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The Journal of Visualized Experiments (JoVE) is a peer reviewed, PubMed-indexed video journal. Our mission is to increase the productivity of scientific research.

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Articles by Bing Song in JoVE

 JoVE Bioengineering

Electric Field-controlled Directed Migration of Neural Progenitor Cells in 2D and 3D Environments


JoVE 3453 2/16/2012

*1, *2,3, 1, 3, 3, 3, 1

1School of Dentistry, Cardiff Institute of Tissue Engineering & Repair, Cardiff University, 2Shandong Qianfoshan Hospital, Shandong University School of Medicine, 3Dermatology and Ophthalmology Research, Institute for Regenerative Cures, University of California at Davis

This protocol demonstrates methods used to establish 2D and 3D environments in custom-designed electrotactic chambers, which can track cells in vivo/ex vivo using time-lapse recording at the single cell level, in order to investigate galvanotaxis/electrotaxis and other cellular responses to direct current (DC) electric fields (EFs).

Other articles by Bing Song on PubMed

Electrical Cues Regulate the Orientation and Frequency of Cell Division and the Rate of Wound Healing in Vivo

Proceedings of the National Academy of Sciences of the United States of America. Oct, 2002  |  Pubmed ID: 12368473

Controlling cell division is fundamental. One environmental cue that exerts profound control over both the orientation and frequency of cell division in vivo is a naturally occurring, wound-induced electric field (EF). Wounds in rat corneas generate endogenous EFs in the plane of the epithelial sheet because the transcorneal potential difference (TCPD; +40 mV internally positive) collapses at the wound edge, but is maintained at normal levels at 0.5 mm back from the wound. We manipulated the endogenous EF this creates by using drugs with differing actions. The wound-induced EF controlled the orientation of cell division; most epithelial cells divided with a cleavage plane parallel to the wound edge and perpendicular to the EF vector. Increasing or decreasing the EF pharmacologically, respectively increased or decreased the extent of oriented cell division. In addition, cells closest to the wound edge, where the EF was highest, were oriented most strongly by the EF. Remarkably, an endogenous EF also enhanced the frequency of cell division. This also was regulated by enhancing or suppressing the EF pharmacologically. Because the endogenous EF also regulated the wound healing rate, it may act as one control of the interplay between cell migration and cell division during healing.

Has Electrical Growth Cone Guidance Found Its Potential?

Trends in Neurosciences. Jul, 2002  |  Pubmed ID: 12079763

Many neurobiologists spurn the existence and use of direct-current (DC) electric fields (EFs) in nervous system development and regeneration. This is despite direct measurement of EFs in embryos and adults, and evidence that EFs are required for normal development, dramatically influence the rate and direction of nerve growth in vitro, and promote nerve regeneration in vivo. The notion that growth cones use EFs as guidance cues was dismissed partly because there was no convincing evidence that naturally occurring EFs influence nerve growth at the single-cell level in vivo. Recent work indicates that growth cones can be guided by EFs in vivo and, intriguingly, that in vitro guidance by chemotropic gradients and EFs might invoke similar mechanisms. Ongoing clinical trials to assess the effectiveness of DC EFs in promoting the regeneration of human spinal cord could allow EFs to achieve their potential.

Direct Visualization of a Stratified Epithelium Reveals That Wounds Heal by Unified Sliding of Cell Sheets

FASEB Journal : Official Publication of the Federation of American Societies for Experimental Biology. Mar, 2003  |  Pubmed ID: 12631579

Observing cells in their original niche is a key link between the information gleaned from planar culture and in vivo physiology and pathology. A new approach combining the transparency of the cornea, Hoffman modulation optics, and digital imaging allowed movements of individual corneal cells to be viewed directly in situ. 3-Dimensional time-lapse movies imaging unstained cells within the stratified corneal epithelium during wound healing were made. Tracking cell movements dynamically provided a definitive answer to the long-standing question: does a stratified epithelium heal by "sliding" of cell sheets as a coherent unit or do individual cells "leap frog" each other at the wound margin? A wound in the corneal epithelium healed primarily by sliding of the whole epithelium, with approximately 95% of cells moving with similar speed and trajectories and with little change in their relative position. Only 5% of cells changed layers, with equal proportions moving up or down. Epithelial healing in situ occurred in three phases: a latency, migration, and reconstruction phase. This model provides a unique system to study the behaviors of individual cells in their original niche. It shows that cells slide into a wound as a unified unit to heal a stratified epithelium.

Nerve Regeneration and Wound Healing Are Stimulated and Directed by an Endogenous Electrical Field in Vivo

Journal of Cell Science. Sep, 2004  |  Pubmed ID: 15371524

Biological roles for naturally occurring, extracellular physiological electric fields have been proposed over the past century. However, in the molecular era, many biologists presume that electric fields have little physiological relevance because there has been no unequivocal demonstration of their importance at the single-cell level in vivo. We have used an in vivo rat corneal model, which generates its own endogenous electric field and show that nerve sprouting, the direction of nerve growth and the rate of epithelial wound healing are controlled coordinately by the wound-induced electric field.

Wound Healing in Rat Cornea: the Role of Electric Currents

FASEB Journal : Official Publication of the Federation of American Societies for Experimental Biology. Mar, 2005  |  Pubmed ID: 15746181

Human corneal epithelial cells respond rapidly following injury to restore the integrity of the ocular surface. What stimulates and guides cells to move into the wound to heal? One candidate is the wound-induced electric field. Using vibrating probe techniques, we provide detailed temporal and spatial mapping of endogenous electric currents at rat corneal wounds. We find Cl- and Na+ are the major components of electric currents in rat corneal wounds. Na+ is the major component of ionic transport in the resting (nonwounded) rat cornea and of the wound center leakage current, whereas Cl- is a more important component of the endogenous electrical current at the wound edges. Enhancing or decreasing Cl- flow with clinically approved pharmacological agents such as aminophylline, ascorbic acid, or furosemide increased or decreased endogenous wound electric currents, respectively. These changes in wound currents correlated directly with the rate of wound healing in vivo. Thus, pharmacologically enhancing or decreasing wound-induced electric currents increased and decreased wound healing rate, respectively. This may have wide-reaching and novel therapeutic potential in the management of wound healing and may help explain some mechanistic aspects of the effects of some clinically used agents.

