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Sample records for ganglion cell apoptosis

  1. Real-Time Imaging of Retinal Ganglion Cell Apoptosis

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    Timothy E. Yap

    2018-06-01

    Full Text Available Monitoring real-time apoptosis in-vivo is an unmet need of neurodegeneration science, both in clinical and research settings. For patients, earlier diagnosis before the onset of symptoms provides a window of time in which to instigate treatment. For researchers, being able to objectively monitor the rates of underlying degenerative processes at a cellular level provides a biomarker with which to test novel therapeutics. The DARC (Detection of Apoptosing Retinal Cells project has developed a minimally invasive method using fluorescent annexin A5 to detect rates of apoptosis in retinal ganglion cells, the key pathological process in glaucoma. Numerous animal studies have used DARC to show efficacy of novel, pressure-independent treatment strategies in models of glaucoma and other conditions where retinal apoptosis is reported, including Alzheimer’s disease. This may forge exciting new links in the clinical science of treating both cognitive and visual decline. Human trials are now underway, successfully demonstrating the safety and efficacy of the technique to differentiate patients with progressive neurodegeneration from healthy individuals. We review the current perspectives on retinal ganglion cell apoptosis, the way in which this can be imaged, and the exciting advantages that these future methods hold in store.

  2. Agmatine protects retinal ganglion cells from hypoxia-induced apoptosis in transformed rat retinal ganglion cell line

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    Kim Chan

    2007-10-01

    Full Text Available Abstract Background Agmatine is an endogenous polyamine formed by the decarboxylation of L-arginine. We investigated the protective effects of agmatine against hypoxia-induced apoptosis of immortalized rat retinal ganglion cells (RGC-5. RGC-5 cells were cultured in a closed hypoxic chamber (5% O2 with or without agmatine. Cell viability was determined by lactate dehydrogenase (LDH assay and apoptosis was examined by annexin V and caspase-3 assays. Expression and phosphorylation of mitogen-activated protein kinases (MAPKs; JNK, ERK p44/42, and p38 and nuclear factor-kappa B (NF-κB were investigated by Western immunoblot analysis. The effects of agmatine were compared to those of brain-derived neurotrophic factor (BDNF, a well-known protective neurotrophin for retinal ganglion cells. Results After 48 hours of hypoxic culture, the LDH assay showed 52.3% cell loss, which was reduced to 25.6% and 30.1% when agmatine and BDNF were administered, respectively. This observed cell loss was due to apoptotic cell death, as established by annexin V and caspase-3 assays. Although total expression of MAPKs and NF-κB was not influenced by hypoxic injury, phosphorylation of these two proteins was increased. Agmatine reduced phosphorylation of JNK and NF-κB, while BDNF suppressed phosphorylation of ERK and p38. Conclusion Our results show that agmatine has neuroprotective effects against hypoxia-induced retinal ganglion cell damage in RGC-5 cells and that its effects may act through the JNK and NF-κB signaling pathways. Our data suggest that agmatine may lead to a novel therapeutic strategy to reduce retinal ganglion cell injury related to hypoxia.

  3. Agmatine protects retinal ganglion cells from hypoxia-induced apoptosis in transformed rat retinal ganglion cell line

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    Hong, Samin; Lee, Jong Eun; Kim, Chan Yun; Seong, Gong Je

    2007-01-01

    Background Agmatine is an endogenous polyamine formed by the decarboxylation of L-arginine. We investigated the protective effects of agmatine against hypoxia-induced apoptosis of immortalized rat retinal ganglion cells (RGC-5). RGC-5 cells were cultured in a closed hypoxic chamber (5% O2) with or without agmatine. Cell viability was determined by lactate dehydrogenase (LDH) assay and apoptosis was examined by annexin V and caspase-3 assays. Expression and phosphorylation of mitogen-activated protein kinases (MAPKs; JNK, ERK p44/42, and p38) and nuclear factor-kappa B (NF-κB) were investigated by Western immunoblot analysis. The effects of agmatine were compared to those of brain-derived neurotrophic factor (BDNF), a well-known protective neurotrophin for retinal ganglion cells. Results After 48 hours of hypoxic culture, the LDH assay showed 52.3% cell loss, which was reduced to 25.6% and 30.1% when agmatine and BDNF were administered, respectively. This observed cell loss was due to apoptotic cell death, as established by annexin V and caspase-3 assays. Although total expression of MAPKs and NF-κB was not influenced by hypoxic injury, phosphorylation of these two proteins was increased. Agmatine reduced phosphorylation of JNK and NF-κB, while BDNF suppressed phosphorylation of ERK and p38. Conclusion Our results show that agmatine has neuroprotective effects against hypoxia-induced retinal ganglion cell damage in RGC-5 cells and that its effects may act through the JNK and NF-κB signaling pathways. Our data suggest that agmatine may lead to a novel therapeutic strategy to reduce retinal ganglion cell injury related to hypoxia. PMID:17908330

  4. Stanniocalcin-1 protects retinal ganglion cells by inhibiting apoptosis and oxidative damage.

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    Sang Jin Kim

    Full Text Available Optic neuropathy including glaucoma is one of the leading causes of irreversible vision loss, and there are currently no effective therapies. The hallmark of pathophysiology of optic neuropathy is oxidative stress and apoptotic death of retinal ganglion cells (RGCs, a population of neurons in the central nervous system with their soma in the inner retina and axons in the optic nerve. We here tested that an anti-apoptotic protein stanniocalcin-1 (STC-1 can prevent loss of RGCs in the rat retina with optic nerve transection (ONT and in cultures of RGC-5 cells with CoCl2 injury. We found that intravitreal injection of STC-1 increased the number of RGCs in the retina at days 7 and 14 after ONT, and decreased apoptosis and oxidative damage. In cultures, treatment with STC-1 dose-dependently increased cell viability, and decreased apoptosis and levels of reactive oxygen species in RGC-5 cells that were exposed to CoCl2. The expression of HIF-1α that was up-regulated by injury was significantly suppressed in the retina and in RGC-5 cells by STC-1 treatment. The results suggested that intravitreal injection of STC-1 might be a useful therapy for optic nerve diseases in which RGCs undergo apoptosis through oxidative stress.

  5. Study on the mechanism of retinal ganglion cell apoptosis in early stage of diabetic rats

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    Rui-Dong Gu

    2014-03-01

    Full Text Available AIM: To investigate the mechanism of retinal ganglion cell apoptosis in early stage of streptozotocin(STZ-induced diabetic rats. METHODS: Sixty SD rats were randomly divided into two groups: control group(CONand diabetes mellitus group(DM. Diabetic rat model was produced by intraperitoneal injection of 1% STZ in 30 adult male SD rats. At 4, 8, 12wk,the rats were killed and eyeballs were enucleated for the HE staining, TUNEL staining, transmission electron microscopy detection respectively, and laser confocal microscope detection was used to detect the calcium ion concentration.RESULTS:At 8wk RGCs decreased gradually and appeared disordered arrangement and got worse at 12wk in DM group. In DM group, mitochondrial swelling was detected at 4wk., and became more obvious, more in number at 8wk with reduction in some cells' volume and the number of organelles decreased. In DM group, few TUNEL positive RGCs were seen at 4wk, and became more and more at 8 and 12wk. The apoptosis index was significantly higher in DM group compared with CON group in different time points(PPPCONCLUSION: The study suggested that RGCs apoptosis occurs in early stage of diabetes, the mechanism might be associated with increased intracellular calcium ion concentration.

  6. Gender difference in the neuroprotective effect of rat bone marrow mesenchymal cells against hypoxia-induced apoptosis of retinal ganglion cells.

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    Yuan, Jing; Yu, Jian-Xiong

    2016-05-01

    Bone marrow mesenchymal stem cells can reduce retinal ganglion cell death and effectively prevent vision loss. Previously, we found that during differentiation, female rhesus monkey bone marrow mesenchymal stem cells acquire a higher neurogenic potential compared with male rhesus monkey bone marrow mesenchymal stem cells. This suggests that female bone marrow mesenchymal stem cells have a stronger neuroprotective effect than male bone marrow mesenchymal stem cells. Here, we first isolated and cultured bone marrow mesenchymal stem cells from female and male rats by density gradient centrifugation. Retinal tissue from newborn rats was prepared by enzymatic digestion to obtain primary retinal ganglion cells. Using the transwell system, retinal ganglion cells were co-cultured with bone marrow mesenchymal stem cells under hypoxia. Cell apoptosis was detected by flow cytometry and caspase-3 activity assay. We found a marked increase in apoptotic rate and caspase-3 activity of retinal ganglion cells after 24 hours of hypoxia compared with normoxia. Moreover, apoptotic rate and caspase-3 activity of retinal ganglion cells significantly decreased with both female and male bone marrow mesenchymal stem cell co-culture under hypoxia compared with culture alone, with more significant effects from female bone marrow mesenchymal stem cells. Our results indicate that bone marrow mesenchymal stem cells exert a neuroprotective effect against hypoxia-induced apoptosis of retinal ganglion cells, and also that female cells have greater neuroprotective ability compared with male cells.

  7. The role of NgR-Rhoa-Rock signal pathway in retinal ganglion cell apoptosis of early diabetic rats

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    Yun-Jie Fu

    2014-09-01

    Full Text Available AIM: To study the function and mechanism of the NgR-Rhoa-Rock signal pathways which exists in the retinal ganglion cells apoptosis in diabetes mellitus(DMrats. METHODS: Some healthy SD rats were operated by means of single intraperitoneal injection of 1% streptozotocin based on the standard of 50mg/kg wight, after that the blood sugar value was greater than 16.7mmol/L as DM model, then randomly divided into 3 groups, each group was 10 rats. In addition to take 10 healthy SD rats as control group. Four groups of rats were bilaterally eyeball intravitreal injection in turn with NgR-siRNA virus 10μL(siRNA group, NgR-siRNA virus diluted 10μL(DM group, NgR-siRNA virus-negative-control solution 10μL(siRNA blank group, NgR-siRNA virus diluted 10μL(normal control group, and fed normally. During that time, some life indexes like blood glucose, body mass, etc. were measured and recorded. After 12wk, the expression of NgR and Rhoa, HE staining, and TUNNEL staining were detected by Western blot analysis. RESULTS: Western blot analysis: compared with normal control group, the expression of NgR and Rhoa in DM group and siRNA blank group increased significantly(PP>0.05; compared with DM group and siRNA blank group, the expression of those proteins significantly lowered in siRNA group. HE staining: compared with normal control group, some extent ganglion cells arranged disorder, irregular shape, spacing not consistent were all found in three groups of model rats; compared with DM group and siRNA blank group, there was some improvement in siRNA group of ganglion cells about the order and shape size. TUNEL staining: compared with normal control group, there were retinal ganglion cells apoptosis in all of three groups of model rats. Compared with DM group and siRNA blank group, the number of retinal ganglion cells apoptotic cells was less, and the shape of cells had improved significantly in siRNA group. CONCLUSION: In the DM phase, the expression of NgR and

  8. c-Jun N-terminal kinase 3 expression in the retina of ocular hypertension mice: a possible target to reduce ganglion cell apoptosis

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    Yue He

    2015-01-01

    Full Text Available Glaucoma, a type of optic neuropathy, is characterized by the loss of retinal ganglion cells. It remains controversial whether c-Jun N-terminal kinase (JNK participates in the apoptosis of retinal ganglion cells in glaucoma. This study sought to explore a possible mechanism of action of JNK signaling pathway in glaucoma-induced retinal optic nerve damage. We established a mouse model of chronic ocular hypertension by reducing the aqueous humor followed by photocoagulation using the laser ignition method. Results showed significant pathological changes in the ocular tissues after the injury. Apoptosis of retinal ganglion cells increased with increased intraocular pressure, as did JNK3 mRNA expression in the retina. These data indicated that the increased expression of JNK3 mRNA was strongly associated with the increase in intraocular pressure in the retina, and correlated positively with the apoptosis of retinal ganglion cells.

  9. Neuroprotection of a novel cyclopeptide C*HSDGIC* from the cyclization of PACAP (1-5 in cellular and rodent models of retinal ganglion cell apoptosis.

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    Huanhuan Cheng

    Full Text Available To investigate the protective effects of a novel cyclopeptide C*HSDGIC* (CHC from the cyclization of Pituitary adenylate cyclase-activating polypeptide (PACAP (1-5 in cellular and rodent models of retinal ganglion cell apoptosis.Double-labeling immunohistochemistry was used to detect the expression of Thy-1 and PACAP receptor type 1 in a retinal ganglion cell line RGC-5. The apoptosis of RGC-5 cells was induced by 0.02 J/cm(2 Ultraviolet B irradiation. MTT assay, flow cytometry, fluorescence microscopy were used to investigate the viability, the level of reactive oxygen species (ROS and apoptosis of RGC-5 cells respectively. CHC attenuated apoptotic cell death induced by Ultraviolet B irradiation and inhibited the excessive generation of ROS. Moreover, CHC treatment resulted in decreased expression of Bax and concomitant increase of Bcl-2, as was revealed by western-blot analysis. The in vivo apoptosis of retinal ganglion cells was induced by injecting 50 mM N-methyl-D-aspartate (NMDA (100 nmol in a 2 µL saline solution intravitreally, and different dosages of CHC were administered. At day 7, rats in CHC+ NMDA-treated groups showed obvious aversion to light when compared to NMDA rats. Electroretinogram recordings revealed a marked decrease in the amplitudes of a-wave, b-wave, and photopic negative response due to NMDA damage. In retina receiving intravitreal NMDA and CHC co-treatment, these values were significantly increased. CHC treatment also resulted in less NMDA-induced cell loss and a decrease in the proportion of dUTP end-labeling-positive cells in ganglion cell line.C*HSDGIC*, a novel cyclopeptide from PACAP (1-5 attenuates apoptosis in RGC-5 cells and inhibits NMDA-induced retinal neuronal death. The beneficial effects may occur via the mitochondria pathway. PACAP derivatives like CHC may serve as a promising candidate for neuroprotection in glaucoma.

  10. Low-Intensity Pulsed Ultrasound Protects Retinal Ganglion Cell From Optic Nerve Injury Induced Apoptosis via Yes Associated Protein

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    Jia-Xing Zhou

    2018-06-01

    Full Text Available Background: Low-intensity pulsed ultrasound (LIPUS has been used in clinical studies. But little is known about its effects on the central nervous system (CNS, or its mechanism of action. Retinal ganglion cells (RGCs are CNS neuronal cells that can be utilized as a classic model system to evaluate outcomes of LIPUS protection from external trauma-induced retinal injury. In this study, we aim to: (1 determine the pulse energy and the capability of LIPUS in RGC viability, (2 ascertain the protective role of LIPUS in optic nerve (ON crush-induced retinal injury, and 3 explore the cellular mechanisms of RGC apoptosis prevention by LIPUS.Methods: An ON crush model was set up to induce RGC death. LIPUS was used to treat mice eyes daily, and the retina samples were dissected for immunostaining and Western blot. The expression of yes-associated protein (YAP and apoptosis-related proteins was detected by immunostaining and Western blot in vitro and in vivo. Apoptosis of RGCs was evaluated by TUNEL staining, the survival of RGCs and retained axons were labeled by Fluoro-gold and Tuj1 antibody, respectively. Rotenone was used to set up an in vitro cellular degenerative model and siYAP was used to interfering the expression of YAP to detect the LIPUS protective function.Results: LIPUS protected RGC from loss and apoptosis in vivo and in vitro. The ratio of cleaved/pro-caspase3 also decreased significantly under LIPUS treatment. As a cellular mechanical sensor, YAP expression increased and YAP translocated to nucleus in LIPUS stimulation group, however, phospho-YAP was found to be decreased. When YAP was inhibited, the LIPUS could not protect RGC from caspase3-dependent apoptosis.Conclusion: LIPUS prevented RGCs from apoptosis in an ON crush model and in vitro cellular degenerative model, which indicates a potential treatment for further traumatic ON injury. The mechanism of protection is dependent on YAP activation and correlated with caspase-3 signaling.

  11. Single-cell resolution imaging of retinal ganglion cell apoptosis in vivo using a cell-penetrating caspase-activatable peptide probe.

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    Xudong Qiu

    Full Text Available Peptide probes for imaging retinal ganglion cell (RGC apoptosis consist of a cell-penetrating peptide targeting moiety and a fluorophore-quencher pair flanking an effector caspase consensus sequence. Using ex vivo fluorescence imaging, we previously validated the capacity of these probes to identify apoptotic RGCs in cell culture and in an in vivo rat model of N-methyl- D-aspartate (NMDA-induced neurotoxicity. Herein, using TcapQ488, a new probe designed and synthesized for compatibility with clinically-relevant imaging instruments, and real time imaging of a live rat RGC degeneration model, we fully characterized time- and dose-dependent probe activation, signal-to-noise ratios, and probe safety profiles in vivo. Adult rats received intravitreal injections of four NMDA concentrations followed by varying TcapQ488 doses. Fluorescence fundus imaging was performed sequentially in vivo using a confocal scanning laser ophthalmoscope and individual RGCs displaying activated probe were counted and analyzed. Rats also underwent electroretinography following intravitreal injection of probe. In vivo fluorescence fundus imaging revealed distinct single-cell probe activation as an indicator of RGC apoptosis induced by intravitreal NMDA injection that corresponded to the identical cells observed in retinal flat mounts of the same eye. Peak activation of probe in vivo was detected 12 hours post probe injection. Detectable fluorescent RGCs increased with increasing NMDA concentration; sensitivity of detection generally increased with increasing TcapQ488 dose until saturating at 0.387 nmol. Electroretinography following intravitreal injections of TcapQ488 showed no significant difference compared with control injections. We optimized the signal-to-noise ratio of a caspase-activatable cell penetrating peptide probe for quantitative non-invasive detection of RGC apoptosis in vivo. Full characterization of probe performance in this setting creates an important in

  12. Effect of lycium barbarum polysaccharides on high glucose-induced retinal ganglion cell apoptosis, gene expression and delayed rectifier potassium current

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    Xiao-Fei Ma

    2017-05-01

    Full Text Available Objective: To study the effect of lycium barbarum polysaccharides (LBP on high glucoseinduced retinal ganglion cell apoptosis, gene expression and delayed rectifier potassium current. Methods: RGC-5 retinal ganglion cell lines were cultured and divided into control group, high glucose group and LBP group that were treated with normal DMEM, highglucose DMEM as well as high-glucose DMEM containing 500 ng/mL LBP respectively. After treatment, the Annexin V-FITC/PI kits were used to measure the number of apoptotic cells, fluorescence quantitative PCR kits were used to determine the expression of apoptosis genes and antioxidant genes, and patch clamp was used to test delayed rectifier potassium current. Results: 12, 24, 36 and 48 h after intervention, the number of apoptotic cells of high glucose group was significantly higher than that of control group, and the number of apoptotic cells of LBP group was significantly lower than that of high glucose group (P<0.05; 24 and 48 h after intervention, c-fos, c-jun, caspase-3, caspase-9, Nrf-2, NQO1 and HO-1 mRNA expression as well as potassium current amplitude (IK and maximum conductance (Gmax of high glucose group were significantly higher than those of control group while half maximum activation voltage (V1/2 was significantly lower than that of control group (P<0.05; c-fos, c-jun, caspase-3 and caspase-9 mRNA expression as well as IK and Gmax of LBP group were significantly lower than those of high glucose group, while Nrf-2, NQO1 and HO-1 mRNA expression as well as V1/2 of LBP group were significantly higher than those of high glucose group (P<0.05. Conclusions: LBP can reduce the high glucose-induced retinal ganglion cell apoptosis and inhibit the delayed rectifier potassium current amplitude.

  13. Crocin prevents retinal ischaemia/reperfusion injury-induced apoptosis in retinal ganglion cells through the PI3K/AKT signalling pathway.

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    Qi, Yun; Chen, Li; Zhang, Lei; Liu, Wen-Bo; Chen, Xiao-Yan; Yang, Xin-Guang

    2013-02-01

    Crocin is a pharmacologically active component of Crocus sativus L. (saffron) and has been reported to be useful in the treatment of neuronal damage. In the present study, we investigated the neuroprotective effect of crocin on retinal ganglion cells (RGCs) after retinal ischaemia/reperfusion (IR) injury, and our results show that crocin acts through the PI3K/AKT signalling pathway. Retinal IR injury was induced by raising the intraocular pressure of Sprague-Dawley rats to 110 mmHg for 60 min. The neuroprotective effect of crocin was determined by quantifying the surviving RGCs and apoptotic RGCs following IR injury by means of retrograde labelling and TUNEL staining, respectively. The phosphorylated AKT protein level was determined by western blot and immunohistochemical analysis. To determine the extent to which the PI3K/AKT pathway contributes to the neuroprotective effect of crocin, experiments were also performed using the PI3K inhibitor LY294002. Compared with the IR + vehicle group, crocin (50 mg/kg) treatment enhanced RGC survival by approximately 36% and decreased RGC apoptosis by 44% after retinal IR injury. Western blot and immunohistochemical analysis demonstrated that the PI3K/AKT pathway was activated by crocin in the ganglion cell layer after retinal IR injury. Intravitreal injection of LY294002 blocked the neuroprotective effect of crocin on IR-induced RGC death. In conclusion, crocin prevents retinal IR-induced apoptosis of RGCs by activating the PI3K/AKT signalling pathway. Copyright © 2012 Elsevier Ltd. All rights reserved.

  14. Age-related hearing loss: prevention of threshold declines, cell loss and apoptosis in spiral ganglion neurons

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    Zhu, Xiaoxia; Walton, Joseph P.

    2016-01-01

    Age-related hearing loss (ARHL) -presbycusis - is the most prevalent neurodegenerative disease and number one communication disorder of our aged population; and affects hundreds of millions of people worldwide. Its prevalence is close to that of cardiovascular disease and arthritis, and can be a precursor to dementia. The auditory perceptual dysfunction is well understood, but knowledge of the biological bases of ARHL is still somewhat lacking. Surprisingly, there are no FDA-approved drugs for treatment. Based on our previous studies of human subjects, where we discovered relations between serum aldosterone levels and the severity of ARHL, we treated middle age mice with aldosterone, which normally declines with age in all mammals. We found that hearing thresholds and suprathreshold responses significantly improved in the aldosterone-treated mice compared to the non-treatment group. In terms of cellular and molecular mechanisms underlying this therapeutic effect, additional experiments revealed that spiral ganglion cell survival was significantly improved, mineralocorticoid receptors were upregulated via post-translational protein modifications, and age-related intrinsic and extrinsic apoptotic pathways were blocked by the aldosterone therapy. Taken together, these novel findings pave the way for translational drug development towards the first medication to prevent the progression of ARHL. PMID:27667674

  15. Protective effects of a composition of Chinese herbs-Gurigumu-13 on retinal ganglion cell apoptosis in DBA/2J glaucoma mouse model

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    Qiu-Li Zhang

    2018-03-01

    Full Text Available AIM: To explore the concrete mechanism of a Mongolian compound medicine-Gurigumu-13 (GRGM for glaucoma treatment. METHODS: DBA/2J mice, as glaucoma models, were intragastric administrated with GRGM to study the effect of GRGM on retinal ganglion cells (RGCs. The loss of RGCs was evaluated with the number of RGCs and axons. The expression of the target protein of RGCs or mouse retinas was determined by Western blot. The relative content of malondialdehyde (MDA was examined by ELISA assay. RESULTS: GRGM distinctly improved retina damage via increasing the number of neurons, RGCs and axons in a concentration dependent manner. Meanwhile, GRGM obviously decreased the high level of MDA and the expression of oxidative stress-related proteins in retinas of DBA/2J mice, but promoted the expression of antioxidant proteins. Additionally, GRGM also significantly inhibited the protein expression of Bip and Chop, which were markers of endoplasmic reticulum stress-induced apoptosis. CONCLUSION: GRGM have obvious protective effects on RGCs in DBA/2J mice, and increase the number of RGCs and axons via inhibiting oxidative stress and endoplasmic reticulum stress.

  16. A novel model for rapid induction of apoptosis in spiral ganglions of mice.

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    Lee, Ji Eun; Nakagawa, Takayuki; Kim, Tae Soo; Iguchi, Fukuichiro; Endo, Tsuyoshi; Dong, Youyi; Yuki, Kazuo; Naito, Yasushi; Lee, Sang Heun; Ito, Juichi

    2003-06-01

    The survival of the spiral ganglion (SG) is a critical issue in preservation of hearing. Research on topics related to this issue requires a mouse experimental model because such a model has advantages including use of genetic information and knockout or "knockin" mice. Thus, the aim of the study was to establish a mouse model for induction of apoptosis of SG neurons with a definite time course. Laboratory study using experimental animals. C57BL/6 mice were used as experimental animals and were subjected to direct application of cisplatin into the inner ear. Terminal deoxynucleotidyl transferase-mediated dUTP nick-end labeling (TUNEL) assay and immunostaining for Neurofilament 200-kD (NF) and peripherin were used for analysis of SG degeneration. In addition, generation of peroxynitrite in affected spiral ganglions was examined by immunostaining for nitrotyrosine. Cellular location of activated caspase-9 and cytochrome-c in dying SG neurons were examined for analysis of cell death pathway. The TUNEL assay and immunohistochemical analysis for NF and peripherin indicated that type I neurons in spiral ganglions were deleted through the apoptotic pathway over time. Spiral ganglion neurons treated with cisplatin exhibited expression of nitrotyrosine, indicating induction of peroxynitrite by cisplatin. In dying SG neurons, expression of activated caspase-9 and translocation of cytochrome-c from mitochondria to cytoplasm were observed, indicating the mitochondrial pathway of apoptosis. The predictable fashion of induction of apoptosis in SG neurons over a well-defined time course in the model in the study will aid studies of the molecular mechanism of cell death and elucidation of a strategy for prevention of SG degeneration.

  17. Inhibition on Apoptosis Induced by Elevated Hydrostatic Pressure in Retinal Ganglion Cell-5 via Laminin Upregulating β1-integrin/Focal Adhesion Kinase/Protein Kinase B Signaling Pathway.

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    Li, Yi; Chen, Yan-Ming; Sun, Ming-Ming; Guo, Xiao-Dan; Wang, Ya-Chen; Zhang, Zhong-Zhi

    2016-04-20

    Glaucoma is a progressive optic neuropathy characterized by degeneration of neurons due to loss of retinal ganglion cells (RGCs). High intraocular pressure (HIOP), the main risk factor, causes the optic nerve damage. However, the precise mechanism of HIOP-induced RGC death is not yet completely understood. This study was conducted to determine apoptosis of RGC-5 cells induced by elevated hydrostatic pressures, explore whether laminin is associated with apoptosis under pressure, whether laminin can protect RGCs from apoptosis and affirm the mechanism that regulates the process of RGCs survival. RGC-5 cells were exposed to 0, 20, 40, and 60 mmHg in a pressurized incubator for 6, 12, and 24 h, respectively. The effect of elevated hydrostatic pressure on RGC-5 cells was measured by Annexin V-fluorescein isothiocyanate/propidium iodide staining, 3-(4,5-dimethylthiazol-2-yl)-2,5-diphenyltetrazolium bromide assay, and Western blotting of cleaved caspase-3 protein. Location and expression of laminin were detected by immunofluorescence. The expression of β1-integrin, phosphorylation of focal adhesion kinase (FAK) and protein kinase B (PKB, or AKT) were investigated with real-time polymerase chain reaction and Western blotting analysis. Elevated hydrostatic pressure induced apoptosis in cultured RGC-5 cells. Pressure with 40 mmHg for 24 h induced a maximum apoptosis. Laminin was declined in RGC-5 cells after exposing to 40 mmHg for 24 h. After pretreating with laminin, RGC-5 cells survived from elevated pressure. Furthermore, β1-integrin and phosphorylation of FAK and AKT were increased compared to 40 mmHg group. The data show apoptosis tendency of RGC-5 cells with elevated hydrostatic pressure. Laminin can protect RGC-5 cells against high pressure via β1-integrin/FAK/AKT signaling pathway. These results suggest that the decreased laminin of RGC-5 cells might be responsible for apoptosis induced by elevated hydrostatic pressure, and laminin or activating β1-integrin

  18. Regenerating reptile retinas: a comparative approach to restoring retinal ganglion cell function.

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    Williams, D L

    2017-02-01

    Transection or damage to the mammalian optic nerve generally results in loss of retinal ganglion cells by apoptosis. This cell death is seen less in fish or amphibians where retinal ganglion cell survival and axon regeneration leads to recovery of sight. Reptiles lie somewhere in the middle of this spectrum of nerve regeneration, and different species have been reported to have a significant variation in their retinal ganglion cell regenerative capacity. The ornate dragon lizard Ctenophoris ornatus exhibits a profound capacity for regeneration, whereas the Tenerife wall lizard Gallotia galloti has a more variable response to optic nerve damage. Some individuals regain visual activity such as the pupillomotor responses, whereas in others axons fail to regenerate sufficiently. Even in Ctenophoris, although the retinal ganglion cell axons regenerate adequately enough to synapse in the tectum, they do not make long-term topographic connections allowing recovery of complex visually motivated behaviour. The question then centres on where these intraspecies differences originate. Is it variation in the innate ability of retinal ganglion cells from different species to regenerate with functional validity? Or is it variances between different species in the substrate within which the nerves regenerate, the extracellular environment of the damaged nerve or the supporting cells surrounding the regenerating axons? Investigations of retinal ganglion cell regeneration between different species of lower vertebrates in vivo may shed light on these questions. Or perhaps more interesting are in vitro studies comparing axon regeneration of retinal ganglion cells from various species placed on differing substrates.

  19. Learning LM Specificity for Ganglion Cells

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    Ahumada, Albert J.

    2015-01-01

    Unsupervised learning models have been proposed based on experience (Ahumada and Mulligan, 1990;Wachtler, Doi, Lee and Sejnowski, 2007) that allow the cortex to develop units with LM specific color opponent receptive fields like the blob cells reported by Hubel and Wiesel on the basis of visual experience. These models used ganglion cells with LM indiscriminate wiring as inputs to the learning mechanism, which was presumed to occur at the cortical level.

  20. A Comparative Analysis of Ganglion Cell Complex Parameters in ...

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    Dr femi Oderinlo

    in the eyes, the optic nerve head, nerve fibre layer and retinal ganglion cells. Retinal ganglion cells encompass three layers ... of the macula in eyes with mild diabetic retinopathy. 8. *Correspondence: O Oderinlo, Eye Foundation ... most sensitive detection of GCC thinning. FLV provides a. 10 quantitative measure of the ...

  1. Caspases in retinal ganglion cell death and axon regeneration

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    Thomas, Chloe N; Berry, Martin; Logan, Ann; Blanch, Richard J; Ahmed, Zubair

    2017-01-01

    Retinal ganglion cells (RGC) are terminally differentiated CNS neurons that possess limited endogenous regenerative capacity after injury and thus RGC death causes permanent visual loss. RGC die by caspase-dependent mechanisms, including apoptosis, during development, after ocular injury and in progressive degenerative diseases of the eye and optic nerve, such as glaucoma, anterior ischemic optic neuropathy, diabetic retinopathy and multiple sclerosis. Inhibition of caspases through genetic or pharmacological approaches can arrest the apoptotic cascade and protect a proportion of RGC. Novel findings have also highlighted a pyroptotic role of inflammatory caspases in RGC death. In this review, we discuss the molecular signalling mechanisms of apoptotic and inflammatory caspase responses in RGC specifically, their involvement in RGC degeneration and explore their potential as therapeutic targets. PMID:29675270

  2. Topography of ganglion cell production in the cat's retina

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    Walsh, C.; Polley, E.H.

    1985-01-01

    The ganglion cells of the cat's retina form several classes distinguishable in terms of soma size, axon diameter, dendritic morphology, physiological properties, and central connections. Labeling with [ 3 H]thymidine shows that the ganglion cells which survive in the adult are produced as several temporally shifted, overlapping waves: medium-sized cells are produced before large cells, whereas the smallest ganglion cells are produced throughout the period of ganglion cell generation. Large cells and medium-sized cells show the same distinctive pattern of production, forming rough spirals around the area centralis. The oldest cells tend to lie superior and nasal to the area centralis, whereas cells in the inferior nasal retina and inferior temporal retina are, in general, progressively younger. Within each retinal quadrant, cells nearer the area centralis tend to be older than cells in the periphery, but there is substantial overlap. The retinal raphe divides the superior temporal quadrant into two zones with different patterns of cell addition. Superior temporal retina near the vertical meridian adds cells only slightly later than superior nasal retina, whereas superior temporal retina near the horizontal meridian adds cells very late, contemporaneously with inferior temporal retina. The broader wave of production of smaller ganglion cells seems to follow this same spiral pattern at its beginning and end. The presence of the area centralis as a nodal point about which ganglion cell production in the retinal quadrants pivots suggests that the area centralis is already an important retinal landmark even at the earliest stages of retinal development

  3. Dorsal raphe nucleus projecting retinal ganglion cells: Why Y cells?

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    Pickard, Gary E.; So, Kwok-Fai; Pu, Mingliang

    2015-01-01

    Retinal ganglion Y (alpha) cells are found in retinas ranging from frogs to mice to primates. The highly conserved nature of the large, fast conducting retinal Y cell is a testament to its fundamental task, although precisely what this task is remained ill-defined. The recent discovery that Y-alpha retinal ganglion cells send axon collaterals to the serotonergic dorsal raphe nucleus (DRN) in addition to the lateral geniculate nucleus (LGN), medial interlaminar nucleus (MIN), pretectum and the superior colliculus (SC) has offered new insights into the important survival tasks performed by these cells with highly branched axons. We propose that in addition to its role in visual perception, the Y-alpha retinal ganglion cell provides concurrent signals via axon collaterals to the DRN, the major source of serotonergic afferents to the forebrain, to dramatically inhibit 5-HT activity during orientation or alerting/escape responses, which dis-facilitates ongoing tonic motor activity while dis-inhibiting sensory information processing throughout the visual system. The new data provide a fresh view of these evolutionarily old retinal ganglion cells. PMID:26363667

  4. Hypoxia-ischemia and retinal ganglion cell damage

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    Charanjit Kaur

    2008-08-01

    Full Text Available Charanjit Kaur1, Wallace S Foulds2, Eng-Ang Ling11Department of Anatomy, Yong Loo Lin School of Medicine, National University of Singapore, Singapore; 2Singapore Eye Research Institute, SingaporeAbstract: Retinal hypoxia is the potentially blinding mechanism underlying a number of sight-threatening disorders including central retinal artery occlusion, ischemic central retinal vein thrombosis, complications of diabetic eye disease and some types of glaucoma. Hypoxia is implicated in loss of retinal ganglion cells (RGCs occurring in such conditions. RGC death occurs by apoptosis or necrosis. Hypoxia-ischemia induces the expression of hypoxia inducible factor-1α and its target genes such as vascular endothelial growth factor (VEGF and nitric oxide synthase (NOS. Increased production of VEGF results in disruption of the blood retinal barrier leading to retinal edema. Enhanced expression of NOS results in increased production of nitric oxide which may be toxic to the cells resulting in their death. Excess glutamate release in hypoxic-ischemic conditions causes excitotoxic damage to the RGCs through activation of ionotropic and metabotropic glutamate receptors. Activation of glutamate receptors is thought to initiate damage in the retina by a cascade of biochemical effects such as neuronal NOS activation and increase in intracellular Ca2+ which has been described as a major contributing factor to RGC loss. Excess production of proinflammatory cytokines also mediates cell damage. Besides the above, free-radicals generated in hypoxic-ischemic conditions result in RGC loss because of an imbalance between antioxidant- and oxidant-generating systems. Although many advances have been made in understanding the mediators and mechanisms of injury, strategies to improve the damage are lacking. Measures to prevent neuronal injury have to be developed.Keywords: retinal hypoxia, retinal ganglion cells, glutamate receptors, neuronal injury, retina

  5. Melanopsin retinal ganglion cell loss in Alzheimer's disease

    DEFF Research Database (Denmark)

    La Morgia, Chiara; Ross-Cisneros, Fred N; Koronyo, Yosef

    2015-01-01

    OBJECTIVE: Melanopsin retinal ganglion cells (mRGCs) are photoreceptors driving circadian photoentrainment, and circadian dysfunction characterizes Alzheimer's disease (AD). We investigated mRGCs in AD, hypothesizing their contribution to circadian dysfunction. METHODS: We assessed retinal nerve...

  6. Troxler Fading, Eye Movements, and Retinal Ganglion Cell Properties

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    Romain Bachy

    2014-12-01

    Full Text Available We present four movies demonstrating the effect of flicker and blur on the magnitude and speed of adaptation for foveal and peripheral vision along the three color axes that isolate retinal ganglion cells projecting to magno, parvo, and konio layers of the LGN. The demonstrations support the eye movement hypothesis for Troxler fading for brightness and color, and demonstrate the effects of flicker and blur on adaptation of each class of retinal ganglion cells.

  7. Dominant inheritance of retinal ganglion cell resistance to optic nerve crush in mice

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    Schlamp Cassandra L

    2007-03-01

    Full Text Available Abstract Background Several neurodegenerative diseases are influenced by complex genetics that affect an individual's susceptibility, disease severity, and rate of progression. One such disease is glaucoma, a chronic neurodegenerative condition of the eye that targets and stimulates apoptosis of CNS neurons called retinal ganglion cells. Since ganglion cell death is intrinsic, it is reasonable that the genes that control this process may contribute to the complex genetics that affect ganglion cell susceptibility to disease. To determine if genetic background influences susceptibility to optic nerve damage, leading to ganglion cell death, we performed optic nerve crush on 15 different inbred lines of mice and measured ganglion cell loss. Resistant and susceptible strains were used in a reciprocal breeding strategy to examine the inheritance pattern of the resistance phenotype. Because earlier studies had implicated Bax as a susceptibility allele for ganglion cell death in the chronic neurodegenerative disease glaucoma, we conducted allelic segregation analysis and mRNA quantification to assess this gene as a candidate for the cell death phenotype. Results Inbred lines showed varying levels of susceptibility to optic nerve crush. DBA/2J mice were most resistant and BALB/cByJ mice were most susceptible. F1 mice from these lines inherited the DBA/2J phenotype, while N2 backcross mice exhibited the BALB/cByJ phenotype. F2 mice exhibited an intermediate phenotype. A Wright Formula calculation suggested as few as 2 dominant loci were linked to the resistance phenotype, which was corroborated by a Punnett Square analysis of the distribution of the mean phenotype in each cross. The levels of latent Bax mRNA were the same in both lines, and Bax alleles did not segregate with phenotype in N2 and F2 mice. Conclusion Inbred mice show different levels of resistance to optic nerve crush. The resistance phenotype is heritable in a dominant fashion involving

  8. THE MODULATORY ROLE OF TAURINE IN RETINAL GANGLION CELLS

    Science.gov (United States)

    Jiang, Zheng; Bulley, Simon; Guzzone, Joseph; Ripps, Harris; Shen, Wen

    2017-01-01

    Taurine (2-aminoethylsuphonic acid) is present in nearly all animal tissues, and is the most abundant free amino acid in muscle, heart, CNS and retina. Although it is known to be a major cytoprotectant and essential for normal retinal development, its role in retinal neurotransmission and modulation is not well understood. We investigated the response of taurine in retinal ganglion cells, and its effect on synaptic transmission between ganglion cells and their pre-synaptic neurons. We find that taurine-elicited currents in ganglion cells could be fully blocked by both strychnine and SR95531, glycine and GABAA receptor antagonists, respectively. This suggests that taurine-activated receptors might share the antagonists with GABA and glycine receptors. The effect of taurine at micromolar concentrations can effectively suppress spontaneous vesicle release from the pre-synaptic neurons, but had limited effects on light-evoked synaptic signals in ganglion cells. We also describe a metabotropic effect of taurine in the suppression of light-evoked response in ganglion cells. Clearly, taurine acts in multiple ways to modulate synaptic signals in retinal output neurons, ganglion cells. PMID:23392924

  9. An Optic Nerve Crush Injury Murine Model to Study Retinal Ganglion Cell Survival

    Science.gov (United States)

    Tang, Zhongshu; Zhang, Shuihua; Lee, Chunsik; Kumar, Anil; Arjunan, Pachiappan; Li, Yang; Zhang, Fan; Li, Xuri

    2011-01-01

    Injury to the optic nerve can lead to axonal degeneration, followed by a gradual death of retinal ganglion cells (RGCs), which results in irreversible vision loss. Examples of such diseases in human include traumatic optic neuropathy and optic nerve degeneration in glaucoma. It is characterized by typical changes in the optic nerve head, progressive optic nerve degeneration, and loss of retinal ganglion cells, if uncontrolled, leading to vision loss and blindness. The optic nerve crush (ONC) injury mouse model is an important experimental disease model for traumatic optic neuropathy, glaucoma, etc. In this model, the crush injury to the optic nerve leads to gradual retinal ganglion cells apoptosis. This disease model can be used to study the general processes and mechanisms of neuronal death and survival, which is essential for the development of therapeutic measures. In addition, pharmacological and molecular approaches can be used in this model to identify and test potential therapeutic reagents to treat different types of optic neuropathy. Here, we provide a step by step demonstration of (I) Baseline retrograde labeling of retinal ganglion cells (RGCs) at day 1, (II) Optic nerve crush injury at day 4, (III) Harvest the retinae and analyze RGC survival at day 11, and (IV) Representative result. PMID:21540827

  10. Retinal ganglion cell topography and spatial resolving power in penguins.

    Science.gov (United States)

    Coimbra, João Paulo; Nolan, Paul M; Collin, Shaun P; Hart, Nathan S

    2012-01-01

    Penguins are a group of flightless seabirds that exhibit numerous morphological, behavioral and ecological adaptations to their amphibious lifestyle, but little is known about the topographic organization of neurons in their retinas. In this study, we used retinal wholemounts and stereological methods to estimate the total number and topographic distribution of retinal ganglion cells in addition to an anatomical estimate of spatial resolving power in two species of penguins: the little penguin, Eudyptula minor, and the king penguin, Aptenodytes patagonicus. The total number of ganglion cells per retina was approximately 1,200,000 in the little penguin and 1,110,000 in the king penguin. The topographic distribution of retinal ganglion cells in both species revealed the presence of a prominent horizontal visual streak with steeper gradients in the little penguin. The little penguin retinas showed ganglion cell density peaks of 21,867 cells/mm², affording spatial resolution in water of 17.07-17.46 cycles/degree (12.81-13.09 cycles/degree in air). In contrast, the king penguin showed a relatively lower peak density of ganglion cells of 14,222 cells/mm², but--due to its larger eye--slightly higher spatial resolution in water of 20.40 cycles/degree (15.30 cycles/degree in air). In addition, we mapped the distribution of giant ganglion cells in both penguin species using Nissl-stained wholemounts. In both species, topographic mapping of this cell type revealed the presence of an area gigantocellularis with a concentric organization of isodensity contours showing a peak in the far temporal retina of approximately 70 cells/mm² in the little penguin and 39 cells/mm² in the king penguin. Giant ganglion cell densities gradually fall towards the outermost isodensity contours revealing the presence of a vertically organized streak. In the little penguin, we confirmed our cytological characterization of giant ganglion cells using immunohistochemistry for microtubule

  11. Patterns of lipofuscin accumulation in ganglionic nerve cells of superior cervical ganglion in humans

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    Živković Vladimir

    2008-01-01

    Full Text Available Background/Aim. Considering available literature lipofuscin is a classical age pigment of postmitotic cells, and a consistently recognized phenomenon in humans and animals. Lipofuscin accumulation is characteristic for nerve cells that are postmitotic. This research was focused on lipofuscin accumulation in ganglionic cells (GC (postganglionic sympathetic cell bodies of superior cervical ganglion in humans during ageing. Methods. We analysed 30 ganglions from cadavers ranging from 20 to over 80 years of age. As material the tissue samples were used from the middle portion of the ganglion, which was separated from the surrounding tissue by the method of macrodissection. The tissue samples were routinely fixed in 10% neutral formalin and embedded in paraffin for classical histological analysis, then three consecutive (successive sections 5 μm thick were made and stained with hematoxylin and eosin method (HE, silver impregnation technique by Masson Fontana and trichrome stain by Florantin. Results. Immersion microscopy was used to analyse patterns of lipofuscin accumulation during ageing making possible to distinguish diffuse type (lipofuscin granules were irregularly distributed and non-confluent, unipolar type (lipofuscin granules were grouped at the end of the cell, bipolar type (lipofuscin granules were concentrated at the two opposite ends of a cell with the nucleus in between at the center of a cell, annular type (lipofuscin granules were in the shape of a complete or incomplete ring around the nucleus and a cell completely filled with lipofuscin (two subtypes distinguishing, one with visible a nucleus, and the other with invisible one. Even at the age of 20 there were cells with lipofuscin granules accumulated in diffuse way, but in smaller numbers; the GC without lipofuscin were dominant. Growing older, especially above 60 years, all of the above mentioned patterns of lipofuscin accumulation were present with the evident increase in cells

  12. Curcumin Attenuates Staurosporine-Mediated Death of Retinal Ganglion Cells

    OpenAIRE

    Burugula, Balabharathi; Ganesh, Bhagyalaxmi S.; Chintala, Shravan K.

    2011-01-01

    The functional effect of curcumin, a free radical scavenger and an herbal medicine from Indian yellow curry spice, Curcuma longa, on protease-mediated retinal ganglion cell death was investigated. These results show, for the first time, that curcumin indeed prevents the protease-mediated death of RGCs, both in vitro and in vivo.

  13. Processing of natural temporal stimuli by macaque retinal ganglion cells

    NARCIS (Netherlands)

    Hateren, J.H. van; Rüttiger, L.; Lee, B.B.

    2002-01-01

    This study quantifies the performance of primate retinal ganglion cells in response to natural stimuli. Stimuli were confined to the temporal and chromatic domains and were derived from two contrasting environments, one typically northern European and the other a flower show. The performance of the

  14. Msx2 alters the timing of retinal ganglion cells fate commitment and differentiation

    Energy Technology Data Exchange (ETDEWEB)

    Jiang, Shao-Yun, E-mail: jiangshaoyun@yahoo.com [School of Dentistry, Tianjin Medical University, 12 Qi Xiang Tai Street, Tianjin 300070 (China); Wang, Jian-Tao, E-mail: wangjiantao65@hotmail.com [Eye Center, Tianjin Medical University, 64 Tongan Road, Tianjin 300070 (China); Dohney Eye Institute, Keck School of Medicine, University of Southern California, 1355 San Pablo Street, DOH 314, Los Angeles, CA 90033 (United States)

    2010-05-14

    Timing of cell fate commitment determines distinct retinal cell types, which is believed to be controlled by a tightly coordinated regulatory program of proliferation, cell cycle exit and differentiation. Although homeobox protein Msx2 could induce apoptosis of optic vesicle, it is unclear whether Msx2 regulates differentiation and cell fate commitment of retinal progenitor cells (RPCs) to retinal ganglion cells (RGCs). In this study, we show that overexpression of Msx2 transiently suppressed the expression of Cyclin D1 and blocked cell proliferation. Meanwhile, overexpression of Msx2 delayed the expression of RGC-specific differentiation markers (Math5 and Brn3b), which showed that Msx2 could affect the timing of RGCs fate commitment and differentiation by delaying the timing of cell cycle exit of retinal progenitors. These results indicate Msx2 possesses dual regulatory functions in controlling cell cycle progression of retinal RPCs and timing of RGCs differentiation.

  15. Veratridine increases the survival of retinal ganglion cells in vitro

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    S.P.F. Pereira

    1997-12-01

    Full Text Available Neuronal cell death is an important phenomenon involving many biochemical pathways. This degenerative event has been studied to understand how the cells activate the mechanisms that lead to self-destruction. Target cells and afferent cells play a relevant role in the regulation of natural cell death. We studied the effect of veratridine (1.5, 3.0, 4.5 and 6.0 µM on the survival of neonatal rat retinal ganglion cells in vitro. Veratridine (3.0 µM, a well-known depolarizing agent that opens the Na+ channel, promoted a two-fold increase in the survival of retinal ganglion cells kept in culture for 48 h. This effect was dose-dependent and was blocked by 1.0 µM tetrodotoxin (a classical voltage-dependent Na+ channel blocker and 30.0 µM flunarizine (a Na+ and Ca2+ channel blocker. These results indicate that electrical activity is also important for the maintenance of retinal ganglion cell survival in vitro

  16. Retinal Ganglion Cell Loss in Diabetes Associated with Elevated Homocysteine

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    Kenneth S. Shindler

    2009-11-01

    Full Text Available A number of studies have suggested that homocysteine may be a contributing factor to development of retinopathy in diabetic patients based on observed correlations between elevated homocysteine levels and the presence of retinopathy. The significance of such a correlation remains to be determined, and potential mechanisms by which homocysteine might induce retinopathy have not been well characterized. Ganapathy and colleagues1 used mutant mice that have endogenously elevated homocysteine levels due to heterozygous deletion of the cystathionine-β-synthase gene to examine changes in retinal pathology following induction of diabetes. Their finding that elevated homocysteine levels hastens loss of cells in the retinal ganglion cell layer suggests that toxicity to ganglion cells may warrant further investigation as a potential mechanism of homocysteine enhanced susceptibility to diabetic retinopathy.

  17. Age-Related Change in Vestibular Ganglion Cell Populations in Individuals With Presbycusis and Normal Hearing.

    Science.gov (United States)

    Gluth, Michael B; Nelson, Erik G

    2017-04-01

    We sought to establish that the decline of vestibular ganglion cell counts uniquely correlates with spiral ganglion cell counts, cochlear hair cell counts, and hearing phenotype in individuals with presbycusis. The relationship between aging in the vestibular system and aging in the cochlea is a topic of ongoing investigation. Histopathologic age-related changes the vestibular system may mirror what is seen in the cochlea, but correlations with hearing phenotype and the impact of presbycusis are not well understood. Vestibular ganglion cells, spiral ganglion cells, and cochlear hair cells were counted in specimens from individuals with presbycusis and normal hearing. These were taken from within a large collection of processed human temporal bones. Correlations between histopathology and hearing phenotype were investigated. Vestibular ganglion cell counts were positively correlated with spiral ganglion cell counts and cochlear hair cell counts and were negatively correlated with hearing phenotype. There was no statistical evidence on linear regression to suggest that the relationship between age and cell populations differed significantly according to whether presbycusis was present or not. Superior vestibular ganglion cells were more negatively correlated with age than inferior ganglion cells. No difference in vestibular ganglion cells was noted based on sex. Vestibular ganglion cell counts progressively deteriorate with age, and this loss correlates closely with changes in the cochlea, as well as hearing phenotype. However, these correlations do not appear to be unique in individuals with presbycusis as compared with those with normal hearing.

  18. Autophagy in retinal ganglion cells in a rhesus monkey chronic hypertensive glaucoma model.

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    Shuifeng Deng

    Full Text Available Primary open angle glaucoma (POAG is a neurodegenerative disease characterized by physiological intraocular hypertension that causes damage to the retinal ganglion cells (RGCs. In the past, RGC damage in POAG was suggested to have been attributed to RGC apoptosis. However, in the present study, we applied a model closer to human POAG through the use of a chronic hypertensive glaucoma model in rhesus monkeys to investigate whether another mode of progressive cell death, autophagy, was activated in the glaucomatous retinas. First, in the glaucomatous retinas, the levels of LC3B-II, LC3B-II/LC3B-I and Beclin 1 increased as demonstrated by Western blot analyses, whereas early or initial autophagic vacuoles (AVi and late or degraded autophagic vacuoles (AVd accumulated in the ganglion cell layer (GCL and in the inner plexiform layer (IPL as determined by transmission electron microscopy (TEM analysis. Second, lysosome activity and autophagosome-lysosomal fusion increased in the RGCs of the glaucomatous retinas, as demonstrated by Western blotting against lysosome associated membrane protein-1 (LAMP1 and double labeling against LC3B and LAMP1. Third, apoptosis was activated in the glaucomatous eyes with increased levels of caspase-3 and cleaved caspase-3 and an increased number of TUNEL-positive RGCs. Our results suggested that autophagy was activated in RGCs in the chronic hypertensive glaucoma model of rhesus monkeys and that autophagy may have potential as a new target for intervention in glaucoma treatment.

  19. Intrinsically photosensitive retinal ganglion cell function in relation to age

    DEFF Research Database (Denmark)

    Herbst, Kristina; Sander, Birgit; Lund-Andersen, Henrik

    2012-01-01

    The activity of melanopsin containing intrinsically photosensitive ganglion retinal cells (ipRGC) can be assessed by a means of pupil responses to bright blue (appr.480 nm) light. Due to age related factors in the eye, particularly, structural changes of the lens, less light reaches retina. The aim...... of this study was to examine how age and in vivo measured lens transmission of blue light might affect pupil light responses, in particular, mediated by the ipRGC....

  20. Melanopsin expressing human retinal ganglion cells

    DEFF Research Database (Denmark)

    Hannibal, Jens; Christiansen, Anders Tolstrup; Heegaard, Steffen

    2017-01-01

    microscopy and 3D reconstruction of melanopsin immunoreactive (-ir) RGCs, we applied the criteria used in mouse on human melanopsin-ir RGCs. We identified M1, displaced M1, M2, and M4 cells. We found two other subtypes of melanopsin-ir RGCs, which were named "gigantic M1 (GM1)" and "gigantic displaced M1...

  1. POSTTREATMENT NEUROBLASTOMA MATURATION TO GANGLIONIC CELL TUMOR

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    M. V. Ryzhova

    2012-01-01

    Full Text Available Tumor cells can differentiate into more mature forms in undifferentiated or poorly differentiated tumors, such as medulloblastomas with increased nodularity, as well as neuroblastomas. The authors describe 2 cases of neuroblastoma maturation into ganglioneuroblastoma 5 months after chemotherapy in a 2-year-old girl and 3 years after radiotherapy in a 16-year-old girl.

  2. Melanopsin-expressing retinal ganglion cells: implications for human diseases

    DEFF Research Database (Denmark)

    La Morgia, Chiara; Ross-Cisneros, Fred N; Hannibal, Jens

    2011-01-01

    In the last decade, there was the seminal discovery of melanopsin-expressing retinal ganglion cells (mRGCs) as a new class of photoreceptors that subserve the photoentrainment of circadian rhythms and other non-image forming functions of the eye. Since then, there has been a growing research...... interest on these cells, mainly focused on animal models. Only recently, a few studies have started to address the relevance of the mRGC system in humans and related diseases. We recently discovered that mRGCs resist neurodegeneration in two inherited mitochondrial disorders that cause blindness, i...

  3. Progranulin deficiency causes the retinal ganglion cell loss during development.

    Science.gov (United States)

    Kuse, Yoshiki; Tsuruma, Kazuhiro; Mizoguchi, Takahiro; Shimazawa, Masamitsu; Hara, Hideaki

    2017-05-10

    Astrocytes are glial cells that support and protect neurons in the central nervous systems including the retina. Retinal ganglion cells (RGCs) are in contact with the astrocytes and our earlier findings showed the reduction of the number of cells in the ganglion cell layer in adult progranulin deficient mice. In the present study, we focused on the time of activation of the astrocytes and the alterations in the number of RGCs in the retina and optic nerve in progranulin deficient mice. Our findings showed that the number of Brn3a-positive cells was reduced and the expression of glial fibrillary acidic protein (GFAP) was increased in progranulin deficient mice. The progranulin deficient mice had a high expression of GFAP on postnatal day 9 (P9) but not on postnatal day 1. These mice also had a decrease in the number of the Brn3a-positive cells on P9. Taken together, these findings indicate that the absence of progranulin can affect the survival of RGCs subsequent the activation of astrocytes during retinal development.

  4. Neuroprotection of the rat’s retinal ganglion cells against glutamate-induced toxicity

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    Kariman M.A El-Gohari

    2016-01-01

    Conclusion Taurine protects the retina against glutamate excitotoxicity and could have clinical implications in protecting the ganglion cells from several ophthalmic diseases such as glaucoma and diabetic retinopathy.

  5. Responses of macaque ganglion cells to far violet lights

    International Nuclear Information System (INIS)

    De Monasterio, F.M.; Gouras, P.

    1977-01-01

    In a sample of 487 colour-opponent ganglion cells recorded in the central retina of the rhesus and cynomolgus monkeys, 9% of these neurones were found to have responses with the same sign at both ends of the visible spectrum mediated by red-sensitive cones and mid-spectral responses of opposite sign mediated by green-sensitive cones. Selective chromatic adaptation showed that the responses to far violet lights (400 to 420 nm) were due to input from red- and not blue-sensitive cones. These responses were enhanced by backgrounds depressing the sensitivity of blue- and green-sensitive cones and they were depressed by backgrounds depressing the sensitivity of red-sensitive cones; the sensitivity of these responses was yoked to that of responses to far red lights. The relative incidence of these ganglion cells was maximal at the foveal region and decreased towards the peripheral retina. The properties of these cells are consistent with some psychophysical observations of human vision at the short wave-lengths. (author)

  6. The circadian response of intrinsically photosensitive retinal ganglion cells.

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    Andrew J Zele

    Full Text Available Intrinsically photosensitive retinal ganglion cells (ipRGC signal environmental light level to the central circadian clock and contribute to the pupil light reflex. It is unknown if ipRGC activity is subject to extrinsic (central or intrinsic (retinal network-mediated circadian modulation during light entrainment and phase shifting. Eleven younger persons (18-30 years with no ophthalmological, medical or sleep disorders participated. The activity of the inner (ipRGC and outer retina (cone photoreceptors was assessed hourly using the pupil light reflex during a 24 h period of constant environmental illumination (10 lux. Exogenous circadian cues of activity, sleep, posture, caffeine, ambient temperature, caloric intake and ambient illumination were controlled. Dim-light melatonin onset (DLMO was determined from salivary melatonin assay at hourly intervals, and participant melatonin onset values were set to 14 h to adjust clock time to circadian time. Here we demonstrate in humans that the ipRGC controlled post-illumination pupil response has a circadian rhythm independent of external light cues. This circadian variation precedes melatonin onset and the minimum ipRGC driven pupil response occurs post melatonin onset. Outer retinal photoreceptor contributions to the inner retinal ipRGC driven post-illumination pupil response also show circadian variation whereas direct outer retinal cone inputs to the pupil light reflex do not, indicating that intrinsically photosensitive (melanopsin retinal ganglion cells mediate this circadian variation.

  7. Regulation of Taurine transporter activity in cultured rat retinal ganglion cells and rat retinal Muller Cells

    International Nuclear Information System (INIS)

    Eissa, Laila A.; Smith, Sylvia B.; El-sherbeny, Amira A.

    2006-01-01

    Diabetic retinopathy is one of the most common complications of diabetes. The amino acid taurine is believed to play an antioxidant protective role in diabetic retinopathy through the scavenging of the reactive species. It is not well established whether taurine uptake is altered in retina cells during diabetic conditions. Thus, the present study was designed to investigate the changes in taurine transport in cultures of rat retinal Muller cells and rat retinal ganglion cells under conditions associated with diabetes. Taurine was abundantly taken up by retinal Muller cells and rat retinal ganglion cells under normal glycemic condition. Taurine was actively transported to rat Muller cells and rat retinal ganglion cells in a Na and Cl dependant manner. Taurine uptake further significantly elevated in both type of cells after the incubation with high glucose concentration. This effect could be attributed to the increase in osmolarity. Because Nitric Oxide (NO) is a molecule implicated in the pathogenesis of diabetes, we also determined the activity of taurine transporter in cultured rat retinal Muller cells and rat retinal ganglion cells in the presence of the NO donors, SIN-1 and SNAP. Taurine uptake was elevated above control value after 24-h incubation with low concentration of NO donors. We finally investigated the ability of neurotoxic glutamate to change taurine transporter activity in both types of cells. Uptake of taurine was significantly increased in rat retinal ganglion cells when only incubated with high concentration of glutamate. Our data provide evidence that taurine transporter is present in cultured rat retinal ganglion and Muller cells and is regulated by hyperosmolarity. The data are relevant to disease such as diabetes and neuronal degeneration where retinal cell volume may dramatically change. (author)

  8. Recovery of cat retinal ganglion cell sensitivity following pigment bleaching.

    Science.gov (United States)

    Bonds, A B; Enroth-Cugell, C

    1979-01-01

    1. The threshold illuminance for small spot stimulation of on-centre cat retinal ganglion cells was plotted vs. time after exposure to adapting light sufficiently strong to bleach significant amounts of rhodopsin. 2. When the entire receptive field of an X- or Y-type ganglion cell is bleached by at most 40%, recovery of the cell's rod-system proceeds in two phases: an early relatively fast one during which the response appears transient, and a late, slower one during which responses become more sustained. Log threshold during the later phase is well fit by an exponential in time (tau = 11.5-38 min). 3. After bleaches of 90% of the underlying pigment, threshold is cone-determined for as long as 40 min. Rod threshold continues to decrease for at least 85 min after the bleach. 4. The rate of recovery is slower after strong than after weak bleaches; 10 and 90% bleaches yield time constants for the later phase of 11.5 and 38 min, respectively. This contrasts with an approximate time constant of 11 min for rhodopsin regeneration following any bleach. 5. The relationship between the initial elevation of log rod threshold extrapolated from the fitted exponential curves and the initial amount of pigment bleached is monotonic, but nonlinear. 6. After a bleaching exposure, the maintained discharge is initially very regular. The firing rate first rises, then falls to the pre-bleach level, with more extended time courses of change in firing rate after stronger exposures. The discharge rate is restored before threshold has recovered fully. 7. The change in the response vs. log stimulus relationship after bleaching is described as a shift of the curve to the right, paired with a decrease in slope of the linear segment of the curve. PMID:521963

  9. Hydrostatic Pressure Does Not Cause Detectable Changes in Survival of Human Retinal Ganglion Cells

    Science.gov (United States)

    Osborne, Andrew; Aldarwesh, Amal; Rhodes, Jeremy D.; Broadway, David C.; Everitt, Claire; Sanderson, Julie

    2015-01-01

    Purpose Elevated intraocular pressure (IOP) is a major risk factor for glaucoma. One consequence of raised IOP is that ocular tissues are subjected to increased hydrostatic pressure (HP). The effect of raised HP on stress pathway signaling and retinal ganglion cell (RGC) survival in the human retina was investigated. Methods A chamber was designed to expose cells to increased HP (constant and fluctuating). Accurate pressure control (10-100mmHg) was achieved using mass flow controllers. Human organotypic retinal cultures (HORCs) from donor eyes (pressure for 24 or 48h caused no loss of structural integrity, LDH release, decrease in RGC marker expression (THY-1) or loss of RGCs compared with controls. In addition, there was no increase in TUNEL-positive NeuN-labelled cells at either time-point indicating no increase in apoptosis of RGCs. OGD increased apoptosis, reduced RGC marker expression and RGC number and caused elevated LDH release at 24h. p38 and JNK phosphorylation remained unchanged in HORCs exposed to fluctuating pressure (10-100mmHg; 1 cycle/min) for 15, 30, 60 and 90min durations, whereas OGD (3h) increased activation of p38 and JNK, remaining elevated for 90min post-OGD. Conclusions Directly applied HP had no detectable impact on RGC survival and stress-signalling in HORCs. Simulated ischemia, however, activated stress pathways and caused RGC death. These results show that direct HP does not cause degeneration of RGCs in the ex vivo human retina. PMID:25635827

  10. Gene transfection mediated by polyethyleneimine-polyethylene glycol nanocarrier prevents cisplatin-induced spiral ganglion cell damage

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    Guan-gui Chen

    2015-01-01

    Full Text Available Polyethyleneimine-polyethylene glycol (PEI-PEG, a novel nanocarrier, has been used for transfection and gene therapy in a variety of cells. In our previous study, we successfully carried out PEI-PEG-mediated gene transfer in spiral ganglion cells. It remains unclear whether PEI-PEG could be used for gene therapy with X-linked inhibitor of apoptosis protein (XIAP in the inner ear. In the present study, we performed PEI-PEG-mediated XIAP gene transfection in the cochlea of Sprague-Dawley rats, via scala tympani fenestration, before daily cisplatin injections. Auditory brainstem reflex tests demonstrated the protective effects of XIAP gene therapy on auditory function. Immunohistochemical staining revealed XIAP protein expression in the cytoplasm of cells in the spiral ganglion, the organ of Corti and the stria vascularis. Reverse transcription-PCR detected high levels of XIAP mRNA expression in the cochlea. The present findings suggest that PEI-PEG nanocarrier-mediated XIAP gene transfection results in XIAP expression in the cochlea, prevents damage to cochlear spiral ganglion cells, and protects hearing.

  11. TOPOGRAPHIC ORGANIZATION AND SPECIALIZED AREAS IN THE RETINA OF Callopistes palluma: GANGLION CELL LAYER

    OpenAIRE

    Inzunza, Oscar; Barros B., Zitta; Bravo, Hermes

    1998-01-01

    In this paper we analyze the topographic distribution and cell body size of neurons (ganglion and displaced amacrine) of layer 8 of the retina in the chilean reptile Callopistes palluma; using whole mount retinaswith nissl stain. Callopistes palluma retina has an area centralis without fovea in which the ganglion cell density amounts 20.000 cells / µm2 while the displaced amacrine neurons is about 7.000 cells / µm2. This neural density decreased gradually towards the peripheral retina. A hor...

  12. THE NISSL SUBSTANCE OF LIVING AND FIXED SPINAL GANGLION CELLS

    Science.gov (United States)

    Deitch, Arline D.; Moses, Montrose J.

    1957-01-01

    Living chick spinal ganglion neurons grown for 19 to 25 days in vitro were photographed with a color-translating ultraviolet microscope (UV-91) at 265, 287, and 310 mµ. This instrument was unique in permitting rapid accumulation of ultraviolet information with minimal damage to the cell. In the photographs taken at 265 mµ of the living neurons, discrete ultraviolet-absorbing cytoplasmic masses were observed which were found to be virtually unchanged in appearance after formalin fixation. These were identical with the Nissl bodies of the same cells seen after staining with basic dyes. The correlation of ultraviolet absorption, ribonuclease extraction, and staining experiments with acid and basic dyes confirmed the ribonucleoprotein nature of these Nissl bodies in the living and fixed cells. No change in distribution or concentration of ultraviolet-absorbing substance was observed in the first 12 ultraviolet photographs of a neuron, and it is concluded that the cells had not been subjected to significant ultraviolet damage during the period of photography. On the basis of these observations, as well as previous findings with phase contrast microscopy, it is concluded that Nissl bodies preexist in the living neuron as discrete aggregates containing high concentrations of nucleoprotein. PMID:13438929

  13. Melanopsin-expressing retinal ganglion cells are resistant to cell injury, but not always

    DEFF Research Database (Denmark)

    Georg, Birgitte; Ghelli, Anna; Giordano, Carla

    2017-01-01

    Melanopsin retinal ganglion cells (mRGCs) are intrinsically photosensitive RGCs deputed to non-image forming functions of the eye such as synchronization of circadian rhythms to light-dark cycle. These cells are characterized by unique electrophysiological, anatomical and biochemical properties...

  14. Cat retinal ganglion cell receptive-field alterations after 6-hydroxydopamine induced dopaminergic amacrine cell lesions

    International Nuclear Information System (INIS)

    Maguire, G.W.; Smith, E.L. III

    1985-01-01

    Optic tract single-unit recordings were used to study ganglion cell response functions of the intact cat eye after 6-hydroxydopamine (6-OHDA) lesioning of the dopaminergic amacrine cell (AC) population of the inner retina. The impairment of the dopaminergic AC was verified by high pressure-liquid chromatography with electrochemical detection of endogenous dopamine content and by [ 3 H]dopamine high-affinity uptake; the dopaminergic ACs of the treated eyes demonstrated reduced endogenous dopamine content and reduced [ 3 H]dopamine uptake compared with that of their matched controls. Normal appearing [ 3 H]GABA and [ 3 H]-glycine uptake in the treated retinas suggests the absence of any nonspecific action of the 6-OHDA on the neural retina. The impairment of the dopaminergic AC population was found to alter a number of response properties in off-center ganglion cells, but this impairment had only a modest effect on the on-center cells. An abnormally high proportion of the off-center ganglion cells in the 6-OHDA treated eyes possessed nonlinear, Y-type receptive fields. These cells also possessed shift-responses of greater than normal amplitude, altered intensity-response functions, reduced maintained activities, and more transient center responses. Of the on-center type cells, only the Y-type on-center cells were affected by 6-OHDA, possessing higher than normal maintained activities and altered intensity-response functions. The on-center X-cells were unaffected by 6-OHDA treatment. The dopaminergic AC of the photopically adapted cat retina therefore modulates a number of ganglion cell response properties and within the limits of this study is most prominent in off-center ganglion cell circuitry

  15. Trimetazidine protects retinal ganglion cells from acute glaucoma via the Nrf2/Ho-1 pathway.

    Science.gov (United States)

    Wan, Peixing; Su, Wenru; Zhang, Yingying; Li, Zhidong; Deng, Caibin; Zhuo, Yehong

    2017-09-15

    Acute glaucoma is one of the leading causes of irreversible vision impairment characterized by the rapid elevation of intraocular pressure (IOP) and consequent retinal ganglion cell (RGC) death. Oxidative stress and neuroinflammation have been considered critical for the pathogenesis of RGC death in acute glaucoma. Trimetazidine (TMZ), an anti-ischemic drug, possesses antioxidative and anti-inflammatory properties, contributing to its therapeutic potential in tissue damage. However, the role of TMZ in acute glaucoma and the underlying molecular mechanisms remain elusive. Here, we report that treatment with TMZ significantly attenuated retinal damage and RGC death in mice with acute glaucoma, with a significant decrease in reactive oxygen species (ROS) and inflammatory cytokine production in the retina. Furthermore, TMZ treatment directly decreased ROS production and rebalanced the intracellular redox state, thus contributing to the survival of RGCs in vitro TMZ treatment also reduced the production of inflammatory cytokines in vitro Mechanistically, the TMZ-mediated inhibition of apoptosis and inflammatory cytokine production in RGCs occurred via the regulation of the nuclear factor erythroid 2-related factor 2/heme oxygenase 1/caspase-8 pathway. Moreover, the TMZ-mediated neuroprotection in acute glaucoma was abrogated when an HO-1 inhibitor, SnPP, was used. Our findings identify potential mechanisms of RGC apoptosis and propose a novel therapeutic agent, TMZ, which exerts a precise neuroprotective effect against acute glaucoma. © 2017 The Author(s).

  16. Retinal Ganglion Cell Distribution and Spatial Resolving Power in Deep-Sea Lanternfishes (Myctophidae)

    KAUST Repository

    De Busserolles, Fanny

    2014-01-01

    Topographic analyses of retinal ganglion cell density are very useful in providing information about the visual ecology of a species by identifying areas of acute vision within the visual field (i.e. areas of high cell density). In this study, we investigated the neural cell distribution in the ganglion cell layer of a range of lanternfish species belonging to 10 genera. Analyses were performed on wholemounted retinas using stereology. Topographic maps were constructed of the distribution of all neurons and both ganglion and amacrine cell populations in 5 different species from Nissl-stained retinas using cytological criteria. Amacrine cell distribution was also examined immunohistochemically in 2 of the 5 species using anti-parvalbumin antibody. The distributions of both the total neuron and the amacrine cell populations were aligned in all of the species examined, showing a general increase in cell density toward the retinal periphery. However, when the ganglion cell population was topographically isolated from the amacrine cell population, which comprised up to 80% of the total neurons within the ganglion cell layer, a different distribution was revealed. Topographic maps of the true ganglion cell distribution in 18 species of lanternfishes revealed well-defined specializations in different regions of the retina. Different species possessed distinct areas of high ganglion cell density with respect to both peak density and the location and/or shape of the specialized acute zone (i.e. elongated areae ventro-temporales, areae temporales and large areae centrales). The spatial resolving power was calculated to be relatively low (varying from 1.6 to 4.4 cycles per degree), indicating that myctophids may constitute one of the less visually acute groups of deep-sea teleosts. The diversity in retinal specializations and spatial resolving power within the family is assessed in terms of possible ecological functions and evolutionary history.

  17. The molecular basis of retinal ganglion cell death in glaucoma.

    Science.gov (United States)

    Almasieh, Mohammadali; Wilson, Ariel M; Morquette, Barbara; Cueva Vargas, Jorge Luis; Di Polo, Adriana

    2012-03-01

    Glaucoma is a group of diseases characterized by progressive optic nerve degeneration that results in visual field loss and irreversible blindness. A crucial element in the pathophysiology of all forms of glaucoma is the death of retinal ganglion cells (RGCs), a population of CNS neurons with their soma in the inner retina and axons in the optic nerve. Strategies that delay or halt RGC loss have been recognized as potentially beneficial to preserve vision in glaucoma; however, the success of these approaches depends on an in-depth understanding of the mechanisms that lead to RGC dysfunction and death. In recent years, there has been an exponential increase in valuable information regarding the molecular basis of RGC death stemming from animal models of acute and chronic optic nerve injury as well as experimental glaucoma. The emerging landscape is complex and points at a variety of molecular signals - acting alone or in cooperation - to promote RGC death. These include: axonal transport failure, neurotrophic factor deprivation, toxic pro-neurotrophins, activation of intrinsic and extrinsic apoptotic signals, mitochondrial dysfunction, excitotoxic damage, oxidative stress, misbehaving reactive glia and loss of synaptic connectivity. Collectively, this body of work has considerably updated and expanded our view of how RGCs might die in glaucoma and has revealed novel, potential targets for neuroprotection. Copyright © 2011. Published by Elsevier Ltd.

  18. Spatial consequences of bleaching adaptation in cat retinal ganglion cells.

    Science.gov (United States)

    Bonds, A B; Enroth-Cugell, C

    1981-01-01

    1. Experiments were conducted to study the effects of localized bleaching on the centre responses of rod-driven cat retinal ganglion cells. 2. Stimulation as far as 2 degrees from the bleaching site yielded responses which were reduced nearly as much as those generated at the bleaching site. Bleaching in the receptive field middle reduced responsiveness at a site 1 degrees peripheral more than bleaching at that peripheral site itself. 3. The effectiveness of a bleach in reducing centre responsiveness is related to the sensitivity of the region in which the bleach is applied. 4. Response reduction after a 0.2 degree bleach followed the same temporal pattern for concentric test spots of from 0.2 to 1.8 degrees in diameter, implying a substantially uniform spread of adaptation within these bounds. 5. A linear trade-off between fraction of rhodopsin and area bleached over a range of 8:1 yields the same pattern of response reduction, implying that the non-linear nature of bleaching adaptation is a property of the adaptation pool rather than independent photoreceptors. PMID:7320894

  19. Spatial distribution of excitatory synapses on the dendrites of ganglion cells in the mouse retina.

    Directory of Open Access Journals (Sweden)

    Yin-Peng Chen

    Full Text Available Excitatory glutamatergic inputs from bipolar cells affect the physiological properties of ganglion cells in the mammalian retina. The spatial distribution of these excitatory synapses on the dendrites of retinal ganglion cells thus may shape their distinct functions. To visualize the spatial pattern of excitatory glutamatergic input into the ganglion cells in the mouse retina, particle-mediated gene transfer of plasmids expressing postsynaptic density 95-green fluorescent fusion protein (PSD95-GFP was used to label the excitatory synapses. Despite wide variation in the size and morphology of the retinal ganglion cells, the expression of PSD95 puncta was found to follow two general rules. Firstly, the PSD95 puncta are regularly spaced, at 1-2 µm intervals, along the dendrites, whereby the presence of an excitatory synapse creates an exclusion zone that rules out the presence of other glutamatergic synaptic inputs. Secondly, the spatial distribution of PSD95 puncta on the dendrites of diverse retinal ganglion cells are similar in that the number of excitatory synapses appears to be less on primary dendrites and to increase to a plateau on higher branch order dendrites. These observations suggest that synaptogenesis is spatially regulated along the dendritic segments and that the number of synaptic contacts is relatively constant beyond the primary dendrites. Interestingly, we also found that the linear puncta density is slightly higher in large cells than in small cells. This may suggest that retinal ganglion cells with a large dendritic field tend to show an increased connectivity of excitatory synapses that makes up for their reduced dendrite density. Mapping the spatial distribution pattern of the excitatory synapses on retinal ganglion cells thus provides explicit structural information that is essential for our understanding of how excitatory glutamatergic inputs shape neuronal responses.

  20. In vivo fluorescence imaging of primate retinal ganglion cells and retinal pigment epithelial cells

    Science.gov (United States)

    Gray, Daniel C.; Merigan, William; Wolfing, Jessica I.; Gee, Bernard P.; Porter, Jason; Dubra, Alfredo; Twietmeyer, Ted H.; Ahamd, Kamran; Tumbar, Remy; Reinholz, Fred; Williams, David R.

    2006-08-01

    The ability to resolve single cells noninvasively in the living retina has important applications for the study of normal retina, diseased retina, and the efficacy of therapies for retinal disease. We describe a new instrument for high-resolution, in vivo imaging of the mammalian retina that combines the benefits of confocal detection, adaptive optics, multispectral, and fluorescence imaging. The instrument is capable of imaging single ganglion cells and their axons through retrograde transport in ganglion cells of fluorescent dyes injected into the monkey lateral geniculate nucleus (LGN). In addition, we demonstrate a method involving simultaneous imaging in two spectral bands that allows the integration of very weak signals across many frames despite inter-frame movement of the eye. With this method, we are also able to resolve the smallest retinal capillaries in fluorescein angiography and the mosaic of retinal pigment epithelium (RPE) cells with lipofuscin autofluorescence.

