bluuebunny/crossref_metadata_2025 huggingface dataset:
High glucose‐mediated alterations of mechanisms important in myogenesis of mouse C2C12 myoblasts
Authors: Grzelkowska‐Kowalczyk K. , Wieteska‐Skrzeczyńska W. , Grabiec K. , Tokarska J.
Published: August 2013

We have examined the progression and regulation of myogenesis, cellular levels of IGFBP‐4, ‐5, ‐6, and several extracellular matrix (ECM) proteins (fibronectin, integrin α5, β1 subunits and a disintegrin metalloprotease ADAM12) in murine C2C12 myoblasts during 3‐day differentiation under high glucose alone or combined with high insulin, factors characteristic for type 1 and 2 diabetes. High ambient glucose inhibited myogenesis of C2C12 myoblasts, an effect manifested by a twofold decrease in myoblast fusion, a drop in intracellular MyoD, myogenin and MHC levels, and increased cellular content of active myostatin isoform. Reduction in myogenesis by high glucose is accompanied by increase in cellular levels of IGFBP‐4 and ‐6 and decrease in IGFBP‐5. High glucose could modify ECM components assembly, by the increase in fibronectin levels and the decrease in metalloprotease ADAM12, without the effect on integrin α5 and β1 subunits. In contrast, high glucose and high insulin activate myoblast differentiation, manifested by an increase in fusion index and myogenin, as well as a drop in myostatin levels. The presence of high insulin prevented high‐glucose‐dependent changes in IGFBPs and ECM proteins. The data indicate the potential mechanisms of the influence of extracellular environment associated with maternal diabetes and insulin resistance on foetal myogenesis.
Directed differentiation of human embryonic stem cell‐line HUES9 to dopaminergic neurons in a serum‐free defined culture niche
Authors: Datta Indrani , Ganapathy Kavina , Tattikota Surendra Mohan , Bhonde Ramesh
Published: August 2013

Although there are several reports on differentiation of human embryonic stem cells to dopaminergic neurons, notable heterogeneity exists in the reported yields of tyrosine hydroxylase (TH)‐positive cells. For benchmarking performance and efficiency standards in future applications of hESC‐derived dopaminergic neurons, there is thus a dire need of well‐defined directed differentiation protocols. Pal et al. [Pal et al. 2009 Exp Biol Med (Maywood) 234:1230–3] demonstrated predisposition of HUES9 towards ectodermal lineage, but the directed differentiation of HUES9 to dopaminergic neurons has not yet been reported. Therefore, we report here a simple two‐step protocol using suitable ECM and serum‐free induction medium for generating dopaminergic cells from HUES9‐derived embryoid bodies. Flow cytometry analysis of the neural progenitors obtained after the first step gave an enriched yield of cells immune‐positive for nestin (99.6 ± 0.1%), musashi12 (98.1 ± 1.5%) and Sox2 (95.4 ± 2.6%). Most of these cells also expressed the proliferation marker Ki67 (83.8 ± 1.5%), whereas the presence of the undifferentiated stem cell marker Oct4 was negligible. In the second step, when these neural progenitors were exposed to midbrain cues sonic hedgehog and fibroblast growth factor 8 along with bFGF, the differentiated cells showed an upregulation of dopaminergic‐related transcription factors Nurr1 and Engrailed1. Immunocytochemistry and flow cytometry analysis showed that these differentiated cells were positive for the mature neuronal marker Map2ab (96.2 ± 1.5%) and dopaminergic neuronal marker TH (71.9 ± 4.4%). Thus, the data demonstrate novel findings of the directed differentiation of HUES9 to dopaminergic neurons using well‐defined serum‐free nutrient supplements.
Drosophila splicing factor SF2 knock‐down mutant shows altered cell‐cycle in vivo
Authors: Dubatolova Tatiana D. , Volkova Elena I. , Omelyanchuk Leonid V.
Published: August 2013

Pioneering single gene study documents pre‐mRNA processing proteins participation in the cell‐cycle regulation of multi‐cellular animals (Andersen and Tapon, 2008, J Biol Chem 283: 31256–67). Whole‐genome RNAi screen in Drosophila tissue‐culture cell lines demonstrates that 17 genes involved in RNA‐processing are required for G2/M check‐point function (Kondo and Perrimon, 2011, Sci Signal 4: rs1). In particular, the silencing of Splicing Factor 2 (SF2) increases the number of G2(M) cells. We have measured the absolute duration of cell‐cycle phases in SF2 depleted flies with the use of flow cytometry and growth parameters of GFP marked mosaic clones. For SF2 mutant cells, G1 = 1.89 h, G2(M) = 7.22 h and S = 1.30 h compared with G1 = 2.25 h, G2(M) = 4.86 h and S = 1.28 h for control normal cells. Thus, G2(M) phase appears to be longer in SF2 silenced cells, supporting the evidence that this splicing protein participates in G2‐M check‐point function.
Effect of treadmill running on the expression of genes that are involved in neuronal differentiation in the hippocampus of adult male rats
Authors: Mojtahedi Shima , Kordi Mohammad Reza , Hosseini Seyed Ebrahim , Omran Simin Fallah , Soleimani Masoud
Published: August 2013