Controlling Cell Behavior Electrically: Current Views and Future Potential

Physiological Reviews. Jul, 2005  |  Pubmed ID: 15987799

Direct-current (DC) electric fields are present in all developing and regenerating animal tissues, yet their existence and potential impact on tissue repair and development are largely ignored. This is primarily due to ignorance of the phenomenon by most researchers, some technically poor early studies of the effects of applied fields on cells, and widespread misunderstanding of the fundamental concepts that underlie bioelectricity. This review aims to resolve these issues by describing: 1) the historical context of bioelectricity, 2) the fundamental principles of physics and physiology responsible for DC electric fields within cells and tissues, 3) the cellular mechanisms for the effects of small electric fields on cell behavior, and 4) the clinical potential for electric field treatment of damaged tissues such as epithelia and the nervous system.

Expression of the Human PAC1 Receptor Leads to Dose-dependent Hydrocephalus-related Abnormalities in Mice

The Journal of Clinical Investigation. Jul, 2006  |  Pubmed ID: 16823490

Hydrocephalus is a common and potentially devastating birth defect affecting the CNS, and its relationship with G protein-coupled receptors (GPCRs) is unknown. We have expressed 2, 4, or 6 copies of a GPCR--the human PAC1 receptor with a 130-kb transgene in the mouse nervous system in a pattern closely resembling that of the endogenous gene. Consistent with PAC1 actions, PKA and PKC activity were elevated in the brains of Tg mice. Remarkably, Tg mice developed dose-dependent hydrocephalus-like characteristics, including enlarged third and lateral ventricles and reduced cerebral cortex, corpus callosum, and subcommissural organ (SCO). Neuronal proliferation and apoptosis were implicated in hydrocephalus, and we observed significantly reduced neuronal proliferation and massively increased neuronal apoptosis in the developing cortex and SCO of Tg embryos, while neurite outgrowth and neuronal migration in vitro remain uncompromised. Ventricular ependymal cilia are crucial for directing cerebrospinal fluid flow, and ependyma of Tg mice exhibited disrupted cilia with increased phospho-CREB immunoreactivity. These data demonstrate that altered neuronal proliferation/apoptosis and disrupted ependymal cilia are the main factors contributing to hydrocephalus in PAC1-overexpressing mice. This is the first report to our knowledge demonstrating that misregulation of GPCRs can be involved in hydrocephalus-related neurodevelopmental disorders.

Electrical Signals Control Wound Healing Through Phosphatidylinositol-3-OH Kinase-gamma and PTEN

Nature. Jul, 2006  |  Pubmed ID: 16871217

Wound healing is essential for maintaining the integrity of multicellular organisms. In every species studied, disruption of an epithelial layer instantaneously generates endogenous electric fields, which have been proposed to be important in wound healing. The identity of signalling pathways that guide both cell migration to electric cues and electric-field-induced wound healing have not been elucidated at a genetic level. Here we show that electric fields, of a strength equal to those detected endogenously, direct cell migration during wound healing as a prime directional cue. Manipulation of endogenous wound electric fields affects wound healing in vivo. Electric stimulation triggers activation of Src and inositol-phospholipid signalling, which polarizes in the direction of cell migration. Notably, genetic disruption of phosphatidylinositol-3-OH kinase-gamma (PI(3)Kgamma) decreases electric-field-induced signalling and abolishes directed movements of healing epithelium in response to electric signals. Deletion of the tumour suppressor phosphatase and tensin homolog (PTEN) enhances signalling and electrotactic responses. These data identify genes essential for electrical-signal-induced wound healing and show that PI(3)Kgamma and PTEN control electrotaxis.

Beta-adrenergic Receptor Agonists Delay While Antagonists Accelerate Epithelial Wound Healing: Evidence of an Endogenous Adrenergic Network Within the Corneal Epithelium

Journal of Cellular Physiology. Apr, 2007  |  Pubmed ID: 17226783

Wound healing is a complex and well-orchestrated biological process. Corneal epithelial cells (CECs) must respond quickly to trauma to rapidly restore barrier function and protect the eye from noxious agents. They express a high level of beta2-adrenergic receptors but their function is unknown. Here, we report the novel finding that they form part of a regulatory network in the corneal epithelium, capable of modulating corneal epithelial wound repair. Beta-adrenergic receptor agonists delay CEC migration via a protein phosphatase 2A-mediated mechanism and decrease both electric field-directed migration and corneal wound healing. Conversely, beta-adrenergic receptor antagonists accelerate CEC migration, enhance electric field-mediated directional migration, and promote corneal wound repair. We demonstrate that CECs express key enzymes required for epinephrine (beta-adrenergic receptor agonist) synthesis in the cytoplasm and can detect epinephrine in cell extracts. We propose that the mechanism for the pro-motogenic effect of the beta-adrenergic antagonist is blockade of the beta2-adrenergic receptor preventing autocrine catecholamine binding. Further investigation of this network will improve our understanding of one of the most frequently prescribed class of drugs.

In Vitro Degradation and Release Profiles for Poly-dl-lactide Film Containing Paracetamol

Journal of Materials Science. Materials in Medicine. Aug, 2007  |  Pubmed ID: 17483888

The biodegradable film containing paracetamol with suitable dimensions was prepared by polymer solution-cast method. The study mainly investigated the in vitro degradation of the films without and with 7.0% and 14% paracetamol in the phosphate-buffer saline (PBS) of pH 7.4 at 37 degrees C. The results showed that the degradation rate for the film with same dimensions containing more drug was faster. The in vitro drug release profiles showed that paracetamol release from film matrix was almost sustained within one month. It suggested that the biodegradable film should be potential in preventing tissue adhesion and local inflammation on operating procedure.