  1. Taurine Provides Neuroprotection against Retinal Ganglion Cell Degeneration

    Science.gov (United States)

    Froger, Nicolas; Cadetti, Lucia; Lorach, Henri; Martins, Joao; Bemelmans, Alexis-Pierre; Dubus, Elisabeth; Degardin, Julie; Pain, Dorothée; Forster, Valérie; Chicaud, Laurent; Ivkovic, Ivana; Simonutti, Manuel; Fouquet, Stéphane; Jammoul, Firas; Léveillard, Thierry; Benosman, Ryad; Sahel, José-Alain; Picaud, Serge

    2012-01-01

    Retinal ganglion cell (RGC) degeneration occurs in numerous retinal diseases leading to blindness, either as a primary process like in glaucoma, or secondary to photoreceptor loss. However, no commercial drug is yet directly targeting RGCs for their neuroprotection. In the 70s, taurine, a small sulfonic acid provided by nutrition, was found to be essential for the survival of photoreceptors, but this dependence was not related to any retinal disease. More recently, taurine deprivation was incriminated in the retinal toxicity of an antiepileptic drug. We demonstrate here that taurine can improve RGC survival in culture or in different animal models of RGC degeneration. Taurine effect on RGC survival was assessed in vitro on primary pure RCG cultures under serum-deprivation conditions, and on NMDA-treated retinal explants from adult rats. In vivo, taurine was administered through the drinking water in two glaucomatous animal models (DBA/2J mice and rats with vein occlusion) and in a model of Retinitis pigmentosa with secondary RGC degeneration (P23H rats). After a 6-day incubation, 1 mM taurine significantly enhanced RGCs survival (+68%), whereas control RGCs were cultured in a taurine-free medium, containing all natural amino-acids. This effect was found to rely on taurine-uptake by RGCs. Furthermore taurine (1 mM) partly prevented NMDA-induced RGC excitotoxicity. Finally, taurine supplementation increased RGC densities both in DBA/2J mice, in rats with vein occlusion and in P23H rats by contrast to controls drinking taurine-free water. This study indicates that enriched taurine nutrition can directly promote RGC survival through RGC intracellular pathways. It provides evidence that taurine can positively interfere with retinal degenerative diseases. PMID:23115615

  2. Ganglion cell complex scan in the early prediction of glaucoma.

    Science.gov (United States)

    Ganekal, S

    2012-01-01

    To compare the macular ganglion cell complex (GCC) with peripapillary retinal fiber layer (RNFL) thickness map in glaucoma suspects and patients. Forty participants (20 glaucoma suspects and 20 glaucoma patients) were enrolled. Macular GCC and RNFL thickness maps were performed in both eyes of each participant in the same visit. The sensitivity and specificity of a color code less than 5% (red or yellow) for glaucoma diagnosis were calculated. Standard Automated Perimetry was performed with the Octopus 3.1.1 Dynamic 24-2 program. The statistical analysis was performed with the SPSS 10.1 (SPSS Inc. Chicago, IL, EUA). Results were expressed as mean +/- standard deviation and a p value of 0.05 or less was considered significant. Provide absolute numbers of these findings with their units of measurement. There was a statistically significant difference in average RNFL thickness (p=0.004), superior RNFL thickness (p=0.006), inferior RNFL thickness (p=0.0005) and average GCC (p=0.03) between the suspects and glaucoma patients. There was no difference in optic disc area (p=0.35) and vertical cup/disc ratio (p=0.234) in both groups. While 38% eyes had an abnormal GCC and 13% had an abnormal RNFL thickness in the glaucoma suspect group, 98% had an abnormal GCC and 90% had an abnormal RNFL thickness in the glaucoma group. The ability to diagnose glaucoma with macular GCC thickness is comparable to that with peripapillary RNFL thickness . Macular GCC thickness measurements may be a good alternative or a complementary measurement to RNFL thickness assessment in the clinical evaluation of glaucoma. © NEPjOPH.

  3. Neuroprotective effect of He-Ying-Qing-Re formula on retinal ganglion cell in diabetic retinopathy.

    Science.gov (United States)

    Zhang, Cheng; Xu, Yu; Tan, Hor-Yue; Li, Sha; Wang, Ning; Zhang, Yinjian; Feng, Yibin

    2018-03-25

    He-Ying-Qing-Re Formula (HF) was empirically modified from Si-Miao-Yong-An Decoction (SD), which was recorded in the literature of Divine Doctor's Secret Transmission, and has been utilized for centuries to treat vasculopathy through clearing heat and accelerating bloodstream. HF has been used as an effective holistic treatment of diabetic retinopathy (DR) for decades and experimentally reported to ameliorate retinal condition in diabetic mice. Our study aims to investigate the effect of HF in preventing sustained hyperglycemia and hyperlipidemia-associated retinal ganglion cell (RGC) cell death and its possible mechanism. Chromatographic fingerprint of HF was obtained upon the UPLC-based analytic system; Diabetic retinopathy was established in streptozotocin (STZ) injection-induced hyperglycemic mice; Alterations of retinal structure was assayed by H&E staining. Expression of PSD-95 and CHOP in retinae was assessed by immunofluorescence; RGC cell line (mRGC) was used for in vitro study. Cell death was analyzed by flow cytometry; Intracellular reactive oxygen species (ROS) was measured by 2',7'-dichlorofluorescin diacetate (DCFDA); Apoptosis-related proteins and signaling were monitored with immunoblotting and colorimetric assay. Chlorogenic acid, ferulic acid, and rutin were identified in HF. HF attenuates the loss of RGCs, thinning of inner retinal layers in diabetic mice. Furthermore, expressions of Brn3a and PSD-95 were restored while CHOP level was downregulated upon HF treatment. In vitro study, HF alleviates H 2 O 2 -induced apoptosis of mRGCs and loss of postsynaptic protein via scavenging ROS and suppressing ATF4/CHOP-mediated endoplasmic reticulum stress and mitochondria-related pro-apoptotic factors, probably as cleaved-caspase-3, and phospho-p38 mitogen-activated protein kinase (MARK). Meanwhile, both pro-survival protein levels like Bcl-2/Bcl-xL and postsynaptic protein of PSD-95 were upregulated upon HF treatment. HF administration was a valid

  4. Hydrostatic pressure does not cause detectable changes in survival of human retinal ganglion cells.

    Directory of Open Access Journals (Sweden)

    Andrew Osborne

    Full Text Available Elevated intraocular pressure (IOP is a major risk factor for glaucoma. One consequence of raised IOP is that ocular tissues are subjected to increased hydrostatic pressure (HP. The effect of raised HP on stress pathway signaling and retinal ganglion cell (RGC survival in the human retina was investigated.A chamber was designed to expose cells to increased HP (constant and fluctuating. Accurate pressure control (10-100 mmHg was achieved using mass flow controllers. Human organotypic retinal cultures (HORCs from donor eyes (<24 h post mortem were cultured in serum-free DMEM/HamF12. Increased HP was compared to simulated ischemia (oxygen glucose deprivation, OGD. Cell death and apoptosis were measured by LDH and TUNEL assays, RGC marker expression by qRT-PCR (THY-1 and RGC number by immunohistochemistry (NeuN. Activated p38 and JNK were detected by Western blot.Exposure of HORCs to constant (60 mmHg or fluctuating (10-100 mmHg; 1 cycle/min pressure for 24 or 48 h caused no loss of structural integrity, LDH release, decrease in RGC marker expression (THY-1 or loss of RGCs compared with controls. In addition, there was no increase in TUNEL-positive NeuN-labelled cells at either time-point indicating no increase in apoptosis of RGCs. OGD increased apoptosis, reduced RGC marker expression and RGC number and caused elevated LDH release at 24 h. p38 and JNK phosphorylation remained unchanged in HORCs exposed to fluctuating pressure (10-100 mmHg; 1 cycle/min for 15, 30, 60 and 90 min durations, whereas OGD (3 h increased activation of p38 and JNK, remaining elevated for 90 min post-OGD.Directly applied HP had no detectable impact on RGC survival and stress-signalling in HORCs. Simulated ischemia, however, activated stress pathways and caused RGC death. These results show that direct HP does not cause degeneration of RGCs in the ex vivo human retina.

  5. Neuroprotective effect of peroxiredoxin 6 against hypoxia-induced retinal ganglion cell damage

    Directory of Open Access Journals (Sweden)

    Kumar Anil

    2010-10-01

    Full Text Available Abstract Background The ability to respond to changes in the extra-intracellular environment is prerequisite for cell survival. Cellular responses to the environment include elevating defense systems, such as the antioxidant defense system. Hypoxia-evoked reactive oxygen species (ROS-driven oxidative stress is an underlying mechanism of retinal ganglion cell (RGC death that leads to blinding disorders. The protein peroxiredoxin 6 (PRDX6 plays a pleiotropic role in negatively regulating death signaling in response to stressors, and thereby stabilizes cellular homeostasis. Results We have shown that RGCs exposed to hypoxia (1% or hypoxia mimetic cobalt chloride display reduced expression of PRDX6 with higher ROS expression and activation of NF-κB. These cells undergo apoptosis, while cells with over-expression of PRDX6 demonstrate resistance against hypoxia-driven RGC death. The RGCs exposed to hypoxia either with 1% oxygen or cobalt chloride (0-400 μM, revealed ~30%-70% apoptotic cell death after 48 and 72 h of exposure. Western analysis and real-time PCR showed elevated expression of PRDX6 during hypoxia at 24 h, while PRDX6 protein and mRNA expression declined from 48 h onwards following hypoxia exposure. Concomitant with this, RGCs showed increased ROS expression and activation of NF-κB with IkB phosphorylation/degradation, as examined with H2DCF-DA and transactivation assays. These hypoxia-induced adverse reactions could be reversed by over-expression of PRDX6. Conclusion Because an abundance of PRDX6 in cells was able to attenuate hypoxia-induced RGC death, the protein could possibly be developed as a novel therapeutic agent acting to postpone RGC injury and delay the progression of glaucoma and other disorders caused by the increased-ROS-generated death signaling related to hypoxia.

  6. Expression of EFR3A in the mouse cochlea during degeneration of spiral ganglion following hair cell loss.

    Directory of Open Access Journals (Sweden)

    Chen Nie

    Full Text Available Retrograde degeneration of spiral ganglion cells in the cochlea following hair cell loss is similar to dying back in pathology. The EFR3A gene has recently been discovered to be involved in the pathogenesis of dying back. The relationship of EFR3A and spiral ganglion degeneration, however, was rarely investigated. In this study, we destroyed the hair cells of the mouse cochlea by co-administration of kanamycin and furosemide and then investigated the EFR3A expression during the induced spiral ganglion cell degeneration. Our results revealed that co-administration of kanamycin and furosemide quickly induced hair cell loss in the C57BL/6J mice and then resulted in progressive degeneration of the spiral ganglion beginning at day 5 following drug administration. The number of the spiral ganglion cells began to decrease at day 15. The expression of EFR3A increased remarkably in the spiral ganglion at day 5 and then decreased to near normal level within the next 10 days. Our study suggested that the change of EFR3A expression in the spiral ganglion was coincident with the time of the spiral ganglion degeneration, which implied that high expression of EFR3A may be important to prompt initiation of spiral ganglion degeneration following hair cell loss.

  7. Melanopsin retinal ganglion cells are resistant to neurodegeneration in mitochondrial optic neuropathies

    DEFF Research Database (Denmark)

    La Morgia, C; Ross-Cisneros, F.N.; Sadun, A.A.

    2010-01-01

    Mitochondrial optic neuropathies, that is, Leber hereditary optic neuropathy and dominant optic atrophy, selectively affect retinal ganglion cells, causing visual loss with relatively preserved pupillary light reflex. The mammalian eye contains a light detection system based on a subset of retinal...... ganglion cells containing the photopigment melanopsin. These cells give origin to the retinohypothalamic tract and support the non-image-forming visual functions of the eye, which include the photoentrainment of circadian rhythms, light-induced suppression of melatonin secretion and pupillary light reflex...... subjects as in controls, indicating that the retinohypothalamic tract is sufficiently preserved to drive light information detected by melanopsin retinal ganglion cells. We then investigated the histology of post-mortem eyes from two patients with Leber hereditary optic neuropathy and one case...

  8. Electromagnetic radiation (Wi-Fi) and epilepsy induce calcium entry and apoptosis through activation of TRPV1 channel in hippocampus and dorsal root ganglion of rats.

    Science.gov (United States)

    Ghazizadeh, Vahid; Nazıroğlu, Mustafa

    2014-09-01

    Incidence rates of epilepsy and use of Wi-Fi worldwide have been increasing. TRPV1 is a Ca(2+) permeable and non-selective channel, gated by noxious heat, oxidative stress and capsaicin (CAP). The hyperthermia and oxidant effects of Wi-Fi may induce apoptosis and Ca(2+) entry through activation of TRPV1 channel in epilepsy. Therefore, we tested the effects of Wi-Fi (2.45 GHz) exposure on Ca(2+) influx, oxidative stress and apoptosis through TRPV1 channel in the murine dorsal root ganglion (DRG) and hippocampus of pentylentetrazol (PTZ)-induced epileptic rats. Rats in the present study were divided into two groups as controls and PTZ. The PTZ groups were divided into two subgroups namely PTZ + Wi-Fi and PTZ + Wi-Fi + capsazepine (CPZ). The hippocampal and DRG neurons were freshly isolated from the rats. The DRG and hippocampus in PTZ + Wi-Fi and PTZ + Wi-Fi + CPZ groups were exposed to Wi-Fi for 1 hour before CAP stimulation. The cytosolic free Ca(2+), reactive oxygen species production, apoptosis, mitochondrial membrane depolarization, caspase-3 and -9 values in hippocampus were higher in the PTZ group than in the control although cell viability values decreased. The Wi-Fi exposure induced additional effects on the cytosolic Ca(2+) increase. However, pretreatment of the neurons with CPZ, results in a protection against epilepsy-induced Ca(2+) influx, apoptosis and oxidative damages. In results of whole cell patch-clamp experiments, treatment of DRG with Ca(2+) channel antagonists [thapsigargin, verapamil + diltiazem, 2-APB, MK-801] indicated that Wi-Fi exposure induced Ca(2+) influx via the TRPV1 channels. In conclusion, epilepsy and Wi-Fi in our experimental model is involved in Ca(2+) influx and oxidative stress-induced hippocampal and DRG death through activation of TRPV1 channels, and negative modulation of this channel activity by CPZ pretreatment may account for the neuroprotective activity against oxidative stress.

  9. Rac1 selective activation improves retina ganglion cell survival and regeneration.

    Directory of Open Access Journals (Sweden)

    Erika Lorenzetto

    Full Text Available In adult mammals, after optic nerve injury, retinal ganglion cells (RGCs do not regenerate their axons and most of them die by apoptosis within a few days. Recently, several strategies that activate neuronal intracellular pathways were proposed to prevent such degenerative processes. The rho-related small GTPase Rac1 is part of a complex, still not fully understood, intracellular signaling network, mediating in neurons many effects, including axon growth and cell survival. However, its role in neuronal survival and regeneration in vivo has not yet been properly investigated. To address this point we intravitreally injected selective cell-penetrating Rac1 mutants after optic nerve crush and studied the effect on RGC survival and axonal regeneration. We injected two well-characterized L61 constitutively active Tat-Rac1 fusion protein mutants, in which a second F37A or Y40C mutation confers selectivity in downstream signaling pathways. Results showed that, 15 days after crush, both mutants were able to improve survival and to prevent dendrite degeneration, while the one harboring the F37A mutation also improved axonal regeneration. The treatment with F37A mutant for one month did not improve the axonal elongation respect to 15 days. Furthermore, we found an increase of Pak1 T212 phosphorylation and ERK1/2 expression in RGCs after F37A treatment, whereas ERK1/2 was more activated in glial cells after Y40C administration. Our data suggest that the selective activation of distinct Rac1-dependent pathways could represent a therapeutic strategy to counteract neuronal degenerative processes in the retina.

  10. Retinal Ganglion Cell Distribution and Spatial Resolving Power in Deep-Sea Lanternfishes (Myctophidae)

    KAUST Repository

    De Busserolles, Fanny; Marshall, N. Justin; Collin, Shaun P.

    2014-01-01

    Topographic analyses of retinal ganglion cell density are very useful in providing information about the visual ecology of a species by identifying areas of acute vision within the visual field (i.e. areas of high cell density). In this study, we

  11. Retinal Ganglion Cell Diversity and Subtype Specification from Human Pluripotent Stem Cells

    Directory of Open Access Journals (Sweden)

    Kirstin B. Langer

    2018-04-01

    Full Text Available Summary: Retinal ganglion cells (RGCs are the projection neurons of the retina and transmit visual information to postsynaptic targets in the brain. While this function is shared among nearly all RGCs, this class of cell is remarkably diverse, comprised of multiple subtypes. Previous efforts have identified numerous RGC subtypes in animal models, but less attention has been paid to human RGCs. Thus, efforts of this study examined the diversity of RGCs differentiated from human pluripotent stem cells (hPSCs and characterized defined subtypes through the expression of subtype-specific markers. Further investigation of these subtypes was achieved using single-cell transcriptomics, confirming the combinatorial expression of molecular markers associated with these subtypes, and also provided insight into more subtype-specific markers. Thus, the results of this study describe the derivation of RGC subtypes from hPSCs and will support the future exploration of phenotypic and functional diversity within human RGCs. : In this article, Langer and colleagues present extensive characterization of RGC subtypes derived from human pluripotent stem cells, with multiple subtypes identified by subtype-specific molecular markers. Their results present a more detailed analysis of RGC diversity in human cells and yield the use of different markers to identify RGC subtypes. Keywords: iPSC, retina, retinal ganglion cell, RGC subtype, stem cell, ipRGC, alpha RGC, direction selective RGC, RNA-seq

  12. Visual Field Defects and Retinal Ganglion Cell Losses in Human Glaucoma Patients

    Science.gov (United States)

    Harwerth, Ronald S.; Quigley, Harry A.

    2007-01-01

    Objective The depth of visual field defects are correlated with retinal ganglion cell densities in experimental glaucoma. This study was to determine whether a similar structure-function relationship holds for human glaucoma. Methods The study was based on retinal ganglion cell densities and visual thresholds of patients with documented glaucoma (Kerrigan-Baumrind, et al.) The data were analyzed by a model that predicted ganglion cell densities from standard clinical perimetry, which were then compared to histologic cell counts. Results The model, without free parameters, produced accurate and relatively precise quantification of ganglion cell densities associated with visual field defects. For 437 sets of data, the unity correlation for predicted vs. measured cell densities had a coefficient of determination of 0.39. The mean absolute deviation of the predicted vs. measured values was 2.59 dB, the mean and SD of the distribution of residual errors of prediction was -0.26 ± 3.22 dB. Conclusions Visual field defects by standard clinical perimetry are proportional to neural losses caused by glaucoma. Clinical Relevance The evidence for quantitative structure-function relationships provides a scientific basis of interpreting glaucomatous neuropathy from visual thresholds and supports the application of standard perimetry to establish the stage of the disease. PMID:16769839

  13. REDUCED GANGLION CELL VOLUME ON OPTICAL COHERENCE TOMOGRAPHY IN PATIENTS WITH GEOGRAPHIC ATROPHY.

    Science.gov (United States)

    Ramkumar, Hema L; Nguyen, Brian; Bartsch, Dirk-Uwe; Saunders, Luke J; Muftuoglu, Ilkay Kilic; You, Qisheng; Freeman, William R

    2017-11-07

    Geographic atrophy (GA) is the sequelae of macular degeneration. Automated inner retinal analysis using optical coherence tomography is flawed because segmentation software is calibrated for normal eyes. The purpose of this study is to determine whether ganglion cell layer (GCL) volume is reduced in GA using manual analysis. Nineteen eyes with subfoveal GA and 22 controls were selected for morphometric analyses. Heidelberg scanning laser ophthalmoscope optical coherence tomography images of the optic nerve and macula were obtained, and the Viewing Module was used to manually calibrate retinal layer segmentation. Retinal layer volumes in the central 3-mm and surrounding 6-mm diameter were measured. Linear mixed models were used for statistics. The GCL volume in the central 3 mm of the macula is less (P = 0.003), and the retinal nerve fiber layer volume is more (P = 0.02) in patients with GA when compared with controls. Ganglion cell layer volume positively correlated with outer nuclear layer volume (P = 0.020). The patients with geographic atrophy have a small significant loss of the GCL. Ganglion cell death may precede axonal loss, and increased macular retinal nerve fiber layer volumes are not indicative of GCL volume. Residual ganglion cell stimulation by interneurons may enable vision in patients with GA.

  14. Loss of Melanopsin-Expressing Retinal Ganglion Cells in Patients With Diabetic Retinopathy

    DEFF Research Database (Denmark)

    Obara, Elisabeth Anne; Hannibal, Jens; Heegaard, Steffen

    2017-01-01

    Purpose: Photo-entrainment of the circadian clock is mediated by melanopsin-expressing retinal ganglion cells (mRGCs) located in the retina. Patients suffering from diabetic retinopathy (DR) show impairment of light regulated circadian activity such as sleep disorders, altered blood pressure...

  15. Mitochondrial disfunction and apoptosis in leukemia cells

    Directory of Open Access Journals (Sweden)

    Annamaria PALLAG

    2008-05-01

    Full Text Available Apoptosis or programmed cell death is a process which involves the intentional degradation of the cell from the inside, the participation of the mitochondria to propagate the apoptotic signal, the alteration of the phospholipid cell membrane composition, the perturbation and alteration of the cell metabolism.The antineoplastic drugs is inducing the apoptotic process in the sensitive cells.It have been studied acute lymphoblastic leukemia cells. Using Annexin V-PE Apoptosis Detection Kit and flow cytometer, the amount of cells undergoing apoptosis, in various stages of the antineoplasic treatment, was detected. At the same time, were monitored, the serum level of malondialdehyde. The results obtained confirm the alteration of the mitochondrial metabolism. We can observed the mitochondrial dysfunction role in cell apoptosis.

  16. Melanopsin-expressing retinal ganglion cells are resistant to cell injury, but not always.

    Science.gov (United States)

    Georg, Birgitte; Ghelli, Anna; Giordano, Carla; Ross-Cisneros, Fred N; Sadun, Alfredo A; Carelli, Valerio; Hannibal, Jens; La Morgia, Chiara

    2017-09-01

    Melanopsin retinal ganglion cells (mRGCs) are intrinsically photosensitive RGCs deputed to non-image forming functions of the eye such as synchronization of circadian rhythms to light-dark cycle. These cells are characterized by unique electrophysiological, anatomical and biochemical properties and are usually more resistant than conventional RGCs to different insults, such as axotomy and different paradigms of stress. We also demonstrated that these cells are relatively spared compared to conventional RGCs in mitochondrial optic neuropathies (Leber's hereditary optic neuropathy and Dominant Optic Atrophy). However, these cells are affected in other neurodegenerative conditions, such as glaucoma and Alzheimer's disease. We here review the current evidences that may underlie this dichotomy. We also present our unpublished data on cell experiments demonstrating that melanopsin itself does not explain the robustness of these cells and some preliminary data on immunohistochemical assessment of mitochondria in mRGCs. Copyright © 2017 Elsevier B.V. and Mitochondria Research Society. All rights reserved.

  17. Relationship between macular ganglion cell complex thickness and macular outer retinal thickness: a spectral-domain optical coherence tomography study.

    Science.gov (United States)

    Kita, Yoshiyuki; Kita, Ritsuko; Takeyama, Asuka; Anraku, Ayako; Tomita, Goji; Goldberg, Ivan

    2013-01-01

    To assess the relationship between macular ganglion cell complex and macular outer retinal thicknesses. Case-control study. Forty-two normal eyes and 91 eyes with primary open-angle glaucoma were studied. Spectral-domain optical coherence tomography (RTVue-100) was used to measure the macular ganglion cell complex and macular outer retinal thickness. Ganglion cell complex to outer retinal thickness ratio was also calculated. The relationships between the ganglion cell complex and outer retinal thicknesses and between the ganglion cell complex to outer retinal thickness ratio and outer retinal thickness were evaluated. There was a positive correlation between ganglion cell complex and outer retinal thicknesses in the normal group and the glaucoma group (r = 0.53, P variation in the outer retinal thickness. Therefore, when determining the ganglion cell complex, it seems necessary to consider the outer retinal thickness as well. We propose the ratio as a suitable parameter to account for individual variations in outer retinal thickness. © 2013 The Authors. Clinical and Experimental Ophthalmology © 2013 Royal Australian and New Zealand College of Ophthalmologists.

  18. Eliminating Glutamatergic Input onto Horizontal Cells Changes the Dynamic Range and Receptive Field Organization of Mouse Retinal Ganglion Cells.

    Science.gov (United States)

    Ströh, Sebastian; Puller, Christian; Swirski, Sebastian; Hölzel, Maj-Britt; van der Linde, Lea I S; Segelken, Jasmin; Schultz, Konrad; Block, Christoph; Monyer, Hannah; Willecke, Klaus; Weiler, Reto; Greschner, Martin; Janssen-Bienhold, Ulrike; Dedek, Karin

    2018-02-21

    In the mammalian retina, horizontal cells receive glutamatergic inputs from many rod and cone photoreceptors and return feedback signals to them, thereby changing photoreceptor glutamate release in a light-dependent manner. Horizontal cells also provide feedforward signals to bipolar cells. It is unclear, however, how horizontal cell signals also affect the temporal, spatial, and contrast tuning in retinal output neurons, the ganglion cells. To study this, we generated a genetically modified mouse line in which we eliminated the light dependency of feedback by deleting glutamate receptors from mouse horizontal cells. This genetic modification allowed us to investigate the impact of horizontal cells on ganglion cell signaling independent of the actual mode of feedback in the outer retina and without pharmacological manipulation of signal transmission. In control and genetically modified mice (both sexes), we recorded the light responses of transient OFF-α retinal ganglion cells in the intact retina. Excitatory postsynaptic currents (EPSCs) were reduced and the cells were tuned to lower temporal frequencies and higher contrasts, presumably because photoreceptor output was attenuated. Moreover, receptive fields of recorded cells showed a significantly altered surround structure. Our data thus suggest that horizontal cells are responsible for adjusting the dynamic range of retinal ganglion cells and, together with amacrine cells, contribute to the center/surround organization of ganglion cell receptive fields in the mouse. SIGNIFICANCE STATEMENT Horizontal cells represent a major neuronal class in the mammalian retina and provide lateral feedback and feedforward signals to photoreceptors and bipolar cells, respectively. The mode of signal transmission remains controversial and, moreover, the contribution of horizontal cells to visual processing is still elusive. To address the question of how horizontal cells affect retinal output signals, we recorded the light

  19. A Learning Model for L/M Specificity in Ganglion Cells

    Science.gov (United States)

    Ahumada, Albert J.

    2016-01-01

    An unsupervised learning model for developing LM specific wiring at the ganglion cell level would support the research indicating LM specific wiring at the ganglion cell level (Reid and Shapley, 2002). Removing the contributions to the surround from cells of the same cone type improves the signal-to-noise ratio of the chromatic signals. The unsupervised learning model used is Hebbian associative learning, which strengthens the surround input connections according to the correlation of the output with the input. Since the surround units of the same cone type as the center are redundant with the center, their weights end up disappearing. This process can be thought of as a general mechanism for eliminating unnecessary cells in the nervous system.

  20. Density, proportion, and dendritic coverage of retinal ganglion cells of the common marmoset (Callithrix jacchus jacchus

    Directory of Open Access Journals (Sweden)

    F.L. Gomes

    2005-06-01

    Full Text Available We performed a quantitative analysis of M and P cell mosaics of the common-marmoset retina. Ganglion cells were labeled retrogradely from optic nerve deposits of Biocytin. The labeling was visualized using horseradish peroxidase (HRP histochemistry and 3-3'diaminobenzidine as chromogen. M and P cells were morphologically similar to those found in Old- and New-World primates. Measurements were performed on well-stained cells from 4 retinas of different animals. We analyzed separate mosaics for inner and outer M and P cells at increasing distances from the fovea (2.5-9 mm of eccentricity to estimate cell density, proportion, and dendritic coverage. M cell density decreased towards the retinal periphery in all quadrants. M cell density was higher in the nasal quadrant than in other retinal regions at similar eccentricities, reaching about 740 cells/mm² at 2.5 mm of temporal eccentricity, and representing 8-14% of all ganglion cells. P cell density increased from peripheral to more central regions, reaching about 5540 cells/mm² at 2.5 mm of temporal eccentricity. P cells represented a smaller proportion of all ganglion cells in the nasal quadrant than in other quadrants, and their numbers increased towards central retinal regions. The M cell coverage factor ranged from 5 to 12 and the P cell coverage factor ranged from 1 to 3 in the nasal quadrant and from 5 to 12 in the other quadrants. These results show that central and peripheral retinal regions differ in terms of cell class proportions and dendritic coverage, and their properties do not result from simply scaling down cell density. Therefore, differences in functional properties between central and peripheral vision should take these distinct regional retinal characteristics into account.

  1. Delayed rectifier K channels contribute to contrast adaptation in mammalian retinal ganglion cells

    Science.gov (United States)

    Weick, Michael; Demb, Jonathan B.

    2011-01-01

    SUMMARY Retinal ganglion cells adapt by reducing their sensitivity during periods of high contrast. Contrast adaptation in the firing response depends on both presynaptic and intrinsic mechanisms. Here, we investigated intrinsic mechanisms for contrast adaptation in OFF Alpha ganglion cells in the in vitro guinea pig retina. Using either visual stimulation or current injection, we show that brief depolarization evoked spiking and suppressed firing during subsequent depolarization. The suppression could be explained by Na channel inactivation, as shown in salamander cells. However, brief hyperpolarization in the physiological range (5–10 mV) also suppressed firing during subsequent depolarization. This suppression was sensitive selectively to blockers of delayed-rectifier K channels (KDR). Somatic membrane patches showed TEA-sensitive KDR currents with activation near −25 mV and removal of inactivation at voltages negative to Vrest. Brief periods of hyperpolarization apparently remove KDR inactivation and thereby increase the channel pool available to suppress excitability during subsequent depolarization. PMID:21745646

  2. Nanosecond laser pulse stimulation of spiral ganglion neurons and model cells.

    Science.gov (United States)

    Rettenmaier, Alexander; Lenarz, Thomas; Reuter, Günter

    2014-04-01

    Optical stimulation of the inner ear has recently attracted attention, suggesting a higher frequency resolution compared to electrical cochlear implants due to its high spatial stimulation selectivity. Although the feasibility of the effect is shown in multiple in vivo experiments, the stimulation mechanism remains open to discussion. Here we investigate in single-cell measurements the reaction of spiral ganglion neurons and model cells to irradiation with a nanosecond-pulsed laser beam over a broad wavelength range from 420 nm up to 1950 nm using the patch clamp technique. Cell reactions were wavelength- and pulse-energy-dependent but too small to elicit action potentials in the investigated spiral ganglion neurons. As the applied radiant exposure was much higher than the reported threshold for in vivo experiments in the same laser regime, we conclude that in a stimulation paradigm with nanosecond-pulses, direct neuronal stimulation is not the main cause of optical cochlea stimulation.

  3. Axonal transmission in the retina introduces a small dispersion of relative timing in the ganglion cell population response.

    Directory of Open Access Journals (Sweden)

    Günther Zeck

    Full Text Available BACKGROUND: Visual stimuli elicit action potentials in tens of different retinal ganglion cells. Each ganglion cell type responds with a different latency to a given stimulus, thus transforming the high-dimensional input into a temporal neural code. The timing of the first spikes between different retinal projection neurons cells may further change along axonal transmission. The purpose of this study is to investigate if intraretinal conduction velocity leads to a synchronization or dispersion of the population signal leaving the eye. METHODOLOGY/PRINCIPAL FINDINGS: We 'imaged' the initiation and transmission of light-evoked action potentials along individual axons in the rabbit retina at micron-scale resolution using a high-density multi-transistor array. We measured unimodal conduction velocity distributions (1.3±0.3 m/sec, mean ± SD for axonal populations at all retinal eccentricities with the exception of the central part that contains myelinated axons. The velocity variance within each piece of retina is caused by ganglion cell types that show narrower and slightly different average velocity tuning. Ganglion cells of the same type respond with similar latency to spatially homogenous stimuli and conduct with similar velocity. For ganglion cells of different type intraretinal conduction velocity and response latency to flashed stimuli are negatively correlated, indicating that differences in first spike timing increase (up to 10 msec. Similarly, the analysis of pair-wise correlated activity in response to white-noise stimuli reveals that conduction velocity and response latency are negatively correlated. CONCLUSION/SIGNIFICANCE: Intraretinal conduction does not change the relative spike timing between ganglion cells of the same type but increases spike timing differences among ganglion cells of different type. The fastest retinal ganglion cells therefore act as indicators of new stimuli for postsynaptic neurons. The intraretinal dispersion

  4. NUTRITION AND VASCULAR SUPPLY OF RETINAL GANGLION CELLS DURING HUMAN DEVELOPMENT

    Directory of Open Access Journals (Sweden)

    Paul eRutkowski

    2016-04-01

    Full Text Available Purpose. To review the roles of the different vascular beds nourishing the inner retina (retinal ganglion cells during normal development of the human eye and using our own tissue specimens to support our conclusions.Methods. An extensive search of the appropriate literature included PubMed, Google scholar, and numerous available textbooks. In addition, choroidal and retinal NADPH-diaphorase stained whole mount preparations were investigated.Results. The first critical interaction between vascular bed and retinal ganglion cell (RGC formation occurs in the 6th-8th month of gestation leading to a massive reduction of RGCs mainly in the peripheral retina. The first three years of age are characterized by an intense growth of the eyeball to near adult size. In the adult eye, the influence of the choroid on inner retinal nutrition was determined by examining the peripheral retinal watershed zones in more detail.Conclusion. This delicately balanced situation of retinal ganglion cell nutrition is described in the different regions of the eye, and a new graphic presentation is introduced to combine morphological measurements and clinical visual field data.

  5. Identification of retinal ganglion cells and their projections involved in central transmission of information about upward and downward image motion.

    Directory of Open Access Journals (Sweden)

    Keisuke Yonehara

    Full Text Available The direction of image motion is coded by direction-selective (DS ganglion cells in the retina. Particularly, the ON DS ganglion cells project their axons specifically to terminal nuclei of the accessory optic system (AOS responsible for optokinetic reflex (OKR. We recently generated a knock-in mouse in which SPIG1 (SPARC-related protein containing immunoglobulin domains 1-expressing cells are visualized with GFP, and found that retinal ganglion cells projecting to the medial terminal nucleus (MTN, the principal nucleus of the AOS, are comprised of SPIG1+ and SPIG1(- ganglion cells distributed in distinct mosaic patterns in the retina. Here we examined light responses of these two subtypes of MTN-projecting cells by targeted electrophysiological recordings. SPIG1+ and SPIG1(- ganglion cells respond preferentially to upward motion and downward motion, respectively, in the visual field. The direction selectivity of SPIG1+ ganglion cells develops normally in dark-reared mice. The MTN neurons are activated by optokinetic stimuli only of the vertical motion as shown by Fos expression analysis. Combination of genetic labeling and conventional retrograde labeling revealed that axons of SPIG1+ and SPIG1(- ganglion cells project to the MTN via different pathways. The axon terminals of the two subtypes are organized into discrete clusters in the MTN. These results suggest that information about upward and downward image motion transmitted by distinct ON DS cells is separately processed in the MTN, if not independently. Our findings provide insights into the neural mechanisms of OKR, how information about the direction of image motion is deciphered by the AOS.

  6. Coenzyme Q10 protects retinal cells from apoptosis induced by radiation in vitro and in vivo

    International Nuclear Information System (INIS)

    Lulli, M.; Witort, E.; Papucci, L.; Torre, E.; Schiavone, N.; Capaccioli, S.; Dal Monte, M.

    2012-01-01

    The key pathogenetic event of many retinopathies is apoptosis of retinal cells. Our previous studies have demonstrated that Coenzyme Q10 (CoQ10) prevents apoptosis of corneal keratocytes both in vitro and in vivo, by virtue of its ability to inhibit mitochondrial depolarization, independently of its free radical scavenger role. The aim of this study was to evaluate whether CoQ10 can protect cultured retinal cells and the retinas of rats from radiation-induced apoptosis, if instilled as eye drops in the cornea. In vitro experiments were carried out on cultured ARPE-19 or retinal ganglion cells (RGC)-5 cells pretreated with CoQ10 before eliciting apoptosis by ultraviolet (UV)- and γ-radiation, chemical hypoxia (Antimycin A) and serum starvation. Cell viability was evaluated by light microscopy and fluorescence activated cell sorting analysis. Apoptotic events were scored by time-lapse videomicroscopy. Mitochondrial permeability transition was evaluated by JC-1. The anti-apoptotic effectiveness of CoQ10 in retina was also evaluated by an in situ end-labeling assay in Wistar albino rats treated with CoQ10 eye drops prior to UV irradiation of the eye. CoQ10 substantially increased cell viability and lowered retinal cell apoptosis in response both to UV- and γ-radiation and to chemical hypoxia or serum starvation by inhibiting mitochondrion depolarization. In the rat, CoQ10, even when applied as eye drops on the cornea, protected all retina layers from ultraviolet radiation (UVR)-induced apoptosis. The ability of CoQ10 to protect retinal cells from radiation-induced apoptosis following its instillation on the cornea suggests the possibility for CoQ10 eye drops to become a future therapeutic countermeasure for radiation-induced retinal lesions. (author)

  7. Spatially and Temporally Regulated NRF2 Gene Therapy Using Mcp-1 Promoter in Retinal Ganglion Cell Injury

    Directory of Open Access Journals (Sweden)

    Kosuke Fujita

    2017-06-01

    Full Text Available Retinal ganglion cell degeneration triggered by axonal injury is believed to underlie many ocular diseases, including glaucoma and optic neuritis. In these diseases, retinal ganglion cells are affected unevenly, both spatially and temporally, such that healthy and unhealthy cells coexist in different patterns at different time points. Herein, we describe a temporally and spatially regulated adeno-associated virus gene therapy aiming to reduce undesired off-target effects on healthy retinal neurons. The Mcp-1 promoter previously shown to be activated in stressed retinal ganglion cells following murine optic nerve injury was combined with the neuroprotective intracellular transcription factor Nrf2. In this model, Mcp-1 promoter-driven NRF2 expression targeting only stressed retinal ganglion cells showed efficacy equivalent to non-selective cytomegalovirus promoter-driven therapy for preventing cell death. However, cytomegalovirus promoter-mediated NRF2 transcription induced cellular stress responses and death of Brn3A-positive uninjured retinal ganglion cells. Such undesired effects were reduced substantially by adopting the Mcp-1 promoter. Combining a stress-responsive promoter and intracellular therapeutic gene is a versatile approach for specifically targeting cells at risk of degeneration. This strategy may be applicable to numerous chronic ocular and non-ocular conditions.

  8. Effect of duration and severity of migraine on retinal nerve fiber layer, ganglion cell layer, and choroidal thickness.

    Science.gov (United States)

    Abdellatif, Mona K; Fouad, Mohamed M

    2018-03-01

    To investigate the factors in migraine that have the highest significance on retinal and choroidal layers' thickness. Ninety patients with migraine and 40 age-matched healthy participants were enrolled in this observational, cross-sectional study. After full ophthalmological examination, spectral domain-optical coherence tomography was done for all patients measuring the thickness of ganglion cell layer and retinal nerve fiber layer. Enhanced depth imaging technique was used to measure the choroidal thickness. There was significant thinning in the superior and inferior ganglion cell layers, all retinal nerve fiber layer quadrants, and all choroidal quadrants (except for the central subfield) in migraineurs compared to controls. The duration of migraine was significantly correlated with ganglion cell layer, retinal nerve fiber layer, and all choroidal quadrants, while the severity of migraine was significantly correlated with ganglion cell layer and retinal nerve fiber layer only. Multiregression analysis showed that the duration of migraine is the most important determinant factor of the superior retinal nerve fiber layer quadrant (β = -0.375, p = 0.001) and in all the choroidal quadrants (β = -0.531, -0.692, -0.503, -0.461, -0.564, respectively, p  layer quadrants (β = -0.256, -0.335, -0.308; p  = 0.036, 0.005, 0.009, respectively) and the inferior ganglion cell layer hemisphere (β = -0.377 and p = 0.001). Ganglion cell layer, retinal nerve fiber layer, and choroidal thickness are significantly thinner in patients with migraine. The severity of migraine has more significant influence in the thinning of ganglion cell layer and retinal nerve fiber layer, while the duration of the disease affected the choroidal thickness more.

  9. Empirical Derivation of Correction Factors for Human Spiral Ganglion Cell Nucleus and Nucleolus Count Units.

    Science.gov (United States)

    Robert, Mark E; Linthicum, Fred H

    2016-01-01

    Profile count method for estimating cell number in sectioned tissue applies a correction factor for double count (resulting from transection during sectioning) of count units selected to represent the cell. For human spiral ganglion cell counts, we attempted to address apparent confusion between published correction factors for nucleus and nucleolus count units that are identical despite the role of count unit diameter in a commonly used correction factor formula. We examined a portion of human cochlea to empirically derive correction factors for the 2 count units, using 3-dimensional reconstruction software to identify double counts. The Neurotology and House Histological Temporal Bone Laboratory at University of California at Los Angeles. Using a fully sectioned and stained human temporal bone, we identified and generated digital images of sections of the modiolar region of the lower first turn of cochlea, identified count units with a light microscope, labeled them on corresponding digital sections, and used 3-dimensional reconstruction software to identify double-counted count units. For 25 consecutive sections, we determined that double-count correction factors for nucleus count unit (0.91) and nucleolus count unit (0.92) matched the published factors. We discovered that nuclei and, therefore, spiral ganglion cells were undercounted by 6.3% when using nucleolus count units. We determined that correction factors for count units must include an element for undercounting spiral ganglion cells as well as the double-count element. We recommend a correction factor of 0.91 for the nucleus count unit and 0.98 for the nucleolus count unit when using 20-µm sections. © American Academy of Otolaryngology—Head and Neck Surgery Foundation 2015.

  10. Cerebellar and basal ganglion involvement in Langerhans cell histiocytosis

    Energy Technology Data Exchange (ETDEWEB)

    Saatci, I.; Baskan, O.; Haliloglu, M.; Aydingoz, U. [Department of Radiology, Hacettepe University Hospital, Sihhiye 06100, Ankara (Turkey)

    1999-06-01

    Langerhans cell histiocytosis (LCH) is a disease of unknown cause characterised by proliferation of histiocytic granulomas in tissues; the primary cerebral manifestation is diabetes insipidus caused by hypothalamic infiltration. We present a patient in whom, except for the absence of high signal on T 1 weighting in the posterior pituitary, consistent with central diabetes insipidus, MRI showed no evidence of hypothalamic involvement by histiocytosis, despite the long duration of the disease. However, there was bilateral, symmetrical involvement of the cerebellum and globus pallidus in addition to a calvarial lesion. High signal in the cerebellar white matter on T 2-weighted images may represent demyelination, gliosis and cell loss, as previously reported on pathologic examination. (orig.) With 5 figs., 22 refs.

  11. Dendritic thickness: a morphometric parameter to classify mouse retinal ganglion cells

    Directory of Open Access Journals (Sweden)

    L.D. Loopuijt

    2007-10-01

    Full Text Available To study the dendritic morphology of retinal ganglion cells in wild-type mice we intracellularly injected these cells with Lucifer yellow in an in vitro preparation of the retina. Subsequently, quantified values of dendritic thickness, number of branching points and level of stratification of 73 Lucifer yellow-filled ganglion cells were analyzed by statistical methods, resulting in a classification into 9 groups. The variables dendritic thickness, number of branching points per cell and level of stratification were independent of each other. Number of branching points and level of stratification were independent of eccentricity, whereas dendritic thickness was positively dependent (r = 0.37 on it. The frequency distribution of dendritic thickness tended to be multimodal, indicating the presence of at least two cell populations composed of neurons with dendritic diameters either smaller or larger than 1.8 µm ("thin" or "thick" dendrites, respectively. Three cells (4.5% were bistratified, having thick dendrites, and the others (95.5% were monostratified. Using k-means cluster analysis, monostratified cells with either thin or thick dendrites were further subdivided according to level of stratification and number of branching points: cells with thin dendrites were divided into 2 groups with outer stratification (0-40% and 2 groups with inner (50-100% stratification, whereas cells with thick dendrites were divided into one group with outer and 3 groups with inner stratification. We postulate, that one group of cells with thin dendrites resembles cat ß-cells, whereas one group of cells with thick dendrites includes cells that resemble cat a-cells.

  12. [Progression of nerve fiber layer defects in retrobulbar optic neuritis by the macular ganglion cell complex].

    Science.gov (United States)

    Hong, D; Bosc, C; Chiambaretta, F

    2017-11-01

    Recent studies with SD OCT had shown early axonal damage to the macular ganglion cell complex (which consists of the three innermost layers of the retina: Inner Plexiform Layer [IPL], Ganglion Cell Layer [GCL], Retinal Nerve Fibre layer [RNFL]) in optic nerve pathology. Retrobulbar optic neuritis (RBON), occurring frequently in demyelinating diseases, leads to atrophy of the optic nerve fibers at the level of the ganglion cell axons, previously described in the literature. The goal of this study is to evaluate the progression of optic nerve fiber defects and macular ganglion cell complex defects with the SPECTRALIS OCT via a reproducible method by calculating a mean thickness in each quadrant after an episode of retrobulbar optic neuritis. This is a prospective monocentric observational study including 8 patients at the Clermont-Ferrand university medical center. All patients underwent ocular examination with macular and disc OCT analysis and a Goldmann visual field at the time of inclusion (onset or recurrence of RBON), at 3 months and at 6 months. Patients were 40-years-old on average at the time of inclusion. After 6 months of follow-up, there was progression of the atrophy of the macular ganglion cell complex in the affected eye on (11.5% or 11μm) predominantly inferonasally (13.9% or 16μm) and superonasally (12.9% or 14μm) while the other eye remained stable. The decrease in thickness occurred mainly in the most internal 3 layers of the retina. On average, the loss in thickness of the peripapillary RNFL was predominantly inferotemporal (24.9% or 39μm) and superotemporal (21.8% or 28μm). In 3 months of progression, the loss of optic nerve fibers is already seen on macular and disc OCT after an episode of RBON, especially in inferior quadrants in spite of the improvement in the Goldmann visual field and visual acuity. Segmentation by quadrant was used here to compare the progression of the defect by region compared to the fovea in a global and reproducible

  13. Cell cycle and apoptosis genes in atherosclerosis

    NARCIS (Netherlands)

    Boesten, Lianne Simone Mirjam

    2006-01-01

    The work described in this thesis was aimed at identifying the role of cell cycle and apoptosis genes in atherosclerosis. Atherosclerosis is the primary cause of cardiovascular disease, a disorder occurring in the large and medium-sized arteries of the body. Although in the beginning 90s promising

  14. Spiral ganglion cell site of excitation I: comparison of scala tympani and intrameatal electrode responses.

    Science.gov (United States)

    Cartee, Lianne A; Miller, Charles A; van den Honert, Chris

    2006-05-01

    To determine the site of excitation on the spiral ganglion cell in response to electrical stimulation similar to that from a cochlear implant, single-fiber responses to electrical stimuli delivered by an electrode positioned in the scala tympani were compared to responses from stimuli delivered by an electrode placed in the internal auditory meatus. The response to intrameatal stimulation provided a control set of data with a known excitation site, the central axon of the spiral ganglion cell. For both intrameatal and scala tympani stimuli, the responses to single-pulse, summation, and refractory stimulus protocols were recorded. The data demonstrated that summation pulses, as opposed to single pulses, are likely to give the most insightful measures for determination of the site of excitation. Single-fiber summation data for both scala tympani and intrameatally stimulated fibers were analyzed with a clustering algorithm. Combining cluster analysis and additional numerical modeling data, it was hypothesized that the scala tympani responses corresponded to central excitation, peripheral excitation adjacent to the cell body, and peripheral excitation at a site distant from the cell body. Fibers stimulated by an intrameatal electrode demonstrated the greatest range of jitter measurements indicating that greater fiber independence may be achieved with intrameatal stimulation.

  15. The retina of the shovel-nosed ray, Rhinobatos batillum (Rhinobatidae): morphology and quantitative analysis of the ganglion, amacrine and bipolar cell populations.

    Science.gov (United States)

    Collin, S P

    1988-01-01

    A light microscopy study of the retina of the shovel-nosed ray, Rhinobatos batillum (Rhinobatidae) has revealed a duplex retina with a rod to cone ratio between 4:1 and 6:1. The inner nuclear layer consists of three layers of large horizontal cells, tightly packed, stellate bipolar cells, and up to three substrata of amacrine cells. The collaterals of the many supporting Müller cells project from the inner to the outer limiting membrane and divide the retina into many subunits. The cells of the ganglion cell layer are distributed into two layers, although a large proportion of ganglion cells are also displaced into the inner plexiform and inner nuclear layers. Topographic analysis of the cells in the ganglion cell layer, inner plexiform and inner nuclear layers reveals a number of regional specializations or "areae centrales". Ganglion cells were retrogradely-labelled with cobalt-lysine from the optic nerve, and three sub-populations of neurons characterized on their soma size and position. Small (20-50 microns2), large (80-300 microns2) and giant (greater than 300 microns2) sub-populations of ganglion cells each revealed distinct retinal specializations with peak densities of 3 x 10(3), 1.25 x 10(3) and 1.57 x 10(3) cells per mm2, respectively. Topographical comparison between Nissl-stained and retrogradely-labelled ganglion cell populations have established that a maximum of 20% in the "area centralis", and 75% in unspecialized, peripheral regions of the retina are non-ganglion cells. Out of a total of 210,566 cells in the ganglion cell layer, 49% were found to be non-ganglion cells. Iso-density contour maps of amacrine and bipolar cell distributions also reveal some specializations. These cell concentrations lie in corresponding regions to areas of increased density in the large and giant ganglion cell populations, suggesting some functional association.

  16. Investigation of retinal ganglion cells and axons of normal rats using fluorogold retrograde labeling

    International Nuclear Information System (INIS)

    Yin Xiaolei; Ye Jian; Chen Chunlin

    2006-01-01

    To investigate the retinal ganglion cells (RGCs) by means of fluorogold retrograde labeling, RGCs were labeled by injecting the fluorogold bilaterally into the superficial superior colliculus and lateral genicutate nucleus in six adult SD rats. One and two weeks (3 rats in each group) after injecting the fluorogold, RGCs FG-labeled were observed and the number of them were counted. The results showed that after a week mean density of fluorogold-labeled RGCs was 2210 ± 128/mm 2 , and it was 2164 ± 117/mm 2 after two weeks. Our conclusion is fluorogold retrograde labeling could be very useful in the research of RGCs. (authors)

  17. Macular retinal ganglion cell-inner plexiform layer thickness in patients on hydroxychloroquine therapy.

    Science.gov (United States)

    Lee, Min Gyu; Kim, Sang Jin; Ham, Don-Il; Kang, Se Woong; Kee, Changwon; Lee, Jaejoon; Cha, Hoon-Suk; Koh, Eun-Mi

    2014-11-25

    We evaluated macular ganglion cell-inner plexiform layer (GC-IPL) thickness using spectral-domain optical coherence tomography (SD-OCT) in patients with chronic exposure to hydroxychloroquine (HCQ). This study included 130 subjects, who were divided into three groups: Group 1A, 55 patients with HCQ use ≥5 years; Group 1B, 46 patients with HCQ use 1000 g), significant correlations were not observed. This study revealed that macular GC-IPL thickness did not show definite correlations with HCQ use. However, some patients, especially with HCQ retinopathy or high cumulative doses, showed thin GC-IPL. Copyright 2015 The Association for Research in Vision and Ophthalmology, Inc.

  18. Distinguishing ischaemic optic neuropathy from optic neuritis by ganglion cell analysis.

    Science.gov (United States)

    Erlich-Malona, Natalie; Mendoza-Santiesteban, Carlos E; Hedges, Thomas R; Patel, Nimesh; Monaco, Caitlin; Cole, Emily

    2016-12-01

    To determine whether a pattern of altitudinal ganglion cell loss, as detected and measured by optical coherence tomography (OCT), can be used to distinguish non-arteritic ischaemic optic neuropathy (NAION) from optic neuritis (ON) during the acute phase, and whether the rate or severity of ganglion cell loss differs between the two diseases. We performed a retrospective, case-control study of 44 patients (50 eyes) with ON or NAION and 44 age-matched controls. Non-arteritic ischaemic optic neuropathy and ON patients had OCT at presentation and four consecutive follow-up visits. Controls had OCT at one point in time. The ganglion cell complex (GCC) was evaluated in the macula, and the retinal nerve fibre layer (RNFL) was evaluated in the peripapillary region. Ganglion cell complex thickness, RNFL thickness and GCC mean superior and inferior hemispheric difference were compared between NAION and ON patients at each time-point using unpaired t-tests and between disease and control subjects at first measurement using paired t-tests. Mean time from onset of symptoms to initial presentation was 10.7 ± 6.6 days in NAION and 11.7 ± 8.6 days in ON (p = 0.67). There was a significantly greater vertical hemispheric difference in GCC thickness in NAION patients than ON patients at all time-points (5.5-10.7 μm versus 3.1-3.6 μm, p = 0.01-0.049). Mean GCC thickness was significantly decreased at less than 2 weeks after onset in NAION compared to age-matched controls (72.1 μm versus 82.1 μm, p < 0.001), as well as in ON compared to age-matched controls (74.3 μm versus 84.5 μm, p < 0.001). Progression and severity of GCC and RNFL loss did not differ significantly between NAION and ON. A quantitative comparison of mean superior and inferior hemispheric GCC thickness with OCT may be used to distinguish NAION from ON. © 2016 Acta Ophthalmologica Scandinavica Foundation. Published by John Wiley & Sons Ltd.

  19. Delayed-rectifier K channels contribute to contrast adaptation in mammalian retinal ganglion cells.

    Science.gov (United States)

    Weick, Michael; Demb, Jonathan B

    2011-07-14

    Retinal ganglion cells adapt by reducing their sensitivity during periods of high contrast. Contrast adaptation in the firing response depends on both presynaptic and intrinsic mechanisms. Here, we investigated intrinsic mechanisms for contrast adaptation in OFF Alpha ganglion cells in the in vitro guinea pig retina. Using either visual stimulation or current injection, we show that brief depolarization evoked spiking and suppressed firing during subsequent depolarization. The suppression could be explained by Na channel inactivation, as shown in salamander cells. However, brief hyperpolarization in the physiological range (5-10 mV) also suppressed firing during subsequent depolarization. This suppression was selectively sensitive to blockers of delayed-rectifier K channels (K(DR)). In somatic membrane patches, we observed tetraethylammonium-sensitive K(DR) currents that activated near -25 mV. Recovery from inactivation occurred at potentials hyperpolarized to V(rest). Brief periods of hyperpolarization apparently remove K(DR) inactivation and thereby increase the channel pool available to suppress excitability during subsequent depolarization. Copyright © 2011 Elsevier Inc. All rights reserved.

  20. Primary culture of glial cells from mouse sympathetic cervical ganglion: a valuable tool for studying glial cell biology.

    Science.gov (United States)

    de Almeida-Leite, Camila Megale; Arantes, Rosa Maria Esteves

    2010-12-15

    Central nervous system glial cells as astrocytes and microglia have been investigated in vitro and many intracellular pathways have been clarified upon various stimuli. Peripheral glial cells, however, are not as deeply investigated in vitro despite its importance role in inflammatory and neurodegenerative diseases. Based on our previous experience of culturing neuronal cells, our objective was to standardize and morphologically characterize a primary culture of mouse superior cervical ganglion glial cells in order to obtain a useful tool to study peripheral glial cell biology. Superior cervical ganglia from neonatal C57BL6 mice were enzymatically and mechanically dissociated and cells were plated on diluted Matrigel coated wells in a final concentration of 10,000cells/well. Five to 8 days post plating, glial cell cultures were fixed for morphological and immunocytochemical characterization. Glial cells showed a flat and irregular shape, two or three long cytoplasm processes, and round, oval or long shaped nuclei, with regular outline. Cell proliferation and mitosis were detected both qualitative and quantitatively. Glial cells were able to maintain their phenotype in our culture model including immunoreactivity against glial cell marker GFAP. This is the first description of immunocytochemical characterization of mouse sympathetic cervical ganglion glial cells in primary culture. This work discusses the uses and limitations of our model as a tool to study many aspects of peripheral glial cell biology. Copyright © 2010 Elsevier B.V. All rights reserved.

  1. Expression of polysialylated neural cell adhesion molecules on adult stem cells after neuronal differentiation of inner ear spiral ganglion neurons

    Energy Technology Data Exchange (ETDEWEB)

    Park, Kyoung Ho [Department of Otolaryngology Head and Neck Surgery, College of Medicine, Catholic University, Seoul (Korea, Republic of); Yeo, Sang Won, E-mail: swyeo@catholic.ac.kr [Department of Otolaryngology Head and Neck Surgery, College of Medicine, Catholic University, Seoul (Korea, Republic of); Troy, Frederic A., E-mail: fatroy@ucdavis.edu [Department of Biochemistry and Molecular Medicine, University of California, School of Medicine, Davis, CA 95616 (United States); Xiamen University, School of Medicine, Xiamen City (China)

    2014-10-17

    Highlights: • PolySia expressed on neurons primarily during early stages of neuronal development. • PolySia–NCAM is expressed on neural stem cells from adult guinea pig spiral ganglion. • PolySia is a biomarker that modulates neuronal differentiation in inner ear stem cells. - Abstract: During brain development, polysialylated (polySia) neural cell adhesion molecules (polySia–NCAMs) modulate cell–cell adhesive interactions involved in synaptogenesis, neural plasticity, myelination, and neural stem cell (NSC) proliferation and differentiation. Our findings show that polySia–NCAM is expressed on NSC isolated from adult guinea pig spiral ganglion (GPSG), and in neurons and Schwann cells after differentiation of the NSC with epidermal, glia, fibroblast growth factors (GFs) and neurotrophins. These differentiated cells were immunoreactive with mAb’s to polySia, NCAM, β-III tubulin, nestin, S-100 and stained with BrdU. NSC could regenerate and be differentiated into neurons and Schwann cells. We conclude: (1) polySia is expressed on NSC isolated from adult GPSG and on neurons and Schwann cells differentiated from these NSC; (2) polySia is expressed on neurons primarily during the early stage of neuronal development and is expressed on Schwann cells at points of cell–cell contact; (3) polySia is a functional biomarker that modulates neuronal differentiation in inner ear stem cells. These new findings suggest that replacement of defective cells in the inner ear of hearing impaired patients using adult spiral ganglion neurons may offer potential hope to improve the quality of life for patients with auditory dysfunction and impaired hearing disorders.

  2. Expression of polysialylated neural cell adhesion molecules on adult stem cells after neuronal differentiation of inner ear spiral ganglion neurons

    International Nuclear Information System (INIS)

    Park, Kyoung Ho; Yeo, Sang Won; Troy, Frederic A.

    2014-01-01

    Highlights: • PolySia expressed on neurons primarily during early stages of neuronal development. • PolySia–NCAM is expressed on neural stem cells from adult guinea pig spiral ganglion. • PolySia is a biomarker that modulates neuronal differentiation in inner ear stem cells. - Abstract: During brain development, polysialylated (polySia) neural cell adhesion molecules (polySia–NCAMs) modulate cell–cell adhesive interactions involved in synaptogenesis, neural plasticity, myelination, and neural stem cell (NSC) proliferation and differentiation. Our findings show that polySia–NCAM is expressed on NSC isolated from adult guinea pig spiral ganglion (GPSG), and in neurons and Schwann cells after differentiation of the NSC with epidermal, glia, fibroblast growth factors (GFs) and neurotrophins. These differentiated cells were immunoreactive with mAb’s to polySia, NCAM, β-III tubulin, nestin, S-100 and stained with BrdU. NSC could regenerate and be differentiated into neurons and Schwann cells. We conclude: (1) polySia is expressed on NSC isolated from adult GPSG and on neurons and Schwann cells differentiated from these NSC; (2) polySia is expressed on neurons primarily during the early stage of neuronal development and is expressed on Schwann cells at points of cell–cell contact; (3) polySia is a functional biomarker that modulates neuronal differentiation in inner ear stem cells. These new findings suggest that replacement of defective cells in the inner ear of hearing impaired patients using adult spiral ganglion neurons may offer potential hope to improve the quality of life for patients with auditory dysfunction and impaired hearing disorders

  3. Xenopus laevis Retinal Ganglion Cell Dendritic Arbors Develop Independently of Visual Stimulation

    Directory of Open Access Journals (Sweden)

    Barbara Lom

    2004-01-01

    Full Text Available Newly formed neurons must locate their appropriate target cells and then form synaptic connections with these targets in order to establish a functional nervous system. In the vertebrate retina, retinal ganglion cell (RGC dendrites extend from the cell body and form synapses with nearby amacrine and bipolar cells. RGC axons, however, exit the retina and synapse with the dendrites of midbrain neurons in the optic tectum. We examined how visual stimulation influenced Xenopus RGC dendritic arborization. Neuronal activity is known to be an important factor in shaping dendritic and axonal arborization. Thus, we reared tadpoles in dark and light environments then used rhodamine dextran retrograde labeling to identify RGCs in the retina. When we compared RGC dendritic arbors from tadpoles reared in dark and light environments, we found no morphological differences, suggesting that physiological visual activity did not contribute to the morphological development of Xenopus RGC dendritic arbors.

  4. A feed-forward regulation of endothelin receptors by c-Jun in human non-pigmented ciliary epithelial cells and retinal ganglion cells.

    Directory of Open Access Journals (Sweden)

    Junming Wang

    Full Text Available c-Jun, c-Jun N-terminal kinase(JNK and endothelin B (ETB receptor have been shown to contribute to the pathogenesis of glaucoma. Previously, we reported that an increase of c-Jun and CCAAT/enhancer binding protein β (C/EBPβ immunohistostaining is associated with upregulation of the ETB receptor within the ganglion cell layer of rats with elevated intraocular pressure (IOP. In addition, both transcription factors regulate the expression of the ETB receptor in human non-pigmented ciliary epithelial cells (HNPE. The current study addressed the mechanisms by which ET-1 produced upregulation of ET receptors in primary rat retinal ganglion cells (RGCs and HNPE cells. Treatment of ET-1 and ET-3 increased the immunocytochemical staining of c-Jun and C/EBPβ in primary rat RGCs and co-localization of both transcription factors was observed. A marked increase in DNA binding activity of AP-1 and C/EBPβ as well as elevated protein levels of c-Jun and c-Jun-N-terminal kinase (JNK were detected following ET-1 treatment in HNPE cells. Overexpression of ETA or ETB receptor promoted the upregulation of c-Jun and also elevated its promoter activity. In addition, upregulation of C/EBPβ augmented DNA binding and mRNA expression of c-Jun, and furthermore, the interaction of c-Jun and C/EBPβ was confirmed using co-immunoprecipitation. Apoptosis of HNPE cells was identified following ET-1 treatment, and overexpression of the ETA or ETB receptor produced enhanced apoptosis. ET-1 mediated upregulation of c-Jun and C/EBPβ and their interaction may represent a novel mechanism contributing to the regulation of endothelin receptor expression. Reciprocally, c-Jun was also found to regulate the ET receptors and C/EBPβ appeared to play a regulatory role in promoting expression of c-Jun. Taken together, the data suggests that ET-1 triggers the upregulation of c-Jun through both ETA and ETB receptors, and conversely c-Jun also upregulates endothelin receptor expression

  5. Retinal vessel diameters decrease with macular ganglion cell layer thickness in autosomal dominant optic atrophy and in healthy subjects

    DEFF Research Database (Denmark)

    Rönnbäck, Cecilia; Grønskov, Karen; Larsen, Michael

    2014-01-01

    diameters (central retinal artery equivalent, CRAE, and central retinal vein equivalent, CRVE). Statistical analysis was corrected for age, gender, spherical equivalent refraction, axial length and mean arterial blood pressure (MABP) in a mixed model analysis. RESULTS: Retinal arteries and veins were...... ganglion cell-inner plexiform layer (GC-IPL) thickness (p = 0.0017 and p = 0.0057, respectively). CONCLUSION: Narrow retinal arteries and veins were associated not only with the severity of ADOA but with ganglion cell volume in patients with ADOA and in healthy subjects. This suggests that narrow vessels...

  6. Adult rat retinal ganglion cells and glia can be printed by piezoelectric inkjet printing

    International Nuclear Information System (INIS)

    Lorber, Barbara; Martin, Keith R; Hsiao, Wen-Kai; Hutchings, Ian M

    2014-01-01

    We have investigated whether inkjet printing technology can be extended to print cells of the adult rat central nervous system (CNS), retinal ganglion cells (RGC) and glia, and the effects on survival and growth of these cells in culture, which is an important step in the development of tissue grafts for regenerative medicine, and may aid in the cure of blindness. We observed that RGC and glia can be successfully printed using a piezoelectric printer. Whilst inkjet printing reduced the cell population due to sedimentation within the printing system, imaging of the printhead nozzle, which is the area where the cells experience the greatest shear stress and rate, confirmed that there was no evidence of destruction or even significant distortion of the cells during jet ejection and drop formation. Importantly, the viability of the cells was not affected by the printing process. When we cultured the same number of printed and non-printed RGC/glial cells, there was no significant difference in cell survival and RGC neurite outgrowth. In addition, use of a glial substrate significantly increased RGC neurite outgrowth, and this effect was retained when the cells had been printed. In conclusion, printing of RGC and glia using a piezoelectric printhead does not adversely affect viability and survival/growth of the cells in culture. Importantly, printed glial cells retain their growth-promoting properties when used as a substrate, opening new avenues for printed CNS grafts in regenerative medicine. (paper)

  7. Transglial transmission at the dorsal root ganglion sandwich synapse: glial cell to postsynaptic neuron communication.

    Science.gov (United States)

    Rozanski, Gabriela M; Li, Qi; Stanley, Elise F

    2013-04-01

    The dorsal root ganglion (DRG) contains a subset of closely-apposed neuronal somata (NS) separated solely by a thin satellite glial cell (SGC) membrane septum to form an NS-glial cell-NS trimer. We recently reported that stimulation of one NS with an impulse train triggers a delayed, noisy and long-lasting response in its NS pair via a transglial signaling pathway that we term a 'sandwich synapse' (SS). Transmission could be unidirectional or bidirectional and facilitated in response to a second stimulus train. We have shown that in chick or rat SS the NS-to-SGC leg of the two-synapse pathway is purinergic via P2Y2 receptors but the second SGC-to-NS synapse mechanism remained unknown. A noisy evoked current in the target neuron, a reversal potential close to 0 mV, and insensitivity to calcium scavengers or G protein block favored an ionotropic postsynaptic receptor. Selective block by D-2-amino-5-phosphonopentanoate (AP5) implicated glutamatergic transmission via N-methyl-d-aspartate receptors. This agent also blocked NS responses evoked by puff of UTP, a P2Y2 agonist, directly onto the SGC cell, confirming its action at the second synapse of the SS transmission pathway. The N-methyl-d-aspartate receptor NR2B subunit was implicated by block of transmission with ifenprodil and by its immunocytochemical localization to the NS membrane, abutting the glial septum P2Y2 receptor. Isolated DRG cell clusters exhibited daisy-chain and branching NS-glial cell-NS contacts, suggestive of a network organization within the ganglion. The identification of the glial-to-neuron transmitter and receptor combination provides further support for transglial transmission and completes the DRG SS molecular transmission pathway. © 2013 Federation of European Neuroscience Societies and Blackwell Publishing Ltd.

  8. The trophic effect of ouabain on retinal ganglion cells is mediated by IL-1β and TNF-α

    International Nuclear Information System (INIS)

    Salles von-Held-Ventura, Juliana; Mázala-de-Oliveira, Thalita; Cândida da Rocha Oliveira, Amanda; Granja, Marcelo Gomes; Gonçalves-de-Albuquerque, Cassiano Felippe; Castro-Faria-Neto, Hugo Caire; Giestal-de-Araujo, Elizabeth

    2016-01-01

    Ouabain is a steroid hormone that binds to the enzyme Na + , K + – ATPase and stimulates different intracellular pathways controlling growth, proliferation and cell survival. IL-1β and TNF-α are pleiotropic molecules, conventionally regarded as pro-inflammatory cytokines with well-known effects in the immune system. In addition, IL-1β and TNF-α also play important roles in the nervous system including neuroprotective effects. Previous data from our group showed that ouabain treatment is able to induce an increase in retinal ganglion cell survival kept in mixed retinal cell cultures. The aim of this work was to investigate if IL-1β and TNF-α could be mediating the trophic effect of ouabain on retinal ganglion cells. Our results show that the trophic effect of ouabain on retinal ganglion cell was inhibited by either anti-IL-1β or anti-TNF-α antibodies. In agreement, IL-1β or TNF-α increased the retinal ganglion cells survival in a dose-dependent manner. Accordingly, ouabain treatment induces a temporal release of TNF-α and IL-1β from retinal cell cultures. Interestingly, TNF-α and IL-1β regulate each other intracellular levels. Our results suggest that ouabain treatment triggers the activation of TNF-α and IL-1β signaling pathways leading to an increase in retinal ganglion cell survival. - Highlights: • Pro-inflammatory cytokines regulates the ouabain effect on RGC survival. • Ouabain treatment modulates the intracellular levels of TNF-α and IL-1β. • Ouabain induces the release of TNF-α and IL-1β in retinal cell cultures.

  9. The trophic effect of ouabain on retinal ganglion cells is mediated by IL-1β and TNF-α

    Energy Technology Data Exchange (ETDEWEB)

    Salles von-Held-Ventura, Juliana; Mázala-de-Oliveira, Thalita; Cândida da Rocha Oliveira, Amanda; Granja, Marcelo Gomes [Departamento de Neurobiologia, Programa de Neurociências, Outeiro de São João Batista s/n CEP: 24020-150, Universidade Federal Fluminense, Niterói, RJ (Brazil); Gonçalves-de-Albuquerque, Cassiano Felippe; Castro-Faria-Neto, Hugo Caire [Fundação Oswaldo Cruz, Instituto Oswaldo Cruz, Departamento de Fisiologia e Farmacodinâmica, Av., no 4365, Manguinhos, 21045-900, Rio de Janeiro, RJ (Brazil); Giestal-de-Araujo, Elizabeth, E-mail: egiestal@vm.uff.br [Departamento de Neurobiologia, Programa de Neurociências, Outeiro de São João Batista s/n CEP: 24020-150, Universidade Federal Fluminense, Niterói, RJ (Brazil)

    2016-09-09

    Ouabain is a steroid hormone that binds to the enzyme Na{sup +}, K{sup +} – ATPase and stimulates different intracellular pathways controlling growth, proliferation and cell survival. IL-1β and TNF-α are pleiotropic molecules, conventionally regarded as pro-inflammatory cytokines with well-known effects in the immune system. In addition, IL-1β and TNF-α also play important roles in the nervous system including neuroprotective effects. Previous data from our group showed that ouabain treatment is able to induce an increase in retinal ganglion cell survival kept in mixed retinal cell cultures. The aim of this work was to investigate if IL-1β and TNF-α could be mediating the trophic effect of ouabain on retinal ganglion cells. Our results show that the trophic effect of ouabain on retinal ganglion cell was inhibited by either anti-IL-1β or anti-TNF-α antibodies. In agreement, IL-1β or TNF-α increased the retinal ganglion cells survival in a dose-dependent manner. Accordingly, ouabain treatment induces a temporal release of TNF-α and IL-1β from retinal cell cultures. Interestingly, TNF-α and IL-1β regulate each other intracellular levels. Our results suggest that ouabain treatment triggers the activation of TNF-α and IL-1β signaling pathways leading to an increase in retinal ganglion cell survival. - Highlights: • Pro-inflammatory cytokines regulates the ouabain effect on RGC survival. • Ouabain treatment modulates the intracellular levels of TNF-α and IL-1β. • Ouabain induces the release of TNF-α and IL-1β in retinal cell cultures.

  10. MicroRNA-1 promotes apoptosis of hepatocarcinoma cells by targeting apoptosis inhibitor-5 (API-5).

    Science.gov (United States)

    Li, Dong; Liu, Yu; Li, Hua; Peng, Jing-Jing; Tan, Yan; Zou, Qiang; Song, Xiao-Feng; Du, Min; Yang, Zheng-Hui; Tan, Yong; Zhou, Jin-Jun; Xu, Tao; Fu, Zeng-Qiang; Feng, Jian-Qiong; Cheng, Peng; chen, Tao; Wei, Dong; Su, Xiao-Mei; Liu, Huan-Yi; Qi, Zhong-Chun; Tang, Li-Jun; Wang, Tao; Guo, Xin; Hu, Yong-He; Zhang, Tao

    2015-01-02

    Although microRNA-1 (miR-1) is a known liver cancer suppressor, the role of miR-1 in apoptosis of hepatoma cells has remained largely unknown. Our study shows that ectopic miR-1 overexpression induced apoptosis of liver hepatocellular carcinoma (HepG2) cells. Apoptosis inhibitor 5 (API-5) was found to be a potential regulator of miR-1 induced apoptosis, using a bioinformatics approach. Furthermore, an inverse relationship between miR-1 and API-5 expression was observed in human liver cancer tissues and adjacent normal liver tissues. Negative regulation of API-5 expression by miR-1 was demonstrated to promote apoptosis of HepG2 cells. Our study provides a novel regulatory mechanism of miR-1 in the apoptosis of hepatoma cells. Copyright © 2014 Federation of European Biochemical Societies. Published by Elsevier B.V. All rights reserved.

  11. Different mechanisms between premitotic apoptosis and postmitotic apoptosis in X-irradiated U937 cells

    International Nuclear Information System (INIS)

    Shinomiya, Nariyoshi; Kuno, Yukie; Yamamoto, Fuyumi; Fukasawa, Masashi; Okumura, Atsushi; Uefuji, Megumi; Rokutanda, Makoto

    2000-01-01

    Purpose: Apoptosis is currently being evaluated for its importance as a pathway of radiation-induced cell death. However, the difference in the mechanisms between premitotic and postmitotic apoptosis following X-irradiation remains not well understood. We show here that the human monoblastoid cell line U937 can be induced to undergo these two different types of apoptosis. Methods and Materials: U937 cells were irradiated at a dose of 5 or 20 Gy, and the DNA fragmentation rate was measured by both flow cytometric analysis and gel electrophoresis. Activation of caspase-3 was detected by Western blot analysis and fluorogenic assay using acetyl-Asp-Glu-Val-Asp-7-amino-4-methyl-coumarin (Ac-DEVD-AMC). Detection of mitochondrial transmembrane potential (no. DELTAno. no. PSIno. ) was performed by using Rho123. Chasing of S-phase fraction following X-irradiation was performed after labeling with 5-bromo-2'-deoxyuridine (BrdU). Thymidine was used for synchronization of the cells. Inhibition of caspase-3 activity was achieved by Acetyl-Asp-Glu-Val-Asp-aldehyde (Ac-DEVD-CHO). Results: Time courses of the apoptotic rates, caspase activation, and no. DELTAno. no. PSIno. indicated that two different types of cell death were induced by the different X-ray doses. High-dose X-ray (20 Gy) induced a rapid and strong apoptosis, whereas low-dose X-ray (5 Gy) induced a slow and mild apoptosis. Cell-cycle analyses revealed that there was cell death before cell division in the former apoptosis but the cells must be dying after cell division in the latter apoptosis. By means of cell-cycle synchronization, the S-phase cells proved to be the most sensitive fraction to premitotic apoptosis, but an obvious difference in the susceptibility to cell death among the cell-cycle phases was not observed in postmitotic apoptosis. Ac-DEVD-CHO treatment effectively blocked caspase activity and premitotic apoptosis, but it failed to block postmitotic apoptosis. Conclusions: Irradiation of U937 cells at

  12. Effect of Extracellular Zinc Chelator on Rat Retinal Ganglion Cell Number, and Taurine and Zinc Transporters in These Cells

    Directory of Open Access Journals (Sweden)

    Asarí Márquez García

    2017-05-01

    Full Text Available Zinc deficiency in humans causes decreased antioxidants in the retina and is related with abnormal darkness adaptation, cataracts, blindness, and macular degeneration. There is little information about the effects of zinc on the taurine system in mammalian retinal cells. Therefore, we studied the effect of zinc on the taurine transporter (TAUT and zinc transporters (ZnT-1 and 3 using the extracellular zinc chelator, diethylenetriaminepentaacetic acid (DTPA by fluorescence immunocytochemistry and immunohistochemistry in the ganglion cells (CG and cell layers of the retina of rats. Three days after administration of DTPA (10µM primary antibodies and secondary antibodies conjugated with rhodamine or fluorescein isothiocyanate (FITC were used as required. For immunocytochemical labeling approximately three hundred cells per condition were counted. For immunohistochemical labeling, the fluorescence intensity was measured as integrated optical density (DOI in four areas for each layer of tissue. DTPA produced a decrease of 32 % and 29 % in GC of the total cells labeled with antibody against glycoprotein Thy 1.1 and γ-synuclein, respectively. It also produced a significant decrease in TAUT localization in 27 and 28 % compared to controls. DTPA produced a decrease in the localization of ZnT-1 and ZnT-3 in the retina layers (ganglion cells, GCC and the outer and inner plexiform, CEP and CIP. The study of these molecules in the retina is relevant to understanding the interactions of taurine and zinc in this structure.

  13. Synchronized Firings in Retinal Ganglion Cells in Response to Natural Stimulation

    International Nuclear Information System (INIS)

    Zhang Ying-Ying; Xiao Lei; Liu Wen-Zhong; Gong Hai-Qing; Liang Pei-Ji

    2011-01-01

    The response of synchronously firing groups of population retinal ganglion cells (RGCs) to natural movies (NMs) and pseudo-random white-noise checker-board flickering (CB, as control) are investigated using an information-theoretic algorithm. The main results are: (1) the population RGCs tend to fire in synchrony far more frequently than expected by chance during both NM and CB stimulation; (2) more synchronous groups could be formed and each group contains more neurons under NM than CB stimulation; (3) the individual neurons also participate in more groups and have more distinct partners in NM than CB stimulation. All these results suggest that the synchronized firings in RGCs are more extensive and diverse, which may account for more effective information processing in representing the natural visual environment. (cross-disciplinary physics and related areas of science and technology)

  14. Rapid and coordinated processing of global motion images by local clusters of retinal ganglion cells.

    Science.gov (United States)

    Matsumoto, Akihiro; Tachibana, Masao

    2017-01-01

    Even when the body is stationary, the whole retinal image is always in motion by fixational eye movements and saccades that move the eye between fixation points. Accumulating evidence indicates that the brain is equipped with specific mechanisms for compensating for the global motion induced by these eye movements. However, it is not yet fully understood how the retina processes global motion images during eye movements. Here we show that global motion images evoke novel coordinated firing in retinal ganglion cells (GCs). We simultaneously recorded the firing of GCs in the goldfish isolated retina using a multi-electrode array, and classified each GC based on the temporal profile of its receptive field (RF). A moving target that accompanied the global motion (simulating a saccade following a period of fixational eye movements) modulated the RF properties and evoked synchronized and correlated firing among local clusters of the specific GCs. Our findings provide a novel concept for retinal information processing during eye movements.