Gene expression in the rodent brain is dependent on exercise. However, our findings regarding this fact are inadequate. The aim of this study was to evaluate the effect of exercise intensity on changes in factors involved in neuronal differentiation and plasticity in the hippocampus of adult male rats. Thus, the effects of 2 weeks treadmill running at of low (11 m/min) and high (25 m/min) intensities, 30 min/day for 14 consecutive days on the expression of two genes, microRNA‐124 and neuron‐restrictive silencer factor/RE1‐silencing transcription factor (REST) as well as changes of two proteins of brain‐derived neurotrophic factor (BDNF) and tyrosine kinase B (TrkB) were analysed. Using quantitative real‐time PCR techniques have shown that both exercise intensities reduced the expression of REST (31% in low exercise intensity, 52% in high exercise intensity) and elevated the expression of miR‐124 (3.8‐fold in low exercise intensity, 11.9‐fold in high exercise intensity) significantly (P ≤ 0.05). This suggests that exercise probably through the changes in these genes is involved in neuronal differentiation in the hippocampus of rats. BDNF and TrkB levels were measured by ELISA. Exercise, at low intensity, increased TrkB (4.51 ng/L vs. 3.73 ng/L) and BDNF (11.97 ng/L vs. 11.08 ng/L) proteins significantly (P ≤ 0.05), while at high intensity, the changes in comparison with the control group were not significant. Expression levels during the exercise programme with high intensity were dramatic for miR‐124 and less dramatic for REST compared with low intensity exercise programme.
Expression and biological function of programmed death ligands in human placenta mesenchymal stem cells
Authors: Wang Guoyan , Zhang Siying , Wang Feifei , Li Guangyun , Zhang Lixia , Luan Xiying
Published: August 2013

Mesenchymal stem cells (MSCs) play important roles in tissue regeneration due to their self‐renewal, multilineage differentiation and immunosuppression abilities. MSCs can be isolated from various kinds of tissue, such as umbilical cord, cord blood and placenta. Human placenta mesenchymal stem cells (hPMSCs) possess stronger immunosuppressive properties, such as the ability to inhibit T‐cell activation and proliferation, than human bone marrow MSCs. We have investigated that the roles of the programmed death ligands 1 and 2 (PDL1 and PDL2) in hPMSC adhesion, migration and immunosuppression were investigated. PDL1 and PDL2 were highly expressed by hPMSCs. Knockdown of PDL1 and/or PDL2 by siRNA increased hPMSC adhesion, but greatly decreasing migration. PDL1 and PDL2 expressed on hPMSCs inhibited T‐cell proliferation by arresting the cell cycle. Knockdown of PDL1 and/or PDL2 in hPMSCs, however, had no effect on the expression of CD69, a T‐cell early activation marker found on both CD4+ and CD8+ T‐cell subsets. In summary, the roles of the negative co‐stimulators PDL1 and PDL2 is on the adhesion, migration and immunosuppression of hPMSCs. These findings may be useful regarding the potential use of hPMSCs in clinical cell.
Adventitial fibroblasts from apoE(−/−) mice exhibit the characteristics of transdifferentiation into myofibroblasts
Authors: Xu Fang , Liu Ying , Hu Weicheng
Published: August 2013

Adventitial fibroblasts (AFs) are the main cell type in the adventitia, however, their role in atherosclerosis remains unclear. We have investigate the role of AFs in atherosclerotic lesion formation by comparing the characteristics of AFs from apoE(−/−) to C57BL/6 mice. A minority of AFs from apoE(−/−) mice expressed α‐SM‐actin, but no α‐SM‐actin‐positive cells were found in AFs from C57BL/6 mice. The content of total collagens, and the mRNA levels of collagen I and collagen III in AFs of apoE(−/−) mice, were higher than in C57BL/6 mice. AFs from apoE(−/−) mice proliferate and migrate faster, and synthesized more TGF‐β1, MCP‐1, and PDGF‐b AFs from apoE(−/−) mice have the characteristics of transdifferentiation into myofibroblasts, including enhanced proliferation and migration, along with synthesis of collagens and cytokines compared to AFs from C57BL/6 mice. The histological and functional characteristics of AFs may contribute to early atherosclerotic lesion formation.
Asymmetric cytokinesis guide the development of pseudomonads inRhynchospora pubera(Cyperaceae)
Authors: San Martin Juca Abramo Barrera , de Jesus Andrade Celia Guadalupe Tardeli , Mastroberti Alexandra Antunes , de Araújo Mariath Jorge Ernesto , Vanzela André Luís Laforga
Published: August 2013

The late stages of microsporogenesis in the family Cyperaceae are marked by the formation of an asymmetrical tetrad, degeneration of three of the four nuclei resulting from meiosis and the formation of pseudomonads. In order to understand the cytological changes involved in the development of pseudomonads, a combination of 11 different techniques (conventional staining, cytochemistry procedures, immunofluorescence, FISH and transmission electron microscopy: TEM) were used to study the later stages of microsporogenesis inRhynchospora pubera. The results demonstrated the occurrence of two cytoplasmic domains in the pseudomonads, one functional and the other degenerative, which are physically and asymmetrically separated by cell plate with an endomembrane system rich in polysaccharides. Other changes associated with endomembrane behaviour were observed, such as a large number of lipid droplets, vacuoles containing electron‐dense material and concentric layers of endoplasmic reticulum. Concomitant with the isolation of degenerative nuclei, the tapetal cells also showed evidence of degeneration, indicating that both tissues under programmed cell death (PCD), as indicated by immunofluorescence and TEM procedures. The results are significant because they associate cellular polarisation and asymmetry with different cytoplasmic domains, and hence open new possibilities for studying cellular compartmentalisation and PCD.
Dopaminergic cells, derived from a high efficiency differentiation protocol from umbilical cord derived mesenchymal stem cells, alleviate symptoms in a Parkinson's disease rodent model
Authors: Shetty Prathibha , Thakur Anirban M. , Viswanathan Chandra
Published: August 2013

