell homeostasis and to control developmental timing and cancer progression. This is not a trivial task because the post-transcriptional regulation of gene expression mediated by miRNAs is rarely resolved by a simple one-to-one interaction between a miRNA and a target gene. It is much more complex, often involving multiple binding of the same miRNA and/or of different miRNAs in a cooperative manner. The combinatorial effects of different miRNAs on the same gene, or on different genes of the same pathway, is an essential part of the mechanism through which they are able to fine-tune signaling pathways (Inui et al., 2010). Indeed, the effect of a miRNA may change depending on which other miRNAs are co-expressed or silenced, which in turn depends on the specific context in which the cell, the tissue or the organism is considered. This makes the interpretation of miRNAs expression profile really difficult and a mere analysis of the list of differentially expressed genes cannot provide enough information to elucidate the multiplicity of potential miRNA:mRNA interactions. In this context, the exploitation of data mining techniques, and in particular of biclustering algorithms, is considered as a useful approach to search the correlations among miRNAs and mRNAs. However, as each miRNA may target hundreds of genes, the selection of the most significant results for further experimental validations still remains a challenging task for many biologists. The proposed method, which is implemented in the system HOCCLUS2, has been designed to analyse data of miRNA:mRNA interactions (derived from expression arrays or from large sets of predictions) in order to detect significant co-regulatory partnerships. In particular, the aim is to provide the biologists with a tool which can support them in two challenging tasks, that is, the detection of actual miRNAs target genes and the identification of the context-specific co-associations of different miRNAs. A further contribution to the considered research consists in the ranking of the extracted biclusters on the basis of the semantic similarity between the target genes, which allows the biologists to easily select the most significant results, from a biological view point. Availability: http://www.di.uniba.it/~ceci/micFiles/ systems/HOCCLUS2/index.html

Abstract B63: Developing Novel STAT3 Inhibitors for the Treatment and Prevention of Cancer

Abstract Signal transducers and activators of transcription (STATs) are a family of transcription factors involved in the regulation of early embryonic development, immune response, cell proliferation, differentiation, and apoptosis. Particularly, STAT3 regulates lobuloalveolar apoptosis during involution in mammary gland development, and its activation is required for the growth of CD44+CD24- stem cell-like breast cancer cells and the development of multiple cancers including breast cancer. Increased STAT3 transcriptional activity was found to correlate with ER-negative phenotype in breast cancer cell lines and in primary human invasive ductal breast carcinomas. Thus, STAT3 represents a promising target for therapeutic and preventive intervention of breast cancer. Yet current strategies of inhibiting STAT3 activity did not sufficiently block STAT3 activation in cancer cells due to their limited potency, poor solubility or bioavailability. Despite several STAT3 inhibitors have been advanced into early clinical trials, there is an urgent need to develop a more successful strategy targeting STAT3 for cancer therapies, and preventive use in particular. We attempted to identify potent and orally active STAT3 inhibitors for such purposes and have designed, synthesized and screened a series of novel STAT3 inhibitors utilizing fragment-based drug design strategy. A number of small molecules with new scaffolds have been discovered with low-micromolar to nanomolar potency in inhibiting the proliferation of various human breast cancer and pancreatic cancer cells. The identified compound HJC0152 significantly suppresses basal and IL-6-stimulated STAT3 promoter activity and STAT3 phosphorylation, reduces total STAT3 protein expression, induces G2/M cell cycle arrest, and induces early apoptosis. HJC0152 has demonstrated a better potency in suppressing the growth of MDA-MB-231 breast xenograft tumors via intraperitoneal and oral administrative routes, with remarkably improved aqueous solubility (3,300 fold improvement) and less toxicity than the lead compounds. In conclusion, we have developed a new class of orally active STAT3 inhibitors with superior drug-like properties for further preclinical therapeutic and preventive evaluation. Citation Format: Zhengduo Yang, Haijun Chen, Lili Chu, Yusong Zhang, Chunyong Ding, Kristin Terry, Jia Zhou, Qiang Shen. Developing novel STAT3 inhibitors for the treatment and prevention of cancer. [abstract]. In: Proceedings of the Eleventh Annual AACR International Conference on Frontiers in Cancer Prevention Research; 2012 Oct 16-19; Anaheim, CA. Philadelphia (PA): AACR; Cancer Prev Res 2012;5(11 Suppl):Abstract nr B63.