Application of Direct Current Electric Fields to Cells and Tissues in Vitro and Modulation of Wound Electric Field in Vivo

Nature Protocols. 2007  |  Pubmed ID: 17545984

It has long been known that cells can be induced to migrate by the application of small d.c. electric fields (EFs), a phenomenon referred to as galvanotaxis. We recently reported some significant effects of electric signals of physiological strength in guiding cell migration and wound healing. We present here protocols to apply an EF to cells or tissues cultured in an electrotactic chamber. The chamber can be built to allow controlled medium flow to prevent the potential development of chemical gradients generated by the EFs. It can accommodate cells on planar culture or tissues in 3D gels. Mounted on an inverted microscope, this setup allows close and well-controlled observation of cellular responses to electric signals. As similar EFs are widely present during development and wound healing, this experimental system can be used to simulate and study cellular and molecular responses to electric signals in these events.

Visualization of Fast-moving Cells in Vivo Using Digital Holographic Video Microscopy

Journal of Biomedical Optics. Jan-Feb, 2008  |  Pubmed ID: 18315365

Digital in-line holography offers some significant advantages over conventional optical holography and microscopy to image biological specimens. By combining holography with digital video microscopy, an in-line holographic video microscope is developed and is capable of recording spatial 3D holographic images of biological specimens, while preserving the time dimension. The system enables high-speed video recording of fast cell movement, such as the rapid movement of blood cells in the blood stream in vivo. This capability is demonstrated with observations of fast 3-D movement of live cells in suspension cultures in response to a gentle shake to the Petri dish. The experimental and numerical procedures are incorporated with a fast reconstruction algorithm for reconstruction of holographic video frames at various planes (z axis) from the hologram and along the time axis. The current system enables both lateral and longitudinal resolutions down to a few micrometers. Postreconstruction processing of background subtraction is utilized to eliminate noise caused by scattered light, thereby enabling visualization of, for example, blood streams of live Xenopos tadpoles. The combination of digital holography and microscopy offers unique advantages for imaging of fast moving cells and other biological particles in three dimensions in vivo with high spatial and temporal resolution.

[Effect of Shuanghuanglian Injection on the Activation and Proliferation of T Lymphocytes in Mice in Vitro]

Xi Bao Yu Fen Zi Mian Yi Xue Za Zhi = Chinese Journal of Cellular and Molecular Immunology. Aug, 2008  |  Pubmed ID: 18687215

To study the effect of shuanghuanglian injection on the activation and proliferation of T lymphocytes in mice in vitro.

[Effects of Forskolin on Proliferation Cell-cycle Distribution and Activation of the Murine T Lymphocytes in Vitro]

Zhong Yao Cai = Zhongyaocai = Journal of Chinese Medicinal Materials. Jul, 2008  |  Pubmed ID: 18973017

To investigate the effects of Forskolin on activation, proliferation, and cell-cycle distribution of murine CD3+ T lymphocytes, and study the mechanisms of its immunosuppressive effect.

[Effect of Icariin on Intermediate and Advanced Activation of Murine T Lymphocytes in Vitro]

Xi Bao Yu Fen Zi Mian Yi Xue Za Zhi = Chinese Journal of Cellular and Molecular Immunology. Nov, 2008  |  Pubmed ID: 18992191

To study the effect of icariin (ICA) on the intermediate and advanced activation of murine T lymphocytes stimulated by concanvalin A (ConA) in vitro.

Impact of Alanyl-glutamine Dipeptide on Severe Acute Pancreatitis in Early Stage

World Journal of Gastroenterology : WJG. Jan, 2008  |  Pubmed ID: 18200673

To evaluate the therapeutic effect of alanyl-glutamine dipeptide (AGD) in the treatment of severe acute pancreatitis (SAP) in early and advanced stage.

Effect of Antibiotic Prophylaxis on Acute Necrotizing Pancreatitis: Results of a Randomized Controlled Trial

Journal of Gastroenterology and Hepatology. May, 2009  |  Pubmed ID: 19220676

This study addresses whether antibiotic prophylaxis is beneficial for acute necrotizing pancreatitis.

[Effects of Hesperetin on Cell Activation and Proliferation of Murine T Lymphocytes in Vitro]

Xi Bao Yu Fen Zi Mian Yi Xue Za Zhi = Chinese Journal of Cellular and Molecular Immunology. Apr, 2009  |  Pubmed ID: 19351499

The effect of Hesperetin (Hes) on activation and proliferation of murine T lymphocytes in vitro as well as its mechanism of action is investigated to provide a theory for developing an immunosuppressive agent.

Infectious Complications in Patients with Severe Acute Pancreatitis

Digestive Diseases and Sciences. Dec, 2009  |  Pubmed ID: 19104931

This study aims to investigate the clinical characteristics of infectious complications in severe acute pancreatitis (SAP). From September 2003 to March 2005, 140 patients with SAP were retrospectively identified. SAP was defined by the diagnostic criteria formulated for SAP at the 2002 Bangkok World Congress of Gastroenterology in Thailand. Clinical data of the infected and non-infected patients was compared and the characteristics of infection were also analyzed. There were 44 patients who developed infectious complications with a rate of 31.4% (44/140). The severity index, the incidence of complications and mortality, was significantly higher in the infected patients than in the non-infected patients (P<0.05). Of 65 episodes of infection, infected (peri) pancreatic necrosis accounted for 47.7% (31/65), pneumonia for 27.7% (18/65), bacteremia for 10.8% (7/65), urinary tract infection for 6.1% (4/65), and gastrointestinal tract infection for 7.7% (5/65). The earliest infection was observed in pneumonia (10.7±2.5 days), followed by bacteremia (13.7±1.5 days), gastrointestinal tract infection (16.8±3.9 days), infected (peri)pancreatic necrosis (17.6±2.9 days), and urinary tract infection (20.5±4.8 days). Gram-negative bacteria were preponderantly found, comprising 56.6% (64/113) of the isolated strains. Gram-positive bacteria and fungus accounted for 22.1% (25/113) and 21.2% (24/113) of the isolated strains, respectively. Infectious complications in patients with SAP occurred in those who had severe episodes, and consequently complicated the clinical courses. Infected (peri)pancreatic necrosis is the most susceptible and pneumonia is the earliest. Gram-negative bacteria were predominant in multi-microorganisms.