  15. Effect of eye NGF administration on two animal models of retinal ganglion cells degeneration

    Directory of Open Access Journals (Sweden)

    Valeria Colafrancesco

    2011-01-01

    Full Text Available The aim of this study was to investigate the effect of nerve growth factor (NGF administration on retinal ganglion cells (RGCs in experimentally induced glaucoma (GL and diabetic retinopathy (DR. GL was induced in adult rats by injection of hypertonic saline into the episcleral vein of the eye and diabetes (DT was induced by administration of streptozoticin. Control and experimental rats were treated daily with either ocular application of NGF or vehicle solution. We found that both animal models present a progressive degeneration of RGCs and changing NGF and VEGF levels in the retina and optic nerve. We then proved that NGF eye drop administration exerts a protective effect on these models of retinal degeneration. In brief, our findings indicate that NGF can play a protective role against RGC degeneration occurring in GL and DR and suggest that ocular NGF administration might be an effective pharmacological approach.

  16. Rhythmic ganglion cell activity in bleached and blind adult mouse retinas.

    Science.gov (United States)

    Menzler, Jacob; Channappa, Lakshmi; Zeck, Guenther

    2014-01-01

    In retinitis pigmentosa--a degenerative disease which often leads to incurable blindness--the loss of photoreceptors deprives the retina from a continuous excitatory input, the so-called dark current. In rodent models of this disease this deprivation leads to oscillatory electrical activity in the remaining circuitry, which is reflected in the rhythmic spiking of retinal ganglion cells (RGCs). It remained unclear, however, if the rhythmic RGC activity is attributed to circuit alterations occurring during photoreceptor degeneration or if rhythmic activity is an intrinsic property of healthy retinal circuitry which is masked by the photoreceptor's dark current. Here we tested these hypotheses by inducing and analysing oscillatory activity in adult healthy (C57/Bl6) and blind mouse retinas (rd10 and rd1). Rhythmic RGC activity in healthy retinas was detected upon partial photoreceptor bleaching using an extracellular high-density multi-transistor-array. The mean fundamental spiking frequency in bleached retinas was 4.3 Hz; close to the RGC rhythm detected in blind rd10 mouse retinas (6.5 Hz). Crosscorrelation analysis of neighbouring wild-type and rd10 RGCs (separation distance rhythmic RGC spiking in these retinas is driven by a network of presynaptic neurons. The inhibition of glutamatergic ganglion cell input or the inhibition of gap junctional coupling abolished the rhythmic pattern. In rd10 and rd1 retinas the presynaptic network leads to local field potentials, whereas in bleached retinas additional pharmacological disinhibition is required to achieve detectable field potentials. Our results demonstrate that photoreceptor bleaching unmasks oscillatory activity in healthy retinas which shares many features with the functional phenotype detected in rd10 retinas. The quantitative physiological differences advance the understanding of the degeneration process and may guide future rescue strategies.

  17. Cobalamin C Deficiency Shows a Rapidly Progressing Maculopathy With Severe Photoreceptor and Ganglion Cell Loss

    Science.gov (United States)

    Bonafede, Lucas; Ficicioglu, Can H.; Serrano, Leona; Han, Grace; Morgan, Jessica I. W.; Mills, Monte D.; Forbes, Brian J.; Davidson, Stefanie L.; Binenbaum, Gil; Kaplan, Paige B.; Nichols, Charles W.; Verloo, Patrick; Leroy, Bart P.; Maguire, Albert M.; Aleman, Tomas S.

    2015-01-01

    Purpose To describe in detail the retinal structure and function of a group of patients with cobalamin C (cblC) disease. Methods Patients (n = 11, age 4 months to 15 years) with cblC disease (9/11, early onset) diagnosed by newborn screening underwent complete ophthalmic examinations, fundus photography, near-infrared reflectance imaging, and spectral-domain optical coherence tomography (SD-OCT). Electroretinograms (ERGs) were performed in a subset of patients. Results Patients carried homozygous or compound heterozygote mutations in the methylmalonic aciduria and homocystinuria type C (MMACHC) gene. Late-onset patients had a normal exam. All early-onset patients showed a maculopathy; older subjects had a retina-wide degeneration (n = 4; >7 years of age). In general, retinal changes were first observed before 1 year of age and progressed within months to a well-established maculopathy. Pseudocolobomas were documented in three patients. Measurable visual acuities ranged from 20/200 to 20/540. Nystagmus was present in 8/11 patients; 5/6 patients had normal ERGs; 1/6 had reduced rod-mediated responses. Spectral-domain OCT showed macular thinning, with severe ganglion cell layer (GCL) and outer nuclear layer (ONL) loss. Inner retinal thickening was observed in areas of total GCL/ONL loss. A normal lamination pattern in the peripapillary nasal retina was often seen despite severe central and/or retina-wide disease. Conclusions Patients with early-onset cblC and MMACHC mutations showed an early-onset, unusually fast-progressing maculopathy with severe central ONL and GCL loss. An abnormally thickened inner retina supports a remodeling response to both photoreceptor and ganglion cell degeneration and/or an interference with normal development in early-onset cblC. PMID:26658511

  18. Coding properties of three intrinsically distinct retinal ganglion cells under periodic stimuli: a computational study

    Directory of Open Access Journals (Sweden)

    Lei Wang

    2016-09-01

    Full Text Available As the sole output neurons in the retina, ganglion cells play significant roles in transforming visual information into spike trains, and then transmitting them to the higher visual centers. However, coding strategies that retinal ganglion cells (RGCs adopt to accomplish these processes are not completely clear yet. To clarify these issues, we investigate the coding properties of three types of RGCs (repetitive spiking, tonic firing, and phasic firing by two different measures (spike-rate and spike-latency. Model results show that for periodic stimuli, repetitive spiking RGC and tonic RGC exhibit similar spike-rate patterns. Their spike-rates decrease gradually with increased stimulus frequency, moreover, variation of stimulus amplitude would change the two RGCs’ spike-rate patterns. For phasic RGC, it activates strongly at medium levels of frequency when the stimulus amplitude is low. While if high stimulus amplitude is applied, phasic RGC switches to respond strongly at low frequencies. These results suggest that stimulus amplitude is a prominent factor in regulating RGCs in encoding periodic signals. Similar conclusions can be drawn when analyzes spike-latency patterns of the three RGCs. More importantly, the above phenomena can be accurately reproduced by Hodgkin’s three classes of neurons, indicating that RGCs can perform the typical three classes of firing dynamics, depending on the distinctions of ion channel densities. Consequently, model results from the three RGCs may be not specific, but can also applicable to neurons in other brain regions which exhibit part(s or all of the Hodgkin’s three excitabilities.

  19. Endothelin B receptors contribute to retinal ganglion cell loss in a rat model of glaucoma.

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    Alena Z Minton

    Full Text Available Glaucoma is an optic neuropathy, commonly associated with elevated intraocular pressure (IOP characterized by optic nerve degeneration, cupping of the optic disc, and loss of retinal ganglion cells which could lead to loss of vision. Endothelin-1 (ET-1 is a 21-amino acid vasoactive peptide that plays a key role in the pathogenesis of glaucoma; however, the receptors mediating these effects have not been defined. In the current study, endothelin B (ET(B receptor expression was assessed in vivo, in the Morrison's ocular hypertension model of glaucoma in rats. Elevation of IOP in Brown Norway rats produced increased expression of ET(B receptors in the retina, mainly in retinal ganglion cells (RGCs, nerve fiber layer (NFL, and also in the inner plexiform layer (IPL and inner nuclear layer (INL. To determine the role of ET(B receptors in neurodegeneration, Wistar-Kyoto wild type (WT and ET(B receptor-deficient (KO rats were subjected to retrograde labeling with Fluoro-Gold (FG, following which IOP was elevated in one eye while the contralateral eye served as control. IOP elevation for 4 weeks in WT rats caused an appreciable loss of RGCs, which was significantly attenuated in KO rats. In addition, degenerative changes in the optic nerve were greatly reduced in KO rats compared to those in WT rats. Taken together, elevated intraocular pressure mediated increase in ET(B receptor expression and its activation may contribute to a decrease in RGC survival as seen in glaucoma. These findings raise the possibility of using endothelin receptor antagonists as neuroprotective agents for the treatment of glaucoma.

  20. Ganglion cell loss in relation to visual disability in multiple sclerosis.

    Science.gov (United States)

    Walter, Scott D; Ishikawa, Hiroshi; Galetta, Kristin M; Sakai, Reiko E; Feller, Daniel J; Henderson, Sam B; Wilson, James A; Maguire, Maureen G; Galetta, Steven L; Frohman, Elliot; Calabresi, Peter A; Schuman, Joel S; Balcer, Laura J

    2012-06-01

    We used high-resolution spectral-domain optical coherence tomography (SD-OCT) with retinal segmentation to determine how ganglion cell loss relates to history of acute optic neuritis (ON), retinal nerve fiber layer (RNFL) thinning, visual function, and vision-related quality of life (QOL) in multiple sclerosis (MS). Cross-sectional study. A convenience sample of patients with MS (n = 122; 239 eyes) and disease-free controls (n = 31; 61 eyes). Among MS eyes, 87 had a history of ON before enrollment. The SD-OCT images were captured using Macular Cube (200×200 or 512×128) and ONH Cube 200×200 protocols. Retinal layer segmentation was performed using algorithms established for glaucoma studies. Thicknesses of the ganglion cell layer/inner plexiform layer (GCL+IPL), RNFL, outer plexiform/inner nuclear layers (OPL+INL), and outer nuclear/photoreceptor layers (ONL+PRL) were measured and compared in MS versus control eyes and MS ON versus non-ON eyes. The relation between changes in macular thickness and visual disability was also examined. The OCT measurements of GCL+IPL and RNFL thickness; high contrast visual acuity (VA); low-contrast letter acuity (LCLA) at 2.5% and 1.25% contrast; on the 25-item National Eye Institute Visual Function Questionnaire (NEI-VFQ-25) and 10-Item Neuro-Ophthalmic Supplement composite score. Macular RNFL and GCL+IPL were significantly decreased in MS versus control eyes (Pvisual function and vision-specific QOL in MS, and may serve as a useful structural marker of disease. Our findings parallel those of magnetic resonance imaging studies that show gray matter disease is a marker of neurologic disability in MS. Proprietary or commercial disclosure may be found after the references. Copyright © 2012 American Academy of Ophthalmology. Published by Elsevier Inc. All rights reserved.

  1. Pathological evaluation of ganglion cells in biopsies from upper side of the dentate line in patients with perianal problems

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    Marjan Joudi

    2014-07-01

    Full Text Available Introduction: Constipation is one of the most common complaints of individuals, which may present with complication like hemorrhoid and fissure. Hirschsprung is a disease presenting with chronic constipation and its diagnosis may be delayed until adulthood. It is diagnosed by biopsies from anorectal transitional zone. This study aimed to evaluate the association between Hirschsprung and anorectal problems. Method: Sixty three patients with anorectal problems who underwent surgery enrolled in this study. Some consecutive biopsies were obtained from anal canal at 2, 4 and 6 cm above the dentate line. Biopsies were assessed for ganglion cells changes. Patients' data and biopsies results were analyzed with SPSS version18. Results: Out of 63 patients 29 (46 % patients were female and 34 (54 % were male with the mean of 32.65 ± 13.73 years. Fifty six (73 % patients complained from constipation with the mean time of 57.65 ± 45.21 months. Aganglionic zone were reported in six patients with the mean length of 43.33 mm. There was not any relation between anal ganglion cells pathology and constipation (p=0.363, but there was a significant relation between duration of constipation and pathologic changes (p=0.001. The ratio of constipation duration to age was related to anal ganglion cell pathology (p=0.001. Hemorrhoid degree was also affected anal ganglion cells pathology (p=0.037. Conclusion: The relation between Hirschsprung's disease and anorectal problems in adults were significant. The pathologic findings were more presented in younger patients, and those with longer history of constipation and lower degree hemorrhoids. Key words: Anal ganglion cells, Hemorrhoids, Constipation  

  2. The ciliary margin zone of the mammalian retina generates retinal ganglion cells

    Science.gov (United States)

    Marcucci, Florencia; Murcia-Belmonte, Veronica; Coca, Yaiza; Ferreiro-Galve, Susana; Wang, Qing; Kuwajima, Takaaki; Khalid, Sania; Ross, M. Elizabeth; Herrera, Eloisa; Mason, Carol

    2016-01-01

    Summary The retina of lower vertebrates grows continuously by integrating new neurons generated from progenitors in the ciliary margin zone (CMZ). Whether the mammalian CMZ provides the neural retina with retinal cells is controversial. Live-imaging of embryonic retina expressing eGFP in the CMZ shows that cells migrate laterally from the CMZ to the neural retina where differentiated retinal ganglion cells (RGCs) reside. As Cyclin D2, a cell-cycle regulator, is enriched in ventral CMZ, we analyzed Cyclin D2−/− mice to test whether the CMZ is a source of retinal cells. Neurogenesis is diminished in Cyclin D2 mutants, leading to a reduction of RGCs in the ventral retina. In line with these findings, in the albino retina, the decreased production of ipsilateral RGCs is correlated with fewer Cyclin D2+ cells. Together, these results implicate the mammalian CMZ as a neurogenic site that produces RGCs and whose proper generation depends on Cyclin D2 activity. PMID:28009286

  3. Method to investigate temporal dynamics of ganglion and other retinal cells in the living human eye

    Science.gov (United States)

    Kurokawa, Kazuhiro; Liu, Zhuolin; Crowell, James; Zhang, Furu; Miller, Donald T.

    2018-02-01

    The inner retina is critical for visual processing, but much remains unknown about its neural circuitry and vulnerability to disease. A major bottleneck has been our inability to observe the structure and function of the cells composing these retinal layers in the living human eye. Here, we present a noninvasive method to observe both structural and functional information. Adaptive optics optical coherence tomography (AO-OCT) is used to resolve the inner retinal cells in all three dimensions and novel post processing algorithms are applied to extract structure and physiology down to the cellular level. AO-OCT captured the 3D mosaic of individual ganglion cell somas, retinal nerve fiber bundles of micron caliber, and microglial cells, all in exquisite detail. Time correlation analysis of the AO-OCT videos revealed notable temporal differences between the principal layers of the inner retina. The GC layer was more dynamic than the nerve fiber and inner plexiform layers. At the cellular level, we applied a customized correlation method to individual GCL somas, and found a mean time constant of activity of 0.57 s and spread of +/-0.1 s suggesting a range of physiological dynamics even in the same cell type. Extending our method to slower dynamics (from minutes to one year), time-lapse imaging and temporal speckle contrast revealed appendage and soma motion of resting microglial cells at the retinal surface.

  4. Dose-dependent effects of ouabain on spiral ganglion neurons and Schwann cells in mouse cochlea.

    Science.gov (United States)

    Zhang, Zhi-Jian; Guan, Hong-Xia; Yang, Kun; Xiao, Bo-Kui; Liao, Hua; Jiang, Yang; Zhou, Tao; Hua, Qing-Quan

    2017-10-01

    This study aimed in fully investigating the toxicities of ouabain to mouse cochlea and the related cellular environment, and providing an optimal animal model system for cell transplantation in the treatment of auditory neuropathy (AN) and sensorineural hearing loss (SNHL). Different dosages of ouabain were applied to mouse round window. The auditory brainstem responses and distortion product otoacoustic emissions were used to evaluate the cochlear function. The immunohistochemical staining and cochlea surface preparation were performed to detect the spiral ganglion neurons (SGNs), Schwann cells and hair cells. Ouabain at the dosages of 0.5 mM, 1 mM and 3 mM selectively and permanently destroyed SGNs and their functions, while leaving the hair cells relatively intact. Ouabain at 3 mM resulted in the most severe SGNs loss and induced significant loss of Schwann cells started as early as 7 days and with further damages at 14 and 30 days after ouabain exposure. The application of ouabain to mouse round window induces damages of SGNs and Schwann cells in a dose- and time-dependent manner, this study established a reliable and accurate animal model system of AN and SNHL.

  5. Hepatocyte growth factor promotes long-term survival and axonal regeneration of retinal ganglion cells after optic nerve injury: comparison with CNTF and BDNF.

    Science.gov (United States)

    Wong, Wai-Kai; Cheung, Anny Wan-Suen; Yu, Sau-Wai; Sha, Ou; Cho, Eric Yu Pang

    2014-10-01

    Different trophic factors are known to promote retinal ganglion cell survival and regeneration, but each had their own limitations. We report that hepatocyte growth factor (HGF) confers distinct advantages in supporting ganglion cell survival and axonal regeneration, when compared to two well-established trophic factors ciliary neurotrophic factor (CNTF) and brain-derived neurotrophic factor (BDNF). Ganglion cells in adult hamster were injured by cutting the optic nerve. HGF, CNTF, or BDNF was injected at different dosages intravitreally after injury. Ganglion cell survival was quantified at 7, 14, or 28 days postinjury. Peripheral nerve (PN) grafting to the cut optic nerve of the growth factor-injected eye was performed either immediately after injury or delayed until 7 days post-injury. Expression of heat-shock protein 27 and changes in microglia numbers were quantified in different growth factor groups. The cellular distribution of c-Met in the retina was examined by anti-c-Met immunostaining. Hepatocyte Growth Factor (HGF) was equally potent as BDNF in promoting short-term survival (up to 14 days post-injury) and also supported survival at 28 days post-injury when ganglion cells treated by CNTF or BDNF failed to be sustained. When grafting was performed without delay, HGF stimulated twice the number of axons to regenerate compared with control but was less potent than CNTF. However, in PN grafting delayed for 7 days after optic nerve injury, HGF maintained a better propensity of ganglion cells to regenerate than CNTF. Unlike CNTF, HGF application did not increase HSP27 expression in ganglion cells. Microglia proliferation was prolonged in HGF-treated retinas compared with CNTF or BDNF. C-Met was localized to both ganglion cells and Muller cells, suggesting HGF could be neuroprotective via interacting with both neurons and glia. Compared with CNTF or BDNF, HGF is advantageous in sustaining long-term ganglion cell survival and their propensity to respond to

  6. The apoptosis of CHO cells induced by X-rays

    International Nuclear Information System (INIS)

    Lu Zhaohong; Zhao Jingyong; Zhu Mingqing; Shi Xijin; Wang Chunlei

    2004-01-01

    The work is to study the mechanism of toxic effects on reproductive system and apoptosis of Chinese hamster ovary (CHO) cells induced by X-rays. CHO cell was exposed to X-rays 2 to 20 Gy. Apoptosis and morphological changes of the cells were observed by fluorescent microscopy and flow cytometry analyzer with double staining with Annexin V/PI. The apoptosis could be observed at 24, 48 and 72h after the exposure, but it was more obvious 48 and 72 h after the exposure. Rate of the apoptosis increased along with radiation dose were elevated. Some morphological changes, such as irregular agglomerate of chromatins, pycnosis and periphery distribution of nuclei, crescent-moon-like cells, small apoptosis body, were observed. Radiation results DNA damage in the CHO cells, and the damage cannot be repaired, hence the induced cell apoptosis. (authors)

  7. Cyclin-dependent kinases regulate apoptosis of intestinal epithelial cells

    Science.gov (United States)

    Bhattacharya, Sujoy; Ray, Ramesh M.; Johnson, Leonard R.

    2014-01-01

    Homeostasis of the gastrointestinal epithelium is dependent upon a balance between cell proliferation and apoptosis. Cyclin-dependent kinases (Cdks) are well known for their role in cell proliferation. Previous studies from our group have shown that polyamine-depletion of intestinal epithelial cells (IEC-6) decreases cyclin-dependent kinase 2 (Cdk2) activity, increases p53 and p21Cip1 protein levels, induces G1 arrest, and protects cells from camptothecin (CPT)-induced apoptosis. Although emerging evidence suggests that members of the Cdk family are involved in the regulation of apoptosis, their roles directing apoptosis of IEC-6 cells are not known. In this study, we report that inhibition of Cdk1, 2, and 9 (with the broad range Cdk inhibitor, AZD5438) in proliferating IEC-6 cells triggered DNA damage, activated p53 signaling, inhibited proliferation, and induced apoptosis. By contrast, inhibition of Cdk2 (with NU6140) increased p53 protein and activity, inhibited proliferation, but had no effect on apoptosis. Notably, AZD5438 sensitized, whereas, NU6140 rescued proliferating IEC-6 cells from CPT-induced apoptosis. However, in colon carcinoma (Caco2) cells with mutant p53, treatment with either AZD5438 or NU6140 blocked proliferation, albeit more robustly with AZD5438. Both Cdk inhibitors induced apoptosis in Caco2 cells in a p53-independent manner. In serum starved quiescent IEC-6 cells, both AZD5438 and NU6140 decreased TNF- /CPT-induced activation of p53 and, consequently, rescued cells from apoptosis, indicating that sustained Cdk activity is required for apoptosis of quiescent cells. Furthermore, AZD5438 partially reversed the protective effect of polyamine depletion whereas NU6140 had no effect. Together, these results demonstrate that Cdks possess opposing roles in the control of apoptosis in quiescent and proliferating cells. In addition, Cdk inhibitors uncouple proliferation from apoptosis in a p53-dependent manner. PMID:24242917

  8. Aspartame-induced apoptosis in PC12 cells

    OpenAIRE

    Horio, Yukari; Sun, Yongkun; Liu, Chuang; Saito, Takeshi; Kurasaki, Masaaki

    2014-01-01

    Aspartame is an artificial sweetner added to many low-calorie foods. The safety of aspartame remains controversial even though there are many studies on its risks. In this study, to understand the physiological effects of trace amounts of artificial sweetners on cells, the effects of aspartame on apoptosis were investigated using a PC12 cell system. In addition, the mechanism of apoptosis induced by aspartame in PC12 cells and effects on apoptotic factors such as cytochrome c, apoptosis-induc...

  9. Changes in intrinsic excitability of ganglion cells in degenerated retinas of RCS rats

    Directory of Open Access Journals (Sweden)

    Yi-Ming Ren

    2018-05-01

    Full Text Available AIM: To evaluate the intrinsic excitability of retinal ganglion cells (RGCs in degenerated retinas. METHODS: The intrinsic excitability of various morphologically defined RGC types using a combination of patch-clamp recording and the Lucifer yellow tracer in retinal whole-mount preparations harvested from Royal College of Surgeons (RCS rats, a common retinitis pigmentosa (RP model, in a relatively late stage of retinal degeneration (P90 were investigated. Several parameters of RGC morphologies and action potentials (APs were measured and compared to those of non-dystrophic control rats, including dendritic stratification, dendritic field diameter, peak amplitude, half width, resting membrane potential, AP threshold, depolarization to threshold, and firing rates. RESULTS: Compared with non-dystrophic control RGCs, more depolarizations were required to reach the AP threshold in RCS RGCs with low spontaneous spike rates and in RCS OFF cells (especially A2o cells, and RCS RGCs maintained their dendritic morphologies, resting membrane potentials and capabilities to generate APs. CONCLUSION: RGCs are relatively well preserved morphologically and functionally, and some cells are more susceptible to decreased excitability during retinal degeneration. These findings provide valuable considerations for optimizing RP therapeutic strategies.

  10. Regulatory mechanisms of apoptosis in regularly dividing cells

    Directory of Open Access Journals (Sweden)

    Ribal S Darwish

    2010-08-01

    Full Text Available Ribal S DarwishDepartment of Anesthesiology, Division of Critical Care Medicine, University of Maryland Medical Center, Baltimore, Maryland, USAAbstract: The balance between cell survival and death is essential for normal development and homeostasis of organisms. Apoptosis is a distinct type of cell death with ultrastructural features that are consistent with an active, inherently controlled process. Abnormalities and ­dysregulation of apoptosis contribute to the pathophysiology of multiple disease processes. Apoptosis is strictly regulated by several positive and negative feedback mechanisms that regulate cell death and determine the final outcome after cell exposure to apoptotic stimuli. Mitochondria and caspases are central components of the regulatory mechanisms of ­apoptosis. Recently, noncaspase pathways of apoptosis have been explored through the studies of ­apoptosis-inducing factor and endonuclease G. Multiple difficulties in the apoptosis research relate to apoptosis detection and imaging. This article reviews current understanding of the regulatory mechanisms of apoptosis.Keywords: caspases, apoptosis-inducing factor, apoptosis inhibitory proteins, cytochrome c, mitochondria 

  11. Melanopsin expressing human retinal ganglion cells: Subtypes, distribution, and intraretinal connectivity.

    Science.gov (United States)

    Hannibal, Jens; Christiansen, Anders Tolstrup; Heegaard, Steffen; Fahrenkrug, Jan; Kiilgaard, Jens Folke

    2017-06-01

    Intrinsically photosensitive retinal ganglion cells (ipRGCs) expressing the photopigment melanopsin belong to a heterogenic population of RGCs which regulate the circadian clock, masking behavior, melatonin suppression, the pupillary light reflex, and sleep/wake cycles. The different functions seem to be associated to different subtypes of melanopsin cells. In rodents, subtype classification has associated subtypes to function. In primate and human retina such classification has so far, not been applied. In the present study using antibodies against N- and C-terminal parts of human melanopsin, confocal microscopy and 3D reconstruction of melanopsin immunoreactive (-ir) RGCs, we applied the criteria used in mouse on human melanopsin-ir RGCs. We identified M1, displaced M1, M2, and M4 cells. We found two other subtypes of melanopsin-ir RGCs, which were named "gigantic M1 (GM1)" and "gigantic displaced M1 (GDM1)." Few M3 cells and no M5 subtypes were labeled. Total cell counts from one male and one female retina revealed that the human retina contains 7283 ± 237 melanopsin-ir (0.63-0.75% of the total number of RGCs). The melanopsin subtypes were unevenly distributed. Most significant was the highest density of M4 cells in the nasal retina. We identified input to the melanopsin-ir RGCs from AII amacrine cells and directly from rod bipolar cells via ribbon synapses in the innermost ON layer of the inner plexiform layer (IPL) and from dopaminergic amacrine cells and GABAergic processes in the outermost OFF layer of the IPL. The study characterizes a heterogenic population of human melanopsin-ir RGCs, which most likely are involved in different functions. © 2017 Wiley Periodicals, Inc.

  12. Interspike Interval Based Filtering of Directional Selective Retinal Ganglion Cells Spike Trains

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    Aurel Vasile Martiniuc

    2012-01-01

    Full Text Available The information regarding visual stimulus is encoded in spike trains at the output of retina by retinal ganglion cells (RGCs. Among these, the directional selective cells (DSRGC are signaling the direction of stimulus motion. DSRGCs' spike trains show accentuated periods of short interspike intervals (ISIs framed by periods of isolated spikes. Here we use two types of visual stimulus, white noise and drifting bars, and show that short ISI spikes of DSRGCs spike trains are more often correlated to their preferred stimulus feature (that is, the direction of stimulus motion and carry more information than longer ISI spikes. Firstly, our results show that correlation between stimulus and recorded neuronal response is best at short ISI spiking activity and decrease as ISI becomes larger. We then used grating bars stimulus and found that as ISI becomes shorter the directional selectivity is better and information rates are higher. Interestingly, for the less encountered type of DSRGC, known as ON-DSRGC, short ISI distribution and information rates revealed consistent differences when compared with the other directional selective cell type, the ON-OFF DSRGC. However, these findings suggest that ISI-based temporal filtering integrates a mechanism for visual information processing at the output of retina toward higher stages within early visual system.

  13. Loss of Melanopsin-Expressing Retinal Ganglion Cells in Severely Staged Glaucoma Patients

    DEFF Research Database (Denmark)

    Obara, Elisabeth Anne; Hannibal, Jens; Heegaard, Steffen

    2016-01-01

    Purpose: Multiple studies have shown overwhelming evidence supporting the impairment of melanopsin function due to glaucoma. However, few studies have been carried out in humans analyzing the histology of melanopsin-expressing retinal ganglion cells (mRGCs) in retinas with glaucoma. The aim...... of this study was to analyze the pattern of expression of mRGCs relative to RGCs in the normal retina and retinas harboring varying stages of glaucoma. Methods: Paraffin-embedded human donor eyes with glaucoma (n = 11) and age-matched controls (n = 10) were obtained from Department of Pathology at Rigshospital...... difference was observed in mRGC expression in the normal retinas and mild-staged retinas with glaucoma; the densities of mRGCs were 3.08 ± 0.47 and 3.00 ± 0.13 cell counts/mm2, respectively. However, the severely staged retinas with glaucoma showed a significant loss in mRGCs density, 1.09 ± 0.35 cell counts...

  14. Isolation of Primary Murine Retinal Ganglion Cells (RGCs) by Flow Cytometry.

    Science.gov (United States)

    Chintalapudi, Sumana R; Patel, Need N; Goldsmith, Zachary K; Djenderedjian, Levon; Wang, Xiang Di; Marion, Tony N; Jablonski, Monica M; Morales-Tirado, Vanessa M

    2017-07-05

    Neurodegenerative diseases often have a devastating impact on those affected. Retinal ganglion cell (RGC) loss is implicated in an array of diseases, including diabetic retinopathy and glaucoma, in addition to normal aging. Despite their importance, RGCs have been extremely difficult to study until now due in part to the fact that they comprise only a small percentage of the wide variety of cells in the retina. In addition, current isolation methods use intracellular markers to identify RGCs, which produce non-viable cells. These techniques also involve lengthy isolation protocols, so there is a lack of practical, standardized, and dependable methods to obtain and isolate RGCs. This work describes an efficient, comprehensive, and reliable method to isolate primary RGCs from mice retinae using a protocol based on both positive and negative selection criteria. The presented methods allow for the future study of RGCs, with the goal of better understanding the major decline in visual acuity that results from the loss of functional RGCs in neurodegenerative diseases.

  15. Intraocular Injection of ES Cell-Derived Neural Progenitors Improve Visual Function in Retinal Ganglion Cell-Depleted Mouse Models

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    Mundackal S. Divya

    2017-09-01

    Full Text Available Retinal ganglion cell (RGC transplantation is a promising strategy to restore visual function resulting from irreversible RGC degeneration occurring in glaucoma or inherited optic neuropathies. We previously demonstrated FGF2 induced differentiation of mouse embryonic stem cells (ESC to RGC lineage, capable of retinal ganglion cell layer (GCL integration upon transplantation. Here, we evaluated possible improvement of visual function by transplantation of ES cell derived neural progenitors in RGC depleted glaucoma mice models. ESC derived neural progenitors (ES-NP were transplanted into N-Methyl-D-Aspartate (NMDA injected, RGC-ablated mouse models and a pre-clinical glaucoma mouse model (DBA/2J having sustained higher intra ocular pressure (IOP. Visual acuity and functional integration was evaluated by behavioral experiments and immunohistochemistry, respectively. GFP-expressing ES-NPs transplanted in NMDA-injected RGC-depleted mice differentiated into RGC lineage and possibly integrating into GCL. An improvement in visual acuity was observed after 2 months of transplantation, when compared to the pre-transplantation values. Expression of c-Fos in the transplanted cells, upon light induction, further suggests functional integration into the host retinal circuitry. However, the transplanted cells did not send axonal projections into optic nerve. Transplantation experiments in DBA/2J mouse showed no significant improvement in visual functions, possibly due to both host and transplanted retinal cell death which could be due to an inherent high IOP. We showed that, ES NPs transplanted into the retina of RGC-ablated mouse models could survive, differentiate to RGC lineage, and possibly integrate into GCL to improve visual function. However, for the survival of transplanted cells in glaucoma, strategies to control the IOP are warranted.

  16. Hyperthermia: an effective strategy to induce apoptosis in cancer cells.

    Science.gov (United States)

    Ahmed, Kanwal; Tabuchi, Yoshiaki; Kondo, Takashi

    2015-11-01

    Heat has been used as a medicinal and healing modality throughout human history. The combination of hyperthermia (HT) with radiation and anticancer agents has been used clinically and has shown positive results to a certain extent. However, the clinical results of HT treatment alone have been only partially satisfactory. Cell death following HT treatment is a function of both temperature and treatment duration. HT induces cancer cell death through apoptosis; the degree of apoptosis and the apoptotic pathway vary in different cancer cell types. HT-induced reactive oxygen species production are responsible for apoptosis in various cell types. However, the underlying mechanism of signal transduction and the genes related to this process still need to be elucidated. In this review, we summarize the molecular mechanism of apoptosis induced by HT, enhancement of heat-induced apoptosis, and the genetic network involved in HT-induced apoptosis.

  17. Aspartame-induced apoptosis in PC12 cells.

    Science.gov (United States)

    Horio, Yukari; Sun, Yongkun; Liu, Chuang; Saito, Takeshi; Kurasaki, Masaaki

    2014-01-01

    Aspartame is an artificial sweetner added to many low-calorie foods. The safety of aspartame remains controversial even though there are many studies on its risks. In this study, to understand the physiological effects of trace amounts of artificial sweetners on cells, the effects of aspartame on apoptosis were investigated using a PC12 cell system. In addition, the mechanism of apoptosis induced by aspartame in PC12 cells and effects on apoptotic factors such as cytochrome c, apoptosis-inducing factor, and caspase family proteins were studied by Western blotting and RT-PCR. Aspartame-induced apoptosis in PC12 cells in a dose-dependent manner. In addition, aspartame exposure increased the expressions of caspases 8 and 9, and cytochrome c. These results indicate that aspartame induces apoptosis mainly via mitochondrial pathway involved in apoptosis due to oxigen toxicity. Copyright © 2013 Elsevier B.V. All rights reserved.

  18. Retinal ganglion cells in the eastern newt Notophthalmus viridescens: topography, morphology, and diversity.

    Science.gov (United States)

    Pushchin, Igor I; Karetin, Yuriy A

    2009-10-20

    The topography and morphology of retinal ganglion cells (RGCs) in the eastern newt were studied. Cells were retrogradely labeled with tetramethylrhodamine-conjugated dextran amines or horseradish peroxidase and examined in retinal wholemounts. Their total number was 18,025 +/- 3,602 (mean +/- SEM). The spatial density of RGCs varied from 2,100 cells/mm(2) in the retinal periphery to 4,500 cells/mm(2) in the dorsotemporal retina. No prominent retinal specializations were found. The spatial resolution estimated from the spatial density of RGCs varied from 1.4 cycles per degree in the periphery to 1.95 cycles per degree in the region of the peak RGC density. A sample of 68 cells was camera lucida drawn and subjected to quantitative analysis. A total of 21 parameters related to RGC morphology and stratification in the retina were estimated. Partitionings obtained by using different clustering algorithms combined with automatic variable weighting and dimensionality reduction techniques were compared, and an effective solution was found by using silhouette analysis. A total of seven clusters were identified and associated with potential cell types. Kruskal-Wallis ANOVA-on-Ranks with post hoc Mann-Whitney U tests showed significant pairwise between-cluster differences in one or more of the clustering variables. The average silhouette values of the clusters were reasonably high, ranging from 0.52 to 0.79. Cells assigned to the same cluster displayed similar morphology and stratification in the retina. The advantages and limitations of the methodology adopted are discussed. The present classification is compared with known morphological and physiological RGC classifications in other salamanders.

  19. Cellular response after irradiation: Cell cycle control and apoptosis

    International Nuclear Information System (INIS)

    Siles, E.; Valenzuela, M.T.; Nunez, M.I.; Guerrero, R.; Villalobos, M.; Ruiz de Almodovar, J.M.

    1997-01-01

    The importance of apoptotic death was assessed in a set of experiments, involving eight human tumour cell lines (breast cancer, bladder carcinoma, medulloblastoma). Various aspects of the quantitative study of apoptosis and methods based on the detection of DNA fragmentation (in situ tailing and comet assay) are described and discussed. Data obtained support the hypothesis that apoptosis is not crucial for cellular radiosensitivity and that the relationship between p53 functionality or clonogenic survival and apoptosis may bee cell type specific. (author)

  20. Retinal ganglion cells with distinct directional preferences differ in molecular identity, structure, and central projections.

    Science.gov (United States)

    Kay, Jeremy N; De la Huerta, Irina; Kim, In-Jung; Zhang, Yifeng; Yamagata, Masahito; Chu, Monica W; Meister, Markus; Sanes, Joshua R

    2011-05-25

    The retina contains ganglion cells (RGCs) that respond selectively to objects moving in particular directions. Individual members of a group of ON-OFF direction-selective RGCs (ooDSGCs) detect stimuli moving in one of four directions: ventral, dorsal, nasal, or temporal. Despite this physiological diversity, little is known about subtype-specific differences in structure, molecular identity, and projections. To seek such differences, we characterized mouse transgenic lines that selectively mark ooDSGCs preferring ventral or nasal motion as well as a line that marks both ventral- and dorsal-preferring subsets. We then used the lines to identify cell surface molecules, including Cadherin 6, CollagenXXVα1, and Matrix metalloprotease 17, that are selectively expressed by distinct subsets of ooDSGCs. We also identify a neuropeptide, CART (cocaine- and amphetamine-regulated transcript), that distinguishes all ooDSGCs from other RGCs. Together, this panel of endogenous and transgenic markers distinguishes the four ooDSGC subsets. Patterns of molecular diversification occur before eye opening and are therefore experience independent. They may help to explain how the four subsets obtain distinct inputs. We also demonstrate differences among subsets in their dendritic patterns within the retina and their axonal projections to the brain. Differences in projections indicate that information about motion in different directions is sent to different destinations.

  1. A Pixel-Encoder Retinal Ganglion Cell with Spatially Offset Excitatory and Inhibitory Receptive Fields

    Directory of Open Access Journals (Sweden)

    Keith P. Johnson

    2018-02-01

    Full Text Available The spike trains of retinal ganglion cells (RGCs are the only source of visual information to the brain. Here, we genetically identify an RGC type in mice that functions as a pixel encoder and increases firing to light increments (PixON-RGC. PixON-RGCs have medium-sized dendritic arbors and non-canonical center-surround receptive fields. From their receptive field center, PixON-RGCs receive only excitatory input, which encodes contrast and spatial information linearly. From their receptive field surround, PixON-RGCs receive only inhibitory input, which is temporally matched to the excitatory center input. As a result, the firing rate of PixON-RGCs linearly encodes local image contrast. Spatially offset (i.e., truly lateral inhibition of PixON-RGCs arises from spiking GABAergic amacrine cells. The receptive field organization of PixON-RGCs is independent of stimulus wavelength (i.e., achromatic. PixON-RGCs project predominantly to the dorsal lateral geniculate nucleus (dLGN of the thalamus and likely contribute to visual perception.

  2. Edaravone suppresses retinal ganglion cell death in a mouse model of normal tension glaucoma

    Science.gov (United States)

    Akaiwa, Kei; Namekata, Kazuhiko; Azuchi, Yuriko; Guo, Xiaoli; Kimura, Atsuko; Harada, Chikako; Mitamura, Yoshinori; Harada, Takayuki

    2017-01-01

    Glaucoma, one of the leading causes of irreversible blindness, is characterized by progressive degeneration of optic nerves and retinal ganglion cells (RGCs). In the mammalian retina, excitatory amino-acid carrier 1 (EAAC1) is expressed in neural cells, including RGCs. Loss of EAAC1 leads to RGC degeneration without elevated intraocular pressure (IOP) and exhibits glaucomatous pathology including glutamate neurotoxicity and oxidative stress. In the present study, we found that edaravone, a free radical scavenger that is used for treatment of acute brain infarction and amyotrophic lateral sclerosis (ALS), reduces oxidative stress and prevents RGC death and thinning of the inner retinal layer in EAAC1-deficient (KO) mice. In addition, in vivo electrophysiological analyses demonstrated that visual impairment in EAAC1 KO mice was ameliorated with edaravone treatment, clearly establishing that edaravone beneficially affects both histological and functional aspects of the glaucomatous retina. Our findings raise intriguing possibilities for the management of glaucoma by utilizing a widely prescribed drug for the treatment of acute brain infarction and ALS, edaravone, in combination with conventional treatments to lower IOP. PMID:28703795

  3. Retinal ganglion cell survival and axon regeneration after optic nerve injury in naked mole-rats.

    Science.gov (United States)

    Park, Kevin K; Luo, Xueting; Mooney, Skyler J; Yungher, Benjamin J; Belin, Stephane; Wang, Chen; Holmes, Melissa M; He, Zhigang

    2017-02-01

    In the adult mammalian central nervous system (CNS), axonal damage often triggers neuronal cell death and glial activation, with very limited spontaneous axon regeneration. In this study, we performed optic nerve injury in adult naked mole-rats, the longest living rodent, with a maximum life span exceeding 30 years, and found that injury responses in this species are quite distinct from those in other mammalian species. In contrast to what is seen in other mammals, the majority of injured retinal ganglion cells (RGCs) survive with relatively high spontaneous axon regeneration. Furthermore, injured RGCs display activated signal transducer and activator of transcription-3 (STAT3), whereas astrocytes in the optic nerve robustly occupy and fill the lesion area days after injury. These neuron-intrinsic and -extrinsic injury responses are reminiscent of those in "cold-blooded" animals, such as fish and amphibians, suggesting that the naked mole-rat is a powerful model for exploring the mechanisms of neuronal injury responses and axon regeneration in mammals. J. Comp. Neurol. 525:380-388, 2017. © 2016 Wiley Periodicals, Inc. © 2016 Wiley Periodicals, Inc.

  4. Nervus terminalis ganglion of the bonnethead shark (Sphyrna tiburo): evidence for cholinergic and catecholaminergic influence on two cell types distinguished by peptide immunocytochemistry.

    Science.gov (United States)

    White, J; Meredith, M

    1995-01-16

    The nervus terminalis is a ganglionated vertebrate cranial nerve of unknown function that connects the brain and the peripheral nasal structures. To investigate its function, we have studied nervus terminalis ganglion morphology and physiology in the bonnethead shark (Sphyrna tiburo), where the nerve is particularly prominent. Immunocytochemistry for gonadotropin-releasing hormone (GnRH) and Leu-Pro-Leu-Arg-Phe-NH2 (LPLRFamide) revealed two distinct populations of cells. Both were acetylcholinesterase positive, but LPLR-Famide-immunoreactive cells consistently stained more darkly for acetylcholinesterase activity. Tyrosine hydroxylase immunocytochemistry revealed fibers and terminal-like puncta in the ganglion, primarily in areas containing GnRH-immunoreactive cells. Consistent with the anatomy, in vitro electrophysiological recordings provided evidence for cholinergic and catecholaminergic actions. In extracellular recordings, acetylcholine had a variable effect on baseline ganglion cell activity, whereas norepinephrine consistently reduced activity. Electrical stimulation of the nerve trunks suppressed ganglion activity, as did impulses from the brain in vivo. During electrical suppression, acetylcholine consistently increased activity, and norepinephrine decreased activity. Muscarinic and, to a lesser extent, alpha-adrenergic antagonists both increased activity during the electrical suppression, suggesting involvement of both systems. Intracellular recordings revealed two types of ganglion cells that were distinguishable pharmacologically and physiologically. Some cells were hyperpolarized by cholinergic agonists and unaffected by norepinephrine; these cells did not depolarize with peripheral nerve trunk stimulation. Another group of cells did depolarize with peripheral trunk stimulation; a representative of this group was depolarized by carbachol and hyperpolarized by norepinephrine. These and other data suggest that the bonnethead nervus terminalis ganglion

  5. Correspondence between visual and electrical input filters of ON and OFF mouse retinal ganglion cells

    Science.gov (United States)

    Sekhar, S.; Jalligampala, A.; Zrenner, E.; Rathbun, D. L.

    2017-08-01

    Objective. Over the past two decades retinal prostheses have made major strides in restoring functional vision to patients blinded by diseases such as retinitis pigmentosa. Presently, implants use single pulses to activate the retina. Though this stimulation paradigm has proved beneficial to patients, an unresolved problem is the inability to selectively stimulate the on and off visual pathways. To this end our goal was to test, using white noise, voltage-controlled, cathodic, monophasic pulse stimulation, whether different retinal ganglion cell (RGC) types in the wild type retina have different electrical input filters. This is an important precursor to addressing pathway-selective stimulation. Approach. Using full-field visual flash and electrical and visual Gaussian noise stimulation, combined with the technique of spike-triggered averaging (STA), we calculate the electrical and visual input filters for different types of RGCs (classified as on, off or on-off based on their response to the flash stimuli). Main results. Examining the STAs, we found that the spiking activity of on cells during electrical stimulation correlates with a decrease in the voltage magnitude preceding a spike, while the spiking activity of off cells correlates with an increase in the voltage preceding a spike. No electrical preference was found for on-off cells. Comparing STAs of wild type and rd10 mice revealed narrower electrical STA deflections with shorter latencies in rd10. Significance. This study is the first comparison of visual cell types and their corresponding temporal electrical input filters in the retina. The altered input filters in degenerated rd10 retinas are consistent with photoreceptor stimulation underlying visual type-specific electrical STA shapes in wild type retina. It is therefore conceivable that existing implants could target partially degenerated photoreceptors that have only lost their outer segments, but not somas, to selectively activate the on and off

  6. Apoptosis of acinar cells in pancreas allograft rejection

    NARCIS (Netherlands)

    Boonstra, J. G.; Wever, P. C.; Laterveer, J. C.; Bruijn, J. A.; van der Woude, F. J.; ten Berge, I. J.; Daha, M. R.

    1997-01-01

    BACKGROUND: Recently it has been recognized that apoptosis of target cells may occur during liver and kidney allograft rejection and is probably induced by infiltrating cells. Pancreas rejection is also characterized by a cellular infiltrate, however, the occurrence of apoptosis has not been

  7. Apoptosis-Dependent and Apoptosis-Independent Functions Bim in Prostate Cancer Cells

    National Research Council Canada - National Science Library

    Liu, Junwei

    2004-01-01

    ... (death, survival, proliferation/division, etc). Our hypothesis is that, under normal, unstimulated conditions, with its apoptotic function blocked, the upregulated Bim in PCa cells play an apoptosis-independent function...

  8. Apoptosis-Dependent and Apoptosis-Independent Functions of Bim in Prostate Cancer Cells

    National Research Council Canada - National Science Library

    Tang, Dean; Liu, Junwei

    2005-01-01

    ... (death, survival, proliferation/division, etc.). Our hypothesis is that, under normal, unstimulated conditions, with its apoptotic function blocked, the upregulated Bim in PCa cells plays an apoptosis-independent function...

  9. Analysis the macular ganglion cell complex thickness in monocular strabismic amblyopia patients by Fourier-domain OCT

    Directory of Open Access Journals (Sweden)

    Hong-Wei Deng

    2014-11-01

    Full Text Available AIM: To detect the macular ganglion cell complex thickness in monocular strabismus amblyopia patients, in order to explore the relationship between the degree of amblyopia and retinal ganglion cell complex thickness, and found out whether there is abnormal macular ganglion cell structure in strabismic amblyopia. METHODS: Using a fourier-domain optical coherence tomography(FD-OCTinstrument iVue®(Optovue Inc, Fremont, CA, Macular ganglion cell complex(mGCCthickness was measured and statistical the relation rate with the best vision acuity correction was compared Gman among 26 patients(52 eyesincluded in this study. RESULTS: The mean thickness of the mGCC in macular was investigated into three parts: centrial, inner circle(3mmand outer circle(6mm. The mean thicknesses of mGCC in central, inner and outer circle was 50.74±21.51μm, 101.4±8.51μm, 114.2±9.455μm in the strabismic amblyopia eyes(SAE, and 43.79±11.92μm,92.47±25.01μm, 113.3±12.88μm in the contralateral sound eyes(CSErespectively. There was no statistically significant difference among the eyes(P>0.05. But the best corrected vision acuity had a good correlation rate between mGcc thicknesses, which was better relative for the lower part than the upper part.CONCLUSION:There is a relationship between the amblyopia vision acuity and the mGCC thickness. Although there has not statistically significant difference of the mGCC thickness compared with the SAE and CSE. To measure the macular center mGCC thickness in clinic may understand the degree of amblyopia.

  10. Role of heat shock proteins in cell apoptosis

    Directory of Open Access Journals (Sweden)

    Arleta Kaźmierczuk

    2010-06-01

    Full Text Available Apoptosis is, apart from necrosis and autophagy, one of the possible cell death mechanisms eliminating needless, not normal or infected cells. This process ensures quantitative and qualitative cell control of organisms. Apoptosis is tightly regulated, it requires both activation of a large number of genes and energy input. Up-to-date two main apoptotic pathways have been recognized – external/receptor and internal, processed with the participation of mitochondria. Heat shock proteins HSPs, the molecules known from their chaperone activity and molecular conservatism, play essential functions in the course of apoptosis. Among that proteins family, i.e. HSP100, 90, 70, 60, 40 and small molecular (sHSP, there are agents mainly protective against programmed cell death. However, in some conditions some of these proteins may promote apoptosis. This review describes different key apoptotic proteins interacting with main members of HSP family and the consequence of these events for cell survival or apoptosis.

  11. Ezh2 does not mediate retinal ganglion cell homeostasis or their susceptibility to injury.

    Directory of Open Access Journals (Sweden)

    Lin Cheng

    Full Text Available Epigenetic predisposition is thought to critically contribute to adult-onset disorders, such as retinal neurodegeneration. The histone methyltransferase, enhancer of zeste homolog 2 (Ezh2, is transiently expressed in the perinatal retina, particularly enriched in retinal ganglion cells (RGCs. We previously showed that embryonic deletion of Ezh2 from retinal progenitors led to progressive photoreceptor degeneration throughout life, demonstrating a role for embryonic predisposition of Ezh2-mediated repressive mark in maintaining the survival and function of photoreceptors in the adult. Enrichment of Ezh2 in RGCs leads to the question if Ezh2 also mediates gene expression and function in postnatal RGCs, and if its deficiency changes RGC susceptibility to cell death under injury or disease in the adult. To test this, we generated mice carrying targeted deletion of Ezh2 from RGC progenitors driven by Math5-Cre (mKO. mKO mice showed no detectable defect in RGC development, survival, or cell homeostasis as determined by physiological analysis, live imaging, histology, and immunohistochemistry. Moreover, RGCs of Ezh2 deficient mice revealed similar susceptibility against glaucomatous and acute optic nerve trauma-induced neurodegeneration compared to littermate floxed or wild-type control mice. In agreement with the above findings, analysis of RNA sequencing of RGCs purified from Ezh2 deficient mice revealed few gene changes that were related to RGC development, survival and function. These results, together with our previous report, support a cell lineage-specific mechanism of Ezh2-mediated gene repression, especially those critically involved in cellular function and homeostasis.

  12. Spontaneous oscillatory rhythms in the degenerating mouse retina modulate retinal ganglion cell responses to electrical stimulation

    Directory of Open Access Journals (Sweden)

    Yong Sook eGoo

    2016-01-01

    Full Text Available Characterization of the electrical activity of the retina in the animal models of retinal degeneration has been carried out in part to understand the progression of retinal degenerative diseases like age-related macular degeneration (AMD and retinitis pigmentosa (RP, but also to determine optimum stimulus paradigms for use with retinal prosthetic devices. The models most studied in this regard have been the two lines of mice deficient in the β-subunit of phosphodiesterase (rd1 and rd10 mice, where the degenerating retinas exhibit characteristic spontaneous hyperactivity and oscillatory local field potentials (LFPs. Additionally, there is a robust ~10 Hz rhythmic burst of retinal ganglion cell (RGC spikes on the trough of the oscillatory LFP. In rd1 mice, the rhythmic burst of RGC spikes is always phase-locked with the oscillatory LFP and this phase-locking property is preserved regardless of postnatal ages. However, in rd10 mice, the frequency of the oscillatory rhythm changes according to postnatal age, suggesting that this rhythm might be a marker of the stage of degeneration. Furthermore when a biphasic current stimulus is applied to rd10 mice degenerate retina, distinct RGC response patterns that correlate with the stage of degeneration emerge. This review also considers the significance of these response properties.

  13. Activation of Satellite Glial Cells in Rat Trigeminal Ganglion after Upper Molar Extraction

    International Nuclear Information System (INIS)

    Gunjigake, Kaori K.; Goto, Tetsuya; Nakao, Kayoko; Kobayashi, Shigeru; Yamaguchi, Kazunori

    2009-01-01

    The neurons in the trigeminal ganglion (TG) are surrounded by satellite glial cells (SGCs), which passively support the function of the neurons, but little is known about the interactions between SGCs and TG neurons after peripheral nerve injury. To examine the effect of nerve injury on SGCs, we investigated the activation of SGCs after neuronal damage due to the extraction of the upper molars in rats. Three, 7, and 10 days after extraction, animals were fixed and the TG was removed. Cryosections of the ganglia were immunostained with antibodies against glial fibrillary acidic protein (GFAP), a marker of activated SGCs, and ATF3, a marker of damaged neurons. After tooth extraction, the number of ATF3-immunoreactive (IR) neurons enclosed by GFAP-IR SGCs had increased in a time-dependent manner in the maxillary nerve region of the TG. Although ATF3-IR neurons were not detected in the mandibular nerve region, the number of GFAP-IR SGCs increased in both the maxillary and mandibular nerve regions. Our results suggest that peripheral nerve injury affects the activation of TG neurons and the SGCs around the injured neurons. Moreover, our data suggest the existence of a neuronal interaction between maxillary and mandibular neurons via SGC activation

  14. Retinal nerve fiber layer and ganglion cell complex thickness in patients with type 2 diabetes mellitus

    Directory of Open Access Journals (Sweden)

    Mehmet Demir

    2014-01-01

    Full Text Available Aim: The aim of the following study is to evaluate the retinal nerve fiber layer (RNFL and ganglion cell complex (GCC thickness in patients with type 2 diabetes mellitus (DM. Materials and Methods: Average, inferior, and superior values of RNFL and GCC thickness were measured in 123 patients using spectral domain optical coherence tomography. The values of participants with DM were compared to controls. Diabetic patients were collected in Groups 1, 2 and 3. Group 1 = 33 participants who had no diabetic retinopathy (DR; Group 2 = 30 participants who had mild nonproliferative DR and Group 3 = 30 participants who had moderate non-proliferative DR. The 30 healthy participants collected in Group 4. Analysis of variance test and a multiple linear regression analysis were used for statistical analysis. Results: The values of RNFL and GCC in the type 2 diabetes were thinner than controls, but this difference was not statistically significant. Conclusions: This study showed that there is a nonsignificant loss of RNFL and GCC in patients with type 2 diabetes.

  15. Losartan Treatment Protects Retinal Ganglion Cells and Alters Scleral Remodeling in Experimental Glaucoma

    Science.gov (United States)

    Pitha, Ian F.; Nguyen, Cathy; Steinhart, Matthew R.; Nguyen, Thao D.; Pease, Mary Ellen; Oglesby, Ericka N.; Berlinicke, Cynthia A.; Mitchell, Katherine L.; Kim, Jessica; Jefferys, Joan J.

    2015-01-01

    Purpose To determine if oral losartan treatment decreases the retinal ganglion cell (RGC) death caused by experimental intraocular pressure (IOP) elevation in mice. Methods We produced IOP increase in CD1 mice and performed unilateral optic nerve crush. Mice received oral losartan, spironolactone, enalapril, or no drug to test effects of inhibiting angiotensin receptors. IOP was monitored by Tonolab, and blood pressure was monitored by tail cuff device. RGC loss was measured in masked axon counts and RGC bodies by β-tubulin labeling. Scleral changes that could modulate RGC injury were measured including axial length, scleral thickness, and retinal layer thicknesses, pressure-strain behavior in inflation testing, and study of angiotensin receptors and pathways by reverse transcription polymerase chain reaction, Western blot, and immunohistochemistry. Results Losartan treatment prevented significant RGC loss (median loss = 2.5%, p = 0.13), while median loss with water, spironolactone, and enalapril treatments were 26%, 28% and 43%; p glaucoma eyes (p = 0.007). Losartan inhibited effects of glaucoma, including reduction in extracellular signal-related kinase activity and modification of glaucoma-related changes in scleral thickness and creep under controlled IOP. Conclusions The neuroprotective effect of losartan in mouse glaucoma is associated with adaptive changes in the sclera expressed at the optic nerve head. PMID:26505191

  16. Image quality of the cat eye measured during retinal ganglion cell experiments.

    Science.gov (United States)

    Bonds, A B; Enroth-Cugell, C; Pinto, L H

    1972-01-01

    1. The modulation transfer function (MTF) of the dioptrics of fifteen cat eyes was determined. The aerial image, formed by the eye of a standard object (a 0.5-1.0 degrees annulus), was photographed. The transmission of the film negative was measured with a scanning microdensitometer to yield the light distribution within the aerial image. Correcting for the double passage, this experimentally determined light distribution and the known object light distribution were used to obtain the MTF, applying Fourier methods. Each MTF was used to calculate the light distribution within the retinal image of stimuli of various geometry used in experiments on retinal ganglion cells in the same eye.2. When the eye was equipped with an artificial pupil of the same size as that used in the neurophysiological experiments (4.0-4.8 mm diam.) the MTF had fallen to 0.5 at 2.43 c/deg. When the pupil was removed the MTF had fallen to 0.5 at a much lower spatial frequency (1.0 c/deg). This shows that even when one uses an artificial pupil too large to provide optimal image quality there is a vast improvement over using no pupil.3. These image quality measurements were prompted by the need to know the actual stimulus image in experiments on the functional organization of the receptive field, a need exemplified in this paper by a few specific physiological results. The full neurophysiological results appear in the next two papers.

  17. Apoptosis and cancer stem cells : Implications for apoptosis targeted therapy

    NARCIS (Netherlands)

    Kruyt, Frank A. E.; Schuringa, Jan Jacob

    2010-01-01

    Evidence is accumulating showing that cancer stem cells or tumor-initiating cells are key drivers of tumor formation and progression. Successful therapy must therefore eliminate these cells, which is hampered by their high resistance to commonly used treatment modalities. Thus far, only a limited

  18. [Study on thaspine in inducing apoptosis of A549 cell].

    Science.gov (United States)

    Zhang, Yan-min; He, Lang-chong

    2007-04-01

    To investigate the effect of thaspine on the cellular proliferation, apoptosis and cell cycle in A549 cell line. A549 cell was cultured with different concentrations of thaspine. Cellular proliferation was detected with MTT, apoptosis and cell cycle were checked with Flow Cytometer, and change of microstructure was observed by transmission electron microscope. Thaspine could inhibit the proliferation and induce apoptosis of A549 cell in a time-dose dependent manner. Cell cycle was significantly stopped at the S phase by thaspine with FCM technology. Under electronic microscope, the morphology of A549 cell showed nuclear karyopycnosis, chromatin agglutination and typical apoptotic body when the cell was treated with thaspine. Thaspine has the effects of anti-tumor and inducing apoptosis.

  19. Electric stimulus duration alters network-mediated responses depending on retinal ganglion cell type

    Science.gov (United States)

    Im, Maesoon; Werginz, Paul; Fried, Shelley I.

    2018-06-01

    Objective. To improve the quality of artificial vision that arises from retinal prostheses, it is important to bring electrically-elicited neural activity more in line with the physiological signaling patterns that arise normally in the healthy retina. Our previous study reported that indirect activation produces a closer match to physiological responses in ON retinal ganglion cells (RGCs) than in OFF cells (Im and Fried 2015 J. Physiol. 593 3677-96). This suggests that a preferential activation of ON RGCs would shape the overall retinal response closer to natural signaling. Recently, we found that changes to the rate at which stimulation was delivered could bias responses towards a stronger ON component (Im and Fried 2016a J. Neural Eng. 13 025002), raising the possibility that changes to other stimulus parameters can similarly bias towards stronger ON responses. Here, we explore the effects of changing stimulus duration on the responses in ON and OFF types of brisk transient (BT) and brisk sustained (BS) RGCs. Approach. We used cell-attached patch clamp to record RGC spiking in the isolated rabbit retina. Targeted RGCs were first classified as ON or OFF type by their light responses, and further sub-classified as BT or BS types by their responses to both light and electric stimuli. Spiking in targeted RGCs was recorded in response to electric pulses with durations varying from 5 to100 ms. Stimulus amplitude was adjusted at each duration to hold total charge constant for all experiments. Main results. We found that varying stimulus durations modulated responses differentially for ON versus OFF cells: in ON cells, spike counts decreased significantly with increasing stimulus duration while in OFF cells the changes were more modest. The maximum ratio of ON versus OFF responses occurred at a duration of ~10 ms. The difference in response strength for BT versus BS cells was much larger in ON cells than in OFF cells. Significance. The stimulation rates preferred by

  20. Imaging and quantifying ganglion cells and other transparent neurons in the living human retina.

    Science.gov (United States)

    Liu, Zhuolin; Kurokawa, Kazuhiro; Zhang, Furu; Lee, John J; Miller, Donald T

    2017-11-28

    Ganglion cells (GCs) are fundamental to retinal neural circuitry, processing photoreceptor signals for transmission to the brain via their axons. However, much remains unknown about their role in vision and their vulnerability to disease leading to blindness. A major bottleneck has been our inability to observe GCs and their degeneration in the living human eye. Despite two decades of development of optical technologies to image cells in the living human retina, GCs remain elusive due to their high optical translucency. Failure of conventional imaging-using predominately singly scattered light-to reveal GCs has led to a focus on multiply-scattered, fluorescence, two-photon, and phase imaging techniques to enhance GC contrast. Here, we show that singly scattered light actually carries substantial information that reveals GC somas, axons, and other retinal neurons and permits their quantitative analysis. We perform morphometry on GC layer somas, including projection of GCs onto photoreceptors and identification of the primary GC subtypes, even beneath nerve fibers. We obtained singly scattered images by: ( i ) marrying adaptive optics to optical coherence tomography to avoid optical blurring of the eye; ( ii ) performing 3D subcellular image registration to avoid motion blur; and ( iii ) using organelle motility inside somas as an intrinsic contrast agent. Moreover, through-focus imaging offers the potential to spatially map individual GCs to underlying amacrine, bipolar, horizontal, photoreceptor, and retinal pigment epithelium cells, thus exposing the anatomical substrate for neural processing of visual information. This imaging modality is also a tool for improving clinical diagnosis and assessing treatment of retinal disease. Copyright © 2017 the Author(s). Published by PNAS.

  1. Apoptosis and T cell depletion during feline infectious peritonitis

    OpenAIRE

    Horzinek, M.C.; Haagmans, B.L.; Egberink, H.F.

    1996-01-01

    Cats that have succumbed to feline infectious peritonitis, an immune- mediated disease caused by variants of feline coronaviruses, show apoptosis and T-cell depletion in their lymphoid organs. The ascitic fluid that develops in the course of the condition causes apoptosis in vitro but only in activated T cells. Since feline infectious peritonitis virus does not infect T cells, and viral proteins did not inhibit T-cell proliferation, we postulate that soluble mediators released during the infe...

  2. Apoptosis transcriptional mechanism of feline infectious peritonitis virus infected cells.

    Science.gov (United States)

    Shuid, Ahmad Naqib; Safi, Nikoo; Haghani, Amin; Mehrbod, Parvaneh; Haron, Mohd Syamsul Reza; Tan, Sheau Wei; Omar, Abdul Rahman

    2015-11-01

    Apoptosis has been postulated to play an important role during feline infectious peritonitis virus (FIPV) infection; however, its mechanism is not well characterized. This study is focused on apoptosis and transcriptional profiling of FIPV-infected cells following in vitro infection of CRFK cells with FIPV 79-1146 WSU. Flow cytometry was used to determine mode of cell death in first 42 h post infection (hpi). FIPV infected cells underwent early apoptosis at 9 hpi (p apoptosis at 12 hpi (p apoptosis cluster (80 down-regulated and 51 up-regulated) along with increase of apoptosis, p53, p38 MAPK, VEGF and chemokines/cytokines signaling pathways were probably involved in apoptosis process. Six of the de-regulated genes expression (RASSF1, BATF2, MAGEB16, PDCD5, TNFα and TRAF2) and TNFα protein concentration were analyzed by RT-qPCR and ELISA, respectively, at different time-points. Up-regulations of both pro-apoptotic (i.e. PDCD5) and anti-apoptotic (i.e. TRAF2) were detected from first hpi and continuing to deregulate during apoptosis process in the infected cells.

  3. Beta-irradiation used for systemic radioimmunotherapy induces apoptosis and activates apoptosis pathways in leukaemia cells

    International Nuclear Information System (INIS)

    Friesen, Claudia; Lubatschofski, Annelie; Debatin, Klaus-Michael; Kotzerke, Joerg; Buchmann, Inga; Reske, Sven N.

    2003-01-01

    Beta-irradiation used for systemic radioimmunotherapy (RIT) is a promising treatment approach for high-risk leukaemia and lymphoma. In bone marrow-selective radioimmunotherapy, beta-irradiation is applied using iodine-131, yttrium-90 or rhenium-188 labelled radioimmunoconjugates. However, the mechanisms by which beta-irradiation induces cell death are not understood at the molecular level. Here, we report that beta-irradiation induced apoptosis and activated apoptosis pathways in leukaemia cells depending on doses, time points and dose rates. After beta-irradiation, upregulation of CD95 ligand and CD95 receptor was detected and activation of caspases resulting in apoptosis was found. These effects were completely blocked by the broad-range caspase inhibitor zVAD-fmk. In addition, irradiation-mediated mitochondrial damage resulted in perturbation of mitochondrial membrane potential, caspase-9 activation and cytochrome c release. Bax, a death-promoting protein, was upregulated and Bcl-x L , a death-inhibiting protein, was downregulated. We also found higher apoptosis rates and earlier activation of apoptosis pathways after gamma-irradiation in comparison to beta-irradiation at the same dose rate. Furthermore, irradiation-resistant cells were cross-resistant to CD95 and CD95-resistant cells were cross-resistant to irradiation, indicating that CD95 and irradiation used, at least in part, identical effector pathways. These findings demonstrate that beta-irradiation induces apoptosis and activates apoptosis pathways in leukaemia cells using both mitochondrial and death receptor pathways. Understanding the timing, sequence and molecular pathways of beta-irradiation-mediated apoptosis may allow rational adjustment of chemo- and radiotherapeutic strategies. (orig.)

  4. ["Point by point" approach to structure-function correlation of glaucoma on the ganglion cell complex in the posterior pole].

    Science.gov (United States)

    Zeitoun, M

    2017-01-01

    To try to establish a "point by point" relationship between the local thickness of the retinal ganglion cell complex and its sensitivity. In total, 104 glaucomatous eyes of 89 patients with a confirmed 24-2 visual field, were measured by superimposing the visual field, using imaging software, with the Wide 40° by 30° measurements of retinal ganglion cell complex obtained from the Topcon © 3D 2000 OCT, after upward adjustment, inversion and scaling. Visual fields were classified into two groups according to the extent of the disease: 58 mild to moderate (MD up to -12dB), and 46 severe (MD beyond -12dB). The 6mm by 6mm central region, equipped with a normative database, was studied, corresponding to 16 points in the visual field. These points were individually matched one by one to the local ganglion cell complex, which was classified into 2 groups depending on whether it was greater or less than 70 microns. The normative database confirmed the pathological nature of the thin areas, with a significance of 95 to 99%. Displacement of central retinal ganglion cells was compensated for. Of 1664 points (16 central points for 104 eyes), 283 points were found to be "borderline" and excluded. Of the 1381 analyzed points, 727 points were classified as "over 70 microns" and 654 points "under 70 microns". (1) For all stages combined, 85.8% of the 727 points which were greater than 70 microns had a deviation between -3 and +3dB: areas above 70 microns had no observable loss of light sensitivity. (2) In total, 92.5% of the 428 points having a gap ranging from -6 to -35dB were located on ganglion cell complex areas below 70 microns: functional visual loss was identified in thin areas, which were less than 70 microns. (3) Areas which were less than 70 microns, that is 654 points, had quite variable sensitivity and can be divided into three groups: the first with preserved sensitivity, another with obliterated sensitivity, and an intermediate group connecting

  5. Melatonin potentiates glycine currents through a PLC/PKC signalling pathway in rat retinal ganglion cells.

    Science.gov (United States)

    Zhao, Wen-Jie; Zhang, Min; Miao, Yanying; Yang, Xiong-Li; Wang, Zhongfeng

    2010-07-15

    In vertebrate retina, melatonin regulates various physiological functions. In this work we investigated the mechanisms underlying melatonin-induced potentiation of glycine currents in rat retinal ganglion cells (RGCs). Immunofluorescence double labelling showed that rat RGCs were solely immunoreactive to melatonin MT(2) receptors. Melatonin potentiated glycine currents of RGCs, which was reversed by the MT(2) receptor antagonist 4-P-PDOT. The melatonin effect was blocked by intracellular dialysis of GDP-beta-S. Either preincubation with pertussis toxin or application of the phosphatidylcholine (PC)-specific phospholipase C (PLC) inhibitor D609, but not the phosphatidylinositol (PI)-PLC inhibitor U73122, blocked the melatonin effect. The protein kinase C (PKC) activator PMA potentiated the glycine currents and in the presence of PMA melatonin failed to cause further potentiation of the currents, whereas application of the PKC inhibitor bisindolylmaleimide IV abolished the melatonin-induced potentiation. The melatonin effect persisted when [Ca(2+)](i) was chelated by BAPTA, and melatonin induced no increase in [Ca(2+)](i). Neither cAMP-PKA nor cGMP-PKG signalling pathways seemed to be involved because 8-Br-cAMP or 8-Br-cGMP failed to cause potentiation of the glycine currents and both the PKA inhibitor H-89 and the PKG inhibitor KT5823 did not block the melatonin-induced potentiation. In consequence, a distinct PC-PLC/PKC signalling pathway, following the activation of G(i/o)-coupled MT(2) receptors, is most likely responsible for the melatonin-induced potentiation of glycine currents of rat RGCs. Furthermore, in rat retinal slices melatonin potentiated light-evoked glycine receptor-mediated inhibitory postsynaptic currents in RGCs. These results suggest that melatonin, being at higher levels at night, may help animals to detect positive or negative contrast in night vision by modulating inhibitory signals largely mediated by glycinergic amacrine cells in the inner

  6. Losartan Treatment Protects Retinal Ganglion Cells and Alters Scleral Remodeling in Experimental Glaucoma.

    Directory of Open Access Journals (Sweden)

    Harry A Quigley

    Full Text Available To determine if oral losartan treatment decreases the retinal ganglion cell (RGC death caused by experimental intraocular pressure (IOP elevation in mice.We produced IOP increase in CD1 mice and performed unilateral optic nerve crush. Mice received oral losartan, spironolactone, enalapril, or no drug to test effects of inhibiting angiotensin receptors. IOP was monitored by Tonolab, and blood pressure was monitored by tail cuff device. RGC loss was measured in masked axon counts and RGC bodies by β-tubulin labeling. Scleral changes that could modulate RGC injury were measured including axial length, scleral thickness, and retinal layer thicknesses, pressure-strain behavior in inflation testing, and study of angiotensin receptors and pathways by reverse transcription polymerase chain reaction, Western blot, and immunohistochemistry.Losartan treatment prevented significant RGC loss (median loss = 2.5%, p = 0.13, while median loss with water, spironolactone, and enalapril treatments were 26%, 28% and 43%; p < 0.0001. The lower RGC loss with losartan was significantly less than the loss with spironolactone or enalapril (regression model p = 0.001; drug treatment group term p = 0.01. Both losartan and enalapril significantly lowered blood pressure (p< 0.001, but losartan was protective, while enalapril led to worse than water-treated RGC loss. RGC loss after crush injury was unaffected by losartan treatment (difference from control p = 0.9. Survival of RGC in cell culture was not prolonged by sartan treatment. Axonal transport blockade after 3 day IOP elevations was less in losartan-treated than in control glaucoma eyes (p = 0.007. Losartan inhibited effects of glaucoma, including reduction in extracellular signal-related kinase activity and modification of glaucoma-related changes in scleral thickness and creep under controlled IOP.The neuroprotective effect of losartan in mouse glaucoma is associated with adaptive changes in the sclera expressed at

  7. Periosteal ganglion

    International Nuclear Information System (INIS)

    Kolar, J.; Zidkova, H.; Matejovsky, Z.

    1986-01-01

    Ganglionic cysts are a common myxomatous degenerative disorder in periarticular connective tissues particularly in the hand and foot as well as within the subchondral bone adjacent to osteoarthritic joints. Compared with them, periosteal ganglia are only rarely reported in the literature. Their radiologic features are quite typical as documented by the following observation. (orig.) [de

  8. Ganglion Cysts

    Science.gov (United States)

    ... All Topics A-Z Videos Infographics Symptom Picker Anatomy Bones Joints Muscles Nerves Vessels Tendons About Hand Surgery What is a Hand Surgeon? What is a Hand Therapist? Media Find a Hand Surgeon Home Anatomy Ganglion Cysts Email to a friend * required fields ...

  9. 5-Fluorouracil-induced apoptosis in cultured oral cancer cells.

    Science.gov (United States)

    Tong, D; Poot, M; Hu, D; Oda, D

    2000-03-01

    Chemotherapy is commonly used to treat advanced oral squamous cell carcinoma (SCC) and is known to kill cancer cells through apoptosis. Our hypothesis states that 5-fluorouracil (5FU) also kills cultured oral epithelial cells through programmed cell death or apoptosis. Cultured oral cancer cells were exposed to an optimum dose of 20 mg/ml of 5FU. Cells were analyzed for changes in cell cycle distribution and induction of cell death including apoptosis. Normal control, human papilloma virus-immortalized (PP), ATCC SCC cell line (CA1) and two primary oral SCC cell lines (CA3 and -4) were studied. Inhibition of apoptosis by a pan-caspase inhibitor was used. SYTO 11 flow cytometry showed increased apoptosis in all 5FU-treated cell cultures compared to untreated controls. The results show biological variation in apoptotic response. CA1 had the lowest apoptotic rate of the cancer cell lines at 1.5%. Next lowest was CA3, followed by CA4 and PP. In addition, alteration in the G1 and S phase fractions were found. Untreated CA1 showed 28% G1, 53% S compared to 43% G1, and 40% S of treated. We investigated the pathway of apoptosis using the pan-caspase inhibitor IDN-1529 by methylthiazolyl diphenyl tetrazolium bromide (MTT) colorimetric analysis. Results showed mild inhibition of cell death when cells were incubated with 50 microM IDN-1529 for 24 h. This suggests a probable caspase-dependent apoptotic pathway. In conclusion, our data suggest that 5FU induces oral cancer cell death through apoptosis and that biological variation exists between normal and cancer cells and between different types of cancer cells themselves. Our data indicate that cultures of a useful in vitro model for chemosensitivity assays are possible. Our results also suggest a caspase-dependent pathway for chemocytotoxicity in oral SCC.

  10. Apoptosis and T cell depletion during feline infectious peritonitis

    NARCIS (Netherlands)

    Horzinek, M.C.; Haagmans, B.L.; Egberink, H.F.

    1996-01-01

    Cats that have succumbed to feline infectious peritonitis, an immune- mediated disease caused by variants of feline coronaviruses, show apoptosis and T-cell depletion in their lymphoid organs. The ascitic fluid that develops in the course of the condition causes apoptosis in vitro but only in

  11. Characterization of radiation-induced Apoptosis in rodent cell lines

    International Nuclear Information System (INIS)

    Guo, Min; Chen, Changhu; Ling, C.C.

    1997-01-01

    For REC:myc(ch1), Rat1 and Rat1:myc b cells, we determined the events in the development of radiation-induced apoptosis to be in the following order: cell division followed by chromatin condensation, membrane blebbing, loss of adhesion and the uptake of vital dye. Experimental data which were obtained using 4 He ions of well defined energies and which compared the dependence of apoptosis and clonogenic survival on 4 He range strongly suggested that in our cells both apoptosis and loss of clonogenic survival resulted from radiation damage to the cell nucleus. Corroboratory evidence was that BrdU incorporation sensitized these cells to radiation-induced apoptosis. Comparing the dose response for apoptosis and the clonogenic survival curves for Rat1 and Rat1:myc b cells, we concluded that radiation-induced cell inactivation as assayed by clonogenic survival, and that a modified linear-quadratic model, proposed previously, modeled such a contribution effectively. In the same context, the selective increase in radiation-induced apoptosis. Comparing the dose response for apoptosis and the clonogenic survival curves for Rat1 and Rat1:myc b cells, we concluded that radiation-induced apoptosis contributed to the overall radiation-induced cell inactivation as assayed by clonogenic survival, and that a modified linear-quadratic model, proposed previously, modeled such a contribution effectively. In the same context, the selective increase in radiation-induced apoptosis during late S and G 2 phases reduced the relative radioresistance observed for clonogenic survival during late S and G 2 phases. 30 refs., 8 figs

  12. Human amniotic fluid promotes retinal pigmented epithelial cells' trans-differentiation into rod photoreceptors and retinal ganglion cells.