VEGF-mediated phosphorylation of eNOS regulates angioblast and embryonic endothelial cell proliferation

To evaluate potential roles of nitric oxide (NO) in the regulation of the endothelial lineage and neovascular processes (vasculogenesis and angiogenesis) we evaluated endothelial nitric oxide synthase (eNOS) and phosphorylated eNOS (p-eNOS) expression in 7.2–8.5 days post-coitum (dpc) mouse embryos. Analysis revealed that p-eNOS(S1177) but not P-eNOS(S617) or P-eNOS(T495) was expressed in a subpopulation of angioblasts (TAL-1+/Flk-1+/CD31−/CD34−/VE-Cadherin−) at 7.2 dpc. A role of the VEGF/Akt1/eNOS signaling pathway in the regulation of the endothelial cell (EC) lineage was suggested by the strong correlation observed between cell division and p-eNOS(S1177) expression in both angioblasts and embryonic endothelial cells (EECs, TAL-1+/Flk-1+/CD31+/CD34+/VE-Cadherin+). Our studies using Akt1 null mouse embryos show a reduction in p-eNOS(S1177) expression in angioblast and EECs that is correlated with a decrease in endothelial cell proliferation and results in changes in VEGF-induced vascular patterning. Further, we show that VEGF-mediated cell proliferation in Flk-1+ cells in allantoic cultures is decreased by pharmacological inhibitors of the VEGF/Akt1/eNOS signaling pathways. Taken together, our findings suggest that VEGF-mediated eNOS phosphorylation on Ser1177 regulates angioblast and EEC division, which underlies the formation of blood vessels and vascular networks.

9-Cis-Retinoic Acid Induces Growth Inhibition in Retinoid-Sensitive Breast Cancer and Sea Urchin Embryonic Cells via Retinoid X Receptor α and Replication Factor C3

There is widespread interest in defining factors and mechanisms that suppress the proliferation of cancer cells. Retinoic acid (RA) is a potent suppressor of mammary cancer and developmental embryonic cell proliferation. However, the molecular mechanisms by which 9-cis-RA signaling induces growth inhibition in RA-sensitive breast cancer and embryonic cells are not apparent. Here, we provide evidence that the inhibitory effect of 9-cis-RA on cell proliferation depends on 9-cis-RA-dependent interaction of retinoid X receptor α (RXRα) with replication factor C3 (RFC3), which is a subunit of the RFC heteropentamer that opens and closes the circular proliferating cell nuclear antigen (PCNA) clamp on DNA. An RFC3 ortholog in a sea urchin cDNA library was isolated by using the ligand-binding domain of RXRα as bait in a yeast two-hybrid screening. The interaction of RFC3 with RXRα depends on 9-cis-RA and bexarotene, but not on all-trans-RA or an RA receptor (RAR)-selective ligand. Truncation and mutagenesis experiments demonstrated that the C-terminal LXXLL motifs in both human and sea urchin RFC3 are critical for the interaction with RXRα. The transient interaction between 9-cis-RA-activated RXRα and RFC3 resulted in reconfiguration of the PCNA-RFC complex. Furthermore, we found that knockdown of RXRα or overexpression of RFC3 impairs the ability of 9-cis-RA to inhibit proliferation of MCF-7 breast cancer cells and sea urchin embryogenesis. Our results indicate that 9-cis-RA-activated RXRα suppresses the growth of RA-sensitive breast cancer and embryonic cells through RFC3.

