bluuebunny/crossref_metadata_2025 huggingface dataset:
Pharmacological doses of melatonin induce alterations in mitochondrial mass and potential, bcl‐2 levels and K+ currents in UVB‐exposed U937 cells
Authors: Canonico Barbara , Luchetti Francesca , Ambrogini Patrizia , Arcangeletti Marcella , Betti Michele , Cesarini Erica , Lattanzi Davide , Ciuffoli Stefano , Palma Fulvio , Cuppini Riccardo , Papa Stefano
Published: August 2013

Apoptosis is observed in ‘actively’ dying cells after the exposure to cell stressors such as ultraviolet light irradiation. Since melatonin has been proposed to act under stressful conditions as cell protection factor, in this study we examined the potential of this molecule when used at pharmacological concentrations to control mitochondrial damage and apoptotic signalling of UVB irradiated U937 human leukaemic cells. Moreover, the effect of melatonin treatment on electrophysiological properties and membrane K+ currents of irradiated U937 cells was investigated as functional aspects relevant to the anti‐apoptotic role of melatonin. The general effect is associated with the restoration of mass, number and membrane potential of mitochondria, with a lower caspase activation and bcl‐2 upregulation. In the presence of the caspase inhibitor ZVAD‐Fmk, melatonin seems to drive UVB stressed cells to follow the mitochondrial intrinsic pathway, interfering just at the mitochondrial level. Moreover, treatment with melatonin, as well as ZVAD‐Fmk, prevented the K+ current reduction observed late following the UVB insult application, by sparing cells from death; this result also indicates that the decrease of K+ leakage currents could represent a functional feature of apoptotic process in UV‐exposed U937 cells.
Evidence of chondrocyte turnover in lung cartilage, with the probable participation of nestin‐positive cells
Authors: Ortega‐Martínez Marta , Romero‐Núñez Edgar , Niderhauser‐García Alberto , de‐la‐Garza‐González Carlos , Ancer‐Rodríguez Jesús , Jaramillo‐Rangel Gilberto
Published: August 2013

Healthy adult cartilage is thought to have little or no capacity to renewal, and cell turnover has not been reported in lung cartilage. We report that chondrocyte turnover occurs in lung cartilage, found in an unrelated study. Lung specimens from CD1 mice of 2, 6, 12, 18 or 24 months were fixed in 10% neutral‐buffered formalin and paraffin‐embedded. Apoptosis was analysed by in situ end‐labelling of fragmented DNA. Proliferating cell nuclear antigen (PCNA) and nestin were examined by immunohistochemistry. Apoptosis and PCNA were detected in lung chondrocytes. Serial section analysis showed that cells in apoptosis were different from PCNA‐positive cells, indicating that turnover was occurring. Chondrocytes were negative for nestin. Nestin‐positive cells were present in connective tissue associated with cartilage, in some specimens in close proximity of it and in perivascular cells. Thus cell turnover in lung cartilage is possible, which may be mediated by nestin‐positive cells.
Comparison of human first and third trimester placental mesenchymal stem cell
Authors: Park Saeyoung , Koh Seong‐Eun , Hur Chang Young , Lee Won‐Don , Lim Jinho , Lee Young‐Jay
Published: August 2013

Placenta mesenchymal stem cells (PMSCs) have the characteristic features of stem cells including renewability in vitro, surface expression, differentiation potency and ability to adhere to the culture surface. PMSCs expressed genes are normally found in the embryonic tissues before the onset of gastrulation, indicating multipotency. However, the stemness can depend on the stages of the placenta from which the cells were isolated. PMSCs were isolated from two different stages of placenta for comparison, that is the first and third trimesters. Both sets had very similar patterns of surface expression as CD44, CD73, CD90 and CD105, and of self renewability in vitro. Expressions of pluripotency‐coupled genes were also confirmed in both sets of cells; however, there was a significant difference in the expression levels: fPMSC (mesenchymal stem cells isolated from the first trimester human placenta) being 2–11‐fold higher than tPMSC (mesenchymal stem cells isolated from the third trimester human placenta). Possibly due to the difference in the expression levels of the pluripotency‐related genes, induction of genes specific to the ectodermal tissues were more prominent in fPMSC than tPMSC after induced differentiation.
Role of HMGB1 in propofol protection of rat intestinal epithelial cells injured by heat shock
Authors: Tang Jing , Deng Peng , Jiang Yu , Tang Youqing , Chen Bin , Su Lei , Liu Zhifeng
Published: August 2013

Gut‐derived endotoxin and pathogenic bacteria may be important causative factors of morbidity and death during heat stroke. However, as the key component of intestinal mucosal barrier, the molecular mechanism of how intestinal epithelial cells are injured by heat shock is remains unclear. After rat intestinal epithelial cells (IEC‐6) had been exposed to heat shock, their viability was measured. Propofol, which plays an important role in anti‐inflammation and organ protection, was investigated to see how it affected viability under this stress. Changes of high mobility group box 1 (HMGB1) in IEC‐6 cells were measured with RT‐PCR and Western blot assay at transcription and translational levels, respectively. Ethyl pyruvate (EP), a specific inhibitor of HMGB1 that can inhibit the release of HMGB1 without affecting its intracellular synthesis, was also investigated. Heat shock significantly reduced the intracellular level of HMGB1, and propofol inhibit its reduction. Propofol protected the heat shock‐injured cells, at least partly through inhibiting the release of intracellular HMGB1 to reduce the direct or indirect cell damage caused by HMGB1. Pretreatment with high concentrations of EP also attenuated heat‐shock injury.
Polypeptide chain release factor eRF3 is a novel molecular partner of survivin
Authors: Xiao Ruilin , Gao Yan , Shen Quan , Li Cui , Chang Wenjuan , Chai Baofeng
Published: August 2013