Mesenchymal stem cells (MSCs) could be an alternative to foetal cells in the treatment of several neurodegenerative diseases, especially Parkinson's disease (PD). We have previously demonstrated the functional efficacy of the undifferentiated bone marrow MSCs (BMMSCs) cultured in a xenofree conditions in PD animal models. We now demonstrate isolation of MSCs from the umbilical cord matrix tissue and assess their safety and efficacy to improve Parkinsonian symptoms in an in vivo animal model. The efficacy of MSCs from BM and umbilical cord in the PD animal mode has also been studied, and more importantly the efficacy of using differentiated UCMSC (D‐UCMSCs) to dopaminergic phenotype. Phenotypically, UCMSCs expressed higher levels of SSEA4 compared to BMMSCs. Analysis of differentiated cells showed that D‐UCMSCs expressed significant levels of Tyrosine Hydroxyalse and Nurr1 compared to D‐BMMSCs. The in vivo efficacy of the differentiated and undifferentiated cell types in the Parkinsonian rats showed that D‐UCMSCs improved the symptoms throughout a year of study. Differentiated cell types are potentially better for clinical use than the undifferentiated type, provided they are made available at the site of action in adequate numbers. MSCs are less immunogenic and immunomodulatory, which opens up the further possibility of using these cells in allogeneic settings. This could be a novel cell therapy application, especially when getting autochthonous cells is difficult.
Propofol induces apoptosis of hepatocellular carcinoma cells by upregulation of microRNA‐199a expression
Authors: Zhang Jian , Wu Guo‐qing , Zhang Yan , Feng Zhi‐ying , Zhu Sheng‐mei
Published: August 2013

Propofol is one of the extensively commonly used intravenous anaesthetic agents. The effects of Propofol on hepatocellular carcinoma (HCC) growth inhibition and apoptosis have been examined. The techniques used were the MTT assay, flow cytometry, real‐time PCR to assess miR‐199a expression, as also caspase‐8 and caspase‐9 activity in HepG2 cells treated with Propofol. Finally, we evaluated the effect of miR‐199a on Propofol‐induced anti‐tumour activity using anti‐miR‐199a. Propofol efficiently inhibited the growth of HCC cells, but was less toxic to normal hepatic cells. It induced apoptosis and increased expression of miR‐199a. Activation of caspase‐8 and caspase‐9 suggested that both extrinsic and intrinsic pathways are involved in Propofol‐induced apoptosis. Anti‐miR‐199a reversed the effect of Propofol on apoptosis and activation of caspase‐8 and caspase‐9 in HepG2 cells. Propofol can effectively induce apoptosis of HCC cells and modulation of miR‐199a possibly contributes to the anti‐tumour action of Propofol. Hence, Propofol might be an effective drug for HCC.
Detection and comparison of cut‐off values for total AgNOR area/nuclear area and AgNOR number/nucleus in benign thyroid nodules and normal thyroid tissue
Authors: Eroz Recep , Cucer Nurhan , Unluhizarci Kursad , Ozturk Figen
Published: August 2013

Argyrophilic nucleolar organising region associated proteins (AgNORs) are of interest in a variety of diseases including thyroid disorders. We have investigated the cut‐off values for AgNOR count and with a new approach, Total AgNOR area/nuclear area (TNORa/Na) proportions to discriminate thyrocytes obtained from benign thyroid nodules and normal thyroid tissue. Thirty patients whose fine needle aspiration (FNA) materials were compatible with a benign lesion (named as goiter group) and 30 controls (subjects with normal thyroid tissue) were included. In the control group, biopsy material was obtained from histologically normal thyroid gland operated on because FNA material was compatible with papillary thyroid carcinoma (PTC). These samples were stained for AgNOR and 100 nuclei per individual were examined. Both AgNOR values for individual cells and cut‐off values were detected for each group. Patients with goiter had significantly (P < 0.001) higher AgNOR count (2.1 ± 0.6%) and TNORa/Na (6.1 ± 1.5%) than the control groups (1.4 ± 0.2 and 3.0 ± 0.4, respectively). The cut‐off values were <1.5 for AgNOR count and <3.8 for TNORa/Na in thyrocytes of the controls. This modified method is an easy and reliable method for discriminating various thyroid disorders, including the differentiation of benign thyroid nodules from malignant ones. It also helps to discriminate thyrocytes obtained from benign nodules from normal thyroid tissue, aiding accurate localisation of sampling in FNA material.
Pigment epithelium‐derived factor delays cellular senescence of human mesenchymal stem cells in vitro by reducing oxidative stress
Authors: Cao Yukun , Yang Ting , Gu Chunhu , Yi Dinghua
Published: August 2013

Mesenchymal stem cells (MSCs) are multipotent progenitor cells that represent a promising approach in the field of regenerative medicine; however, this potential diminishes with senescence. Pigment epithelium‐derived factor (PEDF) gives some protection by reducing oxidative stress, which is known to accelerate cellular senescence. Thus we hypothesized that PEDF could delay senescence during MSC expansion by reducing oxidative stress. Proliferation and differentiation potentials, oxidative stress, senescence and p53/p16 expressions have been examined. In MSCs cultured under normoxic conditions treated with PEDF, proliferative lifespan in vitro was significantly increased compared with control group not given PEDF, with ∼10 additional population doublings (PD) occurring before terminal growth arrest. Most of the MSCs cultured under normoxic conditions ceased to proliferate after 20–28 PD, while few senescent cells were found in the hypoxic, PEDF‐hypoxic and PEDF‐normoxic cultures; this was associated with downregulation of p53 and p16 expression and decreased oxidative stress. PEDF also preserved differentiation potentials of MSCs compared with the control group. Thus PEDF suppression of oxidative stress delays cellular senescence and allows greater expansion of MSCs.
Mutation of Hof1 PEST motif phosphorylation sites leads to retention of Hof1 at the bud neck and a decrease in the rate of myosin contraction
Authors: Stockstill Katherine E. , Park Jungeun , Wille Rachel , Bay Grace , Joseph Avery , Shannon Katie B.
Published: August 2013