    Science.gov (United States)

    Ghaderi, Shima; Soheili, Zahra-Soheila; Ahmadieh, Hamid; Davari, Maliheh; Jahromi, Fatemeh Sanie; Samie, Shahram; Rezaie-Kanavi, Mozhgan; Pakravesh, Jalil; Deezagi, Abdolkhalegh

    2011-09-01

    To evaluate the effect of human amniotic fluid (HAF) on retinal pigmented epithelial cells growth and trans-differentiation into retinal neurons, retinal pigmented epithelium (RPE) cells were isolated from neonatal human cadaver eye globes and cultured in Dulbecco's modified Eagle's medium-F12 supplemented with 10% fetal bovine serum (FBS). Confluent monolayer cultures were trypsinized and passaged using FBS-containing or HAF-containing media. Amniotic fluid samples were received from pregnant women in the first trimester of gestation. Cell proliferation and death enzyme-linked immunosorbent assays were performed to assess the effect of HAF on RPE cell growth. Trans-differentiation into rod photoreceptors and retinal ganglion cells was also studied using immunocytochemistry and real-time polymerase chain reaction techniques. Primary cultures of RPE cells were successfully established under FBS-containing or HAF-containing media leading to rapid cell growth and proliferation. When RPE cells were moved to in vitro culture system, they began to lose their differentiation markers such as pigmentation and RPE65 marker and trans-differentiated neural-like cells followed by spheroid colonies pertaining to stem/progenitor cells were morphologically detected. Immunocytochemistry (ICC) analysis of HAF-treated cultures showed a considerable expression of Rhodopsin gene (30% Rhodopsin-positive cells) indicating trans-differentiation of RPE cells to rod photoreceptors. Real-time polymerase chain reaction revealed an HAF-dose-dependant expression of Thy-1 gene (RGC marker) and significant promoting effect of HAF on RGCs generation. The data presented here suggest that HAF possesses invaluable stimulatory effect on RPE cells growth and trans-differentiation into retinal neurons. It can be regarded as a newly introduced enriched supplement in serum-free kinds of media used in neuro-retinal regeneration studies.

  13. Retinal Astrocytes and GABAergic Wide-Field Amacrine Cells Express PDGFRα: Connection to Retinal Ganglion Cell Neuroprotection by PDGF-AA.

    Science.gov (United States)

    Takahama, Shokichi; Adetunji, Modupe O; Zhao, Tantai; Chen, Shan; Li, Wei; Tomarev, Stanislav I

    2017-09-01

    Our previous experiments demonstrated that intravitreal injection of platelet-derived growth factor-AA (PDGF-AA) provides retinal ganglion cell (RGC) neuroprotection in a rodent model of glaucoma. Here we used PDGFRα-enhanced green fluorescent protein (EGFP) mice to identify retinal cells that may be essential for RGC protection by PDGF-AA. PDGFRα-EGFP mice expressing nuclear-targeted EGFP under the control of the PDGFRα promoter were used. Localization of PDGFRα in the neural retina was investigated by confocal imaging of EGFP fluorescence and immunofluorescent labeling with a panel of antibodies recognizing different retinal cell types. Primary cultures of mouse RGCs were produced by immunopanning. Neurobiotin injection of amacrine cells in a flat-mounted retina was used for the identification of EGFP-positive amacrine cells in the inner nuclear layer. In the mouse neural retina, PDGFRα was preferentially localized in the ganglion cell and inner nuclear layers. Immunostaining of the retina demonstrated that astrocytes in the ganglion cell layer and a subpopulation of amacrine cells in the inner nuclear layer express PDGFRα, whereas RGCs (in vivo or in vitro) did not. PDGFRα-positive amacrine cells are likely to be Type 45 gamma-aminobutyric acidergic (GABAergic) wide-field amacrine cells. These data indicate that the neuroprotective effect of PDGF-AA in a rodent model of glaucoma could be mediated by astrocytes and/or a subpopulation of amacrine cells. We suggest that after intravitreal injection of PDGF-AA, these cells secrete factors protecting RGCs.

  14. Human retinal pigment epithelial cell-induced apoptosis in activated T cells

    DEFF Research Database (Denmark)

    Jørgensen, A; Wiencke, A K; la Cour, M

    1998-01-01

    human retinal pigment epithelial (RPE) cells can induce apoptosis in activated T cells. METHODS: Fas ligand (FasL) expression was detected by flow cytometry and immunohistochemistry. Cultured RPE cells were cocultured with T-cell lines and peripheral blood lymphocytes for 6 hours to 2 days. Induction...... of apoptosis was detected by 7-amino-actinomycin D and annexin V staining. RESULTS: Retinal pigment epithelial cells expressed FasL and induced apoptosis in activated Fas+ T cells. Blocking of Fas-FasL interaction with antibody strongly inhibited RPE-mediated T-cell apoptosis. Retinal pigment epithelial cells...... induced apoptosis in several activated T-cell populations and T-cell lines, including T-cell antigen receptor (TCR)-CD3-negative T-cell lines. In contrast, RPE cells induced little or no apoptosis in resting peripheral T cells. Major histocompatibility complex (MHC) class II monoclonal antibodies, which...

  15. Apoptosis signal-regulating kinase 1 mediates denbinobin-induced apoptosis in human lung adenocarcinoma cells

    Directory of Open Access Journals (Sweden)

    Pan Shiow-Lin

    2009-05-01

    Full Text Available Abstract In the present study, we explore the role of apoptosis signal-regulating kinase 1 (ASK1 in denbinobin-induced apoptosis in human lung adenocarcinoma (A549 cells. Denbinobin-induced cell apoptosis was attenuated by an ASK1 dominant-negative mutant (ASK1DN, two antioxidants (N-acetyl-L-cysteine (NAC and glutathione (GSH, a c-Jun N-terminal kinase (JNK inhibitor (SP600125, and an activator protein-1 (AP-1 inhibitor (curcumin. Treatment of A549 cells with denbinobin caused increases in ASK1 activity and reactive oxygen species (ROS production, and these effects were inhibited by NAC and GSH. Stimulation of A549 cells with denbinobin caused JNK activation; this effect was markedly inhibited by NAC, GSH, and ASK1DN. Denbinobin induced c-Jun phosphorylation, the formation of an AP-1-specific DNA-protein complex, and Bim expression. Bim knockdown using a bim short interfering RNA strategy also reduced denbinobin-induced A549 cell apoptosis. The denbinobin-mediated increases in c-Jun phosphorylation and Bim expression were inhibited by NAC, GSH, SP600125, ASK1DN, JNK1DN, and JNK2DN. These results suggest that denbinobin might activate ASK1 through ROS production to cause JNK/AP-1 activation, which in turn induces Bim expression, and ultimately results in A549 cell apoptosis.

  16. Osteoblasts Protect AML Cells from SDF-1-Induced Apoptosis

    Science.gov (United States)

    Kremer, Kimberly N.; Dudakovic, Amel; McGee-Lawrence, Meghan E.; Philips, Rachael L.; Hess, Allan D.; Smith, B. Douglas; van Wijnen, Andre J.; Karp, Judith E.; Kaufmann, Scott H.; Westendorf, Jennifer J.; Hedin, Karen E.

    2014-01-01

    The bone marrow provides a protective environment for acute myeloid leukemia (AML) cells that often allows leukemic stem cells to survive standard chemotherapeutic regimens. Targeting these leukemic stem cells within the bone marrow is critical for preventing relapse. We recently demonstrated that SDF-1, a chemokine abundant in the bone marrow, induces apoptosis in AML cell lines and in patient samples expressing high levels of its receptor, CXCR4. Here we show that a subset of osteoblast lineage cells within the bone marrow can protect AML cells from undergoing apoptosis in response to the SDF-1 naturally present in that location. In co-culture systems, osteoblasts at various stages of differentiation protected AML cell lines and patient isolates from SDF-1-induced apoptosis. The differentiation of the osteoblast cell lines, MC3T3 and W-20-17, mediated this protection via a cell contact-independent mechanism. In contrast, bone marrow-derived mesenchymal cells, the precursors of osteoblasts, induced apoptosis in AML cells via a CXCR4-dependent mechanism and failed to protect AML cells from exogenously added SDF-1. These results indicate that osteoblasts in the process of differentiation potently inhibit the SDF-1-driven apoptotic pathway of CXCR4-expressing AML cells residing in the bone marrow. Drugs targeting this protective mechanism could potentially provide a new approach to treating AML by enhancing the SDF-1-induced apoptosis of AML cells residing within the bone marrow microenvironment. PMID:24851270

  17. Apoptosis in ovarian cells in postmenopausal women.

    Directory of Open Access Journals (Sweden)

    Maria Laszczyńska

    2007-06-01

    Full Text Available Apoptosis is a natural process which accompanies human ovary from the moment of birth until old age. While it is a well-known process at the reproductive age, it still needs to be thoroughly examined when referring to the postmenopausal age. The study involved 30 postmenopausal women who had their ovaries removed by laparotomy due to nonneoplastic diseases of the uterus. The women were divided into 3 groups depending on the time that had passed since the last menstruation. Group A consisted of women who had their last menstruation no more than 5 years earlier. In group B menopause occurred 5 to 10 years earlier. Group C was composed of patients who had the last menstruation over 10 years earlier. In all the patients concentrations of follitropin (FSH and estradiol (E2 in blood plasma were measured. Ovarian tissue was obtained during surgery. For morphological studies, ovaries were fixed in Bouin's solution and 4% formalin and embedded in paraffin. Morphological analysis was carried out after hematoxylin-eosin (H-E staining. For histochemical detection of apoptotic cells (in situ localization of fragment DNA, the TUNEL method was used. The expression of caspase-3 positive cells was determined immunohistochemically in paraffin-embedded specimens. Comparing to groups A and B, the ovaries in group C contained small number of corpora albicantia located in the medullary part as well as thinned blood vessels and few lymphatic vessels and nerves. In contrast to group A where the number of TUNEL-positive cells was high and caspase-3 expression was observed, no TUNEL-positive nuclei and caspase-3 expression were found in the examined ovaries of group C women.

  18. Apoptosis in HEp-2 cells infected with Ureaplasma diversum.

    Science.gov (United States)

    Amorim, Aline Teixeira; Marques, Lucas Miranda; Santos, Angelita Maria Oliveira Gusmão; Martins, Hellen Braga; Barbosa, Maysa Santos; Rezende, Izadora Souza; Andrade, Ewerton Ferraz; Campos, Guilherme Barreto; Lobão, Tássia Neves; Cortez, Beatriz Araujo; Monezi, Telma Alvez; Machado-Santelli, Glaucia Maria; Timenetsky, Jorge

    2014-09-04

    Bacterial pathogens have many strategies for infecting and persisting in host cells. Adhesion, invasion and intracellular life are important features in the biology of mollicutes. The intracellular location of Ureaplasma diversum may trigger disturbances in the host cell. This includes activation or inhibition of pro and anti-apoptotic factors, which facilitate the development of host damage. The aim of the present study was to associate U. diversum infection in HEp-2 cells and apoptosis induction. Cells were infected for 72hs with four U. diversum clinical isolates and an ATCC strain. The U. diversum invasion was analyzed by Confocal Laser Scanning Microscopy and gentamicin invasion assay. The apoptosis was evaluated using pro-apoptotic and anti-apoptotic gene expression, and FITC Annexin V/Dead Cell Apoptosis Kit. The number of internalized ureaplasma in HEp-2 cells increased significantly throughout the infection. The flow cytometry analysis with fluorochromes to detect membrane depolarization and gene expression for caspase 2, 3 and 9 increased in infected cells after 24 hours. However, after 72 hours a considerable decrease of apoptotic cells was observed. The data suggests that apoptosis may be initially induced by some isolates in association with HEp-2 cells, but over time, there was no evidence of apoptosis in the presence of ureaplasma and HEp-2 cells. The initial increase and then decrease in apoptosis could be related to bacterial pathogen-associated molecular pattern (PAMPS). Moreover, the isolates of U. diversum presented differences in the studied parameters for apoptosis. It was also observed that the amount of microorganisms was not proportional to the induction of apoptosis in HEp-2 cells.

  19. Regional Retinal Ganglion Cell Axon Loss in a Murine Glaucoma Model.

    Science.gov (United States)

    Schaub, Julie A; Kimball, Elizabeth C; Steinhart, Matthew R; Nguyen, Cathy; Pease, Mary E; Oglesby, Ericka N; Jefferys, Joan L; Quigley, Harry A

    2017-05-01

    To determine if retinal ganglion cell (RGC) axon loss in experimental mouse glaucoma is uniform in the optic nerve. Experimental glaucoma was induced for 6 weeks with a microbead injection model in CD1 (n = 78) and C57BL/6 (B6, n = 68) mice. From epoxy-embedded sections of optic nerve 1 to 2 mm posterior to the globe, total nerve area and regional axon density (axons/1600 μm2) were measured in superior, inferior, nasal, and temporal zones. Control eyes of CD1 mice have higher axon density and more total RGCs than control B6 mice eyes. There were no significant differences in control regional axon density in all mice or by strain (all P > 0.2, mixed model). Exposure to elevated IOP caused loss of RGC in both strains. In CD1 mice, axon density declined without significant loss of nerve area, while B6 mice had less density loss, but greater decrease in nerve area. Axon density loss in glaucoma eyes was not significantly greater in any region in either mouse strain (both P > 0.2, mixed model). In moderately damaged CD1 glaucoma eyes, and CD1 eyes with the greatest IOP elevation exposure, density loss differed by region (P = 0.05, P = 0.03, mixed model) with the greatest loss in the temporal and superior regions, while in severely injured B6 nerves superior loss was greater than inferior loss (P = 0.01, mixed model, Bonferroni corrected). There was selectively greater loss of superior and temporal optic nerve axons of RGCs in mouse glaucoma at certain stages of damage. Differences in nerve area change suggest non-RGC responses differ between mouse strains.

  20. Effects of p-xylene inhalation on axonal transport in the rat retinal ganglion cells

    Energy Technology Data Exchange (ETDEWEB)

    Padilla, S.S.; Lyerly, D.P. (Environmental Protection Agency, Research Triangle Park, NC (USA))

    1989-12-01

    Although the solvent xylene is suspected of producing nervous system dysfunction in animals and humans, little is known regarding the neurochemical consequences of xylene inhalation. The intent of this study was to determine the effect of intermittent, acute, and subchronic p-xylene exposure on the axonal transport of proteins and glycoproteins within the rat retinofugal tract. A number of different exposure regimens were tested ranging from 50 ppm for a single 6-hr exposure to 1600 ppm 6 hr/day, 5 days/week, for a total of 8 exposure days. Immediately following removal from the inhalation chambers rats were injected intraocularly with (35S)methionine and (3H)fucose (to label retinal proteins and glycoproteins, respectively) and the axonal transport of labeled macromolecules to axons (optic nerve and optic tract) and nerve endings (lateral geniculate body and superior colliculus) was examined 20 hr after precursor injection. Only relatively severe exposure regimens (i.e., 800 or 1600 ppm 6 hr/day, 5 days/week, for 1.5 weeks) produced significant reductions in axonal transport; there was a moderate reduction in the axonal transport of 35S-labeled proteins in the 800-ppm-treated group which was more widespread in the 1600 ppm-treated group. Transport of 3H-labeled glycoproteins was less affected. Assessment of retinal metabolism immediately after isotope injection indicated that the rate of precursor uptake was not reduced in either treatment group. Furthermore, rapid transport was still substantially reduced in animals exposed to 1600 ppm p-xylene and allowed a 13-day withdrawal period. These data indicate that p-xylene inhalation decreases rapid axonal transport supplied to the projections of the rat retinal ganglion cells immediately after cessation of inhalation exposure and that this decreased transport is still apparent 13 days after the last exposure.

  1. Effects of p-xylene inhalation on axonal transport in the rat retinal ganglion cells

    International Nuclear Information System (INIS)

    Padilla, S.S.; Lyerly, D.P.

    1989-01-01

    Although the solvent xylene is suspected of producing nervous system dysfunction in animals and humans, little is known regarding the neurochemical consequences of xylene inhalation. The intent of this study was to determine the effect of intermittent, acute, and subchronic p-xylene exposure on the axonal transport of proteins and glycoproteins within the rat retinofugal tract. A number of different exposure regimens were tested ranging from 50 ppm for a single 6-hr exposure to 1600 ppm 6 hr/day, 5 days/week, for a total of 8 exposure days. Immediately following removal from the inhalation chambers rats were injected intraocularly with [35S]methionine and [3H]fucose (to label retinal proteins and glycoproteins, respectively) and the axonal transport of labeled macromolecules to axons (optic nerve and optic tract) and nerve endings (lateral geniculate body and superior colliculus) was examined 20 hr after precursor injection. Only relatively severe exposure regimens (i.e., 800 or 1600 ppm 6 hr/day, 5 days/week, for 1.5 weeks) produced significant reductions in axonal transport; there was a moderate reduction in the axonal transport of 35S-labeled proteins in the 800-ppm-treated group which was more widespread in the 1600 ppm-treated group. Transport of 3H-labeled glycoproteins was less affected. Assessment of retinal metabolism immediately after isotope injection indicated that the rate of precursor uptake was not reduced in either treatment group. Furthermore, rapid transport was still substantially reduced in animals exposed to 1600 ppm p-xylene and allowed a 13-day withdrawal period. These data indicate that p-xylene inhalation decreases rapid axonal transport supplied to the projections of the rat retinal ganglion cells immediately after cessation of inhalation exposure and that this decreased transport is still apparent 13 days after the last exposure

  2. Mice deficient of glutamatergic signaling from intrinsically photosensitive retinal ganglion cells exhibit abnormal circadian photoentrainment.

    Directory of Open Access Journals (Sweden)

    Nicole Purrier

    Full Text Available Several aspects of behavior and physiology, such as sleep and wakefulness, blood pressure, body temperature, and hormone secretion exhibit daily oscillations known as circadian rhythms. These circadian rhythms are orchestrated by an intrinsic biological clock in the suprachiasmatic nuclei (SCN of the hypothalamus which is adjusted to the daily environmental cycles of day and night by the process of photoentrainment. In mammals, the neuronal signal for photoentrainment arises from a small subset of intrinsically photosensitive retinal ganglion cells (ipRGCs that send a direct projection to the SCN. ipRGCs also mediate other non-image-forming (NIF visual responses such as negative masking of locomotor activity by light, and the pupillary light reflex (PLR via co-release of neurotransmitters glutamate and pituitary adenylate cyclase-activating peptide (PACAP from their synaptic terminals. The relative contribution of each neurotransmitter system for the circadian photoentrainment and other NIF visual responses is still unresolved. We investigated the role of glutamatergic neurotransmission for circadian photoentrainment and NIF behaviors by selective ablation of ipRGC glutamatergic synaptic transmission in mice. Mutant mice displayed delayed re-entrainment to a 6 h phase shift (advance or delay in the light cycle and incomplete photoentrainment in a symmetrical skeleton photoperiod regimen (1 h light pulses between 11 h dark periods. Circadian rhythmicity in constant darkness also was reduced in some mutant mice. Other NIF responses such as the PLR and negative masking responses to light were also partially attenuated. Overall, these results suggest that glutamate from ipRGCs drives circadian photoentrainment and negative masking responses to light.

  3. Virally delivered, constitutively active NFκB improves survival of injured retinal ganglion cells.

    Science.gov (United States)

    Dvoriantchikova, Galina; Pappas, Steve; Luo, Xueting; Ribeiro, Marcio; Danek, Dagmara; Pelaez, Daniel; Park, Kevin K; Ivanov, Dmitry

    2016-12-01

    As axon damage and retinal ganglion cell (RGC) loss lead to blindness, therapies that increase RGC survival and axon regrowth have direct clinical relevance. Given that NFκB signaling is critical for neuronal survival and may regulate neurite growth, we investigated the therapeutic potential of NFκB signaling in RGC survival and axon regeneration. Although both NFκB subunits (p65 and p50) are present in RGCs, p65 exists in an inactive (unphosphorylated) state when RGCs are subjected to neurotoxic conditions. In this study, we used a phosphomimetic approach to generate DNA coding for an activated (phosphorylated) p65 (p65mut), then employed an adeno-associated virus serotype 2 (AAV2) to deliver the DNA into RGCs. We tested whether constitutive p65mut expression prevents death and facilitates neurite outgrowth in RGCs subjected to transient retinal ischemia or optic nerve crush (ONC), two models of neurotoxicity. Our data indicate that RGCs treated with AAV2-p65mut displayed a significant increase in survival compared to controls in ONC model (77 ± 7% vs. 25 ± 3%, P-value = 0.0001). We also found protective effect of modified p65 in RGCs of ischemic retinas (55 ± 12% vs. 35 ± 6%), but not to a statistically significant degree (P-value = 0.14). We did not detect a difference in axon regeneration between experimental and control animals after ONC. These findings suggest that increased NFκB signaling in RGCs attenuates retinal damage in animal models of neurodegeneration, but insignificantly impacts axon regeneration. © 2016 Federation of European Neuroscience Societies and John Wiley & Sons Ltd.

  4. α-Lipoic acid antioxidant treatment limits glaucoma-related retinal ganglion cell death and dysfunction.

    Directory of Open Access Journals (Sweden)

    Denise M Inman

    Full Text Available Oxidative stress has been implicated in neurodegenerative diseases, including glaucoma. However, due to the lack of clinically relevant models and expense of long-term testing, few studies have modeled antioxidant therapy for prevention of neurodegeneration. We investigated the contribution of oxidative stress to the pathogenesis of glaucoma in the DBA/2J mouse model of glaucoma. Similar to other neurodegenerative diseases, we observed lipid peroxidation and upregulation of oxidative stress-related mRNA and protein in DBA/2J retina. To test the role of oxidative stress in disease progression, we chose to deliver the naturally occurring, antioxidant α-lipoic acid (ALA to DBA/2J mice in their diet. We used two paradigms for ALA delivery: an intervention paradigm in which DBA/2J mice at 6 months of age received ALA in order to intervene in glaucoma development, and a prevention paradigm in which DBA/2J mice were raised on a diet supplemented with ALA, with the goal of preventing glaucoma development. At 10 and 12 months of age (after 4 and 11 months of dietary ALA respectively, we measured changes in genes and proteins related to oxidative stress, retinal ganglion cell (RGC number, axon transport, and axon number and integrity. Both ALA treatment paradigms showed increased antioxidant gene and protein expression, increased protection of RGCs and improved retrograde transport compared to control. Measures of lipid peroxidation, protein nitrosylation, and DNA oxidation in retina verified decreased oxidative stress in the prevention and intervention paradigms. These data demonstrate the utility of dietary therapy for reducing oxidative stress and improving RGC survival in glaucoma.

  5. Retinal ganglion cell complex changes using spectral domain optical coherence tomography in diabetic patients without retinopathy

    Institute of Scientific and Technical Information of China (English)

    Ahmed; I.Hegazy; Rasha; H.Zedan; Tamer; A.Macky; Soheir; M.Esmat

    2017-01-01

    AIM:To assess the ganglion cell complex(GCC)thickness in diabetic eyes without retinopathy. METHODS:Two groups included 45 diabetic eyes without retinopathy and 21 non diabetic eyes. All subjects underwent full medical and ophthalmological history,full ophthalmological examination,measuring GCC thickness and central foveal thickness(CFT)using the RTVue~? spectral domainoptical coherence tomography(SD-OCT),and HbA1C level.RESULTS:GCC focal loss volume(FLV%)was significantly more in diabetic eyes(22.2% below normal)than normal eyes(P=0.024). No statistically significant difference was found between the diabetic group and the control group regarding GCC global loss volume(GLV%)(P=0.160). CFT was positively correlated to the average,superior and inferior GCC(P=0.001,0.000 and 0.001 respectively)and negatively correlated to GLV% and FLV%(P=0.002 and0.031 respectively)in diabetic eyes. C/D ratio in diabetic eyes was negatively correlated to average,superior and inferior GCC(P=0.015,0.007 and 0.017 respectively). The FLV% was negatively correlated to the refraction and level of Hb A1c(P=0.019 and 0.013 respectively)and positively correlated to the best corrected visual acuity(BCVA)in log MAR in diabetic group(P=0.004).CONCLUSION:Significant GCC thinning in diabetes predates retinal vasculopathy,which is mainly focal rather than diffuse. It has no preference to either the superior or inferior halves of the macula. Increase of myopic error is significantly accompanied with increased focal GCC loss. GCC loss is accompanied with increased C/D ratio in diabetic eyes.

  6. Retinal ganglion cell complex changes using spectral domain optical coherence tomography in diabetic patients without retinopathy

    Directory of Open Access Journals (Sweden)

    Ahmed I. Hegazy

    2017-03-01

    Full Text Available AIM: To assess the ganglion cell complex (GCC thickness in diabetic eyes without retinopathy. METHODS: Two groups included 45 diabetic eyes without retinopathy and 21 non diabetic eyes. All subjects underwent full medical and ophthalmological history, full ophthalmological examination, measuring GCC thickness and central foveal thickness (CFT using the RTVue® spectral domain-optical coherence tomography (SD-OCT, and HbA1C level. RESULTS: GCC focal loss volume (FLV% was significantly more in diabetic eyes (22.2% below normal than normal eyes (P=0.024. No statistically significant difference was found between the diabetic group and the control group regarding GCC global loss volume (GLV% (P=0.160. CFT was positively correlated to the average, superior and inferior GCC (P=0.001, 0.000 and 0.001 respectively and negatively correlated to GLV% and FLV% (P=0.002 and 0.031 respectively in diabetic eyes. C/D ratio in diabetic eyes was negatively correlated to average, superior and inferior GCC (P=0.015, 0.007 and 0.017 respectively. The FLV% was negatively correlated to the refraction and level of HbA1c (P=0.019 and 0.013 respectively and positively correlated to the best corrected visual acuity (BCVA in logMAR in diabetic group (P=0.004. CONCLUSION: Significant GCC thinning in diabetes predates retinal vasculopathy, which is mainly focal rather than diffuse. It has no preference to either the superior or inferior halves of the macula. Increase of myopic error is significantly accompanied with increased focal GCC loss. GCC loss is accompanied with increased C/D ratio in diabetic eyes.

  7. Apoptosis in fish: environmental factors and programmed cell death.

    Science.gov (United States)

    AnvariFar, Hossein; Amirkolaie, Abdolsamad Keramat; Miandare, Hamed Kolangi; Ouraji, Hossein; Jalali, M Ali; Üçüncü, Sema İşisağ

    2017-06-01

    Apoptosis, a form of programmed cell death, is a critical component in maintaining homeostasis and growth in all tissues and plays a significant role in immunity and cytotoxicity. In contrast to necrosis or traumatic cell death, apoptosis is a well-controlled and vital process characterized mainly by cytoplasmic shrinkage, chromatin condensation, DNA fragmentation, membrane blebbing and apoptotic bodies. Our understanding of apoptosis is partly based on observations in invertebrates but mainly in mammals. Despite the great advantages of fish models in studying vertebrate development and diseases and the tremendous interest observed in recent years, reports on apoptosis in fish are still limited. Although apoptotic machinery is well conserved between aquatic and terrestrial organisms throughout the history of evolution, some differences exist in key components of apoptotic pathways. Core parts of apoptotic machinery in fish are virtually expressed as equivalent to the mammalian models. Some differences are, however, evident, such as the extrinsic and intrinsic pathways of apoptosis including lack of a C-terminal region in the Fas-associated protein with a death domain in fish. Aquatic species inhabit a complex and highly fluctuating environment, making these species good examples to reveal features of apoptosis that may not be easily investigated in mammals. Therefore, in order to gain a wider view on programmed cell death in fish, interactions between the main environmental factors, chemicals and apoptosis are discussed in this review. It is indicated that apoptosis can be induced in fish by exposure to environmental stressors during different stages of the fish life cycle.

  8. High expression of markers of apoptosis in Langerhans cell histiocytosis

    DEFF Research Database (Denmark)

    Petersen, Bodil Laub; Lundegaard, Pia Rengtved; Bank, M I

    2003-01-01

    53 and the number of cells in apoptosis detected with TUNEL. Langerhans cell histiocytosis cells showed strong expression of p53 and in some cases co-expression of Fas and Fas-L. The expression of Fas-L was significantly higher in infiltrates from patients with single-system disease. The actual...... number of pathological Langerhans cells in apoptosis as estimated by TUNEL was low. CONCLUSIONS: The low number of TUNEL-reactive cells can be explained by the rapid turnover of apoptotic cells in the tissue, not leaving the apoptotic cells long enough in the tissue to be detected. The co......-expression of Fas and Fas-L in some Langerhans cells can lead to an autocrine apoptotic shortcut, mediating the death of the double-positive cells. Our findings suggest that apoptosis mediated through the Fas/Fas-L pathway may contribute to the spontaneous regression of lesions in single-system disease. A delicate...

  9. Transcriptome of Atoh7 retinal progenitor cells identifies new Atoh7-dependent regulatory genes for retinal ganglion cell formation.

    Science.gov (United States)

    Gao, Zhiguang; Mao, Chai-An; Pan, Ping; Mu, Xiuqian; Klein, William H

    2014-11-01

    The bHLH transcription factor ATOH7 (Math5) is essential for establishing retinal ganglion cell (RGC) fate. However, Atoh7-expressing retinal progenitor cells (RPCs) can give rise to all retinal cell types, suggesting that other factors are involved in specifying RGCs. The basis by which a subpopulation of Atoh7-expressing RPCs commits to an RGC fate remains uncertain but is of critical importance to retinal development since RGCs are the earliest cell type to differentiate. To better understand the regulatory mechanisms leading to cell-fate specification, a binary genetic system was generated to specifically label Atoh7-expressing cells with green fluorescent protein (GFP). Fluorescence-activated cell sorting (FACS)-purified GFP(+) and GFP(-) cells were profiled by RNA-seq. Here, we identify 1497 transcripts that were differentially expressed between the two RPC populations. Pathway analysis revealed diminished growth factor signaling in Atoh7-expressing RPCs, indicating that these cells had exited the cell cycle. In contrast, axon guidance signals were enriched, suggesting that axons of Atoh7-expressing RPCs were already making synaptic connections. Notably, many genes enriched in Atoh7-expressing RPCs encoded transcriptional regulators, and several were direct targets of ATOH7, including, and unexpectedly, Ebf3 and Eya2. We present evidence for a Pax6-Atoh7-Eya2 pathway that acts downstream of Atoh7 but upstream of differentiation factor Pou4f2. EYA2 is a protein phosphatase involved in protein-protein interactions and posttranslational regulation. These properties, along with Eya2 as an early target gene of ATOH7, suggest that EYA2 functions in RGC specification. Our results expand current knowledge of the regulatory networks operating in Atoh7-expressing RPCs and offer new directions for exploring the earliest aspects of retinogenesis. © 2014 Wiley Periodicals, Inc.

  10. Inhibition of microparticle release triggers endothelial cell apoptosis and detachment

    NARCIS (Netherlands)

    Abid Hussein, Mohammed N.; Böing, Anita N.; Sturk, Augueste; Hau, Chi M.; Nieuwland, Rienk

    2007-01-01

    Endothelial cell cultures contain caspase 3-containing microparticles (EMP), which are reported to form during or after cell detachment. We hypothesize that also adherent endothelial cells release EMP, thus protecting these cells from caspase 3 accumulation, detachment and apoptosis. Human umbilical

  11. CDB-4124 does not cause apoptosis in cultured fibroid cells.

    Science.gov (United States)

    Roeder, Hilary; Jayes, Friederike; Feng, Liping; Leppert, Phyllis C

    2011-09-01

    Selective progesterone receptor modulators (SPRMs), such as asoprisnil (J867) and ulipristal (CDB-2914), have been shown to reduce fibroid volume in vivo and to induce apoptosis in vitro. CDB-4124 (telapristone), a SPRM with different side groups, also reduced fibroid volume in vivo, and we hypothesized that this SPRM would also cause apoptosis in cultured fibroid cells. Immortalized, progesterone receptor-positive fibroid cells, known to be capable of apoptosis, were grown to 80% confluence in serum-containing media. Cells were then treated for 48 hours in serum-free media with 0, 10, 100, or 1000 nmol/L CDB-4124. Actinomycin-D and staurosporine were used as positive controls to induce apoptosis. Apoptosis was quantified using a TUNEL-fluorescein kit. Images were captured with a widefield-fluorescence microscope and analyzed using MetaMorph image analysis software. To validate results, Western blots of total cell lysates were probed for cleaved caspase-3 (c-CASP3). Experiments were repeated 3 times using independent cell batches. Analysis of 19 712 nuclei indicated 14.8% ± 10.9% (mean ± SEM), 8.4% ± 4.6%, 8.2% ± 4.7%, and 9.3% ± 6.3% apoptosis in 0, 10, 100, and 1000 nmol/L CDB-4124-treated cells, respectively. There was no evidence of elevated c-CASP3 over vehicle control after treatment with CDB-4124. CDB-4124 did not significantly induce apoptosis in cultured fibroid cells under the conditions described suggesting apoptosis may not be the main pathway responsible for CDB-4124-induced fibroid shrinkage. Variations in SPRM biological effects may be due to differences in fibroid source cells, binding kinetics, or extracellular matrix characteristics, and can be exploited in further investigations of the mechanisms of action of SPRMs in fibroid biology.

  12. Apoptosis (programmed cell death) as an indicator of xenobiotic toxicity

    International Nuclear Information System (INIS)

    Bond, G.P.

    1989-01-01

    Xenobiotics alter the frequency and pattern of apoptosis (programmed cell death). Preliminary studies identified the mouse liver, with normally low levels of apoptosis, as a preferable test system to the chicken embryo limb, with normally high levels of apoptosis. The major purposes of these investigations, using the apoptogen and necrogen 1,1-dichloroethylene (DCE), were to determine if increases in apoptosis, (1) could be quantified as a direct result of treatment, (2) were dose- and time-dependent, (3) were independent of necrosis, (4) were associated with mitosis in the control of cell numbers and (5) were limited to specific areas of the liver. To these ends, food-deprived female, CF-1 mice were administered DCE ip under varying experimental conditions. Increased apoptosis occurred in a dose- and time-dependent manner after treatment with 12.5, 40, and 125 mg/kg for 0.5, 1, 2, 4 and 8 hr. Peak effects were observed at 4 hr. Apoptosis occurred only in the midzonal/pericentral areas of the liver. At 12.5 mg/kg, there were no effects on biochemical (alanine transaminase) and morphological indices of necrosis, establishing apoptosis as a separate phenomenon from necrosis. Increased 3 H-thymidine incorporation (DNA synthesis), mitosis and the percentage of octaploid hepatocytes occurred from 24-48 hr after treatment with the apoptotic but non-necrotic dose of 40 mg/kg. Apoptosis only occurred in the midzonal/pericentral areas of the liver after multiple doses with DCE, indicating the zonal selectivity of the response. In conclusion, apoptosis, a normally occurring homeostatic process associated with mitosis in the control of cell numbers, is affected by selected xenobiotics in a dose-dependent manner. Xenobiotic-induced apoptosis in the liver occurs at low doses of xenobiotics which cause no other effects on tissue structure or function

  13. Orexin-A potentiates L-type calcium/barium currents in rat retinal ganglion cells.

    Science.gov (United States)

    Liu, F; Weng, S-J; Yang, X-L; Zhong, Y-M

    2015-10-01

    Two neuropeptides, orexin-A and orexin-B (also called hypocretin-1 and -2), have been implicated in sleep/wake regulation, feeding behaviors via the activation of two subtypes of G-protein-coupled receptors: orexin 1 and orexin 2 receptors (OX1R and OX2R). While the expression of orexins and orexin receptors is immunohistochemically revealed in retinal neurons, the function of these peptides in the retina is largely unknown. Using whole-cell patch-clamp recordings in rat retinal slices, we demonstrated that orexin-A increased L-type-like barium currents (IBa,L) in ganglion cells (GCs), and the effect was blocked by the selective OX1R antagonist SB334867, but not by the OX2R antagonist TCS OX2 29. The orexin-A effect was abolished by intracellular dialysis of GDP-β-S/GPAnt-2A, a Gq protein inhibitor, suggesting the mediation of Gq. Additionally, during internal dialysis of the phosphatidylinositol (PI)-phospholipase C (PLC) inhibitor U73122, orexin-A did not change the IBa,L of GCs, whereas the orexin-A effect persisted in the presence of the phosphatidylcholine (PC)-PLC inhibitor D609. The orexin-A-induced potentiation was not seen with internal infusion of Ca(2+)-free solution or when inositol 1,4,5-trisphosphate (IP3)-sensitive Ca(2+) release from intracellular stores was blocked by heparin/xestospongins-C. Moreover, the orexin-A effect was mimicked by the protein kinase C (PKC) activator phorbol 12-myristate 13-acetate, but was eliminated when PKC was inhibited by bisindolylmaleimide IV (Bis-IV)/Gö6976. Neither adenosine 3',5'-cyclic monophosphate (cAMP)-protein kinase A (PKA) nor guanosine 3',5'-cyclic monophosphate (cGMP)-protein kinase G (PKG) signaling pathway was likely involved, as orexin-A persisted to potentiate the IBa,L of GCs no matter these two pathways were activated or inhibited. These results suggest that, by activating OX1R, orexin-A potentiates the IBa,L of rat GCs through a distinct Gq/PI-PLC/IP3/Ca(2+)/PKC signaling pathway. Copyright

  14. Coatings of Different Carbon Nanotubes on Platinum Electrodes for Neuronal Devices: Preparation, Cytocompatibility and Interaction with Spiral Ganglion Cells.

    Science.gov (United States)

    Burblies, Niklas; Schulze, Jennifer; Schwarz, Hans-Christoph; Kranz, Katharina; Motz, Damian; Vogt, Carla; Lenarz, Thomas; Warnecke, Athanasia; Behrens, Peter

    2016-01-01

    Cochlear and deep brain implants are prominent examples for neuronal prostheses with clinical relevance. Current research focuses on the improvement of the long-term functionality and the size reduction of neural interface electrodes. A promising approach is the application of carbon nanotubes (CNTs), either as pure electrodes but especially as coating material for electrodes. The interaction of CNTs with neuronal cells has shown promising results in various studies, but these appear to depend on the specific type of neurons as well as on the kind of nanotubes. To evaluate a potential application of carbon nanotube coatings for cochlear electrodes, it is necessary to investigate the cytocompatibility of carbon nanotube coatings on platinum for the specific type of neuron in the inner ear, namely spiral ganglion neurons. In this study we have combined the chemical processing of as-delivered CNTs, the fabrication of coatings on platinum, and the characterization of the electrical properties of the coatings as well as a general cytocompatibility testing and the first cell culture investigations of CNTs with spiral ganglion neurons. By applying a modification process to three different as-received CNTs via a reflux treatment with nitric acid, long-term stable aqueous CNT dispersions free of dispersing agents were obtained. These were used to coat platinum substrates by an automated spray-coating process. These coatings enhance the electrical properties of platinum electrodes, decreasing the impedance values and raising the capacitances. Cell culture investigations of the different CNT coatings on platinum with NIH3T3 fibroblasts attest an overall good cytocompatibility of these coatings. For spiral ganglion neurons, this can also be observed but a desired positive effect of the CNTs on the neurons is absent. Furthermore, we found that the well-established DAPI staining assay does not function on the coatings prepared from single-wall nanotubes.

  15. Time-Dependent Nerve Growth Factor Signaling Changes in the Rat Retina During Optic Nerve Crush-Induced Degeneration of Retinal Ganglion Cells

    Directory of Open Access Journals (Sweden)

    Louise A. Mesentier-Louro

    2017-01-01

    Full Text Available Nerve growth factor (NGF is suggested to be neuroprotective after nerve injury; however, retinal ganglion cells (RGC degenerate following optic-nerve crush (ONC, even in the presence of increased levels of endogenous NGF. To further investigate this apparently paradoxical condition, a time-course study was performed to evaluate the effects of unilateral ONC on NGF expression and signaling in the adult retina. Visually evoked potential and immunofluorescence staining were used to assess axonal damage and RGC loss. The levels of NGF, proNGF, p75NTR, TrkA and GFAP and the activation of several intracellular pathways were analyzed at 1, 3, 7 and 14 days after crush (dac by ELISA/Western Blot and PathScan intracellular signaling array. The progressive RGC loss and nerve impairment featured an early and sustained activation of apoptotic pathways; and GFAP and p75NTR enhancement. In contrast, ONC-induced reduction of TrkA, and increased proNGF were observed only at 7 and 14 dac. We propose that proNGF and p75NTR contribute to exacerbate retinal degeneration by further stimulating apoptosis during the second week after injury, and thus hamper the neuroprotective effect of the endogenous NGF. These findings might aid in identifying effective treatment windows for NGF-based strategies to counteract retinal and/or optic-nerve degeneration.

  16. Time-Dependent Nerve Growth Factor Signaling Changes in the Rat Retina During Optic Nerve Crush-Induced Degeneration of Retinal Ganglion Cells.

    Science.gov (United States)

    Mesentier-Louro, Louise A; De Nicolò, Sara; Rosso, Pamela; De Vitis, Luigi A; Castoldi, Valerio; Leocani, Letizia; Mendez-Otero, Rosalia; Santiago, Marcelo F; Tirassa, Paola; Rama, Paolo; Lambiase, Alessandro

    2017-01-05

    Nerve growth factor (NGF) is suggested to be neuroprotective after nerve injury; however, retinal ganglion cells (RGC) degenerate following optic-nerve crush (ONC), even in the presence of increased levels of endogenous NGF. To further investigate this apparently paradoxical condition, a time-course study was performed to evaluate the effects of unilateral ONC on NGF expression and signaling in the adult retina. Visually evoked potential and immunofluorescence staining were used to assess axonal damage and RGC loss. The levels of NGF, proNGF, p75 NTR , TrkA and GFAP and the activation of several intracellular pathways were analyzed at 1, 3, 7 and 14 days after crush (dac) by ELISA/Western Blot and PathScan intracellular signaling array. The progressive RGC loss and nerve impairment featured an early and sustained activation of apoptotic pathways; and GFAP and p75 NTR enhancement. In contrast, ONC-induced reduction of TrkA, and increased proNGF were observed only at 7 and 14 dac. We propose that proNGF and p75 NTR contribute to exacerbate retinal degeneration by further stimulating apoptosis during the second week after injury, and thus hamper the neuroprotective effect of the endogenous NGF. These findings might aid in identifying effective treatment windows for NGF-based strategies to counteract retinal and/or optic-nerve degeneration.

  17. Isolation and Molecular Profiling of Primary Mouse Retinal Ganglion Cells: Comparison of Phenotypes from Healthy and Glaucomatous Retinas

    OpenAIRE

    Chintalapudi, Sumana R.; Djenderedjian, Levon; Stiemke, Andrew B.; Steinle, Jena J.; Jablonski, Monica M.; Morales-Tirado, Vanessa M.

    2016-01-01

    Loss of functional retinal ganglion cells (RGC) is an element of retinal degeneration that is poorly understood. This is in part due to the lack of a reliable and validated protocol for the isolation of primary RGCs. Here we optimize a feasible, reproducible, standardized flow cytometry-based protocol for the isolation and enrichment of homogeneous RGC with the Thy1.2hiCD48negCD15negCD57neg surface phenotype. A three-step validation process was performed by: (1) genomic profiling of 25-genes ...

  18. Retinal ganglion cells: mechanisms underlying depolarization block and differential responses to high frequency electrical stimulation of ON and OFF cells

    Science.gov (United States)

    Kameneva, T.; Maturana, M. I.; Hadjinicolaou, A. E.; Cloherty, S. L.; Ibbotson, M. R.; Grayden, D. B.; Burkitt, A. N.; Meffin, H.

    2016-02-01

    Objective. ON and OFF retinal ganglion cells (RGCs) are known to have non-monotonic responses to increasing amplitudes of high frequency (2 kHz) biphasic electrical stimulation. That is, an increase in stimulation amplitude causes an increase in the cell’s spike rate up to a peak value above which further increases in stimulation amplitude cause the cell to decrease its activity. The peak response for ON and OFF cells occurs at different stimulation amplitudes, which allows differential stimulation of these functional cell types. In this study, we investigate the mechanisms underlying the non-monotonic responses of ON and OFF brisk-transient RGCs and the mechanisms underlying their differential responses. Approach. Using in vitro patch-clamp recordings from rat RGCs, together with simulations of single and multiple compartment Hodgkin-Huxley models, we show that the non-monotonic response to increasing amplitudes of stimulation is due to depolarization block, a change in the membrane potential that prevents the cell from generating action potentials. Main results. We show that the onset for depolarization block depends on the amplitude and frequency of stimulation and reveal the biophysical mechanisms that lead to depolarization block during high frequency stimulation. Our results indicate that differences in transmembrane potassium conductance lead to shifts of the stimulus currents that generate peak spike rates, suggesting that the differential responses of ON and OFF cells may be due to differences in the expression of this current type. We also show that the length of the axon’s high sodium channel band (SOCB) affects non-monotonic responses and the stimulation amplitude that leads to the peak spike rate, suggesting that the length of the SOCB is shorter in ON cells. Significance. This may have important implications for stimulation strategies in visual prostheses.

  19. Overexpression of Pax6 results in microphthalmia, retinal dysplasia and defective retinal ganglion cell axon guidance

    Directory of Open Access Journals (Sweden)

    Jeffery Glen

    2008-05-01

    Full Text Available Abstract Background The transcription factor Pax6 is expressed by many cell types in the developing eye. Eyes do not form in homozygous loss-of-function mouse mutants (Pax6Sey/Sey and are abnormally small in Pax6Sey/+ mutants. Eyes are also abnormally small in PAX77 mice expressing multiple copies of human PAX6 in addition to endogenous Pax6; protein sequences are identical in the two species. The developmental events that lead to microphthalmia in PAX77 mice are not well-characterised, so it is not clear whether over- and under-expression of Pax6/PAX6 cause microphthalmia through similar mechanisms. Here, we examined the consequences of over-expression for the eye and its axonal connections. Results Eyes form in PAX77+/+ embryos but subsequently degenerate. At E12.5, we found no abnormalities in ocular morphology, retinal cell cycle parameters and the incidence of retinal cell death. From E14.5 on, we observed malformations of the optic disc. From E16.5 into postnatal life there is progressively more severe retinal dysplasia and microphthalmia. Analyses of patterns of gene expression indicated that PAX77+/+ retinae produce a normal range of cell types, including retinal ganglion cells (RGCs. At E14.5 and E16.5, quantitative RT-PCR with probes for a range of molecules associated with retinal development showed only one significant change: a slight reduction in levels of mRNA encoding the secreted morphogen Shh at E16.5. At E16.5, tract-tracing with carbocyanine dyes in PAX77+/+ embryos revealed errors in intraretinal navigation by RGC axons, a decrease in the number of RGC axons reaching the thalamus and an increase in the proportion of ipsilateral projections among those RGC axons that do reach the thalamus. A survey of embryos with different Pax6/PAX6 gene dosage (Pax6Sey/+, Pax6+/+, PAX77+ and PAX77+/+ showed that (1 the total number of RGC axons projected by the retina and (2 the proportions that are sorted into the ipsilateral and

  20. Ionizing radiation induces apoptosis in hematopoietic stem and progenitor cells

    International Nuclear Information System (INIS)

    Meng, A.; Zhou, D.; Geiger, H.; Zant, G.V.

    2003-01-01

    The aims of this study was to determine if ionizing radiation (IR) induces apoptosis in hematopoietic stem (HSC) and progenitor cells. Lin-cells were isolated from mouse bone marrow (BM) and pretreated with vehicle or 100 μM z-VAD 1 h prior to exposure to 4 Gy IR. The apoptotic and/or necrotic responses of these cells to IR were analyzed by measuring the annexin V and/or 7-AAD staining in HSC and progenitor populations using flow cytometry, and hematopoietic function of these cells was determined by CAFC assay. Exposure of Lin-cells to IR selectively decreased the numbers of HSC and progenitors in association with an increase in apoptosis in a time-dependent manner. Pretreatment of Lin- cells with z-VAD significantly inhibited IR-induced apoptosis and the decrease in the numbers of HSC and progenitors. However, IR alone or in combination with z-VAD did not lead to a significant increase in necrotic cell death in either HSC or progenitors. In addition, pretreatment of BM cells with z-VAD significantly attenuated IR-induced reduction in the frequencies of day-7, -28 and -35 CAFC. Exposure of HSC and progenitors to IR induces apoptosis. The induction of HSC and progenitor apoptosis contributes to IR-induced suppression of their hematopoietic function

  1. Activation of the ζ receptor 1 suppresses NMDA responses in rat retinal ganglion cells.

    Science.gov (United States)

    Zhang, X-J; Liu, L-L; Jiang, S-X; Zhong, Y-M; Yang, X-L

    2011-03-17

    The sigma receptor 1 (σR1) has been shown to modulate the activity of several voltage- and ligand-gated channels. Using patch-clamp techniques in rat retinal slice preparations, we demonstrated that activation of σR1 by SKF10047 (SKF) or PRE-084 suppressed N-methyl-D-aspartate (NMDA) receptor-mediated current responses from both ON and OFF type ganglion cells (GCs), dose-dependently, and the effect could be blocked by the σR1 antagonist BD1047 or the σR antagonist haloperidol. The suppression by SKF of NMDA currents was abolished with pre-incubation of the G protein inhibitor GDP-β-S or the Gi/o activator mastoparan. We further explored the intracellular signaling pathway responsible for the SKF-induced suppression of NMDA responses. Application of either cAMP/the PKA inhibitor Rp-cAMP or cGMP/the PKG inhibitor KT5823 did not change the SKF-induced effect, suggesting the involvement of neither cAMP/PKA nor cGMP/PKG pathway. In contrast, suppression of NMDA responses by SKF was abolished by internal infusion of the phosphatidylinostiol-specific phospholipase C (PLC) inhibitor U73122, but not by the phosphatidylcholine-PLC inhibitor D609. SKF-induced suppression of NMDA responses was dependent on intracellular Ca2+ concentration ([Ca2+]i), as evidenced by the fact that the effect was abolished when [Ca2+]i was buffered with 10 mM BAPTA. The SKF effect was blocked by xestospongin-C/heparin, IP3 receptor antagonists, but unchanged by ryanodine/caffeine, ryanodine receptor modulators. Furthermore, application of protein kinase C inhibitors Bis IV and Gö6976 eliminated the SKF effect. These results suggest that the suppression of NMDA responses of rat retinal GCs caused by the activation of σR1 may be mediated by a distinct [Ca2+]i-dependent PLC-PKC pathway. This effect of SKF could help ameliorate malfunction of GCs caused by excessive stimulation of NMDA receptors under pathological conditions. Copyright © 2011 IBRO. Published by Elsevier Ltd. All rights

  2. Protective effects of triptolide on retinal ganglion cells in a rat model of chronic glaucoma

    Directory of Open Access Journals (Sweden)

    Yang F

    2015-11-01

    Full Text Available Fan Yang, Dongmei Wang, Lingling Wu, Ying Li Ophthalmology Department, Peking University Third Hospital, Beijing, People’s Republic of China Purpose: To study the effects of triptolide, a Chinese herb extract, on retinal ganglion cells (RGCs in a rat model of chronic glaucoma.Methods: Eighty Wistar rats were randomly divided into triptolide group (n=40 and normal saline (NS group (n=40. Angle photocoagulation was used to establish the model of glaucoma, with right eye as laser treated eye and left eye as control eye. Triptolide group received triptolide intraperitoneally daily, while NS group received NS. Intraocular pressure (IOP, anti-CD11b immunofluorescent stain in retina and optic nerve, RGCs count with Nissel stain and microglia count with anti-CD11b immunofluorescence stain in retina flat mounts, retinal tumor necrosis factor (TNF-α mRNA detection by reverse transcription–polymerase chain reaction, and double immunofluorescent labeling with anti-TNF-α and anti-CD11b in retinal frozen section were performed.Results: Mean IOP of the laser treated eyes significantly increased 3 weeks after photocoagulation (P<0.05, with no statistical difference between the two groups (P>0.05. RGCs survival in the laser treated eyes was significantly improved in the triptolide group than the NS group (P<0.05. Microglia count in superficial retina of the laser treated eyes was significantly less in the triptolide group (30.40±4.90 than the NS group (35.06±7.59 (P<0.05. TNF-α mRNA expression in the retina of the laser treated eyes in the triptolide group decreased by 60% compared with that in the NS group (P<0.01. The double immunofluorescent labeling showed that TNF-α was mainly distributed around the microglia.Conclusion: Triptolide improved RGCs survival in this rat model of chronic glaucoma, which did not depend on IOP decrease but might be exerted by inhibiting microglia activities and reducing TNF-α secretion. Keywords: glaucoma, triptolide

  3. Retinal ganglion cell-inner plexiform and nerve fiber layers in neuromyelitis optica.

    Science.gov (United States)

    Hu, Sai-Jing; Lu, Pei-Rong

    2018-01-01

    To determine the thickness of the retinal ganglion cell-inner plexiform layer (GCIPL) and the retinal nerve fiber layer (RNFL) in patients with neuromyelitis optica (NMO). We conducted a cross-sectional study that included 30 NMO patients with a total of 60 eyes. Based on the presence or absence of optic neuritis (ON), subjects were divided into either the NMO-ON group (30 eyes) or the NMO-ON contra group (10 eyes). A detailed ophthalmologic examination was performed for each group; subsequently, the GCIPL and the RNFL were measured using high-definition optical coherence tomography (OCT). In the NMO-ON group, the mean GCIPL thickness was 69.28±21.12 µm, the minimum GCIPL thickness was 66.02±10.02 µm, and the RNFL thickness were 109.33±11.23, 110.47±3.10, 64.92±12.71 and 71.21±50.22 µm in the superior, inferior, temporal and nasal quadrants, respectively. In the NMO-ON contra group, the mean GCIPL thickness was 85.12±17.09 µm, the minimum GCIPL thickness was 25.39±25.1 µm, and the RNFL thicknesses were 148.33±23.22, 126.36±23.45, 82.21±22.30 and 83.36±31.28 µm in the superior, inferior, temporal and nasal quadrants, respectively. In the control group, the mean GCIPL thickness was 86.98±22.37 µm, the minimum GCIPL thickness was 85.28±10.75 µm, and the RNFL thicknesses were 150.22±22.73, 154.79±60.23, 82.33±7.01 and 85.62±13.81 µm in the superior, inferior, temporal and nasal quadrants, respectively. The GCIPL and RNFL were thinner in the NMO-ON contra group than in the control group ( P deviation (MD) and corrected pattern standard deviation (PSD) in the NMO-ON group ( P <0.05). The thickness of the GCIPL and RNFL, as measured using OCT, may indicate optic nerve damage in patients with NMO.

  4. Neuroprotection of rat retinal ganglion cells mediated through alpha7 nicotinic acetylcholine receptors.

    Science.gov (United States)

    Iwamoto, K; Mata, D; Linn, D M; Linn, C L

    2013-05-01

    Glutamate-induced excitotoxicity is thought to play an important role in several neurodegenerative diseases in the central nervous system (CNS). In this study, neuroprotection against glutamate-induced excitotoxicity was analyzed using acetylcholine (ACh), nicotine and the α7 specific nicotinic acetylcholine receptor (α7 nAChR) agonist, N-[(3R)-1-azabicyclo[2.2.2]oct-3-yl]-4-chlorobenzamide hydrochloride (PNU-282987), in cultured adult rat retinal neurons. Adult Long Evans rat retinas were dissociated and retinal ganglion cells (RGCs) were isolated from all other retinal tissue using a two-step panning technique. Once isolated, RGCs were cultured under various pharmacological conditions to demonstrate excitotoxicity and neuroprotection against excitotoxicity. After 3 days, RGCs were immunostained with antibodies against the glycoprotein, Thy 1.1, counted and cell survival was assessed relative to control untreated conditions. 500 μM glutamate induced excitotoxicity in large and small RGCs in an adult rat dissociated culture. After 3 days in culture with glutamate, the cell survival of large RGCs decreased by an average of 48.16% while the cell survival of small RGCs decreased by an average of 42.03%. Using specific glutamate receptor agonists and antagonists, we provide evidence that the excitotoxic response was mediated through α-amino-3-hydroxy-5-methyl-4-isoxazolepropionic acid (AMPA)/kainic acid (KA) and N-methyl-d-aspartate (NMDA) glutamate receptors through an apoptotic mechanism. However, the excitotoxic effect of glutamate on all RGCs was eliminated if cells were cultured for an hour with 10 μM ACh, 100 μM nicotine or 100 nM of the α7 nAChR agonist, PNU-282987, before the glutamate insult. Inhibition studies using 10nM methyllycaconitine (MLA) or α-bungarotoxin (α-Bgt) supported the hypothesis that neuroprotection against glutamate-induced excitotoxicity on rat RGCs was mediated through α7 nAChRs. In immunocytochemical studies, double

  5. Hypoxia Induces a Metabolic Shift and Enhances the Stemness and Expansion of Cochlear Spiral Ganglion Stem/Progenitor Cells

    Directory of Open Access Journals (Sweden)

    Hsin-Chien Chen

    2015-01-01

    Full Text Available Previously, we demonstrated that hypoxia (1% O2 enhances stemness markers and expands the cell numbers of cochlear stem/progenitor cells (SPCs. In this study, we further investigated the long-term effect of hypoxia on stemness and the bioenergetic status of cochlear spiral ganglion SPCs cultured at low oxygen tensions. Spiral ganglion SPCs were obtained from postnatal day 1 CBA/CaJ mouse pups. The measurement of oxygen consumption rate, extracellular acidification rate (ECAR, and intracellular adenosine triphosphate levels corresponding to 20% and 5% oxygen concentrations was determined using a Seahorse XF extracellular flux analyzer. After low oxygen tension cultivation for 21 days, the mean size of the hypoxia-expanded neurospheres was significantly increased at 5% O2; this correlated with high-level expression of hypoxia-inducible factor-1 alpha (Hif-1α, proliferating cell nuclear antigen (PCNA, cyclin D1, Abcg2, nestin, and Nanog proteins but downregulated expression of p27 compared to that in a normoxic condition. Low oxygen tension cultivation tended to increase the side population fraction, with a significant difference found at 5% O2 compared to that at 20% O2. In addition, hypoxia induced a metabolic energy shift of SPCs toward higher basal ECARs and higher maximum mitochondrial respiratory capacity but lower proton leak than under normoxia, where the SPC metabolism was switched toward glycolysis in long-term hypoxic cultivation.

  6. Co-expression of two subtypes of melatonin receptor on rat M1-type intrinsically photosensitive retinal ganglion cells.

    Directory of Open Access Journals (Sweden)

    Wen-Long Sheng

    Full Text Available Intrinsically photosensitive retinal ganglion cells (ipRGCs are involved in circadian and other non-image forming visual responses. An open question is whether the activity of these neurons may also be under the regulation mediated by the neurohormone melatonin. In the present work, by double-staining immunohistochemical technique, we studied the expression of MT1 and MT2, two known subtypes of mammalian melatonin receptors, in rat ipRGCs. A single subset of retinal ganglion cells labeled by the specific antibody against melanopsin exhibited the morphology typical of M1-type ipRGCs. Immunoreactivity for both MT1 and MT2 receptors was clearly seen in the cytoplasm of all labeled ipRGCs, indicating that these two receptors were co-expressed in each of these neurons. Furthermore, labeling for both the receptors were found in neonatal M1 cells as early as the day of birth. It is therefore highly plausible that retinal melatonin may directly modulate the activity of ipRGCs, thus regulating non-image forming visual functions.

  7. Verocytotoxin-induced apoptosis of human microvascular endothelial cells.

    Science.gov (United States)

    Pijpers, A H; van Setten, P A; van den Heuvel, L P; Assmann, K J; Dijkman, H B; Pennings, A H; Monnens, L A; van Hinsbergh, V W

    2001-04-01

    The pathogenesis of the epidemic form of hemolytic uremic syndrome is characterized by endothelial cell damage. In this study, the role of apoptosis in verocytotoxin (VT)-mediated endothelial cell death in human glomerular microvascular endothelial cells (GMVEC), human umbilical vein endothelial cells, and foreskin microvascular endothelial cells (FMVEC) was investigated. VT induced apoptosis in GMVEC and human umbilical vein endothelial cells when the cells were prestimulated with the inflammatory mediator tumor necrosis factor-alpha (TNF-alpha). FMVEC displayed strong binding of VT and high susceptibility to VT under basal conditions, which made them suitable for the study of VT-induced apoptosis without TNF-alpha interference. On the basis of functional (flow cytometry and immunofluorescence microscopy using FITC-conjugated annexin V and propidium iodide), morphologic (transmission electron microscopy), and molecular (agarose gel electrophoresis of cellular DNA fragments) criteria, it was documented that VT induced programmed cell death in microvascular endothelial cells in a dose- and time-dependent manner. Furthermore, whereas partial inhibition of protein synthesis by VT was associated with a considerable number of apoptotic cells, comparable inhibition of protein synthesis by cycloheximide was not. This suggests that additional pathways, independent of protein synthesis inhibition, may be involved in VT-mediated apoptosis in microvascular endothelial cells. Specific inhibition of caspases by Ac-Asp-Glu-Val-Asp-CHO, but not by Ac-Tyr-Val-Ala-Asp-CHO, was accompanied by inhibition of VT-induced apoptosis in FMVEC and TNF-alpha-treated GMVEC. These data indicate that VT can induce apoptosis in human microvascular endothelial cells.

  8. Induction of apoptosis by eugenol in human breast cancer cells

    International Nuclear Information System (INIS)

    Vidhya, N.; Niranjali Devaraj, S.

    2011-01-01

    In the present study, potential anticancer effect of eugenol on inhibition of cell proliferation and induction of apoptosis in human MCF-7 breast cancer cells was investigated. Induction of cell death by eugenol was evaluated following MTT assay and monitoring lactate dehydrogenase released into the culture medium for cell viability and cytotoxicity, giemsa staining for morphological alterations, fluorescence microscopy analysis of cells using ethidium bromide and acridine orange and quantitation of DNA fragments for induction of apoptosis. Effect of eugenol on intracellular redox status of the human breast cancer cells was assessed by determining the level of glutathione and lipid peroxidation products (TBARS). Eugenol treatment inhibited the growth and proliferation of human MCF-7 breast cancer cells through induction of cell death, which was dose and time dependent. Microscopic examination of eugenol treated cells showed cell shrinkage, membrane blebbing and apoptotic body formation. Further, eugenol treatment also depleted the level of intracellular glutathione and increased the level of lipid peroxidation. The dose dependent increase in the percentage of apoptotic cells and DNA fragments suggested that apoptosis was involved in eugenol induced cell death and apoptosis might have played a role in the chemopreventive action of eugenol. (author)

  9. Renal cell apoptosis in human lupus nephritis: a histological study

    DEFF Research Database (Denmark)

    Faurschou, M; Penkowa, Milena; Andersen, C B

    2009-01-01

    Nuclear autoantigens from apoptotic cells are believed to drive the immunological response in systemic lupus erythematosus (SLE). Conflicting data exist as to the possible renal origin of apoptotic cells in SLE patients with nephritis. We assessed the level of renal cell apoptosis in kidney...

  10. Andrographolide sensitizes prostate cancer cells to TRAIL-induced apoptosis

    Directory of Open Access Journals (Sweden)

    Ruo-Jing Wei

    2018-01-01

    Full Text Available Tumor necrosis factor-related apoptosis-inducing ligand (TRAIL is a promising agent for anticancer therapy. The identification of small molecules that can establish the sensitivity of prostate cancer (PCa cells to TRAIL-induced apoptosis is crucial for the targeted treatment of PCa. PC3, DU145, JAC-1, TsuPr1, and LNCaP cells were treated with Andrographolide (Andro and TRAIL, and the apoptosis was measured using the Annexin V/PI double staining method. Real time-polymerase chain reaction (PCR and Western blot analysis were performed to measure the expression levels of target molecules. RNA interference technique was used to down-regulate the expression of the target protein. We established a nude mouse xenograft model of PCa, which was used to measure the caspase-3 activity in the tumor cells using flow cytometry. In this research study, our results demonstrated that Andro preferentially increased the sensitivity of PCa cells to TRAIL-induced apoptosis at subtoxic concentrations, and the regulation mechanism was related to the up-regulation of DR4. In addition, it also increased the p53 expression and led to the generation of reactive oxygen species (ROS in the cells. Further research revealed that the DR4 inhibition, p53 expression, and ROS generation can significantly reduce the apoptosis induced by the combination of TRAIL and Andro in PCa cells. In conclusion, Andro increases the sensitivity of PCa cells to TRAIL-induced apoptosis through the generation of ROS and up-regulation of p53 and then promotes PCa cell apoptosis associated with the activation of DR4.

  11. Efficiency of lipofection of adherent cells is limited by apoptosis.

    Science.gov (United States)

    Bednarek, I; Czajka, M; Wilczok, T

    2002-01-01

    Stability of gene expression and transfection efficiency plays the main role in the application of gene transfer method. In somatic cell gene delivery, expression of the gene product is limited by the function of the cell to which it is delivered. In the present study analyzing the lipofected adherent cells, we have shown that lower level of transgene: beta-galactosidase activity at later time period correlated with decrease in cell viability, which was shown to be due to apoptosis. Apoptosis following DNA uptake occurred only when DNA was present during lipofection.

  12. Transglutaminase induction by various cell death and apoptosis pathways.

    Science.gov (United States)

    Fesus, L; Madi, A; Balajthy, Z; Nemes, Z; Szondy, Z

    1996-10-31

    Clarification of the molecular details of forms of natural cell death, including apoptosis, has become one of the most challenging issues of contemporary biomedical sciences. One of the effector elements of various cell death pathways is the covalent cross-linking of cellular proteins by transglutaminases. This review will discuss the accumulating data related to the induction and regulation of these enzymes, particularly of tissue type transglutaminase, in the molecular program of cell death. A wide range of signalling pathways can lead to the parallel induction of apoptosis and transglutaminase, providing a handle for better understanding the exact molecular interactions responsible for the mechanism of regulated cell death.

  13. Physiology and pathophysiology of apoptosis in epithelial cells of the liver, pancreas, and intestine.

    Science.gov (United States)

    Jones, B A; Gores, G J

    1997-12-01

    Cell death of gastrointestinal epithelial cells occurs by a process referred to as apoptosis. In this review, we succinctly define apoptosis and summarize the role of apoptosis in the physiology and pathophysiology of epithelial cells in the liver, pancreas, and small and large intestine. The physiological mediators regulating apoptosis in gastrointestinal epithelial cells, when known, are discussed. Selected pathophysiological consequences of excessive apoptosis and inhibition of apoptosis are used to illustrate the significance of apoptosis in disease processes. These examples demonstrate that excessive apoptosis may result in epithelial cell atrophy, injury, and dysfunction, whereas inhibition of apoptosis results in hyperplasia and promotes malignant transformation. The specific cellular mechanisms responsible for dysregulation of epithelial cell apoptosis during pathophysiological disturbances are emphasized. Potential future areas of physiological research regarding apoptosis in gastrointestinal epithelia are highlighted when appropriate.

  14. Tumor Response to Radiotherapy Regulated by Endothelial Cell Apoptosis

    Science.gov (United States)

    Garcia-Barros, Monica; Paris, Francois; Cordon-Cardo, Carlos; Lyden, David; Rafii, Shahin; Haimovitz-Friedman, Adriana; Fuks, Zvi; Kolesnick, Richard

    2003-05-01

    About 50% of cancer patients receive radiation therapy. Here we investigated the hypothesis that tumor response to radiation is determined not only by tumor cell phenotype but also by microvascular sensitivity. MCA/129 fibrosarcomas and B16F1 melanomas grown in apoptosis-resistant acid sphingomyelinase (asmase)-deficient or Bax-deficient mice displayed markedly reduced baseline microvascular endothelial apoptosis and grew 200 to 400% faster than tumors on wild-type microvasculature. Thus, endothelial apoptosis is a homeostatic factor regulating angiogenesis-dependent tumor growth. Moreover, these tumors exhibited reduced endothelial apoptosis upon irradiation and, unlike tumors in wild-type mice, they were resistant to single-dose radiation up to 20 grays (Gy). These studies indicate that microvascular damage regulates tumor cell response to radiation at the clinically relevant dose range.

  15. Fisetin induces apoptosis through mitochondrial apoptosis pathway in human uveal melanoma cells.

    Science.gov (United States)

    Wang, Kai; Hu, Dan-Ning; Lin, Hui-Wen; Yang, Wei-En; Hsieh, Yi-Hsien; Chien, Hsiang-Wen; Yang, Shun-Fa

    2018-05-01

    Fisetin, a diatery flavonoid, been reported that possess anticancer effects in various cancers. The purpose of the study was to investigate the antitumor effects of fisetin in cultured uveal melanoma cell lines and compared with normal retinal pigment epithelial (RPE) cells. MTT assay was used for evaluating cytotoxic effects of fisetin. Flow cytometry study was used for the determination of apoptosis. JC-1 fluorescent reader was used to determine mitochondrial transmembrane potential changes. The results shown that fisetin dose-dependently decreased the cell viability of uveal melanoma cells but not influenced the cell viability of RPE cells. Apoptosis of uveal melanoma cells was induced by fisetin efficiently. Fisetin inhibited antiapoptotic Bcl-2 family proteins and damaged the mitochondrial transmembrane potential. The levels of proapoptotic Bcl-2 proteins, cytochrome c, and various caspase activities were increased by fisetin. In conclusion, fisetin induces apoptosis of uveal melanoma cells selectively and may be a promising agent to be explored for the treatment of uveal melanoma. © 2018 Wiley Periodicals, Inc.

  16. More sensitive correlation of afferent pupillary defect with ganglion cell complex

    Directory of Open Access Journals (Sweden)

    Eulogio Besada

    2018-04-01

    Full Text Available Purpose: This study investigated the correlation between the relative afferent pupillary defect (RAPD and retinal nerve fiber layer thickness (RNFLT in optic neuropathy. Methods: RAPD assessment was performed using a log unit neutral density filter bar. Spectral domain optical coherence tomography RTVue-100 (Optovue was used to examine the subjects. The optic nerve head pattern (ONH was subdivided and identified for the purpose of the study into circumpapillary RNFLT (cpRNFLT and peripheral circumpapillary RNFLT (pcpRNFLT. The cpRNFLT, pcpRNFLT and ganglion cell complex (GCC parameters were analyzed. Results: Eighteen females and twenty three males with asymmetric optic neuropathy and a RAPD participated. Thirty-three subjects had glaucoma and eight had optic neuropathy other than glaucoma. Significant correlations (p < 0.02 were obtained for the RAPD and the percentage difference loss of the GCC and RNFLT parameters. The grouped mean percentage difference loss for RNFLT was significantly different from that of the GCC (p < 0.001. At a 0.6 log unit RAPD, the average mean percentage difference loss was 23% for the CRNFLT, 15% for the GCC, 12% for the global loss volume percentage and 6% for the focal loss volume percentage (FLV%. Conclusions: Significant correlations between RNFLT loss for cpRNFLT, pcpRNFLT and GCC parameters with RAPD were observed. Approximately a 35% higher sensitivity was obtained using GCC compared to CRNFL parameters. The expected change in GCC average for every 0.3 log unit increment was approximately 8.49 μm. The FLV% corresponded more sensitively to a RAPD but appeared to be influenced by disease severity. Resumen: Objetivo: Este estudio investigó la correlación entre el defecto pupilar aferente relativo (DPAR y el grosor de la capa de fibras nerviosas de la retina (RNFLT en la neuropatía óptica. Métodos: La valoración del DPAR se realizó utilizando una barra de filtro de densidad neutra de unidades logar

  17. The influence of venous blood flow on the retinal ganglion cell complex in patients with primary open angle glaucoma

    Directory of Open Access Journals (Sweden)

    N. I. Kurysheva

    2014-07-01

    Full Text Available Purpose: To study the influence of venous blood flow on the ganglion cell complex (GCC in patients with preperimetric and perimetric open angle glaucoma.Methods: 74 patients were included in the research. 59 eyes and 62 eyes were diagnosed with preperimetric and perimetric open angle glaucoma respectively. The mean age was 56.5±10.5 years. 22 (12 female and 10 male healthy individuals constituted the control group. The ganglion cell complex and retinal nerve fibre layer were evaluated with the help of optical coherence tomography (RTVue-100 OCT, Optovue, Inc., Fremont, CA. Ocular blood flow was measured by Color Doppler Imaging (multifunctional VOLUSON 730 ProSystem. The statistical analysis included correlation between GCC and RNFL thickness in both glaucoma groups.Results: The results showed a statistically significant reduction of venous blood flow velocity in both glaucoma groups compared to the control group. No difference in venous blood flow parameters between two glaucoma groups was found, except resistance index, which was higher in perimetric group in comparison to preperimetric group. A correlation was also obtained between venous blood flow parameters and GCC and RNFL thickness in both glaucoma groups.Conclusion: Early GCC damage in glaucoma might occur due to venous blood flow reduction. This fact may be of great value in understanding glaucoma pathogenesis and search for novel treatment options.

  18. RdgB2 is required for dim-light input into intrinsically photosensitive retinal ganglion cells.

    Science.gov (United States)

    Walker, Marquis T; Rupp, Alan; Elsaesser, Rebecca; Güler, Ali D; Sheng, Wenlong; Weng, Shijun; Berson, David M; Hattar, Samer; Montell, Craig

    2015-10-15

    A subset of retinal ganglion cells is intrinsically photosensitive (ipRGCs) and contributes directly to the pupillary light reflex and circadian photoentrainment under bright-light conditions. ipRGCs are also indirectly activated by light through cellular circuits initiated in rods and cones. A mammalian homologue (RdgB2) of a phosphoinositide transfer/exchange protein that functions in Drosophila phototransduction is expressed in the retinal ganglion cell layer. This raised the possibility that RdgB2 might function in the intrinsic light response in ipRGCs, which depends on a cascade reminiscent of Drosophila phototransduction. Here we found that under high light intensities, RdgB2(-/-) mutant mice showed normal pupillary light responses and circadian photoentrainment. Consistent with this behavioral phenotype, the intrinsic light responses of ipRGCs in RdgB2(-/-) were indistinguishable from wild-type. In contrast, under low-light conditions, RdgB2(-/-) mutants displayed defects in both circadian photoentrainment and the pupillary light response. The RdgB2 protein was not expressed in ipRGCs but was in GABAergic amacrine cells, which provided inhibitory feedback onto bipolar cells. We propose that RdgB2 is required in a cellular circuit that transduces light input from rods to bipolar cells that are coupled to GABAergic amacrine cells and ultimately to ipRGCs, thereby enabling ipRGCs to respond to dim light. © 2015 Walker et al. This article is distributed by The American Society for Cell Biology under license from the author(s). Two months after publication it is available to the public under an Attribution–Noncommercial–Share Alike 3.0 Unported Creative Commons License (http://creativecommons.org/licenses/by-nc-sa/3.0).

  19. DNA repair synthesis in rat retinal ganglion cells treated with chemical carcinogens or ultraviolet light in vitro, with special reference to aging and repair level

    International Nuclear Information System (INIS)

    Ishikawa, T.; Takayama, S.; Kitagawa, T.

    1978-01-01

    A system in which the retinal tissues of noninbred Wistar rats were used in combination with autoradiography was developed for measurement of DNA repair synthesis in ganglion cells of the central nervous system. Retinal tissues in short-term organ culture were treated with various carcinogens plus tritiated thymidine ([methyl -3 H]dThd) or were irradiated with uv light and then treated with [methyl -3 H]dThd. Preliminary study with retinal tissues from rats at various ages revealed no age-associated changes in the levels of unscheduled DNA synthesis in ganglion cells

  20. Spironolactone induces apoptosis in human mononuclear cells. Association between apoptosis and cytokine suppression

    DEFF Research Database (Denmark)

    Mikkelsen, Martin; Sønder, S U; Nersting, J

    2006-01-01

    Spironolactone (SPIR) has been described to suppress accumulation of pro-inflammatory cytokines. Here, the suppression of TNF-alpha in lipopolysaccharide (LPS)-stimulated mononuclear cell cultures was confirmed. However, SPIR was also found to induce apoptosis, prompting the investigations...... of a possible association between the two effects: The apoptosis-inducing and the cytokine-suppressive effects of SPIR correlated with regard to the effective concentration range. Also, pre-incubation experiments demonstrated a temporal separation of the two effects of ... preceding apoptosis. An association between the two effects was also seen when testing several SPIR analogues. Contrary to TNF-alpha, the levels of IL-1beta increased in SPIR-treated cultures. However, the amount of IL-1beta in the supernatants depended upon the order of SPIR and LPS addition, as IL-1beta...

  1. The effects of canine bone marrow stromal cells on neuritogenesis from dorsal root ganglion neurons in vitro.

    Science.gov (United States)

    Kamishina, Hiroaki; Cheeseman, Jennifer A; Clemmons, Roger M

    2009-10-01

    The present in vitro study was designed to evaluate whether canine bone marrow stromal cells (BMSCs) promote neurite outgrowth from dorsal root ganglion (DRG) neurons. Bone marrow aspirates were collected from iliac crests of three young adult dogs. DRG neurons were cultured on BMSCs, fibroblasts, or laminin substrates. DRG neurons were also cultured in BMSC- or fibroblast-conditioned media. DRG neurons grown on BMSCs extended longer neurites and developed a much more elaborate conformation of branching neurites compared to those on fibroblasts or laminin. Quantitative analysis revealed that these effects were associated with the emergence of increased numbers of primary and branching neurites. The effect appears to be dependent upon cell-cell interactions rather than by elaboration of diffusible molecules. With more extensive investigations into the basic biology of canine BMSCs, their ability for promoting neurite outgrowth may be translated into a novel therapeutic strategy for dogs with a variety of neurological disorders.