The eukaryotic class II polypeptide chain release factor (eRF3) is an eRF1‐ and ribosome‐dependent GTPase involved in translation termination of protein biosynthesis. eRF3 is a multifunctional protein that is also involved in chromosomal segregation and cytokinesis during mitosis. Survivin is a member of the inhibitor of apoptosis protein (IAP) family that is involved in the organisation of spindle and cell apoptosis. Interaction between survivin and eRF3a‐F3 or eRF3b, encoded by the GSPT1 and GSPT2 genes, respectively, was confirmed using yeast two‐hybrid (Y2H) and pull‐down assays in vitro, and co‐immunoprecipitation in vivo. The domains involved in the formation of the survivin–eRF3s complex have been identified. The sites on survivin that interact with eRF3 are located in the baculovirus IAP repeat domain (residues 65–76), which forms a beta‐strand structure with an overall negative charge. The sites on eRF3 that interact with survivin were localised to the N‐terminal domain(NTD; residues 131–200). Cell localisation experiments indicate that both factors are in the nucleus, suggesting that they cooperatively function in nuclear processes.
Cytokine profiles, signalling pathways and effects of fluticasone propionate in respiratory syncytial virus‐infected human foetal lung fibroblasts
Authors: Seki Erina , Yoshizumi Masakazu , Tanaka Ryota , Ryo Akihide , Ishioka Taisei , Tsukagoshi Hiroyuki , Kozawa Kunihisa , Okayama Yoshimichi , Okabe‐Kado Junko , Goya Tomoyuki , Kimura Hirokazu
Published: August 2013

To examine cytokine production in response to RSV infection, we assessed the levels of 29 cytokines released from RSV‐infected human foetal lung fibroblasts. We also examined the relationships between the effects of fluticasone propionate and various signalling pathways in the cells. Twenty‐four hours after infection (1MOI), RSV‐infected cells released cytokines, for example proinflammatory cytokines (IL‐1β, IL‐6 and TNF‐α), anti‐inflammatory (IL‐1ra), Th1 (IFN‐γ, IFN‐λ1a, IL‐2 and IL‐12), Th2 (IL‐4, IL‐5, IL‐10 and IL‐13), granulopoiesis‐inducing (G‐CSF and GM‐CSF), eosinophil recruitment‐inducing (eotaxin and RANTES) and neutrophil recruitment‐inducing cytokines (IL‐8, IP‐10, MCP‐1 and MIP‐1α). Aberrant release of most was significantly suppressed by fluticasone propionate. Twelve hours after RSV infection, increased phosphorylation of Akt, p38 MAPK, ERK1/2 and IκB‐α was noted. Fluticasone propionate suppressed the phosphorylation of Akt, p38 MAPK, and ERK1/2, but not IκB‐α, in virus‐infected cells. TLR‐4 expression was unchanged in control and RSV‐infected cells, and TLR‐3 and RIG‐I expression was not detected. The results indicate that RSV infection induces aberrant production and release of certain cytokines through these signalling pathways in human lung fibroblasts. Overproduction and imbalance of these cytokines may be associated with the pathophysiology of RSV‐induced excessive and allergic inflammation.
Death inducing and cytoprotective autophagy in T‐47D cells by two common antibacterial drugs: sulphathiazole and sulphacetamide
Authors: Mohammadpour Raziye , Safarian Shahrokh , Sheibani Nader , Norouzi Saeed , Razazan Atefeh
Published: August 2013

The broad spectrum of the pharmacological effects of sulphonamide family of drugs motivated us to investigate the cellular mechanisms for anti‐cancer effects of sulphathiazole and sulphacetamide on T‐47D breast cancer cells. Fluorescent microscopy, flow cytometric analysis, caspase‐3 activity and DNA fragmentation assays were used to detect apoptosis. The distribution of the cells among different phases of the cell cycle was measured by flow cytometry. The expression of several genes with important roles in some critical cellular pathways including apoptosis, mTOR/AKT pathway and autophagy were determined by real‐time RT‐PCR analysis. Sulphathiazole and sulphacetamide induced anti‐proliferative effects on T‐47D cells were independent of apoptosis and cell cycle arrest. The overexpression of critical genes involved in autophagy including ATG5, p53 and DRAM indicated that the main effect of the drug‐induced anti‐proliferative effects was through induction of autophagy. This process was induced in two different forms, including death inducing and cytoprotective autophagy. Sulphathiazole treatment was followed by higher expression of p53/DRAM and downregulation of Akt/mTOR pathway resulting in death autophagy. In contrast, sulphacetamide treatment lowered expression of p53/DRAM pathway in parallel with upregulation of Akt/mTOR pathway promoting cytoprotective autophagy. The results indicated that autophagy is the main mechanism mediating the anti‐cancer effects of sulphathiazole and sulphacetamide on T‐47D cells. Alignment of the p53 and DRAM expression along with activation level of Akt survival pathway therefore determines the type of autophagy that occurs.
Intestinal stem cells – types and markers
Authors: Zhang Zhigang , Huang Jian
Published: August 2013

Intestinal epithelium is a rapidly cycling tissue, always renewing every 4–5 days under normal conditions, which is maintained by intestinal stem cells (ISCs). Using the fluorescence labelling trace, ISCs can be divided into two different types: active intestinal stem cells (A‐ISCs) located in bottom of the intestinal crypt and the quiescent intestinal stem cells (Q‐ISCs) in the +4 position of the crypt. There is a complex signal regulation net between the ISCs and other intestinal cells, such as Wnt and Notch pathways. ISCs have an intimate relationship with the colorectal cancer (CRC). However, a deficiency of stem cells markers severely limits research on the biological characteristics of ISCs. We have reviewed several ISCs markers, including Lgr5, PHLDA1, Bmi1 and Lrig1. These markers have widely different biological functions, but also have a close relationship with cancers, especially CRC. Our hypothesis concerns the reasons for ISCs having two distinct types and why endless ISCs markers have emerged.
Insulin at a unicellular eukaryote level
Authors: Csaba György
Published: August 2013