Regulation of actomyosin ring contraction is important for the coordination of cytokinesis with mitosis. Hof1, a member of the Pombe Cdc15 homology (PCH) family of proteins, is required for efficient cytokinesis in budding yeast. Phosphorylation of Hof1 depends on the mitotic exit network (MEN), and its degradation at the end of mitosis depends on its PEST motif and interaction with the E3 ligase Grr1. To test the hypothesis that targeted destruction of Hof1 temporally couples mitotic exit with contraction of the actomyosin ring, we mutated the Hof1 PEST motif to prevent phosphorylation and subsequent degradation. These mutations increased the amount of Hof1 at the bud neck during cytokinesis, resulted in smaller bud neck diameter, and slowed the rate of myosin contraction. However, Hof1 PEST motif phosphorylation site mutants did not have cytokinesis defects, indicating that regulation of Hof1 levels does not control the onset of actomyosin ring contraction as predicted.
Enhanced maintenance of rat islets of Langerhans on laminin‐coated electrospun nanofibrillar matrix in vitro
Authors: Sojoodi Mozhdeh , Farrokhi Ali , Moradmand Azadeh , Baharvand Hossein
Published: August 2013

Understanding the extracellular matrix (ECM) effect on pancreatic β cells is critical to optimise the derivation of functional β cells for transplantation and understand mechanisms that control islet neogenesis and glucose homeostasis. We assessed the effect of natural ECMs [collagen I, collagen IV, laminin and fibronectin (FN)] on rat islets of Langerhans' morphology, adhesion, viability, functionality and islet specific genes expression after 7 days in vitro culture. However, we could not detect a significant difference on the other parameters in these ECMs and islets interaction. To examine islets interactions, we used a synthetic three‐dimensional surface composed of electrospun polyamide nanofibres. Laminin‐coated nanofibrillar surfaces, but not laminin or nanosurface alone, induced comparable expression of the Ins1 and Ins2 genes in adult β cells. Using a glucose challenge test, a marked response of insulin secretion by islets occurred that were cultured on laminin‐coated nanofibrillar surfaces. We contend that the reestablishment of cellular interactions by the combination of nanomaterials and natural ECMs can be useful in maintaining in vitro islet functions.
Repeated versus single transplantation of mesenchymal stem cells in carbon tetrachloride‐induced liver injury in mice
Authors: Miryounesi Maryam , Piryaei Abbas , Pournasr Behshad , Aghdami Nasser , Baharvand Hossein
Published: August 2013

Despite its numerous limitations, liver transplants are the only definite cure for end‐stage liver disease. Various stem cell populations may contribute to liver regeneration, of which there is accumulating evidence of the contribution of mesenchymal stem cells (MSCs). This study examines the hypothesis that repeated infusions of human bone marrow‐derived MSCs (hBM‐MSCs) can improve liver injury in an experimental model. MSCs were intravenously transplanted into immunosuppressed mice with carbon tetrachloride (CCl4)‐induced liver fibrosis. Transplanting 3 × 106 MSCs in three divided doses improved survival, liver fibrosis and necrosis compared with injection of the same number of MSCs in a single dose. This was accompanied by increased influence on the expression of the fibrogenic/fibrolytic related genes Col1a1, Timp1 and Mmp13 in the repeated transplant group. Repeat administration of MSCs was three times more effective in homing of PKH‐tagged transplanted cells 3 weeks post‐transplant compared with the single transplant group. The benefits of repeated transplants may be of considerable significance in clinical trials on liver failure.
PAMAM dendrimer roles in gene delivery methods and stem cell research
Authors: Daneshvar Nasibeh , Abdullah Rasedee , Shamsabadi Fatemeh T. , How Chee Wun , MH M. Aizat , Mehrbod Parvaneh
Published: August 2013

Nanotechnology has provided new technological opportunities, which could help in challenges confronting stem cell research. Polyamidoamine (PAMAM) dendrimers, a new class of macromolecular polymers with high molecular uniformity, narrow molecular distribution specific size and shape and highly functionalised terminal surface have been extensively explored for biomedical application. PAMAM dendrimers are also nanospherical, hyperbranched and monodispersive molecules exhibiting exclusive properties which make them potential carriers for drug and gene delivery.
Atomic force microscopy of chronic lymphatic leukaemia cells activation induced by Staphylococcus aureus
Authors: Dong Shisong , Wang Qiulan , Sun Shaoyang , Liang Yuanwei , Jiang Jinhuan , Liu Li , Hu Anbin , Cai Jiye
Published: August 2013

Activation of lymphatic cells is associated with changes in morphology, ultrastructure and adhesion force. We have investigated the activation efficiency of Staphylococcus aureus (SAC) on B‐cell chronic lymphatic leukaemia (B‐CLL) cells using atomic force microscopy (AFM), and found changes in the above properties. Cell viability and proliferation were measured using Cell Counting Kit‐8 (CCK‐8) and enzyme‐linked immunosorbent assay (ELISA). AFM clearly showed that the volume and nuclear–cytoplasm ratio of cells increased significantly with activated time. It also showed that pseudopodia and immunological synapses began to appear at 24 h. In the activation process, nano‐structures of the cell surface became aggregated, and adhesion increased. In conclusion, the results indicate a close relationship between membrane reconstruction and multiplication process of B‐CLL cells.
A novel regulation of PSMA and PSA expression by Q640X AR in 22Rv1 and LNCaP prostate cancer cells
Authors: Ben Jemaa Awatef , Sallami Sataa , Céraline Jocelyn , Oueslati Ridha
Published: August 2013