SEPA-1 Mediates the Specific Recognition and Degradation of P Granule Components by Autophagy in C. Elegans

Cell. Jan, 2009  |  Pubmed ID: 19167332

How autophagy, an evolutionarily conserved intracellular catabolic system for bulk degradation, selectively degrades protein aggregates is poorly understood. Here, we show that several maternally derived germ P granule components are selectively eliminated by autophagy in somatic cells during C. elegans embryogenesis. The activity of sepa-1 is required for the degradation of these P granule components and for their accumulation into aggregates, termed PGL granules, in autophagy mutants. SEPA-1 forms protein aggregates and is also a preferential target of autophagy. SEPA-1 directly binds to the P granule component PGL-3 and also to the autophagy protein LGG-1/Atg8. SEPA-1 aggregates consistently colocalize with PGL granules and with LGG-1 puncta. Thus, SEPA-1 functions as a bridging molecule in mediating the specific recognition and degradation of P granule components by autophagy. Our study reveals a mechanism for preferential degradation of protein aggregates by autophagy and emphasizes the physiological significance of selective autophagy during animal development.

[Effects of Kaempferol on Activation, Proliferation and Cell Cycle of Mouse T Lymphocytes in Vitro.]

Xi Bao Yu Fen Zi Mian Yi Xue Za Zhi = Chinese Journal of Cellular and Molecular Immunology. Dec, 2009  |  Pubmed ID: 19961795

To study the effects of Kaempferol on activation, proliferation and cell cycle of murine T lymphocytes, and to elucidate the mechanism of the immunosuppressive effect of Kaempferol.

[Value of Nitrite in Exhaled Breath Condensate in Estimating the Severity of Asthma]

Nan Fang Yi Ke Da Xue Xue Bao = Journal of Southern Medical University. Feb, 2009  |  Pubmed ID: 19246312

To explore the relationship between nitrite levels in the exhaled breath condensates (EBC) and the severity of asthma.

Electric Currents in Xenopus Tadpole Tail Regeneration

Developmental Biology. Nov, 2009  |  Pubmed ID: 19733557

Xenopus laevis tadpoles can regenerate tail, including spinal cord, after partial amputation, but lose this ability during a specific period around stage 45. They regain this ability after stage 45. What happens during this "refractory period" might hold the key to spinal cord regeneration. We hypothesize that electric currents at amputated stumps play significant roles in tail regeneration. We measured electric current at tail stumps following amputation at different developmental stages. Amputation induced large outward currents leaving the stump. In regenerating stumps of stage 40 tadpoles, a remarkable reversal of the current direction occurred around 12-24 h post-amputation, while non-regenerating stumps of stage 45 tadpole maintained outward currents. This reversal of electric current at tail stumps correlates with whether tails regenerate or not (regenerating stage 40-inward current; non-regenerating stage 45-outward current). Reduction of tail stump current using sodium-free solution decreased the rate of regeneration and percentage regeneration. Fin punch wounds healed normally at stages 45 and 48, and in sodium-free solution, suggesting that the absence of tail re-growth at stage 45 is regeneration-specific rather than a general inhibition of wound healing. These data suggest that electric signals might be one of the key players regulating regeneration.

An Alternative Technique for Tricuspid Valve Repair Using Autologous Pericardium

Journal of Cardiac Surgery. Sep-Oct, 2009  |  Pubmed ID: 19740287

We developed an alternative technique to integrate the replacement of tricuspid septal leaflet and chordae using a fan-shaped fresh autologous pericardial patch. Echocardiographic evaluation showed normal mobility, and no progression of tricuspid regurgitation or evidence of calcification early and late after operation. This alternative approach appears to be an easy, rapid, and effective technique for tricuspid valve repair.

Electrical Dimensions in Cell Science

Journal of Cell Science. Dec, 2009  |  Pubmed ID: 19923270

Cells undergo a variety of physiological processes, including division, migration and differentiation, under the influence of endogenous electrical cues, which are generated physiologically and pathologically in the extracellular and sometimes intracellular spaces. These signals are transduced to regulate cell behaviours profoundly, both in vitro and in vivo. Bioelectricity influences cellular processes as fundamental as control of the cell cycle, cell proliferation, cancer-cell migration, electrical signalling in the adult brain, embryonic neuronal cell migration, axon outgrowth, spinal-cord repair, epithelial wound repair, tissue regeneration and establishment of left-right body asymmetry. In addition to direct effects on cells, electrical gradients interact with coexisting extracellular chemical gradients. Indeed, cells can integrate and respond to electrical and chemical cues in combination. This Commentary details how electrical signals control multiple cell behaviours and argues that study of the interplay between combined electrical and chemical gradients is underdeveloped yet necessary.