Involvement of the cAMP response element binding protein, CREB, and cyclin D1 in LPA-induced proliferation of P19 embryonic carcinoma cells.

Lysophosphatidic acid (LPA) is a lipid growth factor that induces proliferation of fibroblasts by activating the cAMP response element binding protein (CREB). Here, we further investigated whether LPA induces proliferation of P19 cells, a line of pluripotent embryonic carcinoma cells. 5'-Bromo-2-deoxyuridine incorporation and cell viability assays showed that LPA stimulated proliferation of P19 cells. Immunoblot experiments with P19 cells revealed that the mitogen activated protein kinases, including p-ERK, p38, pAKT, glycogen synthase kinase 3β, and CREB were phosphorylated by treatment with 10 μM LPA. LPA-induced phosphorylation of CREB was efficiently blocked by U0126 and H89, inhibitors of the MAP kinases ERK1/2 and mitogen- and stress-activated protein kinase 1, respectively. Involvement of cyclin D1 in LPA-induced P19 cell proliferation was verified by immunoblot analysis in combination with pharmacological inhibitor treatment. Furthermore, LPA up-regulated CRE-harboring cyclin D1 promoter activity, suggesting that CREB and cyclin D1 play significant roles in LPA-induced proliferation of P19 embryonic carcinoma cells.

Oocyte Family Trees: Old Branches or New Stems?

Oocyte Family Trees: Old Branches or New Stems?

Highlights of This Issue

The apparent parallels between embryonic stem cell (ESC) self-renewal and cancer cell proliferation have raised speculation that a common network of genes may regulate these traits. Jung and colleagues identified a set of microRNAs as promoters of ESC self-renewal and liver cancer cell proliferation. These microRNAs are frequently upregulated in solid liver tumor tissues, and their overexpression in quiescent human primary hepatocytes induces EMT and activates multiple cancer-associated signaling pathways. These findings suggest that microRNAs that are essential in normal development as promoters of ESC self-renewal may harbor neoplastic transformation when they escape silencing in quiescent cellular lineages.Mast cells are recruited to many solid tumors via the stem cell factor/KIT receptor signaling axis. Within tumors, mast cell–derived mediators enhance growth of tumor vasculature and metastasis. Kwok and colleagues used a combination of in vitro and in vivo assays to show that FES kinase signaling in mast cells is required for migration toward mammary tumors and promotes tumor progression in mice. These results identify FES as a potential therapeutic target in cancers with stromal mast cell involvement.The Hippo pathway has emerged as an important component of the intricate cellular signaling network regulating cell proliferation and apoptosis. Despite the distinct evidence of loss of the NF2 gene in the majority of sporadic meningiomas, little is known about the contribution of the Hippo pathway to the development of meningeal neoplasia. Baia and colleagues examined 70 primary tumors by immunohistochemistry and found that YAP1 was highly expressed and localized to the nucleus, highlighting its activation in meningiomas. Using human cells lines and an in vivo mouse model, they investigated the role of YAP1 in meningiomas and its effects on cell proliferation, migration, apoptosis, and tumorigenesis. These findings strongly suggest that deregulation of the Hippo pathway is largely observed and that YAP1 functions as an oncogene in these tumors.Radioresistance is a major feature of erbB-overexpressing tumors with activated phosphoinositide 3-kinase/Akt signaling, stimulating DNA double-strand break repair. Toulany and colleagues describe the specific steps of how Akt interacts with DNA–protein kinase complex (DNA-PKcs), the major enzyme of the nonhomologous end-joining repair. Radiation-induced Akt activates DNA-PKcs through direct protein–protein interaction and promotes its accumulation at the DNA damage site. Subsequently, the Akt-dependent release of DNA-PKcs facilitates effective repair of radiation-induced DNA double-strand breaks and thus stimulates tumor cell survival. This mechanistic study provides the rationale to target Akt in combination with radiotherapy for resistant tumors.