The unicellular ciliate, Tetrahymena, has been the main model for studying the hormonal system of unicellular animals. Tetrahymena produce, store, secrete and take up insulin, the hormone being similar to that of mammals, both immunocytochemically and functionally. The plasma membrane and nuclear envelope of Tetrahymena have insulin receptors, which are structurally similar to the mammalian receptor, as it their binding capacity. The cell has also second messengers and signal pathways for insulin. Insulin influences the synthesis of other hormones. The first short encounter between the cell and insulin provokes the hormonal imprinting that alters the function of the cells and is transmitted to the progeny, and can persist for over 1,000 generations, in hormone binding, hormone content, phagocytosis, cell growth and movement. Insulin has a survival function in Tetrahymena and during stress insulin production and binding are elevated. Other protozoa also react to insulin, and the evolutionary aspects are discussed in this review since it is still not appreciated that the hormones are of great antiquity in the animal kingdom.
Higher propensity of Wharton's jelly derived mesenchymal stromal cells towards neuronal lineage in comparison to those derived from adipose and bone marrow
Authors: Balasubramanian Sudha , Thej Charan , Venugopal Parvathy , Priya Nancy , Zakaria Zubaidah , SundarRaj Swathi , Majumdar Anish Sen
Published: August 2013

Mesenchymal stromal cells (MSCs) derived from different tissue sources are capable of differentiating into neural and glial cell types. However, the efficiency of differentiation varies between MSCs derived from different tissues. We compared the efficiency of neural progenitor population generation between adipose (AD), bone marrow (BM) and Wharton's jelly (WJ) derived MSCs. MSCs isolated from the three sources were induced to form primary neurospheres using epidermal growth factor (20 ng/mL) and bFGF (20 ng/mL). The self‐renewal potential of the primary neurospheres was assessed by secondary neurosphere assay. Primary neurospheres were differentiated to neuronal lineage on fibronectin‐coated dishes. The neurospheres and the resulting differentiated cells were characterized by immunocytochemistry and the RT‐PCR analyses. We have also investigated the secretome profile of neuronal‐related growth factors using Ray biotech cytokine array. The results show that MSCs from the three sources can be induced to generate neurospheres and they expressed neural progenitor markers nestin, Sox2 and Pax6 transcription factors. When differentiated on fibronectin coated dishes in mitogen free culture conditions, the primary spheres from all three sources were able to generate neuron/glial ‐ like cells which expressed Nfl, Map2 and GFAP with varied efficiency. Self‐renewal potential of these progenitors was determined by secondary sphere formation. WJ‐ and BM‐derived neurospheres were able to self‐renew, while AD derived progenitors failed to do so. Comparison of the secretome profile suggested that WJ derived MSCs secrete more neurotrophic factors. The data suggest that human WJ derived MSCs can be induced to make neural progenitors with higher efficiency compared to BM and AD derived MSCs.
Aurora B kinase is required for cytokinesis through effecting spindle structure
Authors: Wu Qiang , Zhang Wei , Mu Tianhao , Song Tianqiang , Li Dengwen
Published: August 2013

The spindle, composed of microtubules and associated proteins, serves as major machinery for cell division. Aurora B is a chromosome passenger protein that has important functions in mitosis. Its dynamic distribution at mitosis goes along with spindle structure and dynamics. We used the siRNA technique to knockdown protein expression and immunofluorescence technique to follow Aurora B during mitosis. Aurora B regulates microtubule plus ends as mitosis progresses, including both kinetochore and polar microtubules. Interactions between Aurora B and polar microtubule plus ends lead to failure in cytokinesis and abnormal midbody structure. We think Aurora B may not only play a role as kinase, but regulate microtubule plus ends.
Novel interactions of GRP78: UPR and estrogen responses in the brain
Authors: Avila Marco Fidel , Cabezas Ricardo , Torrente Daniel , Gonzalez Janneth , Morales Ludis , Alvarez Lisandro , Capani Francisco , Barreto George E.
Published: August 2013

Glucose‐regulated protein 78 (GRP78; 78 kDa) belongs to a group of highly conserved heat shock proteins (Hsp) with important functions at the cellular level. The emerging interest for GRP78 relies on its different functions, both in normal and pathological circumstances. GRP78 regulates intracellular calcium, protein shaping, endoplasmic reticulum (ER) stress and cell survival by an immediate response to insults, and that its expression may also be regulated by estrogens. Although these roles are well explored, the mechanisms by which GRP78 induces these changes are not completely understood. In this review, we highlight various aspects related to the GRP78 functioning in cellular protection and repair in response to ER stress and unfolded protein response by the regulation of intracellular Ca2+ and other mechanisms. In this respect, the novel interactions between GRP78 and estrogens, such as estradiol and others, are analyzed in the context of the central nervous system (CNS). We also discuss the importance of GRP78 and estrogens in brain diseases including ischemia, Alzheimer's and Huntington's disorders. Finally, the main protective mechanisms of GRP78 and estrogens during ER dysfunction in the brain are described, and the prospective roles of GRP78 in therapeutic processes.
Coexpression of recombinant adenovirus carrying GDNF and EDNRB genes in neural stem cells in vitro
Authors: Sun Nian‐Feng , Zhong Wen‐Yu , Lu Sheng‐Ai , Tian Yu‐Ling , Chen Jing‐Bo , Hu San‐Yuan , Tian Ai‐Ling
Published: August 2013