Electric Currents and Lens Regeneration in the Rat

Experimental Eye Research. Feb, 2010  |  Pubmed ID: 19931246

We studied the process of lens regeneration in the rat following an extracapsular lens extraction preserving the anterior lens capsule and anterior lens epithelium. We assessed clinically the clarity of the newly regenerated lens, evaluated changes in the lens electrical currents following surgery and during the regeneration process and correlated these changes with findings on light microscopy. Protein analysis of the regenerated lens was also undertaken. Experiments were performed in 41 Sprague-Dawley rats, sacrificed at 0, 2, 4 and 8 weeks postoperatively. Our results showed that complete lens regeneration occurred 8 weeks postoperatively only if the anterior epithelium was preserved and the lens capsule was closed surgically. Lens electrical currents, altered following surgery, recovered in parallel with the process of regeneration of the lens. The newly regenerated lens was optically clear and biochemical analysis revealed a pattern of protein expression resembling that observed during lens development. In conclusion, complete lens regeneration occurs in the rat and it is possible that lens electrical signals, together with other cues, may play an important role in this process.

Effects of Physiological Electric Fields on Migration of Human Dermal Fibroblasts

The Journal of Investigative Dermatology. Sep, 2010  |  Pubmed ID: 20410911

Endogenous electric currents generated instantly at skin wounds direct migration of epithelial cells and are likely to be important in wound healing. Migration of fibroblasts is critical in wound healing. It remains unclear how wound electric fields guide migration of dermal fibroblasts. We report here that mouse skin wounds generated endogenous electric currents for many hours. Human dermal fibroblasts of both primary and cell-line cultures migrated directionally but slowly toward the anode in an electric field of 50-100 mV mm(-1). This is different from keratinocytes, which migrate quickly to the cathode. It took more than 1 hour for dermal fibroblasts to manifest detectable directional migration. Larger field strength (400 mV mm(-1)) was required to induce directional migration within 1 hour after onset of the field. Phosphatidylinositol-3-OH kinase (PI3 kinase) mediates cathode-directed migration of keratinocytes. We tested the role of PI3 kinase in anode-directed migration of fibroblasts. An applied electric field activated PI3 kinase/Akt in dermal fibroblasts. Dermal fibroblasts from p110gamma (a PI3 kinase catalytic subunit) null mice showed significantly decreased directional migration. These results suggest that physiological electric fields may regulate motility of dermal fibroblasts and keratinocytes differently, albeit using similar PI3 kinase-dependent mechanisms.

Polo-like Kinase 1 Phosphorylation of G2 and S-phase-expressed 1 Protein is Essential for P53 Inactivation During G2 Checkpoint Recovery

EMBO Reports. Aug, 2010  |  Pubmed ID: 20577264

In response to G2 DNA damage, the p53 pathway is activated to lead to cell-cycle arrest, but how p53 is eliminated during the subsequent recovery process is poorly understood. It has been established that Polo-like kinase 1 (Plk1) controls G2 DNA-damage recovery. However, whether Plk1 activity contributes to p53 inactivation during this process is unknown. In this study, we show that G2 and S-phase-expressed 1 (GTSE1) protein, a negative regulator of p53, is required for G2 checkpoint recovery and that Plk1 phosphorylation of GTSE1 at Ser 435 promotes its nuclear localization, and thus shuttles p53 out of the nucleus to lead to its degradation during the recovery.

[Clinical Effects of Modified Ultrafiltration During Pediatric Cardiac Surgery: a Systematic Review]

Nan Fang Yi Ke Da Xue Xue Bao = Journal of Southern Medical University. Jul, 2010  |  Pubmed ID: 20650795

To assess the clinical effects and safety of modified ultrafiltration during pediatric cardiac surgery.

Polo-like Kinase 1 Phosphorylation of P150Glued Facilitates Nuclear Envelope Breakdown During Prophase

Proceedings of the National Academy of Sciences of the United States of America. Aug, 2010  |  Pubmed ID: 20679239

Nuclear envelope breakdown (NEBD) is an essential step during the G2/M transition in higher eukaryotic cells. Increasing evidence supports the notion that both microtubules and microtubule-associated motor proteins are critical regulators of NEBD. Although it has been described that p150(Glued), the major component of the dynein/dynactin complex, localizes in the nuclear envelope during prophase, the exact role of p150(Glued) and its regulation during NEBD are largely elusive. Polo-like kinase 1 (Plk1), the best characterized Ser/Thr kinase, is involved in mitotic entry in several systems; however, the targets of Plk1 during NEBD are unknown. Herein, we show that in mammalian cells both Plk1 and p150(Glued) regulate NEBD and that Plk1 interacts with and phosphoryates p150(Glued) during NEBD at prophase. Using various approaches, we showed that Plk1 phosphorylates p150(Glued) at Ser-179 and that the pS179 epitope is generated at the nuclear envelope of prophase cells. Significantly, Plk1-mediated phosphorylation of p150(Glued) at Ser-179 positively regulates its accumulation at the nuclear envelope during prophase. Finally, we found that cells expressing the Plk1-unphosphorylatable mutant (p150(Glued)-S179A) arrest at G2, as indicated by reduced NEBD, increased levels of cyclin B and phospho-H3, but a decreased level of Cdc2 kinase activity. Taking these data together, we conclude that Plk1 phosphorylation of p150(Glued) might be one major pathway of NEBD regulation.

A Time-lapse and Quantitative Modelling Analysis of Neural Stem Cell Motion in the Absence of Directional Cues and in Electric Fields

Journal of Neuroscience Research. Nov, 2010  |  Pubmed ID: 20890991

Neural stem cell (NSC) migration is an important component of their developmental function and therapeutic potential. Understanding their mode of migration and their response to guidance cues can contribute to improved therapies for CNS repair, in which appropriate homing to sites of injury is essential. Using time-lapse imaging, we have analyzed the NSC mode of migration in vitro, both in the absence of directional cues and in the presence of applied electric fields (EFs), previously shown to constitute a strong directional signal for these cells. Without EFs, NSCs displayed an amoeboid motion, characterized by small lamellipodial-like protrusions with changing orientations, leading to highly tortuous migration. In EFs, tortuosity diminished as electrotaxis toward the cathode occurred. EFs suppressed the formation of protrusions oriented toward the anode, suggesting that restriction of protrusions with opposing orientation could underlie the change from tortuous motion to directed migration. Treatment with LY294002, a phosphatidylinositol-3-OH kinase (Pi3K) inhibitor, reduced the cathodal bias of protrusions in EFs and the frequency of changes in direction. We generated a model of NSC migration with only two key parameters, which could accurately reproduce experimental migration patterns, and we used it to show that both effects of LY294002 contribute to impair electrotaxis, although decreased protrusion bias is the most important. Our results show that control of protrusion orientation by EFs is an important component of the electrotactic response. A simple modelling approach might be useful in understanding how diverse pharmacological treatments or genetic deletions affect different kinds of directional cell migration.