Spinal epidural vascular tumors and malformations

Dear Editor, We read the article “Pure spinal epidural cavernous hemangioma” by Zhong et al. [4] published recently in Acta Neurochirurgica with great interest and found that some discussion is pertinent. In 1982, Mulliken and Gloweki [3] classified vascular anomalies into two categories of hemangiomas (infantile hemangioma, congenital hemangioma, kaposiform hemangioendothelioma, and tufted angioma) and vascular malformation (arteriovenous, venous, lymphatic, and capillary malformations). In 1996, the International Society for the Study of Vascular Anomalies approved this classification system to establish a common language for the many different medical specialists who are involved in the management of these lesions. Previously, there was no precise classification and naming of hemangioma and vascular malformation. They were mostly referred to as hemangioma or lymphangioma together, and were primarily named according to the lesion’s morphology. For example, hemangioma included capillary hemangioma, cavernous hemangioma, racemose hemangioma; lymphangioma included capillary, cavernous and cystic. Since patients with hemangioma or vascular malformations can be treated in different clinical departments, even the same lesion can also get different names, as wine-color stains are referred as PWS in some departments. In 1982, Mulliken and Gloweki put forward that there should be a re-classification according to the cell biological and pathological characteristics, and made it clear that the two types of lesions as tumor and deformity should be distinguished. They were divided into two categories of hemangiomas and vascular malformations according to whether the proliferations of endothelial cells were found in the pathologic tissues. Hemangioma was the embryonic vascular endothelial cell proliferation characterized by benign tumor. This is quite different from the original traditional classification that was based on morphology and pathology. The occurance of hemangioma in neonates is about 3 %, which often occurs in the neonatal period and then enters the proliferative phase. Generally, it should get timely treatment once it is found. Ninety percent of vascular malformations are there at birth, but with no proliferation phase, they will grow proportionally with the increase in age [1]. Previous traditional classification rules defined wine-color stains, cavernous hemangioma, and racemose hemangioma as all belonging to vascular malformation, but they are now named as capillary malformations, venous malformation and arteriovenous malformations respectively. Due to errors in classification, many vascular malformations are mistaken as hemangioma, and many patients received false and ineffective treatment. The term “cavernous hemangioma” refers to a venous malformation. It consists of dilated sinusoidal channels lined by quiescent endothelium. The magnetic resonance imaging features of vascular malformations include: T2-weighted images that performed as high-intensity signal, T1-weighted images performed as the iso-intensity signal, just as that recorded in the article; magnetic resonance imaging can reveal the spinal epidural venous malformation, intra-lesion hemorrhage and thrombosis. Magnetic resonance imaging can demonstrate the breadth of lesions and the relationship between lesions and adjacent tissues; it can also help to provide the potential treatment plans. M. Zhu :Y. Luo : L. Qiu (*) Department of Diagnostic Ultrasound, West China Hospital of Sichuan University, No.37 Guo Xue Xiang, Chengdu 610041, Sichuan, China e-mail: vigour_sky@163.com

The ubiquitin ligase mLin41 temporally promotes neural progenitor cell maintenance through FGF signaling

How self-renewal versus differentiation of neural progenitor cells is temporally controlled during early development remains ill-defined. We show that mouse Lin41 (mLin41) is highly expressed in neural progenitor cells and its expression declines during neural differentiation. Loss of mLin41 function in mice causes reduced proliferation and premature differentiation of embryonic neural progenitor cells. mLin41 was recently implicated as the E3 ubiquitin ligase that mediates degradation of Argonaute 2 (AGO2), a key effector of the microRNA pathway. However, our mechanistic studies of neural progenitor cells indicate mLin41 is not required for AGO2 ubiquitination or stability. Instead, mLin41-deficient neural progenitors exhibit hyposensitivity for fibroblast growth factor (FGF) signaling. We show that mLin41 promotes FGF signaling by directly binding to and enhancing the stability of Shc SH2-binding protein 1 (SHCBP1) and that SHCBP1 is an important component of FGF signaling in neural progenitor cells. Thus, mLin41 acts as a temporal regulator to promote neural progenitor cell maintenance, not via the regulation of AGO2 stability, but through FGF signaling.