Gene therapy and nerve stem cells isolated from the developing human enteric nervous system (ENS) are significant. They may open the route for the cell therapy of Hirschsprung's disease (HD). We have constructed the recombinant adenovirus‐carrying glial cell line‐derived neurotrophic factor (GDNF) and endothelin receptor B (EDNRB) gene, and investigated the exosomatic coexpression in neural stem cells. The recombinant adenovirus Ad‐GE coexpressing GDNF and EDNRB gene was constructed by the AdEasy system and confirmed by the reverse transcription polymerase chain reaction (RT‐PCR) method. Expression of exogenous genes in neural stem cells after transfection was confirmed by the flow cytometry and real‐time fluorescence quantitative PCR. Fragments of pAd Track‐CMV‐GE were consistent with GDNF and EDNRB. The green fluorescence of the positive cells was followed by fluorescence microscopy at 24 h after NSCs had been transfected, reaching a peak at 72 h after transfection. Flow cytometry showed that the efficiency of transfection was 15.0, 23.6, and 25.4% at 24, 48 and 72 h respectively. Real‐time fluorescence quantitative PCR showed the expression levels of mRNA of GDNF and EDNRB in 48 and 72 h groups were obviously higher than that in 24 and 96 h groups. Recombinant adenovirus carrying GDNF and EDNRB genes are coexpressed in neural stem cells, which may offer the possibility of a novel approach to local combination gene therapy for Hirschsprung's disease.
Vinculin, cell mechanics and tumour cell invasion
Authors: Goldmann Wolfgang H. , Auernheimer Vera , Thievessen Ingo , Fabry Ben
Published: August 2013

The focal adhesion protein, vinculin, is important for transmitting mechanical forces and orchestrating mechanical signalling events. Deregulation of vinculin results in altered cell adhesion, contractility, motility and growth, all of which are important processes in cancer metastasis. This review summarises recent reports on the role of vinculin in cellular force generation and signalling, and discusses implications for a role of vinculin in promoting cancer cell migration in 3D environments.
Association between a functional variant at PTGS2 gene 3′UTR and its mRNA expression in lymphoblastoid cell lines
Authors: Wang Xueting , Zhao Yunzhao , Wang Yanru , Wang Zexing , Guan Xiaoxiang
Published: August 2013

PTGS2 genetic 3′ untranslated region (3′UTR) miRNA binding sites variants are significantly associated with cancer risk; however, the roles of genetic variants in PTGS2 gene 3′UTR and post‐transcriptional regulation have not been elucidated. We report that rs689470 and rs5275 in the PTGS2 3′UTR have potential miRNA‐binding sites by using bioinformatics analysis. However, only the rs689470 was significantly associated with PTGS2 mRNA expression in lymphoblastoid cell lines (P = 0.026), but not for rs5275 (P = 0.626). rs689470 might be putative variants mediating the post‐transcriptional regulation of target PTGS2 gene. Better understanding of how 3′UTR variants regulate PTGS2 activity will pave the way to targeting the PTGS2 pathway in cancer therapy.
Direct TGF‐β1 signaling between activated platelets and pancreatic cancer cells primes cisplatin insensitivity
Authors: Chen Hongxu , Lan Xiang , Liu Menggang , Zhou Bo , Wang Baoling , Chen Ping
Published: August 2013

The exact mechanisms underlying chemotherapy insensitivity in pancreatic cancer remain largely unclear. The dynamic cross talk between tumors and their microenvironment is an important determinant of cancer chemosensitivity. However, whether additional signals provided during the intravascular transit of tumor cells affect the sensitivity of pancreatic cancer cells to chemotherapy is unknown. We have found that activated platelet–cancer cell interactions are sufficient to prime cisplatin (CDDP) insensitivity in pancreatic cancer cells. Activated platelet‐derived TGF‐β1 rather than direct platelet–tumor cell contacts stimulates PI3K/Akt and MEK/Erk signaling in pancreatic cancer cells, resulting in reduction of CDDP sensitivity in these cells. Therefore, the platelet–tumor cell interactions and the relevant signaling pathways that prime CDDP insensitivity may be potential targets for adjuvant chemotherapy for pancreatic cancer.
Inhibition of ATP‐induced calcium influx by homocysteine in human umbilical vein endothelial cells
Authors: Cortés Magdalena P. , Becerra Juan P. , Vinet Raúl , Álvarez Rocío , Quintana Irene
Published: August 2013

Mechanisms involved in the association between hyperhomocysteinemia and vascular occlusive diseases remain unclear. Homocysteine (Hcy) may disturb calcium (Ca2+) cytosolic regulation in endothelial cells, a process that can directly affect the synthesis of vasoactive substances, such as nitric oxide (NO). We have investigated the effect of acute and chronic incubation with high concentrations of Hcy (100 and 500 μmol/L) on the changes in the intracellular Ca2+ concentration ([Ca2+]i) induced by ATP, using primary cultures of human umbilical vein endothelial cells (HUVEC). The changes in [Ca2+]i, expressed as ΔFt/Fb, were measured using the microspectrofluorimetric technique with Fluo‐3 as Ca2+ indicator. HUVEC acutely exposed to Hcy did not produce significant effects on any of the parameters studied. However, chronic exposition (24 h) caused a significant decrease in the speed of store‐mediated Ca2+ entry, expressed as (ΔFt/Fb)/t (s−1). Exposure of HUVEC to 100 and 500 µmol/L Hcy gave significantly lower values (0.019 ± 0.002 s−1, n = 5 and 0.021 ± 0.004 s−1, n = 6, respectively) compared to the controls (0.046 ± 0.004 s−1, n = 8, P < 0.003). This was detected only when the sustained phase of the ATP‐induced [Ca+2]i increase was isolated. These results demonstrate that high concentrations of Hcy can affect the mechanisms involved in [Ca2+]i regulation of HUVEC, and that alteration occurs specifically in the sustained phase, which has been directly associated with NO synthesis.
Rho kinase pathway is likely responsible for the profibrotic actions of aldosterone in renal epithelial cells via inducing epithelial–mesenchymal transition and extracellular matrix excretion
Authors: Wei Jiali , Li Zhuori , Ma Chunyang , Zhan Feng , Wu Wei , Han Hui , Huang Yun , Li Wenning , Chen Daojun , Peng Youming
Published: August 2013