We have investigated the expression of prostate‐specific membrane antigen (PSMA) and prostate‐specific antigen (PSA) transcripts in androgen‐dependent (LNCaP) and androgen‐independent (22Rv1) prostate cancer cell lines. We also enquired whether Q640X CTE‐truncated androgen receptor (AR) has an impact on transcription of mRNA for PSMA and PSA in transfected androgen‐sensitive prostate cancer LNCaP cells. Wild type LNCaP, 22Rv1 prostate cancer cells, prostate stromal cells (PrSC) and LNCaP cells transfected with p‐Q640X AR, p‐WT AR or p‐C3 empty plasmids were studied. The expression of PSMA and PSA were detected by real‐time PCR after transfection for 4 and 7 days. Expression of mRNAs for PSA was sixfold greater than PSMA in wild type LNCaP cells. In contrast, the wild type androgen refractory 22Rv1 cell line reacted almost exactly the opposite way reverse to LNCaP cells, since the transcription of mRNA for PSMA almost twofold greater than PSA. Non‐transfected human PrSC responded similarly to PSMA mRNA and PSA mRNA was not detected in these cells. Q640X AR transfected LNCaP cells downregulated the expression of PSMA and PSA genes after 7 days. Our results demonstrate that Q640X mutated AR may have an important regulatory role in mediating the PSMA and PSA genes expression during the progression of prostate cancer from androgen‐dependence to androgen‐independence. Understanding their functional properties and mechanisms by which ARs involved in regulation of PSMA and PSA expression will allow the identification of new target therapies for the treatment of hormone‐resistant prostate cancer.
Attenuation of Smad2 activity shows resistance to TGF‐β signalling in mammary adenocarcinoma (MCF‐7) cells
Authors: Sengupta Suman , Kundu Subhadip , Bhattacharyya Arindam
Published: August 2013

Transforming growth factor‐β (TGF‐β) is a potent inhibitor of the growth of normal mammary epithelial cells, and has a pleiotropic, context‐dependent, concentration‐dependent action. We found attenuation of TGF‐β signalling in mammary adenoma carcinoma cells. Phosphorylation at the linker site of Smad2 occurred in a cooperative way during the attenuation of TGF‐β signalling, and was associated with upregulation of CDK2 and cyclin D1. CDK2 inhibitor restored the anti‐proliferative effect of TGF‐β by upregulating p21, with inhibition of linker phosphorylation of Smad2. CDK2‐mediated linker phosphorylation of Smad2 may be a plausible mechanism for the attenuation of TGF‐β signalling in breast cancer.
microRNAs regulate adipocyte differentiation
Authors: Chen Lin , Song Jinlin , Cui Junhui , Hou Jia , Zheng Xiaohui , Li Cai , Liu Lei
Published: August 2013

The number of adipocytes is relevant to the extent of differentiation from pluripotent stem cells into pre‐adipocytes, whereas the size of adipocytes relates to the extent of differentiation from pre‐adipocytes into mature fat cells and the accumulation of triglyceride. Investigation of the molecular regulatory mechanism of adipocyte differentiation is not only essential for understanding the physiological processes of adipogenesis, but it is also important for identifying new biomarkers and therapeutic targets for some metabolic diseases, such as obesity and diabetes. microRNAs (miRNAs) appear to play important roles in adipocyte differentiation. During adipogenesis, miRNAs can accelerate or inhibit adipocyte differentiation by acting on transcription factors, regulating signalling pathways related to adipogenesis, or blocking the mitotic clonal expansion stage, thus regulating adipocyte development. The regulatory role of some miRNAs varies in different species or different cells. In this review, the biological characteristics of miRNA and the adipocyte differentiation process are concisely discussed. Recent advances in our understanding of the role of miRNAs in adipocytes development or adipogenesis are discussed.
Role of adipose‐derived stem cells in enhancing angiogenesis early after aspirated fat transplantation: induction or differentiation?
Authors: Yuan Yi , Gao Jianhua , Liu Linqi , Lu Feng
Published: August 2013

Autologous fat tissue has been used as a potential filler for soft‐tissue defects, despite unpredictable clinical outcomes and low graft survival. Co‐transplantation of adipose‐derived stem cells (ASCs) is an alternative therapeutic approach to effectively enhance the survival and quality of transplanted fat tissue by increasing neovascularization. Nevertheless, the mechanisms by which ASCs exerted their angiogenic effects remain obscure. ASCs can secrete several angiogenic growth factors, for example vascular endothelial growth factor, hepatocyte growth factor and basic fibroblast growth factor. Hypoxic conditions may promote the proliferation of ASCs and their secretion. However, the differentiation of ASCs into endothelial cells (ECs), pericytes and smooth muscular cells in vivo has not been confirmed. The role of ASCs early after aspirated fat transplantation may be to induce new vessels from the recipient region to grow around and into the graft by releasing significant amounts of angiogenic growth factors rather than to differentiate into ECs, pericytes or smooth muscular cells forming new vessels, an effect that might be enhanced by hypoxia.
MiR‐34c oligonucleotide enhances chemosensitivity of Ishikawa cell to cisplatin by inducing apoptosis
Authors: Jiang Ling , Meng Wei , Zeng Jun , Hu Haiyan , Lu Lin
Published: August 2013

Expression of miR‐34c is significantly reduced in human endometrial cancer cells and miR‐34c regulates multiple genes associated with cancer cell proliferation, apoptosis and invasiveness. Thus, miR‐34c can theoretically become a target that enhances chemotherapy. Upregulation of miR‐34c to enhance the chemotherapeutic effect of cisplatin (DDP) has not been studied in human endometrial cancer cells before. In an in vitro study, the human endometrial cancer cell line, Ishikawa, was treated with DDP and miR‐34c mimics, alone or in combination. IC50 values were dramatically decreased in cells treated with miR‐34c mimics when combined with DDP, to a greater extent than those treated with DDP alone. Furthermore, miR‐34c mimics significantly enhanced apoptosis in Ishikawa cells by inhibiting IL‐6R expression. Therefore, a combination of miR‐34c mimics and DDP could be an effective therapeutic strategy for controlling Ishikawa cell proliferation.
Induced autologous stem cell transplantation for treatment of rabbit type 1 diabetes
Authors: Hu Mei‐Jun , Ruan Guang‐Ping , Yao Xiang , Ruan Guang‐Hong , Wang Jin‐Xiang , Pang Rong‐Qing , Cai Xue‐Min , Zhu Xiang‐Qing , He Jie , Pan Xing‐Hua
Published: August 2013