  2. Asymmetry between ON and OFF α ganglion cells of mouse retina: integration of signal and noise from synaptic inputs.

    Science.gov (United States)

    Freed, Michael A

    2017-11-15

    Bipolar and amacrine cells presynaptic to the ON sustained α cell of mouse retina provide currents with a higher signal-to-noise power ratio (SNR) than those presynaptic to the OFF sustained α cell. Yet the ON cell loses proportionately more SNR from synaptic inputs to spike output than the OFF cell does. The higher SNR of ON bipolar cells at the beginning of the ON pathway compensates for losses incurred by the ON ganglion cell, and improves the processing of positive contrasts. ON and OFF pathways in the retina include functional pairs of neurons that, at first glance, appear to have symmetrically similar responses to brightening and darkening, respectively. Upon careful examination, however, functional pairs exhibit asymmetries in receptive field size and response kinetics. Until now, descriptions of how light-adapted retinal circuitry maintains a preponderance of signal over the noise have not distinguished between ON and OFF pathways. Here I present evidence of marked asymmetries between members of a functional pair of sustained α ganglion cells in the mouse retina. The ON cell exhibited a proportionately greater loss of signal-to-noise power ratio (SNR) from its presynaptic arrays to its postsynaptic currents. Thus the ON cell combines signal and noise from its presynaptic arrays of bipolar and amacrine cells less efficiently than the OFF cell does. Yet the inefficiency of the ON cell is compensated by its presynaptic arrays providing a higher SNR than the arrays presynaptic to the OFF cell, apparently to improve visual processing of positive contrasts. Dynamic clamp experiments were performed that introduced synaptic conductances into ON and OFF cells. When the amacrine-modulated conductance was removed, the ON cell's spike train exhibited an increase in SNR. The OFF cell, however, showed the opposite effect of removing amacrine input, which was a decrease in SNR. Thus ON and OFF cells have different modes of synaptic integration with direct effects on

  3. BIGH3 protein and macrophages in retinal endothelial cell apoptosis.

    Science.gov (United States)

    Mondragon, Albert A; Betts-Obregon, Brandi S; Moritz, Robert J; Parvathaneni, Kalpana; Navarro, Mary M; Kim, Hong Seok; Lee, Chi Fung; LeBaron, Richard G; Asmis, Reto; Tsin, Andrew T

    2015-01-01

    Diabetes is a pandemic disease with a higher occurrence in minority populations. The molecular mechanism to initiate diabetes-associated retinal angiogenesis remains largely unknown. We propose an inflammatory pathway of diabetic retinopathy in which macrophages in the diabetic eye provide TGFβ to retinal endothelial cells (REC) in the retinal microvasculature. In response to TGFβ, REC synthesize and secrete a pro-apoptotic BIGH3 (TGFβ-Induced Gene Human Clone 3) protein, which acts in an autocrine loop to induce REC apoptosis. Rhesus monkey retinal endothelial cells (RhREC) were treated with dMCM (cell media of macrophages treated with high glucose and LDL) and assayed for apoptosis (TUNEL), BIGH3 mRNA (qPCR), and protein (Western blots) expressions. Cells were also treated with ΤGFβ1 and 2 for BIGH3 mRNA and protein expression. Inhibition assays were carried out using antibodies for TGFβ1 and for BIGH3 to block apoptosis and mRNA expression. BIGH3 in cultured RhREC cells were identified by immunohistochemistry (IHC). Distribution of BIGH3 and macrophages in the diabetic mouse retina was examined with IHC. RhRECs treated with dMCM or TGFβ showed a significant increase in apoptosis and BIGH3 protein expression. Recombinant BIGH3 added to RhREC culture medium led to a dose-dependent increase in apoptosis. Antibodies (Ab) directed against BIGH3 and TGFβ, as well as TGFβ receptor blocker resulted in a significant reduction in apoptosis induced by either dMCM, TGFβ or BIGH3. IHC showed that cultured RhREC constitutively expressed BIGH3. Macrophage and BIGH3 protein were co-localized to the inner retina of the diabetic mouse eye. Our results support a novel inflammatory pathway for diabetic retinopathy. This pathway is initiated by TGFβ released from macrophages, which promotes synthesis and release of BIGH3 protein by REC and REC apoptosis.

  4. Effect of Rosmarinic acid on sertoli cells apoptosis and serum ...

    African Journals Online (AJOL)

    inflammatory and antimicrobial activities and help to prevent cell damage caused by free radicals. The objective was to study the effect of Rosmarinic acid on sertolli cells apoptosis and serum antioxidant levels in rats after they were exposed to ...

  5. Chloroquinone Inhibits Cell Proliferation and Induces Apoptosis in ...

    African Journals Online (AJOL)

    Purpose: To demonstrate the role of chloroquinone (CQ) in inducing apoptosis in HONE-1 and HNE-1, nasopharyngeal carcinoma (NPC) cell lines. Methods: Water-soluble tetrazolium salt (WST)-1 assay was used for the determination of cell proliferation while an inverted microscope was employed for the analysis of ...

  6. Vaccination with apoptosis colorectal cancer cell pulsed autologous ...

    African Journals Online (AJOL)

    To investigate vaccination with apoptosis colorectal cancer (CRC) cell pulsed autologous dendritic cells (DCs) in advanced CRC, 14 patients with advanced colorectal cancer (CRC) were enrolled and treated with DCs vaccine to assess toxicity, tolerability, immune and clinical responses to the vaccine. No severe toxicity ...

  7. The single-cell gel electrophoresis assay to determine apoptosis ...

    African Journals Online (AJOL)

    When the frequency of appearance of apoptotic cells following was observed over a period of time, there was a significant increase in appearance of apoptosis when using single cell gel electrophoresis assay. The present report demonstrates that the characteristic pattern of apoptotic comets detected by the comet assay ...

  8. Retinal ganglion cell-inner plexiform and nerve fiber layers in neuromyelitis optica

    Directory of Open Access Journals (Sweden)

    Sai-Jing Hu

    2018-01-01

    Full Text Available AIM: To determine the thickness of the retinal ganglion cell-inner plexiform layer (GCIPL and the retinal nerve fiber layer (RNFL in patients with neuromyelitis optica (NMO. METHODS: We conducted a cross-sectional study that included 30 NMO patients with a total of 60 eyes. Based on the presence or absence of optic neuritis (ON, subjects were divided into either the NMO-ON group (30 eyes or the NMO-ON contra group (10 eyes. A detailed ophthalmologic examination was performed for each group; subsequently, the GCIPL and the RNFL were measured using high-definition optical coherence tomography (OCT. RESULTS: In the NMO-ON group, the mean GCIPL thickness was 69.28±21.12 μm, the minimum GCIPL thickness was 66.02±10.02 μm, and the RNFL thickness were 109.33±11.23, 110.47±3.10, 64.92±12.71 and 71.21±50.22 μm in the superior, inferior, temporal and nasal quadrants, respectively. In the NMO-ON contra group, the mean GCIPL thickness was 85.12±17.09 μm, the minimum GCIPL thickness was 25.39±25.1 μm, and the RNFL thicknesses were 148.33±23.22, 126.36±23.45, 82.21±22.30 and 83.36±31.28 μm in the superior, inferior, temporal and nasal quadrants, respectively. In the control group, the mean GCIPL thickness was 86.98±22.37 μm, the minimum GCIPL thickness was 85.28±10.75 μm, and the RNFL thicknesses were 150.22±22.73, 154.79±60.23, 82.33±7.01 and 85.62±13.81 μm in the superior, inferior, temporal and nasal quadrants, respectively. The GCIPL and RNFL were thinner in the NMO-ON contra group than in the control group (P<0.05; additionally, the RNFL was thinner in the inferior quadrant in the NMO-ON group than in the control group (P<0.05. Significant correlations were observed between the GCIPL and RNFL thickness measurements as well as between thickness measurements and the two visual field parameters of mean deviation (MD and corrected pattern standard deviation (PSD in the NMO-ON group (P<0.05. CONCLUSION: The thickness of the GCIPL

  9. Molecular Cell Biology of Apoptosis and Necroptosis in Cancer.

    Science.gov (United States)

    Dillon, Christopher P; Green, Douglas R

    Cell death is a major mechanism to eliminate cells in which DNA is damaged, organelles are stressed, or oncogenes are overexpressed, all events that would otherwise predispose cells to oncogenic transformation. The pathways that initiate and execute cell death are complex, genetically encoded, and subject to significant regulation. Consequently, while these pathways are often mutated in malignancy, there is considerable interest in inducing cell death in tumor cells as therapy. This chapter addresses our current understanding of molecular mechanisms contributing to two cell death pathways, apoptotic cell death and necroptosis, a regulated form of necrotic cell death. Apoptosis can be induced by a wide variety of signals, leading to protease activation that dismantles the cell. We discuss the physiological importance of each apoptosis pathway and summarize their known roles in cancer suppression and the current efforts at targeting each pathway therapeutically. The intricate mechanistic link between death receptor-mediated apoptosis and necroptosis is described, as well as the potential opportunities for utilizing necroptosis in the treatment of malignancy.

  10. Apoptosis and signalling in acid sphingomyelinase deficient cells

    Directory of Open Access Journals (Sweden)

    Sillence Dan J

    2001-11-01

    Full Text Available Abstract Background Recent evidence suggests that the activation of a non-specific lipid scramblase during apoptosis induces the flipping of sphingomyelin from the cell surface to the cytoplasmic leaftet of the plasma membrane. Inner leaflet sphingomyelin is then cleaved to ceramide by a neutral sphingomyelinase. The production of this non-membrane forming lipid induces blebbing of the plasma membrane to aid rapid engulfment by professional phagocytes. However contrary evidence suggests that cells which are deficient in acid sphingomyelinase are defective in apoptosis signalling. This data has been interpreted as support for the activation of acid sphingomyelinase as an early signal in apoptosis. Hypothesis An alternative explanation is put forward whereby the accumulation of intracellular sphingomyelin in sphingomyelinase deficient cells leads to the formation of intracellular rafts which lead to the sequestration of important signalling molecules that are normally present on the cell surface where they perform their function. Testing the hypothesis It is expected that the subcellular distribution of important signalling molecules is altered in acid sphingomyelinase deficient cells, leading to their sequestration in late endosomes / lysosomes. Other sphingolipid storage diseases such as Niemann-Pick type C which have normal acid sphingomyelinase activity would also be expected to show the same phenotype. Implications of the hypothesis If true the hypothesis would provide a mechanism for the pathology of the sphingolipid storage diseases at the cellular level and also have implications for the role of ceramide in apoptosis.

  11. Role of endoplasmic reticulum stress in the loss of retinal ganglion cells in diabetic retinopathy

    Institute of Scientific and Technical Information of China (English)

    Liping Yang; Lemeng Wu; Dongmei Wang; Ying Li; Hongliang Dou; Mark OMTso; Zhizhong Ma

    2013-01-01

    Endoplasmic reticulum stress is closely involved in the early stage of diabetic retinopathy. In the present study, a streptozotocin-induced diabetic animal model was given an intraperitoneal injection of tauroursodeoxycholic acid. Results from immunofluorescent co-localization experiments showed that both caspase-12 protein and c-Jun N-terminal kinase 1 phosphorylation levels significantly in-creased, which was associated with retinal ganglion celldeath in diabetic retinas. The C/ERB ho-mologous protein pathway directly contributed to glial reactivity, and was subsequently responsible for neuronal loss and vascular abnormalities in diabetic retinopathy. Our experimental findings in-dicate that endoplasmic reticulum stress plays an important role in diabetes-induced retinal neu-ronal loss and vascular abnormalities, and that inhibiting the activation of the endoplasmic reticulum stress pathway provides effective protection against diabetic retinopathy.

  12. Cuprous oxide nanoparticles selectively induce apoptosis of tumor cells

    Science.gov (United States)

    Wang, Ye; Zi, Xiao-Yuan; Su, Juan; Zhang, Hong-Xia; Zhang, Xin-Rong; Zhu, Hai-Ying; Li, Jian-Xiu; Yin, Meng; Yang, Feng; Hu, Yi-Ping

    2012-01-01

    In the rapid development of nanoscience and nanotechnology, many researchers have discovered that metal oxide nanoparticles have very useful pharmacological effects. Cuprous oxide nanoparticles (CONPs) can selectively induce apoptosis and suppress the proliferation of tumor cells, showing great potential as a clinical cancer therapy. Treatment with CONPs caused a G1/G0 cell cycle arrest in tumor cells. Furthermore, CONPs enclosed in vesicles entered, or were taken up by mitochondria, which damaged their membranes, thereby inducing apoptosis. CONPs can also produce reactive oxygen species (ROS) and initiate lipid peroxidation of the liposomal membrane, thereby regulating many signaling pathways and influencing the vital movements of cells. Our results demonstrate that CONPs have selective cytotoxicity towards tumor cells, and indicate that CONPs might be a potential nanomedicine for cancer therapy. PMID:22679374

  13. Effect of bFGF on radiation-induced apoptosis of vascular endothelial cells

    International Nuclear Information System (INIS)

    Gu Qingyang; Wang Dewen; Li Yuejuan; Peng Ruiyun; Dong Bo; Wang Zhaohai; Liu Jie; Deng Hua; Jiang Tao

    2003-01-01

    Objective: To study the effect of bFGF on radiation-induced apoptosis vascular endothelial cells. Methods: A cell line PAE (porcine aortic endothelial cells) and primary cultured HUVEC (human umbilical vein endothelial cells) were irradiated with 60 Co γ-rays to establish cell apoptosis models. Flow cytometry with annexin-V-FITC + PI labeling was used to evaluate cell apoptosis. Different amounts of bFGF were used to study their effects on radiation-induced endothelial cell apoptosis. Results and Conclusions: It is found that bFGF could inhibit radiation-induced endothelial cell apoptosis in a considerable degree

  14. Prevalence and Distribution of Segmentation Errors in Macular Ganglion Cell Analysis of Healthy Eyes Using Cirrus HD-OCT.

    Directory of Open Access Journals (Sweden)

    Rayan A Alshareef

    Full Text Available To determine the frequency of different types of spectral domain optical coherence tomography (SD-OCT scan artifacts and errors in ganglion cell algorithm (GCA in healthy eyes.Infrared image, color-coded map and each of the 128 horizontal b-scans acquired in the macular ganglion cell-inner plexiform layer scans using the Cirrus HD-OCT (Carl Zeiss Meditec, Dublin, CA macular cube 512 × 128 protocol in 30 healthy normal eyes were evaluated. The frequency and pattern of each artifact was determined. Deviation of the segmentation line was classified into mild (less than 10 microns, moderate (10-50 microns and severe (more than 50 microns. Each deviation, if present, was noted as upward or downward deviation. Each artifact was further described as per location on the scan and zones in the total scan area.A total of 1029 (26.8% out of total 3840 scans had scan errors. The most common scan error was segmentation error (100%, followed by degraded images (6.70%, blink artifacts (0.09% and out of register artifacts (3.3%. Misidentification of the inner retinal layers was most frequent (62%. Upward Deviation of the segmentation line (47.91% and severe deviation (40.3% were more often noted. Artifacts were mostly located in the central scan area (16.8%. The average number of scans with artifacts per eye was 34.3% and was not related to signal strength on Spearman correlation (p = 0.36.This study reveals that image artifacts and scan errors in SD-OCT GCA analysis are common and frequently involve segmentation errors. These errors may affect inner retinal thickness measurements in a clinically significant manner. Careful review of scans for artifacts is important when using this feature of SD-OCT device.

  15. Moringa oleifera: An apoptosis inducer in cancer cells

    African Journals Online (AJOL)

    Advanced Medical and Dental Institute, Universiti Sains Malaysia, Bertam, 13200 ... ability of M. oleifera to trigger apoptosis in cancer cells largely depends on its ... These compounds act by activating pro-apoptotic protein such as caspases, .... are safe for drinking [37]. .... the authors named in this article and all liabilities.

  16. Apoptosis induction of epifriedelinol on human cervical cancer cell line

    African Journals Online (AJOL)

    Background: Present investigation evaluates the antitumor activity of epifriedelinol for the management of cervical cancer by inducing process of apoptosis. Methods: Human Cervical Cancer Cell Line, C33A and HeLa were selected for study and treated with epifriedelinol at a concentration of (50-1000 μg/ml). Cytotoxicity of ...

  17. Chloroquinone Inhibits Cell Proliferation and Induces Apoptosis in ...

    African Journals Online (AJOL)

    Apoptosis in Nasopharyngeal Carcinoma Cell Lines. Xin-Qing ... However, this requires clinical investigation to ascertain its ... results to restrict the growth of tumors locally in addition ..... MEK1/ERK1/2/iNOS/sGC/PKG pathway associated with.

  18. What history tells us XXI. Apoptosis and programmed cell death

    Indian Academy of Sciences (India)

    2010-04-30

    Apr 30, 2010 ... Home; Journals; Journal of Biosciences; Volume 35; Issue 2. What history tells us XXI. Apoptosis and programmed cell death: when biological categories are blurred. Michel Morange. Series Volume 35 Issue 2 June 2010 pp 177-181 ...

  19. Ulinastatin Reduces T Cell Apoptosis in Rats with Severe Acute ...

    African Journals Online (AJOL)

    T cell apoptosis was determined by Annexin-V/PI double-staining. Oxidative stress was evaluated by examining changes in the levels of reactive oxygen species (ROS). Total superoxide dismutase (SOD) in serum was tested by hydroxylamine colorimetric assay, and malondialdehyde levels were examined by thiobarbituric ...

  20. Succinobucol induces apoptosis in vascular smooth muscle cells

    Czech Academy of Sciences Publication Activity Database

    Midwinter, R.G.; Maghzal, G.; Dennis, J.M.; Wu, B.J.; Cai, H.; Kapralov, A.A.; Belikova, N.A.; Tyurina, Y.Y.; Dong, L. F.; Khachigian, L.; Neužil, Jiří; Kagan, V.E.; Stocker, R.

    2012-01-01

    Roč. 52, č. 5 (2012), s. 871-879 ISSN 0891-5849 R&D Projects: GA ČR(CZ) GAP301/10/1937 Institutional research plan: CEZ:AV0Z50520701 Keywords : reactive oxygen species * free radicals * apoptosis Subject RIV: EA - Cell Biology Impact factor: 5.271, year: 2012

  1. Effect of quercetin on apoptosis of PANC-1 cells.

    Science.gov (United States)

    Lee, Joo Hyun; Lee, Han-Beom; Jung, Gum O; Oh, Jung Taek; Park, Dong Eun; Chae, Kwon Mook

    2013-12-01

    To investigate the chemotherapeutic effect of quercetin against cancer cells, signaling pathway of apoptosis was explored in human pancreatic cells. Various anticancer drugs including adriamycin, cisplatin, 5-fluorouracil (5-FU) and gemcitabine were used. Cell viability was measured by 3-[4,5-dimethylthiazol-2-yl]-2,5-diphe-nyltetra zolium bromide assay. Apoptosis was determined by 4'-6-diamidino-2-phenylindole nuclei staining and flow cytometry in PANC-1 cells treated with 50 µg/mL quercetin for 24 hours. Expression of endoplas mic reticulum (ER) stress mediators including, Grp78/Bip, p-PERK, PERK, ATF4, ATF6 and GADD153/CHOP proteins were measured by Western blot analysis. Mitochondrial membrane potential was measured by fluorescence staining with JC-1, rhodamine 123. Quercetin induced the apoptosis of PANC-1, which was characterized as nucleic acid and genomic DNA fragmentation, chromatin condensation, and sub-G0/G1 fraction of cell cycle increase. But not adriamycin, cisplatin, gemcitabine, and 5-FU. PANC-1 cells were markedly sensitive to quercetin. Treatment with quercetin resulted in the increased accumulation of intracellular Ca(2+) ion. Treatment with quercetin also increased the expression of Grp78/Bip and GADD153/CHOP protein and induced mitochondrial dysfunction. Quercetin exerted cytotoxicity against human pancreatic cancer cells via ER stress-mediated apoptotic signaling including reactive oxygen species production and mitochondrial dysfunction. These data suggest that quercetin may be an important modulator of chemosensitivity of cancer cells against anticancer chemotherapeutic agents.

  2. Study of the neuroprotective effects and mechanisms of Tianma Gouteng Decoction on retinal ganglion cells in rat optic nerve crush model

    Directory of Open Access Journals (Sweden)

    Fan-Tao Lyu

    2018-01-01

    Full Text Available AIM: To observe the mechanism of Tianma Gouteng Decoction on the protein molecular level in the optic nerve crush model rats. METHODS: Totally 36 participants 36 male Wistar rats were divided randomly into six groups(6 in every group: normal control group, negative control group, Tianma Gouteng Decoction treatment groups(con-centrations were 0.6g/mL, 1.2g/mL, 2.4g/mL respictivelyand ginkgo biloba tablets positive control group(concentrations was 1.2mg/mL. Nothing was done in the normal control group. The optic nerve of right eye in the other groups was done with the optic nerve crush model. Normal control group and negative control group was treated only with water. The average grey scale values of the N-methyl-D-aspartic acid receptor 2B(NMDA2Breceptor protein, beta - amyloid protein(Aβin the average grey scale values were detected. RESULTS: The average grey scale value of Tianma Gouteng Decoction in low, medium and high dose groups about NMDA2B receptor protein was significantly less than that of the negative control group(all PP=0.092, 0.411, 0.676, the difference between normal control group and negative control group was significant(PP=0.030, 0.001. The low dose group than the negative control group was not obviously(P=0.614. The high dose group was not significantly different from the positive control group(P=0.927, the difference between normal control group and negative control group was significant(PCONCLUSION: Tianma Gouteng Decoction can go through the decrease of the NMDA2B receptor protein expression and the control of beta-amyloid deposition to reduce the retinal ganglion cell injury and apoptosis.

  3. SIRT6 knockout cells resist apoptosis initiation but not progression: a computational method to evaluate the progression of apoptosis.

    Science.gov (United States)

    Domanskyi, Sergii; Nicholatos, Justin W; Schilling, Joshua E; Privman, Vladimir; Libert, Sergiy

    2017-11-01

    Apoptosis is essential for numerous processes, such as development, resistance to infections, and suppression of tumorigenesis. Here, we investigate the influence of the nutrient sensing and longevity-assuring enzyme SIRT6 on the dynamics of apoptosis triggered by serum starvation. Specifically, we characterize the progression of apoptosis in wild type and SIRT6 deficient mouse embryonic fibroblasts using time-lapse flow cytometry and computational modelling based on rate-equations and cell distribution analysis. We find that SIRT6 deficient cells resist apoptosis by delaying its initiation. Interestingly, once apoptosis is initiated, the rate of its progression is higher in SIRT6 null cells compared to identically cultured wild type cells. However, SIRT6 null cells succumb to apoptosis more slowly, not only in response to nutrient deprivation but also in response to other stresses. Our data suggest that SIRT6 plays a role in several distinct steps of apoptosis. Overall, we demonstrate the utility of our computational model to describe stages of apoptosis progression and the integrity of the cellular membrane. Such measurements will be useful in a broad range of biological applications.

  4. Ouabain enhances ADPKD cell apoptosis via the intrinsic pathway

    Directory of Open Access Journals (Sweden)

    Gustavo eBlanco

    2016-03-01

    Full Text Available Progression of autosomal dominant polycystic kidney disease (ADPKD is highly influenced by factors circulating in blood. We have shown that the hormone ouabain enhances several characteristics of the ADPKD cystic phenotype, including the rate of cell proliferation, fluid secretion and the capacity of the cells to form cysts. In this work, we found that physiological levels of ouabain (3nM also promote programmed cell death of renal epithelial cells obtained from kidney cysts of patients with ADPKD (ADPKD cells. This was determined by Alexa Fluor 488 labeled-Annexin-V staining and TUNEL assay, both biochemical markers of apoptosis. Ouabain-induced apoptosis also takes place when ADPKD cell growth is blocked; suggesting that the effect is not secondary to the stimulatory actions of ouabain on cell proliferation. Ouabain alters the expression of BCL family of proteins, reducing BCL-2 and increasing BAX expression levels, anti- and pro-apoptotic mediators respectively. In addition, ouabain caused the release of cytochrome c from mitochondria. Moreover, ouabain activates caspase-3, a key executioner caspase in the cell apoptotic pathway, but did not affect caspase-8. This suggests that ouabain triggers ADPKD cell apoptosis by stimulating the intrinsic, but not the extrinsic pathway of programmed cell death. The apoptotic effects of ouabain are specific for ADPKD cells and do not occur in normal human kidney cells (NHK cells. Taken together with our previous observations, these results show that ouabain causes an imbalance in cell growth/death, to favor growth of the cystic cells. This event, characteristic of ADPKD, further suggests the importance of ouabain as a circulating factor that promotes ADPKD progression.

  5. Activation of Neuropeptide Y Receptors Modulates Retinal Ganglion Cell Physiology and Exerts Neuroprotective Actions In Vitro

    DEFF Research Database (Denmark)

    Martins, João; Elvas, Filipe; Brudzewsky, Dan

    2015-01-01

    -reperfusion injury, pretreatment with NPY or (Leu31, Pro34)-NPY was not able to prevent apoptosis or rescue RGCs. In conclusion, we found modulatory effects of NPY application that for the first time were detected at the level of RGCs. However, further studies are needed to evaluate whether NPY neuroprotective...

  6. Induction of apoptosis by opium in some tumor cell lines.

    Science.gov (United States)

    Khaleghi, M; Farsinejad, A; Dabiri, S; Asadikaram, G

    2016-09-30

    The current study is aimed at investigation of the opium effects on the apoptosis of different cell lines in culture medium and compares such effects with one another. The study is carried out on over 8 cell lines (AA8, AGS, Hela, HepG2, MCF7, N2a, PC12, WEHI). A 2.86 x 10-4 g/ml opium concentration was prepared and added to the culture medium of the cell lines for 48 hours. Cytotoxicity was tested by 3-[4,5-dimethylthiazol-2-yl]-2,5-diphenyltetrazolium bromide (MTT) assay. The apoptotic effect of opium on the cell lines was analyzed by Annexin-PI test. Opium with concentration of 2.86 x 10-4 g/ml in 48 hours significantly induces apoptosis in certain cell lines (i.e. AA8, N2a, WEHI), apoptosis and necrosis in some others (i.e. Hela, HepG2, MCF7, and PC12), and also solely necrosis in the AGS cell line. One could infer that the usage of opium with different levels in different tissues leads to certain disorders in some tissues and may have therapeutic effects under distinctive conditions (i.e. unchecked growth of cells) as confirmed by the results.

  7. Radiation-induced apoptosis in F9 teratocarcinoma cells

    International Nuclear Information System (INIS)

    Langley, R.E.; Palayoor, S.T.; Coleman, C.N.; Bump, E.A.

    1994-01-01

    We have found that F9 murine teratocarcinoma cells undergo morphological changes and internucleosomal DNA fragmentation characteristic of apoptosis after exposure to ionizing radiation. We studied the time course, radiation dose-response, and the effects of protein and RNA synthesis inhibitors on this process. The response is dose dependent in the range 2-12 Gy. Internucleosomal DNA fragmentation can be detected as early as 6 h postirradiation and is maximal by 48 h. Cycloheximide, a protein synthesis inhibitor, and 5,6-dichloro-1-β-D-ribofuranosylbenzimidazole, an RNA synthesis inhibitor, both induced internucleosomal DNA fragmentation in the unirradiated cells and enhanced radiation-induced DNA fragmentation. F9 cells can be induced to differentiate into cells resembling endoderm with retinoic acid. After irradiation, differentiated F9 cells exhibit less DNA fragmentation than stem cells. This indicates that ionizing radiation can induce apoptosis in non-lymphoid tumours. We suggest that embryonic tumour cells may be particularly susceptible to agents that induce apoptosis. (Author)

  8. Radiation-induced apoptosis in F9 teratocarcinoma cells

    Energy Technology Data Exchange (ETDEWEB)

    Langley, R E; Palayoor, S T; Coleman, C N; Bump, E A [Joint Center for Radiation Therapy and Dana Farber Cancer Inst., Boston (United States)

    1994-05-01

    We have found that F9 murine teratocarcinoma cells undergo morphological changes and internucleosomal DNA fragmentation characteristic of apoptosis after exposure to ionizing radiation. We studied the time course, radiation dose-response, and the effects of protein and RNA synthesis inhibitors on this process. The response is dose dependent in the range 2-12 Gy. Internucleosomal DNA fragmentation can be detected as early as 6 h postirradiation and is maximal by 48 h. Cycloheximide, a protein synthesis inhibitor, and 5,6-dichloro-1-[beta]-D-ribofuranosylbenzimidazole, an RNA synthesis inhibitor, both induced internucleosomal DNA fragmentation in the unirradiated cells and enhanced radiation-induced DNA fragmentation. F9 cells can be induced to differentiate into cells resembling endoderm with retinoic acid. After irradiation, differentiated F9 cells exhibit less DNA fragmentation than stem cells. This indicates that ionizing radiation can induce apoptosis in non-lymphoid tumours. We suggest that embryonic tumour cells may be particularly susceptible to agents that induce apoptosis. (Author).

  9. Cuprous oxide nanoparticles selectively induce apoptosis of tumor cells

    Directory of Open Access Journals (Sweden)

    Wang Y

    2012-05-01

    Full Text Available Ye Wang,1,2,* Xiao-Yuan Zi,1,* Juan Su,1 Hong-Xia Zhang,1 Xin-Rong Zhang,3 Hai-Ying Zhu,1 Jian-Xiu Li,1 Meng Yin,3 Feng Yang,3 Yi-Ping Hu,11Department of Cell Biology, 2School of Clinical Medicine, 3Department of Pharmaceuticals, Second Military Medical University, Shanghai, People's Republic of China*Authors contributed equally.Abstract: In the rapid development of nanoscience and nanotechnology, many researchers have discovered that metal oxide nanoparticles have very useful pharmacological effects. Cuprous oxide nanoparticles (CONPs can selectively induce apoptosis and suppress the proliferation of tumor cells, showing great potential as a clinical cancer therapy. Treatment with CONPs caused a G1/G0 cell cycle arrest in tumor cells. Furthermore, CONPs enclosed in vesicles entered, or were taken up by mitochondria, which damaged their membranes, thereby inducing apoptosis. CONPs can also produce reactive oxygen species (ROS and initiate lipid peroxidation of the liposomal membrane, thereby regulating many signaling pathways and influencing the vital movements of cells. Our results demonstrate that CONPs have selective cytotoxicity towards tumor cells, and indicate that CONPs might be a potential nanomedicine for cancer therapy.Keywords: nanomedicine, selective cytotoxicity, apoptosis, cell cycle arrest, mitochondrion-targeted nanomaterials

  10. Renal cell apoptosis in human lupus nephritis: a histological study

    DEFF Research Database (Denmark)

    Faurschou, M; Penkowa, Milena; Andersen, C B

    2009-01-01

    Nuclear autoantigens from apoptotic cells are believed to drive the immunological response in systemic lupus erythematosus (SLE). Conflicting data exist as to the possible renal origin of apoptotic cells in SLE patients with nephritis. We assessed the level of renal cell apoptosis in kidney...... biopsies from 35 patients with lupus nephritis by means of terminal deoxynucleotidyl-transferase (TdT)-mediated deoxyuridine triphosphate (dUTP)-digoxigenin nick end labeling (TUNEL). Five samples of normal kidney tissue served as control specimens. We did not observe apoptotic glomerular cells in any...... cells constitute a quantitatively important source of auto-antibody-inducing nuclear auto-antigens in human lupus nephritis....

  11. Interferon-gamma (IFN-γ-mediated retinal ganglion cell death in human tyrosinase T cell receptor transgenic mouse.

    Directory of Open Access Journals (Sweden)

    Shahid Husain

    Full Text Available We have recently demonstrated the characterization of human tyrosinase TCR bearing h3T-A2 transgenic mouse model, which exhibits spontaneous autoimmune vitiligo and retinal dysfunction. The purpose of current study was to determine the role of T cells and IFN-γ in retina dysfunction and retinal ganglion cell (RGC death using this model. RGC function was measured by pattern electroretinograms (ERGs in response to contrast reversal of patterned visual stimuli. RGCs were visualized by fluorogold retrograde-labeling. Expression of CD3, IFN-γ, GFAP, and caspases was measured by immunohistochemistry and Western blotting. All functional and structural changes were measured in 12-month-old h3T-A2 mice and compared with age-matched HLA-A2 wild-type mice. Both pattern-ERGs (42%, p = 0.03 and RGC numbers (37%, p = 0.0001 were reduced in h3T-A2 mice when compared with wild-type mice. The level of CD3 expression was increased in h3T-A2 mice (h3T-A2: 174 ± 27% vs. HLA-A2: 100%; p = 0.04. The levels of effector cytokine IFN-γ were also increased significantly in h3T-A2 mice (h3T-A2: 189 ± 11% vs. HLA-A2: 100%; p = 0.023. Both CD3 and IFN-γ immunostaining were increased in nerve fiber (NF and RGC layers of h3T-A2 mice. In addition, we have seen a robust increase in GFAP staining in h3T-A2 mice (mainly localized to NF layer, which was substantially reduced in IFN-γ ((-/- knockout h3T-A2 mice. We also have seen an up-regulation of caspase-3 and -9 in h3T-A2 mice. Based on our data we conclude that h3T-A2 transgenic mice exhibit visual defects that are mostly associated with the inner retinal layers and RGC function. This novel h3T-A2 transgenic mouse model provides opportunity to understand RGC pathology and test neuroprotective strategies to rescue RGCs.

  12. Staurosporine induces ganglion cell differentiation in part by stimulating urokinase-type plasminogen activator expression and activation in the developing chick retina

    International Nuclear Information System (INIS)

    Kim, Yeoun-Hee; Chang, Yongmin; Jung, Jae-Chang

    2012-01-01

    Highlights: ► Staurosporine mediates stimulation of RGC differentiation in vitro cultured retinal neuroblasts. ► Staurosporine mediates uPA activation during RGC differentiation in vitro. ► Inhibition of uPA blocks the staurosporine mediated RGC differentiation both in vitro and in ovo. ► Thus, uPA may play a role in the staurosporine-mediated stimulation of RGC differentiation. -- Abstract: Here, we investigated whether staurosporine-mediated urokinase-type plasminogen activator (uPA) activation is involved in retinal ganglion cell (RGC) differentiation. Retinal cells were isolated from developing chick retinas at embryonic day 6 (E6). Relatively few control cells grown in serum-free medium started to form processes by 12 h. In contrast, staurosporine-treated cells had processes within 3 h, and processes were evident at 8 h. Immunofluorescence staining showed that Tuj-1-positive cells with shorter neurites could be detected in control cultures at 18 h, whereas numerous Tuj-1 positive ganglion cells with longer neuritic extensions were seen in staurosporine-treated cultures. BrdU-positive proliferating cells were more numerous in control cultures than in staurosporine-treated cultures, and the BrdU staining was not detected in post-mitotic Tuj-1 positive ganglion cells. Western blotting of cell lysates showed that staurosporine induced high levels of the active form of uPA. The staurosporine-induced uPA signal was localized predominantly in the soma, neurites and axons of Tuj-1-positive ganglion cells. Amiloride, an inhibitor of uPA, markedly reduced staurosporine-induced Tuj-1 staining, neurite length, neurite number, and uPA staining versus controls. In developing retinas in ovo, amiloride administration remarkably reduced the staurosporine-induced uPA staining and RGC differentiation. Taken together, our in vitro and in vivo data collectively indicate that uPA plays a role in the staurosporine-mediated stimulation of RGC differentiation.

  13. Human retinal pigment epithelial cell-induced apoptosis in activated T cells

    DEFF Research Database (Denmark)

    Jørgensen, A; Wiencke, A K; la Cour, M

    1998-01-01

    PURPOSE: The immune privilege of the eye has been thought to be dependent on physical barriers and absence of lymphatic vessels. However, the immune privilege may also involve active immunologic processes, as recent studies have indicated. The purpose of the present study was to investigate whether...... human retinal pigment epithelial (RPE) cells can induce apoptosis in activated T cells. METHODS: Fas ligand (FasL) expression was detected by flow cytometry and immunohistochemistry. Cultured RPE cells were cocultured with T-cell lines and peripheral blood lymphocytes for 6 hours to 2 days. Induction...... of apoptosis was detected by 7-amino-actinomycin D and annexin V staining. RESULTS: Retinal pigment epithelial cells expressed FasL and induced apoptosis in activated Fas+ T cells. Blocking of Fas-FasL interaction with antibody strongly inhibited RPE-mediated T-cell apoptosis. Retinal pigment epithelial cells...

  14. Cytokines and Pancreatic β-Cell Apoptosis

    DEFF Research Database (Denmark)

    Berchtold, L A; Prause, M; Størling, J

    2016-01-01

    The discovery 30 years ago that inflammatory cytokines cause a concentration, activity, and time-dependent bimodal response in pancreatic β-cell function and viability has been a game-changer in the fields of research directed at understanding inflammatory regulation of β-cell function and survival...... and the causes of β-cell failure and destruction in diabetes. Having until then been confined to the use of pathophysiologically irrelevant β-cell toxic chemicals as a model of β-cell death, researchers could now mimic endocrine and paracrine effects of the cytokine response in vitro by titrating concentrations...... of local, chronic islet inflammation. Since then, numerous studies have clarified how these bimodal responses depend on discrete signaling pathways. Most interest has been devoted to the proapoptotic response dependent upon mainly nuclear factor κ B and mitogen-activated protein kinase activation, leading...

  15. Sun Ginseng Protects Endothelial Progenitor Cells From Senescence Associated Apoptosis

    Science.gov (United States)

    Im, Wooseok; Chung, Jin-Young; Bhan, Jaejun; Lim, Jiyeon; Lee, Soon-Tae; Chu, Kon; Kim, Manho

    2012-01-01

    Endothelial progenitor cells (EPC) are a population of cells that circulate in the blood stream. They play a role in angiogenesis and, therefore, can be prognostic markers of vascular repair. Ginsenoside Rg3 prevents endothelial cell apoptosis through the inhibition of the mitochondrial caspase pathway. It also affects estrogen activity, which reduces EPC senescence. Sun ginseng (SG), which is heat-processed ginseng, has a high content of ginsenosides. The purpose of this study was to investigate the protective effects of SG on senescence-associated apoptosis in EPCs. In order to isolate EPCs, mononuclear cells of human blood buffy coats were cultured and characterized by their uptake of acetylated low-density lipoprotein (acLDL) and their binding of Ulex europaeus agglutinin I (ulex-lectin). Flow cytometry with annexin-V staining was performed in order to assess early and late apoptosis. Senescence was determined by β-galactosidase (β-gal) staining. Staining with 4′-6-Diamidino-2-phenylindole verified that most adherent cells (93±2.7%) were acLDL-positive and ulex-lectin-positive. The percentage of β-gal-positive EPCs was decreased from 93.8±2.0% to 62.5±3.6% by SG treatment. A fluorescence-activated cell sorter (FACS) analysis showed that 4.9% of EPCs were late apoptotic in controls. Sun ginseng decreased the apoptotic cell population by 39% in the late stage of apoptosis from control baseline levels. In conclusion, these results show antisenescent and antiapoptotic effects of SG in human-derived EPCs, indicating that SG can enhance EPC-mediated repair mechanisms. PMID:23717107

  16. Isolation and Molecular Profiling of Primary Mouse Retinal Ganglion Cells: Comparison of Phenotypes from Healthy and Glaucomatous Retinas.

    Science.gov (United States)

    Chintalapudi, Sumana R; Djenderedjian, Levon; Stiemke, Andrew B; Steinle, Jena J; Jablonski, Monica M; Morales-Tirado, Vanessa M

    2016-01-01

    Loss of functional retinal ganglion cells (RGC) is an element of retinal degeneration that is poorly understood. This is in part due to the lack of a reliable and validated protocol for the isolation of primary RGCs. Here we optimize a feasible, reproducible, standardized flow cytometry-based protocol for the isolation and enrichment of homogeneous RGC with the Thy1.2(hi)CD48(neg)CD15(neg)CD57(neg) surface phenotype. A three-step validation process was performed by: (1) genomic profiling of 25-genes associated with retinal cells; (2) intracellular labeling of homogeneous sorted cells for the intracellular RGC-markers SNCG, brain-specific homeobox/POU domain protein 3A (BRN3A), TUJ1, and RNA-binding protein with multiple splicing (RBPMS); and (3) by applying the methodology on RGC from a mouse model with elevated intraocular pressure (IOP) and optic nerve damage. Use of primary RGC cultures will allow for future careful assessment of important cell specific pathways in RGC to provide mechanistic insights into the declining of visual acuity in aged populations and those suffering from retinal neurodegenerative diseases.

  17. Isolation and Molecular Profiling of Primary Mouse Retinal Ganglion Cells: Comparison of Phenotypes from Healthy and Glaucomatous Retinas

    Science.gov (United States)

    Chintalapudi, Sumana R.; Djenderedjian, Levon; Stiemke, Andrew B.; Steinle, Jena J.; Jablonski, Monica M.; Morales-Tirado, Vanessa M.

    2016-01-01

    Loss of functional retinal ganglion cells (RGC) is an element of retinal degeneration that is poorly understood. This is in part due to the lack of a reliable and validated protocol for the isolation of primary RGCs. Here we optimize a feasible, reproducible, standardized flow cytometry-based protocol for the isolation and enrichment of homogeneous RGC with the Thy1.2hiCD48negCD15negCD57neg surface phenotype. A three-step validation process was performed by: (1) genomic profiling of 25-genes associated with retinal cells; (2) intracellular labeling of homogeneous sorted cells for the intracellular RGC-markers SNCG, brain-specific homeobox/POU domain protein 3A (BRN3A), TUJ1, and RNA-binding protein with multiple splicing (RBPMS); and (3) by applying the methodology on RGC from a mouse model with elevated intraocular pressure (IOP) and optic nerve damage. Use of primary RGC cultures will allow for future careful assessment of important cell specific pathways in RGC to provide mechanistic insights into the declining of visual acuity in aged populations and those suffering from retinal neurodegenerative diseases. PMID:27242509

  18. Quinuclidine compounds differently act as agonists of Kenyon cell nicotinic acetylcholine receptors and induced distinct effect on insect ganglionic depolarizations.

    Science.gov (United States)

    Mathé-Allainmat, Monique; Swale, Daniel; Leray, Xavier; Benzidane, Yassine; Lebreton, Jacques; Bloomquist, Jeffrey R; Thany, Steeve H

    2013-12-01

    We have recently demonstrated that a new quinuclidine benzamide compound named LMA10203 acted as an agonist of insect nicotinic acetylcholine receptors. Its specific pharmacological profile on cockroach dorsal unpaired median neurons (DUM) helped to identify alpha-bungarotoxin-insensitive nAChR2 receptors. In the present study, we tested its effect on cockroach Kenyon cells. We found that it induced an inward current demonstrating that it bounds to nicotinic acetylcholine receptors expressed on Kenyon cells. Interestingly, LMA10203-induced currents were completely blocked by the nicotinic antagonist α-bungarotoxin. We suggested that LMA10203 effect occurred through the activation of α-bungarotoxin-sensitive receptors and did not involve α-bungarotoxin-insensitive nAChR2, previously identified in DUM neurons. In addition, we have synthesized two new compounds, LMA10210 and LMA10211, and compared their effects on Kenyon cells. These compounds were members of the 3-quinuclidinyl benzamide or benzoate families. Interestingly, 1 mM LMA10210 was not able to induce an inward current on Kenyon cells compared to LMA10211. Similarly, we did not find any significant effect of LMA10210 on cockroach ganglionic depolarization, whereas these three compounds were able to induce an effect on the central nervous system of the third instar M. domestica larvae. Our data suggested that these three compounds could bind to distinct cockroach nicotinic acetylcholine receptors.

  19. Zinc oxide nanoparticles decrease the expression and activity of plasma membrane calcium ATPase, disrupt the intracellular calcium homeostasis in rat retinal ganglion cells.

    Science.gov (United States)

    Guo, Dadong; Bi, Hongsheng; Wang, Daoguang; Wu, Qiuxin

    2013-08-01

    Zinc oxide nanoparticle is one of the most important materials with diverse applications. However, it has been reported that zinc oxide nanoparticles are toxic to organisms, and that oxidative stress is often hypothesized to be an important factor in cytotoxicity mediated by zinc oxide nanoparticles. Nevertheless, the mechanism of toxicity of zinc oxide nanoparticles has not been completely understood. In this study, we investigated the cytotoxic effect of zinc oxide nanoparticles and the possible molecular mechanism involved in calcium homeostasis mediated by plasma membrane calcium ATPase in rat retinal ganglion cells. Real-time cell electronic sensing assay showed that zinc oxide nanoparticles could exert cytotoxic effect on rat retinal ganglion cells in a concentration-dependent manner; flow cytometric analysis indicated that zinc oxide nanoparticles could lead to cell damage by inducing the overproduction of reactive oxygen species. Furthermore, zinc oxide nanoparticles could also apparently decrease the expression level and their activity of plasma membrane calcium ATPase, which finally disrupt the intracellular calcium homeostasis and result in cell death. Taken together, zinc oxide nanoparticles could apparently decrease the plasma membrane calcium ATPase expression, inhibit their activity, cause the elevated intracellular calcium ion level and disrupt the intracellular calcium homeostasis. Further, the disrupted calcium homeostasis will trigger mitochondrial dysfunction, generate excessive reactive oxygen species, and finally initiate cell death. Thus, the disrupted calcium homeostasis is involved in the zinc oxide nanoparticle-induced rat retinal ganglion cell death. Copyright © 2013 Elsevier Ltd. All rights reserved.

  20. Gallic Acid Induces Apoptosis in Human Gastric Adenocarcinoma Cells.

    Science.gov (United States)

    Tsai, Chung-Lin; Chiu, Ying-Ming; Ho, Tin-Yun; Hsieh, Chin-Tung; Shieh, Dong-Chen; Lee, Yi-Ju; Tsay, Gregory J; Wu, Yi-Ying

    2018-04-01

    Gastric cancer is one of the most common malignant cancers with a poor prognosis and high mortality rate worldwide. Current treatment of gastric cancer includes surgery and chemotherapy as the main modalities, but the potentially severe side-effects of chemotherapy present a considerable challenge. Gallic acid is a trihydroxybenzoic acid found to exert an anticancer effect against a variety of cancer cells. The purpose of this study was to determine the anti-cancer activity of Galla chinensis and its main component gallic acid on human gastric adenocarcinoma cells. MTT assay and cell death ELISA were used to determine the apoptotic effect of Gallic Chinensis and gallic acid on human gastric adenocarcinoma cells. To determine the pathway and relevant components by which gallic acid-induced apoptosis is mediated through, cells were transfected with siRNA (Fas, FasL, DR5, p53) using Lipofectamine 2000. Reults: Gallic Chinensis and gallic acid induced apoptosis of human gastric adenocarcinoma cells. Gallic acid induced up-regulation of Fas, FasL, and DR5 expression in AGS cells. Transfection of cells with Fas, FasL, or DR5 siRNA reduced gallic acid-induced cell death. In addition, p53 was shown to be involved in gallic acid-mediated Fas, FasL, and DR5 expression as well as cell apoptosis in AGS cells. These results suggest that gallic acid has a potential role in the treatment of gastric cancer. Copyright© 2018, International Institute of Anticancer Research (Dr. George J. Delinasios), All rights reserved.

  1. Bystander apoptosis in human cells mediated by irradiated blood plasma

    Energy Technology Data Exchange (ETDEWEB)

    Vinnikov, Volodymyr, E-mail: vlad.vinnikov@mail.ru [Grigoriev Institute for Medical Radiology of the National Academy of Medical Science of Ukraine (Ukraine); Lloyd, David; Finnon, Paul [Centre for Radiation, Chemical and Environmental Hazards of the Health Protection Agency of the United Kingdom (United Kingdom)

    2012-03-01

    Following exposure to high doses of ionizing radiation, due to an accident or during radiotherapy, bystander signalling poses a potential hazard to unirradiated cells and tissues. This process can be mediated by factors circulating in blood plasma. Thus, we assessed the ability of plasma taken from in vitro irradiated human blood to produce a direct cytotoxic effect, by inducing apoptosis in primary human peripheral blood mononuclear cells (PBM), which mainly comprised G{sub 0}-stage lymphocytes. Plasma was collected from healthy donors' blood irradiated in vitro to 0-40 Gy acute {gamma}-rays. Reporter PBM were separated from unirradiated blood with Histopaque and held in medium with the test plasma for 24 h at 37 Degree-Sign C. Additionally, plasma from in vitro irradiated and unirradiated blood was tested against PBM collected from blood given 4 Gy. Apoptosis in reporter PBM was measured by the Annexin V test using flow cytometry. Plasma collected from unirradiated and irradiated blood did not produce any apoptotic response above the control level in unirradiated reporter PBM. Surprisingly, plasma from irradiated blood caused a dose-dependent reduction of apoptosis in irradiated reporter PBM. The yields of radiation-induced cell death in irradiated reporter PBM (after subtracting the respective values in unirradiated reporter PBM) were 22.2 {+-} 1.8% in plasma-free cultures, 21.6 {+-} 1.1% in cultures treated with plasma from unirradiated blood, 20.2 {+-} 1.4% in cultures with plasma from blood given 2-4 Gy and 16.7 {+-} 3.2% in cultures with plasma from blood given 6-10 Gy. These results suggested that irradiated blood plasma did not cause a radiation-induced bystander cell-killing effect. Instead, a reduction of apoptosis in irradiated reporter cells cultured with irradiated blood plasma has implications concerning oncogenic risk from mutated cells surviving after high dose in vivo irradiation (e.g. radiotherapy) and requires further study.

  2. Bystander apoptosis in human cells mediated by irradiated blood plasma

    International Nuclear Information System (INIS)

    Vinnikov, Volodymyr; Lloyd, David; Finnon, Paul

    2012-01-01

    Following exposure to high doses of ionizing radiation, due to an accident or during radiotherapy, bystander signalling poses a potential hazard to unirradiated cells and tissues. This process can be mediated by factors circulating in blood plasma. Thus, we assessed the ability of plasma taken from in vitro irradiated human blood to produce a direct cytotoxic effect, by inducing apoptosis in primary human peripheral blood mononuclear cells (PBM), which mainly comprised G 0 -stage lymphocytes. Plasma was collected from healthy donors’ blood irradiated in vitro to 0–40 Gy acute γ-rays. Reporter PBM were separated from unirradiated blood with Histopaque and held in medium with the test plasma for 24 h at 37 °C. Additionally, plasma from in vitro irradiated and unirradiated blood was tested against PBM collected from blood given 4 Gy. Apoptosis in reporter PBM was measured by the Annexin V test using flow cytometry. Plasma collected from unirradiated and irradiated blood did not produce any apoptotic response above the control level in unirradiated reporter PBM. Surprisingly, plasma from irradiated blood caused a dose-dependent reduction of apoptosis in irradiated reporter PBM. The yields of radiation-induced cell death in irradiated reporter PBM (after subtracting the respective values in unirradiated reporter PBM) were 22.2 ± 1.8% in plasma-free cultures, 21.6 ± 1.1% in cultures treated with plasma from unirradiated blood, 20.2 ± 1.4% in cultures with plasma from blood given 2–4 Gy and 16.7 ± 3.2% in cultures with plasma from blood given 6–10 Gy. These results suggested that irradiated blood plasma did not cause a radiation-induced bystander cell-killing effect. Instead, a reduction of apoptosis in irradiated reporter cells cultured with irradiated blood plasma has implications concerning oncogenic risk from mutated cells surviving after high dose in vivo irradiation (e.g. radiotherapy) and requires further study.

  3. Knockdown of HIF-1α and IL-8 induced apoptosis of hepatocellular carcinoma triggers apoptosis of vascular endothelial cells.

    Science.gov (United States)

    Choi, Sung Hoon; Park, Jun Yong; Kang, Wonseok; Kim, Seung Up; Kim, Do Young; Ahn, Sang Hoon; Ro, Simon Wonsang; Han, Kwang-Hyub

    2016-01-01

    A local hypoxic microenvironment is one of the most important characteristics of solid tumors. Hypoxia inducible factor-1α (HIF-1α) and Interleukin-8 (IL-8) activate tumor survival from hypoxic-induced apoptosis in each pathway. This study aimed to evaluate whether knockdown of HIF-1α and IL-8 induced apoptosis of the hepatocellular carcinoma (HCC) and endothelial cell lines. HCC cell lines were infected with adenovirus-expressing shRNA for HIF-1α and IL-8 and maintained under hypoxic conditions (1% O2, 24 h). The expression levels of HIF-1α and both apoptotic and growth factors were examined by real-time quantitative PCR and western blot. We also investigated apoptosis by TUNEL assay (FACS and Immunofluorescence) and measured the concentration of cytochrome C. Inhibition of HIF-1α and IL-8 up-regulated the expression of apoptotic factors while downregulating anti-apoptotic factors simultaneously. Knockdown of HIF-1α and IL-8 increased the concentration of cytochrome C and enhanced DNA fragmentation in HCC cell lines. Moreover, culture supernatant collected from the knockdown of HIF-1α and IL-8 in HCC cell lines induced apoptosis in human umbilical vein endothelial cells under hypoxia, and the expression of variable apoptotic ligand increased from HCC cell lines, time-dependently. These data suggest that adenovirus-mediated knockdown of HIF-1α and IL-8 induced apoptosis in HCC cells and triggered apoptosis of vascular endothelial cells.

  4. Selective deletion of cochlear hair cells causes rapid age-dependent changes in spiral ganglion and cochlear nucleus neurons.

    Science.gov (United States)

    Tong, Ling; Strong, Melissa K; Kaur, Tejbeer; Juiz, Jose M; Oesterle, Elizabeth C; Hume, Clifford; Warchol, Mark E; Palmiter, Richard D; Rubel, Edwin W

    2015-05-20

    During nervous system development, critical periods are usually defined as early periods during which manipulations dramatically change neuronal structure or function, whereas the same manipulations in mature animals have little or no effect on the same property. Neurons in the ventral cochlear nucleus (CN) are dependent on excitatory afferent input for survival during a critical period of development. Cochlear removal in young mammals and birds results in rapid death of target neurons in the CN. Cochlear removal in older animals results in little or no neuron death. However, the extent to which hair-cell-specific afferent activity prevents neuronal death in the neonatal brain is unknown. We further explore this phenomenon using a new mouse model that allows temporal control of cochlear hair cell deletion. Hair cells express the human diphtheria toxin (DT) receptor behind the Pou4f3 promoter. Injections of DT resulted in nearly complete loss of organ of Corti hair cells within 1 week of injection regardless of the age of injection. Injection of DT did not influence surrounding supporting cells directly in the sensory epithelium or spiral ganglion neurons (SGNs). Loss of hair cells in neonates resulted in rapid and profound neuronal loss in the ventral CN, but not when hair cells were eliminated at a more mature age. In addition, normal survival of SGNs was dependent on hair cell integrity early in development and less so in mature animals. This defines a previously undocumented critical period for SGN survival. Copyright © 2015 the authors 0270-6474/15/357878-14$15.00/0.

  5. Establishment of a long-term spiral ganglion neuron culture with reduced glial cell number: Effects of AraC on cell composition and neurons.

    Science.gov (United States)

    Schwieger, Jana; Esser, Karl-Heinz; Lenarz, Thomas; Scheper, Verena

    2016-08-01

    Sensorineural deafness is mainly caused by damage to hair cells and degeneration of the spiral ganglion neurons (SGN). Cochlear implants can functionally replace lost hair cells and stimulate the SGN electrically. The benefit from cochlear implantation depends on the number and excitability of these neurons. To identify potential therapies for SGN protection, in vitro tests are carried out on spiral ganglion cells (SGC). A glial cell-reduced and neuron-enhanced culture of neonatal rat SGC under mitotic inhibition (cytarabine (AraC)) for up to seven days is presented. Serum containing and neurotrophin-enriched cultures with and without AraC-addition were analyzed after 4 and 7 days. The total number of cells was significantly reduced, while the proportion of neurons was greatly increased by AraC-treatment. Cell type-specific labeling demonstrated that nearly all fibroblasts and most of the glial cells were removed. Neither the neuronal survival, nor the neurite outgrowth or soma diameter were negatively affected. Additionally neurites remain partly free of surrounding non-neuronal cells. Recent culture conditions allow only for short-term cultivation of neonatal SGC and lack information on the influence of non-neuronal cells on SGN and of direct contact of neurites with test-materials. AraC-addition reduces the number of non-neuronal cells and increases the ratio of SGN in culture, without negative impact on neuronal viability. This treatment allows longer-term cultivation of SGC and provides deeper insight into SGN-glial cell interaction and the attachment of neurites on test-material surfaces. Copyright © 2016 The Authors. Published by Elsevier B.V. All rights reserved.

  6. Direct Reprogramming of Spiral Ganglion Non-neuronal Cells into Neurons: Toward Ameliorating Sensorineural Hearing Loss by Gene Therapy

    Directory of Open Access Journals (Sweden)

    Teppei Noda

    2018-02-01

    Full Text Available Primary auditory neurons (PANs play a critical role in hearing by transmitting sound information from the inner ear to the brain. Their progressive degeneration is associated with excessive noise, disease and aging. The loss of PANs leads to permanent hearing impairment since they are incapable of regenerating. Spiral ganglion non-neuronal cells (SGNNCs, comprised mainly of glia, are resident within the modiolus and continue to survive after PAN loss. These attributes make SGNNCs an excellent target for replacing damaged PANs through cellular reprogramming. We used the neurogenic pioneer transcription factor Ascl1 and the auditory neuron differentiation factor NeuroD1 to reprogram SGNNCs into induced neurons (iNs. The overexpression of both Ascl1 and NeuroD1 in vitro generated iNs at high efficiency. Transcriptome analyses revealed that iNs displayed a transcriptome profile resembling that of endogenous PANs, including expression of several key markers of neuronal identity: Tubb3, Map2, Prph, Snap25, and Prox1. Pathway analyses indicated that essential pathways in neuronal growth and maturation were activated in cells upon neuronal induction. Furthermore, iNs extended projections toward cochlear hair cells and cochlear nucleus neurons when cultured with each respective tissue. Taken together, our study demonstrates that PAN-like neurons can be generated from endogenous SGNNCs. This work suggests that gene therapy can be a viable strategy to treat sensorineural hearing loss caused by degeneration of PANs.

  7. A Cell Culture Model of Latent and Lytic Herpes Simplex Virus Type 1 Infection in Spiral Ganglion.

    Science.gov (United States)

    Liu, Yuehong; Li, Shufeng

    2015-01-01

    Reactivation of latent herpes simplex virus type 1 (HSV-1) in spiral ganglion neurons (SGNs) is supposed to be one of the causes of idiopathic sudden sensorineural hearing loss. This study aims to establish a cell culture model of latent and lytic HSV-1 infection in spiral ganglia. In the presence of acyclovir, primary cultures of SGNs were latently infected with HSV-1 expressing green fluorescent protein. Four days later, these cells were treated with trichostatin A (TSA), a known chemical reactivator of HSV-1. TCID50 was used to measure the titers of virus in cultures on Vero cells. RNA from cultures was detected for the presence of transcripts of ICP27 and latency-associated transcript (LAT) using reverse transcription polymerase chain reaction. There is no detectable infectious HSV-1 in latently infected cultures, whereas they could be observed in both lytically infected and latently infected/TSA-treated cultures. LAT was the only detectable transcript during latent infection, whereas lytic ICP27 transcript was detected in lytically infected and latently infected/TSA-treated cultures. Cultured SGNs can be both latently and lytically infected with HSV-1. Furthermore, latently infected SGNs can be reactivated using TSA, yielding infectious virus.

  8. Action potentials in retinal ganglion cells are initiated at the site of maximal curvature of the extracellular potential.

    Science.gov (United States)

    Eickenscheidt, Max; Zeck, Günther

    2014-06-01

    The initiation of an action potential by extracellular stimulation occurs after local depolarization of the neuronal membrane above threshold. Although the technique shows remarkable clinical success, the site of action and the relevant stimulation parameters are not completely understood. Here we identify the site of action potential initiation in rabbit retinal ganglion cells (RGCs) interfaced to an array of extracellular capacitive stimulation electrodes. We determine which feature of the extracellular potential governs action potential initiation by simultaneous stimulation and recording RGCs interfaced in epiretinal configuration. Stimulation electrodes were combined to areas of different size and were presented at different positions with respect to the RGC. Based on stimulation by electrodes beneath the RGC soma and simultaneous sub-millisecond latency measurement we infer axonal initiation at the site of maximal curvature of the extracellular potential. Stimulation by electrodes at different positions along the axon reveals a nearly constant threshold current density except for a narrow region close to the cell soma. These findings are explained by the concept of the activating function modified to consider a region of lower excitability close to the cell soma. We present a framework how to estimate the site of action potential initiation and the stimulus required to cross threshold in neurons tightly interfaced to capacitive stimulation electrodes. Our results underscore the necessity of rigorous electrical characterization of the stimulation electrodes and of the interfaced neural tissue.

  9. Occurance of apoptosis during ischemia in porcine pancreas islet cells.

    Science.gov (United States)

    Stadlbauer, V; Schaffellner, S; Iberer, F; Lackner, C; Liegl, B; Zink, B; Kniepeiss, D; Tscheliessnigg, K H

    2003-03-01

    Pancreas islet transplantation is a potential treatment of diabetes mellitus and porcine organs provide an easily available source of cells. Unfortunately quality and quantity of isolated islets are still not satisfactory. Apoptosis occurs in freshly isolated islets and plays a significant role in early graft loss. We evaluated the influence of four storage solutions on porcine pancreas islets. After warm ischemia of 15-20 minutes 12 organs were stored in 4 cold preservation solutions: Histidine-Tryptophan-Ketoglutarate solution (HTK), Hank's buffered saline solution (HBSS), University of Wisconsin (UW) solution and Ringer-Lactate (R). After cold ischemia for 100 minutes, organs were fixed in 3% formalin. Apoptotic cells were counted on hematocylin-eosin stainings. Most apoptotic cells were found in organs stored in R. Low numbers were found in the other groups. The difference between organs stored in R and organs stored in UW, HTK, or HBSS was highly significant. No significant difference could be found between UW, HTK and HBSS. Cold and warm ischemia of the pancreas seems to induce apoptosis in islet cells. Preservation solutions cause less apoptosis than electrolyte solution. No significant differences could be found among the preservation solutions.

  10. Effects of electrical stimulation on cell proliferation and apoptosis.

    Science.gov (United States)

    Love, Maria R; Palee, Siripong; Chattipakorn, Siriporn C; Chattipakorn, Nipon

    2018-03-01

    The application of exogenous electrical stimulation (ES) to cells in order to manipulate cell apoptosis and proliferation has been widely investigated as a possible method of treatment in a number of diseases. Alteration of the transmembrane potential of cells via ES can affect various intracellular signaling pathways which are involved in the regulation of cellular function. Controversially, several types of ES have proved to be effective in both inhibiting or inducing apoptosis, as well as increasing proliferation. However, the mechanisms through which ES achieves this remain fairly unclear. The aim of this review was to comprehensively summarize current findings from in vitro and in vivo studies on the effects of different types of ES on cell apoptosis and proliferation, highlighting the possible mechanisms through which ES induced these effects and define the optimum parameters at which ES can be used. Through this we hope to provide a greater insight into how future studies can most effectively use ES at the clinical trial stage. © 2017 Wiley Periodicals, Inc.

  11. A Single Nucleotide Polymorphism in the Bax Gene Promoter Affects Transcription and Influences Retinal Ganglion Cell Death

    Directory of Open Access Journals (Sweden)

    Sheila J Semaan

    2010-03-01

    Full Text Available Pro-apoptotic Bax is essential for RGC (retinal ganglion cell death. Gene dosage experiments in mice, yielding a single wild-type Bax allele, indicated that genetic background was able to influence the cell death phenotype. DBA/2J Bax+/− mice exhibited complete resistance to nerve damage after 2 weeks (similar to Bax −/− mice, but 129B6 Bax+/− mice exhibited significant cell loss (similar to wild-type mice. The different cell death phenotype was associated with the level of Bax expression, where 129B6 neurons had twice the level of endogenous Bax mRNA and protein as DBA/2J neurons. Sequence analysis of the Bax promoters between these strains revealed a single nucleotide polymorphism (T129B6 to CDBA/2J at position −515. A 1.5- to 2.5-fold increase in transcriptional activity was observed from the 129B6 promoter in transient transfection assays in a variety of cell types, including RGC5 cells derived from rat RGCs. Since this polymorphism occurred in a p53 half-site, we investigated the requirement of p53 for the differential transcriptional activity. Differential transcriptional activity from either 129B6 or DBA/2J Bax promoters were unaffected in p53−/− cells, and addition of exogenous p53 had no further effect on this difference, thus a role for p53 was excluded. Competitive electrophoretic mobility-shift assays identified two DNA-protein complexes that interacted with the polymorphic region. Those forming Complex 1 bound with higher affinity to the 129B6 polymorphic site, suggesting that these proteins probably comprised a transcriptional activator complex. These studies implicated quantitative expression of the Bax gene as playing a possible role in neuronal susceptibility to damaging stimuli.

  12. Apoptosis and radiosensitization of Hodgkin cells by proteasome inhibition

    International Nuclear Information System (INIS)

    Pajonk, Frank; Pajonk, Katja; McBride, William H.

    2000-01-01

    Purpose: Malignant cells from Hodgkin's disease have been reported to be defective in regulation of NF-κB activity. Ionizing radiation is known to activate NF-κB, and it has been suggested that this pathway may protect cells from apoptosis following exposure to radiation and other therapeutic agents. Defective NF-κB regulation in Hodgkin cells could therefore dictate the response of this disease to therapy, as well as be responsible for maintaining the malignant phenotype. The purpose of this study was to explore whether NF-κB activity could be modulated in Hodgkin cells and whether it determines the response of these cells to treatment with ionizing radiation and/or dexamethasone. Methods and Materials: Activation of NF-κB in cells is accomplished in large part by degradation of its inhibitor IκB through the 26s proteasome. HD-My-Z Hodgkin cells were treated with the proteasome inhibitor MG-132 or transduced with a dominant negative super-repressor IκBα. Clonogenic survival, apoptosis, proteasome activity, and NF-κB binding activity were monitored in response to ionizing radiation and/or dexamethasone treatment. Results: HD-My-Z Hodgkin cells had modest NF-κB levels but, unlike other cell types, did not decrease their level of constitutively active NF-κB in response to proteasome inhibition with MG-132. In contrast, transduction with a non-phosphorable IκBα construct abolished expression. MG-132 did, however, induce apoptosis in HD-My-Z cells and sensitized them to ionizing radiation. Dexamethasone treatment had no effect on NF-κB activity or clonogenic survival of Hodgkin cells, but protected them from irradiation. Conclusion: We conclude that inhibition of 26s proteasome activity can induce apoptosis in HD-My-Z Hodgkin cells and radiosensitize them, in spite of the fact that their constitutively active NF-κB levels are unaltered. The proteasome may be a promising new therapeutic target for intervention in this disease. In contrast, the use of

  13. Apoptosis and Vocal Fold Disease: Clinically Relevant Implications of Epithelial Cell Death

    Science.gov (United States)

    Novaleski, Carolyn K.; Carter, Bruce D.; Sivasankar, M. Preeti; Ridner, Sheila H.; Dietrich, Mary S.; Rousseau, Bernard

    2017-01-01

    Purpose: Vocal fold diseases affecting the epithelium have a detrimental impact on vocal function. This review article provides an overview of apoptosis, the most commonly studied type of programmed cell death. Because apoptosis can damage epithelial cells, this article examines the implications of apoptosis on diseases affecting the vocal fold…

  14. PDT-induced apoptosis in bladder carcinoma cells

    Science.gov (United States)

    Bachor, Ruediger; Reich, Ella D.; Kleinschmidt, Klaus; Repassy, Denes; Hautmann, Richard E.

    1999-02-01

    Photodynamic therapy (PDT) is a highly efficient inducer of apoptosis in EY-28 bladder carcinoma cells, resulting in extensive DNA fragmentation. Bladder carcinoma cells EY-28 (Tumorbank Heidelberg, Germany) were incubated for 1 h with 1 (mu) g AamTPPn/ml or 2 (mu) g AamTPPn/ml. After incubation cells were refed with complete medium and irradiated with 0.75 J/cm2. To identify apoptotic cells, a in situ cell death detection kit POD (Boehringer Mannheim, Germany) was used. The chromatin condensation characteristic to apoptotic cells was detected by transmission electron microscopy. Using 1 (mu) g AamTPPn/ml and 2 (mu) g AamTPPn/ml (9-Acetamido-2,7,12,17- tetra-n-Porpylporphycene), respectively, and irradiation at 0.75 J/cm2, a percentage of 36.9% and 54.7%, respectively, of apoptotic cells was detected.

  15. Adhesion-Dependent Regulation of Cell Growth and Apoptosis in Human Breast Cancer

    National Research Council Canada - National Science Library

    Helfman, David

    2002-01-01

    .... Normal epithelial cells require attachment to the extracellular matrix (ECM) for survival, and disruption of cell-ECM interactions results in induction of apoptosis, a phenomenon termed "anoikis...

  16. Cloning and Characterization of Genes that Inhibit TRAIL-Induced Apoptosis of Breast Cancer Cells

    National Research Council Canada - National Science Library

    Shu, Hong-Bing

    2003-01-01

    ...). However, some cancer cells are resistant to TRAIL-induced apoptosis (3, 4, 6-13). The purpose of this proposed study is to clone and characterize such inhibitory genes of TRAIL-induced apoptosis...

  17. Influence of vitamin D on cell cycle, apoptosis, and some apoptosis related molecules in systemic lupus erythematosus

    Directory of Open Access Journals (Sweden)

    Nafise Tabasi

    2015-11-01

    Full Text Available Objective(s:Genetic and environmental factors are involved in the pathogenesis of systemic lupus erythematosus (SLE. Autoreactive lymphocytes are cleared through apoptosis and any disturbance in the apoptosis or clearance of apoptotic cells may disturb tolerance and lead to autoimmunity. Vitamin D has anti-proliferative effects and controls cell cycle progression. In this study we investigated the effects of vitamin D on cell cycle and apoptosis induction in lupus patients. Materials and Methods:Isolated peripheral blood mononuclear cells (PBMCs from 25 SLE patients were cultured in the presence of 50 nM of 1,25(OH2D3; then one part of the cells were stained with FITC labeled Annexin V and PI and were analyzed for apoptosis determination. For gene expression assessment of FasL, Bcl-2 and Bax, RNA was extracted from one another part of the cells, cDNA was synthesized and gene expression analysis was performed using Real time PCR. An additional part of the cells were treated with PI and the cell cycle was analyzed using flowcytometer. Results: The mean number of early apoptotic cells in vitamin D treated cells decreased significantly (18.48±7.9% compared to untreated cells (22.02±9.4% (P=0.008. Cell cycle analysis showed a significant increase in G1 phase in vitamin D treated cells (67.33±5.2% compared to non treated ones (60.77±5.7% (P =0.02. Vitamin D up-regulated the expression levels of Bcl-2 by (18.87 fold increase, and down-regulated expression of Bax (23% and FasL (25%. Conclusion:Vitamin D has regulatory effects on cell cycle progression, apoptosis and apoptosis related molecules in lupus patients.

  18. Apoptosis induced by radionuclide 153Sm and expression of relevant genes in three different cancer cells

    International Nuclear Information System (INIS)

    Zou Baomin; Duan Xiaoyi; Chen Wei; Hu Guoying

    2003-01-01

    To study apoptosis of PC-3, ER-75-30 and A549 cells induced by radionuclide 153 Sm and the expression of bcl-2, bax in apoptosis cells, MTT assay was used to detect the anti-tumor effect, light microscope, transmission electron microscope, flow cytometer were used to detect apoptosis, while image analysis was used to detect the expression of bcl-2 and bax. 153 Sm showed anti-tumor effect and could induce tumor cell apoptosis. Both bcl-2 and bax played an important role in apoptosis. Different kind of cells had different sensitivity to 153 Sm

  19. Development of a cell-based treatment for long-term neurotrophin expression and spiral ganglion neuron survival.

    Science.gov (United States)

    Zanin, M P; Hellström, M; Shepherd, R K; Harvey, A R; Gillespie, L N

    2014-09-26

    Spiral ganglion neurons (SGNs), the target cells of the cochlear implant, undergo gradual degeneration following loss of the sensory epithelium in deafness. The preservation of a viable population of SGNs in deafness can be achieved in animal models with exogenous application of neurotrophins such as brain-derived neurotrophic factor (BDNF) and neurotrophin-3. For translation into clinical application, a suitable delivery strategy that provides ongoing neurotrophic support and promotes long-term SGN survival is required. Cell-based neurotrophin treatment has the potential to meet the specific requirements for clinical application, and we have previously reported that Schwann cells genetically modified to express BDNF can support SGN survival in deafness for 4 weeks. This study aimed to investigate various parameters important for the development of a long-term cell-based neurotrophin treatment to support SGN survival. Specifically, we investigated different (i) cell types, (ii) gene transfer methods and (iii) neurotrophins, in order to determine which variables may provide long-term neurotrophin expression and which, therefore, may be the most effective for supporting long-term SGN survival in vivo. We found that fibroblasts that were nucleofected to express BDNF provided the most sustained neurotrophin expression, with ongoing BDNF expression for at least 30 weeks. In addition, the secreted neurotrophin was biologically active and elicited survival effects on SGNs in vitro. Nucleofected fibroblasts may therefore represent a method for safe, long-term delivery of neurotrophins to the deafened cochlea to support SGN survival in deafness. Copyright © 2014 IBRO. Published by Elsevier Ltd. All rights reserved.

  20. Quantitative and Topographical Analysis of the Losses of Cone Photoreceptors and Retinal Ganglion Cells Under Taurine Depletion.

    Science.gov (United States)

    Hadj-Saïd, Wahiba; Froger, Nicolas; Ivkovic, Ivana; Jiménez-López, Manuel; Dubus, Élisabeth; Dégardin-Chicaud, Julie; Simonutti, Manuel; Quénol, César; Neveux, Nathalie; Villegas-Pérez, María Paz; Agudo-Barriuso, Marta; Vidal-Sanz, Manuel; Sahel, Jose-Alain; Picaud, Serge; García-Ayuso, Diego

    2016-09-01

    Taurine depletion is known to induce photoreceptor degeneration and was recently found to also trigger retinal ganglion cell (RGC) loss similar to the retinal toxicity of vigabatrin. Our objective was to study the topographical loss of RGCs and cone photoreceptors, with a distinction between the two cone types (S- and L- cones) in an animal model of induced taurine depletion. We used the taurine transporter (Tau-T) inhibitor, guanidoethane sulfonate (GES), to induce taurine depletion at a concentration of 1% in the drinking water. Spectral-domain optical coherence tomography (SD-OCT) and electroretinograms (ERG) were performed on animals after 2 months of GES treatment administered through the drinking water. Retinas were dissected as wholemounts and immunodetection of Brn3a (RGC), S-opsin (S-cones), and L-opsin (L-cones) was performed. The number of Brn3a+ RGCs, and L- and S-opsin+ cones was automatically quantified and their retinal distribution studied using isodensity maps. The treatment resulted in a significant reduction in plasma taurine levels and a profound dysfunction of visual performance as shown by ERG recordings. Optical coherence tomography analysis revealed that the retina was thinner in the taurine-depleted group. S-opsin+cones were more affected (36%) than L-opsin+cones (27%) with greater cone cell loss in the dorsal area whereas RGC loss (12%) was uniformly distributed. This study confirms that taurine depletion causes RGC and cone loss. Electroretinograms results show that taurine depletion induces retinal dysfunction in photoreceptors and in the inner retina. It establishes a gradient of cell loss depending on the cell type from S-opsin+cones, L-opsin+cones, to RGCs. The greater cell loss in the dorsal retina and of the S-cone population may underline different cellular mechanisms of cellular degeneration and suggests that S-cones may be more sensitive to light-induced retinal toxicity enhanced by the taurine depletion.

  1. Optimization of micropatterned poly(lactic-co-glycolic acid films for enhancing dorsal root ganglion cell orientation and extension

    Directory of Open Access Journals (Sweden)

    Ching-Wen Li

    2018-01-01

    Full Text Available Nerve conduits have been a viable alternative to the ‘gold standard’ autograft for treating small peripheral nerve gap injuries. However, they often produce inadequate functional recovery outcomes and are ineffective in large gap injuries. Ridge/groove surface micropatterning has been shown to promote neural cell orientation and guide growth. However, optimization of the ratio of ridge/groove parameters to promote orientation and extension for dorsal root ganglion (DRG cells on poly(lactic-co-glycolic acid (PLGA films has not been previously conducted. Photolithography and micro-molding were used to define various combinations of ridge/groove dimensions on PLGA films. The DRG cells obtained from chicken embryos were cultured on micropatterned PLGA films for cell orientation and migration evaluation. Biodegradation of the films occurred during the test period, however, this did not cause deformation or distortion of the micropatterns. Results from the DRG cell orientation test suggest that when the ridge/groove ratio equals 1 (ridge/groove width parameters are equal, i.e., 10 μm/10 μm (even, the degree of alignment depends on the size of the ridges and grooves, when the ratio is smaller than 1 (groove controlled the alignment increases as the ridge size decreases, and when the ratio is larger than 1 (ridge controlled, the alignment is reduced as the width of the grooves decreases. The migration rate and neurite extension of DRG neurons were greatest on 10 μm/10 μm and 30 μm/30 μm micropatterned PLGA films. Based on the data, the 10 μm/10 μm and 30 μm/30 μm micropatterned PLGA films are the optimized ridge/groove surface patterns for the construction of nerve repair devices.

  2. Metformin prevents methylglyoxal-induced apoptosis of mouse Schwann cells

    International Nuclear Information System (INIS)

    Ota, Kimiko; Nakamura, Jiro; Li, Weiguo; Kozakae, Mika; Watarai, Atsuko; Nakamura, Nobuhisa; Yasuda, Yutaka; Nakashima, Eirtaro; Naruse, Keiko; Watabe, Kazuhiko; Kato, Koichi; Oiso, Yutaka; Hamada, Yoji

    2007-01-01

    Methylglyoxal (MG) is involved in the pathogenesis of diabetic complications via the formation of advanced glycation end products (AGEs) and reactive oxygen species (ROS). To clarify whether the antidiabetic drug metformin prevents Schwann cell damage induced by MG, we cultured mouse Schwann cells in the presence of MG and metformin. Cell apoptosis was evaluated using Hoechst 33342 nuclear staining, caspase-3 activity, and c-Jun-N-terminal kinase (JNK) phosphorylation. Intracellular ROS formation was determined by flow cytometry, and AMP-activated kinase (AMPK) phosphorylation was also examined. MG treatment resulted in blunted cell proliferation, an increase in the number of apoptotic cells, and the activation of caspase-3 and JNK along with enhanced intracellular ROS formation. All of these changes were significantly inhibited by metformin. No significant activation of AMPK by MG or metformin was observed. Taken together, metformin likely prevents MG-induced apoptotic signals in mouse Schwann cells by inhibiting the formation of AGEs and ROS

  3. Rhein Induces Apoptosis in Human Breast Cancer Cells

    Directory of Open Access Journals (Sweden)

    Ching-Yao Chang

    2012-01-01

    Full Text Available Human breast cancers cells overexpressing HER2/neu are more aggressive tumors with poor prognosis, and resistance to chemotherapy. This study investigates antiproliferation effects of anthraquinone derivatives of rhubarb root on human breast cancer cells. Of 7 anthraquinone derivatives, only rhein showed antiproliferative and apoptotic effects on both HER2-overexpressing MCF-7 (MCF-7/HER2 and control vector MCF-7 (MCF-7/VEC cells. Rhein induced dose- and time-dependent manners increase in caspase-9-mediated apoptosis correlating with activation of ROS-mediated activation of NF-κB- and p53-signaling pathways in both cell types. Therefore, this study highlighted rhein as processing anti-proliferative activity against HER2 overexpression or HER2-basal expression in breast cancer cells and playing important roles in apoptotic induction of human breast cancer cells.