[Effect of Dexamethasone on Atrial Fibrillation After Cardiac Surgery: a Meta-analysis]

Zhonghua Wai Ke Za Zhi [Chinese Journal of Surgery]. Aug, 2010  |  Pubmed ID: 21055014

To assess the effect of preoperative dexamethasone on the occurrence of postoperative atrial fibrillation.

The Substrates of Plk1, Beyond the Functions in Mitosis

Protein & Cell. Nov, 2010  |  Pubmed ID: 21153517

Polo-like kinase 1 (Plk1) is a key regulator of cell division in eukaryotic cells. In this short review, we briefly summarized the well-established functions modulated by Plk1 during mitosis. Beyond mitosis, we focused mainly on the unexpected processes in which Plk1 emerges as a critical player, including microtubule dynamics, DNA replication, chromosome dynamics, p53 regulation, and recovery from the G2 DNA-damage checkpoint. Our discussion is mainly based on the critical substrates targeted by Plk1 during these cellular events and the functional significance associated with each phosphorylation event.

[Isoflurance-based Intravenous and Inhalation Combined Anesthesia Versus Low-dose-ketamine-based Total Intravenous Anesthesia for Valvuloplasty in Minipigs]

Sichuan Da Xue Xue Bao. Yi Xue Ban = Journal of Sichuan University. Medical Science Edition. Sep, 2010  |  Pubmed ID: 21302458

To compare the effectiveness of anesthesia between Isoflurane-based intravenous and inhalant combined approach and low-dose-ketamine-based total intravenous approach for valvuloplasty in minipigs.

[The Effect of Ginsenoside Rb1 on Phagocytosis, the Production of Cytokines and NO of Murine Macrophage in Vitro]

Xi Bao Yu Fen Zi Mian Yi Xue Za Zhi = Chinese Journal of Cellular and Molecular Immunology. Mar, 2011  |  Pubmed ID: 21419038

Investigate the effects of Rb1 on phagocytosis and the production of the pro-inflammatory cytokines and NO of murine macrophage, to elucidate the machanism of the immuno-regulating effect of Rb1.

Liposome Impaired the Adhesion and Spreading of HEK293 Cells: an AFM Study

Scanning. Nov-Dec, 2011  |  Pubmed ID: 21773977

Gene transfer has been proven to be a promising approach for treatment of several diseases. The cytotoxicity of transfection reagents is one of the key factors for clinical applications. The cytotoxicity of liposome has been extensively studied. However, its effects on the adhesion and spreading of transformed cells are still unclear. In this study, the cytotoxic effects of liposome on cell viability and mitochondrial membrane potential of HEK293 cells were first evaluated. Then, an atomic force microscope (AFM) was recruited to investigate the effects of liposome on the adhesion and spreading of HEK293 cells. AFM data indicated that liposome induced a significant decrease in number of cellular pseudopodia and cell-surface particles, in cell-surface roughness, and in average adhesion force of cell membranes. The AFM data implied that liposome impaired the adhesion and spreading of HEK293 cells.

Polo-like Kinase 1 Facilitates Loss of Pten Tumor Suppressor-induced Prostate Cancer Formation

The Journal of Biological Chemistry. Oct, 2011  |  Pubmed ID: 21890624

Loss of the tumor suppressor Pten (phosphatase and tensin homolog deleted on chromosome 10) is thought to mediate the majority of prostate cancers, but the molecular mechanism remains elusive. In this study, we demonstrate that Pten-depleted cells suffer from mitotic stress and that nuclear function of Pten, but not its phosphatase activity, is required to reverse this stress phenotype. Further, depletion of Pten results in elevated expression of Polo-like kinase 1 (Plk1), a critical regulator of the cell cycle. We show that overexpression of Plk1 correlates with genetic inactivation of Pten during prostate neoplasia formation. Significantly, we find that elevated Plk1 is critical for Pten-depleted cells to adapt to mitotic stress for survival and that reintroduction of wild-type Pten into Pten-null prostate cancer cells reduces the survival dependence on Plk1. We further show that Plk1 confers the tumorigenic competence of Pten-deleted prostate cancer cells in a mouse xenograft model. These findings identify a role of Plk1 in facilitating loss of Pten-induced prostate cancer formation, which suggests that Plk1 might be a promising target for prostate cancer patients with inactivating Pten mutations.

Inhibition of Polo-like Kinase 1 Reduces Beta-amyloid-induced Neuronal Cell Death in Alzheimer's Disease

Aging. Sep, 2011  |  Pubmed ID: 21931181

Alzheimer's disease (AD) is a progressive and fatal brain disease, but the pathogenesis of AD is still not understood. Aberrant cell-cycle re-entry of neuronal cells is emerging as a potential pathological mechanism in AD. Polo-like kinase 1 (Plk1) is an established regulator of many cell cycle-related events. Interestingly, Plk1 is present in susceptible hippocampal and cortical neurons of AD patients but not age-matched controls. However, whether Plk1 is involved in the pathogenesis of AD remains elusive. In this study, we showed that Plk1 activity is elevated in AD patient brain as indicated by the increased phosphorylation signal of p150Glued, a Plk1-specific substrate. Furthermore, we demonstrated that Plk1 is elevated during the cell-cycle re-entry of neuronal cells in an in vitro cell-culture model. Significantly, inhibition of Plk1 kinase activity or depletion of Plk1 by RNAi reduces β-amyloid (Aβ)-induced neuronal cell death. These results validate Plk1 as a possible target for AD therapy.