We have examined the effects of induced autologous stem cells on blood sugar levels in a rabbit model of type 1 diabetes. Rabbit skin fibroblasts were induced to dedifferentiate into multipotent stem cells, and were transplanted into the treatment group via the pancreatic artery. After the fibroblasts had been induced for 72 h, some of them became multipotent stem cells. Four weeks after cell transplantation, blood glucose levels of the induced stem cell treatment group were significantly lower. The plasma insulin and plasma C‐peptide levels of the treated group were significantly increased (P < 0.05). The shape and number of islets was different. In the control group, induced cell treatment group and non‐induced cell treatment group. In the control group, islet β‐cell nucleoli were obvious, and cell volumes were larger with more abundant cytoplasm. The rough endoplasmic reticulum was well‐developed and a large number of secretory granules could be seen within the cytoplasm. In the induced cell treatment group, islet β cells were scattered, and their nuclei were oval and slightly irregular in shape. The cytoplasm of these cells contained a nearly normal number of secretory granules. In the non‐induced cell treatment group, islet β‐cells were atrophied and cell volumes were reduced. Cytoplasmic endocrine granules were significantly reduced or absent. In conclusion, treatment with induced multipotent stem cells can reduce blood sugar levels, improve islet cell function, and repair damaged pancreas in a rabbit model of type 1 diabetes.
Enhanced activity of lysosomal proteases in activated rat hepatic stellate cells is associated with a concomitant increase in the number of the mannose‐6‐phosphate/insulin‐like growth factor II receptor
Authors: Mousavi Seyed Ali , Fønhus Marita Sporstøl , Kindberg Grete Mørk , Tolleshaug Helge , Berg Trond
Published: August 2013

Activated hepatic stellate cells (HSCs) play a central role during hepatic tissue repair through their influence on extracellular matrix remodelling. We have determined whether the activity levels of cathepsin B and D are affected by in vitro activation of rat HSCs, and whether the enzymes were released from the cells. Furthermore, given the important role of the mannose‐6‐phosphate/insulin‐like growth factor II receptor (M6P/IGF‐IIR) in the intracellular transport of lysosomal enzymes, we have examined whether changes in the activity of these proteases were associated with parallel changes in the level of the M6P/IGF‐IIR. The activity of cathepsin B and D increased ∼4 times between 2 and 8 days of HSC culture. This result was supported by analysing mRNA expression by RT‐PCR. The cells released the enzymes into the culture medium, amounting to ∼10% of the cell‐associated activity over 24 h. The release of enzymes was not affected by reducing medium pH from 7.4 to 6.2, indicating that the enzymes were transported to the medium independently of the M6P/IGF‐II‐R. The released cathepsin B was mostly in the inactive proenzyme form. HSC activation led to a particularly large increase in M6P/IGF‐IIR expression. A large proportion of the receptors was located on the cell surface and was found to be very suitable for measuring endocytosis of 125I‐IGF‐II. The results show that the endocytic activity increased in parallel with the increase in surface receptors and activity of lysosomal enzymes. Degradation of the ligand was reduced by inhibitors of lysosomal proteases and therefore took place in lysosomes.
Elongated cell morphology and uniaxial mechanical stretch contribute to physical attributes of niche environment for MSC tenogenic differentiation
Authors: Wang Wenbo , Deng Dan , Li Jie , Liu Wei
Published: August 2013

Specific lineage differentiation of mesenchymal stem cells (MSCs), such as osteogenic and chondrogenic differentiation, is the major subject of MSC‐based musculoskeletal tissue regeneration. Nevertheless, an ideal induction regime for MSC tenogenic differentiation is less explored. While induced differentiation of MSCs using chemical signalling, such as growth factors, has been the mainstream methodology in the past decades, recent studies show that a physical signal is one of the major attributes that composes in vivo niches of MSCs, and is a potent factor in determining MSC fate in several in vitro models; for example, control of cell shape with surface topography can drive MSCs towards a specific lineage differentiation. Mechanical signals also play important roles in the differentiation of MSCs towards certain lineages. Tendon is a connective tissue which is subjected to dynamic uniaxial mechanical stretch by physical activity. Additionally, tenocytes have a unique elongated cell morphology and are aligned in parallel with collagen fibres. We thus would hypothesize that both enforced elongated cell morphology and uniaxial mechanical stretch signal contribute to the major physical niche attributes of tenocytes' in vivo microenvironment, and mimicking these physical signals may be sufficient to induce tenogenic differentiation of MSCs.
Identification and analysis of glycogen synthase kinase 3 beta1 interactome
Authors: Gao Xuejuan , Wang Jian‐Ying , Gao Ling‐Mei , Yin Xing‐Feng , Liu Langxia
Published: August 2013

Glycogen synthase kinase‐3 beta (GSK3β) was initially identified as a key protein in glucose metabolism. GSK3β might be involved in cell growth, motility and apoptosis. Systematic identification of GSK3β‐associated proteins is crucial for the exhaustive understanding of its functions. Using GST pull‐down experiment and LCMS/MS analysis coupled to bioinformatics tools, we have identified 114 proteins that interacted with GSK3β1 in hepatocellular carcinoma HepG2 cells. Most of the identified proteins are implicated in metabolic process, whereas other proteins are important for cell proliferation or migration, and have been associated with cancer development and metastasis. Several representative proteins, such as hnRNPK, PCNA, Ezrin and STAT1, have been confirmed to interact with GSK‐3β1 by co‐immunoprecipitation in HepG2 cells. Further studies of these interactions may discover the precise roles and the underlying mechanisms of GSK‐3β1 in tumour growth and metastasis.
Hypoxia preconditioned human adipose derived mesenchymal stem cells enhance angiogenic potential via secretion of increased VEGF and bFGF
Authors: Liu Linqi , Gao Jianhua , Yuan Yi , Chang Qiang , Liao Yunjun , Lu Feng
Published: August 2013