MiR‐210: an important player in the pathogenesis of preeclampsia?

MiR‐210: an important player in the pathogenesis of preeclampsia?

Jmjd5, an H3K36me2 histone demethylase, modulates embryonic cell proliferation through the regulation of Cdkn1a expression

Covalent modifications of histones play an important role in chromatin architecture and dynamics. In particular, histone lysine methylation is important for transcriptional control during diverse biological processes. The nuclear protein Jmjd5 (also called Kdm8) is a histone lysine demethylase that contains a JmjC domain in the C-terminal region. In this study, we have generated Jmjd5-deficient mice (Jmjd5(Δ)(/)(Δ)) to investigate the in vivo function of Jmjd5. Jmjd5(Δ)(/)(Δ) embryos showed severe growth retardation, resulting in embryonic lethality at the mid-gestation stage. Mouse embryonic fibroblasts (MEFs) derived from Jmjd5 hypomorphic embryos (Jmjd5(neo/neo)) also showed the growth defect. Quantitative PCR analysis of various cell cycle regulators indicated that only Cdkn1a expression was upregulated in Jmjd5(neo/neo) MEFs and Jmjd5(Δ)(/)(Δ) embryos. A knockdown assay with Cdkn1a-specific small interfering RNAs revealed that the growth defect of Jmjd5(neo/neo) MEFs was significantly rescued. In addition, a genetic study using Jmjd5(Δ)(/)(Δ); Cdkn1a(Δ)(/)(Δ) double-knockout mice showed that the growth retardation of Jmjd5(Δ)(/)(Δ) embryos was partially rescued by Cdkn1a deficiency. Chromatin immunoprecipitation analysis showed that increased di-methylated lysine 36 of histone H3 (H3K36me2) and reduced recruitment of endogenous Jmjd5 were detected in the transcribed regions of Cdkn1a in Jmjd5(neo/neo) MEFs. Taken together, these results suggest that Jmjd5 physiologically moderates embryonic cell proliferation through the epigenetic control of Cdkn1a expression.

Inhibitor of DNA binding 1 (Id1) induces differentiation and proliferation of mouse embryonic carcinoma P19CL6 cells

The inhibitor of DNA binding (Id) family of genes encodes negative regulators of basic helix–loop–helix transcription factors and has been implicated in such diverse cellular processes as differentiation, proliferation, apoptosis and migration. Id knockout mouse embryos display multiple cardiac defects but the specific role of Id1 in cardiac differentiation is unclear. In the present study, we investigated the function of Id1 in DMSO-induced P19CL6 cells, a widely-accepted cell model of cardiac differentiation.We found that Id1 was upregulated during the cardiac differentiation of P19CL6 cells. The expression of cardiac specific marker genes, Gata4, α-MHC and ISL1, was upregulated in P19CL6 cells stably transfected with Id1 (P19CL6-Id1) during cardiac differentiation. The overexpression of Id1 reduced the number of cells in G1 phase and increased the cell population in G2, M and S phases, while knockdown of Id1 increased the number of cells in G1 phase from 48.6±2.51% to 62.2±1.52% at day 0 of cardiac induction, and from 52.5±3.41% to 63.7±1.02% at day 3 after cardiac induction, indicating that Id1 promoted proliferation of P19CL6 cells. Luciferase assays showed that the activity of TOP flash was higher in P19CL6-Id1 cells than wildtype P19CL6 cells, while Id1 expression was also upregulated in P19CL6 cells treated with Wnt3a or LiCl. This indicates that there may be positive feedback between Id1 and Wnt signaling which plays an important role in cardiac differentiation.