Plk1 Phosphorylation of Orc2 Promotes DNA Replication Under Conditions of Stress

Molecular and Cellular Biology. Dec, 2011  |  Pubmed ID: 21947279

Polo-like kinase 1 (Plk1) plays pivotal roles in mitosis; however, little is known about its function in S phase. In this study, we show that inhibition of Plk1 impairs DNA replication and results in slow S-phase progression in cultured cancer cells. We have identified origin recognition complex 2 (Orc2), a member of the DNA replication machinery, as a Plk1 substrate and have shown that Plk1 phosphorylates Orc2 at Ser188 in vitro and in vivo. Furthermore, Orc2-S188 phosphorylation is enhanced when DNA replication is under challenge induced by ultraviolet, hydroxyurea, gemcitabine, or aphidicolin treatment. Cells expressing the unphosphorylatable mutant (S188A) of Orc2 had defects in DNA synthesis under stress, suggesting that this phosphorylation event is critical to maintain DNA replication under stress. To dissect the mechanism pertinent to this observation, we showed that Orc2-S188 phosphorylation associates with DNA replication origin and that cells expressing Orc2-S188A mutant fail to maintain the functional pre-replicative complex (pre-RC) under DNA replication stress. Furthermore, the intra-S-phase checkpoint is activated in Orc2-S188A-expressing cells to cause delay of S-phase progress. Our study suggests a novel role of Plk1 in facilitating DNA replication under conditions of stress to maintain genomic integrity.

Synthesis and Development of Poly(N-hydroxyethyl Acrylamide)-ran-3-acrylamidophenylboronic Acid Polymer Fluid for Potential Application in Affinity Sensing of Glucose

Journal of Diabetes Science and Technology. Sep, 2011  |  Pubmed ID: 22027298

In previous work, we described viscosity and permittivity microelectromechanical systems (MEMS) sensors for continuous glucose monitoring (CGM) using poly[acrylamide-ran-3-acrylamidophenylboronic acid (PAA-ran-PAAPBA). In order to enhance our MEMS device antifouling properties, a novel, more hydrophilic polymer-sensing fluid was developed.

[Sperm Chromatin Integrity Test for Predicting the Outcomes of IVF and ICSI]

Zhonghua Nan Ke Xue = National Journal of Andrology. Dec, 2011  |  Pubmed ID: 22235675

To explore the predictive value of sperm chromatin integrity test (SCIT) in assisted reproductive technology (ART) by analyzing the relationship of sperm chromatin integrity (SCI) with the outcomes of IVF-ET and ICSI.

Magnetic CoPt Nanoparticles As MRI Contrast Agent for Transplanted Neural Stem Cells Detection

Nanoscale. Mar, 2011  |  Pubmed ID: 21293831

Neural stem cells (NSCs) exhibit features that make them suitable candidates for stem cell replacement therapy and spinal cord reconstruction. Magnetic resonance imaging (MRI) offers the potential to track cells in vivo using innovative approaches to cell labeling and image acquisition. In this study, experiments were carried out to optimize the loading condition of magnetic CoPt hollow nanoparticles (CoPt NPs) into neural stem cells and to define appropriate MRI parameters. Both cell viability and multipotency analysis showed that CoPt NPs at a concentration of 16 µg ml(-1) reduced T2 relaxation times in labeled rat NSCs, producing greater contrast on spin echo acquisitions at 4.7 T, yet did not affect cell viability and in vitro differentiation potential compared to controls. After optimizing nanoparticle loading concentrations and labeled cell numbers for MRI detection, CoPt-loaded NSCs were transplanted into organotypic spinal cord slices. The results showed that MRI could efficiently detect low numbers of CoPt-labeled NSCs with the enhanced image contrast. Our study demonstrated that MRI of grafted NSCs labeled with CoPt NPs is a useful tool to evaluate organotypic spinal cord slice models and has potential applications in other biological systems.

[Effects of Exercise and Nilestriol on Bone Size and Bone Mass in Ovariectomized Rats]

Sichuan Da Xue Xue Bao. Yi Xue Ban = Journal of Sichuan University. Medical Science Edition. Jul, 2011  |  Pubmed ID: 21866639

To investigate the influence of nilestriol (CCE3) and exercise on bone size and bone mass in ovariectomized (OVX) rats.

PI3K Mediated Electrotaxis of Embryonic and Adult Neural Progenitor Cells in the Presence of Growth Factors

Experimental Neurology. Jan, 2011  |  Pubmed ID: 21092738

Correct guidance of the migration of neural progenitor cells (NPCs) is essential for the development and repair of the central nervous system (CNS). Electric field (EF)-guided migration, electrotaxis, has been observed in many cell types. We report here that, in applied EFs of physiological magnitude, embryonic and adult NPCs show marked electrotaxis, which is dependent on the PI3K/Akt pathway. The electrotaxis was also evidenced by ex vivo investigation that transplanted NPCs migrated directionally towards cathode in organotypic spinal cord slice model when treated with EFs. Genetic disruption or pharmacological inhibition of phosphoinositide 3-kinase (PI3K) impaired electrotaxis, whereas EF exposure increased Akt phosphorylation in a growth factor-dependent manner and increased phosphatidylinositol-3,4,5-trisphosphate (PIP3) levels. EF treatments also induced asymmetric redistribution of PIP3, growth factor receptors, and actin cytoskeleton. Electrotaxis in both embryonic and adult NPCs requires epidermal growth factor (EGF) and fibroblast growth factor (FGF). Our results demonstrate the importance of the PI3K/Akt pathway in directed migration of NPCs driven by EFs and growth factors and highlight the potential of EFs to enhance the guidance of various NPC populations in CNS repair therapies.