  4. Cord blood stem cell-mediated induction of apoptosis in glioma downregulates X-linked inhibitor of apoptosis protein (XIAP.

    Directory of Open Access Journals (Sweden)

    Venkata Ramesh Dasari

    2010-07-01

    Full Text Available XIAP (X-linked inhibitor of apoptosis protein is one of the most important members of the apoptosis inhibitor family. XIAP is upregulated in various malignancies, including human glioblastoma. It promotes invasion, metastasis, growth and survival of malignant cells. We hypothesized that downregulation of XIAP by human umbilical cord blood mesenchymal stem cells (hUCBSC in glioma cells would cause them to undergo apoptotic death.We observed the effect of hUCBSC on two malignant glioma cell lines (SNB19 and U251 and two glioma xenograft cell lines (4910 and 5310. In co-cultures of glioma cells with hUCBSC, proliferation of glioma cells was significantly inhibited. This is associated with increased cytotoxicity of glioma cells, which led to glioma cell death. Stem cells induced apoptosis in glioma cells, which was evaluated by TUNEL assay, FACS analyses and immunoblotting. The induction of apoptosis is associated with inhibition of XIAP in co-cultures of hUCBSC. Similar results were obtained by the treatment of glioma cells with shRNA to downregulate XIAP (siXIAP. Downregulation of XIAP resulted in activation of caspase-3 and caspase-9 to trigger apoptosis in glioma cells. Apoptosis is characterized by the loss of mitochondrial membrane potential and upregulation of mitochondrial apoptotic proteins Bax and Bad. Cell death of glioma cells was marked by downregulation of Akt and phospho-Akt molecules. We observed similar results under in vivo conditions in U251- and 5310-injected nude mice brains, which were treated with hUCBSC. Under in vivo conditions, Smac/DIABLO was found to be colocalized in the nucleus, showing that hUCBSC induced apoptosis is mediated by inhibition of XIAP and activation of Smac/DIABLO.Our results indicate that downregulation of XIAP by hUCBSC treatment induces apoptosis, which led to the death of the glioma cells and xenograft cells. This study demonstrates the therapeutic potential of XIAP and hUCBSC to treat malignant

  5. Protection by an oral disubstituted hydroxylamine derivative against loss of retinal ganglion cell differentiation following optic nerve crush.

    Directory of Open Access Journals (Sweden)

    James D Lindsey

    Full Text Available Thy-1 is a cell surface protein that is expressed during the differentiation of retinal ganglion cells (RGCs. Optic nerve injury induces progressive loss in the number of RGCs expressing Thy-1. The rate of this loss is fastest during the first week after optic nerve injury and slower in subsequent weeks. This study was undertaken to determine whether oral treatment with a water-soluble N-hydroxy-2,2,6,6-tetramethylpiperidine derivative (OT-440 protects against loss of Thy-1 promoter activation following optic nerve crush and whether this effect targets the earlier quick phase or the later slow phase. The retina of mice expressing cyan fluorescent protein under control of the Thy-1 promoter (Thy1-CFP mice was imaged using a blue-light confocal scanning laser ophthalmoscope (bCSLO. These mice then received oral OT-440 prepared in cream cheese or dissolved in water, or plain vehicle, for two weeks and were imaged again prior to unilateral optic nerve crush. Treatments and weekly imaging continued for four more weeks. Fluorescent neurons were counted in the same defined retinal areas imaged at each time point in a masked fashion. When the counts at each time point were directly compared, the numbers of fluorescent cells at each time point were greater in the animals that received OT-440 in cream cheese by 8%, 27%, 52% and 60% than in corresponding control animals at 1, 2, 3 and 4 weeks after optic nerve crush. Similar results were obtained when the vehicle was water. Rate analysis indicated the protective effect of OT-440 was greatest during the first two weeks and was maintained in the second two weeks after crush for both the cream cheese vehicle study and water vehicle study. Because most of the fluorescent cells detected by bCSLO are RGCs, these findings suggest that oral OT-440 can either protect against or delay early degenerative responses occurring in RGCs following optic nerve injury.

  6. Protection by an oral disubstituted hydroxylamine derivative against loss of retinal ganglion cell differentiation following optic nerve crush.

    Science.gov (United States)

    Lindsey, James D; Duong-Polk, Karen X; Dai, Yi; Nguyen, Duy H; Leung, Christopher K; Weinreb, Robert N

    2013-01-01

    Thy-1 is a cell surface protein that is expressed during the differentiation of retinal ganglion cells (RGCs). Optic nerve injury induces progressive loss in the number of RGCs expressing Thy-1. The rate of this loss is fastest during the first week after optic nerve injury and slower in subsequent weeks. This study was undertaken to determine whether oral treatment with a water-soluble N-hydroxy-2,2,6,6-tetramethylpiperidine derivative (OT-440) protects against loss of Thy-1 promoter activation following optic nerve crush and whether this effect targets the earlier quick phase or the later slow phase. The retina of mice expressing cyan fluorescent protein under control of the Thy-1 promoter (Thy1-CFP mice) was imaged using a blue-light confocal scanning laser ophthalmoscope (bCSLO). These mice then received oral OT-440 prepared in cream cheese or dissolved in water, or plain vehicle, for two weeks and were imaged again prior to unilateral optic nerve crush. Treatments and weekly imaging continued for four more weeks. Fluorescent neurons were counted in the same defined retinal areas imaged at each time point in a masked fashion. When the counts at each time point were directly compared, the numbers of fluorescent cells at each time point were greater in the animals that received OT-440 in cream cheese by 8%, 27%, 52% and 60% than in corresponding control animals at 1, 2, 3 and 4 weeks after optic nerve crush. Similar results were obtained when the vehicle was water. Rate analysis indicated the protective effect of OT-440 was greatest during the first two weeks and was maintained in the second two weeks after crush for both the cream cheese vehicle study and water vehicle study. Because most of the fluorescent cells detected by bCSLO are RGCs, these findings suggest that oral OT-440 can either protect against or delay early degenerative responses occurring in RGCs following optic nerve injury.

  7. Gene therapy with brain-derived neurotrophic factor as a protection: retinal ganglion cells in a rat glaucoma model.

    Science.gov (United States)

    Martin, Keith R G; Quigley, Harry A; Zack, Donald J; Levkovitch-Verbin, Hana; Kielczewski, Jennifer; Valenta, Danielle; Baumrind, Lisa; Pease, Mary Ellen; Klein, Ronald L; Hauswirth, William W

    2003-10-01

    To develop a modified adenoassociated viral (AAV) vector capable of efficient transfection of retinal ganglion cells (RGCs) and to test the hypothesis that use of this vector to express brain-derived neurotrophic factor (BDNF) could be protective in experimental glaucoma. Ninety-three rats received one unilateral, intravitreal injection of either normal saline (n = 30), AAV-BDNF-woodchuck hepatitis posttranscriptional regulatory element (WPRE; n = 30), or AAV-green fluorescent protein (GFP)-WPRE (n = 33). Two weeks later, experimental glaucoma was induced in the injected eye by laser application to the trabecular meshwork. Survival of RGCs was estimated by counting axons in optic nerve cross sections after 4 weeks of glaucoma. Transgene expression was assessed by immunohistochemistry, Western blot analysis, and direct visualization of GFP. The density of GFP-positive cells in retinal wholemounts was 1,828 +/- 299 cells/mm(2) (72,273 +/- 11,814 cells/retina). Exposure to elevated intraocular pressure was similar in all groups. Four weeks after initial laser treatment, axon loss was 52.3% +/- 27.1% in the saline-treated group (n = 25) and 52.3% +/- 24.2% in the AAV-GFP-WPRE group (n = 30), but only 32.3% +/- 23.0% in the AAV-BDNF-WPRE group (n = 27). Survival in AAV-BDNF-WPRE animals increased markedly and the difference was significant compared with those receiving either AAV-GFP-WPRE (P = 0.002, t-test) or saline (P = 0.006, t-test). Overexpression of the BDNF gene protects RGC as estimated by axon counts in a rat glaucoma model, further supporting the potential feasibility of neurotrophic therapy as a complement to the lowering of IOP in the treatment of glaucoma.

  8. Retinal nerve fiber layer and ganglion cell complex thickness assessment in patients with Alzheimer disease and mild cognitive impairment. Preliminary results

    Directory of Open Access Journals (Sweden)

    A. S. Tiganov

    2014-07-01

    Full Text Available Purpose: to investigate the retinal nerve fiber layer (RNFL and the macular ganglion cell complex (GCC in patients with Alzheimer`s disease and mild cognitive impairment.Methods: this study included 10 patients (20 eyes with Alzheimer`s disease, 10 patients with mild cognitive impairment and 10 age- and sex-matched healthy controls that had no history of dementia. All the subjects underwent psychiatric examination, including the Mini-Mental State Examination (MMSE, and complete ophthalmological examination, comprising optical coherence tomography and scanning laser polarimetry.Results: there was a significant decrease in GCC thickness in patients with Alzheimer`s disease compared to the control group, global loss volume of ganglion cells was higher than in control group. there was no significant difference among the groups in terms of RNFL thickness. Weak positive correlation of GCC thickness and MMSE results was observed.Conclusion: Our data confirm the retinal involvement in Alzheimer`s disease, as reflected by loss of ganglion cells. Further studies will clear up the role and contribution of dementia in pathogenesis of optic neuropathy.

  9. Relationship between macular ganglion cell complex parameters and visual field parameters after tumor resection in chiasmal compression.

    Science.gov (United States)

    Ohkubo, Shinji; Higashide, Tomomi; Takeda, Hisashi; Murotani, Eiji; Hayashi, Yasuhiko; Sugiyama, Kazuhisa

    2012-01-01

    To evaluate the relationship between macular ganglion cell complex (GCC) parameters and visual field (VF) parameters in chiasmal compression and the potential for GCC parameters in order to predict the short-term postsurgical VF. Twenty-three eyes of 12 patients with chiasmal compression and 33 control eyes were studied. All patients underwent transsphenoidal tumor resection. Before surgery a 3D scan of the macula was taken using spectral-domain optical coherence tomography. All patients underwent Humphrey 24-2 VF testing after surgery. Spearman's rank correlation coefficients were used to evaluate the relationship between the GCC parameters and VF parameters [mean deviation (MD), pattern standard deviation]. Coefficients of determination (R2) were calculated using linear regression. Average thickness in the patients was significantly thinner than that of controls. Average thickness, global loss volume and focal loss volume (FLV) significantly correlated with the MD. We observed the greatest R2 between FLV and MD. Examining the macular GCC was useful for evaluating structural damage in patients with chiasmal compression. Preoperative GCC parameters, especially FLV, may be useful in predicting visual function following surgical decompression of chiasmal compression.

  10. Sustained Dorzolamide Release Prevents Axonal and Retinal Ganglion Cell Loss in a Rat Model of IOP-Glaucoma.

    Science.gov (United States)

    Pitha, Ian; Kimball, Elizabeth C; Oglesby, Ericka N; Pease, Mary Ellen; Fu, Jie; Schaub, Julie; Kim, Yoo-Chun; Hu, Qi; Hanes, Justin; Quigley, Harry A

    2018-04-01

    To determine if one injection of a sustained release formulation of dorzolamide in biodegradable microparticles (DPP) reduces retinal ganglion cell (RGC) loss in a rat model of glaucoma. We injected either DPP or control microparticles intravitreally in rats. Two days later, unilateral ocular hypertension was induced by translimbal, diode laser treatment by a surgeon masked to treatment group. IOP and clinical exams were performed until sacrifice 6 weeks after laser treatment. RGC loss was measured by masked observers in both optic nerve cross-sections and RGC layer counts from retinal whole mounts. Cumulative IOP exposure was significantly reduced by DPP injection (49 ± 48 mm Hg × days in treated versus 227 ± 191 mm Hg × days in control microparticle eyes; P = 0.012, t -test). While control-injected eyes increased in axial length by 2.4 ± 1.7%, DPP eyes did not significantly enlarge (0.3 ± 2.2%, difference from control, P = 0.03, t -test). RGC loss was significantly less in DPP eyes compared with control microparticle injection alone (RGC axon count reduction: 21% vs. 52%; RGC body reduction: 25% vs. 50% [beta tubulin labeling]; P = 0.02, t -test). A single injection of sustained release DPP protected against RGC loss and axial elongation in a rat model of IOP glaucoma. Sustained release IOP-lowering medications have the potential to stop glaucoma progression.

  11. Glutamatergic neurotransmission from melanopsin retinal ganglion cells is required for neonatal photoaversion but not adult pupillary light reflex.

    Directory of Open Access Journals (Sweden)

    Anton Delwig

    Full Text Available Melanopsin-expressing retinal ganglion cells (mRGCs in the eye play an important role in many light-activated non-image-forming functions including neonatal photoaversion and the adult pupillary light reflex (PLR. MRGCs rely on glutamate and possibly PACAP (pituitary adenylate cyclase-activating polypeptide to relay visual signals to the brain. However, the role of these neurotransmitters for individual non-image-forming responses remains poorly understood. To clarify the role of glutamatergic signaling from mRGCs in neonatal aversion to light and in adult PLR, we conditionally deleted vesicular glutamate transporter (VGLUT2 selectively from mRGCs in mice. We found that deletion of VGLUT2 in mRGCs abolished negative phototaxis and light-induced distress vocalizations in neonatal mice, underscoring a necessary role for glutamatergic signaling. In adult mice, loss of VGLUT2 in mRGCs resulted in a slow and an incomplete PLR. We conclude that glutamatergic neurotransmission from mRGCs is required for neonatal photoaversion but is complemented by another non-glutamatergic signaling mechanism for the pupillary light reflex in adult mice. We speculate that this complementary signaling might be due to PACAP neurotransmission from mRGCs.

  12. Nuclear orphan receptor TLX affects gene expression, proliferation and cell apoptosis in beta cells

    Energy Technology Data Exchange (ETDEWEB)

    Shi, Xiaoli; Xiong, Xiaokan; Dai, Zhe; Deng, Haohua; Sun, Li; Hu, Xuemei; Zhou, Feng; Xu, Yancheng, E-mail: oxyccc@163.com

    2015-12-04

    Nuclear orphan receptor TLX is an essential regulator of the growth of neural stem cells. However, its exact function in pancreatic islet cells is still unknown. In the present study, gene expression profiling analysis revealed that overexpression of TLX in beta cell line MIN6 causes suppression of 176 genes and upregulation of 49 genes, including a cadre of cell cycle, cell proliferation and cell death control genes, such as Btg2, Ddit3 and Gadd45a. We next examined the effects of TLX overexpression on proliferation, apoptosis and insulin secretion in MIN6 cells. Proliferation analysis using EdU assay showed that overexpression of TLX increased percentage of EdU-positive cells. Cell cycle and apoptosis analysis revealed that overexpression of TLX in MIN6 cells resulted in higher percentage of cells exiting G1 into S-phase, and a 58.8% decrease of cell apoptosis induced by 0.5 mM palmitate. Moreover, TLX overexpression did not cause impairment of insulin secretion. Together, we conclude that TLX is among factors capable of controlling beta cell proliferation and survival, which may serve as a target for the development of novel therapies for diabetes. - Highlights: • TLX overexpression in MIN6 cell causes significant expression changes of 225 genes. • TLX overexpression promotes MIN6 cell proliferation and decreases cell apoptosis. • TLX overexpression does not cause impairment of insulin secretion.

  13. Nuclear orphan receptor TLX affects gene expression, proliferation and cell apoptosis in beta cells

    International Nuclear Information System (INIS)

    Shi, Xiaoli; Xiong, Xiaokan; Dai, Zhe; Deng, Haohua; Sun, Li; Hu, Xuemei; Zhou, Feng; Xu, Yancheng

    2015-01-01

    Nuclear orphan receptor TLX is an essential regulator of the growth of neural stem cells. However, its exact function in pancreatic islet cells is still unknown. In the present study, gene expression profiling analysis revealed that overexpression of TLX in beta cell line MIN6 causes suppression of 176 genes and upregulation of 49 genes, including a cadre of cell cycle, cell proliferation and cell death control genes, such as Btg2, Ddit3 and Gadd45a. We next examined the effects of TLX overexpression on proliferation, apoptosis and insulin secretion in MIN6 cells. Proliferation analysis using EdU assay showed that overexpression of TLX increased percentage of EdU-positive cells. Cell cycle and apoptosis analysis revealed that overexpression of TLX in MIN6 cells resulted in higher percentage of cells exiting G1 into S-phase, and a 58.8% decrease of cell apoptosis induced by 0.5 mM palmitate. Moreover, TLX overexpression did not cause impairment of insulin secretion. Together, we conclude that TLX is among factors capable of controlling beta cell proliferation and survival, which may serve as a target for the development of novel therapies for diabetes. - Highlights: • TLX overexpression in MIN6 cell causes significant expression changes of 225 genes. • TLX overexpression promotes MIN6 cell proliferation and decreases cell apoptosis. • TLX overexpression does not cause impairment of insulin secretion.

  14. Studies on hematopoietic cell apoptosis and the relative gene expression in irradiated mouse bone marrow

    International Nuclear Information System (INIS)

    Peng Ruiyun; Wang Dewen; Xiong Chengqi; Gao Yabing; Yang Hong; Cui Yufang; Wang Baozhen

    2001-01-01

    Objective: To study apoptosis and expressions bcl-2 and p53 in irradiated mouse bone marrow. Methods: LACA mice were irradiated with 60 Co γ-rays. By means of in situ terminal labelling, in situ hybridization and image analysis, the authors studied radiation-induced apoptosis of hematopoietic cells and the expressions of bcl-2 and p53. Results: The characteristics of apoptosis appeared in hematopoietic cells at 6 hrs after irradiation. The expression of bcl-2 was obviously decreased when apoptosis of hematopoietic cells occurred, whereas it increased in the early recovery phase; p53 protein increased during both apoptosis of hematopoietic cells and the recovery phase, and mutant type p53 DNA was positive only in the recovery phase. Conclusion: Radiation may induced apoptosis of hematopoietic cells in a dose-dependent manner; Both bcl-2 and p53 genes play an important role in apoptosis and recovery phase

  15. Suppression of ICE and Apoptosis in Mammary Epithelial Cells by Extracellular Matrix

    Energy Technology Data Exchange (ETDEWEB)

    Boudreau, Nancy; Sympson, C. J.; Werb, Zena; Bissell, Mina J.

    1994-12-01

    Apoptosis (programmed cell death) plays a major role in development and tissue regeneration. Basement membrane extracellular matrix (ECM), but not fibronectin or collagen, was shown to suppress apoptosis of mammary epithelial cells in tissue culture and in vivo. Apoptosis was induced by antibodies to beta 1 integrins or by overexpression of stromelysin-1, which degrades ECM. Expression of interleukin-1 beta converting enzyme (ICE) correlated with the loss of ECM, and inhibitors of ICE activity prevented apoptosis. These results suggest that ECM regulates apoptosis in mammary epithelial cells through an integrin-dependent negative regulation of ICE expression.

  16. Curcumin induces apoptosis-independent death in oesophageal cancer cells.

    LENUS (Irish Health Repository)

    O'Sullivan-Coyne, G

    2012-01-31

    BACKGROUND: Oesophageal cancer incidence is increasing and survival rates remain extremely poor. Natural agents with potential for chemoprevention include the phytochemical curcumin (diferuloylmethane). We have examined the effects of curcumin on a panel of oesophageal cancer cell lines. METHODS: MTT (3-(4,5-dimethyldiazol-2-yl)-2,5 diphenyl tetrazolium bromide) assays and propidium iodide staining were used to assess viability and DNA content, respectively. Mitotic catastrophe (MC), apoptosis and autophagy were defined by both morphological criteria and markers such as MPM-2, caspase 3 cleavage and monodansylcadaverine (MDC) staining. Cyclin B and poly-ubiquitinated proteins were assessed by western blotting. RESULTS: Curcumin treatment reduces viability of all cell lines within 24 h of treatment in a 5-50 muM range. Cytotoxicity is associated with accumulation in G2\\/M cell-cycle phases and distinct chromatin morphology, consistent with MC. Caspase-3 activation was detected in two out of four cell lines, but was a minor event. The addition of a caspase inhibitor zVAD had a marginal or no effect on cell viability, indicating predominance of a non-apoptotic form of cell death. In two cell lines, features of both MC and autophagy were apparent. Curcumin-responsive cells were found to accumulate poly-ubiquitinated proteins and cyclin B, consistent with a disturbance of the ubiquitin-proteasome system. This effect on a key cell-cycle checkpoint regulator may be responsible for the mitotic disturbances and consequent cytotoxicity of this drug. CONCLUSION: Curcumin can induce cell death by a mechanism that is not reliant on apoptosis induction, and thus represents a promising anticancer agent for prevention and treatment of oesophageal cancer.

  17. Curcumin induces apoptosis-independent death in oesophageal cancer cells.

    LENUS (Irish Health Repository)

    O'Sullivan-Coyne, G

    2009-10-06

    Background:Oesophageal cancer incidence is increasing and survival rates remain extremely poor. Natural agents with potential for chemoprevention include the phytochemical curcumin (diferuloylmethane). We have examined the effects of curcumin on a panel of oesophageal cancer cell lines.Methods:MTT (3-(4,5-dimethyldiazol-2-yl)-2,5 diphenyl tetrazolium bromide) assays and propidium iodide staining were used to assess viability and DNA content, respectively. Mitotic catastrophe (MC), apoptosis and autophagy were defined by both morphological criteria and markers such as MPM-2, caspase 3 cleavage and monodansylcadaverine (MDC) staining. Cyclin B and poly-ubiquitinated proteins were assessed by western blotting.Results:Curcumin treatment reduces viability of all cell lines within 24 h of treatment in a 5-50 muM range. Cytotoxicity is associated with accumulation in G2\\/M cell-cycle phases and distinct chromatin morphology, consistent with MC. Caspase-3 activation was detected in two out of four cell lines, but was a minor event. The addition of a caspase inhibitor zVAD had a marginal or no effect on cell viability, indicating predominance of a non-apoptotic form of cell death. In two cell lines, features of both MC and autophagy were apparent. Curcumin-responsive cells were found to accumulate poly-ubiquitinated proteins and cyclin B, consistent with a disturbance of the ubiquitin-proteasome system. This effect on a key cell-cycle checkpoint regulator may be responsible for the mitotic disturbances and consequent cytotoxicity of this drug.Conclusion:Curcumin can induce cell death by a mechanism that is not reliant on apoptosis induction, and thus represents a promising anticancer agent for prevention and treatment of oesophageal cancer.British Journal of Cancer advance online publication, 6 October 2009; doi:10.1038\\/sj.bjc.6605308 www.bjcancer.com.

  18. Restraining reactive oxygen species in Listeria monocytogenes promotes the apoptosis of glial cells.

    Science.gov (United States)

    Li, Sen; Li, Yixuan; Chen, Guowei; Zhang, Jingchen; Xu, Fei; Wu, Man

    2017-07-01

    Listeria monocytogenes is a facultative anaerobic foodborne pathogen that can traverse the blood-brain barrier and cause brain infection. L. monocytogenes infection induces host cell apoptosis in several cell types. In this study, we investigated the apoptosis of human glioma cell line U251 invaded by L. monocytogenes and evaluated the function of bacterial reactive oxygen species (ROS) during infection. Bacterial ROS level was reduced by carrying out treatment with N-acetyl cysteine (NAC) and diphenyleneiodonium chloride (DPI). After infection, the apoptosis of U251 cells was examined by flow cytometry assay and propidium iodide staining. DPI and NAC efficiently decreased ROS level in L. monocytogenes without affecting bacterial growth. Moreover, the apoptosis of glial cells was enhanced upon invasion of DPI- and NAC-pretreated L. monocytogenes. Results indicate that the apoptosis of glial cells can be induced by L. monocytogenes, and that the inhibition of bacterial ROS increases the apoptosis of host cells.

  19. Vesicular glutamate transporter 2 (VGLUT2) is co-stored with PACAP in projections from the rat melanopsin-containing retinal ganglion cells

    DEFF Research Database (Denmark)

    Engelund, Anna Iversen; Fahrenkrug, Jan; Harrison, Adrian Paul

    2010-01-01

    The retinal ganglion cell layer of the eye comprises a subtype of cells characterized by their intrinsic photosensitivity and expression of melanopsin (ipRGCs). These cells regulate a variety of non-image-forming (NIF) functions such as light entrainment of circadian rhythms, acute suppression......-localized in their projections in the suprachiasmatic nucleus, the intergeniculate leaflet, and the olivary pretectal nucleus. We conclude that there is evidence to support the use of glutamate and PACAP as neurotransmitters in NIF photoperception by rat ipRGCs, and that these neurotransmitters are co-stored and probably...

  20. Molecular events associated with increased regenerative capacity of the goldfish retinal ganglion cells following X-irradiation: decreased level of axonal growth inhibitors

    International Nuclear Information System (INIS)

    Rachailovich, I.; Schwartz, M.

    1984-01-01

    In our previous work we established conditions to study the contribution of non-neuronal cells to the process of goldfish optic nerve regeneration. This issue has been studied successfully by adapting the use of X-irradiation to manipulate division of non-neuronal cells associated with the injured nerve. The regenerative capacity of the goldfish retinal ganglion cells was determined subsequent to the X-ray treatment. The authors present an analysis of the molecular events associated with regeneration and enhanced regenerative capacity which follows X-irradiation. (Auth.)

  1. Molecular events associated with increased regenerative capacity of the goldfish retinal ganglion cells following X-irradiation: decreased level of axonal growth inhibitors

    Energy Technology Data Exchange (ETDEWEB)

    Rachailovich, I.; Schwartz, M. (Weizmann Inst. of Science, Rehovot (Israel). Dept. of Neurobiology)

    1984-07-23

    In our previous work we established conditions to study the contribution of non-neuronal cells to the process of goldfish optic nerve regeneration. This issue has been studied successfully by adapting the use of X-irradiation to manipulate division of non-neuronal cells associated with the injured nerve. The regenerative capacity of the goldfish retinal ganglion cells was determined subsequent to the X-ray treatment. The authors present an analysis of the molecular events associated with regeneration and enhanced regenerative capacity which follows X-irradiation.

  2. Expression of SPIG1 reveals development of a retinal ganglion cell subtype projecting to the medial terminal nucleus in the mouse.

    Directory of Open Access Journals (Sweden)

    Keisuke Yonehara

    Full Text Available Visual information is transmitted to the brain by roughly a dozen distinct types of retinal ganglion cells (RGCs defined by a characteristic morphology, physiology, and central projections. However, our understanding about how these parallel pathways develop is still in its infancy, because few molecular markers corresponding to individual RGC types are available. Previously, we reported a secretory protein, SPIG1 (clone name; D/Bsp120I #1, preferentially expressed in the dorsal region in the developing chick retina. Here, we generated knock-in mice to visualize SPIG1-expressing cells with green fluorescent protein. We found that the mouse retina is subdivided into two distinct domains for SPIG1 expression and SPIG1 effectively marks a unique subtype of the retinal ganglion cells during the neonatal period. SPIG1-positive RGCs in the dorsotemporal domain project to the dorsal lateral geniculate nucleus (dLGN, superior colliculus, and accessory optic system (AOS. In contrast, in the remaining region, here named the pan-ventronasal domain, SPIG1-positive cells form a regular mosaic and project exclusively to the medial terminal nucleus (MTN of the AOS that mediates the optokinetic nystagmus as early as P1. Their dendrites costratify with ON cholinergic amacrine strata in the inner plexiform layer as early as P3. These findings suggest that these SPIG1-positive cells are the ON direction selective ganglion cells (DSGCs. Moreover, the MTN-projecting cells in the pan-ventronasal domain are apparently composed of two distinct but interdependent regular mosaics depending on the presence or absence of SPIG1, indicating that they comprise two functionally distinct subtypes of the ON DSGCs. The formation of the regular mosaic appears to be commenced at the end of the prenatal stage and completed through the peak period of the cell death at P6. SPIG1 will thus serve as a useful molecular marker for future studies on the development and function of ON DSGCs.

  3. The Cytotoxic Role of Intermittent High Glucose on Apoptosis and Cell Viability in Pancreatic Beta Cells

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    Zhen Zhang

    2014-01-01

    Full Text Available Objectives. Glucose fluctuations are both strong predictor of diabetic complications and crucial factor for beta cell damages. Here we investigated the effect of intermittent high glucose (IHG on both cell apoptosis and proliferation activity in INS-1 cells and the potential mechanisms. Methods. Cells were treated with normal glucose (5.5 mmol/L, constant high glucose (CHG (25 mmol/L, and IHG (rotation per 24 h in 11.1 or 25 mmol/L for 7 days. Reactive oxygen species (ROS, xanthine oxidase (XOD level, apoptosis, cell viability, cell cycle, and expression of cyclinD1, p21, p27, and Skp2 were determined. Results. We found that IHG induced more significant apoptosis than CHG and normal glucose; intracellular ROS and XOD levels were more markedly increased in cells exposed to IHG. Cells treated with IHG showed significant decreased cell viability and increased cell proportion in G0/G1 phase. Cell cycle related proteins such as cyclinD1 and Skp2 were decreased significantly, but expressions of p27 and p21 were increased markedly. Conclusions. This study suggested that IHG plays a more toxic effect including both apoptosis-inducing and antiproliferative effects on INS-1 cells. Excessive activation of cellular stress and regulation of cyclins might be potential mechanism of impairment in INS-1 cells induced by IHG.

  4. Substituting mouse transcription factor Pou4f2 with a sea urchin orthologue restores retinal ganglion cell development.

    Science.gov (United States)

    Mao, Chai-An; Agca, Cavit; Mocko-Strand, Julie A; Wang, Jing; Ullrich-Lüter, Esther; Pan, Ping; Wang, Steven W; Arnone, Maria Ina; Frishman, Laura J; Klein, William H

    2016-03-16

    Pou domain transcription factor Pou4f2 is essential for the development of retinal ganglion cells (RGCs) in the vertebrate retina. A distant orthologue of Pou4f2 exists in the genome of the sea urchin (class Echinoidea) Strongylocentrotus purpuratus (SpPou4f1/2), yet the photosensory structure of sea urchins is strikingly different from that of the mammalian retina. Sea urchins have no obvious eyes, but have photoreceptors clustered around their tube feet disc. The mechanisms that are associated with the development and function of photoreception in sea urchins are largely unexplored. As an initial approach to better understand the sea urchin photosensory structure and relate it to the mammalian retina, we asked whether SpPou4f1/2 could support RGC development in the absence of Pou4f2. To answer this question, we replaced genomic Pou4f2 with an SpPou4f1/2 cDNA. In Pou4f2-null mice, retinas expressing SpPou4f1/2 were outwardly identical to those of wild-type mice. SpPou4f1/2 retinas exhibited dark-adapted electroretinogram scotopic threshold responses, indicating functionally active RGCs. During retinal development, SpPou4f1/2 activated RGC-specific genes and in S. purpuratus, SpPou4f2 was expressed in photoreceptor cells of tube feet in a pattern distinct from Opsin4 and Pax6. Our results suggest that SpPou4f1/2 and Pou4f2 share conserved components of a gene network for photosensory development and they maintain their conserved intrinsic functions despite vast morphological differences in mouse and sea urchin photosensory structures. © 2016 The Authors.

  5. Multipronged approach to identify and validate a novel upstream regulator of Sncg in mouse retinal ganglion cells.

    Science.gov (United States)

    Chintalapudi, Sumana R; Morales-Tirado, Vanessa M; Williams, Robert W; Jablonski, Monica M

    2016-02-01

    Loss of retinal ganglion cells (RGCs) is one of the hallmarks of retinal neurodegenerative diseases, glaucoma being one of the most common. Mechanistic studies on RGCs are hindered by the lack of sufficient primary cells and consensus regarding their signature markers. Recently, γ-synuclein (SNCG) has been shown to be highly expressed in the somas and axons of RGCs. In various mouse models of glaucoma, downregulation of Sncg gene expression correlates with RGC loss. To investigate the role of Sncg in RGCs, we used a novel systems genetics approach to identify a gene that modulates Sncg expression, followed by confirmatory studies in both healthy and diseased retinae. We found that chromosome 1 harbors an expression quantitative trait locus that modulates Sncg expression in the mouse retina, and identified the prefoldin-2 (PFDN2) gene as the candidate upstream modulator of Sncg expression. Our immunohistochemical analyses revealed similar expression patterns in both mouse and human healthy retinae, with PFDN2 colocalizing with SNCG in RGCs and their axons. In contrast, in retinae from glaucoma subjects, SNCG levels were significantly reduced, although PFDN2 levels were maintained. Using a novel flow cytometry-based RGC isolation method, we obtained viable populations of murine RGCs. Knocking down Pfdn2 expression in primary murine RGCs significantly reduced Sncg expression, confirming that Pfdn2 regulates Sncg expression in murine RGCs. Gene Ontology analysis indicated shared mitochondrial function associated with Sncg and Pfdn2. These data solidify the relationship between Sncg and Pfdn2 in RGCs, and provide a novel mechanism for maintaining RGC health. © 2015 FEBS.

  6. N-acetylphytosphingosine enhances the radiosensitivity of tumor cells by increasing apoptosis

    International Nuclear Information System (INIS)

    Han, Y.; Kim, Y.; Yun, Y.; Jeon, S.; Kim, K.; Song, J.; Hong, S.H.; Park, C.

    2005-01-01

    Ceramides are well-known second messengers which mediate apoptosis, proliferation, differentiation in mammalian cells, but the physiological roles of phytosphingosines are poorly understood. We hypothesized that one of the phytosphingosine derivatives, N-acetylphytosphingosine (NAPS) can induce apoptosis in human leukemia Jurkat cell line and increase apoptosis in irradiated MDA-MB-231 cells. We first examined the effect of NAPS on apoptosis of Jurkat cells. NAPS had a more rapid and stronger apoptotic effect than C 2 -ceramide in Jurkat cells and significant increase of apoptosis was observed at 3 h after treatment. In contrast, the apoptosis induced by C2-ceramide was observed only after 16 h of treatment. NAPS induced apoptosis was mediated by caspase 3 and 8 activation and inhibited by z-VAD-fmk. Ceramide plays a pivotal role in radiation induced apoptosis. We postulated that exogenous treatment of NAPS sensitizes tumor cells to ionizing radiation, since NAPS might be used as a more effective alternative to C2-ceramide. As expected, NAPS decreased clonogenic survival of irradiated MDA-MB-231 cells dose dependently, and apoptosis of irradiated cells in the presence of NAPS was increased through the caspase activation. Taken together, NAPS is an effective apoptosis-inducing agent, which can be readily synthesized from yeast sources, and is a potent alternative to ceramide for the further study of ceramide associated signaling and the development of radiosensitizing agent. (orig.)

  7. Linalool Induces Cell Cycle Arrest and Apoptosis in Leukemia Cells and Cervical Cancer Cells through CDKIs

    Directory of Open Access Journals (Sweden)

    Mei-Yin Chang

    2015-11-01

    Full Text Available Plantaginaceae, a popular traditional Chinese medicine, has long been used for treating various diseases from common cold to cancer. Linalool is one of the biologically active compounds that can be isolated from Plantaginaceae. Most of the commonly used cytotoxic anticancer drugs have been shown to induce apoptosis in susceptible tumor cells. However, the signaling pathway for apoptosis remains undefined. In this study, the cytotoxic effect of linalool on human cancer cell lines was investigated. Water-soluble tetrazolium salts (WST-1 based colorimetric cellular cytotoxicity assay, was used to test the cytotoxic ability of linalool against U937 and HeLa cells, and flow cytometry (FCM and genechip analysis were used to investigate the possible mechanism of apoptosis. These results demonstrated that linalool exhibited a good cytotoxic effect on U937 and HeLa cells, with the IC50 value of 2.59 and 11.02 μM, respectively, compared with 5-FU with values of 4.86 and 12.31 μM, respectively. After treating U937 cells with linalool for 6 h, we found an increased sub-G1 peak and a dose-dependent phenomenon, whereby these cells were arrested at the G0/G1 phase. Furthermore, by using genechip analysis, we observed that linalool can promote p53, p21, p27, p16, and p18 gene expression. Therefore, this study verified that linalool can arrest the cell cycle of U937 cells at the G0/G1 phase and can arrest the cell cycle of HeLa cells at the G2/M phase. Its mechanism facilitates the expression of the cyclin-dependent kinases inhibitors (CDKIs p53, p21, p27, p16, and p18, as well as the non-expression of cyclin-dependent kinases (CDKs activity.

  8. Fractalkine Signaling Regulates Macrophage Recruitment into the Cochlea and Promotes the Survival of Spiral Ganglion Neurons after Selective Hair Cell Lesion.

    Science.gov (United States)

    Kaur, Tejbeer; Zamani, Darius; Tong, Ling; Rubel, Edwin W; Ohlemiller, Kevin K; Hirose, Keiko; Warchol, Mark E

    2015-11-11

    Macrophages are recruited into the cochlea in response to injury caused by acoustic trauma or ototoxicity, but the nature of the interaction between macrophages and the sensory structures of the inner ear remains unclear. The present study examined the role of fractalkine signaling in regulating the injury-evoked behavior of macrophages following the selective ablation of cochlear hair cells. We used a novel transgenic mouse model in which the human diphtheria toxin receptor (huDTR) is selectively expressed under the control of Pou4f3, a hair cell-specific transcription factor. Administration of diphtheria toxin (DT) to these mice resulted in nearly complete ablation of cochlear hair cells, with no evident pathology among supporting cells, spiral ganglion neurons, or cells of the cochlear lateral wall. Hair cell death led to an increase in macrophages associated with the sensory epithelium of the cochlea. Their numbers peaked at 14 days after DT and then declined at later survival times. Increased macrophages were also observed within the spiral ganglion, but their numbers remained elevated for (at least) 56 d after DT. To investigate the role of fractalkine signaling in macrophage recruitment, we crossed huDTR mice to a mouse line that lacks expression of the fractalkine receptor (CX3CR1). Disruption of fractalkine signaling reduced macrophage recruitment into both the sensory epithelium and spiral ganglion and also resulted in diminished survival of spiral ganglion neurons after hair cell death. Our results suggest a fractalkine-mediated interaction between macrophages and the neurons of the cochlea. It is known that damage to the inner ear leads to recruitment of inflammatory cells (macrophages), but the chemical signals that initiate this recruitment and the functions of macrophages in the damaged ear are unclear. Here we show that fractalkine signaling regulates macrophage recruitment into the cochlea and also promotes the survival of cochlear afferents after

  9. Inhibition of apoptosis in T cells expressing human T cell leukemia virus type I Tax

    NARCIS (Netherlands)

    Copeland, K. F.; Haaksma, A. G.; Goudsmit, J.; Krammer, P. H.; Heeney, J. L.

    1994-01-01

    This study set out to determine whether T cell dysfunction associated with HTLV-I led to increased sensitivity of infected cells to apoptosis or, owing to their potential to develop ATL, if infected cells would become resistant to this process. To test this hypothesis we utilized the monoclonal

  10. Cell apoptosis, autophagy and necroptosis in osteosarcoma treatment

    Science.gov (United States)

    Li, Dongqi; Li, Huiling; Ren, Mingyan; Liao, Yedan; Yu, Shunling; Chen, Yanjin; Yang, Yihao; Zhang, Ya

    2016-01-01

    Osteosarcoma is the most common primary bone tumor in children and adolescents. Although combined therapy including surgery and multi-agent chemotherapy have resulted in great improvements in the overall survival of patients, chemoresistance remains an obstacle for the treatment of osteosarcoma. Molecular targets or effective agents that are actively involved in cell death including apoptosis, autophagy and necroptosis have been studied. We summarized how these agents (novel compounds, miRNAs, or proteins) regulate apoptotic, autophagic and necroptotic pathways; and discussed the current knowledge on the role of these new agents in chemotherapy resistance in osteosarcoma. PMID:27007056

  11. Re: Engineered Nanoparticles Induce Cell Apoptosis: Potential for Cancer Therapy

    Directory of Open Access Journals (Sweden)

    Fehmi Narter

    2016-09-01

    Full Text Available Engineered nanoparticles (ENPs have been widely applied in industry, biology and medicine recently (i.e. clothes, sunscreens, cosmetics, foods, diagnostic medicine, imaging and drug delivery. There are many kinds of manufactured nanomaterial products including TiO2, ZnO, CeO2, Fe2O3, and CuO (as metal oxide nanoparticles as well as gold, silver, platinum and palladium (as metal nanoparticles, and other carbon-based ENP’s such as carbon nanotububes and quantum dots. ENPs with their sizes no larger than 100 nm are able to enter the human body and accumulate in organs and cause toxic effects. In many researches, ENP effects on the cancer cells of different organs with related cell apoptosis were noted (AgNP, nano-Cr2O3, Au-Fe2O3 NPs, nano-TiO2, nano-HAP, nano-Se, MoO3 nanoplate, Realgar nanoparticles. ENPs, with their unique properties, such as surface charge, particle size, composition and surface modification with tissue recognition ligands or antibodies, has been increasingly explored as a tool to carry small molecular weight drugs as well as macromolecules for cancer therapy, thus generating the new concept “nanocarrier”. Direct induction of cell apoptosis by ENPs provides an opportunity for cancer treatment. In the century of nanomedicine that depends on development of the nanotechnology, ENPs have a great potential for application in cancer treatment with minimal side effects.

  12. Is radiation-induced cell death in mouse testis apoptosis?

    International Nuclear Information System (INIS)

    Hasegawa, Masatoshi; Wilson, Gene; Yun Zhang; Russell, Lonnie D.; Meistrich, Marvin L.

    1996-01-01

    Purpose: Radiation-induced death of spermatogonia and other germ cells in the testis has been claimed to be by an apoptotic mechanism, but these processes have been incompletely characterized. We investigated irradiated mouse testis by multiple techniques to determine whether the mode of cell death of spermatogonia can be classified as apoptosis. Materials and Methods: Adult male C57BL/6 and p53 knockout mice were irradiated with single doses of 0.5, 2.5 or 5.0 Gy. Four, 6, 8, 12, 18 or 24 hours after irradiation, testes were fixed in Bouin's solution or in 10% formalin. Slides were stained with hematoxylin and eosin or TdT-mediated dUTP-biotin nick end labeling (TUNEL). Some testes were perfusion-fixed with 5% glutaraldehyde for electron microscopy. Gel electrophoresis of DNA was also performed to identify DNA fragmentation. The number of sperm heads was counted 29 days after irradiation to evaluate the effect of radiation on the eventual survival of the differentiated spermatogonia. Results: The earliest sign of histological damage was an increase in the numbers of abnormal spermatogonia in the seminiferous tubules, particularly in stage I-VI of the seminiferous epithelial cycle. The numbers of abnormal spermatogonia began to increase at 6 hours, reached a peak 12 hours after irradiation, and then declined. The total number of spermatogonia began to decrease at 12 hours after irradiation, resulting in a 60% decline in sperm produced 29 days after 0.5 Gy. Although changes were greatest following 5.0 Gy irradiation, even 0.5 Gy induced marked changes. However, these changes were not induced in p53 knockout mice. By both light and electron microscopy, spermatogonia showed some condensation of nuclear chromatin, but margination of chromatin with clear delineation and nuclear fragmentation was rare. Many of the abnormal spermatogonia showed a positive TUNEL reaction, which was also at a maximum at 12 hours after irradiation. In addition, some TUNEL-positive and

  13. VMP1 related autophagy and apoptosis in colorectal cancer cells: VMP1 regulates cell death

    Energy Technology Data Exchange (ETDEWEB)

    Qian, Qinyi [Department of Ultrasonograph, Changshu No. 2 People’s Hospital, Changshu (China); Zhou, Hao; Chen, Yan [Department of General Surgery, The First Affiliated Hospital of Soochow University, Suzhou (China); Shen, Chenglong [Department of General Surgery, Changshu No. 2 People’s Hospital, Changshu (China); He, Songbing; Zhao, Hua; Wang, Liang [Department of General Surgery, The First Affiliated Hospital of Soochow University, Suzhou (China); Wan, Daiwei, E-mail: 372710369@qq.com [Department of Hepatobiliary Surgery, The First Affiliated Hospital of Sun Yat-sen University, Guangzhou (China); Gu, Wen, E-mail: 505339704@qq.com [Department of General Surgery, The First Affiliated Hospital of Soochow University, Suzhou (China)

    2014-01-17

    Highlights: •This research confirmed VMP1 as a regulator of autophagy in colorectal cancer cell lines. •We proved the pro-survival role of VMP1-mediated autophagy in colorectal cancer cell lines. •We found the interaction between VMP1 and BECLIN1 also existing in colorectal cancer cell lines. -- Abstract: Vacuole membrane protein 1 (VMP1) is an autophagy-related protein and identified as a key regulator of autophagy in recent years. In pancreatic cell lines, VMP1-dependent autophagy has been linked to positive regulation of apoptosis. However, there are no published reports on the role of VMP1 in autophagy and apoptosis in colorectal cancers. Therefore, to address this gap of knowledge, we decided to interrogate regulation of autophagy and apoptosis by VMP1. We have studied the induction of autophagy by starvation and rapamycin treatment in colorectal cell lines using electron microscopy, immunofluorescence, and immunoblotting. We found that starvation-induced autophagy correlated with an increase in VMP1 expression, that VMP1 interacted with BECLIN1, and that siRNA mediated down-regulation of VMP1-reduced autophagy. Next, we examined the relationship between VMP1-dependent autophagy and apoptosis and found that VMP1 down-regulation sensitizes cells to apoptosis and that agents that induce apoptosis down-regulate VMP1. In conclusion, similar to its reported role in other cell types, VMP1 is an important regulator of autophagy in colorectal cell lines. However, in contrast to its role in pancreatic cell lines, in colorectal cancer cells, VMP1-dependent autophagy appears to be pro-survival rather than pro-cell death.

  14. Docosahexaenoic acid induces apoptosis in primary chronic lymphocytic leukemia cells

    Directory of Open Access Journals (Sweden)

    Romain Guièze

    2015-12-01

    Full Text Available Chronic lymphocytic leukemia is an indolent disorder with an increased infectious risk remaining one of the main causes of death. Development of therapies with higher safety profile is thus a challenging issue. Docosahexaenoic acid (DHA, 22:6 is an omega-3 fatty acid, a natural compound of normal cells, and has been shown to display antitumor potency in cancer. We evaluated the potential in vitro effect of DHA in primary CLL cells. DHA induces high level of in vitro apoptosis compared to oleic acid in a dose-dependent and time-dependent manner. Estimation of IC50 was only of 4.813 μM, which appears lower than those reported in solid cancers. DHA is highly active on CLL cells in vitro. This observation provides a rationale for further studies aiming to understand its mechanisms of action and its potent in vivo activity.

  15. New cancer cells apoptosis agents: Fluorinated aza-heterocycles

    Science.gov (United States)

    Prima, D. O.; Baev, D. S.; Vorontsova, E. V.; Frolova, T. S.; Bagryanskaya, I. Yu.; Slizhov, Yu. G.; Tolstikova, T. G.; Makarov, A. Yu.; Zibarev, A. V.

    2017-09-01

    Fluorinated benzo-fused 1,3-diazoles, 1,2,3-triazoles, 1,2,5-thia/selenadiazoles and 1,4-diazines were synthesized and tried for cytotoxicity towards the Hep2 (laryngeal epidermoid carcinoma) cells. The diazoles, triazoles and selenadiazoles were cytotoxic with IC50 = 2.2-26.4 µM and induced the cells apoptosis at concentrations C = 1-25 µM. At the same time, they were nontoxic towards normal cells. Due to this, these scaffolds were used in the computer-aided molecular design of new antitumor agents. Particularly, novel 1,2,3-triazole and 1,3-diazole derivatives for the binding site of the PAS domain of the transcription factor HIF were designed and some of them synthesized for further study. Overall, new anticancer agents featuring apoptotic activity are suggested.

  16. Zebrafish diras1 Promoted Neurite Outgrowth in Neuro-2a Cells and Maintained Trigeminal Ganglion Neurons In Vivo via Rac1-Dependent Pathway.

    Science.gov (United States)

    Yeh, Chi-Wei; Hsu, Li-Sung

    2016-12-01

    The small GTPase Ras superfamily regulates several neuronal functions including neurite outgrowth and neuron proliferation. In this study, zebrafish diras1a and diras1b were identified and were found to be mainly expressed in the central nervous system and dorsal neuron ganglion. Overexpression of green fluorescent protein (GFP)-diras1a or GFP-diras1b triggered neurite outgrowth of Neuro-2a cells. The wild types, but not the C terminus truncated forms, of diras1a and diras1b elevated the protein level of Ras-related C3 botulinum toxin substrate 1 (Rac1) and downregulated Ras homologous member A (RhoA) expression. Glutathione S-transferase (GST) pull-down assay also revealed that diras1a and diras1b enhanced Rac1 activity. Interfering with Rac1, Pak1, or cyclin-dependent kinase 5 (CDK5) activity or with the Arp2/3 inhibitor prevented diras1a and diras1b from mediating the neurite outgrowth effects. In the zebrafish model, knockdown of diras1a and/or diras1b by morpholino antisense oligonucleotides not only reduced axon guidance but also caused the loss of trigeminal ganglion without affecting the precursor markers, such as ngn1 and neuroD. Co-injection with messenger RNA (mRNA) derived from mouse diras1 or constitutively active human Rac1 restored the population of trigeminal ganglion. In conclusion, we provided preliminary evidence that diras1 is involved in neurite outgrowth and maintains the number of trigeminal ganglions through the Rac1-dependent pathway.

  17. Demethoxycurcumin Retards Cell Growth and Induces Apoptosis in Human Brain Malignant Glioma GBM 8401 Cells

    Directory of Open Access Journals (Sweden)

    Tzuu-Yuan Huang

    2012-01-01

    Full Text Available Demethoxycurcumin (DMC; a curcumin-related demethoxy compound has been recently shown to display antioxidant and antitumor activities. It has also produced a potent chemopreventive action against cancer. In the present study, the antiproliferation (using the MTT assay, DMC was found to have cytotoxic activities against GBM 8401 cell with IC50 values at 22.71 μM and induced apoptosis effects of DMC have been investigated in human brain malignant glioma GBM 8401 cells. We have studied the mitochondrial membrane potential (MMP, DNA fragmentation, caspase activation, and NF-κB transcriptional factor activity. By these approaches, our results indicated that DMC has produced an inhibition of cell proliferation as well as the activation of apoptosis in GBM 8401 cells. Both effects were observed to increase in proportion with the dosage of DMC treatment, and the apoptosis was induced by DMC in human brain malignant glioma GBM 8401 cells via mitochondria- and caspase-dependent pathways.

  18. Expression of inducible nitric oxide synthase in trigeminal ganglion cells during culture

    DEFF Research Database (Denmark)

    Jansen-Olesen, Inger; Zhou, MingFang; Zinck, Tina Jovanovic

    2005-01-01

    RNA and protein could be detected. The data suggest that iNOS expression may be a molecular mechanism mediating the adaptive response of trigeminal ganglia cells to the serum free stressful stimulus the culture environment provides. It may act as a cellular signalling molecule that is expressed after cell......Nitric oxide (NO) is an important signalling molecule that has been suggested to be a key molecule for induction and maintenance of migraine attacks based on clinical studies, animal experimental studies and the expression of nitric oxide synthase (NOS) immunoreactivity within the trigeminovascular......, reverse transcriptase polymerase chain reaction (RT-PCR) and Western blotting. In trigeminal ganglia cells not subjected to culture, endothelial (e) and neuronal (n) but not inducible (i) NOS mRNA and protein were detected. Culture of rat neurones resulted in a rapid axonal outgrowth of NOS positive...

  19. Effects of P-Glycoprotein and Its Inhibitors on Apoptosis in K562 Cells

    Directory of Open Access Journals (Sweden)

    Yaqiong Zu

    2014-08-01

    Full Text Available P-glycoprotein (P-gp is a major factor in multidrug resistance (MDR which is a serious obstacle in chemotherapy. P-gp has also been implicated in causing apoptosis of tumor cells, which was shown to be another important mechanism of MDR recently. To study the influence of P-gp in tumor cell apoptosis, K562/A cells (P-gp+ and K562/S cells (P-gp− were subjected to doxorubicin (Dox, serum withdrawal, or independent co-incubation with multiple P-gp inhibitors, including valspodar (PSC833, verapamil (Ver and H108 to induce apoptosis. Apoptosis was simultaneously detected by apoptotic rate, cell cycle by flow cytometry and cysteine aspartic acid-specific protease 3 (caspase 3 activity by immunoassay. Cytotoxicity and apoptosis induced by PSC833 were evaluated through an MTT method and apoptosis rate, and cell cycle combined with caspase 3 activity, respectively. The results show that K562/A cells are more resistant to apoptosis and cell cycle arrest than K562/S cells after treatment with Dox or serum deprivation. The apoptosis of K562/A cells increased after co-incubation with each of the inhibitors of P-gp. P-gp inhibitors also enhanced cell cycle arrest in K562/A cell. PSC833 most strikingly decreased viability and led to apoptosis and S phase arrest of cell cycle in K562/A cells. Our study demonstrates that P-gp inhibits the apoptosis of tumor cells in addition to participating in the efflux of intracellular chemotherapy drugs. The results of the caspase 3 activity assay also suggest that the role of P-gp in apoptosis avoidance is caspase-related.

  20. Recombinant Vaccinia Viruses Coding Transgenes of Apoptosis-Inducing Proteins Enhance Apoptosis But Not Immunogenicity of Infected Tumor Cells

    Science.gov (United States)

    Tkachenko, Anastasiya; Richter, Vladimir

    2017-01-01

    Genetic modifications of the oncolytic vaccinia virus (VV) improve selective tumor cell infection and death, as well as activation of antitumor immunity. We have engineered a double recombinant VV, coding human GM-CSF, and apoptosis-inducing protein apoptin (VV-GMCSF-Apo) for comparing with the earlier constructed double recombinant VV-GMCSF-Lact, coding another apoptosis-inducing protein, lactaptin, which activated different cell death pathways than apoptin. We showed that both these recombinant VVs more considerably activated a set of critical apoptosis markers in infected cells than the recombinant VV coding GM-CSF alone (VV-GMCSF-dGF): these were phosphatidylserine externalization, caspase-3 and caspase-7 activation, DNA fragmentation, and upregulation of proapoptotic protein BAX. However, only VV-GMCSF-Lact efficiently decreased the mitochondrial membrane potential of infected cancer cells. Investigating immunogenic cell death markers in cancer cells infected with recombinant VVs, we demonstrated that all tested recombinant VVs were efficient in calreticulin and HSP70 externalization, decrease of cellular HMGB1, and ATP secretion. The comparison of antitumor activity against advanced MDA-MB-231 tumor revealed that both recombinants VV-GMCSF-Lact and VV-GMCSF-Apo efficiently delay tumor growth. Our results demonstrate that the composition of GM-CSF and apoptosis-inducing proteins in the VV genome is very efficient tool for specific killing of cancer cells and for activation of antitumor immunity. PMID:28951871

  1. Oxidative stress in NSC-741909-induced apoptosis of cancer cells

    Directory of Open Access Journals (Sweden)

    Huang Peng

    2010-04-01

    Full Text Available Abstract Background NSC-741909 is a novel anticancer agent that can effectively suppress the growth of several cell lines derived from lung, colon, breast, ovarian, and kidney cancers. We recently showed that NSC-741909-induced antitumor activity is associated with sustained Jun N-terminal kinase (JNK activation, resulting from suppression of JNK dephosphorylation associated with decreased protein levels of MAPK phosphatase-1. However, the mechanisms of NSC-741909-induced antitumor activity remain unclear. Because JNK is frequently activated by oxidative stress in cells, we hypothesized that reactive oxygen species (ROS may be involved in the suppression of JNK dephosphorylation and the cytotoxicity of NSC-741909. Methods The generation of ROS was measured by using the cell-permeable nonfluorescent compound H2DCF-DA and flow cytometry analysis. Cell viability was determined by sulforhodamine B assay. Western blot analysis, immunofluorescent staining and flow cytometry assays were used to determine apoptosis and molecular changes induced by NSC-741909. Results Treatment with NSC-741909 induced robust ROS generation and marked MAPK phosphatase-1 and -7 clustering in NSC-741909-sensitive, but not resistant cell lines, in a dose- and time-dependent manner. The generation of ROS was detectable as early as 30 min and ROS levels were as high as 6- to 8-fold above basal levels after treatment. Moreover, the NSC-741909-induced ROS generation could be blocked by pretreatment with antioxidants, such as nordihydroguaiaretic acid, aesculetin, baicalein, and caffeic acid, which in turn, inhibited the NSC-741909-induced JNK activation and apoptosis. Conclusion Our results demonstrate that the increased ROS production was associated with NSC-741909-induced antitumor activity and that ROS generation and subsequent JNK activation is one of the primary mechanisms of NSC-741909-mediated antitumor cell activity.

  2. [Ca2+]i in exterior of cells effected on apoptosis of HL-60 cells induced by irradiation

    International Nuclear Information System (INIS)

    He Ziyi; Meng Qingyong

    2005-01-01

    Objective: To investigate of the different [Ca 2+ ]i in exterior of cells promotion function on apoptosis of HL-60 cells induced by irradiation. Methods: To put ration dose 32 P and different [Ca 2+ ]i into culture of HL-60 and measure the apoptosis rate with FCM after 24 and 48 hours. Result: Apoptosis rate increased with the increase of [Ca 2+ ]i which shows an obvious function to promote apoptosis, r 24 =0.9001 (P=0.0145); r48=0.9343 (P=0.0063). Conclusion: [Ca 2+ ]i in exterior of cells has a obvious function in promoteing apoptosis induced by irradiation. (authors)

  3. Macular ganglion cell complex and retinal nerve fiber layer comparison in different stages of age-related macular degeneration.

    Science.gov (United States)

    Zucchiatti, Ilaria; Parodi, Maurizio Battaglia; Pierro, Luisa; Cicinelli, Maria Vittoria; Gagliardi, Marco; Castellino, Niccolò; Bandello, Francesco

    2015-09-01

    To employ optical coherence tomography (OCT) to analyze the morphologic changes in the inner retina in different categories of age-related macular degeneration (AMD). Observational cross-sectional study. Single-center study. Inclusion criteria were age over 50, diagnosis of Age-Related Eye Disease Study (AREDS) category 2 and 3, naïve neovascular AMD, and atrophic AMD. Healthy patients of similar age acted as a control group. Primary outcome measures were the changes in ganglion cell complex (GCC) and retinal nerve fiber layer (RNFL). Secondary outcomes included modifications of rim area and cup-to-disc ratio. One hundred and thirty eyes of 130 patients were recruited: 26 eyes for AREDS category 2, 26 for AREDS category 3, 26 for neovascular AMD, 26 with atrophic AMD, and 26 controls. Mean peripapillary RNFL thickness was significantly lower in neovascular AMD, compared to controls (P = .004); peripapillary RNFL did not significantly vary among AREDS category 2 and 3 and atrophic AMD groups, compared to controls. Mean GCC thickness was higher in the control group, becoming progressively thinner up to neovascular and atrophic AMD groups (P < .0001). Rim area was significantly thinner in the neovascular AMD group compared with controls (P = .047); cup-to-disc ratio was higher in the neovascular AMD group compared with the control group (P = .047). This study demonstrates that eyes with neovascular AMD display reduced RNFL and GCC thickness. RNFL is partially spared in atrophic advanced AMD. The identification of alteration in RNFL and GCC thickness may reveal useful for future therapeutic implications. Copyright © 2015 Elsevier Inc. All rights reserved.

  4. Studies of Scleral Biomechanical Behavior Related to Susceptibility for Retinal Ganglion Cell Loss in Experimental Mouse Glaucoma

    Science.gov (United States)

    Nguyen, Cathy; Cone, Frances E.; Nguyen, Thao D.; Coudrillier, Baptiste; Pease, Mary E.; Steinhart, Matthew R.; Oglesby, Ericka N.; Jefferys, Joan L.; Quigley, Harry A.

    2013-01-01

    Purpose. To study anatomical changes and mechanical behavior of the sclera in mice with experimental glaucoma by comparing CD1 to B6 mice. Methods. Chronic experimental glaucoma for 6 weeks was produced in 2- to 4-month-old CD1 (43 eyes) and B6 mice (42 eyes) using polystyrene bead injection into the anterior chamber with 126 control CD1 and 128 control B6 eyes. Intraocular pressure (IOP) measurements were made with the TonoLab at baseline and after bead injection. Axial length and scleral thickness were measured after sacrifice in the CD1 and B6 animals and compared to length data from 78 eyes of DBA/2J mice. Inflation testing of posterior sclera was conducted, and circumferential and meridional strain components were determined from the displacement response. Results. Experimental glaucoma led to increases in axial length and width by comparison to fellow eyes (6% in CD1 and 10% in B6; all P glaucoma, the remainder of the sclera uniformly thinned in CD1, but thickened in B6. Peripapillary sclera in CD1 controls had significantly greater temporal meridional strain than B6 and had differences in the ratios of meridional to effective circumferential strain from B6 mice. In both CD1 and B6 mice, exposure to chronic IOP elevation resulted in stiffer pressure–strain responses for both the effective circumferential and meridional strains (multivariable regression model, P = 0.01–0.03). Conclusions. Longer eyes, greater scleral strain in some directions at baseline, and generalized scleral thinning after glaucoma were characteristic of CD1 mice that have greater tendency to retinal ganglion cell damage than B6 mice. Increased scleral stiffness after glaucoma exposure in mice mimics findings in monkey and human glaucoma eyes. PMID:23404116

  5. Protection of neurons in the retinal ganglion cell layer against excitotoxicity by the N-acylethanolamine, N-linoleoylethanolamine

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    Duncan RS

    2011-04-01

    Full Text Available R. Scott Duncan1,*, Hua Xin1,*, Daryl L Goad1, Kent D Chapman2,3, Peter Koulen1,31Vision Research Center and Departments of Ophthalmology and Basic Medical Science, School of Medicine, University of Missouri, Kansas City, MO, USA; 2Department of Biological Sciences, University of North Texas, Denton, TX, USA; 3Center for Plant Lipid Research, University of North Texas, Denton, TX, USA *Authors contributed equallyAbstract: Retinal ganglion cell (RGC death is a hallmark of neurodegenerative diseases and disease processes of the eye, including glaucoma. The protection of RGCs has been an important strategy for combating glaucoma, but little clinical success has been reported to date. One pathophysiological consequence of glaucoma is excessive extracellular glutamate subsequently leading to excitotoxicity in the retina. Endocannabinoids, such as the N-acylethanolamine (NAE, arachidonylethanolamine (NAE 20:4, exhibit neuroprotective properties in some models of neurodegenerative disease. The majority of NAEs, however, are not cannabinoids, and their physiological function is not clear. Here, we determined whether the noncannabinoid NAE, linoleoylethanolamine (NAE18:2, protects neurons in the RGC layer against glutamate excitotoxicity in ex-vivo retina cultures. Using a terminal deoxynucleotidyl transferase-mediated dUTP (2´-deoxyuridine 5´-triphosphate nick-end labeling (TUNEL assay, we determined that NAE18:2 reduces the number of apoptotic RGC layer neurons in response to glutamate and conclude that NAE18:2 is a neuroprotective compound with potential for treating glaucomatous retinopathy.Keywords: neuroprotection, glutamate, calcium signaling, immunocytochemistry, eye, vision, glaucoma.

  6. Macular Ganglion Cell Inner Plexiform Layer Thickness in Glaucomatous Eyes with Localized Retinal Nerve Fiber Layer Defects.

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    Chunwei Zhang

    Full Text Available To investigate macular ganglion cell-inner plexiform layer (mGCIPL thickness in glaucomatous eyes with visible localized retinal nerve fiber layer (RNFL defects on stereophotographs.112 healthy and 149 glaucomatous eyes from the Diagnostic Innovations in Glaucoma Study (DIGS and the African Descent and Glaucoma Evaluation Study (ADAGES subjects had standard automated perimetry (SAP, optical coherence tomography (OCT imaging of the macula and optic nerve head, and stereoscopic optic disc photography. Masked observers identified localized RNFL defects by grading of stereophotographs.47 eyes had visible localized RNFL defects on stereophotographs. Eyes with visible localized RNFL defects had significantly thinner mGCIPL thickness compared to healthy eyes (68.3 ± 11.4 μm versus 79.2 ± 6.6 μm respectively, P<0.001 and similar mGCIPL thickness to glaucomatous eyes without localized RNFL defects (68.6 ± 11.2 μm, P = 1.000. The average mGCIPL thickness in eyes with RNFL defects was 14% less than similarly aged healthy controls. For 29 eyes with a visible RNFL defect in just one hemiretina (superior or inferior mGCIPL was thinnest in the same hemiretina in 26 eyes (90%. Eyes with inferior-temporal RNFL defects also had significantly thinner inferior-temporal mGCIPL (P<0.001 and inferior mGCIPL (P = 0.030 compared to glaucomatous eyes without a visible RNFL defect.The current study indicates that presence of a localized RNFL defect is likely to indicate significant macular damage, particularly in the region of the macular that topographically corresponds to the location of the RNFL defect.

  7. Endothelial cell energy metabolism, proliferation, and apoptosis in pulmonary hypertension.

    Science.gov (United States)

    Xu, Weiling; Erzurum, Serpil C

    2011-01-01

    Pulmonary arterial hypertension (PAH) is a fatal disease characterized by impaired regulation of pulmonary hemodynamics and excessive growth and dysfunction of the endothelial cells that line the arteries in PAH lungs. Establishment of methods for culture of pulmonary artery endothelial cells from PAH lungs has provided the groundwork for mechanistic translational studies that confirm and extend findings from model systems and spontaneous pulmonary hypertension in animals. Endothelial cell hyperproliferation, survival, and alterations of biochemical-metabolic pathways are the unifying endothelial pathobiology of the disease. The hyperproliferative and apoptosis-resistant phenotype of PAH endothelial cells is dependent upon the activation of signal transducer and activator of transcription (STAT) 3, a fundamental regulator of cell survival and angiogenesis. Animal models of PAH, patients with PAH, and human PAH endothelial cells produce low nitric oxide (NO). In association with the low level of NO, endothelial cells have reduced mitochondrial numbers and cellular respiration, which is associated with more than a threefold increase in glycolysis for energy production. The shift to glycolysis is related to low levels of NO and likely to the pathologic expression of the prosurvival and proangiogenic signal transducer, hypoxia-inducible factor (HIF)-1, and the reduced mitochondrial antioxidant manganese superoxide dismutase (MnSOD). In this article, we review the phenotypic changes of the endothelium in PAH and the biochemical mechanisms accounting for the proliferative, glycolytic, and strongly proangiogenic phenotype of these dysfunctional cells, which consequently foster the panvascular progressive pulmonary remodeling in PAH. © 2011 American Physiological Society.

  8. [Effects of cucurmosin on the cell proliferation and apoptosis in human pancreatic PANC-1 cells].

    Science.gov (United States)

    Xu, Chun-Sen; Huang, He-Guang; Chen, Ming-Huang

    2012-02-01

    To observe the effects of cucurmosin (CUS) on the cell proliferation and apoptosis in pancreatic PANC-1 cells. The inhibition of CUS on the PANC-1 cell growth was observed using MTT assay. The inhibition ratio of CUS on the pancreatic orthotopic transplantation was in vivo observed in the NOD/SCID mouse model. The changes of microstructure of the apoptosis-inducing effect of CUS on PANC-1 was observed under electron microscope. The cell cycle and apoptosis after CUS intervention was detected using flow cytometry. The Caspase-3 activity after CUS treatment was detected using enzyme linked immunospecific assay (ELISA). Treatment with CUS at the dose of 0.125, 0.25, and 0.5 mg/kg inhibited the growth of pancreatic carcinoma PANC-1 xenografs with the ratio of 45.2%, 50.0%, and 59.7%, respectively (P PANC-1 cells in a dose-dependent maner. Being exposed to 40.0 microg/mL of the CUS for 24, 48, and 72 h, the percentage of G0/ G1 phase cells was 56.60% +/- 6.65%, 67.83% +/- 6.76%, and 77.00% +/- 6.73%, respectively (P PANC-1 cells in the G0/G1 phase of the cell cycle in a time-dependent maner. CUS significantly inhibited the growth of PANC-1 cells possibly through the G0/G1 cell cycle arrest and apoptosis.

  9. Transgenic inhibition of astroglial NF-κB protects from optic nerve damage and retinal ganglion cell loss in experimental optic neuritis

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    Brambilla Roberta

    2012-09-01

    Full Text Available Abstract Background Optic neuritis is an acute, demyelinating neuropathy of the optic nerve often representing the first appreciable symptom of multiple sclerosis. Wallerian degeneration of irreversibly damaged optic nerve axons leads to death of retinal ganglion cells, which is the cause of permanent visual impairment. Although the specific mechanisms responsible for triggering these events are unknown, it has been suggested that a key pathological factor is the activation of immune-inflammatory processes secondary to leukocyte infiltration. However, to date, there is no conclusive evidence to support such a causal role for infiltrating peripheral immune cells in the etiopathology of optic neuritis. Methods To dissect the contribution of the peripheral immune-inflammatory response versus the CNS-specific inflammatory response in the development of optic neuritis, we analyzed optic nerve and retinal ganglion cells pathology in wild-type and GFAP-IκBα-dn transgenic mice, where NF-κB is selectively inactivated in astrocytes, following induction of EAE. Results We found that, in wild-type mice, axonal demyelination in the optic nerve occurred as early as 8 days post induction of EAE, prior to the earliest signs of leukocyte infiltration (20 days post induction. On the contrary, GFAP-IκBα-dn mice were significantly protected and showed a nearly complete prevention of axonal demyelination, as well as a drastic attenuation in retinal ganglion cell death. This correlated with a decrease in the expression of pro-inflammatory cytokines, chemokines, adhesion molecules, as well as a prevention of NAD(PH oxidase subunit upregulation. Conclusions Our results provide evidence that astrocytes, not infiltrating immune cells, play a key role in the development of optic neuritis and that astrocyte-mediated neurotoxicity is dependent on activation of a transcriptional program regulated by NF-κB. Hence, interventions targeting the NF-κB transcription

  10. Studying arsenic trioxide-induced apoptosis of Colo-16 cells with ...

    African Journals Online (AJOL)

    Jane

    2011-10-05

    Oct 5, 2011 ... induced apoptosis at the single cell level. Key words: Two-photon laser scanning microscopy, confocal laser scanning microscopy, human skin squamous carcinoma cells (Colo-16 cells), arsenic trioxide, apoptosis. INTRODUCTION. Although arsenic is poisonous and chronic arsenic exposure from ...

  11. Effect of dicycloplatin, a novel platinum chemotherapeutical drug, on inhibiting cell growth and inducing cell apoptosis.

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    Guang-quan Li

    Full Text Available Dicycloplatin, a new supramolecular platinum-based antitumor drug, has been approved by the State Food and Administration (SFDA of China. In this study, we investigated the anticancer activity of dicycloplatin in cancer cells and signaling pathways involved in dicycloplatin-induced apoptosis. Dicycloplatin inhibited the proliferation of cancer cells and increased the percentage of apoptosis in a concentration-dependent manner. Besides, some apoptosis related events were observed after treatment with dicycloplatin, including increase of reactive oxygen species (ROS, collapse of mitochondrial membrane potential (Δψm, release of cytochrome c from the mitochondria to the cytosol, upregulation of p53, which were accompanied by activation of caspase-9, caspase-3, caspase-8, and poly (ADP-ribose polymerase cleavage in a concentration-dependent manner. The role of apoptosis in dicycloplatin-mediated cell death was further confirmed by the concomitant treatment with caspase-8 or caspase-9 inhibitors, which inhibited apoptosis and PARP cleavage. Intracellular glutathione (GSH was also found to inhibit the cytotoxic effect of dicycloplatin. In conclusion, these findings suggest that dicycloplatin induces apoptosis through ROS stress-mediated death receptor pathway and mitochondrial pathway which is similar to carboplatin.

  12. Endonucleases induced TRAIL-insensitive apoptosis in ovarian carcinoma cells

    Energy Technology Data Exchange (ETDEWEB)

    Geel, Tessa M. [Department of Pathology and Medical Biology, Groningen University Institute for Drug Exploration (GUIDE), University Medical Center Groningen (UMCG), Hanzeplein 1, 9713 GZ, Groningen (Netherlands); Meiss, Gregor [Institute of Biochemistry, Justus-Liebig-University Giessen, D-35392 Giessen (Germany); Gun, Bernardina T. van der; Kroesen, Bart Jan; Leij, Lou F. de [Department of Pathology and Medical Biology, Groningen University Institute for Drug Exploration (GUIDE), University Medical Center Groningen (UMCG), Hanzeplein 1, 9713 GZ, Groningen (Netherlands); Zaremba, Mindaugas; Silanskas, Arunas [Institute of Biotechnology, Vilnius LT-02241 (Lithuania); Kokkinidis, Michael [IMBB/FORTH and University of Crete/Department of Biology, GR-71409 Heraklion/Crete (Greece); Pingoud, Alfred [Institute of Biochemistry, Justus-Liebig-University Giessen, D-35392 Giessen (Germany); Ruiters, Marcel H. [Department of Pathology and Medical Biology, Groningen University Institute for Drug Exploration (GUIDE), University Medical Center Groningen (UMCG), Hanzeplein 1, 9713 GZ, Groningen (Netherlands); Synvolux therapeutics, Groningen (Netherlands); McLaughlin, Pamela M. [Department of Pathology and Medical Biology, Groningen University Institute for Drug Exploration (GUIDE), University Medical Center Groningen (UMCG), Hanzeplein 1, 9713 GZ, Groningen (Netherlands); Rots, Marianne G., E-mail: m.g.rots@med.umcg.nl [Department of Pathology and Medical Biology, Groningen University Institute for Drug Exploration (GUIDE), University Medical Center Groningen (UMCG), Hanzeplein 1, 9713 GZ, Groningen (Netherlands)

    2009-09-10

    TRAIL induced apoptosis of tumor cells is currently entering phase II clinical settings, despite the fact that not all tumor types are sensitive to TRAIL. TRAIL resistance in ovarian carcinomas can be caused by a blockade upstream of the caspase 3 signaling cascade. We explored the ability of restriction endonucleases to directly digest DNA in vivo, thereby circumventing the caspase cascade. For this purpose, we delivered enzymatically active endonucleases via the cationic amphiphilic lipid SAINT-18{sup Registered-Sign }:DOPE to both TRAIL-sensitive and insensitive ovarian carcinoma cells (OVCAR and SKOV-3, respectively). Functional nuclear localization after delivery of various endonucleases (BfiI, PvuII and NucA) was indicated by confocal microscopy and genomic cleavage analysis. For PvuII, analysis of mitochondrial damage demonstrated extensive apoptosis both in SKOV-3 and OVCAR. This study clearly demonstrates that cellular delivery of restriction endonucleases holds promise to serve as a novel therapeutic tool for the treatment of resistant ovarian carcinomas.

  13. Effect of triamcinolone in keloids morphological changes and cell apoptosis

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    João Márcio Prazeres dos Santos

    Full Text Available OBJECTIVE:to assess the effects of injectable triamcinolone on keloid scars length, height and thickness, and on the number of cells undergoing apoptosis.METHODS:This study consists in a prospective, controlled, randomized, single-blinded clinical trial, conducted with fifteen patients with ear keloids divided into two groups: group 1 - seven patients undergoing keloid excisions, and group 2 - eight patients undergoing keloid excisions after three sessions of infiltration with one ml of Triamcinolone hexacetonide (20mg/ml with three week intervals between them and between the last session and surgery. The two groups were homogeneous regarding age, gender and evolution of the keloid scar. The keloid scars of patients in group 2 were measured for the length, height and thickness before triamcinolone injection and before surgery. A blinded observer performed morphological detailing and quantification of cells in hematoxylin-eosin-stained surgical specimens. An apoptotic index was created.RESULTS: The apoptotic index in group 1 was 56.82, and in group 2, 68.55, showing no significant difference as for apoptosis (p=0.0971. The reduction in keloid dimensions in Group 2 was 10.12% in length (p=0.6598, 11.94% in height (p=0.4981 and 15.62% in thickness (p=0.4027.CONCLUSION:This study concluded that the infiltration of triamcinolone in keloid scars did not increase the number of apoptosit and did not reduce keloids' size, length, height or thickness.

  14. Sulphamoylated 2-methoxyestradiol analogues induce apoptosis in adenocarcinoma cell lines.

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    Michelle Visagie

    Full Text Available 2-Methoxyestradiol (2ME2 is a naturally occurring estradiol metabolite which possesses antiproliferative, antiangiogenic and antitumor properties. However, due to its limited biological accessibility, synthetic analogues have been synthesized and tested in attempt to develop drugs with improved oral bioavailability and efficacy. The aim of this study was to evaluate the antiproliferative effects of three novel in silico-designed sulphamoylated 2ME2 analogues on the HeLa cervical adenocarcinoma cell line and estrogen receptor-negative breast adenocarcinoma MDA-MB-231 cells. A dose-dependent study (0.1-25 μM was conducted with an exposure time of 24 hours. Results obtained from crystal violet staining indicated that 0.5 μM of all 3 compounds reduced the number of cells to 50%. Lactate dehydrogenase assay was used to assess cytotoxicity, while the mitotracker mitochondrial assay and caspase-6 and -8 activity assays were used to investigate the possible occurrence of apoptosis. Tubulin polymerization assays were conducted to evaluate the influence of these sulphamoylated 2ME2 analogues on tubulin dynamics. Double immunofluorescence microscopy using labeled antibodies specific to tyrosinate and detyrosinated tubulin was conducted to assess the effect of the 2ME2 analogues on tubulin dynamics. An insignificant increase in the level of lactate dehydrogenase release was observed in the compounds-treated cells. These sulphamoylated compounds caused a reduction in mitochondrial membrane potential, cytochrome c release and caspase 3 activation indicating apoptosis induction by means of the intrinsic pathway in HeLa and MDA-MB-231 cells. Microtubule depolymerization was observed after exposure to these three sulphamoylated analogues.