It has been demonstrated that aldosterone (ALD) plays a direct profibrotic role in the kidney but the underlying mechanism remains unclear. We examined the role of Rho kinase signal pathway in epithelial–mesenchymal transition (EMT) process and extracellular matirx (ECM) synthesis induced by ALD in human renal proximal tubular epithelial (HK‐2) cells in vitro. Rho kinase and collagen I, III protein expressions were detected by ELISA. E‐cadherin, α‐smooth muscle actin (SMA), collagen_I and collagen III mRNA expressions were detected by real time PCR. E‐cadherin, and α‐SMA protein expressions were measured by Western blot. Our results showed that ALD could significantly activate the Rho kinase in HK‐2 cells, while in the presence of mineralocorticoid receptor (MR) antagonist eplerenone and Rho kinase inhibitor Y27632, the Rho kinase protein expression were almost completely prevented. Exposure of HK‐2 cells to ALD for 48 h induced EMT as evidenced by loss of E‐cadherin, and de novo expression of α‐SMA. The EMT was completely blocked by eplerenone and Y27632. Meanwhile, ALD could significantly increase the mRNA and protein expressions of collagen I, III in HK‐2 cells when compared with the control group, while eplerenone and Y27632 could almost reverse these effects. These observations suggest that ALD can activate Rho kinase pathway and Rho kinase pathway is likely responsible for the profibrotic actions of ALD in renal proximal tubular epithelial cells via inducing EMT and ECM excretion.
Aberrant expression of microRNAs involved in epithelial–mesenchymal transition of HT‐29 cell line
Authors: Cai Zhi‐Gang , Zhang Shao‐Ming , Zhang Hang , Zhou Yi‐Yong , Wu Hai‐Bo , Xu Xiao‐Ping
Published: August 2013

Epithelial–mesenchymal transition (EMT) is an essential step for cancer metastasis. MicroRNAs (miRNAs) are small non‐coding RNAs that regulate target‐mRNAs post‐transcriptionally. The expression and function of miRNAs in EMT of HT‐29 colonic cells remain elusive. This study looks at expression of miRNAs in EMT and explores the effects of miRNAs on EMT in HT‐29 cell line. HT‐29 was treated with TGF β to establish an EMT model, in which a collection of miRNAs was dynamically regulated by real‐time PCR (qPCR) analysis. Among them, miR‐21 and miR‐27 were significantly upregulated, while miR‐22, miR‐26, miR‐30, miR‐181, miR‐200b, miR‐200c and miR‐214 were markedly downregulated. MiRNA‐inhibitors were used to knockdown miRNAs in HT‐29 and EMT markers were determined by qPCR to monitor the effects of miRNAs on EMT process. Results showed that miR‐22 could not alter the expression of EMT markers, while knockdown of miR‐200b could significantly increase that of epithelial markers, N‐cadherin, Vimentin, α‐Sma and Twist1 and decrease that of mesenchymal marker, E‐cadherin. Bioinformatic analysis and Western blot showed that ZEB1 was directly suppressed by miR‐200b. In conclusion, miRNAs are dynamically regulated in TGF β‐induced EMT of HT‐29 and miR‐200b was essential for EMT by suppressing the expression of ZEB1 in HT‐29.
Canine mesenchymal stem cells (MSCs): characterization in relation to donor age and adipose tissue‐harvesting site
Authors: Guercio Annalisa , Di Bella Santina , Casella Stefania , Di Marco Patrizia , Russo Carmelo , Piccione Giuseppe
Published: August 2013

Adipose tissue as a stem cell source is ubiquitously available and has several advantages compared to other sources, for example it is easily accessible in large quantities with minimal invasive harvesting procedure, and isolation of adipose‐derived mesenchymal stem cells (MSCs) yields a high amount of stem cells, essential for stem cell‐based therapies and tissue engineering. We have explored the effect of donor age, and the anatomical origin of the adipose tissue on several aspects of MSCs in dogs, such as cell yield, proliferative ability, multi‐differentiation potential, colony‐forming capacity, stemness marker expression. We also assessed the effect of cell passaging on the MSCs stemness. We found that the anatomical origin of the adipose tissue and the age of donors have effects only on the proliferative capacity of the MSCs. Moreover, cells show a progressive loss of the stemness characteristics with passages. Cell therapies need a suitable number of cells to use in clinical applications. Characterization of MSCs at different passages, allowed us to demonstrate that, under our culture conditions, the best quantitative and qualitative characteristics are obtained at early passages. Adult MSCs are of particular interest for the therapeutic approach to musculoskeletal diseases, and the dog provides an excellent preclinical model for the development of new approaches in regenerative medicine that might be applied to humans.
Histone H3 is absent from organelle nucleoids in BY‐2 cultured tobacco cells
Authors: Takusagawa Mari , Tamotsu Satoshi , Sakai Atsushi
Published: August 2013

The core histone proteins (H2A, H2B, H3 and H4) are nuclear‐localised proteins that play a central role in the formation of nucleosome structure. They have long been considered to be absent from extra‐nuclear, DNA‐containing organelles; that is plastids and mitochondria. Recently, however, the targeting of core histone H3 to mitochondria, and the presence of nucleosome‐like structures in mitochondrial nucleoids, were proposed in cauliflower and tobacco respectively. Thus, we examined whether histone H3 was present in plant organelles and participated in the organisation of nucleoid structure, using highly purified organelles and organelle nucleoids isolated from BY‐2 cultured tobacco cells. Immunofluorescence microscopic observations and Western blotting analyses demonstrated that histone H3 was absent from organelles and organelle nucleoids, consistent with the historical hypothesis. Thus, the organisation of organelle nucleoids, including putative nucleosome‐like repetitive structures, should be constructed and maintained without participation of histone H3.
Synaptic adhesion molecules in Cadm family at the neuromuscular junction
Authors: Tanabe Yuko , Fujita Eriko , Hayashi Yukiko K. , Zhu Xinran , Lubbert Hermann , Mezaki Yoshihiro , Senoo Haruki , Momoi Takashi
Published: August 2013