Proteomic Analysis of Secretion from Human Transplanted Submandibular Gland Replacing Lacrimal Gland with Severe Keratoconjunctivitis Sicca

Biochimica Et Biophysica Acta. Jan, 2012  |  Pubmed ID: 22306964

Purpose: Proteomic analysis of secretions from transplanted or non-transplanted submandibular glands in patients with severe keratoconjunctivitis sicca and tears from normal eyes. Experimental design: Secretions from submandibular glands transplanted to replace lacrimal glands and non-transplanted submandibular glands were collected at 1year from 5 patients with severe keratoconjunctivitis sicca undergoing transplantation, and tears were collected from 3 normal subjects. 2-D electrophoresis (2-DE), then mass spectrometry was used to identify proteins. Western blot analysis was used to confirm protein expression. Results: We identified 34 and 11 distinct proteins in the saliva from transplanted submandibular glands and tears, respectively. The saliva from transplanted submandibular glands contained almost all the proteins abundant in tear fluid. The functions of identified proteins in the saliva from transplanted submandibular gland were mainly immune response and anti-bacterial. In total, 7 proteins showed differential expression between the saliva of transplanted and non-transplanted submandibular glands. The upregulation of short palate, lung and nasal epithelium carcinoma-associated protein 2 and carbonic anhydrase VI was confirmed by Western blot analysis. Conclusions: Identified proteins in saliva from transplanted submandibular glands may protect ocular structures. These findings can help in understanding the functional status of transplanted submandibular glands.

Polo-like Kinase 1 (Plk1): an Unexpected Player in DNA Replication

Cell Division. Feb, 2012  |  Pubmed ID: 22309699

ABSTRACT: Regulation of cell cycle progression is important for the maintenance of genome integrity, and Polo-like kinases (Plks) have been identified as key regulators of this process. It is well established that Polo-like kinase 1 (Plk1) plays critical roles in mitosis but little is known about its functions at other stages of the cell cycle. Here we summarize the functions of Plk1 during DNA replication, focusing on the molecular events related to Origin Recognition Complex (ORC), the complex that is essential for the initiation of DNA replication. Within the context of Plk1 phosphorylation of Orc2, we also emphasize regulation of Orc2 in different organisms. This review is intended to provide some insight into how Plk1 coordinates DNA replication in S phase with chromosome segregation in mitosis, and orchestrates the cell cycle as a whole.

Cinobufacini Induced MDA-MB-231 Cell Apoptosis-associated Cell Cycle Arrest and Cytoskeleton Function

Bioorganic & Medicinal Chemistry Letters. Feb, 2012  |  Pubmed ID: 22225634

Cinobufacini is a traditional Chinese anti-tumor drug and widely used in clinic experiences. But little is known about its effect on the cells. In this study, the effects of cinobufacini on breast cancer MDA-MB-231 cell were evaluated by CCK-8 assay, and the data showed cinobufacini could inhibit the MDA-MB-231 cells growth effectively in dose-dependent and time-dependent manners. Cell apoptosis and cell cycle were detected by flow cytometry analysis. After the cells being treated with 50μg/mL cinobufacini for 48h, the early apoptosis percentage (20.45±1.46%) is much higher than the normal group (7.73±1.21%). The cell cycle data indicated that cinobufacini caused a cell cycle arrest at S phase. What's more, cinobufacini can affect the disruption of cytoskeleton, and these alterations changed the cell-surface ultrastructure and the cell morphology which were detected by atomic force microscopy (AFM) at nanoscale level. It indicated that the cell membrane structure and cytoskeleton networks were destroyed and the cell tails were narrowed after the cell being treated with cinobufacini. The present study is to provide valuable new insights to understand the mechanism of the drug in anti-tumor process. Furthermore, the knowledge concerning the signaling of cell cycle is potentially important to clinical utility.

[Effects of Ginkgolide B on Proliferation, Phagocytosis, NO and ROS Production of Murine Peritoneal Macrophages in Vitro]

Xi Bao Yu Fen Zi Mian Yi Xue Za Zhi = Chinese Journal of Cellular and Molecular Immunology. Jan, 2012  |  Pubmed ID: 22230494

To explore the potential immunomodulatory effects and related mechanisms of ginkgolide B (GB), a known potent antagonist of platelet-activating factor receptor, we investigated the proliferation, phagocytosis, NO and ROS production of macrophage.

The Cytotoxicity and Anticancer Mechanisms of Alterporriol L, a Marine Bianthraquinone, Against MCF-7 Human Breast Cancer Cells

Applied Microbiology and Biotechnology. Jan, 2012  |  Pubmed ID: 21779847

Alterporriol L, a new bianthraquinone derivative, was isolated from a marine fungus Alternaria sp. ZJ9-6B. The cytotoxic activity and anticancer mechanisms of alterporriol L towards breast cancer cells lines were detected using MTT assay, immunofluorescence, and flow cytometry. Simultaneously, the changes in morphological properties of cells were detected before and after treatment with alterporriol L by atomic force microscope (AFM) at a nanometer scale. MTT assay showed that alterporriol L could effectively inhibit the growth and proliferation, and there was a dose-dependent manner of cell death. Moreover, the alterporriol L could induce cancer cell apoptosis or necrosis. Furthermore, the reactive oxygen species, mitochondrial membrane potential, and cytosolic free calcium level were changed after treatment with alterporriol L, suggesting that alterporriol L played vital roles in breast cancer cells through destroying the mitochondrial. And all these alterations are in accord with changes of morphology detected by AFM, which suggested that the AFM is a useful tool to detect the morphological changes of the cancer cells.

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