  15. Capsaicin induces cell cycle arrest and apoptosis in human KB cancer cells.

    Science.gov (United States)

    Lin, Chia-Han; Lu, Wei-Cheng; Wang, Che-Wei; Chan, Ya-Chi; Chen, Mu-Kuan

    2013-02-25

    Capsaicin, a pungent phytochemical in a variety of red peppers of the genus Capsicum, has shown an anti-proliferative effect on various human cancer cell lines. In contrast, capsaicin has also been considered to promote the growth of cancer cells. Thus, the effects of capsaicin on various cell types need to be explored. The anti-proliferative effects of capsaicin on human KB cancer cells are still unknown. Therefore, we examined the viability, cell cycle progression, and factors associated with apoptosis in KB cells treated with capsaicin. The cell proliferation/viability and cytotoxicity of KB cells exposed to capsaicin were determined by a sulforhodamine B colorimetric assay and trypan blue exclusion. Apoptosis was detected by Hoechst staining and confirmed by western blot analysis of poly-(ADP-ribose) polymerase cleavage. Cell cycle distribution and changes of the mitochondrial membrane potential were analyzed by flow cytometry. Furthermore, the expression of caspase 3, 8 and 9 was evaluated by immunoblotting. We found that treatment of KB cells with capsaicin significantly reduced cell proliferation/viability and induced cell death in a dose-dependent manner compared with that in the untreated control. Cell cycle analysis indicated that exposure of KB cells to capsaicin resulted in cell cycle arrest at G2/M phase. Capsaicin-induced growth inhibition of KB cells appeared to be associated with induction of apoptosis. Moreover, capsaicin induced disruption of the mitochondrial membrane potential as well as activation of caspase 9, 3 and poly-(ADP-ribose) polymerase in KB cells. Our data demonstrate that capsaicin modulates cell cycle progression and induces apoptosis in human KB cancer cells through mitochondrial membrane permeabilization and caspase activation. These observations suggest an anti-cancer activity of capsaicin.

  16. Cell responses to FGFR3 signalling: growth, differentiation and apoptosis

    International Nuclear Information System (INIS)

    L'Hote, Corine G.M.; Knowles, Margaret A.

    2005-01-01

    FGFR3 is a receptor tyrosine kinase (RTK) of the FGF receptor family, known to have a negative regulatory effect on long bone growth. Fgfr3 knockout mice display longer bones and, accordingly, most germline-activating mutations in man are associated with dwarfism. Somatically, some of the same activating mutations are associated with the human cancers multiple myeloma, cervical carcinoma and carcinoma of the bladder. How signalling through FGFR3 can lead to either chondrocyte apoptosis or cancer cell proliferation is not fully understood. Although FGFR3 can be expressed as two main splice isoforms (IIIb or IIIc), there is no apparent link with specific cell responses, which may rather be associated with the cell type or its differentiation status. Depending on cell type, differential activation of STAT proteins has been observed. STAT1 phosphorylation seems to be involved in inhibition of chondrocyte proliferation while activation of the ERK pathway inhibits chondrocyte differentiation and B-cell proliferation (as in multiple myeloma). The role of FGFR3 in epithelial cancers (bladder and cervix) is not known. Some of the cell specificity may arise via modulation of signalling by crosstalk with other signalling pathways. Recently, inhibition of the ERK pathway in achondroplastic mice has provided hope for an approach to the treatment of dwarfism. Further understanding of the ability of FGFR3 to trigger different responses depending on cell type and cellular context may lead to treatments for both skeletal dysplasias and cancer

  17. Foodborne cereulide causes beta-cell dysfunction and apoptosis.

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    Roman Vangoitsenhoven

    Full Text Available To study the effects of cereulide, a food toxin often found at low concentrations in take-away meals, on beta-cell survival and function.Cell death was quantified by Hoechst/Propidium Iodide in mouse (MIN6 and rat (INS-1E beta-cell lines, whole mouse islets and control cell lines (HepG2 and COS-1. Beta-cell function was studied by glucose-stimulated insulin secretion (GSIS. Mechanisms of toxicity were evaluated in MIN6 cells by mRNA profiling, electron microscopy and mitochondrial function tests.24 h exposure to 5 ng/ml cereulide rendered almost all MIN6, INS-1E and pancreatic islets apoptotic, whereas cell death did not increase in the control cell lines. In MIN6 cells and murine islets, GSIS capacity was lost following 24 h exposure to 0.5 ng/ml cereulide (P<0.05. Cereulide exposure induced markers of mitochondrial stress including Puma (p53 up-regulated modulator of apoptosis, P<0.05 and general pro-apoptotic signals as Chop (CCAAT/-enhancer-binding protein homologous protein. Mitochondria appeared swollen upon transmission electron microscopy, basal respiration rate was reduced by 52% (P<0.05 and reactive oxygen species increased by more than twofold (P<0.05 following 24 h exposure to 0.25 and 0.50 ng/ml cereulide, respectively.Cereulide causes apoptotic beta-cell death at low concentrations and impairs beta-cell function at even lower concentrations, with mitochondrial dysfunction underlying these defects. Thus, exposure to cereulide even at concentrations too low to cause systemic effects appears deleterious to the beta-cell.

  18. Study of cell cycle and apoptosis after radiation with electron linear accelerator injury

    International Nuclear Information System (INIS)

    Xu Lan; Zhou Yinghui; Shi Ning; Peng Miao; Wu Shiliang

    2002-01-01

    Purpose: To determine the cell cycle and apoptosis of the injured cells after radiation with the electron linear accelerator. Methods: NIH 3T3 cells were irradiated by the radiation with the electron linear accelerator. In the experiment the condition of the cell cycle and apoptosis of the injured cells were measured. The expression of p53 was also tested. Results: After exposure to radiation, the number of apoptotic cells as well as the expression of p53 increased. Conclusion: The electron linear accelerator radiation injury can induce cell apoptosis

  19. Isolation and identification of gene mediating radiation-induced apoptosis in human leukemia U937 cells

    International Nuclear Information System (INIS)

    Tong Xin; Luo Ying; Dong Yan; Sun Zhixian

    1998-01-01

    Objective: Increasing evidences suggest that Caspase family proteases play an important role in the effector mechanism of apoptotic cell death. Radiation (IR) can induce apoptosis in tumor cells, so it is very important to isolate and identify the member of the Caspase family proteases involved in IR-induced apoptosis, and this would contribute to the understanding of the mechanism responsible for apoptosis execution. Methods: A PCR approach to isolate genes for IR-induced apoptosis was developed. The approach used degenerated oligonucleotide encoding the highly conserved peptides that were present in all known Caspases. Results: Protease inhibitors special for Caspases could block the apoptotic cell death caused by IR, and Caspase-3 was isolated from irradiated human leukemia U937 cells. Conclusion: Caspases involve in IR-induced apoptosis, and Caspase-3 is the pivotal element of IR-induced apoptosis

  20. A protocol to assess cell cycle and apoptosis in human and mouse pluripotent cells

    Directory of Open Access Journals (Sweden)

    Edel Michael J

    2011-04-01

    Full Text Available Abstract Embryonic stem cells (ESC and induced pluripotent stem cells (iPSCs present a great opportunity to treat and model human disease as a cell replacement therapy. There is a growing pressure to understand better the signal transduction pathways regulating pluripotency and self-renewal of these special cells in order to deliver a safe and reliable cell based therapy in the near future. Many signal transduction pathways converge on two major cell functions associated with self-renewal and pluripotency: control of the cell cycle and apoptosis, although a standard method is lacking across the field. Here we present a detailed protocol to assess the cell cycle and apoptosis of ESC and iPSCs as a single reference point offering an easy to use standard approach across the field.

  1. Neuronal injury external to the retina rapidly activates retinal glia, followed by elevation of markers for cell cycle re-entry and death in retinal ganglion cells.

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    Alba Galan

    Full Text Available Retinal ganglion cells (RGCs are neurons that relay visual signals from the retina to the brain. The RGC cell bodies reside in the retina and their fibers form the optic nerve. Full transection (axotomy of the optic nerve is an extra-retinal injury model of RGC degeneration. Optic nerve transection permits time-kinetic studies of neurodegenerative mechanisms in neurons and resident glia of the retina, the early events of which are reported here. One day after injury, and before atrophy of RGC cell bodies was apparent, glia had increased levels of phospho-Akt, phospho-S6, and phospho-ERK1/2; however, these signals were not detected in injured RGCs. Three days after injury there were increased levels of phospho-Rb and cyclin A proteins detected in RGCs, whereas these signals were not detected in glia. DNA hyperploidy was also detected in RGCs, indicative of cell cycle re-entry by these post-mitotic neurons. These events culminated in RGC death, which is delayed by pharmacological inhibition of the MAPK/ERK pathway. Our data show that a remote injury to RGC axons rapidly conveys a signal that activates retinal glia, followed by RGC cell cycle re-entry, DNA hyperploidy, and neuronal death that is delayed by preventing glial MAPK/ERK activation. These results demonstrate that complex and variable neuro-glia interactions regulate healthy and injured states in the adult mammalian retina.

  2. Casticin impairs cell growth and induces cell apoptosis via cell cycle arrest in human oral cancer SCC-4 cells.

    Science.gov (United States)

    Chou, Guan-Ling; Peng, Shu-Fen; Liao, Ching-Lung; Ho, Heng-Chien; Lu, Kung-Wen; Lien, Jin-Cherng; Fan, Ming-Jen; La, Kuang-Chi; Chung, Jing-Gung

    2018-02-01

    Casticin, a polymethoxyflavone, present in natural plants, has been shown to have biological activities including anti-cancer activities. Herein, we investigated the anti-oral cancer activity of casticin on SCC-4 cells in vitro. Viable cells, cell cycle distribution, apoptotic cell death, reactive oxygen species (ROS) production, and Ca 2+ production, levels of ΔΨ m and caspase activity were measured by flow cytometric assay. Cell apoptosis associated protein expressions were examined by Western blotting and confocal laser microscopy. Results indicated that casticin induced cell morphological changes, DNA condensation and damage, decreased the total viable cells, induced G 2 /M phase arrest in SCC-4 cells. Casticin promoted ROS and Ca 2+ productions, decreases the levels of ΔΨ m , promoted caspase-3, -8, and -9 activities in SCC-4 cells. Western blotting assay demonstrated that casticin affect protein level associated with G2/M phase arrest and apoptosis. Confocal laser microscopy also confirmed that casticin increased the translocation of AIF and cytochrome c in SCC-4 cells. In conclusion, casticin decreased cell number through G 2 /M phase arrest and the induction of cell apoptosis through caspase- and mitochondria-dependent pathways in SCC-4 cells. © 2017 Wiley Periodicals, Inc.

  3. SAMHD1 controls cell cycle status, apoptosis and HIV-1 infection in monocytic THP-1 cells

    International Nuclear Information System (INIS)

    Bonifati, Serena; Daly, Michele B.; St Gelais, Corine; Kim, Sun Hee; Hollenbaugh, Joseph A.; Shepard, Caitlin; Kennedy, Edward M.; Kim, Dong-Hyun; Schinazi, Raymond F.; Kim, Baek; Wu, Li

    2016-01-01

    SAMHD1 limits HIV-1 infection in non-dividing myeloid cells by decreasing intracellular dNTP pools. HIV-1 restriction by SAMHD1 in these cells likely prevents activation of antiviral immune responses and modulates viral pathogenesis, thus highlighting a critical role of SAMHD1 in HIV-1 physiopathology. Here, we explored the function of SAMHD1 in regulating cell proliferation, cell cycle progression and apoptosis in monocytic THP-1 cells. Using the CRISPR/Cas9 technology, we generated THP-1 cells with stable SAMHD1 knockout. We found that silencing of SAMHD1 in cycling cells stimulates cell proliferation, redistributes cell cycle population in the G_1/G_0 phase and reduces apoptosis. These alterations correlated with increased dNTP levels and more efficient HIV-1 infection in dividing SAMHD1 knockout cells relative to control. Our results suggest that SAMHD1, through its dNTPase activity, affects cell proliferation, cell cycle distribution and apoptosis, and emphasize a key role of SAMHD1 in the interplay between cell cycle regulation and HIV-1 infection.

  4. Nuclear orphan receptor TLX affects gene expression, proliferation and cell apoptosis in beta cells.

    Science.gov (United States)

    Shi, Xiaoli; Xiong, Xiaokan; Dai, Zhe; Deng, Haohua; Sun, Li; Hu, Xuemei; Zhou, Feng; Xu, Yancheng

    Nuclear orphan receptor TLX is an essential regulator of the growth of neural stem cells. However, its exact function in pancreatic islet cells is still unknown. In the present study, gene expression profiling analysis revealed that overexpression of TLX in beta cell line MIN6 causes suppression of 176 genes and upregulation of 49 genes, including a cadre of cell cycle, cell proliferation and cell death control genes, such as Btg2, Ddit3 and Gadd45a. We next examined the effects of TLX overexpression on proliferation, apoptosis and insulin secretion in MIN6 cells. Proliferation analysis using EdU assay showed that overexpression of TLX increased percentage of EdU-positive cells. Cell cycle and apoptosis analysis revealed that overexpression of TLX in MIN6 cells resulted in higher percentage of cells exiting G1 into S-phase, and a 58.8% decrease of cell apoptosis induced by 0.5 mM palmitate. Moreover, TLX overexpression did not cause impairment of insulin secretion. Together, we conclude that TLX is among factors capable of controlling beta cell proliferation and survival, which may serve as a target for the development of novel therapies for diabetes. Copyright © 2015 Elsevier Inc. All rights reserved.

  5. SAMHD1 controls cell cycle status, apoptosis and HIV-1 infection in monocytic THP-1 cells

    Energy Technology Data Exchange (ETDEWEB)

    Bonifati, Serena [Center for Retrovirus Research, Department of Veterinary Biosciences, The Ohio State University, Columbus, OH (United States); Daly, Michele B. [Center for Drug Discovery, Department of Pediatrics, School of Medicine, Emory University, Atlanta, GA (United States); St Gelais, Corine; Kim, Sun Hee [Center for Retrovirus Research, Department of Veterinary Biosciences, The Ohio State University, Columbus, OH (United States); Hollenbaugh, Joseph A.; Shepard, Caitlin [Center for Drug Discovery, Department of Pediatrics, School of Medicine, Emory University, Atlanta, GA (United States); Kennedy, Edward M. [Department of Molecular Genetics and Microbiology, Duke University, Durham, NC (United States); Kim, Dong-Hyun [Department of Pharmacy, School of Pharmacy, Kyung-Hee University, Seoul (Korea, Republic of); Schinazi, Raymond F. [Center for Drug Discovery, Department of Pediatrics, School of Medicine, Emory University, Atlanta, GA (United States); Kim, Baek, E-mail: baek.kim@emory.edu [Center for Drug Discovery, Department of Pediatrics, School of Medicine, Emory University, Atlanta, GA (United States); Department of Pharmacy, School of Pharmacy, Kyung-Hee University, Seoul (Korea, Republic of); Wu, Li, E-mail: wu.840@osu.edu [Center for Retrovirus Research, Department of Veterinary Biosciences, The Ohio State University, Columbus, OH (United States)

    2016-08-15

    SAMHD1 limits HIV-1 infection in non-dividing myeloid cells by decreasing intracellular dNTP pools. HIV-1 restriction by SAMHD1 in these cells likely prevents activation of antiviral immune responses and modulates viral pathogenesis, thus highlighting a critical role of SAMHD1 in HIV-1 physiopathology. Here, we explored the function of SAMHD1 in regulating cell proliferation, cell cycle progression and apoptosis in monocytic THP-1 cells. Using the CRISPR/Cas9 technology, we generated THP-1 cells with stable SAMHD1 knockout. We found that silencing of SAMHD1 in cycling cells stimulates cell proliferation, redistributes cell cycle population in the G{sub 1}/G{sub 0} phase and reduces apoptosis. These alterations correlated with increased dNTP levels and more efficient HIV-1 infection in dividing SAMHD1 knockout cells relative to control. Our results suggest that SAMHD1, through its dNTPase activity, affects cell proliferation, cell cycle distribution and apoptosis, and emphasize a key role of SAMHD1 in the interplay between cell cycle regulation and HIV-1 infection.

  6. Imiquimod activates p53-dependent apoptosis in a human basal cell carcinoma cell line.

    Science.gov (United States)

    Huang, Shi-Wei; Chang, Shu-Hao; Mu, Szu-Wei; Jiang, Hsin-Yi; Wang, Sin-Ting; Kao, Jun-Kai; Huang, Jau-Ling; Wu, Chun-Ying; Chen, Yi-Ju; Shieh, Jeng-Jer

    2016-03-01

    The tumor suppressor p53 controls DNA repair, cell cycle, apoptosis, autophagy and numerous other cellular processes. Imiquimod (IMQ), a synthetic toll-like receptor (TLR) 7 ligand for the treatment of superficial basal cell carcinoma (BCC), eliminates cancer cells by activating cell-mediated immunity and directly inducing apoptosis and autophagy in cancer cells. To evaluate the role of p53 in IMQ-induced cell death in skin cancer cells. The expression, phosphorylation and subcellular localization of p53 were detected by real-time PCR, luciferase reporter assay, cycloheximide chase analysis, immunoblotting and immunocytochemistry. Using BCC/KMC1 cell line as a model, the upstream signaling of p53 activation was dissected by over-expression of TLR7/8, the addition of ROS scavenger, ATM/ATR inhibitors and pan-caspase inhibitor. The role of p53 in IMQ-induced apoptosis and autophagy was assessed by genetically silencing p53 and evaluated by a DNA content assay, immunoblotting, LC3 puncta detection and acridine orange staining. IMQ induced p53 mRNA expression and protein accumulation, increased Ser15 phosphorylation, promoted nuclear translocation and up-regulated its target genes in skin cancer cells in a TLR7/8-independent manner. In BCC/KMC1 cells, the induction of p53 by IMQ was achieved through increased ROS production to stimulate the ATM/ATR-Chk1/Chk2 axis but was not mediated by inducing DNA damage. The pharmacological inhibition of ATM/ATR significantly suppressed IMQ-induced p53 activation and apoptosis. Silencing of p53 significantly decreased the IMQ-induced caspase cascade activation and apoptosis but enhanced autophagy. Mutant p53 skin cancer cell lines were more resistant to IMQ-induced apoptosis than wildtype p53 skin cancer cell lines. IMQ induced ROS production to stimulate ATM/ATR pathways and contributed to p53-dependent apoptosis in a skin basal cell carcinoma cell line BCC/KMC1. Copyright © 2015 Japanese Society for Investigative Dermatology

  7. Long-term gene therapy causes transgene-specific changes in the morphology of regenerating retinal ganglion cells.

    Directory of Open Access Journals (Sweden)

    Jennifer Rodger

    Full Text Available Recombinant adeno-associated viral (rAAV vectors can be used to introduce neurotrophic genes into injured CNS neurons, promoting survival and axonal regeneration. Gene therapy holds much promise for the treatment of neurotrauma and neurodegenerative diseases; however, neurotrophic factors are known to alter dendritic architecture, and thus we set out to determine whether such transgenes also change the morphology of transduced neurons. We compared changes in dendritic morphology of regenerating adult rat retinal ganglion cells (RGCs after long-term transduction with rAAV2 encoding: (i green fluorescent protein (GFP, or (ii bi-cistronic vectors encoding GFP and ciliary neurotrophic factor (CNTF, brain-derived neurotrophic factor (BDNF or growth-associated protein-43 (GAP43. To enhance regeneration, rats received an autologous peripheral nerve graft onto the cut optic nerve of each rAAV2 injected eye. After 5-8 months, RGCs with regenerated axons were retrogradely labeled with fluorogold (FG. Live retinal wholemounts were prepared and GFP positive (transduced or GFP negative (non-transduced RGCs injected iontophoretically with 2% lucifer yellow. Dendritic morphology was analyzed using Neurolucida software. Significant changes in dendritic architecture were found, in both transduced and non-transduced populations. Multivariate analysis revealed that transgenic BDNF increased dendritic field area whereas GAP43 increased dendritic complexity. CNTF decreased complexity but only in a subset of RGCs. Sholl analysis showed changes in dendritic branching in rAAV2-BDNF-GFP and rAAV2-CNTF-GFP groups and the proportion of FG positive RGCs with aberrant morphology tripled in these groups compared to controls. RGCs in all transgene groups displayed abnormal stratification. Thus in addition to promoting cell survival and axonal regeneration, vector-mediated expression of neurotrophic factors has measurable, gene-specific effects on the morphology of injured

  8. Ganglion cell-inner plexiform layer and retinal nerve fibre layer changes within the macula in retinitis pigmentosa: a spectral domain optical coherence tomography study.

    Science.gov (United States)

    Yoon, Chang Ki; Yu, Hyeong Gon

    2018-03-01

    To investigate how macular ganglion cell-inner plexiform layer (GCIPL) and retinal nerve fibre layer (RNFL) thicknesses within the macula change with retinitis pigmentosa (RP) severity. Spectral domain optical coherence tomography (SD-OCT) was used to examine 177 patients with RP and 177 normal controls. An optical coherence tomography (OCT) line scan was used to grade RP severity. Retinitis pigmentosa (RP) was categorized as more advanced if there was no identifiable inner segment ellipsoid (ISe) band (NISE) and as less advanced if an ISe band could be identified and peripheral loss of ISe was apparent (IISE). Ganglion cell-inner plexiform layer (GCIPL) and RNFL thicknesses were manually measured on OCT images and analysed. Pearson's correlation analyses were used to examine correlations between GCIPL thickness, RNFL thickness, visual acuity (VA) and visual field extent in patients and controls. Ganglion cell-inner plexiform layer (GCIPL) was significantly thicker in IISE than in control eyes (p layer (RNFL) was significantly thicker in eyes with IISE and NISE than in control eyes in both horizontal and vertical meridians (all p layer (GCIPL) thickness showed a weak positive correlation with vision, and RNFL thickness showed a weak negative correlation with vision and visual field extent. Based on these results, the inner retina, including the GCIPL and RNFL, maintains its gross integrity longer than the photoreceptor layer in RP. Additionally, thickening of the inner retina may have some functional implications in patients with RP. © 2017 Acta Ophthalmologica Scandinavica Foundation. Published by John Wiley & Sons Ltd.

  9. EFFECT OF INTRAVITREAL RANIBIZUMAB ON GANGLION CELL COMPLEX AND PERIPAPILLARY RETINAL NERVE FIBER LAYER IN NEOVASCULAR AGE-RELATED MACULAR DEGENERATION USING SPECTRAL DOMAIN OPTICAL COHERENCE TOMOGRAPHY.

    Science.gov (United States)

    Zucchiatti, Ilaria; Cicinelli, Maria V; Parodi, Maurizio Battaglia; Pierro, Luisa; Gagliardi, Marco; Accardo, Agostino; Bandello, Francesco

    2017-07-01

    To analyze the changes in ganglion cell complex and peripapillary retinal nerve fiber layer thickness, in central macular thickness and choroidal thickness on spectral domain optical coherence tomography in patients with neovascular age-related macular degeneration treated with intravitreal ranibizumab injections. All consecutive patients with untreated neovascular age-related macular degeneration received loading phase of three monthly intravitreal ranibizumab, followed by retreatments on a pro re nata protocol for 12 months. changes in ganglion cell complex and retinal nerve fiber layer at the end of follow-up. Secondary outcome: changes in best-corrected visual acuity, central macular thickness, and choroidal thickness at the end of follow-up. Choroidal thickness was measured at 500 μm, 1000 μm, and 1,500 μm intervals nasally, temporally, superiorly, and inferiorly to the fovea, respectively, on horizontal and vertical line scans centered on the fovea. Twenty-four eyes were included. Ganglion cell complex and peripapillary retinal nerve fiber layer thickness did not show statistically significant changes through 12 months (55.6 ± 18.5 and 81.9 ± 9.9 μm at baseline, 52.7 ± 19.3 and 84.6 ± 15.5 μm at month 12, P > 0.05). Central macular thickness showed progressive decrease from baseline to month 12, with maximum reduction at month 3 (P macular thickness was significantly reduced at the end of treatment. Further studies, with larger sample, longer follow-up, and greater number of injections, are warranted.

  10. Apoptosis in vascular cells induced by cold atmospheric plasma treatment

    NARCIS (Netherlands)

    Sladek, R.E.J.; Stoffels - Adamowicz, E.

    2006-01-01

    Apoptosis is a natural mechanism of cellular self-destruction. It can be triggered by moderate, yet irreversible damage. Apoptosis plays a major role in tissue renewal. Artificial apoptosis induction will become a novel therapy that meets all requirements for tissue-saving surgery. Diseased tissues

  11. Differentiation of retinal ganglion cells and photoreceptor precursors from mouse induced pluripotent stem cells carrying an Atoh7/Math5 lineage reporter.

    Directory of Open Access Journals (Sweden)

    Bin-Bin Xie

    Full Text Available The neural retina is a critical component of the visual system, which provides the majority of sensory input in humans. Various retinal degenerative diseases can result in the permanent loss of retinal neurons, especially the light-sensing photoreceptors and the centrally projecting retinal ganglion cells (RGCs. The replenishment of lost RGCs and the repair of optic nerve damage are particularly challenging, as both RGC specification and their subsequent axonal growth and projection involve complex and precise regulation. To explore the developmental potential of pluripotent stem cell-derived neural progenitors, we have established mouse iPS cells that allow cell lineage tracing of progenitors that have expressed Atoh7/Math5, a bHLH transcription factor required for RGC production. These Atoh7 lineage reporter iPS cells encode Cre to replace one copy of the endogenous Atoh7 gene and a Cre-dependent YFP reporter in the ROSA locus. In addition, they express pluripotent markers and are capable of generating teratomas in vivo. Under anterior neural induction and neurogenic conditions in vitro, the Atoh7-Cre/ROSA-YFP iPS cells differentiate into neurons that co-express various RGC markers and YFP, indicating that these neurons are derived from Atoh7-expressing progenitors. Consistent with previous in vivo cell lineage studies, the Atoh7-Cre/ROSA-YFP iPS cells also give rise to a subset of Crx-positive photoreceptor precursors. Furthermore, inhibition of Notch signaling in the iPSC cultures results in a significant increase of YFP-positive RGCs and photoreceptor precursors. Together, these results show that Atoh7-Cre/ROSA-YFP iPS cells can be used to monitor the development and survival of RGCs and photoreceptors from pluripotent stem cells.

  12. Cannabidiol rather than Cannabis sativa extracts inhibit cell growth and induce apoptosis in cervical cancer cells.

    Science.gov (United States)

    Lukhele, Sindiswa T; Motadi, Lesetja R

    2016-09-01

    Cervical cancer remains a global health related issue among females of Sub-Saharan Africa, with over half a million new cases reported each year. Different therapeutic regimens have been suggested in various regions of Africa, however, over a quarter of a million women die of cervical cancer, annually. This makes it the most lethal cancer amongst black women and calls for urgent therapeutic strategies. In this study we compare the anti-proliferative effects of crude extract of Cannabis sativa and its main compound cannabidiol on different cervical cancer cell lines. To achieve our aim, phytochemical screening, MTT assay, cell growth analysis, flow cytometry, morphology analysis, Western blot, caspase 3/7 assay, and ATP measurement assay were conducted. Results obtained indicate that both cannabidiol and Cannabis sativa extracts were able to halt cell proliferation in all cell lines at varying concentrations. They further revealed that apoptosis was induced by cannabidiol as shown by increased subG0/G1 and apoptosis through annexin V. Apoptosis was confirmed by overexpression of p53, caspase 3 and bax. Apoptosis induction was further confirmed by morphological changes, an increase in Caspase 3/7 and a decrease in the ATP levels. In conclusion, these data suggest that cannabidiol rather than Cannabis sativa crude extracts prevent cell growth and induce cell death in cervical cancer cell lines.

  13. Both systemic and local application of Granulocyte-colony stimulating factor (G-CSF is neuroprotective after retinal ganglion cell axotomy

    Directory of Open Access Journals (Sweden)

    Dietz Gunnar PH

    2009-05-01

    Full Text Available Abstract Background The hematopoietic Granulocyte-Colony Stimulating Factor (G-CSF plays a crucial role in controlling the number of neutrophil progenitor cells. Its function is mediated via the G-CSF receptor, which was recently found to be expressed also in the central nervous system. In addition, G-CSF provided neuroprotection in models of neuronal cell death. Here we used the retinal ganglion cell (RGC axotomy model to compare effects of local and systemic application of neuroprotective molecules. Results We found that the G-CSF receptor is robustly expressed by RGCs in vivo and in vitro. We thus evaluated G-CSF as a neuroprotectant for RGCs and found a dose-dependent neuroprotective effect of G-CSF on axotomized RGCs when given subcutaneously. As stem stell mobilization had previously been discussed as a possible contributor to the neuroprotective effects of G-CSF, we compared the local treatment of RGCs by injection of G-CSF into the vitreous body with systemic delivery by subcutaneous application. Both routes of application reduced retinal ganglion cell death to a comparable extent. Moreover, G-CSF enhanced the survival of immunopurified RGCs in vitro. Conclusion We thus show that G-CSF neuroprotection is at least partially independent of potential systemic effects and provide further evidence that the clinically applicable G-CSF could become a treatment option for both neurodegenerative diseases and glaucoma.

  14. Curcumin Induces Autophagy, Apoptosis, and Cell Cycle Arrest in Human Pancreatic Cancer Cells

    Directory of Open Access Journals (Sweden)

    Yaping Zhu

    2017-01-01

    Full Text Available Objective. Curcumin is an active extract from turmeric. The aim of this study was to identify the underlying mechanism of curcumin on PCa cells and the role of autophagy in this process. Methods. The inhibitory effect of curcumin on the growth of PANC1 and BxPC3 cell lines was detected by CCK-8 assay. Cell cycle distribution and apoptosis were tested by flow cytometry. Autophagosomes were tested by cell immunofluorescence assay. The protein expression was detected by Western blot. The correlation between LC3II/Bax and cell viability was analyzed. Results. Curcumin inhibited the cell proliferation in a dose- and time-dependent manner. Curcumin could induce cell cycle arrest at G2/M phase and apoptosis of PCa cells. The autophagosomes were detected in the dosing groups. Protein expression of Bax and LC3II was upregulated, while Bcl2 was downregulated in the high dosing groups of curcumin. There was a significant negative correlation between LC3II/Bax and cell viability. Conclusions. Autophagy could be triggered by curcumin in the treatment of PCa. Apoptosis and cell cycle arrest also participated in this process. These findings imply that curcumin is a multitargeted agent for PCa cells. In addition, autophagic cell death may predominate in the high concentration groups of curcumin.

  15. Multimodal quantitative phase and fluorescence imaging of cell apoptosis

    Science.gov (United States)

    Fu, Xinye; Zuo, Chao; Yan, Hao

    2017-06-01

    Fluorescence microscopy, utilizing fluorescence labeling, has the capability to observe intercellular changes which transmitted and reflected light microscopy techniques cannot resolve. However, the parts without fluorescence labeling are not imaged. Hence, the processes simultaneously happen in these parts cannot be revealed. Meanwhile, fluorescence imaging is 2D imaging where information in the depth is missing. Therefore the information in labeling parts is also not complete. On the other hand, quantitative phase imaging is capable to image cells in 3D in real time through phase calculation. However, its resolution is limited by the optical diffraction and cannot observe intercellular changes below 200 nanometers. In this work, fluorescence imaging and quantitative phase imaging are combined to build a multimodal imaging system. Such system has the capability to simultaneously observe the detailed intercellular phenomenon and 3D cell morphology. In this study the proposed multimodal imaging system is used to observe the cell behavior in the cell apoptosis. The aim is to highlight the limitations of fluorescence microscopy and to point out the advantages of multimodal quantitative phase and fluorescence imaging. The proposed multimodal quantitative phase imaging could be further applied in cell related biomedical research, such as tumor.

  16. Spironolactone induces apoptosis in human mononuclear cells. Association between apoptosis and cytokine suppression

    DEFF Research Database (Denmark)

    Mikkelsen, Martin; Sønder, S U; Nersting, J

    2006-01-01

    preceding apoptosis. An association between the two effects was also seen when testing several SPIR analogues. Contrary to TNF-alpha, the levels of IL-1beta increased in SPIR-treated cultures. However, the amount of IL-1beta in the supernatants depended upon the order of SPIR and LPS addition, as IL-1beta....... In conclusion, suppression of cytokine production by SPIR may be associated with its apoptotic potential, either directly (apoptosis is a consequence of suppressed cytokine production, or vice-versa) or indirectly (suppressed cytokine production and apoptosis are parallel but otherwise unrelated phenomena)....

  17. α-Mangostin Induces Apoptosis and Cell Cycle Arrest in Oral Squamous Cell Carcinoma Cell

    Directory of Open Access Journals (Sweden)

    Hyun-Ho Kwak

    2016-01-01

    Full Text Available Mangosteen has long been used as a traditional medicine and is known to have antibacterial, antioxidant, and anticancer effects. Although the effects of α-mangostin, a natural compound extracted from the pericarp of mangosteen, have been investigated in many studies, there is limited data on the effects of the compound in human oral squamous cell carcinoma (OSCC. In this study, α-mangostin was assessed as a potential anticancer agent against human OSCC cells. α-Mangostin inhibited cell proliferation and induced cell death in OSCC cells in a dose- and time-dependent manner with little to no effect on normal human PDLF cells. α-Mangostin treatment clearly showed apoptotic evidences such as nuclear fragmentation and accumulation of annexin V and PI-positive cells on OSCC cells. α-Mangostin treatment also caused the collapse of mitochondrial membrane potential and the translocation of cytochrome c from the mitochondria into the cytosol. The expressions of the mitochondria-related proteins were activated by α-mangostin. Treatment with α-mangostin also induced G1 phase arrest and downregulated cell cycle-related proteins (CDK/cyclin. Hence, α-mangostin specifically induces cell death and inhibits proliferation in OSCC cells via the intrinsic apoptosis pathway and cell cycle arrest at the G1 phase, suggesting that α-mangostin may be an effective agent for the treatment of OSCC.

  18. Induction of apoptosis by Citrus paradisi essential oil in human leukemic (HL-60) cells.

    Science.gov (United States)

    Hata, Tomona; Sakaguchi, Ikuyo; Mori, Masahiro; Ikeda, Norikazu; Kato, Yoshiko; Minamino, Miki; Watabe, Kazuhito

    2003-01-01

    Limonene is a primary component of citrus essential oils (EOs) and has been reported to induce apoptosis on tumor cells. Little is known about induction of apoptosis by citrus EOs. In this study, we examined induction of apoptosis by Citrus aurantium var. dulcis (sweet orange) EO, Citrus paradisi (grapefruit) EO and Citrus limon (lemon) EO. These EOs induced apoptosis in HL-60 cells and the apoptosis activities were related to the limonene content of the EOs. Moreover, sweet orange EO and grapefruit EO may contain components besides limonene that have apoptotic activity. To identify the components with apoptotic activity, grapefruit EO was fractionated using silica gel columns, and the components were analyzed by GC-MS. The n-hexane fraction contained limonene, and the dichloromethane fraction (DF) contained aldehyde compounds and nootkatone. Decanal, octanal and citral in the DF showed strong apoptotic activity, suggesting that the aldehyde compounds induced apoptosis strongly in HL-60 cells.

  19. Surfactant protein D delays Fas- and TRAIL-mediated extrinsic pathway of apoptosis in T cells.

    Science.gov (United States)

    Djiadeu, Pascal; Kotra, Lakshmi P; Sweezey, Neil; Palaniyar, Nades

    2017-05-01

    Only a few extracellular soluble proteins are known to modulate apoptosis. We considered that surfactant-associated protein D (SP-D), an innate immune collectin present on many mucosal surfaces, could regulate apoptosis. Although SP-D is known to be important for immune cell homeostasis, whether SP-D affects apoptosis is unknown. In this study we aimed to determine the effects of SP-D on Jurkat T cells and human T cells dying by apoptosis. Here we show that SP-D binds to Jurkat T cells and delays the progression of Fas (CD95)-Fas ligand and TRAIL-TRAIL receptor induced, but not TNF-TNF receptor-mediated apoptosis. SP-D exerts its effects by reducing the activation of initiator caspase-8 and executioner caspase-3. SP-D also delays the surface exposure of phosphatidylserine. The effect of SP-D was ablated by the presence of caspase-8 inhibitor, but not by intrinsic pathway inhibitors. The binding ability of SP-D to dying cells decreases during the early stages of apoptosis, suggesting the release of apoptotic cell surface targets during apoptosis. SP-D also delays FasL-induced death of primary human T cells. SP-D delaying the progression of the extrinsic pathway of apoptosis could have important implications in regulating immune cell homeostasis at mucosal surfaces.

  20. Radotinib Induces Apoptosis of CD11b+ Cells Differentiated from Acute Myeloid Leukemia Cells.

    Directory of Open Access Journals (Sweden)

    Sook-Kyoung Heo

    Full Text Available Radotinib, developed as a BCR/ABL tyrosine kinase inhibitor (TKI, is approved for the second-line treatment of chronic myeloid leukemia (CML in South Korea. However, therapeutic effects of radotinib in acute myeloid leukemia (AML are unknown. In the present study, we demonstrate that radotinib significantly decreases the viability of AML cells in a dose-dependent manner. Kasumi-1 cells were more sensitive to radotinib than NB4, HL60, or THP-1 cell lines. Furthermore, radotinib induced CD11b expression in NB4, THP-1, and Kasumi-1 cells either in presence or absence of all trans-retinoic acid (ATRA. We found that radotinib promoted differentiation and induced CD11b expression in AML cells by downregulating LYN. However, CD11b expression induced by ATRA in HL60 cells was decreased by radotinib through upregulation of LYN. Furthermore, radotinib mainly induced apoptosis of CD11b+ cells in the total population of AML cells. Radotinib also increased apoptosis of CD11b+ HL60 cells when they were differentiated by ATRA/dasatinib treatment. We show that radotinib induced apoptosis via caspase-3 activation and the loss of mitochondrial membrane potential (ΔΨm in CD11b+ cells differentiated from AML cells. Our results suggest that radotinib may be used as a candidate drug in AML or a chemosensitizer for treatment of AML by other therapeutics.

  1. Curcumin enhances TRAIL-induced apoptosis of breast cancer cells by regulating apoptosis-related proteins

    Czech Academy of Sciences Publication Activity Database

    Park, S.; Cho, D. J.; Anděra, Ladislav; Suh, N.; Kim, I.

    2013-01-01

    Roč. 383, 1-2 (2013), s. 39-48 ISSN 0300-8177 Institutional support: RVO:68378050 Keywords : TRAIL * curcumin * apoptosis * breast cancer Subject RIV: EB - Genetics ; Molecular Biology Impact factor: 2.388, year: 2013

  2. [The neuroprotective effect of erigeron breviscapus (vant) hand-mazz on retinal ganglion cells after optic nerve crush injury].

    Science.gov (United States)

    Jiang, Bing; Jiang, You-qin

    2003-08-01

    To investigate whether a Chinese herbal medicine, erigeron breviscapus (vant) hand-mazz (EBHM), can protect the retinal ganglion cells (RGC) damaged by calibrated optic nerve crush injury. Forty-two Sprague-Dawley rats were randomly divided into two groups. Calibrated optic nerve crush injury model was induced in the right eyes by a special designed optic nerve clip. The left eyes served as a control. All 42 rats were randomly divided into 2 groups. Group A consisted of the rats with calibrated optic nerve crush injury and group B consisted of rats with calibrated optic nerve crush injury treated with EBHM. In group B, EBHM solution was given once after the crush injury. According to the time interval between the optic nerve crush and the sacrifice, both groups A and B were further divided into three subgroups (day 4, day 14 and day 21). Therefore, there were 7 rats in each subgroup. Three days before sacrifice, 3% fast blue was injected into superior colliculi bilaterally. The eyes were enucleated after the rat was sacrificed, and flat mounts of the retina from both eyes were prepared on a slide and observed under a fluorescence microscope. Four photos with 400 x magnification were taken from each of the four quadrants of the retina 1 mm away from the optic disc. The labeled RGC were counted by a computerized image analyzer. The labeled RGC rate was used for statistical analysis (the labeled RGC rate = number of RGC in injured eye/control eye x 100%). In group A, the labeled RGC rate was (77.79 +/- 7.11)%, (63.76 +/- 3.79)% and (54.66 +/- 4.75)% on day 4, day 14 and day 21, respectively. In group B, the labeled RGC rate was (80.13 +/- 12.03)%, (78.17 +/- 9.19)% and (83.59 +/- 12.61)% on day 4, day 14 and day 21, respectively. In group B, which was treated with EBHM after injury, the labeled RGC rate was significantly higher than that of group A on day 14 and day 21. In the experimental optic nerve crush model in rats, EBHM therapy can increase the survival rate of

  3. Assessment of Rod, Cone, and Intrinsically Photosensitive Retinal Ganglion Cell Contributions to the Canine Chromatic Pupillary Response.

    Science.gov (United States)

    Yeh, Connie Y; Koehl, Kristin L; Harman, Christine D; Iwabe, Simone; Guzman, José M; Petersen-Jones, Simon M; Kardon, Randy H; Komáromy, András M

    2017-01-01

    The purpose of this study was to evaluate a chromatic pupillometry protocol for specific functional assessment of rods, cones, and intrinsically photosensitive retinal ganglion cells (ipRGCs) in dogs. Chromatic pupillometry was tested and compared in 37 dogs in different stages of primary loss of rod, cone, and combined rod/cone and optic nerve function, and in 5 wild-type (WT) dogs. Eyes were stimulated with 1-s flashes of dim (1 cd/m2) and bright (400 cd/m2) blue light (for scotopic conditions) or bright red (400 cd/m2) light with 25-cd/m2 blue background (for photopic conditions). Canine retinal melanopsin/Opn4 was cloned, and its expression was evaluated using real-time quantitative reverse transcription-PCR and immunohistochemistry. Mean ± SD percentage of pupil constriction amplitudes induced by scotopic dim blue (scDB), scotopic bright blue (scBB), and photopic bright red (phBR) lights in WT dogs were 21.3% ± 10.6%, 50.0% ± 17.5%, and 19.4% ± 7.4%, respectively. Melanopsin-mediated responses to scBB persisted for several minutes (7.7 ± 4.6 min) after stimulus offset. In dogs with inherited retinal degeneration, loss of rod function resulted in absent scDB responses, followed by decreased phBR responses with disease progression and loss of cone function. Primary loss of cone function abolished phBR responses but preserved those responses to blue light (scDB and scBB). Although melanopsin/Opn4 expression was diminished with retinal degeneration, melanopsin-expressing ipRGCs were identified for the first time in both WT and degenerated canine retinas. Pupil responses elicited by light stimuli of different colors and intensities allowed differential functional assessment of canine rods, cones, and ipRGCs. Chromatic pupillometry offers an effective tool for diagnosing retinal and optic nerve diseases.

  4. Retina ganglion cell/inner plexiform layer and peripapillary nerve fiber layer thickness in patients with acromegaly.

    Science.gov (United States)

    Şahin, Muhammed; Şahin, Alparslan; Kılınç, Faruk; Yüksel, Harun; Özkurt, Zeynep Gürsel; Türkcü, Fatih Mehmet; Pekkolay, Zafer; Soylu, Hikmet; Çaça, İhsan

    2017-06-01

    Increased secretion of growth hormone and insulin-like growth factor-1 in acromegaly has various effects on multiple organs. However, the ocular effects of acromegaly have yet to be investigated in detail. The aim of the present study was to compare retina ganglion cell/inner plexiform layer (GCIPL) and peripapillary nerve fiber layer thickness (pRNFL) between patients with acromegaly and healthy control subjects using spectral domain optical coherence tomography (SD-OCT). This cross-sectional, comparative study included 18 patients with acromegaly and 20 control subjects. All participants underwent SD-OCT to measure pRNFL (in the seven peripapillary areas), GCIPL (in the nine ETDRS areas), and central macular thickness (CMT). Visual field (VF) examinations were performed using a Humphrey field analyzer in acromegalic patients. Measurements were compared between patients with acromegaly and control subjects. A total of 33 eyes of 18 patients with acromegaly and 40 eyes of 20 control subjects met the inclusion criteria of the present study. The overall calculated average pRNFL thickness was significantly lower in patients with acromegaly than in control subjects (P = 0.01), with pRNFL thickness significantly lower in the temporal superior and temporal inferior quadrants. Contrary to our expectations, pRNFL thickness in the nasal quadrant was similar between acromegalic and control subjects. The mean overall pRNFL thickness and superonasal, nasal, inferonasal, and inferotemporal quadrant pRNFL thicknesses were found to correlate with the mean deviation (MD) according to Spearman's correlation. However, other quadrants were not correlated with VF sensitivity. No significant difference in CMT values was observed (P = 0.6). GCIPL thickness was significantly lower in all quadrants of the inner and outer macula, except for central and inferior outer quadrants, in the acromegaly group than that in the control group (P acromegaly compared with that in control subjects

  5. Expression of hepatitis C virus envelope protein 2 induces apoptosis in cultured mammalian cells

    Institute of Scientific and Technical Information of China (English)

    Li-Xin Zhu; Jing Liu; You-Hua Xie; Yu-Ying Kong; Ye Ye; Chun-Lin Wang; Guang-Di Li; Yuan Wang

    2004-01-01

    AIM: To explore the role of hepatitis C virus (HCV) envelope protein 2 (E2) in the induction of apoptosis.METHODS: A carboxyterminal truncated E2 (E2-661) was transiently expressed in several cultured mammalian cell lines or stably expressed in Chinese hamster ovary (CHO)cell line. Cell proliferation was assessed by 3H thymidine uptake. Apoptosis was examined by Hoechst 33258staining, flow cytometry and DNA fragmentation analysis.RESULTS: Reduced proliferation was readily observed in the E2-661 expressing cells. These cells manifested the typical features of apoptosis, including cell shrinkage,chromatin condensation and hypodiploid genomic DNA content. Similar apoptotic cell death was observed in an E2-661 stably expressing cell line.CONCLUSION: HCV E2 can induce apoptosis in cultured mammalian cells.

  6. Apoptosis induced by cold shock in vitro is dependent on cell growth phase.

    Science.gov (United States)

    Soloff, B L; Nagle, W A; Moss, A J; Henle, K J; Crawford, J T

    1987-06-15

    Chinese hamster V79 fibroblast cells were exposed to brief periods of cold but non-freezing temperatures at different points on the population growth curve. Upon rewarming, cells at the transition from logarithmic to stationary growth exhibited apoptosis (programmed cell death). Cells in other stages of growth, or after reentry into logarithmic growth by refeeding, did not exhibit apoptosis. Apoptosis was expressed by marked cytoplasmic blebbing, by a characteristic non-random fragmentation of DNA into nucleosomal-sized pieces, and by loss of colony-forming ability. The data suggest that cold shock served as a stimulus for susceptible cells to undergo apoptosis. Thus, the experiments describe a new in vitro system for studying the mechanisms of apoptosis.

  7. Dihydromyricetin induces cell cycle arrest and apoptosis in melanoma SK-MEL-28 cells.

    Science.gov (United States)

    Zeng, Guofang; Liu, Jie; Chen, Hege; Liu, Bin; Zhang, Qingyu; Li, Mingyi; Zhu, Runzhi

    2014-06-01

    Dihydromyricetin (DHM) exhibits multiple pharmacological activities; however, the role of DHM in anti-melanoma activities and the underlying molecular mechanisms are unclear. The aim of the present study was to evaluate the effects of DHM on cell proliferation, cell cycle distribution and apoptosis in the human melanoma SK-MEL-28 cell line, and to explore the related mechanisms. The effect of DHM on cell proliferation was investigated by MTT assay, and cell cycle distribution was determined by flow cytometry. TUNEL assay was used to evaluate DHM-mediated apoptosis, and western blotting was applied to examine expression levels of p53, p21, Cdc25A, Cdc2, P-Cdc2, Bax, IKK-α, NF-κB p65, p38 and P-p38 proteins. The results revealed that DHM suppressed cell proliferation of SK-MEL-28 cells in a concentration- and time-dependent manner, and caused cell cycle arrest at the G1/S phase. DHM increased the production of p53 and p21 proteins and downregulated the production of Cdc25A, Cdc2 and P-Cdc2 proteins, which induced cell cycle arrest. Additionally, DHM significantly induced the apoptosis of SK-MEL-28 cells, and enhanced the expression levels of Bax proteins and decreased the protein levels of IKK-α, NF-κB (p65) and P-p38. The results suggest that DHM may be a novel and effective candidate agent to inhibit the growth of melanoma.

  8. Human Adipose Derived Stem Cells Induced Cell Apoptosis and S Phase Arrest in Bladder Tumor

    Directory of Open Access Journals (Sweden)

    Xi Yu

    2015-01-01

    Full Text Available The aim of this study was to determine the effect of human adipose derived stem cells (ADSCs on the viability and apoptosis of human bladder cancer cells. EJ and T24 cells were cocultured with ADSCs or cultured with conditioned medium of ADSCs (ADSC-CM, respectively. The cell counting and colony formation assay showed ADSCs inhibited the proliferation of EJ and T24 cells. Cell viability assessment revealed that the secretions of ADSCs, in the form of conditioned medium, were able to decrease cancer cell viability. Wound-healing assay suggested ADSC-CM suppressed migration of T24 and EJ cells. Moreover, the results of the flow cytometry indicated that ADSC-CM was capable of inducing apoptosis of T24 cells and inducing S phase cell cycle arrest. Western blot revealed ADSC-CM increased the expression of cleaved caspase-3 and cleaved PARP, indicating that ADSC-CM induced apoptosis in a caspase-dependent way. PTEN/PI3K/Akt pathway and Bcl-2 family proteins were involved in the mechanism of this reaction. Our study indicated that ADSCs may provide a promising and practicable manner for bladder tumor therapy.

  9. Study of radionuclide 90Sr-90Y on cell proliferation and apoptosis in benign prostatic hyperplasia

    International Nuclear Information System (INIS)

    Zhang Tong; Wei Wei; Zou Benjie; Liu Fang; Xu Zhishun

    2003-01-01

    Objective: To investigate the effect of 90 Sr- 90 Yon cell proliferation and apoptosis in benign prostatic hyperplasia. Methods: The apoptosis and expression of Ki-67 in benign prostatic hyperplasia (BPH) before and after irradiation 90 Sr- 90 Y were detected by transferase-mediated dUTP-biotin nick end labeling (TUNEL) method and immunohistochemical technique, respectively. Results: The proliferation index (PI) of BPH after 90 Sr- 90 Y irradiation was much lower than that before irradiation, but there was no significant change in apoptosis index (AI). Conclusion: Irradiation with 90 Sr- 90 Y could restrain cell proliferation of BPH, but could not induce apoptosis

  10. Anaplasma phagocytophilum Manipulates Host Cell Apoptosis by Different Mechanisms to Establish Infection

    Directory of Open Access Journals (Sweden)

    Pilar Alberdi

    2016-07-01

    Full Text Available Anaplasma phagocytophilum is an emerging zoonotic pathogen that causes human and animal granulocytic anaplasmosis and tick-borne fever of ruminants. This obligate intracellular bacterium evolved to use common strategies to establish infection in both vertebrate hosts and tick vectors. Herein, we discuss the different strategies used by the pathogen to modulate cell apoptosis and establish infection in host cells. In vertebrate neutrophils and human promyelocytic cells HL-60, both pro-apoptotic and anti-apoptotic factors have been reported. Tissue-specific differences in tick response to infection and differential regulation of apoptosis pathways have been observed in adult female midguts and salivary glands in response to infection with A. phagocytophilum. In tick midguts, pathogen inhibits apoptosis through the Janus kinase/signal transducers and activators of transcription (JAK/STAT pathway, while in salivary glands, the intrinsic apoptosis pathways is inhibited but tick cells respond with the activation of the extrinsic apoptosis pathway. In Ixodes scapularis ISE6 cells, bacterial infection down-regulates mitochondrial porin and manipulates protein processing in the endoplasmic reticulum and cell glucose metabolism to inhibit apoptosis and facilitate infection, whereas in IRE/CTVM20 tick cells, inhibition of apoptosis appears to be regulated by lower caspase levels. These results suggest that A. phagocytophilum uses different mechanisms to inhibit apoptosis for infection of both vertebrate and invertebrate hosts.

  11. [Gene Expression Profile of Apoptosis in Leukemia Cells Induced by Hsp90 Selective inhibitor 17-AAG].

    Science.gov (United States)

    Wang, Na-Na; Li, Zhi-Heng; Tao, Yan-Fang; Xu, Li-Xiao; Pan, Jian; Hu, Shao-Yan

    2016-06-01

    To investigate the apoptotic effects of Hsp90 selective inhibitor 17-AAG on human leukemia HL-60 and NB4 cells and analyse its possible mechanism. CCK-8 assay was used to quantify the growth inhibition of cells after exposure to 17-AAG for 24 hours. Flow cytometrve with annexin V/propidium iodide staining was used to detect apoptosis of leukemia cells. Then Western blot was used to detect the activation of apoptosis related protein caspase-3 and PARP level. Gene expression profile of NB4 cells treated with 17-AAG was analyzed with real-time PCR arrays. The inhibition of leukemia cell proliferation displayed a dose-dependent manner. Annexin V assay, cell cycle analysis and activation of PARP demonstrate that 17-AAG induced apoptosis leukemia cells. Real-time PCR array analysis showed that expression of 56 genes significantly up-regulated and expression of 23 genes were significantly down-regulated after 17-AAG treatment. The 17-AAG can inhibit the proliferation and induce the apoptosis of leukemia cells. After leukemia cells are treated with 17-AAG, the significant changes of apoptosis-related genes occured, and the cell apoptosis occurs via activating apoptosis related signaling pathway.

  12. Sensing lymphoma cells based on a cell-penetrating/apoptosis-inducing/electron-transfer peptide probe

    International Nuclear Information System (INIS)

    Sugawara, Kazuharu; Shinohara, Hiroki; Kadoya, Toshihiko; Kuramitz, Hideki

    2016-01-01

    To electrochemically sense lymphoma cells (U937), we fabricated a multifunctional peptide probe that consists of cell-penetrating/apoptosis-inducing/electron-transfer peptides. Electron-transfer peptides derive from cysteine residue combined with the C-terminals of four tyrosine residues (Y_4). A peptide whereby Y_4C is bound to the C-terminals of protegrin 1 (RGGRLCYCRRRFCVCVGR-NH_2) is known to be an apoptosis-inducing agent against U937 cells, and is referred to as a peptide-1 probe. An oxidation response of the peptide-1 probe has been observed due to a phenolic hydroxyl group, and this response is decreased by the uptake of the peptide probe into the cells. To improve the cell membrane permeability against U937 cells, the RGGR at the N-terminals of the peptide-1 probe was replaced by RRRR (peptide-2 probe). In contrast, RNRCKGTDVQAWY_4C (peptide-3 probe), which recognizes ovalbumin, was constructed as a control. Compared with the other probes, the change in the peak current of the peptide-2 probe was the greatest at low concentrations and occurred in a short amount of time. Therefore, the cell membrane permeability of the peptide-2 probe was increased based on the arginine residues and the apoptosis-inducing peptides. The peak current was linear and ranged from 100 to 1000 cells/ml. The relative standard deviation of 600 cells/ml was 5.0% (n = 5). Furthermore, the membrane permeability of the peptide probes was confirmed using fluorescent dye. - Highlights: • We constructed a multifunctional peptide probe for the electrochemical sensing of lymphoma cells. • The peptide probe consists of cell-penetrating/apoptosis-inducing/electron-transfer peptides. • The electrode response of the peptide probe changes due to selective uptake into the cells.

  13. Adipose-derived stromal cells inhibit prostate cancer cell proliferation inducing apoptosis

    Energy Technology Data Exchange (ETDEWEB)

    Takahara, Kiyoshi [Department of Urology, Faculty of Medicine, Osaka Medical College, Osaka (Japan); Ii, Masaaki, E-mail: masaii@art.osaka-med.ac.jp [Department of Pharmacology, Faculty of Medicine, Osaka Medical College, Osaka (Japan); Inamoto, Teruo; Komura, Kazumasa; Ibuki, Naokazu; Minami, Koichiro; Uehara, Hirofumi; Hirano, Hajime; Nomi, Hayahito; Kiyama, Satoshi [Department of Urology, Faculty of Medicine, Osaka Medical College, Osaka (Japan); Asahi, Michio [Department of Pharmacology, Faculty of Medicine, Osaka Medical College, Osaka (Japan); Azuma, Haruhito [Department of Urology, Faculty of Medicine, Osaka Medical College, Osaka (Japan)

    2014-04-18

    Highlights: • AdSC transplantation exhibits inhibitory effect on tumor progressions of PCa cells. • AdSC-induced PCa cell apoptosis may occur via the TGF-β signaling pathway. • High expression of the TGF-β1 gene in AdSCs. - Abstract: Mesenchymal stem cells (MSCs) have generated a great deal of interest in the field of regenerative medicine. Adipose-derived stromal cells (AdSCs) are known to exhibit extensive proliferation potential and can undergo multilineage differentiation, sharing similar characteristics to bone marrow-derived MSCs. However, as the effect of AdSCs on tumor growth has not been studied sufficiently, we assessed the degree to which AdSCs affect the proliferation of prostate cancer (PCa) cell. Human AdSCs exerted an inhibitory effect on the proliferation of androgen-responsive (LNCaP) and androgen-nonresponsive (PC3) human PCa cells, while normal human dermal fibroblasts (NHDFs) did not, and in fact promoted PCa cell proliferation to a degree. Moreover, AdSCs induced apoptosis of LNCaP cells and PC3 cells, activating the caspase3/7 signaling pathway. cDNA microarray analysis suggested that AdSC-induced apoptosis in both LNCaP and PC3 cells was related to the TGF-β signaling pathway. Consistent with our in vitro observations, local transplantation of AdSCs delayed the growth of tumors derived from both LNCaP- and PC3-xenografts in immunodeficient mice. This is the first preclinical study to have directly demonstrated that AdSC-induced PCa cell apoptosis may occur via the TGF-β signaling pathway, irrespective of androgen-responsiveness. Since autologous AdSCs can be easily isolated from adipose tissue without any ethical concerns, we suggest that therapy with these cells could be a novel approach for patients with PCa.

  14. Sensing lymphoma cells based on a cell-penetrating/apoptosis-inducing/electron-transfer peptide probe

    Energy Technology Data Exchange (ETDEWEB)

    Sugawara, Kazuharu, E-mail: kzsuga@maebashi-it.ac.jp [Maebashi Institute of Technology, Gunma 371-0816 (Japan); Shinohara, Hiroki; Kadoya, Toshihiko [Maebashi Institute of Technology, Gunma 371-0816 (Japan); Kuramitz, Hideki [Department of Environmental Biology and Chemistry, Graduate School of Science and Engineering for Research, University of Toyama, Toyama 930-8555 (Japan)

    2016-06-14

    To electrochemically sense lymphoma cells (U937), we fabricated a multifunctional peptide probe that consists of cell-penetrating/apoptosis-inducing/electron-transfer peptides. Electron-transfer peptides derive from cysteine residue combined with the C-terminals of four tyrosine residues (Y{sub 4}). A peptide whereby Y{sub 4}C is bound to the C-terminals of protegrin 1 (RGGRLCYCRRRFCVCVGR-NH{sub 2}) is known to be an apoptosis-inducing agent against U937 cells, and is referred to as a peptide-1 probe. An oxidation response of the peptide-1 probe has been observed due to a phenolic hydroxyl group, and this response is decreased by the uptake of the peptide probe into the cells. To improve the cell membrane permeability against U937 cells, the RGGR at the N-terminals of the peptide-1 probe was replaced by RRRR (peptide-2 probe). In contrast, RNRCKGTDVQAWY{sub 4}C (peptide-3 probe), which recognizes ovalbumin, was constructed as a control. Compared with the other probes, the change in the peak current of the peptide-2 probe was the greatest at low concentrations and occurred in a short amount of time. Therefore, the cell membrane permeability of the peptide-2 probe was increased based on the arginine residues and the apoptosis-inducing peptides. The peak current was linear and ranged from 100 to 1000 cells/ml. The relative standard deviation of 600 cells/ml was 5.0% (n = 5). Furthermore, the membrane permeability of the peptide probes was confirmed using fluorescent dye. - Highlights: • We constructed a multifunctional peptide probe for the electrochemical sensing of lymphoma cells. • The peptide probe consists of cell-penetrating/apoptosis-inducing/electron-transfer peptides. • The electrode response of the peptide probe changes due to selective uptake into the cells.

  15. Adipose-derived stromal cells inhibit prostate cancer cell proliferation inducing apoptosis

    International Nuclear Information System (INIS)

    Takahara, Kiyoshi; Ii, Masaaki; Inamoto, Teruo; Komura, Kazumasa; Ibuki, Naokazu; Minami, Koichiro; Uehara, Hirofumi; Hirano, Hajime; Nomi, Hayahito; Kiyama, Satoshi; Asahi, Michio; Azuma, Haruhito

    2014-01-01

    Highlights: • AdSC transplantation exhibits inhibitory effect on tumor progressions of PCa cells. • AdSC-induced PCa cell apoptosis may occur via the TGF-β signaling pathway. • High expression of the TGF-β1 gene in AdSCs. - Abstract: Mesenchymal stem cells (MSCs) have generated a great deal of interest in the field of regenerative medicine. Adipose-derived stromal cells (AdSCs) are known to exhibit extensive proliferation potential and can undergo multilineage differentiation, sharing similar characteristics to bone marrow-derived MSCs. However, as the effect of AdSCs on tumor growth has not been studied sufficiently, we assessed the degree to which AdSCs affect the proliferation of prostate cancer (PCa) cell. Human AdSCs exerted an inhibitory effect on the proliferation of androgen-responsive (LNCaP) and androgen-nonresponsive (PC3) human PCa cells, while normal human dermal fibroblasts (NHDFs) did not, and in fact promoted PCa cell proliferation to a degree. Moreover, AdSCs induced apoptosis of LNCaP cells and PC3 cells, activating the caspase3/7 signaling pathway. cDNA microarray analysis suggested that AdSC-induced apoptosis in both LNCaP and PC3 cells was related to the TGF-β signaling pathway. Consistent with our in vitro observations, local transplantation of AdSCs delayed the growth of tumors derived from both LNCaP- and PC3-xenografts in immunodeficient mice. This is the first preclinical study to have directly demonstrated that AdSC-induced PCa cell apoptosis may occur via the TGF-β signaling pathway, irrespective of androgen-responsiveness. Since autologous AdSCs can be easily isolated from adipose tissue without any ethical concerns, we suggest that therapy with these cells could be a novel approach for patients with PCa

  16. The mitochondria-mediate apoptosis of Lepidopteran cells induced by azadirachtin.

    Directory of Open Access Journals (Sweden)

    Jingfei Huang

    Full Text Available Mitochondria have been shown to play an important role in apoptosis using mammalian cell lines. However, this seems not to be the case in Drosophila, an insect model organism; thus more in-depth studies of insect cell apoptosis are necessary. In the present study, mitochondrial involvement during azadirachtin- and camptothecin-induced apoptosis in Spodoptera frugiperda Sf9 cells (isolated from Spodoptera frugiperda pupal ovarian tissue was investigated. The results showed that both azadirachtin and camptothecin could induce apoptosis in Sf9 cells. Reactive oxygen species (ROS generation, activation of mitochondrial permeability transition pores (MPTPs and loss of mitochondrial membrane potential (MMP were observed very early during apoptosis and were followed subsequently by the release of cytochrome-c from the mitochondria. Furthermore, the results also revealed that the opening of MPTPs and the loss of MMP induced by azadirachtin could be significantly inhibited by the permeability transition pore (PTP inhibitor cyclosporin A (CsA, which was used to identify the key role of mitochondria in the apoptosis of Sf9 cells. However, in camptothecin-treated Sf9 cells, CsA could not suppress the opening of MPTPs and the loss of MMP when apoptosis was induced. The data from caspase-3 and caspase-9 activity assays and detection of apoptosis by morphological observation and flow cytometry also uncovered the different effect of CsA on the two botanical apoptosis inducers. Although different mechanisms of apoptosis induction exist, our study revealed that mitochondria play a crucial role in insect cell line apoptosis.

  17. The mitochondria-mediate apoptosis of Lepidopteran cells induced by azadirachtin.

    Science.gov (United States)

    Huang, Jingfei; Lv, Chaojun; Hu, Meiying; Zhong, Guohua

    2013-01-01

    Mitochondria have been shown to play an important role in apoptosis using mammalian cell lines. However, this seems not to be the case in Drosophila, an insect model organism; thus more in-depth studies of insect cell apoptosis are necessary. In the present study, mitochondrial involvement during azadirachtin- and camptothecin-induced apoptosis in Spodoptera frugiperda Sf9 cells (isolated from Spodoptera frugiperda pupal ovarian tissue) was investigated. The results showed that both azadirachtin and camptothecin could induce apoptosis in Sf9 cells. Reactive oxygen species (ROS) generation, activation of mitochondrial permeability transition pores (MPTPs) and loss of mitochondrial membrane potential (MMP) were observed very early during apoptosis and were followed subsequently by the release of cytochrome-c from the mitochondria. Furthermore, the results also revealed that the opening of MPTPs and the loss of MMP induced by azadirachtin could be significantly inhibited by the permeability transition pore (PTP) inhibitor cyclosporin A (CsA), which was used to identify the key role of mitochondria in the apoptosis of Sf9 cells. However, in camptothecin-treated Sf9 cells, CsA could not suppress the opening of MPTPs and the loss of MMP when apoptosis was induced. The data from caspase-3 and caspase-9 activity assays and detection of apoptosis by morphological observation and flow cytometry also uncovered the different effect of CsA on the two botanical apoptosis inducers. Although different mechanisms of apoptosis induction exist, our study revealed that mitochondria play a crucial role in insect cell line apoptosis.

  18. Isthmin targets cell-surface GRP78 and triggers apoptosis via induction of mitochondrial dysfunction

    Science.gov (United States)

    Chen, M; Zhang, Y; Yu, V C; Chong, Y-S; Yoshioka, T; Ge, R

    2014-01-01

    Isthmin (ISM) is a secreted 60-kDa protein that potently induces endothelial cell (EC) apoptosis. It suppresses tumor growth and angiogenesis in mice when stably overexpressed in cancer cells. Although αvβ5 integrin serves as a low-affinity receptor for ISM, the mechanism by which ISM mediates antiangiogenesis and apoptosis in ECs remain to be fully resolved. In this work, we report the identification of cell-surface glucose-regulated protein 78 kDa (GRP78) as a high-affinity receptor for ISM (Kd=8.6 nM). We demonstrated that ISM-GRP78 interaction triggers apoptosis not only in activated ECs but also in cancer cells expressing high level of cell-surface GRP78. Normal cells and benign tumor cells tend to express low level of cell-surface GRP78 and are resistant to ISM-induced apoptosis. Upon binding to GRP78, ISM is internalized into ECs through clathrin-dependent endocytosis that is essential for its proapoptotic activity. Once inside the cell, ISM co-targets with GRP78 to mitochondria where it interacts with ADP/ATP carriers on the inner membrane and blocks ATP transport from mitochondria to cytosol, thereby causing apoptosis. Hence, ISM is a novel proapoptotic ligand that targets cell-surface GRP78 to trigger apoptosis by inducing mitochondrial dysfunction. The restricted and high-level expression of cell-surface GRP78 on cancer cells and cancer ECs make them uniquely susceptible to ISM-targeted apoptosis. Indeed, systemic delivery of recombinant ISM potently suppressed subcutaneous 4T1 breast carcinoma and B16 melanoma growth in mice by eliciting apoptosis selectively in the cancer cells and cancer ECs. Together, this work reveals a novel ISM-GRP78 apoptosis pathway and demonstrates the potential of ISM as a cancer-specific and dual-targeting anticancer agent. PMID:24464222

  19. Isthmin targets cell-surface GRP78 and triggers apoptosis via induction of mitochondrial dysfunction.

    Science.gov (United States)

    Chen, M; Zhang, Y; Yu, V C; Chong, Y-S; Yoshioka, T; Ge, R

    2014-05-01

    Isthmin (ISM) is a secreted 60-kDa protein that potently induces endothelial cell (EC) apoptosis. It suppresses tumor growth and angiogenesis in mice when stably overexpressed in cancer cells. Although αvβ5 integrin serves as a low-affinity receptor for ISM, the mechanism by which ISM mediates antiangiogenesis and apoptosis in ECs remain to be fully resolved. In this work, we report the identification of cell-surface glucose-regulated protein 78 kDa (GRP78) as a high-affinity receptor for ISM (Kd=8.6 nM). We demonstrated that ISM-GRP78 interaction triggers apoptosis not only in activated ECs but also in cancer cells expressing high level of cell-surface GRP78. Normal cells and benign tumor cells tend to express low level of cell-surface GRP78 and are resistant to ISM-induced apoptosis. Upon binding to GRP78, ISM is internalized into ECs through clathrin-dependent endocytosis that is essential for its proapoptotic activity. Once inside the cell, ISM co-targets with GRP78 to mitochondria where it interacts with ADP/ATP carriers on the inner membrane and blocks ATP transport from mitochondria to cytosol, thereby causing apoptosis. Hence, ISM is a novel proapoptotic ligand that targets cell-surface GRP78 to trigger apoptosis by inducing mitochondrial dysfunction. The restricted and high-level expression of cell-surface GRP78 on cancer cells and cancer ECs make them uniquely susceptible to ISM-targeted apoptosis. Indeed, systemic delivery of recombinant ISM potently suppressed subcutaneous 4T1 breast carcinoma and B16 melanoma growth in mice by eliciting apoptosis selectively in the cancer cells and cancer ECs. Together, this work reveals a novel ISM-GRP78 apoptosis pathway and demonstrates the potential of ISM as a cancer-specific and dual-targeting anticancer agent.

  20. Ceramide-Induced Apoptosis in Renal Tubular Cells: A Role of Mitochondria and Sphingosine-1-Phoshate

    Science.gov (United States)

    Ueda, Norishi

    2015-01-01

    Ceramide is synthesized upon stimuli, and induces apoptosis in renal tubular cells (RTCs). Sphingosine-1 phosphate (S1P) functions as a survival factor. Thus, the balance of ceramide/S1P determines ceramide-induced apoptosis. Mitochondria play a key role for ceramide-induced apoptosis by altered mitochondrial outer membrane permeability (MOMP). Ceramide enhances oligomerization of pro-apoptotic Bcl-2 family proteins, ceramide channel, and reduces anti-apoptotic Bcl-2 proteins in the MOM. This process alters MOMP, resulting in generation of reactive oxygen species (ROS), cytochrome C release into the cytosol, caspase activation, and apoptosis. Ceramide regulates apoptosis through mitogen-activated protein kinases (MAPKs)-dependent and -independent pathways. Conversely, MAPKs alter ceramide generation by regulating the enzymes involving ceramide metabolism, affecting ceramide-induced apoptosis. Crosstalk between Bcl-2 family proteins, ROS, and many signaling pathways regulates ceramide-induced apoptosis. Growth factors rescue ceramide-induced apoptosis by regulating the enzymes involving ceramide metabolism, S1P, and signaling pathways including MAPKs. This article reviews evidence supporting a role of ceramide for apoptosis and discusses a role of mitochondria, including MOMP, Bcl-2 family proteins, ROS, and signaling pathways, and crosstalk between these factors in the regulation of ceramide-induced apoptosis of RTCs. A balancing role between ceramide and S1P and the strategy for preventing ceramide-induced apoptosis by growth factors are also discussed. PMID:25751724

  1. Isoalantolactone inhibits UM-SCC-10A cell growth via cell cycle arrest and apoptosis induction.

    Directory of Open Access Journals (Sweden)

    Minjun Wu

    Full Text Available Isoalantolactone is a sesquiterpene lactone compound isolated from the roots of Inula helenium L. Previous studies have demonstrated that isoalantolactone possesses antifungal, anti-bacterial, anti-helminthic and anti-proliferative properties in a variety of cells, but there are no studies concerning its effects on head and neck squamous cell carcinoma (HNSCC. In the present study, an MTT assay demonstrated that isoalantolactone has anti-proliferative activity against the HNSCC cell line (UM-SCC-10A. Immunostaining identified that this compound induced UM-SCC-10A cell apoptosis but not necrosis. To explain the molecular mechanisms underlying its effects, flow cytometry and western blot analysis showed that the apoptosis was associated with cell cycle arrest during the G1 phase, up-regulation of p53 and p21, and down-regulation of cyclin D. Furthermore, our results revealed that induction of apoptosis through a mitochondrial pathway led to up-regulation of pro-apoptotic protein expression (Bax, down-regulation of anti-apoptotic protein expression (Bcl-2, mitochondrial release of cytochrome c (Cyto c, reduction of mitochondrial membrane potential (MMP and activation of caspase-3 (Casp-3. Involvement of the caspase apoptosis pathway was confirmed using caspase inhibitor Z-VAD-FMK pretreatment. Together, our findings suggest that isoalantolactone induced caspase-dependent apoptosis via a mitochondrial pathway and was associated with cell cycle arrest in the G1 phase in UM-SCC-10A cells. Therefore, isoalantolactone may become a potential drug for treating HNSCC.

  2. Distinct protease pathways control cell shape and apoptosis in v-src-transformed quail neuroretina cells

    International Nuclear Information System (INIS)

    Neel, Benjamin D.; Aouacheria, Abdel; Nouvion, Anne-Laure; Ronot, Xavier; Gillet, Germain

    2005-01-01

    Intracellular proteases play key roles in cell differentiation, proliferation and apoptosis. In nerve cells, little is known about their relative contribution to the pathways which control cell physiology, including cell death. Neoplastic transformation of avian neuroretina cells by p60 v-src tyrosine kinase results in dramatic morphological changes and deregulation of apoptosis. To identify the proteases involved in the cellular response to p60 v-src , we evaluated the effect of specific inhibitors of caspases, calpains and the proteasome on cell shape changes and apoptosis induced by p60 v-src inactivation in quail neuroretina cells transformed by tsNY68, a thermosensitive strain of Rous sarcoma virus. We found that the ubiquitin-proteasome pathway is recruited early after p60 v-src inactivation and is critical for morphological changes, whereas caspases are essential for cell death. This study provides evidence that distinct intracellular proteases are involved in the control of the morphology and fate of v-src-transformed cells

  3. Solena amplexicaulis induces cell cycle arrest, apoptosis and inhibits angiogenesis in hepatocarcinoma cells and HUVECs.

    Science.gov (United States)

    Ren, Jie; Xu, Yuan Yuan; Jiang, He Fei; Yang, Meng; Huang, Qian Hui; Yang, Jie; Hu, Kun; Wei, Kun

    2014-01-01

    Solena amplexicaulis (Lam.) Gandhi (SA) has been used as a traditional medicine for the treatment of dysentery, multiple abscess, gastralgia, urethritis, and eczema in the minority area of China. This study was aimed to examine the cell proliferation inhibitory activity of the SA extract (SACE) and its mechanism of action in human hepatoma cell line (HepG2) and evaluate its anti-angiogenesis activity in human umbilical vein endothelial cell line (HUVEC). SACE could inhibit the growth of HepG2 cells in a dose- and time-dependent manner. FCM analysis showed that SACE could induce G2/M phase arrest, cell apoptosis, the mitochondrial membrane potential loss (ΔΨm) and increase the production of intracellular ROS of HepG2 cells. After treatment with SACE, topical morphological changes of apoptotic body formation, obvious increase of apoptosis-related protein expressions, such as Bax, cytochrome c, caspase-3, PARP-1, and decrease of Bcl-2, procaspase-9 protein expressions were observed at the same time. Moreover, SACE caused the significant inhibition of endothelial cell migration and tube formation in HUVEC cells. The results suggested that SACE could act as an angiogenesis inhibitor and induce cell apoptosis via a caspase-dependent mitochondrial pathway. Therefore, SACE could be a potent candidate for the prevention and treatment of liver cancer.

  4. Mitofusin 2 Promotes Apoptosis of CD4+ T Cells by Inhibiting Autophagy in Sepsis

    Directory of Open Access Journals (Sweden)

    Lan Ying

    2017-01-01

    Full Text Available Apoptosis of CD4+ T cells is a primary pathophysiological mechanism of immune dysfunction in the pathogenesis of sepsis. Mitofusin 2 (Mfn2, an integral mitochondrial outer membrane protein, has been confirmed to be associated with cellular metabolism, proliferation, and apoptosis. The function of Mfn2 in CD4+ T cell apoptosis in sepsis is poorly understood. Here, we discovered increased in vivo Mfn2 expression, autophagy deficiency, and elevated cell apoptosis in murine splenic CD4+ T cells after cecal ligation and puncture (CLP. We also observed almost identical results in splenic CD4+ T cells upon lipopolysaccharide (LPS stimulation in vitro. Furthermore, overexpression of Mfn2 resulted in impaired autophagy and increased apoptosis in Jurkat cells. Pharmacological inhibition of autophagy with 3-methyladenine enhanced Mfn2 overexpression-induced cell apoptosis. In addition, overexpression of Mfn2 downregulated phorbol myristate acetate (PMA/ionomycin-, rapamycin- and starvation-induced autophagy in Jurkat T cells. Taken together, these data indicate a critical role of Mfn2 in CD4+ T cell apoptosis in sepsis and the underlying mechanism of autophagy deficiency.