RA175/SynCAM1/Cadm1 (Cadm1), a member of the immunoglobulin superfamily, is a synaptic cell adhesion molecule that has a PDZ‐binding motif at the C‐terminal region. It promotes the formation of presynaptic terminals and induces functional synapses in the central nervous system. Cadm1‐deficient (knockout [KO]) mice show behavioral abnormalities, including excessive aggression and anxiety, but do not show any symptoms of neuromuscular disorder, although neuromuscular junctions (NMJs) have structures similar to synapses. We have examined the expression of members of the Cadm family in the mouse muscle tissues. Cadm4 and Cadm1 were major components of the Cadm family, and Cadm3 was faintly detected, but Cadm2 was not detected by RT‐PCR. Cadm4 as well as Cadm1 colocalized with alpha‐bungarotoxin at the NMJs and interacted with the multiple PDZ domain protein Mupp1. Cadm4 was expressed in Cadm1‐KO mice and might compensate for Cadm1 loss through interactions with Mupp1.
Silencing Cofilin‐1 blocks PDGF‐induced proliferation in eutopic endometrium of endometriosis patients
Authors: Wang Dan‐Bo , Xu Yan‐Li , Chen Peng , Chen Ying‐Han , Li Yan
Published: August 2013

Platelet‐derived growth factor (PDGF) is well known to induce proliferation in variety of cells. The aim of this study was to investigate whether there is a correlation between the expression of Cofilin‐1 (CFL1) and the proliferation of eutopic endometrium stromal cells (ESC) in response to PDGF stimulation. Results show that PDGF induced the expression of CFL1 in ESC and promoted proliferation of ESC significantly in a time‐ or dose‐dependent manner. After silencing CFL1, the effect of PDGF on promoting ESC proliferation was significantly decreased. These data demonstrate that PDGF can induce CFL1 expression in ESC. Silencing CFL1 blocks PDGF‐induced proliferation in ESC.
Myocardial structural protein expression in umbilical cord blood mesenchymal stem cells after myogenic induction
Authors: Ma Nan , Ding Fangbao , Zhang Junwen , Bao Chunrong , Zhong Hong , Mei Ju
Published: August 2013

To assess the effects of three methods of inducing myogenic cells differentiation, umbilical blood mesenchymal stem cells (UCMSCs) from nearly full‐term pregnancy mongrel dogs were purified and cultured. Fourth‐passage UCMSCs were used to detect surface antigens, including CD11a, CD11b, CD29, CD34 and CD71. The cells were induced by 5‐azacytidine (5‐aza), myocardial lysates and myocardial induced fluid. Positive expression of Nkx2.5, α‐actin, desmin, β‐MHC and troponin‐I (TN I) were detected after 3 weeks. The immunohistochemical results were CD11a (−), CD11b (−), CD34 (−), CD29 (+) and CD71 (+). Nkx2.5 was detected in 5‐aza group, myocardial lysates group and myocardial induced fluid group. Semi‐quantitative analysis showed Nkx2.5 expression significantly higher in myocardial lysates group than in the 5‐aza group or myocardial‐induced fluid group (P < 0.05), but there was no significant difference between the 5‐aza and myocardial‐induced fluid groups for Nkx2.5 expression (P > 0.05). MSCs did not express myocardial structural proteins before differentiation, but α‐actin, desmin, β‐MHC and troponin‐I were present after differentiation. The positive expression of four proteins differed with the differentiation conditions. The UCMSCs can be differentiated into myogenic cells by three methods, but the degrees of differentiation are inconsistent. Our results show that the effects of 5‐aza and myocardial lysates are better than that of myocardial induced fluid.
Homage to Rita Levi‐Montalcini, the queen of modern neuroscience
Authors: Aloe Luigi , Chaldakov George N.
Published: August 2013

The first cell growth factor, nerve growth factor (NGF), was discovered by Rita Levi‐Montalcini (RLM) in the early 1950s. Originally identified as neurite outgrowth‐stimulating factor, later studies revealed that non‐neuronal cells, including immune cells, endothelial cells, cardiomyocytes, pancreatic beta cells, prostate epithelial and adipose tissue cells, were also targets for and/or sources of NGF. Nerve growth factor is well recognised as mediating multiple biological phenomena, ranging from the neurotrophic through immunotrophic and epitheliotrophic to metabotrophic effects. Consequently, NGF and other members of the neurotrophin family are implicated in the pathogenesis of a large spectrum of neuronal and non‐neuronal diseases, ranging from Alzheimer's and other neurodegenerative diseases to atherosclerosis and cardiometabolic disorders. Recent studies have demonstrated the therapeutic potentials of NGF in these conditions, including ocular and cutaneous diseases. NGF TrkA receptor antagonists emerged as novel drugs for pain, prostate and breast cancer, melanoma and urinary bladder syndromes. Here, we briefly describe the ‘unpredictable’ ideogenesis of the discovery of NGF, a eureka in the neuroscience.
Genistein induces apoptotic cell death associated with inhibition of the NF‐κB pathway in adult T‐cell leukemia cells
Authors: Yamasaki Masao , Mine Yoshihiro , Nishimura Misato , Fujita Satoshi , Sakakibara Yoichi , Suiko Masahito , Morishita Kazuhiro , Nishiyama Kazuo
Published: August 2013