Mesenchymal stem cells (MSCs) are adult multipotent cells found in bone marrow, adipose tissue, and other adult tissues. MSCs improve regeneration of injured tissues in vivo, but the mechanisms remain unclear. Typically, MSCs are cultured under ambient or normoxic conditions (21% O2). However, the physiological niches of MSCs have much lower oxygen tension. When used as a therapeutic tool to repair tissue injuries, MSCs cultured in standard conditions must adapt from 21% O2 in culture to <1% O2 in ischemic tissue. We have examined the effects of hypoxia preconditioning (1% O2) in human adipose derived mesenchymal stem cells (AD‐MSCs) to discover the conditions that best enhance their tissue regenerative potential. We demonstrate that AD‐MSCs respond positively to hypoxia compared with normoxia preconditioning, show decreased apoptosis even in severe microenvironmental conditions (such as a low‐serum medium), and an increased expression of the angiogenic factors, vascular endothelial growth factor and basic fibroblast growth factor. Human umbilical vein endothelial cells have higher vitality and lower apoptosis when cultured in medium taken from hypoxia‐preconditioned AD‐MSCs, as well as significantly increased capillary‐like structures with this medium on Matrigel. The data suggest that hypoxia preconditioned AD‐MSCs can improve tissue regeneration.
Stromal vascular fraction (SVF) cells enhance long‐term survival of autologous fat grafting through the facilitation of M2 macrophages
Authors: Dong Ziqing , Fu Rong , Liu Linqi , Lu Feng
Published: August 2013

Optimum perfusion may be the key to the endurance, and hence survival, of autologous adipose tissue transportation. Stromal vascular fraction (SVF) cell therapy can greatly improve the survival of fat grafts by enhancing angiogenesis. However, SVF cells are poorly retained in later stages of SVF‐assisted adipose tissue transplant. Therefore, it hardly defines the angiogenic effect through long‐term transportation. Adipose tissue suffers from acute hypoxia in the early stage of transportation, leading to the recruitment of macrophages. M2 macrophages enhance angiogenesis in adipose transplantation by acting as an angiogenic signal source, promoting tip cell migration and assisting tip cell fusion. Furthermore, the angiogenic and anti‐inflammatory micro‐environment in the graft created by M2 macrophages may stimulate the transformation of infiltrating macrophages to M2 macrophages. These M2 macrophages may enhance the long‐term retention of graft through angiogenesis. Based on these observations, we postulate that the long‐term angiogenic effect of SVF cells may be achieved through the facilitation of the M2 macrophages.
Downregulation of miRNA‐128 sensitises breast cancer cell to chemodrugs by targeting Bax
Authors: Ji Shufeng , Shao Guoli , Lv Xiaobin , Liu Yuan , Fan Yingfang , Wu Aiguo , Hu Haiyan
Published: August 2013

miR‐128 is more highly expressed in drug‐resistant breast cancer samples than in drug‐sensitive samples. We have confirmed that Bax is the target of miR‐128 by negative post‐transcriptional regulation. miR‐128 and Bax were detected in the breast cancer cell line, MDA‐MB‐231, which was then transfected with miR‐128 MIMIC (precursor of miR‐128) or AMO (antisense‐miR‐128 oligonucleotides). After transfection, the chemosensitivity of MDA‐MB‐231 cell was up‐regulated with increasing of Bax and inhibition of miR‐128.
Experimental study on the facilitative effects of miR‐125b on the differentiation of rat bone marrow mesenchymal stem cells into neuron‐like cells
Authors: Wu Rui , Wang Na , Li Min , Zang Wenqiao , Xu Yuming
Published: August 2013

To observe the effects of miR‐125b on the differentiation of rat bone marrow mesenchymal stem cells (BMSCs) into neuron‐like cells, rat BMSCs were isolated and transfected with Syn‐rno‐miR‐125b* miScript miRNA Mimic (Mimic + BME group), with anti‐rno‐miR‐125b* miScript miRNA Inhibitor (Inhibitor + BME group) and BME control was set up without transfection. Blank controls without transfection and induction by BME was also set up. BMSCs of three groups including Mimic + BME group, Inhibitor + BME group and BME group were induced to differentiate into neuron‐like cells by β‐mercaptoethanol (BME). Cells in the Blank group were not treated by BME. mRNA expression of miR‐125b was determined with qRT‐PCR in each group. mRNA and protein expressions of seven nerve cell markers, including β3 tubulin, neural microtubule‐associated protein (MAP‐2), neurofilament protein (NF), neuron‐specific enolase (NSE), glial fibrillary acidic protein (GFAP), Nestin and Vimentin in the four groups were determined by RT‐PCR and Western blots. GFAP and Nestin were detected by immunofluorescent and immunocytochemistry assays. Compared with BME group, mRNA and protein expression of β3 tubulin, MAP‐2, NF, NSE, GFAP, Nestin were significantly increased in the Mimic + BME group (P < 0.01), but significantly decreased in the Inhibitor + BME group (P < 0.01). Cells in the Mimic + BME group were more like nerve cells in morphology than cells in the BME groups. Thus miR‐125b can promote BME to induce rat BMSCs differentiation into neuron‐like cells.
Role of melatonin receptor MT2 and quinone reductase II in the regulation of the redox status of 3T3‐L1 preadipocytes in vitro
Authors: Adamczyk‐Sowa Monika , Sowa Pawel , Zwirska‐Korczala Krystyna , Pierzchala Krystyna , Bartosz Grzegorz , Sadowska‐Bartosz Izabela
Published: August 2013