  5. Ginkgo Biloba Extract Kaempferol Inhibits Cell Proliferation and Induces Apoptosis in Pancreatic Cancer Cells

    Science.gov (United States)

    Zhang, Yuqing; Chen, Aaron Y.; Li, Min; Chen, Changyi; Yao, Qizhi

    2010-01-01

    Background Kaempferol is one of the most important constituents in ginkgo flavonoids. Recent studies indicate kaempferol may have anti-tumor activities. The objective in this study was to determine the effect and mechanisms of kaempferol on pancreatic cancer cell proliferation and apoptosis. Materials and Methods Pancreatic cancer cell lines MIA PaCa-2 and Panc-1 were treated with Kampferol, and the inhibitory effects of kaempferol on pancreatic cancer cell proliferation were examined by direct cell counting, 3H-thymidine incorporation and MTS assay. Lactate dehydrogenase (LDH) release from cells was determined as an index of cytotoxicity. Apoptosis was analyzed by TUNEL assay. Results Upon the treatment with 70 μM kaempferol for 4 days, MIA PaCa-2 cell proliferation was significantly inhibited by 79% and 45.7% as determined by direct cell counting and MTS assay, respectively, compared with control cells (Pkaempferol significantly inhibited Panc-1 cell proliferation. Kaempferol treatment also significantly reduced 3H-thymidine incorporation in both MIA PaCa-2 and Panc-1 cells. Combination treatment of low concentrations of kaempferol and 5-fluorouracil (5-FU) showed an additive effect on the inhibition of MIA PaCa-2 cell proliferation. Furthermore, kaempferol had a significantly less cytotoxicity than 5-FU in normal human pancreatic ductal epithelial cells (P=0.029). In both MIA PaCa-2 and Panc-1 cells, apoptotic cell population was increased when treated with kaempferol in a concentration-dependent manner. Conclusions Ginkgo biloba extract kaempferol effectively inhibits pancreatic cancer cell proliferation and induces cancer cell apoptosis, which may sensitize pancreatic tumor cells to chemotherapy. Kaempferol may have clinical applications as adjuvant therapy in the treatment of pancreatic cancer. PMID:18570926

  6. New castanospermine glycoside analogues inhibit breast cancer cell proliferation and induce apoptosis without affecting normal cells.

    Directory of Open Access Journals (Sweden)

    Ghada Allan

    Full Text Available sp²-Iminosugar-type castanospermine analogues have been shown to exhibit anti-tumor activity. However, their effects on cell proliferation and apoptosis and the molecular mechanism at play are not fully understood. Here, we investigated the effect of two representatives, namely the pseudo-S- and C-octyl glycoside 2-oxa-3-oxocastanospermine derivatives SO-OCS and CO-OCS, on MCF-7 and MDA-MB-231 breast cancer and MCF-10A mammary normal cell lines. We found that SO-OCS and CO-OCS inhibited breast cancer cell viability in a concentration- and time-dependent manner. This effect is specific to breast cancer cells as both molecules had no impact on normal MCF-10A cell proliferation. Both drugs induced a cell cycle arrest. CO-OCS arrested cell cycle at G1 and G2/M in MCF-7 and MDA-MB-231 cells respectively. In MCF-7 cells, the G1 arrest is associated with a reduction of CDK4 (cyclin-dependent kinase 4, cyclin D1 and cyclin E expression, pRb phosphorylation, and an overexpression of p21(Waf1/Cip1. In MDA-MB-231 cells, CO-OCS reduced CDK1 but not cyclin B1 expression. SO-OCS accumulated cells in G2/M in both cell lines and this blockade was accompanied by a decrease of CDK1, but not cyclin B1 expression. Furthermore, both drugs induced apoptosis as demonstrated by the increased percentage of annexin V positive cells and Bax/Bcl-2 ratio. Interestingly, in normal MCF-10A cells the two drugs failed to modify cell proliferation, cell cycle progression, cyclins, or CDKs expression. These results demonstrate that the effect of CO-OCS and SO-OCS is triggered by both cell cycle arrest and apoptosis, suggesting that these castanospermine analogues may constitute potential anti-cancer agents against breast cancer.

  7. [Effects of three Wenyang Jianpi Tang on cell proliferation and apoptosis of nonalcoholic fatty liver cells].

    Science.gov (United States)

    Yang, Jia-Yao; Tao, Dong-Qing; Liu, Song; Zhang, Shu; Ma, Wei; Shi, Zhao-Hong

    2017-04-01

    To investigate the effects of Sijunzi Tang, Lizhong Tang and Fuzi Lizhong Tang on the cell proliferation and apoptosis of nonalcoholic fatty liver cells through the nonalcoholic fatty liver cell model established by inducing L02 cells with oleic acid. Different concentrations of oleic acid were added into L02 cells to induce the nonalcoholic fatty liver cell model. Oil red O staining was used to observe fatty droplets of fatty liver cells. Automatic biochemical analyzer was used to detect the levels of aspartic transaminase(AST), alanine aminotransferase(ALT), total cholesterol(TC), and triglyceride(TG) in the cell supernatants. There were five groups, namely normal group, model group, model and Sijunzi Tang group, model and Lizhong Tang group, and model and Fuzi Lizhong Tang group. The cell proliferation and apoptosis of the five groups were detected by MTT colorimetry test and flow cytometer. The expressions of PCNA, cleaved caspase-3, cleaved caspase-8, cleaved caspase-9, Bax and Bcl-2 proteins of the five groups were detected by Western blot. The oil red O staining results showed that the optimum concentration of oleic acid that was used to induce nonalcoholic fatty liver cell models was 80 mg•L-1. The levels of AST, ALT, TC and TG in the nonalcoholic fatty liver cell supernatants were higher than that in normal liver cell supernatants(PTang, Lizhong Tang and Fuzi Lizhong Tang could effectively promote the cell proliferation, and inhibit the cellular apoptosis of nonalcoholic fatty liver cells(PTang showed the best effect. Western blot results showed that Sijunzi Tang, Lizhong Tang and Fuzi Lizhong Tang could down-regulate the expressions of cleaved caspase-3, cleaved caspase-8, cleaved caspase-9 and Bax proteins, and up-regulate the expressions of PCNA and Bcl-2 proteins of nonalcoholic fatty liver cells. And Fuzi Lizhong Tang showed the best effect. In conclusion, all of Sijunzi Tang, Lizhong Tang and Fuzi Lizhong Tang could effectively promote the cell

  8. Characterization of intravitreally delivered capsid mutant AAV2-Cre vector to induce tissue-specific mutations in murine retinal ganglion cells.

    Science.gov (United States)

    Langouet-Astrie, Christophe J; Yang, Zhiyong; Polisetti, Sraavya M; Welsbie, Derek S; Hauswirth, William W; Zack, Donald J; Merbs, Shannath L; Enke, Raymond A

    2016-10-01

    Targeted expression of Cre recombinase in murine retinal ganglion cells (RGCs) by viral vector is an effective strategy for creating tissue-specific gene knockouts for investigation of genetic contribution to RGC degeneration associated with optic neuropathies. Here we characterize dosage, efficacy and toxicity for sufficient intravitreal delivery of a capsid mutant Adeno-associated virus 2 (AAV2) vector encoding Cre recombinase. Wild type and Rosa26 (R26) LacZ mice were intravitreally injected with capsid mutant AAV2 viral vectors. Murine eyes were harvested at intervals ranging from 2 weeks to 15 weeks post-injection and were assayed for viral transduction, transgene expression and RGC survival. 10(9) vector genomes (vg) were sufficient for effective in vivo targeting of murine ganglion cell layer (GCL) retinal neurons. Transgene expression was observed as early as 2 weeks post-injection of viral vectors and persisted to 11 weeks. Early expression of Cre had no significant effect on RGC survival, while significant RGC loss was detected beginning 5 weeks post-injection. Early expression of viral Cre recombinase was robust, well-tolerated and predominantly found in GCL neurons suggesting this strategy can be effective in short-term RGC-specific mutation studies in experimental glaucoma models such as optic nerve crush and transection experiments. RGC degeneration with Cre expression for more than 4 weeks suggests that Cre toxicity is a limiting factor for targeted mutation strategies in RGCs. Copyright © 2016 Elsevier Ltd. All rights reserved.

  9. Variations of retinal nerve fiber layer thickness and ganglion cell-inner plexiform layer thickness according to the torsion direction of optic disc.

    Science.gov (United States)

    Lee, Kang Hoon; Kim, Chan Yun; Kim, Na Rae

    2014-02-20

    To examine the relationship between the optic disc torsion and peripapillary retinal nerve fiber layer (RNFL) thickness through a comparison with the macular ganglion cell inner plexiform layer complex (GCIPL) thickness measured by Cirrus optical coherence tomography (OCT). Ninety-four eyes of 94 subjects with optic disc torsion and 114 eyes of 114 subjects without optic disc torsion were enrolled prospectively. The participants underwent fundus photography and OCT imaging in peripapillary RNFL mode and macular GCIPL mode. The participants were divided into groups according to the presence or absence of optic disc torsion. The eyes with optic disc torsion were further divided into supranasal torsion and inferotemporal torsion groups according to the direction of optic disc torsion. The mean RNFL and GCIPL thicknesses for the quadrants and subsectors were compared. The superior and inferior peak locations of the RNFL were also measured according to the torsion direction. The temporal RNFL thickness was significantly thicker in inferotemporal torsion, whereas the GCIPL thickness at all segments was unaffected. The inferotemporal optic torsion had more temporally positioned superior peak locations of the RNFL than the nontorsion and supranasal-torted optic disc. Thickening of the temporal RNFL with a temporal shift in the superior peak within the eyes with inferotemporal optic disc torsion can lead to interpretation errors. The ganglion cell analysis algorithm can assist in differentiating eyes with optic disc torsion.

  10. The role of cPLA2 in Methylglyoxal-induced cell apoptosis of HUVECs

    International Nuclear Information System (INIS)

    Yuan, Jie; Zhu, Chao; Hong, Yali; Sun, Zongxing; Fang, Xianjun; Wu, Biao; Li, Shengnan

    2017-01-01

    Methylglyoxal (MGO), a highly reactive dicarbonyl compound, is mainly formed as a byproduct of glycolysis. Elevated MGO level is known to induce apoptosis of vascular endothelial cells, which is implicated with progression of atherosclerosis and diabetic complications. However, the underlying mechanisms have not been exhaustively investigated yet. Here, we further characterized the mechanisms how MGO induced apoptosis in human umbilical vein endothelial cells (HUVECs). Our data revealed that cytosolic phospholipase A2 (cPLA2) played an important role in MGO-induced cell apoptosis. It was found that MGO could increase both the activity and expression of cPLA2. Inhibition of cPLA2 by Pyrrophenone (PYR) or siRNA significantly attenuated the MGO-induced apoptosis. Additionally, MGO time-dependently decreased the phosphorylation of nuclear factor κB (NF-κB). Pretreatment of the cells with NF-κB inhibitor, BAY11-7082, further increased MGO-induced apoptosis of HUVECs, indicating that NF-κB played a survival role in this MGO-induced apoptosis. Furthermore, in the presence of si-cPLA2 or PYR, MGO no longer decreased NF-κB phosphorylation. Beyond that, the antioxidant N-acetyl cysteine (NAC) could reverse the changes of both cPLA2 and NF-κB caused by MGO. p38, the upstream of cPLA2, was also significantly phosphorylated by MGO. However, p38 inhibitor failed to reverse the apoptosis induced by MGO. This study gives an important insight into the downstream signaling mechanisms of MGO, cPLA2-NF-κB, in endothelial apoptosis. - Highlights: • cPLA2 participated in MGO-induced HUVECs apoptosis. • Inhibition of NF-κB was involved in MGO-cPLA2-mediated cell apoptosis. • Antioxidant NAC attenuated MGO-induced cPLA2 activation and cell apoptosis.

  11. The role of cPLA2 in Methylglyoxal-induced cell apoptosis of HUVECs

    Energy Technology Data Exchange (ETDEWEB)

    Yuan, Jie; Zhu, Chao; Hong, Yali; Sun, Zongxing; Fang, Xianjun [Jiangsu Provincial Key Lab of Cardiovascular Diseases and Molecular intervention, Department of Pharmacology, Nanjing Medical University, Nanjing 210029 (China); Wu, Biao, E-mail: wubiao@ncu.edu.cn [Department of Surgery, The First Affiliated Hospital, Nanchang University (China); Li, Shengnan, E-mail: snli@njmu.edu.cn [Jiangsu Provincial Key Lab of Cardiovascular Diseases and Molecular intervention, Department of Pharmacology, Nanjing Medical University, Nanjing 210029 (China)

    2017-05-15

    Methylglyoxal (MGO), a highly reactive dicarbonyl compound, is mainly formed as a byproduct of glycolysis. Elevated MGO level is known to induce apoptosis of vascular endothelial cells, which is implicated with progression of atherosclerosis and diabetic complications. However, the underlying mechanisms have not been exhaustively investigated yet. Here, we further characterized the mechanisms how MGO induced apoptosis in human umbilical vein endothelial cells (HUVECs). Our data revealed that cytosolic phospholipase A2 (cPLA2) played an important role in MGO-induced cell apoptosis. It was found that MGO could increase both the activity and expression of cPLA2. Inhibition of cPLA2 by Pyrrophenone (PYR) or siRNA significantly attenuated the MGO-induced apoptosis. Additionally, MGO time-dependently decreased the phosphorylation of nuclear factor κB (NF-κB). Pretreatment of the cells with NF-κB inhibitor, BAY11-7082, further increased MGO-induced apoptosis of HUVECs, indicating that NF-κB played a survival role in this MGO-induced apoptosis. Furthermore, in the presence of si-cPLA2 or PYR, MGO no longer decreased NF-κB phosphorylation. Beyond that, the antioxidant N-acetyl cysteine (NAC) could reverse the changes of both cPLA2 and NF-κB caused by MGO. p38, the upstream of cPLA2, was also significantly phosphorylated by MGO. However, p38 inhibitor failed to reverse the apoptosis induced by MGO. This study gives an important insight into the downstream signaling mechanisms of MGO, cPLA2-NF-κB, in endothelial apoptosis. - Highlights: • cPLA2 participated in MGO-induced HUVECs apoptosis. • Inhibition of NF-κB was involved in MGO-cPLA2-mediated cell apoptosis. • Antioxidant NAC attenuated MGO-induced cPLA2 activation and cell apoptosis.

  12. Wnt1 from cochlear schwann cells enhances neuronal differentiation of transplanted neural stem cells in a rat spiral ganglion neuron degeneration model.

    Science.gov (United States)

    He, Ya; Zhang, Peng-Zhi; Sun, Dong; Mi, Wen-Juan; Zhang, Xin-Yi; Cui, Yong; Jiang, Xing-Wang; Mao, Xiao-Bo; Qiu, Jian-Hua

    2014-04-01

    Although neural stem cell (NSC) transplantation is widely expected to become a therapy for nervous system degenerative diseases and injuries, the low neuronal differentiation rate of NSCs transplanted into the inner ear is a major obstacle for the successful treatment of spiral ganglion neuron (SGN) degeneration. In this study, we validated whether the local microenvironment influences the neuronal differentiation of transplanted NSCs in the inner ear. Using a rat SGN degeneration model, we demonstrated that transplanted NSCs were more likely to differentiate into microtubule-associated protein 2 (MAP2)-positive neurons in SGN-degenerated cochleae than in control cochleae. Using real-time quantitative PCR and an immunofluorescence assay, we also proved that the expression of Wnt1 (a ligand of Wnt signaling) increases significantly in Schwann cells in the SGN-degenerated cochlea. We further verified that NSC cultures express receptors and signaling components for Wnts. Based on these expression patterns, we hypothesized that Schwann cell-derived Wnt1 and Wnt signaling might be involved in the regulation of the neuronal differentiation of transplanted NSCs. We verified our hypothesis in vitro using a coculture system. We transduced a lentiviral vector expressing Wnt1 into cochlear Schwann cell cultures and cocultured them with NSC cultures. The coculture with Wnt1-expressing Schwann cells resulted in a significant increase in the percentage of NSCs that differentiated into MAP2-positive neurons, whereas this differentiation-enhancing effect was prevented by Dkk1 (an inhibitor of the Wnt signaling pathway). These results suggested that Wnt1 derived from cochlear Schwann cells enhanced the neuronal differentiation of transplanted NSCs through Wnt signaling pathway activation. Alterations of the microenvironment deserve detailed investigation because they may help us to conceive effective strategies to overcome the barrier of the low differentiation rate of transplanted

  13. Wavelength-dependent backscattering measurements for quantitative monitoring of apoptosis, Part 1: early and late spectral changes are indicative of the presence of apoptosis in cell cultures

    Science.gov (United States)

    Mulvey, Christine S.; Zhang, Kexiong; Liu, Wei-Han Bobby; Waxman, David J.; Bigio, Irving J.

    2011-11-01

    Apoptosis, a form of programmed cell death with unique morphological and biochemical features, is dysregulated in cancer and is activated by many cancer chemotherapeutic drugs. Noninvasive assays for apoptosis in cell cultures can aid in screening of new anticancer agents. We have previously demonstrated that elastic scattering spectroscopy can monitor apoptosis in cell cultures. In this report we present data on monitoring the detailed time-course of scattering changes in a Chinese hamster ovary (CHO) and PC-3 prostate cancer cells treated with staurosporine to induce apoptosis. Changes in the backscattering spectrum are detectable within 10 min, and continue to progress up to 48 h after staurosporine treatment, with the magnitude and kinetics of scattering changes dependent on inducer concentration. Similar responses were observed in CHO cells treated with several other apoptosis-inducing protocols. Early and late scattering changes were observed under conditions shown to induce apoptosis via caspase activity assay and were absent under conditions where apoptosis was not induced. Finally, blocking caspase activity and downstream apoptotic morphology changes prevented late scattering changes. These observations demonstrate that early and late changes in wavelength-dependent backscattering correlate with the presence of apoptosis in cell cultures and that the late changes are specific to apoptosis.

  14. Solanine induced apoptosis and increased chemosensitivity to Adriamycin in T-cell acute lymphoblastic leukemia cells.

    Science.gov (United States)

    Yi, Ying-Jie; Jia, Xiu-Hong; Wang, Jian-Yong; Chen, Jie-Ru; Wang, Hong; Li, You-Jie

    2018-05-01

    Solanine is an alkaloid and is the main extract of the traditional Chinese herb, Solanum nigrum Linn . It has been reported that Solanine has anti-inflammatory and antitumor properties. The present study aimed to investigate the antitumor effect of Solanine in Jurkat cells and demonstrate the molecular mechanism of antitumor activity of Solanine. A Cell Counting Kit-8 assay demonstrated that Solanine inhibited the proliferation of Jurkat cells in a dose-and time-dependent manner. Cell apoptosis was measured by flow cytometry. Flow cytometry revealed that Solanine induced apoptosis in a dose-dependent manner in Jurkat cells. Reverse transcription-quantitative polymerase chain reaction demonstrated that Solanine modulated the mRNA levels of B-cell lymphoma-2 (Bcl-2) and Bcl-2-associated X protein (Bax). Additionally, Bcl-2 and Bax expression was measured using western blot analysis. Western blot analysis revealed a significant increase in the expression of Bax and decrease in the expression of Bcl-2. Solanine increased the chemosensitivity of Jurkat cells to Adriamycin. In summary, the present results indicated that the antitumor activity of Solanine was associated with inhibition of cell proliferation, induction of apoptosis and increasing cytotoxicity of Adriamycin. Therefore, Solanine may have potential as a novel agent for the treatment of acute lymphocytic leukemia.

  15. Microsporidia infection impacts the host cell's cycle and reduces host cell apoptosis

    Science.gov (United States)

    Higes, Mariano; Sagastume, Soledad; Juarranz, Ángeles; Dias-Almeida, Joyce; Budge, Giles E.; Meana, Aránzazu; Boonham, Neil

    2017-01-01

    Intracellular parasites can alter the cellular machinery of host cells to create a safe haven for their survival. In this regard, microsporidia are obligate intracellular fungal parasites with extremely reduced genomes and hence, they are strongly dependent on their host for energy and resources. To date, there are few studies into host cell manipulation by microsporidia, most of which have focused on morphological aspects. The microsporidia Nosema apis and Nosema ceranae are worldwide parasites of honey bees, infecting their ventricular epithelial cells. In this work, quantitative gene expression and histology were studied to investigate how these two parasites manipulate their host’s cells at the molecular level. Both these microsporidia provoke infection-induced regulation of genes involved in apoptosis and the cell cycle. The up-regulation of buffy (which encodes a pro-survival protein) and BIRC5 (belonging to the Inhibitor Apoptosis protein family) was observed after infection, shedding light on the pathways that these pathogens use to inhibit host cell apoptosis. Curiously, different routes related to cell cycle were modified after infection by each microsporidia. In the case of N. apis, cyclin B1, dacapo and E2F2 were up-regulated, whereas only cyclin E was up-regulated by N. ceranae, in both cases promoting the G1/S phase transition. This is the first report describing molecular pathways related to parasite-host interactions that are probably intended to ensure the parasite’s survival within the cell. PMID:28152065

  16. Antiproliferative and cell apoptosis-inducing activities of compounds from Buddleja davidii in Mgc-803 cells

    Directory of Open Access Journals (Sweden)

    Wu Jian

    2012-08-01

    Full Text Available Abstract Background Buddleja davidii is widely distributed in the southwestern region of China. We have undertaken a systematic analysis of B. davidii as a Chinese traditional medicine with anticancer activity by isolating natural products for their activity against the human gastric cancer cell line Mgc-803 and the human breast cancer cell line Bcap-37. Results Ten compounds were extracted and isolated from B. davidii, among which colchicine was identified in B. davidii for the first time. The inhibitory activities of these compounds were investigated in Mgc-803, Bcap-37 cells in vitro by MTT [3-(4,5-dimethylthiazol-2-yl-2,5-diphenyltetrazolium bromide] assay, and the results showed that luteolin and colchicine had potent inhibitory activities against the growth of Mgc-803 cells. Subsequent fluorescence staining and flow cytometry analysis indicated that these two compounds could induce apoptosis in Mgc-803 cells. The results also showed that the percentages of early apoptotic cells (Annexin V+/PI-, where PI is propidium iodide and late apoptotic cells (Annexin V+/PI+ increased in a dose- and time-dependent manner. After 36 h of incubation with luteolin at 20 μM, the percentages of cells were approximately 15.4% in early apoptosis and 43.7% in late apoptosis; after 36 h of incubation with colchicine at 20 μM, the corresponding values were 7.7% and 35.2%, respectively. Conclusions Colchicine and luteolin from B. davidii have potential applications as adjuvant therapies for treating human carcinoma cells. These compounds could also induce apoptosis in tumor cells.

  17. Antiproliferative and cell apoptosis-inducing activities of compounds from Buddleja davidii in Mgc-803 cells.

    Science.gov (United States)

    Wu, Jian; Yi, Wenshi; Jin, Linhong; Hu, Deyu; Song, Baoan

    2012-08-31

    Buddleja davidii is widely distributed in the southwestern region of China. We have undertaken a systematic analysis of B. davidii as a Chinese traditional medicine with anticancer activity by isolating natural products for their activity against the human gastric cancer cell line Mgc-803 and the human breast cancer cell line Bcap-37. Ten compounds were extracted and isolated from B. davidii, among which colchicine was identified in B. davidii for the first time. The inhibitory activities of these compounds were investigated in Mgc-803, Bcap-37 cells in vitro by MTT [3-(4,5-dimethylthiazol-2-yl)-2,5-diphenyltetrazolium bromide] assay, and the results showed that luteolin and colchicine had potent inhibitory activities against the growth of Mgc-803 cells. Subsequent fluorescence staining and flow cytometry analysis indicated that these two compounds could induce apoptosis in Mgc-803 cells. The results also showed that the percentages of early apoptotic cells (Annexin V+/PI-, where PI is propidium iodide) and late apoptotic cells (Annexin V+/PI+) increased in a dose- and time-dependent manner. After 36 h of incubation with luteolin at 20 μM, the percentages of cells were approximately 15.4% in early apoptosis and 43.7% in late apoptosis; after 36 h of incubation with colchicine at 20 μM, the corresponding values were 7.7% and 35.2%, respectively. Colchicine and luteolin from B. davidii have potential applications as adjuvant therapies for treating human carcinoma cells. These compounds could also induce apoptosis in tumor cells.

  18. Delayed administration of glial cell line-derived neurotrophic factor (GDNF) protects retinal ganglion cells in a pig model of acute retinal ischemia

    DEFF Research Database (Denmark)

    Kyhn, Maria Voss; Klassen, Henry; Johansson, Ulrica Englund

    2009-01-01

    electroretinography (mfERG), quantification of NeuN positive cells and evaluation of the degree of retinal perivasculitis and inflammation 6 weeks after the insult. In the post-injection eyes (days 14, 28 and 42), the ratios of the iN1 and the iP2 amplitudes were 0.10 (95% CI: 0.05-0.15) and 0.09 (95% CI: 0.......04-0.16) in eyes treated with blank microspheres, and 0.24 (95% CI: 0.18-0.32) and 0.23 (95% CI: 0.15-0.33) in eyes treated with GDNF microspheres. These differences were statistically significant (P eyes...... injected with GDNF microspheres compared to eyes injected with blank microspheres. In eyes injected with GDNF microspheres the ganglion cell count was 9.5/field (s.e.m.: 2.1, n = 8), in eyes injected with blank microspheres it was 3.5/field (s.e.m.: 1.2, n = 7). This difference was statistically...

  19. TGEV nucleocapsid protein induces cell cycle arrest and apoptosis through activation of p53 signaling

    International Nuclear Information System (INIS)

    Ding, Li; Huang, Yong; Du, Qian; Dong, Feng; Zhao, Xiaomin; Zhang, Wenlong; Xu, Xingang; Tong, Dewen

    2014-01-01

    Highlights: • TGEV N protein reduces cell viability by inducing cell cycle arrest and apoptosis. • TGEV N protein induces cell cycle arrest and apoptosis by regulating p53 signaling. • TGEV N protein plays important roles in TGEV-induced cell cycle arrest and apoptosis. - Abstract: Our previous studies showed that TGEV infection could induce cell cycle arrest and apoptosis via activation of p53 signaling in cultured host cells. However, it is unclear which viral gene causes these effects. In this study, we investigated the effects of TGEV nucleocapsid (N) protein on PK-15 cells. We found that TGEV N protein suppressed cell proliferation by causing cell cycle arrest at the S and G2/M phases and apoptosis. Characterization of various cellular proteins that are involved in regulating cell cycle progression demonstrated that the expression of N gene resulted in an accumulation of p53 and p21, which suppressed cyclin B1, cdc2 and cdk2 expression. Moreover, the expression of TGEV N gene promoted translocation of Bax to mitochondria, which in turn caused the release of cytochrome c, followed by activation of caspase-3, resulting in cell apoptosis in the transfected PK-15 cells following cell cycle arrest. Further studies showed that p53 inhibitor attenuated TGEV N protein induced cell cycle arrest at S and G2/M phases and apoptosis through reversing the expression changes of cdc2, cdk2 and cyclin B1 and the translocation changes of Bax and cytochrome c induced by TGEV N protein. Taken together, these results demonstrated that TGEV N protein might play an important role in TGEV infection-induced p53 activation and cell cycle arrest at the S and G2/M phases and apoptosis occurrence

  20. Feedback regulation of mitochondria by caspase-9 in the B cell receptor-mediated apoptosis.

    Science.gov (United States)

    Eeva, J; Nuutinen, U; Ropponen, A; Mättö, M; Eray, M; Pellinen, R; Wahlfors, J; Pelkonen, J

    2009-12-01

    During the germinal centre reaction (GC), B cells with non-functional or self-reactive antigen receptors are negatively selected by apoptosis to generate B cell repertoire with appropriate antigen specificities. We studied the molecular mechanism of Fas/CD95- and B cell receptor (BCR)-induced apoptosis to shed light on the signalling events involved in the negative selection of GC B cells. As an experimental model, we used human follicular lymphoma (FL) cell line HF1A3, which originates from a GC B cell, and transfected HF1A3 cell lines overexpressing Bcl-x(L), c-FLIP(long) or dominant negative (DN) caspase-9. Fas-induced apoptosis was dependent on the caspase-8 activation, since the overexpression of c-FLIP(long), a natural inhibitor of caspase-8 activation, blocked apoptosis induced by Fas. In contrast, caspase-9 activation was not involved in Fas-induced apoptosis. BCR-induced apoptosis showed the typical characteristics of mitochondria-dependent (intrinsic) apoptosis. Firstly, the activation of caspase-9 was involved in BCR-induced DNA fragmentation, while caspase-8 showed only marginal role. Secondly, overexpression of Bcl-x(L) could block all apoptotic changes induced by BCR. As a novel finding, we demonstrate that caspase-9 can enhance the cytochrome-c release and collapse of mitochondrial membrane potential (DeltaPsi(m)) during BCR-induced apoptosis. The requirement of different signalling pathways in apoptosis induced by BCR and Fas may be relevant, since Fas- and BCR-induced apoptosis can thus be regulated independently, and targeted to different subsets of GC B cells.

  1. Effects of HSP27 chaperone on THP-1 tumor cell apoptosis.

    Science.gov (United States)

    Kaigorodova, E V; Ryazantseva, N V; Novitskii, V V; Maroshkina, A N; Belkina, M V

    2012-11-01

    The role of Hsp27 (heat shock protein 27) chaperone in regulation of THP-1 tumor cell apoptosis was studied. Realization of tumor cell apoptosis under conditions of in vitro culturing with Hsp27 specific inhibitor (KRIBB3) was evaluated by fluorescent microscopy with FITC-labeled annexin V and propidium iodide. Measurements of Bcl-2 family proteins (Bcl-2, Bax, Bad) in tumor cells incubated with Hsp27 inhibitor were carried out by Western blotting. Chaperone Hsp27 acted as apoptosis inhibitor in THP-1 tumor cells modulating the proportion of antiapoptotic (Bcl-2) and proapoptotic (Bax and Bad) proteins.

  2. Characterization of chlorpyrifos-induced apoptosis in placental cells

    International Nuclear Information System (INIS)

    Saulsbury, Marilyn D.; Heyliger, Simone O.; Wang, Kaiyu; Round, Dorothy

    2008-01-01

    The mechanism by which chlorpyrifos exerts its toxicity in fetal and perinatal animals has yet to be elucidated. Since the placenta is responsible for transport of nutrients and is a major supplier hormone to the fetus, exposure to xenobiotics that alter the function or viability of placenta cells could ostensibly alter the development of the fetus. In this study, JAR cells were used to determine if CPF and the metabolites 3,5,6-trichloro-2-pyridinol (TCP) and chlorpyrifos-oxon (CPO) are toxic to the placenta. Our results indicate that chlorpyrifos (CPF), and its metabolite chlorpyrifos-oxon (CPO) caused a dose-dependent reduction in cellular viability with CPF being more toxic than its metabolites. Chlorpyrifos-induced toxicity was characterized by the loss of mitochondrial potential, the appearance of nuclear condensation and fragmentation, down-regulation of Bcl-2 as well as up-regulation of TNFα and FAS mRNA. Pharmacological inhibition of FAS, nicotinic and TNF-α receptors did not attenuate CPF-induced toxicity. Atropine exhibited minimal ability to reverse toxicity. Furthermore, signal transduction inhibitors PD98059, SP600125, LY294002 and U0126 failed to attenuate toxicity; however, SB202190 (inhibitor of p38α and p38ss MAPK) sensitized cells to CPF-induced toxicity. Pan-caspase inhibitor Q-VD-OPh produced a slight but significant reversal of CPF-induced toxicity indicating that the major caspase pathways are not integral to CPF-induced toxicity. Taken collectively, these results suggest that chlorpyrifos induces apoptosis in placental cells through pathways not dependent on FAS/TNF signaling, activation of caspases or inhibition of cholinesterase. In addition, our data further indicates that activation of p38 MAPK is integral to the protection cells against CPF-induced injury

  3. Apoptosis in differentiating C2C12 muscle cells selectively targets Bcl-2-deficient myotubes

    Science.gov (United States)

    Schoneich, Christian; Dremina, Elena; Galeva, Nadezhda; Sharov, Victor

    2014-01-01

    Muscle cell apoptosis accompanies normal muscle development and regeneration, as well as degenerative diseases and aging. C2C12 murine myoblast cells represent a common model to study muscle differentiation. Though it was already shown that myogenic differentiation of C2C12 cells is accompanied by enhanced apoptosis in a fraction of cells, either the cell population sensitive to apoptosis or regulatory mechanisms for the apoptotic response are unclear so far. In the current study we characterize apoptotic phenotypes of different types of C2C12 cells at all stages of differentiation, and report here that myotubes of differentiated C2C12 cells with low levels of anti-apoptotic Bcl-2 expression are particularly vulnerable to apoptosis even though they are displaying low levels of pro-apoptotic proteins Bax, Bak and Bad. In contrast, reserve cells exhibit higher levels of Bcl-2 and high resistance to apoptosis. The transfection of proliferating myoblasts with Bcl-2 prior to differentiation did not protect against spontaneous apoptosis accompanying differentiation of C2C12 cell but led to Bcl-2 overexpression in myotubes and to significant protection from apoptotic cell loss caused by exposure to hydrogen peroxide. Overall, our data advocate for a Bcl-2-dependent mechanism of apoptosis in differentiated muscle cells. However, downstream processes for spontaneous and hydrogen peroxide induced apoptosis are not completely similar. Apoptosis in differentiating myoblasts and myotubes is regulated not through interaction of Bcl-2 with pro-apoptotic Bcl-2 family proteins such as Bax, Bak, and Bad. PMID:24129924

  4. Effects of biodegradable Mg–6Zn alloy extracts on apoptosis of intestinal epithelial cells

    International Nuclear Information System (INIS)

    Wang Zhanhui; Yan Jun; Li Jianan; Zheng Qi; Wang Zhigang; Zhang Xiaonong; Zhang Shaoxiang

    2012-01-01

    Highlights: ► We evaluated the effects of Mg–6Zn alloys on apoptosis of IEC-6 cells. ► The apoptosis was evaluated by investigating the expression of caspase-1 and Bcl-2. ► The IEC-6 cells displayed better cell functions in 60% or 20% extract. ► The conspicuous alkaline environment is disadvantageous to apoptosis of IEC cells. ► The excessive Mg concentration is disadvantageous to apoptosis of IEC-6 cells. - Abstract: In this study, intestinal epithelial cells (IEC)-6 were cultured in different concentration extracts of Mg–6Zn alloys for different time periods. To achieve a total of three concentrations (100%, 60% and 20% concentration), the extracts were serially diluted with Dulbecco's modified Eagle medium High Glucose to observe a dose–response relationship. We studied the indirect effects of Mg–6Zn alloys on IEC-6 cells apoptosis. The apoptosis of IEC-6 cells was measured using flow cytometry. And the apoptosis of IEC-6 cells was evaluated by investigating the expression of caspase-1and Bcl-2 using real-time polymerase chain reaction (PCR) and Western blotting tests. It was found that the levels of apoptosis in IEC-6 cells cultured in 100% Mg–6Zn alloy extracts were significantly higher than those in 60% and 20% extracts; the 100% extract can down-regulate expression of Bcl-2 after culture. The in vitro results indicated that the conspicuous alkaline environment and excessive Mg concentration, even Zn concentration caused by rapid corrosion of Mg–6Zn alloys promote IEC-6 cells apoptosis, although further experiments will be necessary to formally prove our conclusions. Therefore, the adjustment of the degradation rate is needed for using Mg–Zn alloy as a surgical suture material.

  5. Interferon-gamma sensitizes colonic epithelial cell lines to physiological and therapeutic inducers of colonocyte apoptosis.

    LENUS (Irish Health Repository)

    O'Connell, J

    2012-02-03

    Homeostasis in the colonic epithelium is achieved by a continuous cycle of proliferation and apoptosis, in which imbalances are associated with disease. Inflammatory bowel disease (IBD) and colon cancer are associated with either excessive or insufficient apoptosis of colonic epithelial cells, respectively. By using two colonic epithelial cell lines, HT29 and SW620, we investigated how the epithelial cell\\'s sensitivity to apoptosis was regulated by the proinflammatory cytokine interferon-gamma (IFN-gamma). We found that IFN-gamma sensitized HT29 cells, and to a lesser extent SW620, to diverse inducers of apoptosis of physiologic or therapeutic relevance to the colon. These apoptosis inducers included Fas (CD95\\/APO-1) ligand (FasL), short-chain fatty acids, and chemotherapeutic drugs. The extent of IFN-gamma-mediated apoptosis sensitization in these two cell lines correlated well with the degree of IFN-gamma-mediated upregulation of the proapoptotic protease caspase-1. Although IFN-gamma alone effectively sensitized HT29 cells to apoptosis, inclusion of the protein synthesis inhibitor cyclohexamide (CHX) during apoptotic challenge was necessary for maximal sensitization of SW620. The requirement of CHX to sensitize SW620 cells to apoptosis implies a need to inhibit translation of antiapoptotic proteins absent from HT29. In particular, the antiapoptotic protein Bcl-2 was strongly expressed in SW620 cells but absent from HT29. Our results indicate that IFN-gamma increases the sensitivity of colonic epithelial cells to diverse apoptotic stimuli in concert, via upregulation of caspase-1. Our findings implicate caspase-1 and Bcl-2 as important central points of control determining the general sensitivity of colonic epithelial cells to apoptosis.

  6. Radiation-induced apoptosis and cell cycle checkpoints in human colorectal tumour cell lines

    International Nuclear Information System (INIS)

    Playle, L.C.

    2001-03-01

    The p53 tumour suppressor gene is mutated in 75% of colorectal carcinomas and is critical for DNA damage-induced G1 cell cycle arrest. Data presented in this thesis demonstrate that after treatment with Ionizing Radiation (IR), colorectal tumour cell lines with mutant p53 are unable to arrest at G1 and undergo cell cycle arrest at G2. The staurosporine derivative, UCN-01, was shown to abrogate the IR-induced G2 checkpoint in colorectal tumour cell lines. Furthermore, in some cell lines, abrogation of the G2 checkpoint was associated with radiosensitisation. Data presented in this study demonstrate that 2 out of 5 cell lines with mutant p53 were sensitised to IR by UCN-01. In order to determine whether radiosensitisation correlated with lack of functional p53, transfected derivatives of an adenoma-derived cell line were studied, in which endogenous wild type p53 was disrupted by expression of a dominant negative p53 mutant protein (and with a vector control). In both these cell lines UCN-01 abrogated the G2 arrest however this was not associated with radiosensitisation, indicating that radiosensitisation is a cell type-specific phenomenon. Although 2 colorectal carcinoma cell lines, with mutant p53, were sensitised to IR by UCN-01, the mechanisms of p53-independent IR-induced apoptosis in the colon are essentially unknown. The mitogen-activated protein kinase (MAPK) pathways (that is the JNK, p38 and ERK pathways) have been implicated in apoptosis in a range of cell systems and in IR-induced apoptosis in some cell types. Data presented in this study show that, although the MAPKs can be activated by the known activator anisomycin, there is no evidence of a role for MAPKs in IR-induced apoptosis in colorectal tumour cell lines, regardless of p53 status. In summary, some colorectal tumour cell lines with mutant p53 can be sensitised to IR-induced cell death by G2 checkpoint abrogation and this may be an important treatment strategy, however mechanisms of IR-induced p53

  7. β-Catenin promotes cell proliferation, migration, and invasion but induces apoptosis in renal cell carcinoma

    Directory of Open Access Journals (Sweden)

    Yang CM

    2017-02-01

    Full Text Available Chun-ming Yang,1 Shan Ji,2 Yan Li,3 Li-ye Fu,3 Tao Jiang,3 Fan-dong Meng31Department of Urology, The First Affiliated Hospital, China Medical University, 2Department of Endocrinology, The Fifth People’s Hospital of Shenyang, 3Department of Biotherapy, Cancer Research Institute, The First Affiliated Hospital, China Medical University, Shenyang, ChinaAbstract: β-Catenin (CTNNB1 gene coding protein is a component of the Wnt signaling pathway that has been shown to play an important role in the formation of certain cancers. Abnormal accumulation of CTNNB1 contributes to most cancers. This research studied the involvement of β-catenin in renal cell carcinoma (RCC cell proliferation, apoptosis, migration, and invasion. Proliferation, cell cycle, and apoptosis were analyzed by using Cell Counting Kit-8 and by flow cytometry. Migration and invasion assays were measured by transwell analysis. Real-time polymerase chain reaction and Western blot analysis were used to detect the expression of CTNNB1, ICAM-1, VCAM-1, CXCR4, and CCL18 in RCC cell lines. It was found that CTNNB1 knockdown inhibited cell proliferation, migration, and invasion and induced apoptosis of A-498 cells. CTNNB1 overexpression promoted cell proliferation, migration, and invasion and inhibited apoptosis of 786-O cells. Moreover, knockdown of CTNNB1 decreased the levels of ICAM-1, VCAM-1, CXCR4, and CCL18 expression, but CTNNB1 overexpression increased the expression of ICAM-1, VCAM-1, CXCR4, and CCL18. Further in vivo tumor formation study in nude mice indicated that inhibition of CTNNB1 delayed the progress of tumor formation through inhibiting PCNA and Ki67 expression. These results indicate that CTNNB1 could act as an oncogene and may serve as a promising therapeutic strategy for RCC.Keywords: kidney cancer, oncogene, β-catenin, survival time, tumor migration-related protein

  8. Intracerebroventricular gene therapy that delays neurological disease progression is associated with selective preservation of retinal ganglion cells in a canine model of CLN2 disease.

    Science.gov (United States)

    Whiting, Rebecca E H; Jensen, Cheryl A; Pearce, Jacqueline W; Gillespie, Lauren E; Bristow, Daniel E; Katz, Martin L

    2016-05-01

    CLN2 disease is one of a group of lysosomal storage disorders called the neuronal ceroid lipofuscinoses (NCLs). The disease results from mutations in the TPP1 gene that cause an insufficiency or complete lack of the soluble lysosomal enzyme tripeptidyl peptidase-1 (TPP1). TPP1 is involved in lysosomal protein degradation, and lack of this enzyme results in the accumulation of protein-rich autofluorescent lysosomal storage bodies in numerous cell types including neurons throughout the central nervous system and the retina. CLN2 disease is characterized primarily by progressive loss of neurological functions and vision as well as generalized neurodegeneration and retinal degeneration. In children the progressive loss of neurological functions typically results in death by the early teenage years. A Dachshund model of CLN2 disease with a null mutation in TPP1 closely recapitulates the human disorder with a progression from disease onset at approximately 4 months of age to end-stage at 10-11 months. Delivery of functional TPP1 to the cerebrospinal fluid (CSF), either by periodic infusion of the recombinant protein or by a single administration of a TPP1 gene therapy vector to the CSF, significantly delays the onset and progression of neurological signs and prolongs life span but does not prevent the loss of vision or modest retinal degeneration that occurs by 11 months of age. In this study we found that in dogs that received the CSF gene therapy treatment, the degeneration of the retina and loss of retinal function continued to progress during the prolonged life spans of the treated dogs. Eventually the normal cell layers of the retina almost completely disappeared. An exception was the ganglion cell layer. In affected dogs that received TPP1 gene therapy to the CSF and survived an average of 80 weeks, ganglion cell axons were present in numbers comparable to those of normal Dachshunds of similar age. The selective preservation of the retinal ganglion cells suggests

  9. Increased autophagy in CD4(+) T cells of rheumatoid arthritis patients results in T-cell hyperactivation and apoptosis resistance

    NARCIS (Netherlands)

    van Loosdregt, Jorg; Rossetti, Maura; Spreafico, Roberto; Moshref, Maryam; Olmer, Merissa; Williams, Gary W; Kumar, Pavanish; Copeland, Dana; Pischel, Ken; Lotz, Martin; Albani, Salvatore

    2016-01-01

    Rheumatoid arthritis (RA) is an autoimmune disease hallmarked by aberrant cellular homeostasis, resulting in hyperactive CD4(+) T cells that are more resistant to apoptosis. Both hyperactivation and resistance to apoptosis may contribute to the pathogenicity of CD4(+) T cells in the autoimmune

  10. Apoptosis in chondrogenesis of human mesenchymal stem cells: effect of serum and medium supplements.

    Science.gov (United States)

    Wang, Chien-Yuan; Chen, Ling-Lan; Kuo, Pei-Yin; Chang, Jia-Ling; Wang, Yng-Jiin; Hung, Shih-Chieh

    2010-04-01

    Apoptosis is an inevitable process during development and is evident in the formation of articular cartilage and endochondral ossification of growth plate. Mesenchymal stem cells (MSCs) can serve as alternative sources for cell therapy in focal chondral lesions or diffuse osteoarthritis. But there are few, if any, studies investigating apoptosis during chondrogenesis by MSCs. The aim of this study was to find the better condition to prevent apoptosis during chondrogenesis by MSCs. Apoptosis were evaluated in MSCs induced in different chondrogenic media by the use of Annexin V, TUNEL staining, lysosomal labeling with lysotracker and immunostaining of apoptotic markers. We found apparent apoptosis was demonstrated by Annexin V, TUNEL staining and lysosomal labeling during chondrogenesis. Meanwhile, the degree of apoptosis was related to the reagents of the defined chondrogenic medium. Adding serum in medium increased apoptosis, however, TGF-beta1 inhibited apoptosis. The apoptosis was associated with the activation of caspase-3, the increase in the Bax/Bcl-2 ratio, the loss of lysosomal integrity, and the increase of PARP-cleavage. Pro-inflammatory cytokines, IL-1alpha, IL-1beta and TNFalpha did not induce any increase in apoptosis. Interestingly, the inhibition of apoptosis by serum free medium supplemented with ITS was also associated with an increase in the expression of type II collagen, and a decrease in the expression of type X collagen, Runx2, and other osteogenic genes, while TGF-beta1 increased the expression of Sox9, type II and type X collagen and decreased the expression of osteogenic genes. These data suggest apoptosis occurs during chondrogenesis by MSCs by cell death intrinsic pathway activation and this process may be modulated by culture conditions.

  11. Evaluation of apoptosis and apoptosis proteins as possible markers of radiation at doses 0.1-2 Gy, in comparison to the micronucleus assay in three cell lines

    International Nuclear Information System (INIS)

    Jaworska, A.; Angelis, P. de

    1997-01-01

    In recent years the interest in apoptosis as possible indicator of radiation damage has increased. Studies have been done to examine the induction of apoptosis after ionizing radiation using morphological criteria, characteristic DNA damage pattern(ladders), early DNA damage using flow cytometry and/or expression of the proteins involved in apoptosis. But the picture which emerges from these investigations is unclear. Some researchers suggest that apoptosis studies can be used as potential assays of biological dosimetry, others doubt if apoptosis can be used as a marker of irradiation at all. Most studies have been done using relatively high doses of radiation. In this study we focus on apoptosis induction after relatively small doses (0,1-2 Gy). We detected apoptosis with the in situ terminal deoxynucleotidyl transferase assay by flow cytometry, and measured the expression of proteins that regulate apoptosis (Bcl-2, Bax, P53) with Western blotting. As comparison we used the cytokinesis-block micronucleus assay as a reference. The studies were carried out in three lymphoid cell lines: the mouse lymphoma L5178Y resistant and sensitive cell lines widely used in radiobiological studies, and the human pre-B cell leukemia Reh cells. Our results indicate that we can not consider the examined parameters of apoptosis as markers of radiation in these cell lines. (author)

  12. Optical imaging the redox status change during cell apoptosis

    Science.gov (United States)

    Su, Ting; Zhang, Zhihong; Lin, Juqiang; Luo, Qingming

    2007-02-01

    Many cellular events involve the alteration in redox equilibrium, globally or locally. In many cases, excessive reactive oxygen species (ROS) production is the underlying cause. Several green fluoresecence protein based indicators are constructed to measure redox status in cells, e.g, rxYFP and roGFPs, which allow real time detection. reduction and oxidization-sensitive GFP (RoGFPs) are more useful due to ratiometric variation by excitation, making the measurement more accurate. Utilizing one of those roGFPs called roGFP1, we establish a mitochondrial redox state probing platform in HeLa cells with laser scan confocal microscopy (LSCM) as detection system. Control experiments confirmed that our platform could produce stable ratiometric values, which made the data more accurately reflect the real environmental changes of redox status that roGFP1 probed. Using exogenous H IIO II and DTT, we evaluated the reactivity and reversibility of roGFP1. The minimal hydrogen peroxide concentration that roGFP1 could show detectable ratiometric changes in our system was about 200μM. Preliminarily applying our platform to exploring the redox status during apoptosis, we observed an increase in ratiometric, suggesting an excessive ROS production.

  13. Graphene-induced apoptosis in lung epithelial cells through EGFR

    Science.gov (United States)

    Tsai, Shih-Ming; Bangalore, Preeti; Chen, Eric Y.; Lu, David; Chiu, Meng-Hsuen; Suh, Andrew; Gehring, Matthew; Cangco, John P.; Garcia, Santiago G.; Chin, Wei-Chun

    2017-07-01

    Expanding interest in nanotechnology applied to electronic and biomedical fields has led to fast-growing development of various nanomaterials. Graphene is a single-atom thick, two-dimensional sheet of hexagonally arranged carbon atoms with unique physical and chemical properties. Recently, graphene has been used in many studies on electronics, photonics, composite materials, energy generation and storage, sensors, and biomedicine. However, the current health risk assessment for graphene has been relatively limited and inconclusive. This study evaluated the toxicity effects of graphene on the airway epithelial cell line BEAS-2B, which represents the first barrier of the human body to interact with airborne graphene particles. Our result showed that graphene can induce the cellular Ca2+ by phospholipase C (PLC) associated pathway by activating epidermal growth factor receptor (EGFR). Subsequently, inositol 1,4,5-triphosphate (IP3) receptors activate the release of Ca2+ from the endoplasmic reticulum (ER) Ca2+ stores. Those Ca2+ signals further trigger the calcium-regulated apoptosis in the cell. Furthermore, the stimulation can cause EGFR upregulation, which have been demonstrated to associate with diseases such as lung cancer, chronic obstructive pulmonary disease (COPD), and cardiovascular diseases. This study highlights the additional health risk considering that it can function as a contributing factor for other respiratory diseases.

  14. Salidroside protects against foam cell formation and apoptosis, possibly via the MAPK and AKT signaling pathways.

    Science.gov (United States)

    Ni, Jing; Li, Yuanmin; Li, Weiming; Guo, Rong

    2017-10-10

    Foam cell formation and apoptosis are closely associated with atherosclerosis pathogenesis. We determined the effect of salidroside on oxidized low-density lipoprotein (ox-LDL)-induced foam cell formation and apoptosis in THP1 human acute monocytic leukemia cells and investigated the associated molecular mechanisms. THP1-derived macrophages were incubated with salidroside for 5 h and then exposed to ox-LDL for 24 h to induce foam cell formation. Cytotoxicity, lipid deposition, apoptosis, and the expression of various proteins were tested using the CCK8 kit, Oil Red O staining, flow cytometry, and western blotting, respectively. Ox-LDL treatment alone promoted macrophage-derived foam cell formation, while salidroside treatment alone inhibited it (p foam cell formation and apoptosis, partly by regulating the MAPK and Akt signaling pathways.

  15. MicroRNAs regulate B-cell receptor signaling-induced apoptosis

    NARCIS (Netherlands)

    Kluiver, J. L.; Chen, C-Z

    Apoptosis induced by B-cell receptor (BCR) signaling is critical for antigen-driven selection, a process critical to tolerance and immunity. Here, we examined the roles of microRNAs (miRNAs) in BCR signaling-induced apoptosis using the widely applied WEHI-231 model. Comparison of miRNA levels in

  16. Sensitivity of cells to apoptosis induced by iron deprivation can be reversibly changed by iron availability

    Czech Academy of Sciences Publication Activity Database

    Koc, Michal; Naďová, Zuzana; Kovář, Jan

    2006-01-01

    Roč. 39, č. 6 (2006), s. 551-561 ISSN 0960-7722 R&D Projects: GA AV ČR(CZ) KJB5052401 Institutional research plan: CEZ:AV0Z50520514 Keywords : apoptosis * iron deprivation * changed cell sensitivity to apoptosis by iron availability Subject RIV: EB - Genetics ; Molecular Biology Impact factor: 4.492, year: 2006

  17. Noxa/Mcl-1 Balance Regulates Susceptibility of Cells to Camptothecin-Induced Apoptosis

    Directory of Open Access Journals (Sweden)

    Yide Mei

    2007-10-01

    Full Text Available Although camptothecin (CPT has been reported to induce apoptosis in various cancer cells, the molecular details of this regulation remain largely unknown. In this study, we demonstrate that 131-113-only protein Noxa is upregulated during CPT-induced apoptosis, which is independent of p53. In addition, we show that phosphatidylinositol 3-kinase (PI3K/Akt signaling pathway is responsible for Noxa's induction. Luciferase assay, cAMP response element binding protein (CREB knockdown experiments further demonstrate that CREB is involved in the transcriptional upregulation of Noxa. Moreover, blocking Noxa expression using specific small interfering ribonucleic acid (siRNA significantly reduces the apoptosis in response to CPT, indicating that Noxa is an essential mediator for CPT-induced apoptosis. Interestingly, antiapoptotic Mcl-1 was also upregulated through PI3K/Akt signaling pathway upon CPT treatment. Using immunoprecipitation assay, Noxa was found to interact with Mcl-1 in the presence or absence of CPT. Knockdown of Mcl-1 expression by short hairpin ribonucleic acid (shRNA was shown to potentiate CPT-induced apoptosis. Consistently, ectopic overexpression of Mcl-1 rescued cells from apoptosis induced by CPT. Cells coexpressing Noxa, Mcl-1 at different ratio correlates well with the extent of apoptosis, suggesting that the balance between Noxa, Mcl-1 may determine the susceptibility of HeLa cells to CPT-induced apoptosis.

  18. Noxa/Mcl-1 Balance Regulates Susceptibility of Cells to Camptothecin-Induced Apoptosis1

    Science.gov (United States)

    Mei, Yide; Xie, Chongwei; Xie, Wei; Tian, Xu; Li, Mei; Wu, Mian

    2007-01-01

    Although camptothecin (CPT) has been reported to induce apoptosis in various cancer cells, the molecular details of this regulation remain largely unknown. In this study, we demonstrate that BH3-only protein Noxa is upregulated during CPT-induced apoptosis, which is independent of p53. In addition, we show that phosphatidylinositol 3-kinase (PI3K)/Akt signaling pathway is responsible for Noxa's induction. Luciferase assay and cAMP response element binding protein (CREB) knockdown experiments further demonstrate that CREB is involved in the transcriptional upregulation of Noxa. Moreover, blocking Noxa expression using specific small interfering ribonucleic acid (siRNA) significantly reduces the apoptosis in response to CPT, indicating that Noxa is an essential mediator for CPT-induced apoptosis. Interestingly, antiapoptotic Mcl-1 was also upregulated through PI3K/Akt signaling pathway upon CPT treatment. Using immunoprecipitation assay, Noxa was found to interact with Mcl-1 in the presence or absence of CPT. Knockdown of Mcl-1 expression by short hairpin ribonucleic acid (shRNA) was shown to potentiate CPT-induced apoptosis. Consistently, ectopic overexpression of Mcl-1 rescued cells from apoptosis induced by CPT. Cells coexpressing Noxa and Mcl-1 at different ratio correlates well with the extent of apoptosis, suggesting that the balance between Noxa and Mcl-1 may determine the susceptibility of HeLa cells to CPT-induced apoptosis. PMID:17971907

  19. The cell on the edge of life and death: Crosstalk between autophagy and apoptosis.

    Science.gov (United States)

    Kasprowska-Liśkiewicz, Daniela

    2017-09-21

    Recently, the crosstalk between autophagy and apoptosis has attracted broader attention. Basal autophagy serves to maintain cell homeostasis, while the upregulation of this process is an element of stress response that enables the cell to survive under adverse conditions. Autophagy may also determine the fate of the cell through its interactions with cell death pathways. The protein networks that control the initiation and the execution phase of these two processes are highly interconnected. Several scenarios for the crosstalk between autophagy and apoptosis exist. In most cases, the activation of autophagy represents an attempt of the cell to cope with stress, and protects the cell from apoptosis or delays its initiation. Generally, the simultaneous activation of pro-survival and pro-death pathways is prevented by the mutual inhibitory crosstalk between autophagy and apoptosis. But in some circumstances, autophagy or the proteins of the core autophagic machinery may promote cellular demise through excessive self-digestion (so-called "autophagic cell death") or by stimulating the activation of other cell death pathways. It is controversial whether cells actually die via autophagy, which is why the term "autophagic cell death" has been under intense debate lately. This review summarizes the recent findings on the multilevel crosstalk between autophagy and apoptosis in aspects of common regulators, mutual inhibition of these processes, the stimulation of apoptosis by autophagy or autophagic proteins and finally the role of autophagy as a death-execution mechanism.

  20. Modeling Glaucoma: Retinal Ganglion Cells Generated from Induced Pluripotent Stem Cells of Patients with SIX6 Risk Allele Show Developmental Abnormalities.

    Science.gov (United States)

    Teotia, Pooja; Van Hook, Matthew J; Wichman, Christopher S; Allingham, R Rand; Hauser, Michael A; Ahmad, Iqbal

    2017-11-01

    Glaucoma represents a group of multifactorial diseases with a unifying pathology of progressive retinal ganglion cell (RGC) degeneration, causing irreversible vision loss. To test the hypothesis that RGCs are intrinsically vulnerable in glaucoma, we have developed an in vitro model using the SIX6 risk allele carrying glaucoma patient-specific induced pluripotent stem cells (iPSCs) for generating functional RGCs. Here, we demonstrate that the efficiency of RGC generation by SIX6 risk allele iPSCs is significantly lower than iPSCs-derived from healthy, age- and sex-matched controls. The decrease in the number of RGC generation is accompanied by repressed developmental expression of RGC regulatory genes. The SIX6 risk allele RGCs display short and simple neurites, reduced expression of guidance molecules, and immature electrophysiological signature. In addition, these cells have higher expression of glaucoma-associated genes, CDKN2A and CDKN2B, suggesting an early onset of the disease phenotype. Consistent with the developmental abnormalities, the SIX6 risk allele RGCs display global dysregulation of genes which map on developmentally relevant biological processes for RGC differentiation and signaling pathways such as mammalian target of rapamycin that integrate diverse functions for differentiation, metabolism, and survival. The results suggest that SIX6 influences different stages of RGC differentiation and their survival; therefore, alteration in SIX6 function due to the risk allele may lead to cellular and molecular abnormalities. These abnormalities, if carried into adulthood, may make RGCs vulnerable in glaucoma. Stem Cells 2017;35:2239-2252. © 2017 AlphaMed Press.

  1. Structural analysis of retinal photoreceptor ellipsoid zone and postreceptor retinal layer associated with visual acuity in patients with retinitis pigmentosa by ganglion cell analysis combined with OCT imaging

    Science.gov (United States)

    Liu, Guodong; Li, Hui; Liu, Xiaoqiang; Xu, Ding; Wang, Fang

    2016-01-01

    Abstract The aim of this study was to examine changes in photoreceptor ellipsoid zone (EZ) and postreceptor retinal layer in retinitis pigmentosa (RP) patients by ganglion cell analysis (GCA) combined with optical coherence tomography (OCT) imaging to evaluate the structure–function relationships between retinal layer changes and best corrected visual acuity (BCVA). Sixty-eight eyes of 35 patients with RP and 65 eyes of 35 normal controls were analyzed in the study. The average length of EZ was 911.1 ± 208.8 μm in RP patients, which was shortened with the progression of the disease on the OCT images. The average ganglion cell–inner plexiform layer thickness (GCIPLT) was 54.7 ± 18.9 μm in RP patients, while in normal controls it was 85.6 ± 6.8 μm. The GCIPLT in all quarters became significantly thinner along with outer retinal thinning. There was a significantly positive correlation between BCVA and EZ (r = −0.7622, P retinal layer changes from a new perspective in RP patients, which suggests that EZ and GCIPLT obtained by GCA combined with OCT imaging are the direct and valid indicators to diagnosis and predict the pathological process of RP. PMID:28033301

  2. Chlorella vulgaris Induces Apoptosis of Human Non-Small Cell Lung Carcinoma (NSCLC) Cells.

    Science.gov (United States)

    Zhang, Zhi-Dong; Liang, Kai; Li, Kun; Wang, Guo-Quan; Zhang, Ke-Wei; Cai, Lei; Zhai, Shui-Ting; Chou, Kuo-Chen

    2017-01-01

    Chlorella vulgaris (C. vulgaris), a unicellular green microalga, has been widely used as a food supplement and reported to have antioxidant and anticancer properties. The current study was designed to assess the cytotoxic, apoptotic, and DNA-damaging effects of C. vulgaris growth factor (CGF), hot water C. vulgaris extracts, inlung tumor A549 and NCI-H460 cell lines. A549 cells, NCI-H460 cells, and normal human fibroblasts were treated with CGF at various concentrations (0-300 μg/ml) for 24 hr. The comet assay and γH2AX assay showed DNA damage in A549 and NCI-H460 cells upon CGF exposure. Evaluation of apoptosis by the TUNEL assay and DNA fragmentation analysis by agarose gel electrophoresis showed that CGF induced apoptosis in A549 and NCI-H460 cells. Chlorella vulgaris hot water extract induced apoptosis and DNA damage in human lung carcinoma cells. CGF can thus be considered a potential cytotoxic or genotoxic drug for treatment of lung carcinoma. Copyright© Bentham Science Publishers; For any queries, please email at epub@benthamscience.org.

  3. The role of heat shock protein 90 in the regulation of tumor cell apoptosis.

    Science.gov (United States)

    Kaigorodova, E V; Ryazantseva, N V; Novitskii, V V; Belkina, M V; Maroshkina, A N

    2011-02-01

    Programmed death of Jurkat tumor cells was studied under conditions of culturing with 17-AAG selective inhibitor of heat shock protein with a molecular weight of 90 kDa and etoposide. Apoptosis realization was evaluated by fluorescent microscopy with FITC-labeled annexin V and propidium iodide. Activity of caspase-3 was evaluated spectrophotometrically. Inhibition of heat shock protein with a molecular weight of 90 kDa activated the apoptotic program in Jurkat tumor cells and etoposide-induced apoptosis. The heat shock protein with a molecular weight of 90 kDa acted as apoptosis inhibitor in tumor cells.

  4. Zinc finger protein 598 inhibits cell survival by promoting UV-induced apoptosis.

    Science.gov (United States)

    Yang, Qiaohong; Gupta, Romi

    2018-01-19

    UV is one of the major causes of DNA damage induced apoptosis. However, cancer cells adopt alternative mechanisms to evade UV-induced apoptosis. To identify factors that protect cancer cells from UV-induced apoptosis, we performed a genome wide short-hairpin RNA (shRNA) screen, which identified Zinc finger protein 598 (ZNF598) as a key regulator of UV-induced apoptosis. Here, we show that UV irradiation transcriptionally upregulates ZNF598 expression. Additionally, ZNF598 knockdown in cancer cells inhibited UV-induced apoptosis. In our study, we observe that ELK1 mRNA level as well as phosphorylated ELK1 levels was up regulated upon UV irradiation, which was necessary for UV irradiation induced upregulation of ZNF598. Cells expressing ELK1 shRNA were also resistant to UV-induced apoptosis, and phenocopy ZNF598 knockdown. Upon further investigation, we found that ZNF598 knockdown inhibits UV-induced apoptotic gene expression, which matches with decrease in percentage of annexin V positive cell. Similarly, ectopic expression of ZNF598 promoted apoptotic gene expression and also increased annexin V positive cells. Collectively, these results demonstrate that ZNF598 is a UV irradiation regulated gene and its loss results in resistance to UV-induced apoptosis.

  5. Dioscin induces caspase-independent apoptosis through activation of apoptosis-inducing factor in breast cancer cells.

    Science.gov (United States)

    Kim, Eun-Ae; Jang, Ji-Hoon; Lee, Yun-Han; Sung, Eon-Gi; Song, In-Hwan; Kim, Joo-Young; Kim, Suji; Sohn, Ho-Yong; Lee, Tae-Jin

    2014-07-01

    Dioscin, a saponin extracted from the roots of Polygonatum zanlanscianense, shows several bioactivities such as antitumor, antifungal, and antiviral properties. Although, dioscin is already known to induce cell death in variety cancer cells, the molecular basis for dioscin-induced cell death was not definitely known in cancer cells. In this study, we found that dioscin treatment induced cell death in dose-dependent manner in breast cancer cells such as MDA-MB-231, MDA-MB-453, and T47D cells. Dioscin decreased expressions of Bcl-2 and cIAP-1 proteins, which were down-regulated at the transcriptional level. Conversely, Mcl-1 protein level was down-regulated by facilitating ubiquitin/proteasome-mediated Mcl-1 degradation in dioscin-treated cells. Pretreatment with z-VAD fails to attenuate dioscin-induced cell death as well as caspase-mediated events such as cleavages of procaspase-3 and PARP. In addition, dioscin treatment increased the population of annexin V positive cells and induced DNA fragmentation in a dose-dependent manner in MDA-MB-231 cells. Furthermore, apoptosis inducing factor (AIF) was released from the mitochondria and translocated to the nucleus. Suppression in AIF expression by siRNA reduced dioscin-induced apoptosis in MDA-MB-231 cells. Taken together, our results demonstrate that dioscin-induced cell death was mediated via AIF-facilitating caspase-independent pathway as well as down-regulating anti-apoptotic proteins such as Bcl-2, cIAP-1, and Mcl-1 in breast cancer cells.

  6. Troglitazone inhibits cell growth and induces apoptosis of B-cell acute lymphoblastic leukemia cells with t(14;18).

    Science.gov (United States)

    Takenokuchi, M; Saigo, K; Nakamachi, Y; Kawano, S; Hashimoto, M; Fujioka, T; Koizumi, T; Tatsumi, E; Kumagai, S

    2006-01-01

    Peroxisome proliferator-activated receptor-gamma (PPARgamma), a member of the nuclear receptor superfamily, has been detected in several human leukemia cells. Recent studies reported that PPARgamma ligands inhibit cell proliferation and induce apoptosis in both normal and malignant B-lineage cells. We investigated the expression of PPARgamma and the effects of PPARgamma ligands on UTree-O2, Bay91 and 380, three B-cell acute lymphoblastic leukemia (B-ALL) cell lines with t(14;18), which show a poor prognosis, accompanying c-myc abnormality. Western blot analysis identified expression of PPARgamma protein and real-time PCR that of PPARgamma mRNA on the three cell lines. Troglitazone (TGZ), a synthetic PPARgamma ligand, inhibited cell growth in these cell lines in a dose-dependent manner, which was associated with G(1) cell cycle arrest and apoptosis. We also found this effect PPARgamma independent since PPARgamma antagonists failed to reverse this effect. We assessed the expression of c-myc, an apoptosis-regulatory gene, since c-myc abnormality was detected in most B-ALL cells with t(14;18). TGZ was found to dose-dependently downregulate the expression of c-myc mRNA and c-myc protein in the three cell lines. These results suggest that TGZ inhibits cell growth via induction of G(1) cell cycle arrest and apoptosis in these cell lines and that TGZ-induced apoptosis, at least in part, may be related to the downregulation of c-myc expression. Moreover, the downregulation of c-myc expression by TGZ may depend on a PPARgamma-independent mechanism. Further studies indicate that PPARgamma ligands may serve as a therapeutic agent in B-ALL with t(14;18).

  7. Apoptosis related genes expressed in cultured Fallopian tube epithelial cells infected in vitro with Neisseria gonorrhoeae

    Directory of Open Access Journals (Sweden)

    PAZ A REYES

    2007-01-01

    Full Text Available Background: Infection of the Fallopian tubes (FT by Neisseria gonorrhoeae (Ngo can lead to acute salpingitis, an inflammatory condition resulting in damage primarily to the ciliated cells, with loss of ciliary activity and sloughing of the cells from the epithelium. Recently, we have shown that Ngo infection induced apoptosis in FT epithelium cells by a TNF-alpha dependent mechanism that could contribute to the cell and tissue damage observed in gonococcal salpingitis. Aim: To investigate the apoptosis-related genes expressed during apoptosis induction in cultured FT epithelial cells infected in vitro by Ngo. Materials and Methods: In the current study, we used cDNA macroarrays and real time PCR to identify and determine the expression levels of apoptosis related genes during the in vitro gonococci infection of FT epithelial cells. Results: Significant apoptosis was induced following infection with Ngo. Macroarray analysis identified the expression of multiple genes of the TNF receptor family (TNFRSF1B, -4, -6, -10A, -10B and -10D and the Bcl-2 family (BAK1, BAX, BLK, HRK and MCL-1 without differences between controls and infected cells. This lack of difference was confirmed by RT-PCR of BAX, Bcl-2, TNFRS1A (TNFR-I and TNFRSF1B (TNFR-II. Conclusion: Several genes related to apoptosis are expressed in primary cultures of epithelial cells of the human Fallopian tube. Infection with Ngo induces apoptosis without changes in the pattern of gene expression of several apoptosis-related genes. Results strongly suggest that Ngo regulates apoptosis in the FT by post-transcriptional mechanisms that need to be further addressed

  8. LP-THAE induced tumor cell apoptosis of rabbit VX2 liver carcinoma

    International Nuclear Information System (INIS)

    Chen Shengli; Quan Yi; Huang Zicheng; Chen Guodong; Zhu Dongliang

    2007-01-01

    Objective: To research tumor cell apoptosis induced by Lp-THAE of rabbit VX2 liver implanted tumor. Methods: 27 New Zealand white rabbits implanted with VX2 tumor at left middle lobe of the liver divided into three groups: Group A(n= 9) Lp-THAE: treated through transhepatic artery catheterization; Group B(n=9) THAI and Group C(n=9) as control. The rabbits were executed at second to fifth day after treatment. HE dye microscopy was taken for counting the typical apoptosis cells and calculating apoptosis index (ApI). FITC-AnnexinV/PI assay was used for measuring apoptosis by flow cytometry. Results: The ApI of tumor central area and marginal area were (17.769±2.417)%, (4.129±1.172)%, P<0.01. The percentages of tumor cell apoptosis and tumor cell necrosis were (16.483±1.404)%, (9.478±0.964)%, P<0.01 and (43.559±5.053)%, (33.460±1.840)%, P=0.093. The total percentages of tumor cell apoptosis and necrosis were (60.042±13.979)%, (42.938±8.979)%, P< 0.01, at tumor center and marginal area in THAE group respectively. The ApI, percentages of tumor cell apoptosis and necrosis in THAE group were significantly higher than those of THAI group (P<0.01). The percentages of tumor cell apoptosis at tumor center area in THAE group were significantly higher than those of tumor marginal area(P<0.01). Conclusion: Induced tumor cell apoptosis and necrosis are two mechanisms of action for Lp-THAE treatment of liver carcinoma. (authors)

  9. Spatiotemporal distribution of PAX6 and MEIS2 expression and total cell numbers in the ganglionic eminence in the early developing human forebrain

    DEFF Research Database (Denmark)

    Larsen, Karen B; Lutterodt, Melissa C; Laursen, Henning

    2010-01-01

    The development of the human neocortex is a complex and highly regulated process involving a time-related expression of many transcription factors including the homeobox genes Pax6 and Meis2. During early development, Pax6 is expressed in nuclei of radial glia cells in the neocortical proliferative...... in the same time window. We demonstrate by in situ hybridization and immunohistochemistry that the two homeobox genes are expressed during early fetal brain development in humans. PAX6 mRNA and protein were located in the proliferative zones of the neocortex and in single cells in the cortical preplate at 7...... in the proliferative zones of the human fetal neocortex and a higher expression of MEIS2 than PAX6 was observed in these areas at 9 fetal weeks. Further, MEIS2 was expressed at a very high level in the developing ganglionic eminence and at a more moderate level in the cortical plate....

  10. Downregulation of Akt1 Inhibits Anchorage-Independent Cell Growth and Induces Apoptosis in Cancer Cells

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    Xuesong Liu

    2001-01-01

    Full Text Available The serine/threonine kinases, Akti/PKBα, Akt2/PKBβ, and Akt3/PKBγ, play a critical role in preventing cancer cells from undergoing apoptosis. However, the function of individual Akt isoforms in the tumorigenicity of cancer cells is still not well defined. In the current study, we used an AM antisense oligonucleotide (AS to specifically downregulate Akti protein in both cancer and normal cells. Our data indicate that AM AS treatment inhibits the ability of MiaPaCa-2, H460, HCT-15, and HT1080 cells to grow in soft agar. The treatment also induces apoptosis in these cancer cells as demonstrated by FRCS analysis and a caspase activity assay. Conversely, Akti AS treatment has little effect on the cell growth and survival of normal human cells including normal human fibroblast (NHF, fibroblast from muscle (FBM, and mammary gland epithelial 184135 cells. In addition, AM AS specifically sensitizes cancer cells to typical chemotherapeutic agents. Thus, Akti is indispensable for maintaining the tumorigenicity of cancer cells. Inhibition of AM may provide a powerful sensitization agent for chemotherapy specifically in cancer cells.

  11. The ganglioside GM3 is associated with cisplatin-induced apoptosis in human colon cancer cells.

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    Tae-Wook Chung

    Full Text Available Cisplatin (cis-diamminedichloroplatinum, CDDP is a well-known chemotherapeutic agent for the treatment of several cancers. However, the precise mechanism underlying apoptosis of cancer cells induced by CDDP remains unclear. In this study, we show mechanistically that CDDP induces GM3-mediated apoptosis of HCT116 cells by inhibiting cell proliferation, and increasing DNA fragmentation and mitochondria-dependent apoptosis signals. CDDP induced apoptosis within cells through the generation of reactive oxygen species (ROS, regulated the ROS-mediated expression of Bax, Bcl-2, and p53, and induced the degradation of the poly (ADP-ribosyl polymerase (PARP. We also checked expression levels of different gangliosides in HCT116 cells in the presence or absence of CDDP. Interestingly, among the gangliosides, CDDP augmented the expression of only GM3 synthase and its product GM3. Reduction of the GM3 synthase level through ectopic expression of GM3 small interfering RNA (siRNA rescued HCT116 cells from CDDP-induced apoptosis. This was evidenced by inhibition of apoptotic signals by reducing ROS production through the regulation of 12-lipoxigenase activity. Furthermore, the apoptotic sensitivity to CDDP was remarkably increased in GM3 synthase-transfected HCT116 cells compared to that in controls. In addition, GM3 synthase-transfected cells treated with CDDP exhibited an increased accumulation of intracellular ROS. These results suggest the CDDP-induced oxidative apoptosis of HCT116 cells is mediated by GM3.

  12. Bupivacaine-induced apoptosis independently of WDR35 expression in mouse neuroblastoma Neuro2a cells

    Science.gov (United States)

    2012-01-01

    Background Bupivacaine-induced neurotoxicity has been shown to occur through apoptosis. Recently, bupivacaine was shown to elicit reactive oxygen species (ROS) production and induce apoptosis accompanied by activation of p38 mitogen-activated protein kinase (MAPK) in a human neuroblastoma cell line. We have reported that WDR35, a WD40-repeat protein, may mediate apoptosis through caspase-3 activation. The present study was undertaken to test whether bupivacaine induces apoptosis in mouse neuroblastoma Neuro2a cells and to determine whether ROS, p38 MAPK, and WDR35 are involved. Results Our results showed that bupivacaine induced ROS generation and p38 MAPK activation in Neuro2a cells, resulting in apoptosis. Bupivacaine also increased WDR35 expression in a dose- and time-dependent manner. Hydrogen peroxide (H2O2) also increased WDR35 expression in Neuro2a cells. Antioxidant (EUK-8) and p38 MAPK inhibitor (SB202190) treatment attenuated the increase in caspase-3 activity, cell death and WDR35 expression induced by bupivacaine or H2O2. Although transfection of Neuro2a cells with WDR35 siRNA attenuated the bupivacaine- or H2O2-induced increase in expression of WDR35 mRNA and protein, in contrast to our previous studies, it did not inhibit the increase in caspase-3 activity in bupivacaine- or H2O2-treated cells. Conclusions In summary, our results indicated that bupivacaine induced apoptosis in Neuro2a cells. Bupivacaine induced ROS generation and p38 MAPK activation, resulting in an increase in WDR35 expression, in these cells. However, the increase in WDR35 expression may not be essential for the bupivacaine-induced apoptosis in Neuro2a cells. These results may suggest the existence of another mechanism of bupivacaine-induced apoptosis independent from WDR35 expression in Neuro2a cells. PMID:23227925

  13. Apoptosis-like yeast cell death in response to DNA damage and replication defects

    Energy Technology Data Exchange (ETDEWEB)

    Burhans, William C.; Weinberger, Martin; Marchetti, Maria A.; Ramachandran, Lakshmi; D' Urso, Gennaro; Huberman, Joel A

    2003-11-27

    In budding (Saccharomyces cerevisiae) and fission (Schizosaccharomyces pombe) yeast and other unicellular organisms, DNA damage and other stimuli can induce cell death resembling apoptosis in metazoans, including the activation of a recently discovered caspase-like molecule in budding yeast. Induction of apoptotic-like cell death in yeasts requires homologues of cell cycle checkpoint proteins that are often required for apoptosis in metazoan cells. Here, we summarize these findings and our unpublished results which show that an important component of metazoan apoptosis recently detected in budding yeast - reactive oxygen species (ROS) - can also be detected in fission yeast undergoing an apoptotic-like cell death. ROS were detected in fission and budding yeast cells bearing conditional mutations in genes encoding DNA replication initiation proteins and in fission yeast cells with mutations that deregulate cyclin-dependent kinases (CDKs). These mutations may cause DNA damage by permitting entry of cells into S phase with a reduced number of replication forks and/or passage through mitosis with incompletely replicated chromosomes. This may be relevant to the frequent requirement for elevated CDK activity in mammalian apoptosis, and to the recent discovery that the initiation protein Cdc6 is destroyed during apoptosis in mammals and in budding yeast cells exposed to lethal levels of DNA damage. Our data indicate that connections between apoptosis-like cell death and DNA replication or CDK activity are complex. Some apoptosis-like pathways require checkpoint proteins, others are inhibited by them, and others are independent of them. This complexity resembles that of apoptotic pathways in mammalian cells, which are frequently deregulated in cancer. The greater genetic tractability of yeasts should help to delineate these complex pathways and their relationships to cancer and to the effects of apoptosis-inducing drugs that inhibit DNA replication.

  14. Apoptosis-like yeast cell death in response to DNA damage and replication defects

    International Nuclear Information System (INIS)

    Burhans, William C.; We