We have shown that genistein inhibits the growth of adult T‐cell leukemia (ATL) cells in vitro and in vivo, and this leads to pronounced G2/M arrest. This report shows that genistein induces apoptotic death in ATL cells. Although the pan‐caspase inhibitor, Z‐VAD‐fmk, did not inhibit genistein‐induced apoptosis, release of apoptosis‐inducing factor (AIF) into the cytosol occurred. Poly‐ADP ribose polymerase inhibition also abrogated genistein‐induced apoptosis. Genistein decreased nuclear p65 translocation and IκBα phosphorylation, and downregulated the anti‐apoptotic proteins, XIAP, cIAP and survivin, NF‐κB‐responsive gene products. Thus, genistein is a promising agent for ATL that induces caspase‐independent apoptosis through inhibition of the NF‐κB pathway.
Effect of bixin and norbixin on the expression of cytochrome P450 in HepG2 cell line
Authors: Matuo Míriam Cristina Sakuragui , de Oliveira Takamoto Rafael Teruiti , Kikuchi Irene Satiko , de Jesus Andreoli Pinto Terezinha
Published: August 2013

Bixin and norbixin are the main components of annatto, which is extracted from Bixa orellana and largely used as natural colorant in the food and pharmaceutical industries. Annatto can enhance CYP1A and CYP2B activity in rats; however, the inducer effect has not been investigated in human cell lines. In this study, the ability of bixin and norbixin to induce the cytochrome P450 (CYP) enzymes was assessed in HepG2 human hepatoma cell line. HepG2 cells were treated with bixin and norbixin, and the expression of the CYP genes quantified by real‐time reverse transcription polymerase chain reaction (RT‐PCR). Expression of CYP1A1 and CYP1A2 was significantly increased by bixin treatment, while CYP2B6, 2C9, 2E1 and 3A4 were unaffected. Cells were treated with norbixin showed no inducer effect. The results suggest that the inducer potential of annatto is attributed to bixin, but not to norbixin, despite their similarities in molecular structure.
TGF‐β1 induces a nucleus pulposus‐like phenotype in Notch 1 knockdown rabbit bone marrow mesenchymal stem cells
Authors: Morigele Morigele , Shao Zengwu , Zhang Zhicai , Kaige Ma , Zhang Yannan , Qiang Wu , Yang Shuhua
Published: August 2013

We have investigated the effects of Notch1 knockdown and treatment with TGF‐β1 on the regulation of the directional differentiation of mesenchymal stem cells (MSCs). MSCs were isolated from the femur bone of New Zealand rabbits and purified by using discontinuous gradient density centrifugation. Notch1 siRNAs were designed, synthesised and transiently transfected into these MSCs, and treated with TGF‐β1. The MSCs were examined for morphology, stained with toluidine blue for proteoglycan analysis and gene and protein expression were measured with qRT‐PCR and Western blotting respectively. They had an ellipse or fusiform shape and gathered in nests or swirls after being cultured for 7 days. Notch1 expression was knocked down with Notch1 siRNA (the silence rate was 47%; P < 0.001). After knockdown and TGF‐β1 treatment, MSCs expressed more proteoglycan (P < 0.01), and higher levels of collagen II mRNA and protein than control cells (P < 0.001). Thus knockdown of Notch1 expression in MSCs may be useful in the treatment of intervertebral disc degeneration.
Potential of co‐culture of nucleus pulposus mesenchymal stem cells and nucleus pulposus cells in hyperosmotic microenvironment for intervertebral disc regeneration
Authors: Tao Yi‐Qing , Liang Cheng‐Zhen , Li Hao , Zhang Yu‐Jie , Li Fang‐Cai , Chen Gang , Chen Qi‐Xin
Published: August 2013

Nucleus pulposus mesenchymal stem cells (NPMSCs) are a potential cell source for intervertebral disc (IVD) regeneration, but little is known about their response to IVD‐like high osmolarity (400 mOsm). This study was to investigate the viability, proliferation and protein biosynthesis of nucleus pulposus cells (NPCs), NPMSCs and co‐cultured NPMSCs‐NPCs under IVD‐like high osmolarity conditions. NPCs and NPMSCs were isolated and cultured under standard and IVD‐like high osmolarity conditions for 1 or 2 weeks. Cell viability was measured by annexin V‐FITC and PI staining, and cell proliferation measured by MTT assay. The expression of SOX‐9, aggrecan and collagen‐II was measured by RT‐PCR and Western blot analyses. IVD‐like high osmolarity condition slightly inhibited cell viability and decreased the expression of SOX‐9, aggrecan and collagen‐II at the mRNA and protein levels in all groups compared with standard condition. NPMSCs could tolerate IVD‐like high osmolarity, and NPCs‐NPMSCs co‐culture increased cell proliferation and the expression of SOX‐9, aggrecan and collagen‐II under both culture conditions, suggesting that co‐culture of NPMSCs‐NPCs has potential application for IVD regeneration.
Mechanotransduction in cancer stem cells
Authors: Hao Jin , Zhang Yueling , Ye Rui , Zheng Yingcheng , Zhao Zhihe , Li Juan
Published: August 2013

The cancer stem cell (CSC) concept, which arose about a decade ago, proposes that tumor growth is sustained by a subpopulation of highly malignant cells. These cells, termed CSCs, are capable of extensive self‐renewal that contributes to metastasis and treatment resistance. Therefore, therapeutic strategies that target CSCs should be developed for improving outcomes of cancer patients. Recent progress has highlighted the importance of physical properties of the extracellular matrix and mechanotransduction pathway in cancer cells during cancer development. On the other hand, the significance of CXCR1, an upstream signal of FAK/PI3K/Akt has been revealed in CSCs. FAK/PI3K/Akt is a key signal mediator in mechanotransduction pathway. Therefore, mechanotransduction could be a new target for CSCs, and would be an innovative way to treat cancer by inhibiting FAK/PI3K/Akt.
SiRNA‐targeted carboxypeptidase D inhibits hepatocellular carcinoma growth
Authors: Jin Tao , Fu Jie , Feng Xiao‐Jun , Wang Shou‐Mei , Huang Xue , Zhu Ming‐Hua , Zhang Shu‐Hui
Published: August 2013