We have examined the role of melatonin receptor MT2 and quinone reductase II in the regulation of the redox status of preadipocytes (3T3‐L1) in vitro. 3T3‐L1 cells were treated with melatonin at a physiological concentration (10−9 mol/L) and a supraphysiological (pharmacological) concentration (10−3 mol/L) for 24 h. Luzindole (10−4 mol/L), an antagonist of MT2 receptor, and prazosin (10−5 mol/L), an inhibitor of quinone reductase II, were added 30 min before subsequent exposure of the cells to melatonin. The level of oxidative stress was determined by the analysis of activities of enzymes neutralising reactive oxygen species, and determination of the malondialdehyde (MDA) content. Melatonin increased activities of manganese and copper‐zinc superoxide dismutase (MnSOD, Cu/ZnSOD) and catalase (CAT) at both a physiological concentration (10−9 mol/L) and a pharmacological concentration (10−3 mol/L). MDA content was unchanged, whereas activities of glutathione peroxidase (GSH‐Px) and glutathione reductase (GSSG‐Rd) were increased only by the physiological concentration. Both effects were partially inhibited by luzindole, but not prazosin. These observations suggest that melatonin, acting at least partially via MT2 receptors, can increase antioxidant enzymes activities in 3T3‐L1 preadipocytes.
A novel therapeutic drug for colon cancer: EpCAM scFv‐truncated protamine (tp)‐siRNA
Authors: Hao Huiwen , Zhen Yansen , Wang Zaicun , Chen Fulin , Xie Xin
Published: August 2013

Colon cancer is a type of malignant tumor that causes considerable mortality worldwide. Epithelial cellular adhesion molecule (EpCAM), a tumor‐associated antigen of colon tumors, is a target for colon cancer therapy. EpCAM‐specific monoclonal antibodies (mAbs) have been applied in human colon cancer since the 1990s; however, the therapeutic effects are limited. EpCAM activates nuclear signaling pathways by releasing its intracellular domain (EpICD). The released EpICD stimulates the Wnt/β‐catenin signaling pathway, which is also strongly associated with tumorigenesis. EpCAM is also a target gene of the Wnt/β‐catenin signaling pathway. EpCAM and the Wnt/β‐catenin signaling pathway form a functional interaction cycle in colon cancer. Thus, we propose a new therapeutic drug for colon cancer: an EpCAM single‐chain fragment variable antibody (scFv)‐truncated protamine‐siRNA. EpCAM scFv can recognize and bind colon cancer cells through its EpCAM antigen activity. Furthermore, the specific siRNA transferred into colon cancer cells specifically inhibits Wnt/β‐catenin signal transmission. Therefore, this new drug may efficiently interrupt the functional cycle between EpCAM and Wnt/β‐catenin signaling and be an effective therapeutic strategy for colon cancer.
Role of CD271 enrichment in the isolation of mesenchymal stromal cells from umbilical cord blood
Authors: Attar Armin , Ghalyanchi Langeroudi Arash , Vassaghi Attyieh , Ahrari Iman , Maharlooei Mohsen Khosravi , Monabati Ahmad
Published: August 2013

Isolation of mesenchymal stromal cells (MSCs) from the umbilical cord blood (UCB) has a success rate of 25% and is frequently contaminated by osteoclast‐like cells (OLCs). CD271 is a well‐known marker for the enrichment of bone marrow (BM) MSCs. We have assessed the effect of CD271 isolation on the isolation rate of MSCs from UCB. Twenty‐one samples of UCB were collected. Ten samples of UCB and five of BM underwent CD271 isolation using magnetic activated cell sorting. The other 11 UCB samples were used as the control. The isolated cells were cultured and MSC isolation was confirmed with respect to morphology, flow cytometry, adipogenic and osteogenic differentiation potentials. CD271‐positive UCB cells did not show outgrowth despite 54.5% MSCs isolation in the non‐enriched portion. No OLC was noted in the CD271‐enriched group, but 66% of the non‐enriched samples were contaminated. All the CD271‐positive BM cells formed MSC colonies. Although the per cent of CD271+ cells showed no difference between BM‐mononuclear cells (MNCs) and UCB‐MNCs, the haematopoietic marker, CD45, was found in a higher percentage of CD271‐positive UCB‐MNCs. The results of our study indicate that, although CD271 is a valuable marker for enrichment of MSCs from BM, it does not contribute to isolation of MSCs from UCB. In this source, most of the CD271+ cells are from haematopoietic origin, and possibly the process of isolation may eliminate the very low frequent MSCs and the isolation therefore fails.
AEG‐1 participates in TGF‐beta1‐induced EMT through p38 MAPK activation
Authors: Wei Jiali , Li Zhuori , Chen Wenfang , Ma Chunyang , Zhan Feng , Wu Wei , Peng Youming
Published: August 2013

Epithelial–mesenchymal transition (EMT) is an important cellular event in organogenesis, cancer and renal tubulointerstitial fibrosis. Transforming growth factor‐beta1 (TGF‐beta1) is the key inducer of EMT and the p38 mitogen‐activated protein kinases (p38 MAPK), an major intracellular signal transduction pathway is involved in TGF‐beta1‐induced EMT. Astrocyte elevated gene‐1 (AEG‐1) represents an chief genetic determinant regulating multiple events in tumorigenesis. Our present study is to explore the role of AEG‐1 in TGF‐beta1‐induced p38 MAPK activation and EMT process in human renal tubular epithelial (HK‐2) cells. The protein expressions of AEG‐1, the markers of EMT and p38 phosphorylation were measured by Western blot. The protein expression of AEG‐1 was increased in HK‐2 cells treated with TGF‐beta1. Knockdown of AEG‐1 potently inhibited phosphorylation of p38 MAPK and reversed TGF‐beta1‐induced EMT. Over‐expression of AEG‐1 via AEG‐1 transfection elicited p38 MAPK phosphorylation and promoted EMT. The effects of AEG‐1 during EMT were blocked by a p38‐specific inhibitor. Our findings suggest that AEG‐1 plays an important role in TGF‐beta1‐induced EMT through activation of p38 MAPK in proximal tubular epithelial cells.