Carboxypeptidase D (CPD), a membrane‐bound metallocarboxypeptidase that functions as a docking receptor for duck hepatitis B virus, is frequently overexpressed in human cancers. We have explored its expression pattern, clinical significance, and biological function of CPD in hepatocellular carcinoma (HCC). CPD expression was markedly elevated in HCCs relative to adjacent non‐tumor liver tissues, as determined by quantitative real‐time polymerase chain reaction and Western blot analysis. Immunohistochemistry showed that 164 of 400 (41%) HCCs had high expression of CPD. CPD overexpression was significantly associated with serum levels of hepatitis B surface antigen and hepatitis B e antigen, liver cirrhosis, pathological grade, and intrahepatic metastasis. Knockdown of endogenous CPD expression in Huh7 HCC cells by RNA interference reduced cell proliferation, blocked the cell cycle at G1 phase, and increased apoptosis. Many genes implicated in cell‐cycle regulation, including P21waf1, P27 Kip1, SKP2, and CDC2, were deregulated by CPD downregulation. Thus CPD is frequently upregulated in HCC, and targeting CPD inhibits HCC cell proliferation through induction of G1 cell‐cycle arrest and apoptosis, thereby providing a potential therapeutic target for this malignancy.
Knockdown of both FoxO1 and C/EBPβ promotes adipogenesis in porcine preadipocytes through feedback regulation
Authors: Yan Xiong , Weijun Pang , Ning Wei , Yu Wang , Wenkai Ren , Gongshe Yang
Published: August 2013

FoxO1 and C/EBPβ are important transcription factors during adipogenic differentiation; however, the potential mechanisms of their inter‐regulation in adipogenesis remain unknown. We found that FoxO1 and C/EBPβ are abundant in adipose tissues of 3‐ and 180‐day pigs at the mRNA and protein levels. During porcine preadipocyte differentiation, C/EBPβ expression increases to the peak at Day 2 and then decreases. In contrast, FoxO1 is lowest on Day 2 and gradually increases. Knockdown of FoxO1 or C/EBPβ with lentivirus‐mediated shRNA enhances or inhibits lipid accumulation and adipogenic maker (C/EBPα and aP2) expression, respectively. FoxO1 depletion causes a mild decrease of C/EBPβ protein level, and C/EBPβ interference also inhibits the expression of FoxO1 protein. Knockdown of both FoxO1 and C/EBPβ promotes lipid accumulation at Day 8, and increases the adipogenic markers during differentiation in comparison with the controls and the FoxO1 knockdown alone. Co‐immunoprecipitation experiments indicate that FoxO1 binds to C/EBPβ in adipose tissues and in vitro. In conclusion, the results suggest that FoxO1 and C/EBPβ regulate preadipocyte adipogenesis possibly through C/EBPβ → FoxO1 → C/EBPβ feedback regulatory loop and FoxO1‐C/EBPβ protein complex.
A plethora of human pluripotent stem cells
Authors: Gao Liyang , Thilakavathy Karuppiah , Nordin Norshariza
Published: August 2013

At the early stages of mammalian development, a number of developmentally plastic cells appear that possess the ability to give rise to all of the differentiated cell types normally derived from the three primary germ layers – unique character known as pluripotency. To date, embryonic stem cells (ESCs) and induced pluripotent stem cells (iPSCs) have been shown to be truly pluripotent. However, recent studies have revealed a variety of other cells that demonstrate pluripotentiality, including very small embryonic‐like stem cells (VSELs), amniotic fluid stem cells (AFSCs), marrow‐isolated adult multilineage inducible cells (MIAMI) and multipotent adult precursor cells (MAPCs). This review summarises key features of these six kinds of pluripotent and potentially pluripotent stem cells (ESCs, iPSCs, VSELs, AFSCs, MIAMI and MAPCs) and the evidence for their pluripotency properties.
Oxidized low‐density lipoprotein induces hematopoietic stem cell senescence
Authors: Zhang Xian‐Ping , Zhang Gui‐Hai , Wang Yu‐Ying , Liu Jun , Wei Qiang , Xu Chun‐Yan , Wang Jian‐Wei , Wang Ya‐Ping
Published: August 2013

We have investigated oxidized low‐density lipoprotein (ox‐LDL) induced senescence in hematopoietic stem cells (HCs). Mouse Sca‐1+ HCs were separated and purified using the magnetic activated cell sorting technique. Ox‐LDL induced significant senescence in HCs measured by SA‐β‐Gal staining, and reduced CFU‐Mix colony‐forming capacity, arresting cells at G0/G1 phase. In agreement with the cell cycle arrest, ox‐LDL markedly reduced the expression of CDK4, cyclin D, and cyclin E. As possible contributing factors for cell senescence, ox‐LDL also induced cellular oxidative stress and reduced telomerase activity.
Enhanced expression of MYF5 and MYOD1 in fibroblast cells via the forced expression of bos taurus MYF5
Authors: Nie Yong Wei , Ding Xiang Bin , Ge Xiu Guo , Fan Han Lu , Liu Zhong Wei , Guo Hong
Published: August 2013

The formation of vertebrate skeletal muscles widely thought to be under the control of hierarchy of regulatory genes. MYF5 is one of the myogenic determination gene expressed in the developing mouse dermomyotome which control skeletal muscle differentiation. In the current work, we had obtained the cDNA sequence including the full coding region of the bos taurus myogenic factor MYF5 by reverse transcription polymerase chain reaction. Furthermore, we examined whether fibroblast cell derived from mouse and bos taurus can be transduced using plasmid vectors carrying bos taurus MYF5. Bos taurus MYF5 activates MYF5 and MYOD1 expression after 1 day culture. The concerted upregulation of the myogenic regulatory factors enhanced myosin (skeletal fast) expression. These observation show that MYF5 is essential for myogenic differentiation and provides candidates for regulation bos taurus skeletal muscle development.